cyclin-d1 and Uterine-Cervical-Neoplasms

Many studies have evaluated the association between cyclin D1 (CCND1) G870A polymorphism and cervical cancer susceptibility. However, these studies showed inconsistent results. The aim of this study was to derive a more precise estimation of this association. We searched PubMed and Embase for related studies that had been published in English, and ten case-control studies with a total of 2,864 cases and 3,898 controls were finally identified to be eligible studies in the meta-analysis. The association was assessed by summarizing the odds ratios (ORs) with the corresponding 95 % confidence intervals (CIs). Overall, there was no significant association between cyclin D1 (CCND1) G870A polymorphism and cervical cancer risk (for the allele model A vs. G: OR = 1.02, 95 % CI 0.88-1.19, p = 0.76; for the co-dominant model AA vs. GG: OR = 1.03, 95 % CI 0.75-1.41, p = 0.85; for the dominant model AA + GA vs. GG: OR = 1.00, 95 % CI 0.78-1.28, p = 0.99; for the recessive comparison AA vs. GA + GG: OR = 1.06, 95 % CI 0.85-1.32, p = 0.62). In subgroup analysis by ethnicity, no significant difference was found in both Asians and Caucasians. In summary, the present meta-analysis provides evidence that genotypes for the cyclin D1 (CCND1) G870A polymorphism may be not associated with genetic susceptibility of cervical cancer.

Topics: Asian People; Case-Control Studies; Cyclin D1; Female; Genetic Association Studies; Genetic Predisposition to Disease; Humans; Polymorphism, Single Nucleotide; Risk Factors; Uterine Cervical Neoplasms; White People

Association between Cyclin D1 (CCND1) polymorphism and cervical cancer risk are conflicting with published articles. We performed a meta-analysis to investigate the association between CCND1 G870A polymorphism and cervical cancer risk.. PubMed, Embase and CNKI data were researched to conduct a meta-analysis on the associations between CCND1 G870A polymorphism and cervical cancer risk. Ten published case-control studies including 2,864 patients with cervical cancer and 3,898 controls were collected in this meta-analysis. Odds ratio (OR) with 95% confidence interval (CI) were applied to assess the relationship; meta-regression, sensitivity analysis and cumulative analysis were also conducted to guarantee the strength of results.. Overall, no significant association between CCND1 G870A polymorphism and cervical cancer risk were found in allele contrast (A vs. G: OR=1.02, 95% CI=0.88-1.19, P=0.76 I2=74.5%), codominant model (GA vs. GG: OR=0.98, 95% CI=0.77-1.26, P=0.90 I2=69.1%; AA vs GG: OR=1.03, 95% CI=0.75-1.41, P=0.85 I2=75.9%), dominant model (GA + AA vs. GG: OR=1.00, 95% CI=0.78-1.28, P=0.99 I2=72.3%) and recessive model (AA vs GG + GA: OR=1.06, 95% CI=0.85-1.23, P=0.62, I2=70.1%). Similarly, in the stratified analysis by ethnicity, study design and genotyping type, no significant association detected in all genetic models either.. Our meta-analysis indicated that CCND1 G870A might be not the crucial risk factor for the development of cervical cancer.. The virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/13000_2014_168.

Topics: Alleles; Case-Control Studies; Cyclin D1; Female; Genetic Predisposition to Disease; Genotype; Humans; Odds Ratio; Polymorphism, Single Nucleotide; Risk; Risk Factors; Uterine Cervical Neoplasms

Cyclin D1 (CCND1) is a key regulatory protein in the G1/S checkpoint of the cell cycle. We hypothesized that the G870A polymorphism of CCND1 is associated with the risk for cervical cancer and performed a meta-analysis of eligible studies to evaluate this relationship.. In a case-control study of 300 patients with newly diagnosed cervical cancer and 312 cancer-free controls who were frequency-matched by age, we genotyped the G870A polymorphism of CCND1 by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). A meta-analysis was performed using five case-control studies, including our results.. No significant association was observed between the G870A polymorphism of CCND1 and cervical cancer risk. Further, in our meta-analysis, there was no significant risk of cervical cancer that correlated with the G870A polymorphism of CCND1. In the stratified analysis by race, however, individuals who harbored the AA or AA/AG genotypes were at significantly greater risk compared with GG carriers in the Asian population.. The G870A polymorphism in CCND1 may not contribute to the etiology of cervical cancer in Chinese populations.

Topics: Adult; Case-Control Studies; Cyclin D1; Female; Genetic Predisposition to Disease; Genotype; Humans; Middle Aged; Polymerase Chain Reaction; Polymorphism, Restriction Fragment Length; Polymorphism, Single Nucleotide; Uterine Cervical Neoplasms

Carcinoma of the uterine cervix is one of the most common malignancies among women worldwide. Human papillomaviruses (HPV) have been identified as the major etiological factor in cervical carcinogenesis. However, the time lag between HPV infection and the diagnosis of cancer indicates that multiple steps, as well as multiple factors, may be necessary for the development of cervical cancer. The development and progression of cervical carcinoma have been shown to be dependent on various genetic and epigenetic events, especially alterations in the cell cycle checkpoint machinery. In mammalian cells, control of the cell cycle is regulated by the activity of cyclin-dependent kinases (CDKs) and their essential activating coenzymes, the cyclins. Generally, CDKs, cyclins, and CDK inhibitors function within several pathways, including the p16(INK4A)-cyclin D1-CDK4/6-pRb-E2F, p21(WAF1)- p27(KIP1)-cyclinE-CDK2, and p14(ARF)-MDM2-p53 pathways. The results from several studies showed aberrant regulation of several cell cycle proteins, such as cyclin D, cyclin E, p16(INK4A), p21(WAF1), and p27(KIP1), as characteristic features of HPV- infected and HPV E6/E7 oncogene-expressing cervical carcinomas and their precursors. These data suggested further that interactions of viral proteins with host cellular proteins, particularly cell cycle proteins, are involved in the activation or repression of cell cycle progression in cervical carcinogenesis.

Topics: Cell Cycle; Cyclin D1; Cyclin E; Cyclin-Dependent Kinase 2; Cyclin-Dependent Kinase 4; Cyclin-Dependent Kinase Inhibitor p16; Cyclin-Dependent Kinase Inhibitor p21; Cyclin-Dependent Kinase Inhibitor p27; E2F Transcription Factors; Female; Humans; Proto-Oncogene Proteins c-mdm2; Retinoblastoma Protein; Tumor Suppressor Protein p14ARF; Tumor Suppressor Protein p53; Uterine Cervical Neoplasms

The quest for strong, safe and cost-effective natural antiproliferative agents that could reduce cancer has been the focus now a days. In this regard, the organosulfur compounds from garlic (Allium sativum L.), like Diallyl Sulfide (DAS) and Diallyl Disulfide (DADS), have been shown to exhibit potent antiproliferative and anticancer properties in many studies. However, the potential of these compounds against viral oncoproteins in cervical cancer has not been fully elucidated yet.. The objective of this study was to analyze the antiproliferative and apoptotic properties of DADS and DAS in HPV16+ human cervical cancer Caski cell line.. Caski (cervical cancer cells) were cultured and followed by the treatment of various concentrations of organosulphur compounds (DADS and DAS), cell viability was measured by MTT assay. The apoptotic assay was performed by DAPI and Hoechst3342 staining. Reactive Oxygen Species (ROS) was estimated by DCFDA staining protocol. The distributions of cell cycle and apoptosis (FITC-Annexin V assay) were analyzed by flow cytometry. Finally, gene expression analysis was performed via quantitative real time PCR.. Our results showed that DAS and DADS exerted a significant antiproliferative effect on Caski cells by reducing the cell viability and inducing a dose-related increment in intracellular ROS production along with apoptosis in Caski cells. DAS and DADS also induced cell cycle arrest in G0/G1 phase, which was supported by the downregulation of cyclin D1 and CDK4 and upregulation of CDK inhibitors p21WAF1/CIP1 and p27KIP1 in Caski cells. Additionally, DAS and DADS lead to the downregulation of viral oncogene E6 and E7 and restoration of p53 function.. Thus, this study confirms the efficacy of both the organosulfur compounds DADS and DAS against cervical cancer cells.

Topics: Antineoplastic Agents; Apoptosis; Cell Cycle Checkpoints; Cell Proliferation; Cell Survival; Cyclin D1; Cyclin-Dependent Kinase 4; Dose-Response Relationship, Drug; Down-Regulation; Drug Screening Assays, Antitumor; Female; Humans; Models, Molecular; Molecular Structure; Structure-Activity Relationship; Sulfur Compounds; Tumor Cells, Cultured; Uterine Cervical Neoplasms

The objective of the study is to investigate the biomarkers for diagnosis and prevention of human papillomavirus (HPV) infection-induced cervical cancer.. Cervical cancer tissues were collected from patients with cervical cancer, while noncancer tissues were collected from patients diagnosed with cervical lesions or uterine fibroids at the Chinese PLA General Hospital 301 and 309, China from December 2017 to June 2018. The cancer tissues were collected from the site of lesion, while the noncancer tissues were collected from similar anatomical locations. Quantitative real-time PCR, Western blot (WB), and immunohistochemistry (IHC) were used to detect the mRNA and protein levels of HPV E6/E7, RPRD1B (regulation of nuclear pre-mRNA domain containing 1B), cyclin D1, and transcription factor 4 (TCF4) between cervical cancer tissues and noncancer tissues. The correlation of HPV E6/E7, RPRD1B, cyclin D1, and TCF4 expressions was analyzed.. Twenty patients with cervical cancer and 27 controls without cervical cancer were included in this study. The mRNA expression of HPV E6/E7and RPRD1B was significantly higher in patients with cervical cancer than controls, while cyclin D1 mRNA expression was significantly lower in patients with cervical carcinoma in situ stage, compared with controls. RPRD1B protein expression was significantly higher in patients compared to controls when analyzed by IHC. TCF4 was significantly lower in clinical stage I and Ib of cervical cancer when analyzed by WB. The mRNA and protein expressions of RPRD1B and cyclin D1 were significantly different between patients younger than 50 years old, compared to patients 50 years and older.. HPV E6/E7 expression was associated with RPRD1B level in cervical cancer. The expression of RPRD1B and cyclin D1 in patients with cervical cancer might be affected by age.

Topics: Adult; Aged; Alphapapillomavirus; Case-Control Studies; Cell Cycle Proteins; China; Cyclin D1; Female; Humans; Middle Aged; Neoplasm Proteins; Oncogene Proteins, Viral; Papillomavirus E7 Proteins; Papillomavirus Infections; Repressor Proteins; RNA, Messenger; Transcription Factor 4; Uterine Cervical Neoplasms

Cervical cancer is the fourth most common female cancer worldwide. Long non-coding RNAs (lncRNAs), such as SOX21-AS1, play pivotal roles in the progression and metastasis of cancer. We previously described that SOX21-AS1 was hypomethylated in cervical cancer (CC) and aimed to further explore the relationship between methylation of the SOX21-AS1 promoter and CC using clinical cervical samples. Pyrosequencing was performed to detect the methylation status of the SOX21-AS1 promoter in 33 cervical specimens. Additionally, expression levels of related genes in 43 clinical cervical specimens were measured using quantitative real-time PCR (qRT-PCR). The SOX21-AS1 promoter was significantly hypomethylated in CC (P < 0.01). SOX21-AS1 hypomethylation was also significantly associated with an advanced Federation of Gynecology and Obstetrics (FIGO) stage (P < 0.01). The expression levels of SOX21-AS1 and SOX21 were noted to be higher in cancer vs. normal cervix (all P < 0.001). Moreover, the expression of SOX21-AS1 was positively correlated with SOX21 in all samples (r = 0.891, P < 0.001). Methylation statue of the SOX21-AS1 promoter region was negatively correlated with the expression levels of SOX21-AS1 and SOX21 (SOX21-AS1, r = - 0.628; SOX21, r = - 0.648; both P < 0.001). The methylation status of SOX21-AS1 displayed promising diagnostic potential for CC, exhibiting good sensitivity (100.0%) and specificity (69.2%), with an area under the curve of 0.846. In addition, bioinformatic analyses identified a potential link between SOX21-AS1 and the Wnt signaling pathway. Furthermore, methylation status of SOX21-AS1 was negatively correlated with β-catenin/c-myc/cyclin D1 mRNA levels (r

Topics: Adult; Aged; beta Catenin; Biomarkers, Tumor; Cyclin D1; DNA Methylation; Female; Gene Expression Regulation, Neoplastic; Humans; Middle Aged; Promoter Regions, Genetic; Proto-Oncogene Proteins c-myc; RNA, Long Noncoding; Uterine Cervical Neoplasms; Wnt Signaling Pathway

Oncogenic high-risk human papillomavirus (HR-HPV) infection causes a majority of cases of cervical cancer and pre-cancerous cervical lesions. However, the mechanisms underlying the direct evolution from HPV-16/18-infected epithelium to cervical intraepithelial neoplasia (CIN) III, which can progress to cervical cancer, remain poorly identified. Here, we performed RNA-seq after laser capture microdissection, and found that APOBEC3B was highly expressed in cervical cancer specimens compared with CIN III with HPV-16/18 infection. Furthermore, immunohistochemical analysis confirmed that high levels of APOBEC3B were correlated with lymph node metastasis in cervical cancer. Subsequent experiments revealed that HPV-16 E6 could upregulate APOBEC3B through direct binding to the promoter of APOBEC3B in cervical cancer cells. Silencing of APOBEC3B by stable short hairpin RNA-mediated knockdown reduced the proliferative capacity of Caski and HeLa cells in vitro and in vivo, but had only a small effect on the migration and invasion of two cervical cancer cell lines. Finally, we identified the changes in gene expression following APOBEC3B silencing in Caski cells by microarray, demonstrating a biological link between APOBEC3B and CCND1 in cervical cancer cells. Importantly, through methyl-capture sequencing and pyrosequencing, APOBEC3B was found to affect the levels of the downstream protein Cyclin D1 (which is encoded by the CCND1 gene) through hypomethylation of the CCND1 promoter. In conclusion, our study supports HPV-16 E6-induced APOBEC3B expression associates with proliferation of cervical cancer cells and hypomethylation of Cyclin D1. Thus, APOBEC3B may be a potential therapeutic target in human cervical cancer.

Topics: Animals; Cell Line, Tumor; Cell Proliferation; CpG Islands; Cyclin D1; Cytidine Deaminase; DNA Methylation; DNA-Binding Proteins; Female; Gene Expression Profiling; Gene Expression Regulation, Neoplastic; HeLa Cells; Human papillomavirus 16; Human papillomavirus 18; Humans; Mice; Minor Histocompatibility Antigens; Neoplasm Transplantation; Oncogene Proteins, Viral; Promoter Regions, Genetic; Repressor Proteins; Sequence Analysis, DNA; Sequence Analysis, RNA; Up-Regulation; Uterine Cervical Neoplasms

Topics: Case-Control Studies; Cell Cycle Proteins; Cyclin D1; Female; Humans; Middle Aged; Neoplasm Proteins; Prospective Studies; Transcription Factor 4; Uterine Cervical Neoplasms

HOXA5 is considered a regulator involved in embryonic development and cellular differentiation and a tumor suppressor. Nevertheless, its biological role in cervical carcinoma is still unclear. In the present study, immunohistochemistry showed that HOXA5 expression gradually decreased as the degree of cervical lesions deepened. Ectopic expression of HOXA5 restrained cell proliferation, decreased cell viability, and inhibited tumor formation in vitro and in vivo. Furthermore, the expression of HOXA5 could arrest cell cycle from G0/G1 to S phase. RNA-seq revealed that p21 and cyclinD1 were involved in this process. Moreover, the gene set enrichment analysis and the TOP/FOP reporter assay both suggested that HOXA5 could restrain the activity of the Wnt/β-catenin pathway. Further study using dual-luciferase reporter assay and quantitative chromatin immunoprecipitation assay demonstrated that HOXA5 could directly bind to the TAAT motif within the promoter of TP53 by its HD domain and transactivate TP53, which can upregulate p21. Altogether, our data suggest that HOXA5 inhibits the proliferation and neoplasia via repression activity of the Wnt/β-catenin pathway and transactivating TP53 in cervical cancer.

Topics: Animals; Carcinogenesis; Cell Cycle Checkpoints; Cell Line, Tumor; Cell Proliferation; Cyclin D1; Cyclin-Dependent Kinase Inhibitor p21; Down-Regulation; Female; Gene Expression Regulation, Neoplastic; Homeodomain Proteins; Humans; Mice, Inbred BALB C; Mice, Nude; Models, Biological; Promoter Regions, Genetic; Transcriptional Activation; Tumor Suppressor Protein p53; Up-Regulation; Uterine Cervical Neoplasms; Wnt Signaling Pathway

The deubiquitinating (DUB) enzyme ubiquitin-specific protease 18 (USP18), also known as UBP43, is an ubiquitin-specific protease linked to several human malignancies. However, USP18's underlying function in human cervical cancer remains unclear. In the current study, we aimed to analyse the role of USP18 and its signalling pathways in cervical cancer.. Quantitative real-time polymerase chain reaction (qRT-PCR) and immunohistochemical staining were performed to analyse USP18 levels in cervical cancer and matched to adjacent normal tissues. Moreover, RNA interference (RNAi) and lentiviral-mediated vector transfections were performed to silence and overexpress USP18, respectively, in cervical cancer cells. Further, Cell Counting Kit-8 (CCK-8) and Annexin V/PI staining assays were used to assess its biological function in cell proliferation and apoptosis, respectively. A xenograft model was used to examine USP18's function in vivo.. The present findings demonstrated that USP18 was overexpressed in cervical cancer specimens and cell lines. Silencing USP18 in SiHa and Caski cervical cancer cell lines inhibited cell proliferation, induced apoptosis, and promoted cleaved caspase-3 expression. In contrast, USP18 overexpression showed the opposite effects in human HcerEpic cells. A Gene Set Enrichment Analysis revealed that USP18 was enriched in the PI3K/AKT signalling pathway in cervical cancer. Hence, the PI3K/AKT inhibitor LY294002 was used to determine the relationship between USP18 and AKT in cervical cancer cells. Importantly, LY294002 significantly abolished the effects of USP18 overexpression in cervical cancer cells. In vivo, USP18 silencing inhibited human cervical cancer cells' tumorigenicity.. The current study indicates that USP18 is an oncogenic gene in cervical cancer. Our findings not only deepened the understanding of USP18's biological function in cervical cancer pathogenesis, but we also provided novel insight for cervical cancer therapy.. Retrospectively registered.

Topics: Animals; Apoptosis; Caspase 3; Cell Line, Tumor; Cell Proliferation; Cervix Uteri; Chromones; Cyclin D1; Elafin; Enzyme Inhibitors; Female; Gene Silencing; Humans; Ki-67 Antigen; Mice; Mice, Nude; Morpholines; Neoplasm Transplantation; Proto-Oncogene Proteins c-akt; Real-Time Polymerase Chain Reaction; Signal Transduction; Ubiquitin Thiolesterase; Up-Regulation; Uterine Cervical Neoplasms

The present study is to analyze the difference of gene methylation in early cervical adenocarcinoma and to find molecular markers for predicting the occurrence and development of cervical adenocarcinoma.A total of 15 cases of primary cervical adenocarcinoma and 10 cases of primary cervical squamous cell carcinoma at stages IB1 or IIA1 were included in the study. Infinium MethylationEPIC BeadChip (850K) was used to screen specifically expressed genes in cervical adenocarcinoma tissues. Bisulfite sequencing polymerase chain reaction (BSP) and quantitative real-time polymerase chain reaction (qRT-PCR) were used to verify the methylation levels in cervical adenocarcinoma, cervical squamous cell carcinoma, and normal cervical tissues.Sex determining region Y-box 1 (SOX1) and cyclin D1 (CCND1) genes participated in multiple signaling pathways, being the central nodes of gene regulatory networks. SOX1 gene, but not CCND1 gene, was a specifically methylated gene in cervical adenocarcinoma according to BSP. According to qRT-PCR, methylation level of SOX1 in cervical adenocarcinoma tissues is significantly different from that in cervical squamous cell carcinoma tissues or normal cervical tissues, and the methylation level of CCND1 in cervical adenocarcinoma tissues or cervical squamous cell carcinoma tissues is significantly different from that in normal cervical tissues.The present study demonstrates that tumor-suppressor gene SOX1 is a methylation-specific expression gene of cervical adenocarcinoma and is expected to become a specific molecular marker for the diagnosis of cervical adenocarcinoma. However, CCND1 gene was not proven to be a specific methylation expression gene in cervical adenocarcinoma in the present study.

Topics: Adenocarcinoma; Carcinoma, Squamous Cell; Cervix Uteri; Cyclin D1; DNA Methylation; Female; Gene Expression Regulation, Neoplastic; Gene Regulatory Networks; Genetic Markers; Humans; Real-Time Polymerase Chain Reaction; SOXB1 Transcription Factors; Uterine Cervical Neoplasms

Adenocarcinoma (ADC) of the uterine cervix (UC) is a rare form of cervical cancer (CC) caused due to the infection of Human Papilloma Virus (HPV). Cyclin D1 is one of the downstream targets of aberrantly activated Notch signaling, contribute to the etiology of CC. However, little is known about the role of Cyclin D1 in the modulation of cervical ADC and is controversial. The purpose of this study is to determine the role of Cyclin D1 protein and to elucidate the combined analysis with Notch signaling proteins in HPV associated ADCs of CC. A total of 60 biopsy samples (40 normal and 20 ADCs of CC) were analyzed for the expression of Cyclin D1 in HPV associated ADCs via immunohistochemistry and by immunoblotting. HPV-16 positive ADC patients showed a strong association with the Cyclin D1 expression (p = 0.007). The significant mean difference (p = 0.0001) and the pairwise comparison between Cyclin D1/JAG1 (p = 0.0001), and Cyclin D1/Notch-3 (p = 0.0001) were observed. The above Notch signaling proteins showed their synergistic role in modulating Cyclin D1 which in-turn regulates HPV-16 associated ADC of the uterine cervix (UC), affecting women's global health.

