cyclin-d1 and Inflammation

Ventilator-induced lung injury (VILI) may be caused by incorrect mechanical ventilation (MV), and its progression is mainly related to inflammatory reaction, apoptosis, and oxidative stress. The Wnt/β-catenin pathway can modulate inflammation and apoptosis; however, its role in VILI is unknown. This research aims to explore the role of the Wnt/β-catenin pathway in VILI. VILI models were established using rats and type II alveolar epithelial (ATII) cells. Glycogen synthase kinase 3β (GSK-3β), β-catenin, and cyclin D1 were determined using western blotting and immunofluorescence. Apoptosis of lung tissues was evaluated using TUNEL, flow cytometry, Bax, and Bcl2 protein. Interleukin-1β (IL-1β), interleukin-6 (IL-6), and tumor necrosis factor-α (TNF-α) were detected via enzyme-linked immunosorbent assay (ELISA). Lung pathological injury was evaluated through hematoxylin and eosin (H&E) staining. Lung permeability was evaluated by the ratio of dry to wet weight of lung tissue and the total protein level of bronchoalveolar lavage fluid (BALF). The results showed that GSK-3β expression was enhanced and β-catenin expression was diminished in lung tissue under MV. SB216763 increased β-catenin and cyclin D1 expression by inhibiting GSK-3β expression and inhibited the inflammatory response and apoptosis of lung, alleviated pulmonary edema and lung tissue permeability, and significantly mitigated lung injury. However, inhibition of β-catenin expression by MSAB attenuated the anti-inflammatory and antiapoptotic effects of SB216763 in VILI. Overall, the present study demonstrates that the Wnt/β-catenin pathway activation in MV may play an anti-inflammatory and antiapoptotic role, thereby alleviating lung injury and delaying VILI progression, which may be a key point of intervention in VILI.

Topics: Animals; Anti-Inflammatory Agents; Apoptosis; beta Catenin; Cyclin D1; Glycogen Synthase Kinase 3 beta; Inflammation; Interleukin-6; Lung; Rats; Ventilator-Induced Lung Injury

Circular RNAs (circRNAs) are important regulators on the onset and progression of rheumatoid arthritis (RA). Our purpose is to explore the role and underpin mechanism of circ_0000396 in RA progression.. RA patients (n = 39) and healthy volunteers (n = 33) were recruited from the Affiliated Hospital of Shaanxi University of Chinese Medicine for the present work. Circ_0000396, microRNA-574-5p (miR-574-5p) and R-spondin 1 (RSPO1) RNA levels were analyzed by reverse transcription-quantitative polymerase chain reaction. Cell proliferation was analyzed by 3-(4,5-dimethyl-thiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, colony formation assay, and 5-ethynyl-2'-deoxyuridine (EDU) assay. Cell apoptosis was assessed by flow cytometry. Protein expression levels of proliferating cell nuclear antigen (PCNA), Cyclin D1, Cyclin E1, BCL2-associated × protein (Bax), B-cell lymphoma-2 (Bcl2), interleukin-1β (IL-1β), tumor necrosis factor-α (TNF-α) and RSPO1 were detected by western blot assay. Enzyme-linked immunosorbent assay (ELISA) was conducted to analyze the secretion of pro-inflammatory cytokines including IL-1β and TNF-α. The interaction between miR-574-5p and circ_0000396 or RSPO1 was confirmed by dual-luciferase reporter assay and RNA-pull down assay.. Circ_0000396 expression was notably down-regulated in RA patients compared with healthy controls. Circ_0000396 overexpression suppressed the proliferation and inflammatory response and triggered the apoptosis of RA synovial fibroblasts (RASFs), accompanied by decreases in PCNA, Cyclin D1, Cyclin E1, Bcl2, IL-1β and TNF-α protein expression and an increase in Bax protein expression. Circ_0000396 acted as a molecular sponge for miR-574-5p, and circ_0000396 overexpression-mediated protective effects on RASFs dysfunction were largely reversed by the introduction of miR-574-5p mimics. miR-574-5p interacted with the 3' untranslated region (3'UTR) of RSPO1, and miR-574-5p negatively regulated RSPO1 expression in RASFs. Circ_0000396 could up-regulate the expression of RSPO1 by sponging miR-574-5p in RASFs. RSPO1 interference largely overturned circ_0000396 overexpression-mediated effects in RASFs.. Circ_0000396 restrained the proliferation and inflammation and induced the apoptosis of RASFs by mediating miR-574-5p/RSPO1 axis, which provided novel potential targets for RA treatment.

Topics: 3' Untranslated Regions; Arthritis, Rheumatoid; Cell Proliferation; Cyclin D1; Humans; Inflammation; MicroRNAs; Proliferating Cell Nuclear Antigen; RNA, Circular; Thrombospondins; Tumor Necrosis Factor-alpha

Osteoarthritis (OA) is the most prevalent degenerative joint disease. In vitro experiments are an intuitive method used to investigate its early pathogenesis. Chondrocyte inflammation models in rats and mice are often used as in vitro models of OA. However, similarities and differences between them in the early stages of inflammation have not been reported.. This paper seeks to compare the chondrocyte phenotype of rats and mice in the early inflammatory state and identify chondrocytes suitable for the study of early OA.. Under similar conditions, chondrocytes from rats and mice were stimulated using the same IL-1β concentration for a short period of time. The phenotypic changes of chondrocytes were observed under a microscope. The treated chondrocytes were subjected to RNA-seq to identify similarities and differences in gene expression. Chondrocytes were labelled with EdU for proliferation analysis. Cell proliferation-associated proteins, including minichromosome maintenance 2 (MCM2), minichromosome maintenance 5 (MCM5), Lamin B1, proliferating cell nuclear antigen (PCNA), and Cyclin D1, were analysed by immunocytochemical staining, cell immunofluorescence, and Western blots to verify the RNA-seq results.. RNA-seq revealed that the expression patterns of cytokines, chemokines, matrix metalloproteinases, and collagen were similar between the rat and mouse chondrocyte inflammation models. Nonetheless, the expression of proliferation-related genes showed the opposite pattern. The RNA-seq results were further verified by subsequent experiments. The expression levels of MCM2, MCM5, Lamin B1, PCNA, and Cyclin D1 were significantly upregulated in rat chondrocytes (P < 0.05) and mouse chondrocytes (P < 0.05).. Based on the findings, the rat chondrocyte inflammation model may help in the study of the early pathological mechanism of OA.

Topics: Animals; Cell Proliferation; Chondrocytes; Cyclin D1; Disease Models, Animal; Gene Expression; Immunoblotting; Immunohistochemistry; Inflammation; Interleukin-1beta; Mice; Osteoarthritis; Proliferating Cell Nuclear Antigen; Rats; RNA-Seq

The current study investigates the anti-inflammatory and hepatoprotective effects of selenium (Se) formulated as nanoparticles (SeNPs) and in combination with quercetin (QCT) against thioacetamide (TAA)-induced hepatocellular carcinoma (HCC) in rats. ​​​​​​Seventy-two male Sprague-Dawley rats were divided into six groups (n = 12). Three control groups; normal, SeNPs; group received SeNPs only and HCC; group received TAA. In addition, three preventive groups; SeNPs + TAA, QCT + TAA, and QCT + SeNPs + TAA. Induction of HCC was detected histopathologically and by the raise of the serum level of alpha-fetoprotein (AFP). Oxidative stress was evaluated by the hepatic levels of reduced glutathione (GSH), glutathione peroxidase (GPx), and malondialdehyde (MDA) spectrophotometrically. The oncogenic pathway of p53/β-catenin/cyclin D1 was assessed by immunohistochemistry. The inflammatory markers; interleukin-33 (IL-33), IL-6, and IL-1β were assessed by enzyme-linked immune sorbent assay. SeNPs prevented the elevation of serum AFP and hepatic IL-33, IL-1β, and IL-6 in comparison to HCC or QCT + TAA groups. SeNPs + TAA exhibited a lower positive hepatic staining of p53, β-catenin, and cyclin D1 in comparison to HCC or QCT + TAA groups. Moreover, SeNPs improved the overall oxidative balance indicated by low hepatic MDA and enhanced GSH and GPx when compared to HCC or QCT + TAA groups. ​​SeNPs alone and in combination with QCT were found to suppress the progression of HCC in rats via the enhancement of the oxidative stress and then inflammatory status and the prevention of the deregulation of the oncogenic axis pathway of p53/β-catenin/cyclin D.