Topics: Adenocarcinoma; Adult; Blotting, Western; Cervix Uteri; Cyclin D1; Female; Global Health; Human papillomavirus 16; Humans; Immunohistochemistry; Jagged-1 Protein; Papillomavirus Infections; Receptor, Notch3; Uterine Cervical Neoplasms

p21, an inhibitor of cyclin-dependent kinase, functions as an oncogene or tumor suppressor depending on the context of a variety of extracellular and intracellular signals. The expression of p21 could be regulated at the transcriptional and/or post-translational levels. The p21 gene is well-known to be regulated in both p53-dependent and -independent manners. However, the detailed regulatory mechanisms of p21 messenger RNA and protein expression via statins remain unknown, and the possible application of statins as anticancer reagents remains to be controversial. Our data showed that the statins-fluvastatin and lovastatin-induced p21 expression as general histone deacetylase inhibitors in a p53-independent manner, which is mediated through various pathways, such as apoptosis, autophagy, cell cycle progression, and DNA damage, to be involved in the function of p21 in HeLa cells. The curative effect repositioning of digoxin, a cardiovascular medication, was combined with fluvastatin and lovastatin, and the results further implied that p21 induction is involved in a p53-dependent and p53-independent manner. Digoxin modified the effects of statins on ATF3, p21, p53, and cyclin D1 expression, while fluvastatin boosted its DNA damage effect and lovastatin impeded its DNA damage effect. Fluvastatin and lovastatin combined with digoxin further support the localization specificity of their interactivity with our subcellular localization data. This study will not only clarify the regulatory mechanisms of p21 induction by statins but will also shed light on the repurposing of widely cardiovascular medications for the treatment of cervical cancer.

Topics: Activating Transcription Factor 3; Apoptosis; Autophagy; Cell Cycle; Cyclin D1; Cyclin-Dependent Kinase Inhibitor p21; DNA Damage; Female; Fluvastatin; Gene Expression Regulation, Neoplastic; HeLa Cells; Histone Deacetylase Inhibitors; Humans; Lovastatin; Tumor Suppressor Protein p53; Uterine Cervical Neoplasms

Background: Cervical cancer is one of the most prevalent gynecological cancers worldwide and contributes in high\ mortality of Indonesian women. The efficacy of chemotherapy as a standart therapy for cervical cancer decreases because\ it frequenly rises adverse effects. Recent studies have found that metformin has a potential anticancer effect mostly\ through reduction of cyclin expression and activation of Activated Adenosine Monophosphate Kinase (AMPK). This\ study aimed to investigate the effect of metfomin on expression of cyclin D1 and p53 and apoptosis in HeLa cancer cell\ line. Methods: HeLa cells were treated with various doses of metformin and doxorubicin as a positive control. Cytotoxic\ effect of metformin was determined using the MTT assay. Immunocytochemistry was used to assess cyclin D1 and p53\ expression and apoptosis levels of treated HeLa cells were analyzed using flowcytometry. Data of cyclin D1 expression\ was statistically analyzed using the Kruskal-Wallis test followed by the Tamhane test, whilst ANOVA and Tukey post\ Hoc tests were used to analyze data of p53 and apoptosis level. The significant value was p< 0.05. Results: Metformin\ was able to inhibit proliferation of HeLa cells with IC50 60 mM. HeLa cells treated with 60 and 120 mM metformin\ had lower cyclin D1 expression than HeLa cells treated without metformin and reached a significant difference (p=\ 0.001). Moreover, 30 mM or higher doses of metformin increase significantly p53 expression (p< 0.001). Induction of\ apoptosis was observed in HeLa cells treated with all doses of metformin and reached statistically difference (p= 0.04\ and p < 0.001). Conclusion: Metformin can modulate cyclin D1 and p53 expression in HeLa cancer cell line, leading\ to inhibition of cell proliferation and induction of apoptosis. Other cyclin family members, CDK inhibitors and AMPK\ signaling should be further investigated in order to know mechanism of metformin action.

Topics: Apoptosis; Cell Proliferation; Cyclin D1; Female; Gene Expression Regulation, Neoplastic; HeLa Cells; Humans; Hypoglycemic Agents; Metformin; Signal Transduction; Tumor Suppressor Protein p53; Uterine Cervical Neoplasms

The purpose of this study was to investigate the inhibitory effect of baicalein on the proliferation of cervical carcinoma cells and stimulate cervical carcinoma cells with baicalein. MTT method was used to observe cell proliferation. Flow cytometry was used to observe cell cycle, and gene technology was used to observe the expression of corresponding genes at the level of gene and protein. β-catenin activity was assessed using Western blot and ChIP. Baicalein suppressed cervical carcinoma cell HeLa proliferation by enhancing the activity of caspase-3. Baicalein blocked cell cycle at G0/G1 stage by inhibiting the expression of some genes. At the same time, it can prevent the nuclear translocation of β-catenin and inhibit the activity of Wnt. When the Wnt signaling pathway is increased, the proliferation of HeLa cells is inhibited, and apoptosis is promoted in this way. In conclusion, it indicated that baicalein inhibits cervical carcinoma progression by targeting CCND1 via Wnt/β-catenin signaling pathway.

Topics: Antineoplastic Agents; Apoptosis; Cell Line, Tumor; Cell Proliferation; Cyclin D1; Female; Flavanones; Humans; Molecular Targeted Therapy; Up-Regulation; Uterine Cervical Neoplasms; Wnt Signaling Pathway

This study aimed to investigate the role of lncRNA terminal differentiation-induced ncRNA (TINCR) in cervical squamous cell carcinoma (CSCC). By informatics analysis, we found that miR-302 may bind TINCR. Expression analysis showed that miR-302 was downregulated, while TINCR was upregulated in CSCC. Correlation analysis showed that they were not significantly correlated. In CSCC cells, miR-302 and TINCR failed to affect the expression of each other. However, miR-302 overexpression led to downregulated and TINCR overexpression led to upregulated cyclin D1 expression in CSCC cells. Interestingly, overexpression of cyclin D1 led to upregulated miR-302 and TINCR. Cell proliferation analysis showed that TINCR and cyclin D1 overexpression led to increased, while miR-302 overexpression led to decreased rate of cell proliferation. Moreover, miR-302 overexpression reduced the effects of TINCR overexpression. Therefore, TINCR sponges miR-302 to upregulate cyclin D1 in CSCC, thereby promoting cell proliferation.

Topics: Adult; Aged; Carcinoma, Squamous Cell; Cell Proliferation; Cyclin D1; Female; Gene Expression; Humans; Male; MicroRNAs; Middle Aged; Protein Binding; RNA, Long Noncoding; Up-Regulation; Uterine Cervical Neoplasms

Objective To investigate the role and mechanism of microRNA-182 (miR-182) in the proliferation of cervical cancer cells. Methods With liposome-mediated transient transfection method, the level of miR-182 in HeLa and SiHa cells was increased or decreased. CCK-8 assay and colony formation assay were used to observe the effect of miR-182 on the proliferation of cervical cancer cells. Using bioinformatics predictions, real-time quantitative PCR, and dual luciferase reporter assay, we clarified the role of miR-182 in posttranscriptional regulation of adenomatous polyposis coli (APC) gene and its effect on the downstream molecules (c-Myc and cyclin D1) of Wnt singling pathway. Results Up-regulation of miR-182 significantly promoted the proliferation of cervical cancer cells, while down-regulation of miR-182 significantly inhibited the proliferation of cervical cancer cells. Over-expression of miR-182 inhibited the expression of APC gene in cervical cancer cells and the regulation of miR-182 affected the expression of canonical Wnt signaling pathway downstream molecules in cervical cancer cells. Conclusion The miR-182 stimulates canonical Wnt signaling pathway by targeting APC gene and enhances the proliferation of cervical cancer cells.

Topics: Adenomatous Polyposis Coli Protein; beta Catenin; Cell Line, Tumor; Cell Proliferation; Cyclin D1; Female; Gene Expression Regulation, Neoplastic; Gene Targeting; HeLa Cells; Humans; MicroRNAs; Uterine Cervical Neoplasms; Wnt Signaling Pathway

High-mobility group AT-hook1 (HMGA1, formerly HMG-I/Y), an architectural transcription factor, participates in a number of tumor biological processes. However, its effect on cervical cancer remains largely indistinct. In this study, we found that HMGA1 was generally overexpressed in cervical cancer tissues and was positively correlated with lymph node metastasis and advanced clinical stage. Via exogenously increasing or decreasing the expression of HMGA1, we showed that HMGA1 affected the proliferation, colony formation, migration and invasion of cervical cancer cells in vitro. Rescue experiments suggested that miR-221/222 could partly reverse HMGA1-mediated migration and invasion processes. Mechanistically, we discovered that HMGA1 accelerated the G1/S phase transition by regulating the expression of cyclin D1 and cyclin E1, which was consistent with the results of the in vivo experiment. Furthermore, we found that HMGA1 regulated the expression of the miR-221/222 cluster at the transcriptional level and that miR-221/222 targeted the 3'UTR of tissue inhibitor of metalloproteinases 3(TIMP3). We propose a fresh perspective that HMGA1 participates in the migration and invasion process via the miR-221/222-TIMP3-MMP2/MMP9 axis in cervical cancer. In summary, our study identified a critical role played by HMGA1 in the progression of cervical cancer and the potential mechanisms by which exerts its effects, suggesting that targeting HMGA1-related pathways could be conducive to the therapies for cervical cancer.

Topics: Base Sequence; Cell Cycle; Cell Line, Tumor; Cell Movement; Cell Proliferation; Clone Cells; Cyclin D1; Cyclin E; Epithelium; Female; G1 Phase; Gene Expression Regulation, Neoplastic; HMGA1a Protein; Humans; Matrix Metalloproteinases; MicroRNAs; Middle Aged; Models, Biological; Neoplasm Invasiveness; S Phase; Tissue Inhibitor of Metalloproteinase-3; Tumor Stem Cell Assay; Uterine Cervical Neoplasms

We recently reported that TPD7 suppressed tumor cell proliferation, and inhibited invasion, through the suppression of C-X-C chemokine receptor type 4 (CXCR4). In the present study, we investigated the anticancer effect of TPD7 on apoptosis and invasion of cervical cancer HeLa cells. Cell cycle analysis revealed that TPD7 decreased cyclin-dependent kinase (CDK)1 and cyclin D1 expression, and increased cyclin A expression, following S phase blockade. TPD7 induced chromatin condensation and significantly elevated the number of apoptotic cells, suggesting that its inhibitory effect on HeLa cells was due to the induction of cell cycle blockade and apoptosis. Mechanistically, TPD7 altered the extrinsic apoptosis pathway by upregulating Fas expression, and the intrinsic pathway by modulating Bcl-2 family proteins, p53, and NF-κB p65, leading to enhanced apoptosis. TPD7 inhibited HeLa cell invasion by downregulating the expression of matrix metalloproteinase (MMP)-9 and CXCR4 proteins. In vivo experiments revealed that TPD7 inhibited tumor growth in HeLa cell xenografted mice. These findings indicated that TPD7 may be a potential chemoprevention agent for the management of cervical carcinoma.

Topics: Animals; Apoptosis; Biphenyl Compounds; Carbanilides; Cell Line, Tumor; Cell Proliferation; Cyclin D1; Down-Regulation; fas Receptor; Female; HeLa Cells; Humans; Hydroxylamines; Matrix Metalloproteinase 9; Mice; Mice, Inbred BALB C; NF-kappa B; Proto-Oncogene Proteins c-bcl-2; Receptors, CXCR4; S Phase; Signal Transduction; Up-Regulation; Urea; Uterine Cervical Neoplasms

Cervical cancer (CC) is the third most common cancer and the fourth leading cause of malignancy-related mortality in women worldwide. In addition, epithelial-mesenchymal transition (EMT) has been generally studied in tumor metastasis researches in recent years. Cysteine-rich intestinal protein 1 (CRIP1) is differently expressed in human cancer cells. However, the role it plays in CC has not been revealed at present. Preliminary experiments have shown that CRIP1 had a higher expression in CC tissues, compared with adjacent noncancerous tissues. Real-time PCR and western blot were performed to analyze CRIP1 expression in CC cell lines. CRIP1 transient transfection vector and siRNA were constructed. Further analysis revealed the promotion effects of CRIP1 on the cell migration and invasion of CC in vitro (P < 0.01). In addition, western blot was performed to show that CRIP1 mediates EMT by means of EMT marker detection. The expression of CRIP1 and β‑catenin in CC tissues was analyzed by immunohistochemistry (IHC). Interestingly, CRIP1 and β‑catenin were both highly expressed in CC tissues (P < 0.01). Furthermore, CRIP1 increased the protein expression level of c-myc, cyclinD-1 and cytoplasmic β‑catenin, which are indicators for activating the Wnt/β‑catenin signaling pathway. In conclusion, CRIP1 promotes cell migration and invasion, mediates EMT and activates the Wnt/β‑catenin signaling pathway in CC.

Topics: Adult; beta Catenin; Carrier Proteins; Cell Line, Tumor; Cell Movement; Cell Proliferation; Cyclin D1; Cytoplasm; Epithelial-Mesenchymal Transition; Female; Gene Expression Profiling; Gene Expression Regulation, Neoplastic; HeLa Cells; Humans; LIM Domain Proteins; Middle Aged; Neoplasm Invasiveness; Proto-Oncogene Proteins c-myc; Signal Transduction; Uterine Cervical Neoplasms; Wnt Proteins; Wnt Signaling Pathway

Circular RNAs (circRNAs) play a significant role in the development and progression of various human cancers. However, the expression and function of circRNAs in cervical cancer (CC) have rarely been explored. The aim of this study was to investigate the biological function of circRNA8924 in CC and elucidate the possible molecular mechanism involved.. Quantitative polymerase chain reaction was used to determine mRNA expression of circRNA8924, miR-518d-5p/519-5p and CBX8 in CC tissues and cells. CBX8 protein expression was measured by Western blotting. The CCK-8 assay was used to evaluate cell proliferation, and the transwell assay to determine cell migration and invasion. The luciferase reporter assay was used to determine the direct regulation of miR-518d-5p/519-5p and circRNA8924 or CBX8 Results: The study demonstrated that the expression level of circRNA8924 in CC was significantly higher than that in the adjacent normal tissues (P < 0.001), and that it was also associated with tumor size, FIGO staging and myometrial invasion. The knockdown of circRNA8924 significantly inhibited the proliferation, migration and invasion of CC cells SiHa and HeLa. The expression level of miR-518d-5p/519-5p was negatively correlated with circRNA8924, and circRNA8924 regulated CBX8 by competitively binding to miR-518d-5p/519-5p.. CircRNA8924 is highly expressed in CC tissue and can be considered a competitive endogenous RNA of the miR-518d-5p/519-5p family to promote the malignant biological behavior of CC cells. It is suggested that it may serve as a new biomarker for CC diagnosis and disease progression and provide potential targets for targeted therapy.

Topics: Antagomirs; bcl-2-Associated X Protein; Cell Movement; Cell Proliferation; Cyclin D1; Female; G1 Phase Cell Cycle Checkpoints; Gene Expression Regulation, Neoplastic; HeLa Cells; Humans; MicroRNAs; Polycomb Repressive Complex 1; RNA; RNA Interference; RNA, Circular; RNA, Small Interfering; Uterine Cervical Neoplasms

DPP8 is a member of the dipeptidyl peptidase IV family, which belongs to the S9b protease subfamily. It regulates cell proliferation, apoptosis, migration and invasion during cancer progression.. To investigate the role of DPP8 in cervical cancer, we examined DPP8 levels in cervical cancer tissues and cells. The localization of DPP8 was determined by immunofluorescence staining. Subsequently, SiHa and HeLa cells were treated with small interfering RNA (siRNA)-DPP8. We used cell cycle analysis, an 5-ethyl-2'-deoxyuridine assay proliferation assay and a cellular apoptosis assay to determine the effect of DPP8 on the proliferation and apoptosis of cervical cancer cells. We used a Transwell assay to assess the number of transfection cancer cells migrating through the matrix. A real-time polymerase chain reaction and western blot analysis were used to analyze the expression of related proteins and to determine the phenotype caused by the depletion or overexpression of DPP8 in cervical cancer cells.. We observed that DPP8 was highly expressed in cervical cancer tissues and cells. DPP8 expression was observed in the cytosol and in the perinuclear area, as well as in the nuclei of cervical cancer cells. Notably, when cells were treated with siRNA-DPP8, the expression of BAX increased, and the expression of cyclin D1, Bcl-2, MMP2 and MMP9 was downregulated. In cervical cancer cell lines, silencing the expression of DPP8 not only suppressed the proliferation, migration and invasion of the cervical cancer cells, but also promoted cervical cancer cell apoptosis.. The data obtained in the present study reveal that DPP8 promotes the progression of cervical cancer.

Topics: Apoptosis; Cell Line, Tumor; Cell Movement; Cell Proliferation; Cyclin D1; Dipeptidases; Female; Gene Expression Regulation, Neoplastic; HeLa Cells; Humans; Matrix Metalloproteinase 2; Matrix Metalloproteinase 9; Neoplasm Invasiveness; RNA Interference; Signal Transduction; Uterine Cervical Neoplasms

RAD51 is one of the pivotal enzymes for DNA double-strand break (DSB) repair by the homologous recombination (HR) pathway, which implies it as a promising and novel target for cancer therapy. Recent findings have indicated RAD51 protein is overexpressed in a variety of tumors. The high-expression of RAD51 is related to poor prognosis. RAD51 is involved in the repair of DNA damage and the generation of genetic diversity by an evolutionarily conserved mechanism. However, the exact mechanism of Rad51 in the progression of cervical cancer remains unclear. RI-1 is a small molecule that inhibits the central recombination protein RAD51. In this study, we found that RAD51 was highly expressed in invasive squamous cervical cancer (SCC). The administration of RI-1 inhibited cell growth in vitro and reduced growth of tumor xenografts in vivo with cervical cancer cells (HeLa and SiHa). Further investigation suggested that RAD51 protein significantly promoted the cell cycle transition from the G0/G1 to S phase. In addition, the inhibition of RAD51 reduced the level of the cell cycle related protein cyclin D1, but increased the levels of p21 mRNA and protein. As a DNA DSB repair enzyme, the expression of RAD51 in tumor cells possibly affects their sensitivity to anti-cancer agents. Additionally, in experiments using cisplatin and ionizing radiation, RI-1 treated cervical cancer cells, HeLa and SiHa, were sensitized to a greater extent than the untreated control. Thus, HR inhibition of RAD51 may provide yet another mechanism of therapeutic target for the chemosensitization and radiosensitization of cervical cancer with RI-1. Collectively, our data demonstrated for the first time that inhibition of RAD51 suppressed the cervical cancer cell proliferation and the growth of cervical cancer xenografts by attenuating cell cycle transition, which could be a functional link between RAD51 and cyclin D1 and p21.

Topics: Cell Proliferation; Cyclin D1; Cyclin-Dependent Kinase Inhibitor p21; DNA Breaks, Double-Stranded; DNA Damage; DNA Repair; Female; HeLa Cells; Homologous Recombination; Humans; Molecular Targeted Therapy; Morpholines; Pyrroles; Rad51 Recombinase; Uterine Cervical Neoplasms

To review the clinicopathological features of 14 cases of CIN 3-like squamous cell carcinoma (SCC) of the cervix and to investigate possible mechanisms of tumour invasion in the CIN 3-like and 'conventional' (infiltrative) tumour components.. The median patient age was 43.4 years. Of the 12 cases with known stage, eight were stage IB and four were stage II. Initial biopsies were often misinterpreted as CIN 3 alone or CIN 3 with only superficial stromal invasion, and eight patients underwent multiple biopsy procedures prior to definitive diagnosis: upon review, an earlier diagnosis was possible in six of these cases. Nine tumours exhibited both CIN 3-like and conventional tumour components. The former usually demonstrated an E-cadherin-positive and cyclin D1-negative immunophenotype, whereas conventional SCC more often showed loss of E-cadherin and cyclin D1 staining at the margin of larger tumour nests and in smaller invasive clusters.. Cervical SCCs demonstrating a CIN 3-like growth pattern continue to present diagnostic difficulty and are often misinterpreted as non-invasive/minimally invasive disease. The morphology and immunophenotype suggest a process of collective cellular invasion in CIN 3-like SCC, whereas corresponding conventional SCC elements show features consistent with epithelial-mesenchymal transition.

Topics: Adult; Aged; Antigens, CD; Biomarkers, Tumor; Cadherins; Carcinoma, Squamous Cell; Cyclin D1; Female; Humans; Immunohistochemistry; Middle Aged; Uterine Cervical Dysplasia; Uterine Cervical Neoplasms

Cervical cancer is one of the most frequent gynecological malignancies in women worldwide. MicroRNA-195 (miR-195) was recently found highly expressed in cervical cancer. However, the role of miR-195 in the pathology of cervical cancer remains poorly understood. In this study, we first confirmed the downregulation of miR-195 in primary cervical cancer tissues. For the functional study, we introduced the sequences of miR-195 or miR-195 inhibitor into Hela and SiHa cervical cancer cell lines. Overexpression of miR-195 inhibited the proliferation of both Hela and SiHa cells. In contrast, reducing the endogenous miR-195 level by miR-195 inhibitor promoted the proliferation of cervical cancer cells. Flow cytometric assay showed that overexpression of miR-195 induced G1 phase arrest, whereas miR-195 inhibitor shortened G1 phase of cervical cancer cells. In addition, the suppressive role of miR-195 in cell cycle was also demonstrated by the western blot results of various cell cycle indicators, such as phosphorylated retinoblastoma (p-Rb) and proliferating cell nuclear antigen (PCNA), in the gain and loss of function experiments. Furthermore, Dual-Luciferase Reporter Assay revealed that miR-195 targeted the 3'-untranslated region of cyclin D1a transcript, such as to regulate cyclin D1 expression. In summary, our results suggest that miR-195 acts as a suppressor in the proliferation and cell cycle of cervical cancer cells by directly targeting cyclin D1a mRNA.

Topics: 3' Untranslated Regions; Base Sequence; Binding Sites; Cell Proliferation; Cyclin D1; Female; Gene Expression; Gene Expression Regulation, Neoplastic; HeLa Cells; Humans; MicroRNAs; RNA Interference; Uterine Cervical Neoplasms

MicroRNAs are important regulators of multiple cellular processes, and aberrant miRNA expression has been observed in human cervical cancer (CC). The present study was to evaluate the level of miR-195 and cyclin D1 in CC tissues and cells. We further investigated the molecular mechanisms of miR-195 and cyclin D1 in CC cell lines HeLa and SiHa. Here, we found that miR-195 expression was down-regulated in CC tissues, and HeLa and SiHa cells (all p < 0.001). By contrast, cyclin D1 was up-regulated. Furthermore, the expression of miR-195 was inversely proportional to that of cyclin D1 mRNA or protein (p = 0.013, p = 0.015, respectively). In vitro studies demonstrated that the overexpression of miR-195 played a suppressor role in the proliferation of HeLa and SiHa cells and promoted cell apoptosis. Luciferase reporter assays confirmed that miR-195 binding to the 3'-UTR regions of cyclin D1 inhibited the expression of cyclin D1 in HeLa and SiHa cells. However, the inhibitor of miR-195 promoted the expression of cyclin D1 and cell proliferation. In conclusion, our data suggest that miR-195 may have the potential role in treatment of CC patients, as well as miR-195 is a novel regulator of invasiveness and tumorigenicity in CC cells by targeting cyclin D1. MiR-195/cyclin D1 pathway may be a useful therapeutic agent in CC patients.