Topics: alpha-Fetoproteins; Animals; beta Catenin; Carcinoma, Hepatocellular; Cyclin D1; Inflammation; Interleukin-33; Interleukin-6; Liver; Liver Neoplasms; Male; Nanoparticles; Oxidative Stress; Quercetin; Rats; Rats, Sprague-Dawley; Selenium; Thioacetamide; Tumor Suppressor Protein p53

Cyclin D1 is a regulatory subunit of -Cyclin Dependent Kinases 4 and 6 (CDK4/6) and regulates progression from G1 to S phase of the cell cycle. Dysregulated cyclin D1-CDK4/6 contributes to abnormal cell proliferation and tumor development. Phosphorylation of threonine 286 of cyclin D1 is necessary for ubiquitin-dependent degradation. Non-phosphorylatable cyclin D1 mutants are stabilized and concentrated in the nucleus, contributing to genomic instability and tumor development. Studies investigating the tumor-promoting functions of cyclin D1 mutants have focused on the use of artificial promoters to drive the expression which unfortunately may not accurately reflect tumorigenic functions of mutant cyclin D1 in cancer development. We have generated a conditional knock-in mouse model where cyclin D1T286A is expressed under the control of its endogenous promoter following Cre-dependent excision of a lox-stop-lox sequence. Acute expression of cyclin D1T286A following tamoxifen-inducible Cre recombinase triggers inflammation, lymphocyte abnormality and ultimately mesenteric tumors in the intestine. Tissue-specific expression of cyclin D1T286A in the uterus and endometrium cooperates with Pten loss to drive endometrial hyperplasia and cancer. Mechanistically, cyclin D1T286A mutant activates NF-κB signaling, augments inflammation, and contributes to tumor development. These results indicate that mutation of cyclin D1 at threonine 286 has a critical role in regulating inflammation and tumor development.

Topics: Animals; Carcinoma; Cyclin D1; Cyclin-Dependent Kinase 4; Endometrial Hyperplasia; Female; Humans; Inflammation; Mice; PTEN Phosphohydrolase; Threonine

Sinonasal inverted papilloma (IP) is a rare and benign epithelial tumor in the sinonasal tract. Recent study suggested the potential role of chronic inflammation in the pathogenesis of IP. This study aims to compare the inflammatory pattern, the capacity of epithelial cell proliferation and. Sixteen patients with unilateral IP and contralateral nasal polyps (NP) were identified through a retrospective chart review. The neutrophil and eosinophil infiltration in IP and NP were assessed by immunostaining for neutrophil elastase and major basic protein (MBP). Immunohistochemistry was also used to assess the expression of FoxM1, Ki67 and cyclin D1 in IP tissue and contralateral NP. Sanger sequencing was used to evaluate the. The neutrophil count in IP was significantly higher than contralateral NP and 68.8% patients presented with neutrophilic inflammation, whereas only 37.5% contralateral NP tissue showed neutrophilic inflammation. The percentage of positive FoxM1-staining cells was significantly increased in IP, and positively correlated with the percentage of cells with positive staining for cyclin D1 and ki67 as well as neutrophil counts.. Our study demonstrated distinct inflammatory pattern between IP and contralateral NP and implied the oncogenic role of neutrophils in the pathogenesis of IP.

Topics: Cell Proliferation; Cyclin D1; Humans; Inflammation; Ki-67 Antigen; Nasal Polyps; Neutrophil Infiltration; Nose Neoplasms; Papilloma, Inverted; Paranasal Sinus Neoplasms; Retrospective Studies

Knee osteoarthritis (KOA) is one of the common age-degenerative diseases. Recent studies have demonstrated that the pathogenesis of KOA is closely related to synovial lesions. Jiawei Duhuo Jisheng mixture (JDJM) has shown great potential in the treatment of KOA. However, the effect and mechanism of JDJM on synovial lesions of KOA remain unclear.. The regulatory effect of JDJM on the Wnt/β-catenin signaling pathway in KOA inflamed synovium was studied via in vitro and in vivo experiments, respectively.. For the in vitro experiment, fibroblasts were isolated from the rabbit synovium with KOA. The fibroblasts were grouped as follows: the vehicle group was given 0.5% FBS; the inhibitor group was treated with 0.5% volume fraction of XAV939; the normal serum groups and JDJM serum groups were treated with 5%, 10%, and 20% volume fractions of normal serum and JDJM-containing serum. The expression levels of Wnt3a, β-catenin, Cyclin D1, metalloproteinase-7(MMP-7) and cyclooxygenase-2(COX-2) were detected by different assays 48 and 72 h after the intervention. For the in vivo experiment, the rabbit KOA model was prepared using the improved Hulth modeling method, whereby all rabbits were randomly divided into normal control, model control, positive control, and traditional Chinese medicine (TCM) groups. The expression levels of Wnt3a, β-catenin, Cyclin D1, MMP-7 and COX-2 were detected by different assays in the 2, 4, and 8 weeks of treatment.. In the two test results of in vitro experiments, the normal serum group was compared with the JDJM-containing serum group with the same volume fraction, demonstrating that mRNA transcription and protein expression levels of Wnt3a, β-catenin, Cyclin D1, MMP-7, and COX-2 in the latter decreased (P < 0.05), with more pronounced effects observed in the group treated with 20% volume fraction of JDJM serum. Compared with the inhibitor group, there was no significant difference (P > 0.05) in the mRNA transcription and protein expression levels, i.e., Wnt3a, β-catenin, Cyclin D1, and MMP-7 were observed in the JDJM serum groups, except for a significant decrease (P < 0.05) in the level of mRNA transcription and protein expression of COX-2. Based on the in vivo experiment, compared to the model control group, articular cartilage, synovial hyperplasia, and the inflammatory reaction of the TCM group at different treatment times were significantly improved. The mRNA transcription level of Wnt3a, β-catenin, Cyclin D1, MMP-7 and COX-2 detected by RT-qPCR and the protein expression level of Wnt3a, β-catenin, Cyclin D1, MMP-7 and COX-2 detected by Western blot were significantly reduced (P < 0.05), and the effect was more evident at the eighth week.. JDJM can regulate the synovial Wnt/β-catenin signaling pathway in KOA models, reduce the mRNA transcription and protein expression levels of Wnt3a, β-catenin, Cyclin D1, MMP-7, and COX-2 in the synovium, thus inhibiting synovial inflammation and protecting articular cartilage, which could be the key mechanism of action in treating this disease.

Topics: Animals; beta Catenin; Cyclin D1; Cyclooxygenase 2; Drugs, Chinese Herbal; Inflammation; Matrix Metalloproteinase 7; Osteoarthritis, Knee; Rabbits; RNA, Messenger; Synovial Membrane; Wnt Signaling Pathway

Colorectal cancer is related to ulcerative colitis. This study aimed to investigate the effects of aspirin on non-specific inflammation developing into cancer.. Ulcerative colitis model was generated by administrating azoxymethane/dextran sulfate sodium to mice. Weight, tumor size/ amount, and intestinal mucositis scores were analyzed. Inflammatory cell infiltration and atypical hyperplasia were determined with hematoxylin-eosin staining. Immunohistochemical assay was used to detect the proliferating cell nuclear antigen. Interleukin-6 and interleukin-10 were detected using enzyme-linked immunosorbent assay. Signal transducer and activator of transcription 3, phosphorylated-STAT3, cyclin D1, and suppressor of cytokine signaling 3 were examined with western blotting.. Aspirin remarkably decreased tumor size/amount compared to those of the ulcerative colitis model group (P < .05). Interleukin-6 was increased and interleukin-10 was decreased in mice of ulcerative colitis model group compared with the control group (P < .05). Aspirin markedly reduced interleukin-6 and enhanced interleukin-10 compared to the ulcerative colitis model group (P < .05) induced Azoxymethane/dextran sulfate sodium inflammation (3 weeks) and atypical hyperplasia (8 weeks). Aspirin predominantly inhibited the "inflammation-atypical hyperplasia-cancer" process and alleviated inflammatory cell infiltration of mice in the ulcerative colitis model group. Aspirin promoted apoptosis and alleviated proliferating cell nuclear antigen of atypical hyperplastic intestinal mucosal cells at 8 weeks post-modeling. The expression of phosphorylated-STAT3, signal transducer and activator of transcription 3, cyclin D1, and suppressor of cytokine signaling 3 was significantly increased in mice of ulcerative colitis model group compared to the control group (P < .05). Aspirin remarkably decreased phosphorylated-STAT3, signal transducer and activator of transcription, and cyclin D1 expression compared with ulcerative colitis model group (P < .05).. Aspirin inhibited carcinogenesis of intestinal mucosal cells in the ulcerative colitis model by inhibiting the interleukin-6/ Janus kinase/signal transducer and activator of transcription 3 signaling pathway and promoted apoptosis, thereby suppressing proliferation.

Topics: Animals; Apoptosis; Aspirin; Azoxymethane; Carcinogenesis; Cell Proliferation; Colitis, Ulcerative; Colorectal Neoplasms; Cyclin D1; Dextran Sulfate; Hyperplasia; Inflammation; Interleukin-10; Interleukin-6; Janus Kinases; Mice; Proliferating Cell Nuclear Antigen; Signal Transduction; STAT3 Transcription Factor

Space radiation-induced gastrointestinal (GI) cancer risk models for future interplanetary astronauts are being developed that primarily rely on quantitative animal model studies to assess radiation-quality effects of heavy-ion space radiation exposure in relation to γ-rays. While current GI-cancer risk estimation efforts are focused on sporadic GI-cancer mouse models, emerging in-vivo data on heavy-ion radiation-induced long-term GI-inflammation are indicative of a higher but undetermined risk of GI-inflammation associated cancers, such as colitis-associated cancer (CAC). Therefore, we aimed to assess radiation quality effects on colonic inflammation, colon cancer incidence, and associated signaling events using an in-vivo CAC model i.e., Il10-/- mice. Male Il10-/- mice (8-10 weeks, n = 12/group) were irradiated with either sham, γ-rays or heavy-ions (28Si or 56Fe), and histopathological assessments for colitis and CAC were conducted at 2.5 months post-exposure. qPCR analysis for inflammation associated gene transcripts (Ptges and Tgfb1), and in-situ staining for markers of cell-proliferation (phospho-histone H3), oncogenesis (active-β-catenin, and cyclin D1), and inflammation (phospho-p65NF-κB, iNOS, and COX2) were performed. Significantly higher colitis and CAC frequency were noted after heavy-ion exposure, relative to γ and control mice. Higher CAC incidence after heavy-ion exposure was associated with greater activation of β-catenin and NF-κB signaling marked by induced expression of common downstream inflammatory (iNOS and COX2) and pro-proliferative (Cyclin D1) targets. In summary, IR-induced colitis and CAC incidence in Il10-/- mice depends on radiation quality and display co-activation of β-catenin and NF-κB signaling.

Topics: Animals; beta Catenin; Carcinogenesis; Colitis; Colonic Neoplasms; Cyclin D1; Cyclooxygenase 2; Disease Models, Animal; Gastritis; Inflammation; Male; Mice; Mice, Inbred C57BL; Neoplasms, Radiation-Induced; NF-kappa B

Circular RNAs have been reported as vital regulators and promising therapeutic targets in multiple human diseases, including atherosclerosis (AS). However, the functional roles of circ_0010283 in AS remain unclear. The real-time quantitative polymerase chain reaction was used to determine the expression levels of circ_0010283, microRNA (miR)-377-3p, and cyclin D1 (CCND1) in serum samples. The vascular smooth muscle cells (VSMCs) were treated with oxidized low-density lipoprotein (ox-LDL) to establish the in vitro cell model of AS. The 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyl-2H-tetrazol-3-ium bromide and clonal colony-forming assays were performed to assess cell proliferation. The apoptosis was determined by flow cytometry assay. The migration of VSMCs was examined by wound healing and transwell assays. Western blot analysis was used to quantify protein expression. The association among circ_0010283, miR-377-3p, and CCND1 was confirmed by dual-luciferase reporter assay. We found that the serum level of circ_0010283 was upregulated in patients with AS and treatment with ox-LDL also increased the expression of circ_0010283 in VSMCs. Treatment with ox-LDL also increased proliferation, migration, and inflammation while inhibited apoptosis in VSMCs, which was overturned by silencing of circ_0010283. Moreover, miR-377-3p was a target of circ_0010283, and downregulation of miR-377-3p counteracted circ_0010283 silencing-induced effects on ox-LDL-stimulated VSMCs. The overexpression of miR-377-3p inhibited proliferation, migration, and inflammation while induced apoptosis of VSMCs by targeting CCND1. CCND1 was a target of miR-377-3p, and circ_0010283 acted as the miR-377-3p sponge to increase CCND1 expression. Circ_0010283 regulated proliferation, apoptosis, migration, and inflammation of ox-LDL-stimulated VSMCs through modulating miR-377-3p and CCND1.

Topics: Aged; Apoptosis; Atherosclerosis; Case-Control Studies; Cell Movement; Cell Proliferation; Cells, Cultured; Cyclin D1; Female; Gene Expression Regulation; Humans; Inflammation; Lipoproteins, LDL; Male; MicroRNAs; Middle Aged; Muscle, Smooth, Vascular; Myocytes, Smooth Muscle; RNA, Circular; Signal Transduction

Although interferon regulatory factor 7 (IRF7) has known roles in regulating the inflammatory response, vascular smooth muscle cell proliferation, and apoptosis, its role in the pathogenesis of pulmonary hypertension (PH) is unclear. We hypothesized that IRF7 overexpression could inhibit pulmonary vascular remodeling and slow the progression of PH.. IRF7 mRNA and protein levels in the lung samples and pulmonary artery smooth muscle cells (PASMCs) isolated from monocrotaline (MCT)-induced PH rats were assessed. We evaluated the effects of IRF7 on inflammation, proliferation, and apoptosis using an in vivo MCT-induced PH rat model and in vitro methods.. We noted decreased IRF7 mRNA and protein levels in the pulmonary vasculature of MCT-induced PH rats. IRF7 upregulation attenuated pulmonary vascular remodeling, decreased the pulmonary artery systolic pressure, and improved the right ventricular (RV) structure and function. Our findings suggest that nuclear factor kappa-Bp65 (NF-κBp65) deactivation could confer pulmonary vasculature protection, reduce proinflammatory cytokine (tumor necrosis factor-α, interleukin 6) release, and decrease PASMC proliferation and resistance to apoptosis via deactivating transcription factor 3 (ATF3) signaling. ATF3 deactivation induced the downregulation of the proliferation-dependent genes proliferating cell nuclear antigen (PCNA), cyclin D1, and survivin, coupled with increased levels of B cell lymphoma-2-associated X protein (Bax)/B cell lymphoma-2 (Bcl2) ratio, and cleaved caspase-3 in PASMCs.. Our findings showed that IRF7 downregulation could initiate inflammation via NF-κBp65 signaling, causing PASMC proliferation via ATF3 signaling pathway activation. Therefore, IRF7 could be a potential molecular target for PH therapy.