Topics: 3' Untranslated Regions; Adult; Aged; Apoptosis; Cell Line, Tumor; Cell Proliferation; Cyclin D1; Female; Gene Expression Regulation, Neoplastic; Humans; MicroRNAs; Middle Aged; RNA Interference; Uterine Cervical Neoplasms; Young Adult

Persistent infection with oncogenic human papillomavirus viruses (HPVs) is a casual factor for cervical cancer and its precursors, and the abnormal constitutive expression of viral oncoprotein E6 is a key event during the malignant transformation. Here, we performed miRNA microarray to identify changes of miRNAs following ectopic HPV16 E6 overexpression in HEK293T cells and found miR-2861 was greatly decreased in both HEK293T and HaCaT cells expressing HPV16 E6 compared to vector control. Further, we demonstrated a biological link among HPV16 E6, miR-2861, EGFR, AKT2, and CCND1 in cervical cancer cells. We showed that miR-2861 was downregulated in cervical cancer tissues and negatively correlated with advanced tumor stage and lymph node metastasis. Overexpression of miR-2861 suppressed cervical cancer cell proliferation and invasion and enhanced apoptosis. Subsequent investigation revealed that EGFR, AKT2, and CCND1 were all the direct targets of miR-2861. Importantly, silencing EGFR, AKT2, and/or CCND1 recapitulated the cellular effects seen upon miR-2861 overexpression. Restoration of EGFR, AKT2, and/or CCND1 counteracted the effects of miR-2861 expression. Thus, we identified a new pathway employing miR-2861, EGFR, AKT2, and CCND1 that may mediate HPV16 E6 induced initiation and progression of cervical cancer.

Topics: Cell Line, Tumor; Cell Transformation, Viral; Cyclin D1; ErbB Receptors; Female; Gene Expression Regulation, Neoplastic; HEK293 Cells; Human papillomavirus 16; Humans; MicroRNAs; Neoplasm Metastasis; Neoplasm Staging; Oligonucleotide Array Sequence Analysis; Oncogene Proteins, Viral; Papillomavirus Infections; Proto-Oncogene Proteins c-akt; Repressor Proteins; Signal Transduction; Uterine Cervical Neoplasms

The human cervical cancer (CC) acts as the most common one of women tumors. However, the pathological changes and molecular alterations of CC are not clear. It has been reported that miR-202 takes part in the development and progression of different tumors. The present study aims to detect the expression of miR-202 in 100 cases of CC tissues and cells, and then we continued to investigate the potential mechanisms of miR-202 in CC cells. In this work, we found that the expression of miR-202 is obviously decreased in both CC cell lines and tissues, and negatively related with the expression of cyclin D1 in SiHa, HeLa and Caski cells. In-vitro assay revealed that the ectopic expression of miR-202 suppressed the proliferation, migration and invasion of SiHa and HeLa cells. Additionally, the over-expression of miR-202 extremely affected the expression of cyclin D1 protein. Notably, the over-expression of cyclin D1 in SiHa and HeLa cells with miR-202 mimics attenuated the inhibitory effects of miR-202 on cell proliferation, migration and invasion. In conclusion, our study identified that miR-202 plays an important role in regulating cell proliferation, migration and invasion of CC by directly targeting cyclin D1, thus miR-202 may represent a potential therapeutic target for patients with cervical cancer.

Topics: Cell Movement; Cell Proliferation; Cervix Uteri; Cyclin D1; Disease Progression; Down-Regulation; Female; Gene Expression Regulation, Neoplastic; HeLa Cells; Humans; Keratinocytes; MicroRNAs; Neoplasm Grading; Neoplasm Invasiveness; Neoplasm Staging; Real-Time Polymerase Chain Reaction; Uterine Cervical Neoplasms

To explore the effects of silencing HERC4 on the proliferation, apoptosis, and migration of cervical cancer cell line Hela and the possible molecular mechanisms.. Three HERC4-specific small interfering RNAs (siRNAs) were transfected into Hela cells, and HERC4 expression in the cells was examined with Western blotting. CCK-8 assay, annexin V-FITC/PI assay, and wound healing assay were used to assess the effect of HERC4 silencing on the proliferation, apoptosis and migration ability of Hela cells. The expression levels of cyclin D1 and Bcl-2 in the cells were detected using Western blotting.. Transfection of siRNA-3 resulted in significantly decreased HERC4 protein expression (P<0.01). HERC4 silencing by siRNA-3 markedly suppressed the proliferation and migration of Hela cells, increased the apoptosis rate (P<0.01) and reduced the expression levels of cyclin D1 and Bcl-2 (P<0.01).. Silencing of HERC4 efficiently inhibits the proliferation, migration, and invasion of Hela cells in vitro, and the underlying mechanisms may involve the down-regulation of cyclin D1 and Bcl-2.

Topics: Apoptosis; Cell Line, Tumor; Cell Movement; Cell Proliferation; Cyclin D1; Down-Regulation; Female; HeLa Cells; Humans; Proto-Oncogene Proteins c-bcl-2; RNA Interference; RNA, Small Interfering; Transfection; Ubiquitin-Protein Ligases; Uterine Cervical Neoplasms

In this study, we found that the total polyphenol and ascorbic acid levels in the fruit of Opuntia humifusa are higher than those in other parts of the plant. We further hypothesized that antioxidants in O. humifusa might affect the growth or survival of cancer cells. Hexane extracts of seeds and ethyl acetate extracts of fruits and stems significantly suppressed the proliferation of HeLa cervical carcinoma cells, but did not affect the proliferation of normal human BJ fibroblasts. Additionally, the extracts of O. humifusa induced G1 phase arrest in HeLa cells. The O. humifusa extracts reduced the levels of G1 phase-associated cyclin D1, cyclin-dependent kinase 4 (Cdk4), and phosphorylated retinoblastoma proteins. Moreover, p21(WAF1/Cip1) and p53 expression significantly increased after treatment. We examined the effects of ethyl acetate extracts of O. humifusa fruit (OHF) on HeLa cells xenograft tumor growth. OHF treatment significantly reduced tumor volume and this decrease was correlated with decreased Cdk4 and cyclin D1 expression. Furthermore, flavonoids, trans Taxifolin, and dihydrokaempferol, were isolated from OHF. Thus, this extract may be a promising candidate for treating human cervical carcinoma.

Topics: Animals; Antineoplastic Agents, Phytogenic; Antioxidants; Ascorbic Acid; Cell Cycle Checkpoints; Cyclin D1; Cyclin-Dependent Kinase 4; Female; Fibroblasts; Fruit; HeLa Cells; Heterografts; Humans; Mice, Inbred BALB C; Opuntia; Phytotherapy; Plant Extracts; Polyphenols; Retinoblastoma Protein; Uterine Cervical Neoplasms

Resveratrol inhibits cervical cancer (CC) cells by blocking STAT3 signaling. However, the mechanism of resveratrol-induced STAT3 inactivation remains largely unknown. SHP2, PIAS3, and SOCS3 are STAT3 negative regulators; therefore, their statuses in cervical adenocarcinoma (HeLa) and squamous cell carcinoma (SiHa and C33A) cell lines without and with resveratrol treatment and their correlation with STAT3 activation in CC specimens were investigated.. MTT and TUNEL assays were used to check the resveratrol sensitivity of CC cells, and immunocytochemical staining, Western blotting, and RT-PCR were used to analyze SHP2, PIAS3, and SOCS3 expression and the intracellular distribution of STAT3. Tissue microarray based immunohistochemical staining was performed to investigate potential correlations between SHP2, PIAS3, and SOCS3 expression and STAT3 activation.. PIAS3 and SOCS3 were found to be weakly expressed in CC cells and upregulated by resveratrol; this was accompanied by inhibition of STAT3 signaling. The SHP2 level remained unchanged in all three cell lines after resveratrol treatment. STAT3 nuclear translocation was more frequent in adenocarcinomas and squamous cell carcinomas than that of their noncancerous counterparts. The SOCS3 level and detection rate were higher in noncancerous squamous cells (but not in glandular epithelia) compared with their cancerous counterparts. The phospho-SHP2 detection rate was similar in noncancerous and tumor tissues of squamous and glandular origins; however, PIAS3 levels were distinct.. Of the three STAT3 negative regulators, PIAS3 correlated most negatively with STAT3 nuclear translocation and may inhibit STAT3 signaling in both histological CC subtypes. PIAS3 responsiveness may reflect greater resveratrol sensitivity and improved therapeutic outcome in CCs.

Topics: Adenocarcinoma; Antineoplastic Agents, Phytogenic; Carcinoma, Squamous Cell; Cell Proliferation; Cell Survival; Cyclin D1; Female; Gene Expression; HeLa Cells; Humans; Inhibitor of Apoptosis Proteins; Molecular Chaperones; Phosphorylation; Protein Inhibitors of Activated STAT; Protein Tyrosine Phosphatase, Non-Receptor Type 11; Proto-Oncogene Proteins c-myc; Resveratrol; RNA, Neoplasm; Signal Transduction; STAT3 Transcription Factor; Stilbenes; Suppressor of Cytokine Signaling 3 Protein; Suppressor of Cytokine Signaling Proteins; Survivin; Uterine Cervical Neoplasms; Vascular Endothelial Growth Factor A

To investigate the correlation of cyclin D1 (CCND1) G870A single nucleotide polymorphism (SNP) with radiotherapy response in patients with high risk human papillomavirus (HR-HPV) related cervical cancer.
. A total of 273 patients with cervical cancer, who were confirmed by histopathology and hybrid capture 2 (HC-2) assay and treated by radiotherapy, were enrolled for this study. The correlation of CCND1 G870A polymorphism with tumor response in patients was assessed.
. Compared with patients with AA genotype, the patients with GG genotype and AA genotype showed lower sensitivity to radio-therapy treatment (adjusted ORGA=2.69, 95% CI 1.28-5.67 and adjusted ORGG=3.28, 95% CI 1.47-7.29, respectively), an increase in risks of recurrence/metastasis (adjusted ORGA=2.52, 95% CI 1.12-5.63 and adjusted ORGG=3.95, 95% CI 1.68-9.26, respectively), and shorter recurrence/metastasis-free survival (PGA=0.010 and PGG=0.045).
. G870A polymorphism is a frequent variation that could be used for evaluate the radio-sensitivity and prognosis for patients with HR-HPV related cervical cancer.. 目的：探讨细胞周期蛋白D1(cyclin D1，CCND1)基因G870A位点单核苷酸多态性(single nucleotide polymorphism，SNP)与高危型人乳头状瘤病毒高危型(high risk human papillomavirus，HR-HPV)相关宫颈癌患者放疗敏感性的关系。方法：选择经组织病理学和第二代杂交捕获法(hybird capture II，HC-2)确诊的HR-HPV相关宫颈癌患者273例；限制性片段长度多态性-聚合酶链反应(restriction fragment length polymorphism-polymerase chain reaction，PCR-RFLP)技术检测CCND1基因G870A位点的基因型频率分布，分析其与宫颈癌放疗敏感性的关系。结果：与AA基因型相比，携带GA基因型和GG基因型患者发生局部放疗失败(ORGA=2.74，95% CI 1.37~5.46；ORGG=3.01，95% CI 1.42~6.39)和复发转移(ORGA=2.52，95% CI 1.12~5.63；ORGG=3.95，95% CI 1.68~9.26)的风险明显增加，且无复发转移生存期明显缩短(PGA=0.010，PGG=0.045)。结论：CCND1基因G870A位点多态性与HR-HPV相关宫颈癌患者放疗敏感性及复发或转移风险相关，可能是宫颈癌放疗疗效及复发或转移的预测指标。.

Topics: Cyclin D1; Female; Genotype; Humans; Papillomaviridae; Polymorphism, Single Nucleotide; Prognosis; Uterine Cervical Neoplasms

Dysregulation of microRNA-150 (miR-150) is commonly observed in solid tumor and has been reported to be involved in multiple important biological processes, such as cell proliferation, apoptosis, and metastasis. Elevated miR-150 level was also detected in cervical carcinoma, whereas its function in cancer progression has not been studied yet.. The expression of miRNA-150 in cervical carcinoma was compared with normal cervical tissue and using qRT-PCR. The effects of miR-150 on cell cycle and apoptosis, as well as the expression of cycle- and apoptosis-related genes, were determined using flow cytometry, TUNEL assay, qRT-PCR, and Western blot, respectively. The direct target of miR-150 was confirmed using 3' untranslated region (UTR) luciferase reporter assay.. miR-150 promotes cervical cancer cell survival and growth, while the inhibition of miR-150 suppresses these actions. miR-150 also induced the cell cycle progression from G1/G0 to S phase, resulting in an enhancement of growth. Several cell cycle- and apoptosis-related genes, CyclinD1, p27, BIM, and FASL were modulated by miR-150. Moreover, miR-150 directly reduced the expression of FOXO4, which regulates the expression of CyclinD1, p27, BIM, and FASL, by targeting its 3' UTR.. Taken together, our data demonstrated that elevated miR-150 targets FOXO4 expression and therefore regulates multiple genes expression, resulting in cervical cancer cell growth and survival.

Topics: Apoptosis; Apoptosis Regulatory Proteins; Bcl-2-Like Protein 11; Cell Cycle; Cell Cycle Proteins; Cell Line, Tumor; Cell Proliferation; Cell Survival; Cyclin D1; Cyclin-Dependent Kinase Inhibitor p27; Fas Ligand Protein; Female; Forkhead Transcription Factors; Gene Expression Regulation, Neoplastic; HEK293 Cells; Humans; In Situ Nick-End Labeling; Membrane Proteins; MicroRNAs; Neoplasm Metastasis; Proto-Oncogene Proteins; Transcription Factors; Uterine Cervical Neoplasms

To investigate the expression of cyclin D1 in cervical intraepithelial neoplasia (CIN) and squamous cell carcinoma and its relationship with human papillomavirus 16 (HPV16) E7 gene expression.. Both SiHa and Hcc94 cell lines were obtained from cervical epithelial cells of squamous cell carcinoma. E6/E7 gene was silent in Hcc94 cell line.Expression levels of cyclin D1 mRNA and protein in CIN and squamous cell carcinoma were detected by QT-PCR and immunohistochemistry (IHC) respectively. SiRNA was constructed for targeting the promoter of HPV16 E7 and then transfected into SiHa cells to establish cm-16 line with stable silencing of E7. Control cell line B3 was obtained by blank plasmid transfection into SiHa cells. RT-PCR and Western blot were used to detect cyclin D1 mRNA and protein expression in the SiHa, B3, and cm-16 cells, respectively.. Cyclin D1 was expressed in the basal cells of normal cervical squamous epithelia and the expression gradually decreased in the progression from CIN1 to CIN3. Squamous cell carcinoma showed negative or scattered expression of cyclin D1 (P<0.05). Both mRNA and protein of cyclin D1 in E7(+) SiHa cells were lower than those in cm-16 and Hcc94 cells.. Squamous cell carcinoma with high HPV E7 expression shows low level of cyclin D1, suggesting that HPV16 E7 gene inhibits the expression of cyclin D1.

Topics: Carcinoma, Squamous Cell; Cell Line, Tumor; Cyclin D1; Female; Human papillomavirus 16; Humans; Immunohistochemistry; Papillomavirus E7 Proteins; Promoter Regions, Genetic; RNA Interference; RNA, Messenger; RNA, Small Interfering; Transfection; Uterine Cervical Dysplasia; Uterine Cervical Neoplasms

The phosphoinositide 3-kinase (PI3K)/Akt signalling pathway appears to be a key regulator in cervical carcinogenesis. However, the downstream regulatory mechanism of PI3K/Akt signalling remains largely unknown.. The expression of miR-196a in cervical cancer cell lines and cervical cancer tissues was examined using real-time PCR. The effects of miR-196a on PI3K/Akt signalling and cellular proliferation were evaluated by bromodeoxyuridine labelling, 3-(4,5-Dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazoliumbromide, colony formation assays and luciferase assays.. The expression level of miR-196a was markedly increased in cervical cancer tissues and cell lines compared with normal cervical tissue and normal cervical squamous cells. Upregulation of miR-196a was correlated with advanced tumour stage and poor overall and recurrence-free survival in cervical cancer patients. Upregulation of miR-196a enhanced G1/S-phase transition and the proliferative ability of cervical cancer cells, whereas suppression of miR-196a had the opposite effect. Using bioinformatics and biological approaches, we showed that FOXO1 and p27(Kip1), two key effectors of PI3K/Akt signalling, were direct targets of miR-196a.. Our findings suggest that miR-196a has an important role in promoting human cervical cancer cell proliferation and may represent a novel therapeutic target of microRNA-mediated suppression of cell proliferation in cervical cancer.

Topics: Cell Growth Processes; Cell Line, Tumor; Cyclin D1; Cyclin-Dependent Kinase Inhibitor p27; Female; Forkhead Box Protein O1; Forkhead Transcription Factors; G1 Phase; HeLa Cells; Humans; MicroRNAs; Phosphatidylinositol 3-Kinases; Proto-Oncogene Proteins c-akt; S Phase; Up-Regulation; Uterine Cervical Neoplasms

Epigenetic regulators like histone deacetylases (1 and 2), and viral onco-proteins (E6/E7) are known to be overexpressed in cervical cancer cells. The present study was designed to investigate the effect of curcumin on HDACs (1 and 2) and HPV E6/E7 in the cervical cancer cell line SiHa and a drug resistant clone SiHaR (derived from SiHa). It was further intended to investigate whether curcumin could sensitize the cells towards cisplatin induced cell killing by modulation of multi drug resistant proteins like MRP1 and Pgp1. Curcumin inhibited HDACs, HPV expression and differentially increased acetylation and up-regulation of p53 in SiHa and SiHaR, leading to cell cycle arrest at G1-S phase. Up-regulation of pRb, p21, p27 and corresponding inhibition of cyclin D1 and CDK4 were observed. Cisplatin resistance in SiHaR due to over-expression of MRP1 and Pgp1 was overcome by curcumin. Curcumin also sensitized both the cervical cancer cells towards cisplatin induced cell killing. Inhibition of HDACs and HPVs led to cell cycle arrest at G1/S phase by alteration of cell cycle regulatory proteins. Suppression of MRP1 and Pgp1 by curcumin resulted in sensitization of cervical cancer cells, lowering the chemotherapeutic dose of the drug cisplatin.

Topics: Acetylation; Antineoplastic Agents; Cell Cycle Proteins; Cell Line, Tumor; Cisplatin; Curcumin; Cyclin D1; Cyclin-Dependent Kinase 4; Cyclin-Dependent Kinase Inhibitor p21; Cyclin-Dependent Kinase Inhibitor p27; Drug Resistance, Neoplasm; Enzyme Inhibitors; Female; Glutathione; Histone Deacetylase 1; Histone Deacetylase 2; Humans; Hyaluronan Receptors; Multidrug Resistance-Associated Proteins; Oncogene Proteins, Viral; Papillomavirus E7 Proteins; Repressor Proteins; Retinoblastoma Protein; S Phase Cell Cycle Checkpoints; Uterine Cervical Neoplasms

Interruption of the cell cycle is accompanied by changes in several related molecules that result in the activation of apoptosis. The present study was performed to verify the apoptotic effects of sequential treatment with bortezomib and celecoxib in TC-1 cells expressing the human papillomavirus (HPV) E6 and E7 proteins. In TC-1 cells sequentially treated with bortezomib and celecoxib, apoptosis was induced through decreased expression of signal transducer and activator of transcription-3 (STAT3), cyclin D1 and cyclin-dependent kinase (CDK) 2, which are major regulators of the G0/G1 cell cycle checkpoint. In addition, increased levels of p21, CHOP, BiP and p-p38 MAPK were identified in these cells. The treatment-induced apoptosis was effectively inhibited by treatment with SB203580, an inhibitor of p-p38. Moreover, the growth of tumors sequentially treated with bortezomib and celecoxib was retarded compared to the growth of tumors exposed to a single treatment with either bortezomib or celecoxib in vivo. We demonstrated that sequential treatment with bortezomib and celecoxib induced apoptosis via p-p38-mediated G0/G1 cell cycle arrest and endoplasmic reticulum (ER) stress. Sequential treatment with these two drugs could therefore be a useful therapy for cervical cancer.

Topics: Animals; Antineoplastic Agents; Antineoplastic Combined Chemotherapy Protocols; Apoptosis; Boronic Acids; Bortezomib; Celecoxib; Cell Line, Tumor; Cell Proliferation; Cyclin D1; Cyclin-Dependent Kinase 2; Cyclin-Dependent Kinase Inhibitor p21; Cyclooxygenase 2 Inhibitors; Down-Regulation; Endoplasmic Reticulum Stress; Female; G1 Phase Cell Cycle Checkpoints; Imidazoles; Mice; Mice, Inbred C57BL; Oncogene Proteins, Viral; p38 Mitogen-Activated Protein Kinases; Papillomavirus E7 Proteins; Pyrazines; Pyrazoles; Pyridines; Repressor Proteins; STAT3 Transcription Factor; Sulfonamides; Transcription Factor CHOP; Uterine Cervical Neoplasms

The purpose of the present study is to evaluate the potent growth inhibitory effects of aqueous wheatgrass extract (AWE) alone and in combination with cisplatin on human breast and cervical cancer cells.. The cytotoxic potential of AWE alone and in combination with cisplatin was evaluated on human breast and cervical cancer cells (MCF-7 and HeLa) by cell viability assay. Further, the mode of cell death induced by AWE was determined by nuclear morphological examination and cell cycle analysis. These effects were then correlated with the expression of genes involved in apoptosis and proliferation (cyclin D1 and Bax) by RT-PCR.. AWE showed dose- and time dependent selective cytotoxicity towards the cancer highlighting its safe profile. Lower dose combinations of AWE and cisplatin induced increased growth inhibition compared with the individual drugs on both cell lines (combination index < 1) indicating strong synergistic interactions. AWE was found to induce apoptosis and arrested the cells at G0-G1 phase of the cell cycle which correlated with the modulation of expression of bax and cyclin D1 in a time-dependent manner in MCF-7 and HeLa cells.. These results suggest that the anti-cancer potential of AWE may be due to apoptosis induction and its anti-proliferative properties. This study also provides the first evidence demonstrating synergism between AWE and cisplatin, which may enhance the therapeutic index of prevention and/or treatment of human breast and cervical cancer.