Topics: Activating Transcription Factor 3; Animals; Apoptosis; bcl-2-Associated X Protein; Caspase 3; Cell Proliferation; Cells, Cultured; Core Binding Factor Alpha 1 Subunit; Cyclin D1; Dependovirus; Heart Ventricles; Hemodynamics; Hypertension, Pulmonary; Inflammation; Interferon Regulatory Factor-7; Lung; Male; Monocrotaline; Muscle, Smooth, Vascular; Myocytes, Smooth Muscle; Proliferating Cell Nuclear Antigen; Rats, Sprague-Dawley; Receptor for Advanced Glycation End Products; Signal Transduction; Survivin; Up-Regulation; Vascular Remodeling

Mesenchymal stem cells (MSCs) are broadly applied in regenerative therapy to replace cells that are lost or impaired during disease. The low survival rate of MSCs after transplantation is one of the major limitations heavily influencing the success of the therapy. Unfavorable microenvironments with inflammation and oxidative stress in the damaged regions contribute to MSCs loss. Most of the strategies developed to overcome this obstacle are aimed to prevent stress-induced apoptosis, with little attention paid to senescence-another common stress reaction of MSCs. Here, we proposed the strategy to prevent oxidative stress-induced senescence of human endometrial stem cells (hMESCs) based on deferoxamine (DFO) application. DFO prevented DNA damage and stress-induced senescence of hMESCs, as evidenced by reduced levels of reactive oxygen species, lipofuscin, cyclin D1, decreased SA-β-Gal activity, and improved mitochondrial function. Additionally, DFO caused accumulation of HIF-1α, which may contribute to the survival of H

Topics: Apoptosis; Cell Differentiation; Cell Line; Cellular Microenvironment; Cellular Senescence; Cyclin D1; Deferoxamine; Endometrium; Female; Gene Expression Regulation, Developmental; Humans; Hydrogen Peroxide; Inflammation; Lipofuscin; Mesenchymal Stem Cells; Oxidative Stress; Reactive Oxygen Species; Regenerative Medicine; Signal Transduction

Porcine pancreatic stem cells (pPSCs) can be induced to insulin-secreting cells and therefore considered the most promising seeding cells for curing human diabetes in future. However, insufficient pPSCs number is one of the bottleneck problems before its clinical application. SerpinB1 is a serine protease inhibitor in neutrophils and can directly promote the proliferation of β cells. Whether SerpinB1 is involved in pPSC proliferation and differentiation remains unknown. The effects of SerpinB1 on pPSCs proliferation were measured by Cell Counting Kit-8, 5-ethynyl-2'-deoxyuridine, qRT-PCR, western blot, and flow cytometry assays. We found that pPSCs did not efficiently reach the S phase when SerpinB1 expression was knocked down with short hairpin RNA (sh-SerpinB1), the expression of Cyclin D1, CDK-2, and PCNA also decreased. Meanwhile, cell viability and proliferation ability were both declined. Further analyses showed that the expression level of phosphorylated STAT3/STAT3was downregulated, along with an upregulation of p53 and p21. We used a two-step induction method to induce pPSCs to insulin-secreting cells and found that SerpinB1 expression in insulin-secreting cells was higher than in pPSCs. Meanwhile, the protein expression level of phosphorylated STAT3/STAT3 was increased while p53 and p21 was decreased in induced insulin-secreting cells in comparison with control cells. The insulin-secreting cells derived from the sh-SerpinB1 cells secreted less insulin and showed poor sensitivity to high glucose than control group. However, the insulin-secreting cells derived from the ov-SerpinB1 cells has a quite contrary tendency. In conclusion, this study demonstrates that SerpinB1 promotes the proliferation of pPSCs through the STAT3 signaling pathway, and SerpinB1 is a key factor for maintaining the viability of pPSCs during the transition to insulin-secreting cells.

Topics: Adult Stem Cells; Animals; Cell Culture Techniques; Cell Differentiation; Cell Line; Cell Proliferation; Cyclin D1; Cyclin-Dependent Kinase 2; Disease Progression; Inflammation; Insulin; Insulin-Secreting Cells; Pancreas; Phosphorylation; Proliferating Cell Nuclear Antigen; Proto-Oncogene Proteins p21(ras); Serpins; Signal Transduction; STAT3 Transcription Factor; Swine; Tumor Suppressor Protein p53

Dickkopf-1 (DKK1), an inhibitor of the most frequently impaired signaling pathway in hepatocellular carcinoma (HCC), the Wnt/beta-catenin pathway, seems to fulfill contradictory functions in the process of tumorigenesis, acting either as an oncogenic promoter of metastasis or as a tumor suppressor. Elevated serum levels of DKK1 have been reported in HCC; however, little is known about its functional significance. In the current study, we treated HepG2/C3A and PLC/PRF/5 with the recombinant protein DKK1. Cytotoxicity was first determined by the WST-8 assay. AFP expression was measured at both the mRNA and protein levels. Expression of the oncogenes MYC, CCND1, hTERT, and MDM2 and the tumor suppressor genes TP53, P21 and RB was assessed. Western blot analysis of non-phosphorylated ẞ-catenin and Sanger sequencing were performed to explain the functional differences between the two cell lines. Subsequently, inflammation, migration and invasion were evaluated by qPCR, ELISA, the Boyden chamber assay, zymography, and MMP-2 and MMP-9 western blot analysis. Knockdown of DKK1 and TGF-β1 were also performed. Our results suggest that DKK1 exerts an oncogenic effect on HepG2/C3A cell line by upregulating the expression of oncogenes and downregulating that of tumor suppressor genes, whereas the opposite effect was demonstrated in PLC/PRF/5 cells. This differential impact of DKK1 can be explained by the mutations that affect the canonical Wnt pathway that were detected in exon 3 of the CTNNB1 gene in the HepG2 cell line. We further confirmed that DKK1 promotes inflammation, tumor invasion and migration in both cell types. The canonical pathway was not responsible for the DKK1 proinvasive effect, as indicated by the active ẞ-catenin levels in the two cell lines upon DKK1 treatment. Interestingly, knockdown of TGF-β1 negatively affected the DKK1 proinvasive effect. Taken together, DKK1 appears to facilitate tumor invasion and migration through TGF- β1 by remodeling the tumor microenvironment and inducing inflammation. This finding endorses the relevance of TGF-β1 as a therapeutic target.

Topics: alpha-Fetoproteins; Cell Line, Tumor; Cell Movement; Cell Proliferation; Cyclin D1; Cyclin-Dependent Kinase Inhibitor p21; Diffusion Chambers, Culture; Gene Expression Regulation, Neoplastic; Hep G2 Cells; Hepatocytes; Humans; Inflammation; Intercellular Signaling Peptides and Proteins; Proto-Oncogene Proteins c-mdm2; Proto-Oncogene Proteins c-myc; Retinoblastoma Protein; RNA, Small Interfering; Telomerase; Transforming Growth Factor beta1; Tumor Suppressor Protein p53; Wnt Signaling Pathway

Psoriasis is a hyperproliferative inflammatory skin disease; therefore, it is highly likely that psoriatic skin lesions may transform into malignancies. However, malignant transformation is not common. We performed immunohistochemical studies using anti‑cyclin D1, anti‑cyclin E, anti‑pRb, anti‑p53, anti‑p16INK4a, and anti‑Ki‑67 antibodies in normal skin, psoriatic epidermal tissue, and squamous cell carcinoma (SCC) tissue. Furthermore, western blot analysis and immunohistochemical staining were performed to ascertain differences in cyclin D1, cyclin E, pRb, and Ki‑67 expression before and after treatment for psoriasis. Cyclin D1 expression was higher in chronic psoriatic lesions than that in normal epidermis. Psoriasis lesions showed a strong intensity of positive nuclear staining for cyclin D1 among several normally stained nuclei in the basal layer. Cyclin E expression in psoriasis was stronger in the granular and spinous layer than in the normal epidermis. Expression levels of pRb and p53 were found to be higher in the psoriasis group compared with the normal epidermis. Total basal layer cell counts for p53WT expression were found to be significantly higher in the psoriasis group compared with the normal group. However, p16 expression was very weak in the normal and psoriasis groups compared with that in the SCC group. Ki‑67 immunoreactivity was significantly higher in psoriasis compared with normal epidermis and was similar with that in the SCC group. According to immunohistochemistry and immunoblot analysis, the expression levels of cyclin D1, cyclin E, pRb, and Ki‑67 in psoriasis lesions decreased after treatment and were similar with those in the normal group. Thus, increased expression of cyclin D1 and cyclin E may be involved in cell cycle progression in psoriatic epidermis, and pRb and p53 may play important roles in the prevention of malignant transformation under the hyperproliferative state in psoriasis.