Topics: Adjuvants, Immunologic; Antineoplastic Combined Chemotherapy Protocols; Apoptosis; bcl-2-Associated X Protein; Breast Neoplasms; Cell Proliferation; Cisplatin; Cyclin D1; Female; Gene Expression Regulation, Neoplastic; HeLa Cells; Humans; MCF-7 Cells; Plant Extracts; Triticum; Uterine Cervical Neoplasms

In response to tumor development, cells initially undergo invasion and metastasis followed by epithelial-mesenchymal transition (EMT, a process by which cells acquire motility) and overriding senescence (an endogenous defense mechanism against tumor progression). Oncogenic activation of Twist1 and Twist2 is essential for EMT and senescence; however, little is known about the specific contributions of Twist1 versus Twist2 to prognosis, metastasis, and the mechanism underlying cervical carcinoma. Here, we investigated the similarities and differences between Twist1 and Twist2 in assessing prognosis and promoting invasion and metastasis of cervical carcinoma as well as their roles in the underlying molecular mechanisms. By monitoring the survival of 144 clinical cervical cancer patients, we demonstrated that Twist2 shows more effective predictive performance compared with Twist1 and is more closely correlated with International Federation of Gynecology and Obstetrics stage and lymph node metastasis. Compared with Twist1, Twist2 more strongly promotes invasivity and motility by inducing EMT and overriding senescence. Differences between Twist1 and Twist2 in regulating senescence and the cell cycle might be due to their individual roles in regulating the cyclin D1/cyclin dependent kinase 4 (Cdk4) pathway. Overall, our data indicate that Twist2 is the key Twist isoform coupling aberrant signals from EMT to senescence and is an important candidate biomarker for cervical cancer prognosis.

Topics: Adenocarcinoma; Adult; Biomarkers; Carcinoma, Squamous Cell; Cell Cycle; Cell Movement; Cellular Senescence; Cyclin D1; Cyclin-Dependent Kinase 4; Epithelial-Mesenchymal Transition; Female; Gene Expression Regulation, Neoplastic; HeLa Cells; Humans; Neoplasm Invasiveness; Nuclear Proteins; Prognosis; Proportional Hazards Models; Protein Isoforms; Repressor Proteins; ROC Curve; Twist-Related Protein 1; Uterine Cervical Neoplasms

Cyclin D1b is one of two proteins translated from cyclin D1 transcripts (isoforms a and b) that are generated due to gene polymorphism. Our previous study has reported low cyclin D1b expression in cervical cancer tissue, with an expression level in moderately or poorly differentiated tissues that was significantly lower than that in well-differentiated tissues. However, the functional role of cyclin D1b in cervical cancer remains to be elucidated. In this study, using a cervical cancer cell line with stable expression of cyclin D1b, we found that upregulation of cyclin D1b initiated cell cycle arrest at the G0/G1 phase and induced apoptosis, thereby inhibiting cell proliferation and colony formation. Furthermore, xenograft transplantation experiments in nude mice demonstrated that cyclin D1b upregulation inhibited cancer growth and induce apoptosis in vivo. In conclusion, the present study indicates anti-tumor effects of cyclin D1b in cervical cancer, suggesting that cyclin D1b may represent a potential therapeutic target for cervical cancer.

Topics: Animals; Apoptosis; Blotting, Western; Cell Proliferation; Cyclin D1; Female; Flow Cytometry; HeLa Cells; Heterografts; Humans; In Situ Nick-End Labeling; In Vitro Techniques; Mice; Mice, Nude; Protein Isoforms; Real-Time Polymerase Chain Reaction; Reverse Transcriptase Polymerase Chain Reaction; Uterine Cervical Neoplasms

Leucine-rich repeat-containing G protein-coupled receptor 5 (LGR5), a seven transmembrane receptor known as a potential stem cell marker for intestinal crypts and hair follicles, has recently been found to be overexpressed in some types of human cancers. However, the role of LGR5 in cervical cancer remains unclear. In this study, the expression of LGR5 gradually increases from normal cervix to cervical cancer in situ and to cervical cancers as revealed by immunohistochemistry and western blot analyses. Through knocking down or overexpressing LGR5 in SiHa and HeLa cells, the expression level of LGR5 was found to be positively related to cell proliferation in vitro and to tumor formation in vivo. Further investigation indicated that LGR5 protein could significantly promote the acceleration of cell cycle. Moreover, the TOP-Flash reporter assay and western blot for β-catenin, cyclinD1, and c-myc proteins, target genes of the Wnt/β-catenin pathway, indicated that LGR5 significantly activated Wnt/β-catenin signaling. Additionally, the blockage of Wnt/β-catenin pathway resulted in a significant inhibition of cell proliferation induced by LGR5. Taken together, these results demonstrate that LGR5 can promote proliferation and tumor formation in cervical cancer cells by activating the Wnt/β-catenin pathway.

Topics: Animals; beta Catenin; Cell Cycle; Cell Line, Tumor; Cell Proliferation; Cell Survival; Cell Transformation, Neoplastic; Cervix Uteri; Cyclin D1; Female; HeLa Cells; Humans; Intercellular Signaling Peptides and Proteins; Mice; Mice, Inbred BALB C; Neoplasm Transplantation; Proto-Oncogene Proteins c-myc; Receptors, G-Protein-Coupled; Transplantation, Heterologous; Uterine Cervical Neoplasms; Wnt Proteins; Wnt Signaling Pathway

We previously reported frequent loss of microRNA‑218 (miR‑218) in human cervical cancer, which was associated with tumor progression and poor prognosis. In this study, we investigated whether restoration of the miR‑218 level is a valid strategy for the treatment of cervical cancer. The expression of miR‑218 in cervical cancer samples and cell lines was quantified by reverse transcription TaqMan quantitative (RT‑q)PCR. Overexpression of miR‑218 was achieved by both transient and stable transfection, using a miR‑218 mimic and a miR‑218‑expressing plasmid, respectively. Alterations in cellular proliferation and cell‑cycle progression were measured by the MTT assay and flow cytometry analysis. Nude mice bearing SiHa xenografts were used to investigate the functions of miR‑218 and carboplatin on tumor growth and weight. The expression of cycle‑related proteins was detected by western blotting and immunohistochemical staining. In vitro, miR‑218 significantly inhibited cellular growth in all four cell lines tested (P=0.021 for CaSki, P=0.009 for HeLa, P=0.016 for SiHa, and P=0.029 for C33A). Overexpression of miR‑218 induced G1 phase arrest and reduced expression of cyclin D1 and CDK4. In vivo, restoration of miR‑218 notably inhibited tumor growth and decreased tumor weight. In primary cultured samples, tumors with high levels of miR‑218 were more sensitive to carboplatin (R2=0.3319, P=0.0026); consistently, miR‑218 overexpression suppressed tumor growth, induced cell‑cycle arrest, and reduced the cyclin D1 level. Based on these and previous results, we conclude that restoration of the miR‑218 level inhibits the growth of cervical cancer cells both in vitro and in vivo; furthermore, overexpression of miR‑218 sensitizes cervical cancer cells to carboplatin. Our findings suggest a novel therapy for cervical cancer based on miR‑218, especially in patients with reduced levels of miR‑218.

Topics: Animals; Cell Line, Tumor; Cell Proliferation; Cyclin D1; Cyclin-Dependent Kinase 4; Female; G1 Phase Cell Cycle Checkpoints; HeLa Cells; Humans; Mice; Mice, Inbred BALB C; Mice, Nude; MicroRNAs; Transfection; Uterine Cervical Neoplasms

To investigate the expression of metastasis-associated in colon cancer-1 (MACC-1) mediated by siRNA, and to study the effects of its downregulation on cell proliferation, cell cycle and invasion ability of cervical cancer SiHa cells.. MACC-1 siRNA and control siRNA were transfected into cervical cancer SiHa cells, and the expression of MACC-1 protein after transfection with MACC-1 siRNA was detected by Western blotting. The changes of cell proliferation, cell cycle and invasion ability of the SiHa cells were determined by CCK-8 kit, flow cytometry and Boyden chamber assay. The expressions of cell cycle- and invasion-related proteins were analyzed by Western blotting.. Compared with the untreated group (0.317 ± 0.023) and control siRNA group (0.309 ± 0.021), the expression of MACC-1 protein was downregulated in the MACC1 siRNA group (0.041 ± 0.006) (P < 0.05), and its downregulation significantly suppressed the cell proliferation, altered the cell cycle distribution and reduced the cell invasion ability of the SiHa cells (P < 0.05). Compared with the untreated group (0.217 ± 0.025 and 0.215 ± 0.024) and the control siRNA group (0.222 ± 0.023 and 0.207 ± 0.027), the expression of cyclin D1 and Cdk2 proteins were significantly decreased in the MACC1 siRNA group (0.076 ± 0.010 and 0.039 ± 0.007) (P < 0.05). Compared with the untreated group (0.099 ± 0.007) and control siRNA group (0.105 ± 0.012), the expression of p21 protein was significantly increased in the MACC1 siRNA group (0.676 ± 0.044) (P < 0.05). The downregulation of MACC-1 expression also evoked a decrease of expressions of MMP-2 and MMP-9 proteins and an increase of E-cadherin protein expression (P < 0.05).. MACC-1 downregulation-mediated inhibition of proliferation and decreased invasion ability of tumor cells may be closely associated with the alterations of expressions of cell cycle- and invasion-related proteins.

Topics: Cadherins; Cell Cycle; Cell Line, Tumor; Cell Movement; Cell Proliferation; Cyclin D1; Cyclin-Dependent Kinase 2; Down-Regulation; Female; Gene Expression Regulation, Neoplastic; Gene Silencing; Humans; Matrix Metalloproteinase 2; Matrix Metalloproteinase 9; RNA, Small Interfering; Trans-Activators; Transcription Factors; Transfection; Uterine Cervical Neoplasms

To investigate the effect and its mechanism of stem cell related transcription factor Sox2 on the proliferation of cervical squamous carcinoma cell line SiHa.. Plasmid pIRES-EGFP-Sox2 or empty plasmid (pIRES-EGFP-empty) was stably transfected into SiHa cells. The expression of Sox2 was detected by both RT-PCT and Western blot. The effects of Sox2 on cellular proliferation and cell cycle were studied by MTT assay and flow cytometry (FCM) respectively. The expression of cell cycle related protein CyclinD1 was detected by Western blot.. Compared to SiHa-EGFP cells, the expression of Sox2 was obviously up-regulated in SiHaSox2 cells (P < 0.01). MTT result showed that SiHa-Sox2 cells grew faster than the control cells. The over expression of Sox2 increased the proportion of transfected cells in phase S. The increased expression of CyclinD1 was further detected after the successful expression of Sox2 (P < 0.05).. Sox2 could enhance the proliferation of cervical squamous cancer cells in the manner of up-regulating CyclinD1 expression.

Topics: Carcinoma, Squamous Cell; Cell Cycle; Cell Line, Tumor; Cell Proliferation; Cyclin D1; Female; Humans; Plasmids; SOXB1 Transcription Factors; Transfection; Up-Regulation; Uterine Cervical Neoplasms

To investigate the expression of La protein in cervical cancer tissues and explore its role in the occurrence and progression of cervical cancer.. The expression of La protein in cervical cancer and normal cervical tissues was detected by immunohistochemical staining. RNA interference technology was used to silence La protein expression in HeLa cells and the changes in cell proliferation, tumor sphere formation and cell cycles were investigated.. The expression of La protein was significantly higher in cervical cancer tissues than in normal cervical tissues (61% vs 9%, P<0.05). Silencing La protein expression in HeLa cells caused significantly reduced the cell proliferation and lowered the tumor sphere formation rate from the control level of (17.1=1.92)% to (6.3=0.45)% (P<0.05), resulting also in G0/G1 cell cycle arrest and reduced cyclin D1 protein expression.. The RNA binding protein La can promote the development of cervical cancer and may play a critical role in the carcinogenesis and progression of cervical cancer.

Topics: Autoantigens; Cell Cycle Checkpoints; Cyclin D1; Female; HeLa Cells; Humans; Ribonucleoproteins; RNA Interference; RNA-Binding Proteins; SS-B Antigen; Uterine Cervical Neoplasms

High-risk human papillomaviruses are closely associated with cervical cancer and its precursor lesions through interactions between the E6 and E7 oncoproteins and the cell-cycle regulatory proteins, such as p53 and pRb, respectively. As other molecules involved in the cell-cycle control seem to be important for human papillomavirus (HPV)-mediated cervical carcinogenesis, we have analyzed the expression of p53, p21, p16, cyclin D1, and Ki-67 and the presence of HPV (HPV pool and HPV-16) by immunohistochemical studies using tissue microarray in low squamous intraepithelial lesions (n=50), high squamous intraepithelial lesions (n=98), and cervical carcinoma (n=18). We have found a significant increase in the expression of p16 and p21 (P<0.001) from low- to high-grade lesions and cancer. In contrast, cyclin D1 expression showed a significant decrease in more severe lesions (P<0.001). p16, Ki-67, p21, and p53 positivity increased with the cell-layer level and the lesion severity, with stronger correlations being observed for p16 and Ki-67. High positivity for HPV pool (96.3%) and HPV-16 (77.5%) immunostaining was detected in all cases, with an association between p16 and cyclin D1 expression and HPV-16 infection. Our tissue microarray results corroborate the usefulness of the immunohistochemical assessment of cell-cycle biomarkers in distinguishing different groups of precursor lesions of the cervix and cervical carcinoma.

Topics: Biomarkers, Tumor; Brazil; Carcinogenesis; Carcinoma in Situ; Cell Cycle; Cervix Uteri; Cyclin D1; Cyclin-Dependent Kinase Inhibitor p16; Cyclin-Dependent Kinase Inhibitor p21; Female; Human papillomavirus 16; Humans; Immunohistochemistry; Ki-67 Antigen; Neoplasm Proteins; Papillomaviridae; Papillomavirus Infections; Precancerous Conditions; Tissue Array Analysis; Tumor Suppressor Protein p53; Uterine Cervical Neoplasms

Abnormal activation of the Wnt/β-catenin signaling pathway is common in human cancers, including cervical cancer. Many papers have shown that SRY (sex-determining region Y)-box (SOX) family genes serve as either tumor suppressor genes (TSGs) or oncogenes by regulating the Wnt signaling pathway in different cancers. We have demonstrated recently that epigenetic silencing of SOX1 gene occurs frequently in cervical cancer. However, the possible role of SOX1 in cervical cancer remains unclear. This study aimed to explore whether SOX1 functions as a TSG in cervical cancer.. We established a constitutive and an inducible system that overexpressed SOX1 and monitored its function by in vitro experiments. To confirm SOX1 function, we manipulated SOX1 using an inducible expression approach in cell lines. The effect of SOX1 on tumorigenesis was also analyzed in animal models.. Overexpression of SOX1 inhibited cell proliferation, anchorage independency, and invasion in vitro. SOX1 suppressed tumor growth in nonobese diabetic/severe combined immunodeficiency mice. After induction of SOX1 by doxycycline (DOX), SOX1 inhibited cell growth and invasion in the inducible system. Repression of SOX1 by withdrawal of DOX partially reversed the malignant phenotype in cervical cells. SOX1 inhibited TCF-dependent transcriptional activity and the Wnt target genes. SOX1 also repressed the invasive phenotype by regulating the expression of invasion-related genes.. Taken together, these data suggest that SOX1 can function as a tumor suppressor partly by interfering with Wnt/β-catenin signaling in cervical cancer.

Topics: Animals; Antigens, CD; Cadherins; Cell Adhesion; Cell Movement; Cell Proliferation; Cyclin D1; Doxycycline; Female; Genetic Vectors; HeLa Cells; Humans; Mice; Mice, SCID; Plasmids; Proto-Oncogene Proteins c-myc; Snail Family Transcription Factors; SOXB1 Transcription Factors; Transcription Factors; Tumor Suppressor Proteins; Uterine Cervical Neoplasms; Wnt Signaling Pathway

The miR-302-367 cluster is specifically expressed in human embryonic stem cells and has been shown to convert human somatic cells into induced pluripotent stem cells. Here, we investigated the role of the miR-302-367 cluster in cervical carcinoma. The cluster was not endogenously expressed in cervical cancer cells, and its ectopic expression did not reprogram the cervical cancer cells to an embryonic stem cell-like state. However, ectopic expression of the miR-302-367 cluster in HeLa and SiHa cervical cancer cells inhibited cell proliferation and tumor formation by blocking the G1/S cell cycle transition. We identified a new cell cycle regulatory pathway in which the miR-302-367 cluster directly down-regulated both cyclin D1 and AKT1 and indirectly up-regulated p27(Kip1) and p21(Cip1), leading to the suppression of cervical cancer cell proliferation. Our findings suggest that the miR-302-367 cluster may be used as a therapeutic reagent for the treatment of cervical carcinoma.

Topics: Animals; Cell Proliferation; Cyclin D1; Cyclin-Dependent Kinase Inhibitor p21; Cyclin-Dependent Kinase Inhibitor p27; Female; Gene Expression Regulation, Neoplastic; Humans; Mice; MicroRNAs; Proto-Oncogene Proteins c-akt; Uterine Cervical Neoplasms

Surgery and radiotherapy have been used for decades to treat cervical cancer; however, high recurrence and lymph node metastasis rates are observed after these procedures. New therapeutic agents are needed to improve survival rates of patients by reducing tumor growth and metastasis. We previously demonstrated that interleukin-8 (IL-8) was associated with lymph node metastasis of early cervical squamous cell carcinoma (SCC). The current study assessed the role of IL-8 in growth and metastasis of cervical SCC and evaluated the effects of targeting IL-8 with small hairpin RNA (shRNA) and a human anti-IL-8 antibody. The human cervical SCC cell lines CaSki (high IL-8 producers), SiHa, HeLa, and SiHa transfected with the IL-8 gene were used for the studies. IL-8 stimulated proliferation, migration, and invasion but prevented apoptosis of SCC cells in vitro. Suppressing IL-8 expression with shRNA reduced cell growth and invasion of SCC cells in vitro. In a xenograft model, SCC cells were inoculated subcutaneously into athymic mice to evaluate the effect of IL-8 and its antibody on tumor growth and metastasis and animal survival. IL-8 enhanced tumor growth and metastasis in vivo concomitant with reduced animal survival. IL-8 antibody treatment of tumor-bearing animals resulted in smaller tumor volume, decreased lymph node metastasis, and longer animal survival. Blockade of IL-8 with an antibody demonstrated significant anti-tumor effects in a xenograft model and may thus provide a potential alternative approach for the treatment of cervical cancer.

Topics: Animals; Antibodies; Antineoplastic Agents; Carcinoma, Squamous Cell; Cell Proliferation; Cyclin D1; Female; Gene Knockdown Techniques; HeLa Cells; Humans; Interleukin-8; Lymphatic Metastasis; Male; Matrix Metalloproteinase 2; Matrix Metalloproteinase 9; Mice; Mice, Nude; Neoplasm Transplantation; RNA, Small Interfering; Tumor Burden; Uterine Cervical Neoplasms; Xenograft Model Antitumor Assays

Cyclin D1, with a common G/A polymorphism in rs9344, is an essential regulator of the G1 phase in cell cycles and plays an important role in several tumor types, and the homology of cyclin D1 with human papillomavirus (HPV)-16 E7 brought our attention to CCND1 gene in cervical cancer. A total of 738 native Chinese subjects consist of 327 cases and 411 controls were enrolled in this study. CCND1 genotyping was analyzed by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) and partially verified by sequencing of genomic DNA and cDNA. The transcription of cyclin D1 mRNA isoforms was analyzed by quantitative PCR; expression of protein isoforms by immunohistochemistry and Western blotting. We observed that the AA genotype had decreased risk of developing cervical cancer (odds ratio [OR] = 0.332; 95% confidence interval [CI] = 0.113-0.978; P = 0.045). The two mRNA isoforms were both transcripted from A and G allele. Transcript b decreased in squamous cell carcinoma of the uterine cervix (SCCUC) group (P = 0.004), especially poorly differentiated group (P = 0.004), and in G allele group of normal subjects (P = 0.001). In immunohistochemistry analysis, cyclins D1, D1a, and D1b failed to correlate with cervical cancer (P = 0.808, 0.445, and 0.095). However, cyclin D1b was downregulated in SCCUC group analyzed by Western blotting (P = 0.039). This study indicates that CCND1 rs9344 polymorphisms confer host susceptibility to cervical cancer. A allele possesses a relative protective effect probably through the cyclin D1b's inhibition on HPV carcinogenesis.

Topics: Adult; China; Cyclin D1; Female; Genotype; Humans; Middle Aged; Polymerase Chain Reaction; Polymorphism, Genetic; Polymorphism, Restriction Fragment Length; Uterine Cervical Neoplasms

Cervical cancer constitutes the second most common cancer in women. It is evident from earlier studies that epidermal growth factor (EGF) is a mitogen, in that it mimics the function of estrogen by mediating cross-talk with other oncoproteins. Although epidermal growth factor receptor (EGFR) is highly expressed in breast and ovarian tumor tissues, its regulation by the exogenous source of its ligand EGF in human papillomavirus (HPV)-associated cervical cancer remains unclear. In this study, we addressed the question of whether EGF is required for the proliferation of HPV-positive cervical cancer cells and what mechanisms are involved. To determine this, we conducted a series of studies using HPV-positive human cervical cancer cells, CaSki and HeLa, and stimulated the cells with EGF. Our findings suggest that 6 h of stimulation with 10 ng/ml of EGF is sufficient to induce cell cycle progression associated with a significant increase in DNA synthesis, EGFR, COX-2 and cyclin D1 levels. Consistently, cellular localization and Western blot analysis for p-EGFR (Try-1045) protein showed an increase after EGF stimulation. Using siRNA gene knockdown assays we have shown that cyclin D1 siRNA has a significant negative effect on EGFR and inhibit cell growth independent of COX-2 levels. In summary, our findings reveal that an exogenous EGF stimulation may enhance HPV-related cervical cancer cell proliferation by activating EGFR and cyclin D1 that is independent of COX-2 levels, suggesting that the inhibitors of EGFR and cyclin D1 may be effective against cervical cancer cell proliferation.