Topics: Carcinoma, Squamous Cell; Cell Cycle Checkpoints; Cyclin D1; Cyclin E; Female; Gene Expression Regulation; Humans; Inflammation; Ki-67 Antigen; Male; Psoriasis; Retinoblastoma Protein; Skin; Tumor Suppressor Protein p53

The predominant role of Wnt family member 5A (Wnt5a) is to induce non-canonical Wnt signalling pathways, including the Wnt‑Ca2+ and Wnt‑planar cell polarity pathways. Enhanced Wnt5a expression is involved in the formation of psoriatic plaques; however, its mechanistic role remains to be determined. In the present study, the effects of Wnt5a expression on HaCaT keratinocytes were investigated. HaCaT cells were cultured in medium supplemented with 0, 40 or 80 ng/ml Wnt5a for 24 h. Cell proliferation, the cell cycle, gene expression and inflammatory responses were investigated using Cell‑Counting Kit‑8 assays, flow cytometry analyses, reverse transcription‑quantitative polymerase chain reaction analyses and enzyme‑linked immunosorbent assays, respectively. Wnt5a treatment was revealed to suppress cell proliferation in HaCaT cells. Furthermore, Wnt5a was also demonstrated to increase the proportion of HaCaT cells arrested at the G2/M phase of the cell cycle, but reduce the proportion of HaCaT cells arrested at G0/G1 phase cells. In addition, the expression levels of the differentiation markers, including filaggrin, keratin 1 and keratin 10 were revealed to be downregulated in HaCaT cells. Expression of the canonical Wnt signalling genes (β‑catenin and cyclin D1) and proliferation markers, such as Ki‑67 and proliferating cell nuclear antigen in HaCaT cells were also revealed to be downregulated. However, the expression levels of inflammatory response markers (interferon‑γ, interleukin‑8 and interleukin‑17A) were revealed to be upregulated in HaCaT cells following Wnt5a treatment. These findings suggest that Wnt5a expression may be involved in the inhibition of cell differentiation and the induction of an inflammatory response in patients with psoriasis.

Topics: beta Catenin; Cell Differentiation; Cell Line, Transformed; Cell Proliferation; Cell Survival; Cyclin D1; Dose-Response Relationship, Drug; Filaggrin Proteins; G2 Phase Cell Cycle Checkpoints; Gene Expression Regulation; Humans; Inflammation; Interferon-gamma; Interleukin-17; Interleukin-8; Intermediate Filament Proteins; Keratin-1; Keratin-10; Keratinocytes; Ki-67 Antigen; Signal Transduction; Wnt-5a Protein

Topics: Alkaloids; Animals; Carcinogenesis; Cell Proliferation; Cyclin D1; ErbB Receptors; Indole Alkaloids; Inflammation; Male; Mesocricetus; Mouth Neoplasms; NF-kappa B; Phosphatidylinositol 3-Kinases; Proto-Oncogene Proteins c-akt; Signal Transduction; STAT3 Transcription Factor; TOR Serine-Threonine Kinases

Rheumatoid arthritis (RA) fibroblast-like synoviocytes (RA-FLS) are reportedly involved in RA initiation, progression, and perpetuation. Previously a study showed that retinoid interferon-induced mortality 19 (GRIM19) improved the clinical and histological features of collagen-induced arthritis (CIA),and also inhibited osteoclast formation. However, the role of GRIM19 in RA-FLS remains unclear. In this study, we explored the biological function and underlying mechanism of GRIM19 in cultured RA-FLS. The expression of GRIM19 in synovial tissues and RA-FLS was detected by real-time quantitative RT-PCR(qRT-PCR).The effects of GRIM19 on proliferation, migration, invasion, apoptosis, and inflammatory cytokines levels in RA-FLS were determined using CCK8, wound healing, transwell invasion, and flow cytometry assays, and enzyme-linked immunosorbent assay (ELISA), respectively.GRIM19 and its related protein expression levels were determined by western blot. We found that GRIM19expression was significantly decreased in synovial tissues and FLS from RA patients.GRIM19significantly inhibited proliferation, migration, and invasion; promoted apoptosis; and suppressed inflammatory cytokine secretion by RA-FLS. Moreover, GRIM19 overexpression significantly decreasedthe expression levels of signal transducer and activator of transcription 3(STAT3)and its downstreamproteins,CyclinD1, Bcl-2, and MMP-9. These data indicate that promoting the expression of GRIM19 may yield therapeutic benefits in the treatment of RA.

Topics: Adult; Aged; Apoptosis; Apoptosis Regulatory Proteins; Arthritis, Rheumatoid; Cell Movement; Cell Proliferation; Cyclin D1; Cytokines; Female; Fibroblasts; Humans; Inflammation; Male; Matrix Metalloproteinase 9; Middle Aged; NADH, NADPH Oxidoreductases; Neoplasm Invasiveness; Proto-Oncogene Proteins c-bcl-2; Signal Transduction; STAT3 Transcription Factor; Synovial Membrane; Synoviocytes; Wound Healing

One long-term complication of chronic intestinal inflammation is the development of colorectal cancer. However, the mechanisms linking inflammation to the colorectal tumorigenesis are poorly defined. Previously, we have demonstrated that galectin-4 is predominantly expressed in the luminal epithelia of the gastrointestinal tract, and its loss of expression plays a key role in the colorectal tumorigenesis. However, the mechanism by which galectin-4 regulates inflammation-induced tumorigenesis is unclear. Here, we show that galectin-4 secreted by the colorectal cancer cell lines was bound to the cell surface. Neutralization of surface-bound galectin-4 with anti-galectin-4 antibody resulted in increased cell proliferation with concomitant secretion of several chemokines into the extracellular medium. Neutralization of the surface-bound galectin-4 also resulted in the up-regulation of transcription of 29 genes, several of which are components of multiple inflammation signaling pathways. In an alternate experiment, binding of recombinant galectin-4 protein to cell surface of the galectin-4-negative colorectal cancer cells resulted in increased p27, and decreased cyclin D1 and c-Myc levels, leading to cell cycle arrest and apoptosis. Together, these data demonstrated that surface-bound galectin-4 is a dual function protein-down-regulating cell proliferation and chemokine secretion in galectin-4-expressing colorectal cancer cells on one hand and inducing apoptosis in galectin-4-negative colorectal cancer cells on the other hand.

Topics: Apoptosis; Carcinogenesis; Cell Cycle Checkpoints; Cell Line, Tumor; Cell Proliferation; Chemokines; Colorectal Neoplasms; Cyclin D1; Galectin 4; Gene Expression Regulation, Neoplastic; HCT116 Cells; HT29 Cells; Humans; Inflammation; Intestinal Mucosa; Proliferating Cell Nuclear Antigen; Proto-Oncogene Proteins c-myc

Epidemiological studies have convincingly suggested that obesity is an important risk factor for postmenopausal breast cancer, but the mechanisms responsible for this relationship are still not fully understood. We hypothesize that obesity creates a low-grade inflammatory microenvironment, which stimulates Wnt-signaling and thereby promotes the development of breast cancer. To test this hypothesis, we evaluated the correlations between expression of multiple inflammatory cytokines and Wnt pathway downstream genes in mammary tissues from women (age ≥ 50) undergoing reduction mammoplasty. Moreover, we specifically examined the role of tumor necrosis factor-α (TNF-α), an important proinflammatory cytokine associated with obesity and a possible modulator of the Wnt pathway. The regulatory effects of TNF-α on Wnt pathway targets were measured in an ex vivo culture of breast tissue treated with anti-TNF-α antibody or TNF-α recombinant protein. We found that BMI was positively associated with the secretion of inflammatory cytokines IL-1β, IL-6 and TNF-α, all of which were negatively correlated with the expression of SFRP1. The transcriptional expression of Wnt-signaling targets, AXIN2 and CYCLIN D1, were higher in mammary tissue from women with BMI ≥ 30 compared to those with BMI < 30. Our ex vivo work confirmed that TNF-α is causally linked to the up-regulation of active β-CATENIN, a key component in the Wnt pathway, and several Wnt-signaling target genes (i.e. CYCLIN D1, AXIN2, P53 and COX-2). Collectively, these findings indicate that obesity-driven inflammation elevates Wnt-signaling in mammary tissue and thereby creates a microenvironment conducive to the development of breast cancer.

Topics: Antibodies, Blocking; Axin Protein; beta Catenin; Breast Neoplasms; Cell Proliferation; Cells, Cultured; Cyclin D1; Female; Gene Expression Regulation, Neoplastic; Humans; Inflammation; Intercellular Signaling Peptides and Proteins; Interleukin-1beta; Interleukin-6; Mammary Glands, Human; Membrane Proteins; Middle Aged; Obesity; Tumor Microenvironment; Tumor Necrosis Factor-alpha; Wnt Signaling Pathway

Hepatopulmonary syndrome (HPS) is a serious complication of advanced liver disease, which markedly increases mortality. Pulmonary vascular remodelling (PVR) induced by circulating mediators plays an important role in the pathogenesis of HPS, while the underlying mechanism remains undefined. In the present study, we reported that endothelin-1 (ET-1) is up-regulated and annexin A1(ANXA1) is down-regulated in HPS rat, and ET-1 decreases the ANXA1 expression in a dose-dependent manner in rat pulmonary arterial smooth muscle cells (PASMCs). Then, we showed that ANXA1 can decrease nuclear p-ERK1/2 accumulation and decrease the cyclin D1 expression, thus resulting in the subsequent inhibition of PASMCs proliferation. As previously reported, we confirmed that ET-1 decreases the ANXA1 protein levels by the carbonylation and degradation of ANXA1. In conclusion, our research links the signaling cascade of ET1-ANXA1-cell proliferation to a potential therapeutic strategy for blocking IPS-associated PVR.

Topics: Animals; Annexin A1; Cell Proliferation; Cells, Cultured; Cyclin D1; Down-Regulation; Endothelin-1; Hepatopulmonary Syndrome; Inflammation; MAP Kinase Signaling System; Muscle, Smooth, Vascular; Myocytes, Smooth Muscle; Phenotype; Protein Carbonylation; Pulmonary Artery; Rats; Rats, Sprague-Dawley; Signal Transduction; Up-Regulation

Topics: 9,10-Dimethyl-1,2-benzanthracene; Animals; Cell Differentiation; Cell Proliferation; Chemistry, Pharmaceutical; Cyclin D1; Cyclooxygenase 2; Disease Models, Animal; Epidermis; Female; Inflammation; Mice; Phytic Acid; Proliferating Cell Nuclear Antigen; Tumor Cells, Cultured

Dimethylarsinic acid (DMA(V) ), the major urinary metabolite of inorganic arsenic, is a urinary bladder carcinogen and bladder tumor promoter in adult rats. Increased urothelial cellular proliferation has been considered as an earlier phenotype in DMA(V) -induced bladder carcinogenesis. The present study examined the ultrastructural changes of bladder epithelial cells and expressions of proliferation factors, as well as the secretion of inflammatory cytokines in rats exposed to DMA(V) for 10 weeks by transmission electron microscopy (TEM), qRT-PCR, immunohistochemical staining and ELISA methods. The results showed that DMA(V) administered in the drinking water produced urothelial cytotoxicity and ultrastructural changes in rats. PCNA, cyclin D1 and COX-2 mRNA expressions and immunoreactivities were elevated in bladder urothelium. In addition, 200 ppm DMA(V) treatment increased the transforming growth factor-beta 1 (TGF-β1) secretion and decreased tumor necrosis factor-alpha (TNF)-α level in the urine of rats. These data suggest that chronic inflammation, bladder epithelium lesions and proliferation might be the basic process of the chronic toxicity effects in DMA(V) -treated rats.

Topics: Animals; Cacodylic Acid; Cell Proliferation; Cyclin D1; Cyclooxygenase 2; Cytokines; Enzyme-Linked Immunosorbent Assay; Female; Inflammation; Interleukin-6; Male; Microscopy, Electron, Transmission; Proliferating Cell Nuclear Antigen; Rats; Rats, Wistar; Reverse Transcriptase Polymerase Chain Reaction; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha; Urinary Bladder; Urinary Bladder Neoplasms; Urothelium

We describe 23 cases of high-grade myxoinflammatory fibroblastic sarcoma (MIFS). The patients were 15 women and 8 men, with the age ranging at the time of diagnosis from 39 to 93 years (mean, 64.3 years; median, 66 years). Follow-up was available for 18 patients, of whom 9 developed metastatic disease; 7 of these died. Most tumors showed a predilection for the soft tissues of the extremities, with 14 cases involving the lower limb and 5 the upper extremity. However, in both sites, the acral parts were affected in only 1 case each. Of the 4 remaining tumors, 2 were found in axilla, 1 was found in sacral area, and 1 developed in the scar on the breast, 14 years after previous excision of a mammary carcinoma and subsequent local irradiation. The tumor size ranged from 1.3 cm to as much as 30 cm in the largest dimension with a mean size of 8.3 cm. Histologically, the tumors were characterized by occurrence of 3 types of characteristic cells, including (1) lipoblast-like cells with an ample, distended, mucin-filled cytoplasm compartmentalized by a variable number of intracytoplasmic septa, thus remotely resembling soccer balls; (2) large, polygonal, bizarre ganglion-like cells similar to those seen in the Hodgkin disease, also called Reed-Sternberg-like cells. Within an ample, deeply eosinophilic cytoplasm, there was an oval nucleus with vesicular chromatin and a large, inclusion-like nucleolus. Binucleated, multinucleated, or more pleomorphic forms of these cells were also present; (3) cells with emperipolesis of variable sizes, ranging from very inconspicuous neoplastic cells containing only one to a few engulfed cells to conspicuous large ones having many inflammatory cells, usually polymorphonuclear leukocytes admixed with various numbers of some lymphoid cells, within the cytoplasm. Quite often, we found elements that combined the histologic features of all the above 3 characteristic tumor cell types. In 2 tumors, we found an additional undifferentiated spindle cell sarcoma component, whereas in another tumor, a chondrosarcomatous moiety was evident. For comparison, we studied 10 cases of pleomorphic hyalinizing angiectatic tumor (PHAT) of soft tissues. Based on the identification of morphological changes typical for MIFS within most of the cases of PHAT, we suggest that most cases of PHAT represent examples of MIFS merely having hyaline ectatic vessels.

Topics: Adult; Aged; Aged, 80 and over; Chondrosarcoma; Cyclin D1; Female; Fibrosarcoma; Follow-Up Studies; Humans; Hyalin; Immunohistochemistry; Inflammation; Male; Middle Aged; Myxosarcoma; Neoplasm Recurrence, Local; Soft Tissue Neoplasms

Chronic inflammation is an important risk factor for lung cancer. Therefore, identification of chemopreventive agents that suppress inflammation-driven lung cancer is indispensable. We studied the efficacy of combinations of indole-3-carbinol (I3C) and silibinin (Sil), 20 µmol/g diet each, against mouse lung tumors induced by 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) and driven by lipopolysaccharide (LPS), a potent inflammatory agent and constituent of tobacco smoke. Mice treated with NNK + LPS developed 14.7±4.1 lung tumors/mouse, whereas mice treated with NNK + LPS and given combinations of I3C and Sil had 7.1±4.5 lung tumors/mouse, corresponding to a significant reduction of 52%. Moreover, the number of largest tumors (>1.0mm) was significantly reduced from 6.3±2.9 lung tumors/mouse in the control group to 1.0±1.3 and 1.6±1.8 lung tumors/mouse in mice given I3C + Sil and I3C alone, respectively. These results were paralleled by significant reductions in the level of proinflammatory and procarcinogenic proteins (pSTAT3, pIκBα and COX-2) and proteins that regulate cell proliferation (pAkt, cyclin D1, CDKs 2, 4, 6 and pRB). Further studies in premalignant bronchial cells showed that the antiproliferative effects of I3C + Sil were higher than the individual compounds and these effects were mediated by targeting cyclin D1, CDKs 2, 4 and 6 and pRB. I3C + Sil suppressed cyclin D1 by reducing its messenger RNA level and by enhancing its proteasomal degradation. Our results showed the potential lung cancer chemopreventive effects of I3C + Sil in smokers/former smokers with chronic pulmonary inflammatory conditions.