Topics: Cell Cycle; Cell Growth Processes; Cell Line, Tumor; Cyclin D1; Cyclooxygenase 2; Epidermal Growth Factor; ErbB Receptors; Female; HeLa Cells; Human papillomavirus 16; Human papillomavirus 18; Humans; Molecular Targeted Therapy; Papillomavirus Infections; Transcription, Genetic; Uterine Cervical Neoplasms

Recently, thioridazine (10-[2-(1-methyl-2-piperidyl) ethyl]-2-methylthiophenothiazine), a well-known anti-psychotic agent was found to have anti-cancer activity in cancer cells. However, the molecular mechanism of the agent in cellular signal pathways has not been well defined. Thioridazine significantly increased early- and late-stage apoptotic fraction in cervical and endometrial cancer cells, suggesting that suppression of cell growth by thioridazine was due to the induction of apoptosis. Cell cycle analysis indicated thioridazine induced the down-regulation of cyclin D1, cyclin A and CDK4, and the induction of p21 and p27, a cyclin-dependent kinase inhibitor. Additionally, we compared the influence of thioridazine with cisplatin used as a control, and similar patterns between the two drugs were observed in cervical and endometrial cancer cell lines. Furthermore, as expected, thioridazine successfully inhibited phosphorylation of Akt, phosphorylation of 4E-BP1 and phosphorylation of p70S6K, which is one of the best characterized targets of the mTOR complex cascade. These results suggest that thioridazine effectively suppresses tumor growth activity by targeting the PI3K/Akt/mTOR/p70S6K signaling pathway.

Topics: Adaptor Proteins, Signal Transducing; Antineoplastic Agents; Apoptosis; Caspase 3; Cell Cycle Proteins; Cell Division; Cell Line, Tumor; Cell Survival; Cyclin A; Cyclin D1; Cyclin-Dependent Kinase 4; Cyclin-Dependent Kinase Inhibitor p21; Cyclin-Dependent Kinase Inhibitor p27; Down-Regulation; Endometrial Neoplasms; Female; HeLa Cells; Humans; Phosphatidylinositol 3-Kinases; Phosphoproteins; Phosphorylation; Proto-Oncogene Proteins c-akt; Ribosomal Protein S6 Kinases, 70-kDa; Signal Transduction; Thioridazine; TOR Serine-Threonine Kinases; Uterine Cervical Neoplasms

We recently characterized the molecular linkage that directs both BCL10 overexpression and nuclear translocation in response to inflammation-related NF-κB signaling pathway. Since NF-κB activation has been shown to occur in the pathogenesis of cervical cancer, we sought to investigate whether BCL10 possesses clinical significance in relation to cervical cancer.. Four cervical cancer cell lines (C33A, SiHa, HeLa, and CaSki) were used in this study. The DNA-binding activity of NF-κB was determined by the luciferase assay. The expression of BCL10, NF-κB, and cyclin D1 in tumor cells from an array of 182 tissue samples was examined using immunohistochemical staining.. We transfected four cervical cancer cell lines with BCL10 small interfering RNA (siRNA), and discovered that the down-regulation of BCL10 inhibited the viability of these cervical cancer cells through G1 arrest. BCL10 siRNA treatment inhibited the expression of p-IKKβ and p-IκB, and also down-regulated both NF-κB activation cyclin D1, its downstream cell cycle protein. Our results reveal that cervical cancer had a higher rate of positive cytoplasmic staining (74.1%, 123/166) than either carcinoma in situ (50.0%, 3/6) or normal cervix (0.0%, 0/10); and that poorly differentiated cancer had a higher rate of cytoplasm staining (80.7%, 71/88) than moderately differentiated (75.4%, 43/57) and well differentiated (40%, 4/10) carcinoma. Furthermore, nuclear expression of BCL10 was closely associated with NF-κB activation (p<0.001) and cyclin D1 expression (p<0.001).. Our findings indicate that BCL10 plays an important role in controlling the growth of cervical cancer cells through NF-κB dependent cyclin D1 regulation.

Topics: Adaptor Proteins, Signal Transducing; B-Cell CLL-Lymphoma 10 Protein; Cell Culture Techniques; Cell Growth Processes; Cell Line, Tumor; Cyclin D1; Female; Gene Expression Regulation, Neoplastic; HeLa Cells; Humans; Immunohistochemistry; NF-kappa B; RNA, Small Interfering; Signal Transduction; Transfection; Uterine Cervical Neoplasms

In HCT116 colorectal cancer cells, HeLa cervical cancer cells and HuH-7 hepatoma cells, miR-223 is expressed at a low level. Through infection with lentivirus containing miR-223 precursor, miR-223 [corrected] was overexpressed in all these cells. Interestingly, the expression levels of FOXO1 mRNA and protein, and phosphorylation levels became significantly lower than those of their control. FOXO1 was down-regulated mainly in the cytoplasm, while the nuclear FOXO1 level became relatively high compared to the cytoplasm. As the unphosphorylated active form of FOXO1 increased in the cells, cyclin D1/p21/p27 were up-regulated at either mRNA or protein level. Proliferation of the cells was also greatly inhibited when miR-223 was over-expressed. Therein, our data suggest that miR-223 regulates FOXO1 expression and cell proliferation.

Topics: Cell Line, Transformed; Cell Line, Tumor; Cell Nucleus; Cell Proliferation; Colorectal Neoplasms; Cyclin D1; Cyclin-Dependent Kinase Inhibitor Proteins; Cytoplasm; Female; Forkhead Box Protein O1; Forkhead Transcription Factors; Gene Expression Regulation, Neoplastic; Humans; Liver Neoplasms; Male; MicroRNAs; Neoplasm Proteins; Neoplasms; Phosphorylation; Protein Processing, Post-Translational; Protein Transport; RNA, Messenger; Uterine Cervical Neoplasms

Oncogene amplification is a key step in cell transformation towards malignancy. Chromosomal aberrations involving the long arm of chromosome 11, including amplifications at 11q13 and 11q22, have been previously reported in cervical cancer. While the role of the CCND1 gene as the driver gene for 11q13 amplification is well established in different tumor types, the significance of the 11q22 amplicon is less clear. The 11q22 amplicon corresponds to several putative target genes including the apoptose inhibitor BIRC2, recently detected as amplified in cervical cancer cell lines. To better understand the distribution and frequency of 11q amplification sites in uterine cervical carcinomas, we analyzed BIRC2 and CCND1 copy number changes using fluorescence in situ hybridization in a tissue microarray containing 238 cervical cancers. High-level amplification of BIRC2 was found in 12.9 % of tumors. Amplification of BIRC2 in cervical carcinomas was homogeneous as shown in corresponding whole tissue sections of amplified tumors at the tissue microarray. BIRC2 amplification was significantly more frequent than CCND1 amplification (2.1 %) in our cohort (p < 0.01), and amplification of both genes were independent from each other. BIRC2 amplification was associated with younger-patient age (p < 0.05) and squamous cell differentiation (p = 0.025) of cervix carcinomas. However, BIRC2 copy number changes were not related to tumor stage, grading and nodal status of cervical cancers. In conclusion, BIRC2 is amplified in a subset of squamous cell carcinoma of the uterine cervix. Further studies are necessary to evaluate possible prognostic effects of BIRC2 copy number gains in cervical carcinomas.

Topics: Adult; Aged; Aged, 80 and over; Carcinoma, Squamous Cell; Cyclin D1; Female; Gene Amplification; Humans; Immunohistochemistry; In Situ Hybridization, Fluorescence; Inhibitor of Apoptosis Proteins; Middle Aged; Neoplasm Grading; Neoplasm Staging; Tissue Array Analysis; Ubiquitin-Protein Ligases; Uterine Cervical Neoplasms; Young Adult

Explore the role of plasminogen activator inhibitor-1 (PAI-1) in cervical cancer and its relationship to hypoxia and the expression of p53, Ku70/80, and cyclin D1.. The expression of PAI-1, cyclin D1, and p53, together with tumor oxygenation, were determined in 43 consecutive patients suffering from localized cervical carcinoma. Oncoprotein expression was determined by immunohistochemistry. Tumor oxygenation was measured using a polarographic probe system, "pO2 histography.". PAI expression was considered negative in 32.6% and overexpressed in 18.6% of cases. Cyclin D1 showed a median expression of 5.0 (range 0-70). We observed a positive association between PAI expression and altered p53 (p = 0.049) and cyclin D1 (p = 0.020). An inverse association was detected between PAI and Ku70/80 expression (p = 0.042). Cyclin D1 staining increased according to tumor volume (r = 0.314, p = 0.009). We did not observe a significant association between PAI and hypoxia or other clinicopathological parameters.. The present results show that PAI-1 overexpression is associated with nonhomologous end-joining DNA repair down-regulation (low Ku70/80 expression) and with increased p53 and cyclin D1 expression, and they suggest that PAI-1 plays a role in the tumor behavior in cervical carcinoma.

Topics: Adult; Aged; Aged, 80 and over; Antigens, Nuclear; Biopsy; Cell Hypoxia; Cervix Uteri; Cyclin D1; DNA-Binding Proteins; Female; Gene Expression Regulation, Neoplastic; Humans; Immunoenzyme Techniques; Ku Autoantigen; Middle Aged; Neoplasm Staging; Plasminogen Activator Inhibitor 1; Prognosis; Prospective Studies; Tumor Suppressor Protein p53; Uterine Cervical Neoplasms

Immunohistochemistry is helpful in distinguishing cervical neoplastic lesions from their histological mimics, but has contributed less towards the sometimes problematic distinction of in-situ and superficially invasive tumours. Epithelial-mesenchymal transition (EMT) may be a mechanism of invasion in cervical neoplasia and expression of EMT-associated proteins could prove useful in this diagnostic setting.. Immunohistochemical expression of cyclin D1, E-cadherin and beta-catenin was assessed in 22 biopsy specimens from FIGO Stage IA cervical squamous carcinomas, all of which also included foci of cervical intraepithelial neoplasia (CIN) 3, nine biopsies of CIN 3 adjacent to carcinoma, and 10 cases of CIN 3 only. Most invasive tumour cells expressed cyclin D1 and showed a reduction in E-cadherin and beta-catenin staining. Nuclear beta-catenin expression was not observed. Cyclin D1 staining was reduced or showed altered distribution in most cases of CIN 3, while adhesion protein expression generally was preserved. However, altered protein expression similar to that of invasion was seen in some CIN lesions.. Most superficially invasive cervical squamous carcinomas show immunophenotypical changes consistent with EMT. These alterations, particularly cyclin D1 expression, may be useful diagnostically. Similar changes in CIN 3 lesions may indicate the acquisition of increased invasive potential.

Topics: beta Catenin; Biomarkers, Tumor; Biopsy; Cadherins; Carcinoma, Squamous Cell; Cervix Uteri; Cyclin D1; Epithelial-Mesenchymal Transition; Female; Humans; Immunophenotyping; Neoplasm Staging; Uterine Cervical Dysplasia; Uterine Cervical Neoplasms

We examined the distribution of the CCND1 A870G (rs9344) polymorphic variant in patients with cervical cancer (n = 129) and healthy individuals (n = 288) in a sample of a Polish cohort. We showed that patients with advanced cervical cancer bearing the CCND1 A/A and A/G genotypes displayed a 1.811-fold increased risk of cervical cancer (95% CI = 1.150-2.852, p = 0.0098). We also found a significantly higher frequency of the CCND1 870A allele in patients with cancer than in controls, p = 0.0116. Our investigation confirmed that the CCND1 870A gene variant may be a genetic risk factor in the incidence of advanced cervical cancer.

Topics: Carcinoma; Cyclin D1; Female; Genetic Predisposition to Disease; Genotype; Humans; Middle Aged; Neoplasm Staging; Polymerase Chain Reaction; Polymorphism, Restriction Fragment Length; Polymorphism, Single Nucleotide; Risk Factors; Uterine Cervical Neoplasms

The La protein is an essential RNA-binding protein implicated in different aspects of RNA metabolism. Herein, we report that small interfering (siRNA)-mediated La depletion reduces cell proliferation of different cell lines concomitant with a reduction in cyclin D1 (CCND1) protein. To exclude off-target effects we demonstrate that exogenous La expression in La-depleted cells restores cell proliferation and CCND1 protein levels. In contrast, proliferation of immortalized CCND1 knockout cells is not affected by La depletion, supporting a functional coherence between La, CCND1 and proliferation. Furthermore, we document by reversible in vivo crosslinking and ribonucleoprotein (RNP) immunoprecipitation an association of the La protein with CCND1 messengerRNA and that CCND1 internal ribosome entry site (IRES)-dependent translation is modulated by La protein level within the cell. In addition, we show elevated La protein expression in cervical cancer tissue and its correlation with aberrant CCND1 protein levels in cervical tumor tissue lysates. In conclusion, this study establishes a role of La in cell proliferation and CCND1 expression and demonstrates for the first time an overexpression of the RNA-binding protein La in solid tumors.

Topics: Autoantigens; Blotting, Western; Carcinoma, Squamous Cell; Cell Proliferation; Cell Separation; Cyclin D1; Female; Flow Cytometry; Gene Expression; Gene Knockout Techniques; HeLa Cells; Humans; Immunohistochemistry; Immunoprecipitation; Reverse Transcriptase Polymerase Chain Reaction; Ribonucleoproteins; RNA, Small Interfering; SS-B Antigen; Tissue Array Analysis; Uterine Cervical Neoplasms

Cervical cancer is the most common cancer in Indian females and is associated with infection with high-risk Human papilloma viruses (HPVs) which encode viral oncoprotein E6 and E7. Estradiol has been established as a risk factor for cervical cancer and has been shown to play a synergistic role with viral oncoproteins. Curcumin (Diferuloyl methane), a chemopreventive agent, is a natural compound extracted from Curcuma longa that allows suppression and retardation of carcinogenesis in many types of cancer and is currently being tested in various human clinical trials as it has been found to be well tolerated at higher doses with a relatively well established safety profile. The objective of this study was to test the effect of curcumin on HPV-positive and negative cervical cancer cell lines HeLa, SiHa, CaSki, and C33A pretreated with estradiol. It was found that HPV-positive cells pretreated with estradiol show reduced apoptosis as compared to curcumin by itself. However, curcumin was able to counteract the proliferative response of estradiol, and induce apoptosis. There was no difference in percentage apoptosis as compared to estradiol pretreatment in HPV-negative cell line C33A. Molecular studies showed elevation of Telomerase, viral oncoproteins E6 and E7, PCNA, p16, Cyclin D1 in HPV-positive cell lines on treatment with estradiol but after treatment with curcumin the level of E7, PCNA, and Cyclin D1 was reduced but the level of E6, Telomerase, and p16 was unaltered. Furthermore, estradiol-pretreated HPV-negative cell line C33A showed reduction in level of Telomerase, PCNA, p16, and activation of both p53 and p73 tumor suppressor proteins, thus, demonstrating the importance of E6 in estradiol-mediated protective effect.

Topics: Antineoplastic Agents; Apoptosis; Blotting, Western; Cell Line, Tumor; Cell Proliferation; Curcumin; Cyclin D1; Cyclin-Dependent Kinase Inhibitor p16; DNA-Binding Proteins; Estradiol; Female; Flow Cytometry; Humans; Models, Biological; Oncogene Proteins, Viral; Papillomaviridae; Papillomavirus E7 Proteins; Proliferating Cell Nuclear Antigen; Signal Transduction; Telomerase; Tumor Suppressor Protein p53; Up-Regulation; Uterine Cervical Neoplasms

Cervical carcinoma is the second most common cause of death from gynecological cancers worldwide. Knowledge of the molecular mechanisms underlying the tumorigenesis of cervical cancer cell, except human papilloma virus infection, is limited.. A microarray was used to study the differential expression of genes in cancerous tissues to identify new molecular markers for diagnosis and prognosis. Their differential expression was confirmed with Western blotting and immunohistochemical analyses. The clinical correlations and prognostic significance of the aberrantly expressed proteins were evaluated to identify novel biomarkers of cervical cancer.. The expression of gelsolin was significantly upregulated in 78% of patients with cervical cancer, and gelsolin was selected for further study. Gelsolin expression was stronger in cervical tumor tissues than in the surrounding noncancerous tissues (P<0.001). Gelsolin expression in the plasma of cervical cancer patients was increased 2.2-fold compared with that of healthy control subjects (P<0.001). The levels of plasma gelsolin in the early and late stages were significantly different (P=0.006). According to immunohistochemical analysis, increased gelsolin expression was associated with histological type and FIGO stage II. The 5-year overall survival and recurrence-free survival rates for the low-expression group (cut-off=115) were significantly higher than those of the high-expression group. Cancer cells with reduced gelsolin expression exhibited reduced migration and proliferation.. These results provide strong evidence that gelsolin plays an important role in cellular proliferation and migration in cervical cancer and suggest that gelsolin is a promising marker for cervical cancer screening and prognosis.

Topics: Adult; Aged; Biomarkers, Tumor; Cadherins; Case-Control Studies; Cell Growth Processes; Cell Movement; Cyclin D1; Cyclin-Dependent Kinase Inhibitor p21; Epithelial-Mesenchymal Transition; Female; Fibronectins; Gelsolin; Gene Expression Profiling; Gene Knockdown Techniques; HeLa Cells; Humans; Matrix Metalloproteinase 2; Middle Aged; Neoplasm Staging; Prognosis; Retrospective Studies; Survival Rate; Up-Regulation; Uterine Cervical Neoplasms; Vimentin

Cervical cancer cells exhibit an increased requirement for ubiquitin-dependent protein degradation associated with an elevated metabolic turnover rate, and for specific signaling pathways, notably HPV E6-targeted degradation of p53 and PDZ proteins. Natural compounds with antioxidant properties including flavonoids and triterpenoids hold promise as anticancer agents by interfering with ubiquitin-dependent protein degradation. An increasing body of evidence indicates that their α-β unsaturated carbonyl system is the molecular determinant for inhibition of ubiquitin-mediated protein degradation up-stream of the catalytic sites of the 20S proteasome. Herein we report the identification and characterization of a new class of chalcone-based, potent and cell permeable chemical inhibitors of ubiquitin-dependent protein degradation, and a lead compound RAMB1. RAMB1 inhibits ubiquitin-dependent protein degradation without compromising the catalytic activities of the 20S proteasome, a mechanism distinct from that of Bortezomib. Treatment of cervical cancer cells with RAMB1 triggers unfolded protein responses, including aggresome formation and Hsp90 stabilization, and increases p53 steady state levels. RAMB1 treatment results in activation of lysosomal-dependent degradation pathways as a mechanism to compensate for increasing levels of poly-ubiquitin enriched toxic aggregates. Importantly, RAMB1 synergistically triggers cell death of cervical cancer cells when combined with the lysosome inhibitor Chloroquine.

Topics: Antineoplastic Agents; Biocatalysis; Cell Adhesion; Cell Death; Cell Line, Tumor; Cell Survival; Chloroquine; Cyclin D1; Drug Screening Assays, Antitumor; Drug Synergism; Female; Heat-Shock Response; HSP90 Heat-Shock Proteins; Humans; Keratinocytes; Papillomaviridae; Polyubiquitin; Proteasome Endopeptidase Complex; Proteasome Inhibitors; Protein Stability; Proteolysis; Stress, Physiological; Tumor Stem Cell Assay; Tumor Suppressor Protein p53; Ubiquitin; Ubiquitination; Uterine Cervical Neoplasms

It may be difficult to distinguish reactive glandular lesions from adenocarcinoma in situ of the uterine cervix, and although several immunohistochemical markers have established value in this diagnostic setting, none is completely reliable. We have noted that neoplastic endocervical lesions often show loss of nuclear cyclin D1 expression in contrast to benign glandular cells. Therefore, we investigated cyclin D1 staining in a series of 64 cervical biopsy specimens including examples of normal and reactive endocervical epithelium, adenocarcinoma in situ, stratified mucin-producing intraepithelial lesions, and invasive adenocarcinoma. Thirteen specimens also included a component of high-grade cervical squamous intraepithelial neoplasia. Normal endocervical epithelium usually expressed cyclin D1, although staining was typically focal, and there was increased immunoreactivity in reactive and metaplastic glandular cells including tubo-endometrioid metaplasia. In contrast, most cases of adenocarcinoma in situ were completely negative and, therefore, cyclin D1 staining distinguished benign from neoplastic epithelial cells. Although focal cyclin D1 expression was observed in 5/19 cases of adenocarcinoma in situ, the staining was associated with more marked cytological atypia precluding confusion with a reactive process. The invasive adenocarcinomas were mainly negative for cyclin D1. However, focal staining was observed in 10/19 cases and was mainly restricted to cells at the deep tumor margin, or to small infiltrative glands and detached cell clusters within the stroma. In conclusion, cyclin D1 can be included within an immunohistochemical panel to aid in the distinction between reactive cervical glandular lesions and adenocarcinoma in situ. The localized distribution of staining within invasive lesions suggests that cyclin D1 up-regulation has a specific role during the progression of some endocervical adenocarcinomas.

Topics: Adenocarcinoma; Biomarkers, Tumor; Cervix Uteri; Cyclin D1; Female; Humans; Immunohistochemistry; Uterine Cervical Dysplasia; Uterine Cervical Neoplasms

To investigate the effects of lanthanum chloride on proliferation and migration activity of human cervical cancer cells in vitro which may be a new anti-cervical cancer drug and provide experimental data for cervical cancer treatment.. HeLa cells cultured in vitro were divided into two groups: experimental group and control group. In experimental group, the cells were respectively treated with lanthanum chloride at different concentrations, 5, 50 and 100 µmol/L, while the cells in the control group were not treated with lanthanum chloride. The cell growth was observed by inverted microscope and the morphology changes of the cells were observed by the laser scanning confocal microscope (LSCM). Proliferation of HeLa cells in the two groups was detected by methyl thiazolyl tetrazolium (MTT) test; apoptosis rate was analyzed by flow cytometry (FCM). Cell migration test was applied to observe the effect of lanthanum chloride on migration. Reverse transcription (RT)-PCR was employed to evaluate the effects of lanthanum chloride on proliferation gene (cyclinD1), anti-apoptosis gene (zinc finger protein A20) and migration-related gene (matrix metalloproteinase 9, MMP-9).. The status of cell growth was observed under the inverted microscope: with the increased of the lanthanum chloride concentrations, the cell density of reduced, the granule in cytoplasm increased, color intensifying and intercellular space enlarged; some cells became rounding and dead, floating in the culture media; the exfoliated cells increased gradually in the experimental groups. While In the control group, the cells grew adherently, with clear morphology and plump cytoplasm, and adjacent cell grew in lamellar. Observed with LSCM: the nuclear chromatin condensated and marginated with the volume of nuclear decreased in experimental groups. With the increase of the lanthanum chloride concentrations, nuclei in the experimental groups became pyknotic and then underwent karyorrhexis. However, the nuclear of the cells in control group were inact. The growth inhibition rates of lanthanum chloride groups (5, 50, 100 µmol/L) were 24%, 51% and 78%, respectively, in which each was significantly higher than that of the control group (P < 0.05); the apoptosis rates of lanthanum chloride group were (4.91 ± 0.39)%, (7.30 ± 0.71)% and (13.48 ± 0.92)%, respectively, which were all significantly higher than that of the control group [(0.89 ± 0.11)%, P < 0.01]. The migration ability of the cells was also decreased by the treatment of lanthanum chloride, the number of migrated cells in lanthanum chloride groups were 22.2 ± 4.3, 12.0 ± 3.2 and 7.8 ± 2.6 respectively, which were all significantly lower than that of the control group (41.2 ± 5.4, P < 0.01). The expression of genes of cyclinD1, A20 and MMP-9, were all decreased by the treatment of lanthanum chloride in a dose-dependent manner.. Lanthanum chloride can inhibit the proliferation and migration of cervical cancer cells, and induce apoptosis by down-regulating cyclinD1, A20, and MMP-9 expressions in vitro.

Topics: Antineoplastic Agents; Apoptosis; Cell Movement; Cell Proliferation; Cyclin D1; Dose-Response Relationship, Drug; Down-Regulation; Endopeptidases; Female; Flow Cytometry; Gene Expression Regulation, Neoplastic; HeLa Cells; Humans; Lanthanum; Matrix Metalloproteinase 9; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Uterine Cervical Neoplasms

Cervical cancer is the most common cancer in Indian females and is associated with infection with high risk Human papillomaviruses (HPVs). Curcumin (Diferuloyl methane), a chemopreventive agent, is a natural compound extracted from Curcuma longa that allows suppression of carcinogenesis. The objective of this study was to identify the molecular mechanism of curcumin induced apoptosis in HPV positive cervical cancer HeLa, SiHa and Ca Ski cells. Curcumin causes distinct inhibition of human telomerase reverse transcriptase (hTERT) the catalytic core of telomerase thereby reducing proliferation of cancer cells. Curcumin mediated apoptosis in these cells appears to be due to upregulation of proapoptotic Bax, AIF, release of cytochrome c and down regulation of antiapoptotic Bcl-2, Bcl-XL in HeLa and SiHa. This was accompanied by an increase in caspase-3 and -9 activity, suggesting the role of mitochondria in curcumin mediated apoptotic cell death. Curcumin acts as an anti-inflammatory and anti-proliferative agent by causing down regulation of COX-2, iNOS and cyclin D1 in all the three cell lines but to different extent.

Topics: Apoptosis; Caspase 3; Caspase 9; Cell Line, Tumor; Cell Proliferation; Curcumin; Cyclin D1; Cytochromes c; Female; Flow Cytometry; HSP70 Heat-Shock Proteins; Humans; Proto-Oncogene Proteins c-bcl-2; Telomerase; Uterine Cervical Neoplasms

The potential association of single nucleotide polymorphisms (SNPs) (G870A and G1722C) of CCND1 with susceptibility to cervical cancer was investigated. The study included 200 cervical cancer cases along with an equal number of healthy controls. Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis and direct sequencing were employed for genotyping. We found that women carrying the 870AA genotype have a 2.49-fold increased risk for the development of cervical cancer (odds ratio (OR) 2.49; 95% confidence interval (CI) 1.51-4.09; p = 0.0004) compared with GG+GA genotypes. For the 1722 locus, the frequency of the polymorphic 'C' allele was strongly associated with a reduced risk of cervical cancer (p = 0.019; OR 0.71; 95% CI 0.54-0.94). Our data suggest that CCND1 G870A polymorphism could act as a risk factor for the development of cervical cancer. And G1722C polymorphism may play a protective role against the development of human papillomavirus-associated cervical cancer among Indian women.

Topics: Adult; Cyclin D1; Female; Genetic Predisposition to Disease; Human papillomavirus 16; Humans; India; Linkage Disequilibrium; Middle Aged; Polymorphism, Restriction Fragment Length; Polymorphism, Single Nucleotide; Uterine Cervical Neoplasms

High-risk human papillomavirus (hrHPV) infection is the major risk factor for cervical cancer (CxCa). The role of genetic susceptibility in the disease has been suggested, but the existing data lack consistency. We conducted a nested case-control study on 973 CxCa cases and 1,763 matched controls, from two Swedish population-based cohorts to examine the association of common genetic variants with CxCa risk. Human leukocyte antigen (HLA) alleles and 24 other polymorphisms in 14 genes were selected on the basis of reported association or mechanistic plausibility with an HPV infection or cervical cancer development. Genotyping was conducted using multiplex PCR and Luminex technology. A significant association of CxCa with various polymorphisms was observed: rs1800797 in the IL-6 gene (odds ratio [OR] = 0.88, 95% confidence intervals [CI]: 0.79-0.99); rs1041981 in the LTA gene (OR = 0.87, 95% CI: 0.78-0.98), and rs9344 in the CCND1 gene (OR = 1.14, 95% CI: 1.02-1.27), for those individuals carrying the rare allele. Additionally, the alleles 0401 and 1501 of the HLA class II DRB1 locus were associated with an increased risk (OR = 1.23, 95% CI: 1.04-1.45 and OR = 1.29, 95% CI: 1.11-1.50, respectively), and allele 1301 was associated with decreased risk (OR = 0.59, 95% CI: 0.47-0.73). The effects of CCND1 and the HLA*DRB1 alleles were independent of the effect of smoking. We did not find any association of risk with polymorphisms in genes related to the innate immune system. In conclusion, our study provides evidence for genetic susceptibility to CxCa due to variations in genes involved in the immune system and in cell cycle.

Topics: Adult; Aged; Case-Control Studies; Cervix Uteri; Cyclin D1; Female; Genotype; HLA-DR Antigens; HLA-DRB1 Chains; Humans; Interleukin-6; Middle Aged; Papillomaviridae; Papillomavirus Infections; Polymerase Chain Reaction; Polymorphism, Single Nucleotide; Risk Factors; Sweden; Uterine Cervical Neoplasms; Young Adult

In-depth study of cell cycle proteins and human papillomavirus (HPV) genotyping can provide useful information about the malignant potential of precursor lesions of cervical carcinoma (CC). Immunostaining of cell cycle-related proteins (p16, cyclin D1, Ki-67, p53, and ProEx C) was evaluated using tissue microarrays, and HPV genotypes were identified in 144 cervical tissue specimens encompassing normal or benign epithelial lesions, low- and high-grade squamous intraepithelial lesions (LSIL and HSIL, respectively), and CC. In addition, 14 cases with atypical immature metaplasia (AIM) were included to compare their immunohistochemical features with those of well-established precursor lesions. Expression of p16, Ki-67, and ProEx C was most associated with the severity of dysplasia. Positive expression of p16, Ki-67, and ProEx C and negative expression of p53 seem to be related to HPV-16 infection. AIM cases show an immunohistochemical pattern more similar to LSIL than to HSIL. Immunohistochemical assessment of cell cycle proteins may help to distinguish normal and benign conditions of the cervix from precursor lesions of CC.

Topics: Antigens, Neoplasm; Biomarkers, Tumor; Cell Cycle; Cell Cycle Proteins; Cyclin D1; Cyclin-Dependent Kinase Inhibitor p16; DNA Topoisomerases, Type II; DNA-Binding Proteins; Female; Humans; Immunohistochemistry; Ki-67 Antigen; Minichromosome Maintenance Complex Component 2; Neoplasm Proteins; Nuclear Proteins; Papillomavirus Infections; Precancerous Conditions; Tissue Array Analysis; Tumor Suppressor Protein p53; Uterine Cervical Neoplasms

The presence of lymph node metastases is associated with poor prognosis in early stage cervical cancer. As of yet, no molecular markers predicting lymph node metastases have been identified. We examined single genetic markers and a composite marker, comprised of three fluorescence in situ hybridization (FISH) probes targeting the genes LAMP3, PROX1, and PRKAA1, in pretreatment cervical biopsies from 16 lymph node positive cases and 15 lymph node negative controls from women with stage IB and IIA cervical cancer. In addition, we determined clonal patterns by including CCND1 to compare the clonal constitution of primary tumors and associated lymph node metastases. The composite FISH marker allowed for classification of patients into those with and without lymph node metastases with a sensitivity and specificity of 75% and 87%, respectively (P = 0.001). The positive predictive value and negative predictive value were 86% and 76%, respectively. Clonal patterns varied among the tumors. In many cases, changes between the primary tumor and lymph node metastases in the most common clones may indicate that certain clones have a growth advantage for establishing metastases in lymph nodes. We conclude that the composite FISH marker may be useful for determining risk for subsequent development of lymph node metastases in patients with cervical cancer.

Topics: Adult; Aged; AMP-Activated Protein Kinases; Biomarkers, Tumor; Cyclin D1; Female; Homeodomain Proteins; Humans; In Situ Hybridization, Fluorescence; Lymphatic Metastasis; Lysosomal Membrane Proteins; Middle Aged; Neoplasm Proteins; Neoplasm Staging; ROC Curve; Sensitivity and Specificity; Tumor Suppressor Proteins; Uterine Cervical Neoplasms

Cyclin D1 (CCND1) is a key regulatory protein, playing a critical role in the transition from G1 to S phase of the cell cycle. We have evaluated the association between CCND1 A870G polymorphism and risk of cervix cancer in north Indian women by using PCR-RFLP method. This association was estimated by computing odds ratio (ORs) and a 95% Confidence Intervals (95% CI) using a Multivariate Logistic Regression Analysis. No significant association was observed between CCND1 genotypes and overall risk of cervix cancer. But when stratified histologically, statistically significant (OR: 3.7, 95% CI: 1.56-8.87, P: 0.001) increased risk of squamous cell carcinoma (SCC) was observed for individuals with AA genotype. Thus our findings suggest that CCND1 (G870A) polymorphism may be associated with increased risk of SCC of the uterine cervix in north Indian women.

Topics: Case-Control Studies; Confidence Intervals; Cooking; Cyclin D1; Female; Genetic Predisposition to Disease; Humans; India; Middle Aged; Odds Ratio; Polymorphism, Restriction Fragment Length; Polymorphism, Single Nucleotide; Tobacco Smoke Pollution; Uterine Cervical Neoplasms; Wood

Estrogen stimulates human papilloma virus oncogene expression, promotes cervical cancer (CC) cell proliferation and prevents apoptosis. Therefore, blockage of estrogen function may have therapeutic application to CC.. CasKi CC cells were transfected with an adenovirus expressing a dominant negative estrogen receptor gene (Ad-ER-DN) and their responses were investigated by RT-PCR, Flow Cytometry and Western blot assays.. Transfected cells showed disturbance of cell colony morphology, reduced HPV E6 and E7 mRNA, interruption of cell proliferation, reduced cyclin D1 protein and expression of apoptosis.. We report, for the first time, the use of Ad-ER-DN to block estrogen receptors which led to dramatic changes in CC cells that are consistent with the possible reactivation of cellular p53 and Rb function. Their reactivation most likely allowed the recognition of existing chromosome abnormalities as a serious stress signal and the initiation of a cascade of cellular events in response to the stress, including the activation of the core apoptotic machinery which led to self-destruction of the CC cells.

Topics: Adenoviridae; Apoptosis; Cell Growth Processes; Cyclin D1; Female; Flow Cytometry; Genetic Therapy; Genetic Vectors; Human papillomavirus 16; Humans; Oncogene Proteins, Viral; Papillomavirus E7 Proteins; Papillomavirus Infections; Receptors, Estrogen; Repressor Proteins; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Transfection; Uterine Cervical Neoplasms

The histological criteria for cervical intraepithelial neoplastic lesions and their follow-ups have been established, but their reproducibility, specificity and sensibility are not certain. Immunohistochemical markers provide more information on each specific case, in order to facilitate its classification and, eventually, its prognosis. Using immunohistochemical techniques, this study analyzes the prognostic value of three markers (Ki-67, c-erbB2 and Cyclin D1) in cases of low grade squamous intraepithelial neoplasia (CIN-I), high grade squamous intraepithelial neoplasia (CIN-III), and infiltrating squamous cell carcinoma (SCC) taken from a group of cervical samples. In situ hybridization was performed in order to detect high-risk HPV. High risk HPV was demonstrated in 82%, 89% and 100% of the LGSIL, HGSIL and SCC cases, respectively. C-erbB2 expression was detected in 9%, 33% and 50% of the LSIL, HGSIL and SCC cases, respectively. The Ki-67 LI was 25%, 68% and 65.5% in the LGSIL, HGSIL and SCC cases, respectively. Nuclear Cyclin D1 expression was seen in 82%, 11% and 30% of the CIN-I,CIN-III and SCC cases, respectively. We observed that the cytoplasmic cyclin D1 expression increased with the severity of the lesion instead of the nuclear expression decreasing with the progression of the pathology. Nuclear and cytoplasmic Cyclin D1 expression seemed to be related to HPV high risk infection. We concluded that Cyclin D1, cerbB2 and The Ki-67 LI expression changed in relation to the severity of the lesion and that they could be helpful in making a differential diagnosis.

Topics: Antigens, CD1; Biomarkers, Tumor; Carcinoma, Squamous Cell; Cyclin D1; Diagnosis, Differential; Female; Humans; Immunohistochemistry; In Situ Hybridization; Ki-67 Antigen; Papillomavirus Infections; Receptor, ErbB-2; Sensitivity and Specificity; Uterine Cervical Dysplasia; Uterine Cervical Neoplasms

Topics: Carcinoma, Small Cell; Cyclin D1; Cyclin-Dependent Kinase Inhibitor p16; Female; Humans; Immunohistochemistry; Retinoblastoma Protein; Uterine Cervical Neoplasms

Cervical cancer is the one of the most common cancers affecting the south Indian women. The objective of the present study was to analyze the pl6/Rb/cyclin D1 pathway in the normal (n=30), dysplastic (n=56), and malignant cervical epithelium (n=142) using immunohistochemistry. The positive immunoreactivity for p16 was as follows: CIN I--1/12 (8.3%), CIN II--2/8 (25%), CIN III--31/36 (86.1%), and in invasive tumors-121/142 (85.1%); for cyclin D1 it was CIN I--4/12 (66.6%), CIN II--5/8 (62.5%), CIN III--0%, and invasive tumors--5/142 (3.5%); and for pRb it was CIN I--9/12 (75%), CIN II--5/8(62.5%), CIN III--1/36 (97.2%), and in invasive tumors--41/142 (28.8%). Expression of cyclin D1 and p16 in the CINs were mutually exclusive and the correlation between the two biomarkers was found to be statistically significant (p = 0.0009). There was downregulation of pRb in invasive cancers, with only 32% (39/121) of the p16-positive tumors being positive for pRb (p = 0.032). Analysis of the pattern of expression of these biomolecules showed increased p16-positive phenotypes and decreased cyclin D1- and pRB-positive phenotype among the invasive tumors compared to low-grade CIN lesions.

Topics: Cervix Uteri; Cyclin D1; Cyclin-Dependent Kinase Inhibitor p16; Down-Regulation; Epithelial Cells; Female; Gene Expression Regulation; Humans; Immunohistochemistry; India; Neoplasm Staging; Phenotype; Retinoblastoma Protein; Uterine Cervical Dysplasia; Uterine Cervical Neoplasms

Peptidyl-prolyl cis/trans isomerase Pin1 is prevalently overexpressed in human cancers. Up-regulation of Pin1 elevates the expression of Cyclin D1, and plays an important role in tumorigenesis and tumor progression. This study was to investigate the expression and clinical significance of Pin1 and Cyclin D1 in cervical cancer cell lines and cervical epithelial tissues.. The expression of Pin1 and Cyclin D1 in cervical cancer cell lines HeLa, SiHa, C33a and Caski were detected by reverse transcription-polymerase chain reaction (RT-PCR) and Western blot. Their expression in 88 samples of cervical tissues, including 10 samples of normal cervix, 21 samples of cervical intraepithelial neoplasia (CIN), and 57 samples of invasive cervical cancer, were detected by immunohistochemistry.. The mRNA and protein levels of Pin1 were significantly higher in HeLa, SiHa, C33a, and Caski cells than in normal cervical epithelial tissues (P<0.05). The expression of Pin1 increased progressively along with the disease process from normal cervix to CIN, and to invasive cervical cancer (0%, 47.62%, 64.91%, P<0.05). Pin1 expression had no relation to disease stage (FIGO), pathologic grade, and pelvic lymph node metastasis status (P>0.05). The positive rate of Pin1 was significantly higher in cervical adenocarcinoma than in cervical squamous cell carcinoma (100% vs. 60.0%, P<0.05). In cervical cancer tissues, the overexpression of Pin1 was positively correlated to that of Cyclin D1 (P<0.05).. Pin1 is overexpressed in HeLa, SiHa, C33a and Caski cell lines as well as in cervical cancer tissues. The overexpression of Pin1 is closely related to Cyclin D1 expression in cervical cancer. The aberrant expression of Pin1 and Cyclin D1 might contribute to tumorigenesis of cervical cancer.

Topics: Adenocarcinoma; Adult; Carcinoma, Squamous Cell; Cell Line, Tumor; Cervix Uteri; Cyclin D1; Female; HeLa Cells; Humans; Lymphatic Metastasis; Middle Aged; Neoplasm Staging; NIMA-Interacting Peptidylprolyl Isomerase; Peptidylprolyl Isomerase; RNA, Messenger; Uterine Cervical Dysplasia; Uterine Cervical Neoplasms

Cyclins are a family of regulatory proteins that play a key role in controlling the cell cycle. Abnormalities of cell cycle regulators, including cyclins and cyclin dependent kinases, have been reported in various malignant tumors. This study was undertaken to quantitatively detect cyclin B1 and D1 in cervical cancer.. A quantitative real-time reverse transcription polymerase chain reaction and Western blot assay were used to analyze the expression of cyclin B1/D1 mRNA and proteins, respectively, in fresh invasive cervical cancer (n = 41) and normal cervical tissues (n = 10).. There was significantly greater cyclin B1 expression in invasive cervical cancer than in normal cervical tissue (P = 0.019). However, cyclin D1 expression was not significantly different. A Western blot assay yielded similar results.. Our results were consistent with the concept that up-regulation of cyclin B1 expression occurred in cervical cancer and an aberrant expression of cyclin B1 might play an important role in cervical carcinogenesis.

Topics: Blotting, Western; Cervix Uteri; Cyclin B; Cyclin B1; Cyclin D1; DNA, Complementary; Female; Gene Expression Profiling; Humans; Neoplasm Invasiveness; Neoplasm Staging; Reference Values; Reverse Transcriptase Polymerase Chain Reaction; RNA, Neoplasm; Uterine Cervical Neoplasms

Small cell carcinomas (SmCCs) of the uterine cervix are rare tumors. The knowledge regarding protein expression of several checkpoint candidates of cell cycle regulation is limited. Surgically treated SmCCs were selected from our files for immunohistochemical staining (neuroendocrine markers, p53, p16, p14, and cyclin D1). Polymerase chain reaction analysis, using general primers, was performed for human papillomavirus analysis. Nine of 677 tumors (1.3%) were classified as SmCCs after Grimelius staining (8/9 tumors positive) and immunohistochemical reaction against neurone-specific enolase, chromogranin A, synaptophysin (7/9 positive tumors), and CD 56 (8/9 positive tumors). All specimens were positive for at least two of the above. Two SmCCs were p53 positive and one case was p14 positive. Cyclin D1 staining was completely negative. All cases showed strong nuclear and/or cytoplasmic p16-immunostaining. Seven tumors represented human papillomavirus positivity for high-risk types. Four patients died of the tumor after a median time of 36.7 months (range, 15-56 months), representing a 5-year survival rate of 56%. The results suggest that p16 is up-regulated or accumulated in the SmCCs of the uterine cervix, probably caused by infection with human papillomavirus. p14 inactivation is of high prevalence in SmCCs and detection rate of p53 is similar to other histologic types of cervical carcinomas.

Topics: Adult; Carcinoma, Small Cell; Cyclin D1; Cyclin-Dependent Kinase Inhibitor p16; Female; Gene Expression Regulation, Neoplastic; Humans; Immunohistochemistry; Middle Aged; Papillomaviridae; Papillomavirus Infections; Polymerase Chain Reaction; Survival Rate; Tumor Suppressor Protein p14ARF; Tumor Suppressor Protein p53; Tumor Suppressor Proteins; Uterine Cervical Neoplasms

To investigate the patterns of expression of key cell cycle proteins targeting on the human papillomavirus (HPV) sites E6 and E7 in cervical carcinoma.. Semiquantitative immunohistochemistry was used to determine the expression of the cell cycle regulatory factor p53, retinoblastoma gene product pRb, cyclin-dependent kinase inhibitors p21(CIP1/WAF1) (p21), p16(INK4A) (p21), and p27(KIP1) (p21), and cyclin D1 targeting on the HPV sites E6 and E7 in 73 HPV 16-positive specimens of cervical squamous cell carcinoma, 35 from Australia patients and 38 age, lymph node status, and grade of tumor-matched Chinese patients.. There were no significant differences in age, lymph node status, and grade of tumor between the Chinese and Australian groups, however, the number of the patients in advanced stage (>Stage IIa) was greater in the Chinese group than in the Australian group (19:7). The positive rate of p53 in the Australian group was 3%, significantly lower than that in the Chinese group (23%, P = 0.028). The upregulation rate of pRb, p21, and p27 in the Australian group were 56%, 63% and 34% respectively, all significantly higher than those in the Chinese group (89%, 97%, and 89% respectively, P = 0.01, P < 0.001, and P < 0.001). There were no significant differences in the upregulation rate of p16 and cyclin D1 expression rate between the Australian group and Chinese group (97% versus 100%, and 3% versus 12%, both P > 0.05). In the combined data of both groups, the positive rate of p53 was 13%; and the upregulation rates of pRb, p16, p21, p27, and cyclin D1 were 74%, 99%, 81%, 63% and 7% respectively.. Cervical carcinoma overexpresses pRb, p16, p21, and p27. Tumors from Chinese patients are significantly more likely to be positive in p53 and to have upregulated pRb, p21, and p27 than their Australian counterparts. The molecular pathways to human papillomavirus-induced cervical cancer may be influenced by ethnicity and further investigation is necessary.