Topics: Animals; Anticarcinogenic Agents; Cell Cycle Proteins; Cell Line, Tumor; Cell Proliferation; Cell Transformation, Neoplastic; Chemoprevention; Cyclin D1; Cyclin-Dependent Kinases; Cyclooxygenase 2; Drug Combinations; Female; Humans; I-kappa B Proteins; Indoles; Inflammation; Interleukin-6; Lipopolysaccharides; Lung; Lung Neoplasms; Mice; Mice, Inbred A; NF-KappaB Inhibitor alpha; Nitrosamines; Proto-Oncogene Proteins c-akt; Random Allocation; Retinoblastoma Protein; Silybin; Silymarin; Smoke; STAT3 Transcription Factor; Tumor Necrosis Factor-alpha

Liver cancer is a major health-care concern and its oncogenic mechanisms are still largely unclear. Persistent hepatocyte cell death is a common feature among various chronic liver diseases, the blocking of which presents as logical treatment. Therefore, we aimed at investigating tumor development in mice with hepatocyte-specific Bid depletion--a BH3-only Bcl-2 family member that amplifies apoptotic death signals. Hepatocyte-specific conditional Bid-knockout mice (Bid(Δhep)) were injected with 25 mg/kg diethylnitrosamine (DEN) at 14 days of age, and liver tumorigenesis was investigated 9 months later. Additionally, different models of acute liver injury were used including: acute high-dose DEN challenge, 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC) diet and carbon tetrachloride (CCL4) injection. Bid(Δhep) mice developed significantly fewer tumors, showed smaller maximal and average tumor size and reduced tumor incidence. In the acute DEN model, 48 h post injection we observed a significant reduction in liver injury in Bid(Δhep) animals, assessed via serum transaminases and liver histopathology. Furthermore, TNF-α, IL-1ß, cJUN and IL-6 mRNA expression was reduced. These findings were accompanied by reduced compensatory hepatocyte proliferation in Bid(Δhep) mice when compared with controls by immunohistochemistry for Ki67 and proliferating cell nuclear antigen 48 h after DEN injection. In the acute CCL4 model, Bid(Δhep) mice displayed reductions in liver injury and inflammation when compared with controls. No differences in liver injury and serum bilirubin levels were detected in Bid(Δhep) and Bid(flo/flo) mice fed with DDC, which induces bile duct injury and a ductular reaction. Our study demonstrates that in DEN-induced hepatocellular carcinoma, the inhibition of hepatocyte death pathways through Bid deletion protects animals from tumorigenesis. These results suggest that reducing hepatocyte cell death, liver inflammation and compensatory proliferation has a stronger beneficial effect than the potential side effect of enhancing tumor cell survival.

Topics: Animals; BH3 Interacting Domain Death Agonist Protein; Carbon Tetrachloride; Cell Proliferation; Cells, Cultured; Chemical and Drug Induced Liver Injury; Cyclin D1; Diethylnitrosamine; Disease Models, Animal; Hepatocytes; Hydrogen Peroxide; Inflammation; Interleukin-6; Liver; Liver Neoplasms; Mice; Mice, Inbred C57BL; Mice, Knockout; Mice, Transgenic; Non-alcoholic Fatty Liver Disease; Pyridines; Tumor Necrosis Factor-alpha

Ameloblastoma is a locally invasive neoplasm often associated with morbidity and facial deformities, showing increased Epidermal Growth Factor Receptor (EGFR) expression. Inhibition of EGFR was suggested as a treatment option for a subset of ameloblastomas. However, there are resistance mechanisms that impair anti-EGFR therapies. One important resistance mechanism for EGFR-inhibition is the EGFR nuclear localization, which activates genes responsible for its mitogenic effects, such as Cyclin D1.. We assessed EGFR nuclear localization in encapsulated (unicystic, n = 3) and infiltrative (multicystic, n = 11) ameloblastomas and its colocalization with Cyclin D1 by using anti-EGFR and anti-lamin B1 double labeling immunofluorescence analyzed by confocal microscopy. Oral inflammatory fibrous hyperplasia and oral squamous cell carcinoma samples were used for comparison.. Twelve cases of ameloblastoma exhibited nuclear EGFR colocalization with lamin B1. This positive staining was mainly observed in the ameloblast-like cells. The EGFR nuclear localization was also observed in control samples. In addition, nuclear EGFR colocalized with Cyclin D1 in ameloblastomas.. Nuclear EGFR occurs in ameloblastomas in association with Cyclin D1 expression, which is important in terms of tumor biology clarification and raises a concern about anti-EGFR treatment resistance in ameloblastomas.

Topics: Ameloblastoma; Biomarkers, Tumor; Carcinoma, Squamous Cell; Cell Nucleus; Cyclin D1; ErbB Receptors; Gene Expression Regulation, Neoplastic; Humans; Inflammation; Jaw Neoplasms; Lamin Type B; Microscopy, Confocal; Microscopy, Fluorescence; Mouth Neoplasms

UVB-induced inflammation, in particular the overexpression of cyclooxygenase-2 (COX-2) and prostaglandin (PG) E2, has been implicated in photocarcinogenesis. UVB-induced COX-2 has been associated with β-catenin signaling in keratinocytes. However, a definitive role for COX-2 in the activation of β-catenin signaling as well as its role in UVB-induced skin tumors has not been established. We report that exposure of the skin to UVB resulted in a time- and dose-dependent activation of β-catenin in C3H/HeN mice. This response was COX-2-dependent as UVB-exposed COX-2-deficient mice exhibited significantly lower levels of UVB-induced activation of β-catenin. Moreover, treatment of mice with indomethacin, a COX-2 inhibitor, and an EP2 antagonist inhibited UVB-induced β-catenin signaling. Exposure of SKH-1 hairless mice to UVB radiation (180 mJ/cm2) 3 times a week for 24 weeks resulted in activation of β-catenin signaling in UVB-irradiated skin as well as UVB-induced skin tumors. Concomitantly, the levels of CK1α and GSK-3β, which are responsible for β-catenin signaling, were reduced while the levels of c-Myc and cyclin D1, which are downstream targets of β-catenin, were increased. To further verify the role of UVB-induced inflammation in activation of β-catenin signaling, a high-fat-diet model was used. Administration of high-fat diet exacerbated UVB-induced inflammation. Administration of the high-fat diet enhanced β-catenin signaling and the levels of its downstream targets (c-Myc, cyclin D1, cyclin D2, MMP-2 and MMP-9) in UVB-exposed skin and skin tumors in SKH-1 mice. These data suggest that UV-induced COX-2/PGE2 stimulates β-catenin signaling, and that β-catenin activation may contribute to skin carcinogenesis.

Topics: Animals; beta Catenin; Casein Kinase I; Cyclin D1; Cyclin D2; Cyclooxygenase 2; Cyclooxygenase 2 Inhibitors; Diet, High-Fat; Female; Glycogen Synthase Kinase 3; Glycogen Synthase Kinase 3 beta; Indomethacin; Inflammation; Matrix Metalloproteinase 2; Matrix Metalloproteinase 9; Mice; Mice, Inbred C3H; Mice, Knockout; Prostaglandins E; Proto-Oncogene Proteins c-myc; Receptors, Prostaglandin E, EP2 Subtype; Receptors, Prostaglandin E, EP4 Subtype; Signal Transduction; Skin; Skin Neoplasms; Ultraviolet Rays

The goal of this study was to elucidate the action of the CD28 mimetic peptide p2TA (AB103) that attenuates an excessive inflammatory response in mitigating radiation-induced inflammatory injuries. BALB/c and A/J mice were divided into four groups: Control (C), Peptide (P; 5 mg/kg of p2TA peptide), Radiation (R; total body irradiation with 8 Gy γ-rays), and Radiation + Peptide (RP; irradiation followed by p2TA peptide 24 h later). Gastrointestinal tissue damage was evaluated by analysis of jejunum histopathology and immunohistochemistry for cell proliferation (Cyclin D1) and inflammation (COX-2) markers, as well as the presence of macrophages (F4/80). Pro-inflammatory cytokines IL-6 and KC as well as fibrinogen were quantified in plasma samples obtained from the same mice. Our results demonstrated that administration of p2TA peptide significantly reduced the irradiation-induced increase of IL-6 and fibrinogen in plasma 7 days after exposure. Seven days after total body irradiation with 8 Gy of gamma rays numbers of intestinal crypt cells were reduced and villi were shorter in irradiated animals compared to the controls. The p2TA peptide delivery 24 h after irradiation led to improved morphology of villi and crypts, increased Cyclin D1 expression, decreased COX-2 staining and decreased numbers of macrophages in small intestine of irradiated mice. Our study suggests that attenuation of CD28 signaling is a promising therapeutic approach for mitigation of radiation-induced tissue injury.