Topics: Australia; Biomarkers; Carcinoma, Squamous Cell; Cell Cycle Proteins; China; Cyclin D1; Cyclin-Dependent Kinase Inhibitor p21; Cyclin-Dependent Kinase Inhibitor p27; Female; Human papillomavirus 16; Humans; Immunohistochemistry; Oncogene Proteins, Viral; Papillomavirus E7 Proteins; Repressor Proteins; Retinoblastoma Protein; Tumor Suppressor Protein p53; Uterine Cervical Neoplasms

The prolyl isomerase Pin1, which specifically catalyzes conformational changes in certain proline-directed phosphorylation sites, is thought to be a critical catalyst for multiple oncogenic pathways. However, little is known about the role of Pin1 in human cervical cancer. Our previous study showed that Pin1 was overexpressed in cervical cancer tissues as well as cell lines. In this study, whether Pin1 is involved in cervical oncogenesis by regulating cyclin D1 was explored and the potential of Pin1-targeted gene silencing in inhibiting cellular growth and tumorigenicity in cervical cancer was investigated. A Pin1-directed shRNA and a sense Pin1 plasmid were constructed, and then the effects of the shRNA and the sense plasmid on HeLa cells were evaluated. The results showed that Pin1 directly regulated cyclin D1 levels. In addition, silencing Pin1 with RNAi significantly reduced cancer cell proliferation, colony formation, and strongly enhanced the apoptosis of HeLa cells. It is suggested that Pin1 may contribute to cervical tumorigenesis by regulating cyclin D1 expression and Pin1 may serve as a promising molecular target for diagnostics and therapeutics in cervical cancer.

Topics: Apoptosis; Cell Proliferation; Colony-Forming Units Assay; Cyclin D1; Female; Gene Expression Regulation, Neoplastic; HeLa Cells; Humans; NIMA-Interacting Peptidylprolyl Isomerase; Peptidylprolyl Isomerase; RNA, Small Interfering; Uterine Cervical Neoplasms

Interaction of human papilloma virus oncoproteins E6 and E7 with cell cycle proteins leads to disturbances of the cell cycle mechanism and subsequent alteration in the expression of some proteins, such as p16INK4a, cyclin D1, p53 and KI67. In this study, we compared alterations in the expression of these proteins during several stages of intraepithelial cervical carcinogenesis. Accordingly, an immunohistochemical study was performed on 50 cervical biopsies, including negative cases and intraepithelial neoplasias. The expression patterns of these markers were correlated with the histopathological diagnosis and infection with HPV. The p16INK4a, followed by Ki67, showed better correlation with cancer progression than p53 and cyclin D1, which recommends their use in the evaluation of cervical carcinogenesis. These monoclonal antibodies can be applied to cervical biopsy specimens to identify lesions transformed by oncogenic HPV, separating CIN 1 (p16INK4a positive) and identifying high-grade lesions by an increase in the cellular proliferation index (Ki67). In this way, we propose immunomarkers that can be applied in clinical practice to separate patients who need a conservative therapeutic approach from those who require a more aggressive treatment.

Topics: Adult; Aged; Aged, 80 and over; Biomarkers, Tumor; Biopsy; Cell Cycle Proteins; Cell Proliferation; Cervix Uteri; Cyclin D1; Cyclin-Dependent Kinase Inhibitor p16; Female; Humans; Ki-67 Antigen; Middle Aged; Tumor Suppressor Protein p53; Uterine Cervical Dysplasia; Uterine Cervical Neoplasms

Topics: Carcinoma, Small Cell; Cyclin D1; Cyclin-Dependent Kinase Inhibitor p16; Female; Humans; Papillomaviridae; Retinoblastoma Protein; Tumor Suppressor Protein p14ARF; Uterine Cervical Neoplasms

To investigate the effect of human papillomavirus 16 E7 gene on cell cycle of cervical cancer HeLa cell, through construction and expression of human papillomavirus 16 E7 gene with adenovirus vector.. Recombinant adenovirus which expressed E7 gene was constructed and packed. Flow cytometry (FCM) was used to detect the changes of cell cycle phase and cyclin D1 between cells infected and uninfected by recombinant adenovirus. The 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphennyltetrazolium bromide (MTT) colorimetric assay was used in the detection of the alteration of cell growth.. The recombinant adenovirus had stable efficiency of infection and E7 genes could be expressed stably. The result of MTT showed the multiplication of HeLa cells was accelerated from 0.27 +/- 0.03 before infection to 0.38 +/- 0.02 after infection (P < 0.01). FCM showed the number of cells in S phase increased from (26.0 +/- 0.4)% to (36.0 +/- 2.0)% at 12 hours and (49.9 +/- 4.2)% at 24 hours after infection (P < 0.05). Cyclin D1 expression was 22.4% before infection, and decreased to 55.2% after infection (P < 0.01).. The recombinant adenovirus expressing E7 gene could infect target cells. E7 gene can influence cell-cycle of HeLa cells, which can be used to restrain cervical cancer.

Topics: Adenoviridae; Cell Cycle; Cell Proliferation; Cyclin D1; Female; Genes, Viral; Genetic Vectors; HeLa Cells; Humans; Oncogene Proteins, Viral; Papillomavirus E7 Proteins; Plasmids; Transfection; Uterine Cervical Neoplasms

Electrochemical treatment is among the most effective therapies in the management of cervical malignancy. However, the mechanism of action of this treatment remains largely unknown. Therefore, the purpose of the current project was to establish a suitable eletrochemotherapy regimen for cervical cancer and to investigate the mechanism of the therapy in an in vitro model for human cervical carcinoma. HeLa cells were used as a model for cervical cancer in this study, and the effect of electrochemical treatment on these cells was examined in four different dosage groups (5 V + 5 C, 10 V + 5 C, 5 V + 10 C and 10 V + 10 C). Our results showed that the combinations of lower voltage and higher current (5 V/10 V + 10 C) had a greater anticancer effect in this model as compared to other groups. In addition, we compared the cytotoxic effect between electrochemical treatment and different pH condition treatments in this system, and found that the efficacy of electrochemical treatment in cell killing was better than that of acidic or basic medium treatment. Moreover, we demonstrated that the efficacy of electrochemical treatment was correlated with the degree of ionization and alteration in pH scale. The electrodes were basic on the cathode side which elevated the cations K+, Ca2+ and Mg2+, while the electrodes were acidic on the anode side which reduced the anion Cl-. We also assessed the effect of electrochemical treatment on cell cycle distribution in HeLa cells and showed that the percentage of cells in the G1 phase of the cell cycle was increased (G1 arrest), while the cell population in the S phase was decreased. Furthermore, we demonstrated that the levels of the cell cycle regulator cyclin D1 expression were dramatically reduced when 5 V/10 V + 10 C treatments were applied to these cells, as determined by RT-PCR analysis. By contrast, no significant changes in the levels of cyclin B1, CDK1 or CDK4 were detected. Based on these observations, we conclude that the combination of lower voltage and higher current may be a potentially effective eletrochemotherapy regimen for cervical cancer in the clinic, and that the antitumor effect of electrochemical treatment on cervical carcinoma cells is mediated partly via regulating ionization degree, pH state and cell cycle control.

Topics: Carcinoma; Cations; Cell Cycle; Cell Death; Cyclin D1; Electric Stimulation Therapy; Electrochemistry; Electrodes; Female; Flow Cytometry; Gene Expression Profiling; HeLa Cells; Humans; Hydrogen-Ion Concentration; Reverse Transcriptase Polymerase Chain Reaction; Uterine Cervical Neoplasms

Though many investigators have reported relationships between the CCND1 polymorphism and susceptibility to various carcinomas, to our knowledge, no report has been issued concerning its relationship with uterine cervical cancer. Thus, we undertook this study to investigate the association between CCND1 polymorphisms and susceptibility to cervical cancer in Korean women. This study was carried on 222 patients with squamous cell carcinoma of uterine cervix and on 314 normal controls. CCND1 genotyping was determined by polymerase chain reaction and restriction fragment length polymorphism. The allelic frequencies of the cases (A, 0.53; G, 0.47) were not significantly different from those of the controls (A, 0.49; G, 0.51) (P=0.238). Regression analysis after adjusting for age showed that the CCND1 G870A genotypes are not related to the risk of squamous cell carcinoma of the uterine cervix. Our findings suggest that the CCND1 polymorphism is not associated with an increased risk of squamous cell carcinoma of uterine cervix in Korean women.

Topics: Adult; Aged; Carcinoma, Squamous Cell; Case-Control Studies; Cyclin D1; Female; Genetic Predisposition to Disease; Humans; Korea; Middle Aged; Polymorphism, Genetic; Uterine Cervical Neoplasms

The implications of Survivin, CyclinD1, p21(WAF1), Caspase-3 in the development, progression and prognosis in cervical cancer were investigated. By using immunohistochemical SP method, the expression of Survivin, CyclinD1, p21(WAF1), Caspase-3 was detected in 41 cases of cervical cancer, 17 cases of cervical intraepithelial neoplasia (CIN) and 10 cases of normal tissues, and their relation with pathological grade, clinical stage, metastasis and survival time was analyzed. The results showed that the positive expression rate of Survivin, CyclinDl in cervical cancer was significantly higher than in CIN group and normal control group (P < 0.05). The median survival time in the patients with cervical cancer positive for Survivin and CyclinD1 was significantly shorter than in those with negative expression (P < 0.05). The expression of both Survivin and CyclinD1 was not related with tumor grade, clinical stage and metastasis (P > 0.05). The positive expression rate of p21(WAF1) , Caspase-3 in cervical cancer was significantly lower than in CIN group and normal control group (P < 0.05), and had a close relation with tumor grade (P <0.05). The expression of Survivin in cervical cancer in cervical cancer was negatively associated with that of Caspase-3 (P < 0.01), but positively with that of CyclinD1 (P < 0.01). Cox Multivariate analysis revealed that Survivin was the independent prognostic indicator influencing the survival time of the patients with cervical cancer (P < 0.05). It was suggested that the high expression of Survivin or CyclinD1, and low expression of p21(WAF1) or Caspase-3 was closely correlated with the development of cervical cancer. Survivin and CyclinD1 could be used as a useful indicator to predict the prognosis of cervical cancer.

Topics: Adenocarcinoma; Adult; Carcinoma, Squamous Cell; Caspase 3; Caspases; Cyclin D1; Cyclin-Dependent Kinase Inhibitor p21; Female; Humans; Inhibitor of Apoptosis Proteins; Microtubule-Associated Proteins; Middle Aged; Neoplasm Proteins; Prognosis; Survivin; Uterine Cervical Dysplasia; Uterine Cervical Neoplasms

Human papillomavirus (HPV) plays a major role in the etiology of cervical cancer. However, a complex correlation between viral and cellular genes is necessary for cell cycle control deregulation in the progression to invasive cervical cancer (ICC). Cyclin D1 (CCND1) is an important positive regulator of the G1/S phase of the cell cycle. The CCND1 gene is located at 11q13 and is often altered in human cancers. We analyzed the A870G CCND1 polymorphism by polymerase chain reaction/restriction fragment length polymorphism analysis in 246 women including 50 cases with high-grade squamous intraepithelial lesions of the cervix (HSIL), 93 with ICC, and 103 healthy women. The GG genotype was associated with a 4.32-fold higher risk for the development of HSIL [adjusted odds ratio (aOR)=4.32, 95% confidence interval (CI) 1.50-12.46, P=0.0067), and a 3.26-fold increased risk for the development of ICC (aOR=3.26, 95% CI 1.42-7.53, P=0.006). The proportion of cervical cancer cases attributable to the GG CCND1 genotype was 17.26%. This study indicates that the A870G CCND1 polymorphism could act as a cofactor of HPV in the initiation of cervical carcinogenesis, particularly in the transformation zone of HPV-infected women, supporting evidence for a genetic factor in ICC risk.

Topics: Adult; Aged; Cyclin D1; Female; Genotype; Humans; Middle Aged; Papillomaviridae; Papillomavirus Infections; Polymorphism, Genetic; Risk; Uterine Cervical Neoplasms

Most cervical high-grade squamous intraepithelial lesions (HSILs) persist, but approximately one third regress (ie, no HSIL in follow-up biopsies). To identify factors related to histologic proven persistence or regression. Twenty-eight small histologic (marker) biopsies with adequate follow-up were analyzed for human papillomavirus (HPV) genotypes and different immunoquantitative proliferation, cell cycle regulation, and differentiation markers. All cases had a biopsy-interval between the marker and first follow-up biopsy of at least 100 days (median, 8.2 months; range, 3.4-22.5 months). Follow-up was classified as regression or persistence. All lesions were high-risk (hr) HPV and p16 positive, 63% for HPV-16 or HPV-16 mixed with other hr genotypes, while 37% had other hrHPV types. The marker biopsies of the persistent HSILs had lower p53 and retinoblastoma protein (pRb) detected in the deep half of the epithelium (P = 0.001 and 0.02, respectively) than nonpersistent HSILs. The degree of positivity of p16, Ki-67, cyclin D1, lesion extent, positivity of the resection margins, and patient age were all unrelated to persistence or regression. Lesions with HPV-16 or mixed-16 genotypes had a significantly lower percentage of pRb (P = 0.02), p53 (P = 0.02), and cyclin D (P = 0.04) positive nuclei in the deep epithelial layers. In agreement with this, type-16 positive HSILs had a lower regression percentage than those with other HPV types, but the difference was not significant. HSILs with combined negativity/low positivity for p53 and pRb protein in small histologic biopsies are highly likely to persist, contrasting those in which one of these cell cycle regulators is strongly positive (p53 > 15%; pRb > 40%).

Topics: Adult; Biomarkers, Tumor; Biopsy; Carcinoma in Situ; Cyclin D1; Cyclin-Dependent Kinase Inhibitor p16; Epithelium; Female; Follow-Up Studies; Humans; Ki-67 Antigen; Papillomaviridae; Retinoblastoma Protein; Tumor Suppressor Protein p53; Uterine Cervical Neoplasms

Epidemiologic studies have implicated estrogenic exposure as well as human papilloma virus (HPV) infection in cervical carcinogenesis, and some studies have suggested that estrogen and HPV may play synergistic roles in cervical tumorigenesis. In this study, we report a novel finding that approximately 35% of cervical carcinomas tested (n = 19) express aromatase, the enzyme responsible for converting androgen to estrogen, the rate-limiting and final step in estrogen biosynthesis. On the other hand, no aromatase expression was detected in precancerous (n = 42) or normal cervical (n = 17) tissue samples. Increased aromatase was associated with increases in estrogen receptors (ER-alpha and ER-beta) and a decrease in progesterone receptor levels, suggesting that in situ estrogen signaling via ER may be involved in tumor growth. Stable overexpression of aromatase in HPV+ cervical cancer cells resulted in increased cellular proliferation, anchorage-independent growth, and ER expression and activity. In contrast, little change in ER was observed in HPV- cells. Steroid hormone receptor expression observed in vitro paralleled that seen in cervical carcinomas expressing aromatase. Aromatase overexpression also induced the expression of cyclin D1, proliferating cell nuclear antigen, and the HPV oncogenes, E6 and E7. Furthermore, the data underscores the importance of steroid receptor (estrogen and progesterone receptors) regulation in cervical carcinogenesis. To our knowledge, this is the first report demonstrating the induction of aromatase expression in cervical carcinomas, and opens the possibility that aromatase inhibitors may be potential therapeutic agents in cervical carcinomas expressing aromatase.

Topics: Aromatase; Cell Growth Processes; Cell Line, Tumor; Cervix Uteri; Cyclin D1; Enzyme Induction; Estrogen Receptor alpha; Estrogen Receptor beta; Estrogens; Female; Humans; Neoplastic Stem Cells; Papillomaviridae; Papillomavirus E7 Proteins; Papillomavirus Infections; Proliferating Cell Nuclear Antigen; Receptors, Progesterone; Uterine Cervical Neoplasms

High-risk human papillomaviruses are the causative agents of cervical cancer and are also believed to be aetiologically involved in a subset of squamous cell carcinomas of the head and neck region, especially the tonsil. Cervical cancers arise through disruption of the pathways of p53 and the product of the retinoblastoma gene by the human papillomavirus oncoproteins E6 and E7. It is generally assumed that the same pathways are involved in human papillomavirus-induced carcinogenesis at other mucosal surfaces. However, the patterns of expression of cell cycle proteins targeted by human papillomavirus E6 and E7 in cancers from different anatomic sites have been inconsistent, due to either biologic or technological factors. In this study, 73 human papillomavirus, 16-positive cervical squamous cell carcinomas (35 from Australian and 38 from Chinese women) were analysed for the expression of p53, pRb, p16(INK4A), p21(CIP1/WAF1), p27(KIP1) and cyclin D1 by semiquantitative immunohistochemistry. Cervical cancers from Chinese women were found to be significantly more likely to overexpress p53, pRb, p21 and p27 than their Australian counterparts. These findings were compared with those from 31 human papillomavirus 16-positive tonsillar squamous cell carcinomas, all of Australian origin, tested using the same methodology. Comparisons of the tonsillar and combined cervical data showed that tonsillar cancers were significantly more likely to be p53-positive, whereas cervical cancers were significantly more likely to overexpress pRb, p16 and p27. When the tonsillar data were compared with cervical data from Australian women, the associations for p53 and pRb remained. These findings represent new evidence that the molecular pathways to human papillomavirus-induced mucosal cancer may be influenced by anatomic location and ethnicity.

Topics: Adult; Aged; Aged, 80 and over; Australia; Carcinoma, Squamous Cell; Cell Cycle Proteins; China; Cyclin D1; Cyclin-Dependent Kinase Inhibitor p16; Cyclin-Dependent Kinase Inhibitor p21; Cyclin-Dependent Kinase Inhibitor p27; Female; Humans; Immunohistochemistry; Middle Aged; Neoplasm Staging; Papillomaviridae; Papillomavirus Infections; Retinoblastoma Protein; Tonsillar Neoplasms; Tumor Suppressor Protein p53; Tumor Suppressor Proteins; Uterine Cervical Neoplasms

Because the incidence of adenocarcinoma of the uterine cervix has recently risen, the evaluation of radiotherapy (RT) for this disease has become an increasingly urgent matter. We analyzed the expression of the cell cycle-associated proteins p53, p27, p21/waf1/cip1, and cyclin D1 in cervical adenocarcinomas in correlation with the prognostic significance in tumors treated with RT alone.. The expression of p53, p27, p21/waf1/cip1, and cyclin D1 was studied using an immunohistochemical method in 53 cases of cervical adenocarcinoma treated only with RT. Patients received RT alone between 1965 and 1994. The mean patient age was 61.8 +/- 12.6 years (range, 36-82 years). The number of patients with Stage I, II, III, and IVA disease was 6, 16, 28, and 3, respectively.. The number of patients with p53, p27, p21/waf1/cip1, and cyclin D1 positive tumors was 24, 18, 22, and 8, respectively; no statistically significant correlation was noted. The 5-year disease-free survival rate of p53-positive patients was 30%, significantly lower than the 62% for the p53-negative patients (p = 0.02); no statistically significant correlation was noted between disease-free survival and p27, p21/waf1/cip1, and cyclin D1 expression. No statistically significant correlation was observed between local control and expression of any of the proteins.. Expression of p53 protein has a statistically significant impact on disease-free survival in adenocarcinoma of the uterine cervix treated with RT alone. However, the clinical significance of p27, p21/waf1/cip1, and cyclin D1 protein expression was not obvious.

Topics: Adenocarcinoma; Adult; Aged; Aged, 80 and over; Carcinoma, Adenosquamous; Cell Cycle Proteins; Cyclin D1; Cyclin-Dependent Kinase Inhibitor p21; Cyclin-Dependent Kinase Inhibitor p27; Cyclins; Female; Humans; Immunohistochemistry; Middle Aged; Multivariate Analysis; Neoplasm Proteins; Survival Rate; Tumor Suppressor Protein p53; Tumor Suppressor Proteins; Uterine Cervical Neoplasms

Cervical cancer is one of the most common cancers affecting a woman's reproductive organs. Despite its frequency and recurrence, the death rate has been declining over the past 40 years, due to early detection and treatment. In a previous report [Shehata Marlene, Shehata Marian, Shehata Fady, Pater Alan. Apoptosis effects of Xrel3 c-Rel/Nuclear factor-kappa B homolog in human cervical cancer cells. Cell Biology International, in press], we studied the role of the NF-kappaB gene family in HeLa human cervical cancer cells, using the Xrel3 c-Rel homologue of Xenopus laevis. These results showed that the expression of Xrel3/c-Rel slowed cell growth, consistent with an upregulated expression of the cell cycle inhibitor p21 and the activated poly(ADP-ribose) polymerase (PARP) apoptosis effector. However, in this report, we examined more apoptotic and anti-apoptotic factors acting upstream and downstream in apoptosis pathways after cisplatin treatment of HeLa cervical cancer cells. After 1 microM cisplatin treatment, Xrel3 had an anti-apoptotic effect, based on significantly lower levels of apoptotic proteins, including caspase-8, caspase-3 and p21. Anti-apoptotic BAG-1 isoforms were upregulated. After 5 microM cisplatin treatment, expression of HeLa Xrel3 had an apoptotic effect, based on significantly increased expression of the cell cycle inhibitor p21 and apoptotic proteins, including cleaved PARP, caspase-8, and caspase-3. However, anti-apoptotic Bcl-2 and Bcl-X(L) were elevated and the cell cycle regulator cyclin D1 was slightly upregulated with both 1 and 5 microM cisplatin treatment. The HPV E6 oncoprotein showed no significant changes. These results support previous conclusions on the potential anti-apoptotic effects of c-Rel/NF-kappaB in mild stress environments, as opposed to the apoptotic effects associated with high stress conditions [Lake BB, Ford R, Kao KR. Xrel3 is required for head development in Xenopus laevis. Development 2001; 128(2), 263-73.]. Thus, c-Rel/NF-kappaB may potentially be of clinical significance in chemotherapy.

Topics: Antineoplastic Agents; Apoptosis; bcl-X Protein; Carcinoma; Carrier Proteins; Caspases; Cell Cycle Proteins; Cell Proliferation; Cisplatin; Cyclin D1; Cyclin-Dependent Kinase Inhibitor p21; DNA Damage; DNA-Binding Proteins; Dose-Response Relationship, Drug; Female; HeLa Cells; Humans; NF-kappa B; Poly(ADP-ribose) Polymerases; Proto-Oncogene Proteins c-bcl-2; Proto-Oncogene Proteins c-rel; Transcription Factors; Up-Regulation; Uterine Cervical Neoplasms; Xenopus Proteins

In tumorigenesis, loss of function of the G1 pathway (p16-CDK4/cyclinD1-pRB pathway (RB pathway) and p14-MDM2-p53 pathway (p53 pathway)) is a theoretically essential event. The simultaneous analysis of all components of the RB and p53 pathway may be able to explain cervical tumorigenesis. However, there are no reports in which all components of the G1 pathway and HPV typing were examined simultaneously in cervical cancer.. We examined HPV typing and the status of the G1 pathways simultaneously by PCR-SSCP, multiplex PCR, methylation-specific PCR, and immunohistochemical techniques in cervical neoplasia. A total of 105 samples (normal, 10; cervical intraepithelial neoplasm (CIN), 42; invasive cancer (IC), 53) were included.. Abnormality of the RB pathway tended to be more frequent in ICs (60.4%) than in CINs (31.0%) (P = 0.069). The primary target was p16 (CIN, 14.3%; IC, 43.4%; P = 0.032). Abnormality of the p53 pathway was detected in ICs (56.6%) and in CINs (40.5%) (P = 0.1494). In particular, strong expression of MDM2 was higher in ICs (32.1%) than in CINs (7.1%) (P = 0.0045). Abnormalities of the RB and p53 pathways were higher in low-risk and negative HPV than in high-risk HPV (81.3% vs 51.4%, P = 0.0657; 81.3% vs 45.9%, P = 0.0328). Seven HPV-negative cases had abnormalities in the RB or p53 pathways.. In conclusion, abnormality of the G1 pathway may be one of the important mechanisms for carcinogenesis of low-risk and negative HPV cases.

Topics: Cyclin D1; Cyclin-Dependent Kinase 4; Cyclin-Dependent Kinase Inhibitor p16; Cyclin-Dependent Kinases; Female; G1 Phase; Humans; Immunohistochemistry; Papillomaviridae; Polymerase Chain Reaction; Polymorphism, Single-Stranded Conformational; Proto-Oncogene Proteins; Retinoblastoma Protein; Tumor Suppressor Protein p53; Uterine Cervical Dysplasia; Uterine Cervical Neoplasms

We have studied the effects of purvalanol A on the cell cycle progression, proliferation and viability. In synchronized cells, purvalanol A induced a reversible arrest the progression in G1 and G2 phase of the cell cycle, but did not prevent the completion of DNA synthesis in S-phase cells. The specificity of action of the drug was supported by the selective inhibition of the phosphorylation of cyclin-dependent kinase (cdk) substrates such as Rb and cyclin E. The cell contents of cyclins D1 and E were lower in cells incubated with purvalanol A compared to controls, but the level of the cdk inhibitory protein p21(WAF1/CIP1) was increased, indicating that the drug did not cause a general inhibition of gene expression. Purvalanol A did not inhibit transcription under cell-free conditions. This compound, however, caused an inhibition of the estradiol-induced expression of an integrated luciferase gene, suggesting that cdk or related enzymes may participate in the regulation of the activity of certain promoters. When exponentially growing cells, both mouse fibroblasts and human cancer cell lines, were incubated with purvalanol A for prolonged periods of time (24 hr), a lasting inhibition of cell proliferation as well as cell death were observed. In contrast, a 24 hr incubation of quiescent (non-transformed) cells with purvalanol A did not prevent their resumption of cell cycle after removal of the drug.

Topics: Breast Neoplasms; Cell Cycle; Cell Division; Cell Survival; Colonic Neoplasms; Cyclin D1; Cyclin E; Cyclin-Dependent Kinase Inhibitor p21; Cyclin-Dependent Kinases; Cyclins; Enzyme Inhibitors; Female; HeLa Cells; HT29 Cells; Humans; Methionine; Microfilament Proteins; Muscle Proteins; Phosphorylation; Retinoblastoma Protein; Transcription, Genetic; Tumor Cells, Cultured; Uterine Cervical Neoplasms

The purpose of the present study was to analyse clinical correlation between HPV type 16 and 18 infection, expression of p53, cyclin D1, Ki-67, c-erbB2 and EGFr gene products in cervical cancer cells as well as their nuclear ploidy. The morphological parameters evaluated, such as differentiation of carcinomas, vascular invasion, ploidy and expression of oncogenic proteins, indicate the increased biological malignancy of HPV 16/18-positive carcinomas. The majority of them were poorly differentiated, revealed significantly higher frequence of vascular invasion (p<0.05), were more frequently aneuploid and showed overexpression of cyclin D1. The comparison of the data obtained with the mortality rate of the patients suggests that the overexpression of EGFr and moderate expression of Ki-67 seem to be unfavorable prognostic factors, regardless of the presence of HPV 16/18.

Topics: Biomarkers, Tumor; Carcinoma; Causality; Cell Differentiation; Cell Nucleus; Cyclin D1; Disease Progression; DNA; ErbB Receptors; Female; Humans; Immunohistochemistry; Ki-67 Antigen; Neoplasm Invasiveness; Papillomaviridae; Ploidies; Prognosis; Receptor, ErbB-2; Survival Rate; Tumor Suppressor Protein p53; Uterine Cervical Neoplasms

Amplification and overexpression of cyclin D1 and CDK4 genes in cervical carcinoma were studied by semi-quantitative differential polymerase chain reaction assay and an immunostaining technique, respectively. Amplifications of cyclin D1 and CDK4 genes were found in 24% (27/113) and 26% (29/112) of tumors, respectively. Overexpression of cyclin D1 and CDK4 was demonstrated in 32% (21/66) and 73% (45/62) of tumors, respectively. No tumor showed CDK4 gene mutation on single strand conformational polymorphism. Sixteen percent (8/49) of the tumor specimens showed neither amplification nor overexpression. Disease stage, tumor grade and overexpression of cyclin D1 were found to be independent poor prognostic factors in cervical carcinoma.

Topics: Adult; Aged; Aged, 80 and over; Cyclin D1; Cyclin-Dependent Kinases; Female; Gene Amplification; Genes, Retinoblastoma; Humans; Immunohistochemistry; Middle Aged; Neoplasm Staging; Prognosis; Survival Analysis; Uterine Cervical Neoplasms

Many investigators have studied the expression of G1 phase regulatory protein in uterine cervical cancer. However, it is unclear which step of the genetic expression participates in cyclin D1 expression and what its prognostic meaning is. The aims of this study were to evaluate the regulatory level of cyclin D1 expression and the relationship between the expression of cyclin D1 and its inhibitor, p21WAF1/CIP1, and to evaluate their impact on the prognosis of early stage cervical cancer.. The presence of cyclin D1 mRNA was studied using Northern blot in 6 normal cervices and 7 invasive cervical cancer specimens. Western blot was used to detect the cyclin D1 protein in 8 normal cervices and 8 invasive cancer specimens. Thirty-two cases of FIGO stage Ib-IIa cervical cancers (28 squamous cell carcinomas, 3 adenocarcinomas, 1 adenosquamous cell carcinoma), 31 cases of cervical intraepithelial neoplasia 3 (CIN 3), and 28 normal cervices were stained for cyclin D1 and p21(WAF1/CIP1) using monoclonal antibody. Statistical analysis was performed to assess the differences in expression and their prognostic significance. RESULTS. Cyclin D1 mRNA was found to be underexpressed in cervical cancer. Western blot also revealed underexpression of cyclin D1 protein in cervical cancer compared to normal controls. Positive immunohistochemical staining of cyclin D1 was noted in 28/28 (100%) of the normal controls, 1/31 (3%) cases of CIN 3, and 9/32 (28%) cases of invasive cancer. The number of positively stained specimens was lower than that of normal controls in CIN 3 and cervical cancer specimens (P = 0.005). Fifteen of 28 (54%) normal controls, 15/31 cases (48%) of CIN 3, and 27/32 cases (84%) of invasive cancer were proved positive for p21WAF1/CIP1 immunohistochemistry. p21WAF1/CIP1 was more highly expressed in cervical cancer than in that of either normal controls or CIN specimens (P = 0.001). Positive immunostaining of cyclin D1 and p21WAF1/CIP1 was not related to high-risk factors (pelvic lymph node metastasis, deep cervical stromal invasion, parametrial invasion, large tumor size, and unusual histologic type) and human papilloma virus infection. Positive cyclin D1 immunostaining was associated with decreased disease-free survival and a lower overall survival (P = 0.0175 and 0.0189, respectively). On multivariate analysis, positive cyclin D1 expression was a significant prognostic variable for recurrence (P = 0.0004).. Underexpression of cyclin D1 was regulated at the level of transcription in cervical cancer. Although cyclin D1 was underexpressed in cervical neoplasias, it was more frequently expressed in malignant lesions. p21WAF1/CIP1 was more highly expressed in cervical cancers than in either normal cervices or CIN 3 specimens. Unfavorable prognoses were associated with cyclin D1 expression, and not with the expression of its inhibitor, p21WAF1/CIP1.We conclude that immunohistochemical assessment of cyclin D1 can be a useful molecular marker for predicting prognosis in early stage cervical cancer of the uterus.

Topics: Blotting, Northern; Blotting, Western; Cyclin D1; Cyclin-Dependent Kinase Inhibitor p21; Cyclins; Female; Gene Expression; Humans; Immunohistochemistry; Neoplasm Recurrence, Local; Neoplasm Staging; Papillomaviridae; Papillomavirus Infections; Prognosis; RNA, Messenger; Survival Analysis; Tumor Virus Infections; Uterine Cervical Dysplasia; Uterine Cervical Neoplasms

The aims of this study were to study aberrant expression and coexpression of the cell cycle associated proteins TP53, p21, p27, cyclin D1, cdk4, RB, EGFR, and MDM2 in cervical carcinomas, to correlate protein alterations with histopathological and clinical parameters, and to evaluate whether these alterations provide prognostic information.. Seventy-four cervical carcinomas and 10 cases of normal cervical epithelium from patients with benign uterine leiomyomas were investigated immunohistochemically for aberrant expression of the cell cycle associated proteins using the biotin-streptavidin-peroxidase method and the OptiMax Plus automated cell staining system.. In normal cervical epithelium p27 immunostaining was identified in more than 50% of the cells, cdk4 in 5-50% of the cells, and EGFR in less than 5% of the cells, whereas no immunostaining for TP53, p21, MDM2, or cyclin D1 was detected. Positive RB protein staining was identified in all cases of normal cervical epithelium. RB protein staining was also identified in all carcinomas of the cervix uteri. Overexpression of p21 was found in 96% of the tumors, MDM2 in 35%, cdk4 in 69%, cyclin D1 in 3%, and EGFR in 20% of the tumors. A low level of p27 was observed in 65% of the cases. In a previous study, the TP53 protein level has been found to be elevated in 41 of the 74 (55%) cases included in this work. Significant coexpression was seen for TP53 and MDM2 (P = 0. 001); concording results were observed in 67% of the cases. There was no difference in aberrant expression or coexpression of any of the cell cycle regulatory proteins related to histological type, grade of differentiation, FIGO stage, or relapse-free survival.. The high number of cases showing increased levels of p21 and cdk4 and decreased levels of p27 suggests that these proteins may be important in the pathogenesis of cervical carcinoma. Furthermore aberrant expression of MDM2 in a smaller but significant fraction of cases indicates that these proteins could also be involved in the development of these cancers. Finally our results indicate that MDM2 may protect against HPV-induced TP53 protein degradation.

Topics: Adult; Aged; Aged, 80 and over; Cell Cycle Proteins; Cyclin D1; Cyclin-Dependent Kinase 4; Cyclin-Dependent Kinases; ErbB Receptors; Female; Gene Expression Regulation, Neoplastic; Humans; Microfilament Proteins; Middle Aged; Muscle Proteins; Neoplasm Proteins; Nuclear Proteins; Oncogene Protein p21(ras); Proto-Oncogene Proteins; Proto-Oncogene Proteins c-mdm2; Retinoblastoma Protein; Tumor Suppressor Protein p53; Uterine Cervical Neoplasms

Cyclin D1 plays an essential regulatory role in the G1 phase of the cell cycle. The cyclin D1 gene is amplified in 20-50% of squamous cell carcinomas (SCCs), and the protein is overexpressed in up to 80% of SCCs. Our hypothesis was that gene transduction of antisense (AS) cyclin D1 in human SCCs in vivo would result in tumor reduction. A cyclin D1 cDNA was inserted into an E1/E3-deficient serotype 5 adenovirus (AS cyclin D1) in an AS orientation using homologous recombination. AS cyclin D1 transduction suppressed cyclin D1 protein expression in both cultured cells and tumors. AS cyclin D1 significantly inhibited cell proliferation by both [3H]thymidine incorporation in six SCC cell lines (P = 0.01-0.001) and the conversion of tetrazolium salt to formazan in four SCC cell lines (P = 0.01-0.004). Apoptosis detected in >25% of cells in each cell line 48 h after AS cyclin D1 transduction paralleled the reduction in cyclin D1 protein. Preformed SCCs transduced with AS cyclin D1 were significantly inhibited (P = 0.002-0.005), and apoptosis was prominent in the AS cyclin D1-treated tumors, but not in tumors treated with the control vector. These data extend prior in vitro and ex vivo results and indicate that AS cyclin D1 suppresses SCC growth both in vitro and in vivo through suppression of cyclin D1 protein expression, leading to cellular apoptosis. Our findings suggest that cyclin D1 may have a role in cell survival and that cyclin D1 AS therapy may be useful as an adjunct to standard treatment for SCC.

Topics: Adenoviruses, Human; Apoptosis; Carcinoma, Squamous Cell; Cell Cycle; Cell Division; Cyclin D1; DNA, Antisense; Female; Gene Amplification; Genetic Vectors; Head and Neck Neoplasms; Humans; Open Reading Frames; Skin Neoplasms; Tumor Cells, Cultured; Uterine Cervical Neoplasms; Vulvar Neoplasms

In this study, we have demonstrated the overexpression of cyclin D1 protein in 24 (92%) of 26 low-grade squamous intraepithelial lesions infected with low-risk human papillomaviruses (HPV 6, 11, 42, 43, and 44) and the absence of cyclin D1 expression in 25 (87%) of 29 lesions infected with high-risk HPVs (HPV 16, 18, 31, 33, 39, 45, 51, 52, 56, 58, and 66). The expression of cyclins E, A, and B proteins was increased in both low-risk and high-risk HPV infections. Tetrasomy of chromosomes 1, 3, 11, 17, 18, and X was present in nine lesions, all of which were infected with high-risk HPVs, but was not related to the pattern of cyclin expression. These data provide in vivo evidence that low- and high-risk HPV types alter cell cycle control by different mechanisms and that cell cycle checkpoint abnormalities are induced by high-risk, but not low-risk, HPV infection.

Topics: Cell Cycle; Chromosome Aberrations; Chromosome Disorders; Cyclin D1; Epithelium; Female; Humans; Papillomaviridae; Papillomavirus Infections; Tumor Virus Infections; Uterine Cervical Dysplasia; Uterine Cervical Neoplasms; Warts

G1 cyclin, as a candidate proto-oncogene, plays a key role as a cellular regulator through direct interaction with retinoblastoma gene product (pRB), p107, and cyclin-dependent kinase. Human papillomavirus (HPV) 16/18 E6/7 proteins may bind the unphosphorylated pRB and act as a down-regulator of cyclin D. However, this theory has not been proven and the relationship between cyclin E and HPV also remains unclear. Using formalin-fixed and paraffin-embedded cervical tissues, we examined for HPV types 16 and 18 by nested polymerase chain reaction (PCR), and for the accumulation of cyclin D1 and E by immunohistochemistry in 22 cases of normal cervix, 23 cases of cervical intraepithelial neoplasia (CIN), 39 cases of invasive squamous cell carcinoma (SCC), and 6 cases of adenocarcinoma. Cyclin index (CI) was defined as the percentage of positively labeled nuclei per 1000 cells. The accumulation of cyclin D1 was detected in the normal basal and parabasal epithelium and was markedly decreased in neoplastic epithelium (0.87% CI in CIN, 5.88% in SCC). However, cyclin E was absent in the normal cervix but increased in neoplastic epithelium (7.17% CI in CIN, 10.32% in SCC). Cyclin D1 expression was significantly low (p = 0.004), but cyclin E expression was significantly high (p = 0.026) in HPV-positive cases. Cyclin E plays a leading role in neoplastic transformation whereas cyclin D is down-regulated, especially when associated with HPV.

Topics: Adenocarcinoma; Carcinoma, Squamous Cell; Cervix Uteri; Cyclin D1; Cyclin E; Cyclin G; Cyclin G1; Cyclins; Female; Humans; Immunohistochemistry; Papillomaviridae; Polymerase Chain Reaction; Proto-Oncogene Mas; Uterine Cervical Dysplasia; Uterine Cervical Neoplasms

Amplification of chromosome 11q13 leads to overexpression of a G1 cyclin gene, cyclin D1 (PRAD-1, CCND-1), in many non-cervical human carcinomas. Homology between cyclin D1 and human papillomavirus oncoprotein E7 binding sites for the retinoblastoma tumor-suppressor protein suggests that human papillomavirus oncoproteins cyclin D1, and other cell cycle regulatory proteins may act through a common mechanism in the pathogenesis of human cervical squamous cell carcinoma. We have examined 48 cases of cervical neoplasia by RNA-mRNA in situ hybridization for human papillomavirus mRNA and cyclin D1 mRNA and by immunohistochemistry for cyclin D1 protein expression. Hybridization demonstrated human papillomavirus RNA in all 48 cases with types 6, 16, or 18 in 2, 26, and 20 cases, respectively. Immunohistochemical detection using anti-cyclin D1 rabbit polyclonal antibody 19 demonstrated appropriate cyclin D1 expression at constitutive low levels in normal squamous epithelium and low-grade intraepithelial lesions. Immunohistochemical staining failed to demonstrate significant protein expression in any of the high-grade or invasive lesions. In contrast to the immunohistochemical results, in situ hybridization demonstrated cyclin D1 mRNA overexpression in three of five cases of low-grade squamous intraepithelial lesion, one of eight cases of high-grade squamous intraepithelial lesion, 14 of 18 cases of invasive squamous cell carcinoma, two of five cases of adenocarcinoma in situ, one of seven cases of invasive adenocarcinoma, and two of five cases of small cell undifferentiated carcinoma. Transcriptional activation of cyclin D1 can occur in vivo in human papillomavirus-associated invasive cervical carcinoma, but it does not seem to result in a steady state, increased level of cyclin D1 protein expression. These data indicate a limited role for cyclin D1 protein in the pathogenesis of human papillomavirus-associated invasive cervical squamous carcinoma. They support a model in which human papillomavirus proteins can circumvent cellular requirements for cyclin D1 in human cervical neoplasia.

Topics: Carcinoma; Cyclin D1; Cyclins; Female; Gene Expression Regulation, Neoplastic; Humans; Immunohistochemistry; Oncogene Proteins; Uterine Cervical Neoplasms

CYCLIN D1, a cell-cycle control gene, recently has been shown to be identical to an oncogene alternatively known as BCL-1 and PRAD1 and implicated in centrocytic lymphomas and parathyroid adenomas, respectively. PRAD1 complexes to the product of the retinoblastoma (Rb) tumor suppressor gene, an event followed by Rb inactivation. Squamous cell carcinomas of the cervix and vulva are gynecologic tumors in which human papillomaviruses have been implicated as an initiating event, and proteins derived from these viruses also complex with an inactivate Rb. Because of the overlap in some of the molecular processes mediated by human papillomaviruses and by the PRAD1 oncogene, the authors analyzed the PRAD1 (CYCLIN D1/BCL-1) genomic structure and expression in vulvar and cervical squamous cell carcinoma cell lines.. PRAD1 DNA and PRAD1 mRNA expression were assessed by Southern and Northern blotting, respectively, in 13 squamous cell carcinoma cell lines of gynecologic origin (10, cervical cancer; 3, vulvar cancer).. We found low baseline levels of a 4.5-kb PRAD1 transcript in a series of control cell lines, which were derived from normal fibroblasts, various hematologic malignancies, and a choriocarcinoma. PRAD1 mRNA overexpression (> or = 10-fold greater than that in control lines) was seen in all three vulvar carcinoma cell lines, two of which also showed amplification (5-fold and > 10-fold) of PRAD1 genomic sequences. Abnormalities of PRAD1 also were seen in 4 of the 10 cervical cancer cell lines and included overexpression of PRAD1 transcripts (3-9-fold) in 3 lines and rearrangement of PRAD1 DNA in an additional line that, however, did not shown any aberration in PRAD1 mRNA as discernible by Northern blotting. PRAD1 abnormalities were observed in three of the four cervical cell lines derived from metastatic sites and in one of the six cervical lines derived from primary tissue.. Seven of 13 squamous cell lines of gynecologic origin showed abnormalities of PRAD1. These abnormalities included amplification and rearrangement of DNA and overexpression of mRNA. The role of PRAD1 as a cell-cycle regulatory gene and its interactions with the Rb tumor suppressor gene suggests that PRAD1 deregulation may be a significant molecular event in the evolution of these tumors.

Topics: Blotting, Northern; Blotting, Southern; Carcinoma, Squamous Cell; Cyclin D1; Cyclins; Female; Gene Amplification; Gene Rearrangement; Humans; Oncogene Proteins; RNA, Messenger; Tumor Cells, Cultured; Uterine Cervical Neoplasms; Vulvar Neoplasms

Cyclins D1, D2 and D3 are thought to function in the G1 phase of the cell division cycle by regulating the activity of cyclin-dependent protein kinases. All three D-type cyclins can be shown to associate with two specific kinases, cdk4 and cdk6, providing at least six possible combinations. To establish whether different cell types require different subsets of these complexes and whether they are altered in tumours where D-cyclin expression is perturbed, we surveyed a series of tumour cell lines and compared them where possible to non-tumorigenic counterparts. Although complexes involving cdk4 or cdk6 were readily observed in many of the cell lines, no complexes were detectable in human cells harbouring DNA tumour virus oncoproteins or in which the retinblastoma gene product (pRb) is mutated or missing. These data suggest that as well as being a potential substrate for D-cyclin-kinases, functional pRb contributes to the formation or stability of the complexes, at least in human cells.

Topics: Breast Neoplasms; Cyclin D1; Cyclins; Female; Humans; Keratinocytes; Oncogene Proteins; Protein Kinases; Retinoblastoma Protein; Tumor Cells, Cultured; Uterine Cervical Neoplasms