Topics: Amino Acid Sequence; Animals; Antigens, Differentiation; CD28 Antigens; Cell Proliferation; Cyclin D1; Cyclooxygenase 2; Fibrinogen; Gamma Rays; Gastrointestinal Tract; Immunohistochemistry; Inflammation; Interleukin-6; Jejunum; Macrophages; Male; Mice, Inbred BALB C; Mice, Inbred Strains; Peptides; Radiation Injuries, Experimental; Radiation-Protective Agents; Thrombosis; Whole-Body Irradiation

The therapeutic potential of mesenchymal stem cells (MSCs) has been highlighted recently for treatment of acute or chronic liver injury, by possibly differentiating into hepatocyte-like cells, reducing inflammation, and enhancing tissue repair. Despite recent progress, exact mechanisms of action are not clearly elucidated. In this study, we attempted to explore whether and how MSCs protected hepatocytes and stimulated allograft regeneration in small-for-size liver transplantation (SFSLT).. SFSLT model was established with a 30% partial liver transplantation (30PLT) in rats. The differentiation potential and characteristics of bone marrow derived MSCs were explored in vitro. MSCs were infused transvenously immediately after graft implantation in therapy group. Expressions of apoptosis-, inflammatory-, anti-inflammatory-, and growth factor-related genes were measured by RT-PCR, activities of transcription factors AP-1 and NF-κB were analyzed by EMSA, and proliferative responses of the hepatic graft were evaluated by immunohistochemistry and western blot.. MSCs were successfully induced into hepatocyte-like cells, osteoblasts and adipocytes in vitro. MSCs therapy could not only alleviate ischemia reperfusion injury and acute inflammation to promote liver regeneration, but also profoundly improve one week survival rate. It markedly up-regulated the mRNA expressions of HGF, Bcl-2, Bcl-XL, IL-6, IL-10, IP-10, and CXCR2, however, down-regulated TNF-α. Increased activities of AP-1 and NF-κB, as well as elevated expressions of p-c-Jun, cyclin D1, and proliferating cell nuclear antigen (PCNA), were also found in MSCs therapy group.. These data suggest that MSCs therapy promotes hepatocyte proliferation and prolongs survival in SFSLT by reducing ischemia reperfusion injury and acute inflammation, and sustaining early increased expressions of c-Jun N-terminal Kinase, Cyclin D1, and NF-κB.

Topics: Animals; Cell Differentiation; Cell- and Tissue-Based Therapy; Cyclin D1; Hepatocytes; Humans; Inflammation; JNK Mitogen-Activated Protein Kinases; Liver Regeneration; Liver Transplantation; Mesenchymal Stem Cells; NF-kappa B; Rats; Reperfusion Injury

Obesity is associated with increased risk in hepatocellular carcinoma (HCC) development and mortality. An important disease control strategy is the prevention of obesity-related hepatic inflammation and tumorigenesis by dietary means. Here, we report that apo-10'-lycopenoic acid (APO10LA), a cleavage metabolite of lycopene at its 9',10'-double bond by carotene-9',10'-oxygenase, functions as an effective chemopreventative agent against hepatic tumorigenesis and inflammation. APO10LA treatment on human liver THLE-2 and HuH7 cells dose dependently inhibited cell growth and upregulated sirtuin 1 (SIRT1), a NAD(+)-dependent protein deacetylase that may suppress hepatic carcinogenesis. This observed SIRT1 induction was associated with decreased cyclin D1 protein, increased cyclin-dependent kinase inhibitor p21 protein expression, and induced apoptosis. APO10LA supplementation (10 mg/kg diet) for 24 weeks significantly reduced diethylnitrosamine-initiated, high fat diet (HFD)-promoted hepatic tumorigenesis (50% reduction in tumor multiplicity; 65% in volume) and lung tumor incidence (85% reduction) in C57Bl/6J mice. The chemopreventative effects of APO10LA were associated with increased hepatic SIRT1 protein and deacetylation of SIRT1 targets, as well as with decreased caspase-1 activation and SIRT1 protein cleavage. APO10LA supplementation in diet improved glucose intolerance and reduced hepatic inflammation [decreased inflammatory foci, TNFα, interleukin (IL)-6, NF-κB p65 protein expression, and STAT3 activation] in HFD-fed mice. Furthermore, APO10LA suppressed Akt activation, cyclin D1 gene, and protein expression and promoted PARP protein cleavage in transformed cells within liver tumors. Taken together, these data indicate that APO10LA can effectively inhibit HFD-promoted hepatic tumorigenesis by stimulating SIRT1 signaling while reducing hepatic inflammation.

Topics: Alkylating Agents; Animals; Apoptosis; Blotting, Western; Carcinoma, Hepatocellular; Carotenoids; Cell Proliferation; Cell Transformation, Neoplastic; Cyclin D1; Diet, High-Fat; Diethylnitrosamine; Fatty Acids, Unsaturated; Female; Glucose Tolerance Test; Humans; Inflammation; Liver Neoplasms; Lycopene; Mice; Mice, Inbred C57BL; Poly (ADP-Ribose) Polymerase-1; Poly(ADP-ribose) Polymerases; Proto-Oncogene Proteins c-akt; Real-Time Polymerase Chain Reaction; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Tumor Cells, Cultured; Xenograft Model Antitumor Assays

To evaluate the effects of oligonol administration on experimentally induced colitis and colonic adenoma formation.. Oral administration of oligonol protected against mouse colitis induced by dextran sulfate sodium (DSS). Under the same experimental conditions, oligonol administration significantly inhibited the activation of nuclear factor-kappa B and signal transducer and activator of transcription (STAT) 3 and expression of cyclooxygenase-2 (COX-2), inducible nitric oxide synthase (iNOS), and cyclin D1 in the mouse colon. Further, oligonol inhibited azoxymethane-initiated and DSS-promoted adenoma formation in the mouse colon. Oligonol administration also attenuated lipid peroxidation (malondialdehyde) and protein oxidation (4-hydroxy-2-nonenal), thereby preventing oxidative stress-induced apoptosis of colonic epithelial cells. In vitro studies demonstrated that oligonol treatment reduced lipopolysaccharide-induced expression of interleukin (IL)-1β, tumor necrosis factor α, il-6, cox-2, and inos in murine macrophage RAW 264.7 cells. In another study, oligonol upregulated the antioxidant gene expression in the intestinal epithelial CCD841CoN cells and in the mouse colon.. Oligonol, an innovative formulation of catechin-type oligomers derived from the lychee fruit extract, was tested in this study for the first time to evaluate its effects on experimentally induced colitis and colonic adenoma formation in mice.. Oligonol is effective in protecting against DSS-induced mouse colitis and colon carcinogenesis, suggesting that this polyphenol formulation may have a potential for the amelioration of inflammatory bowel disease and related disorders.

Topics: Adenoma; Animals; Antioxidants; Apoptosis; Carcinogenesis; Catechin; Cell Line; Colitis; Colonic Neoplasms; Cyclin D1; Cyclooxygenase 2; Dextran Sulfate; Humans; Inflammation; Interleukin-1beta; Lipid Peroxidation; Male; Mice; Mice, Inbred ICR; NF-kappa B; Nitric Oxide Synthase Type II; Oxidative Stress; Phenols; STAT3 Transcription Factor; Tumor Necrosis Factor-alpha

Spinal cord injury (SCI) is a traumatic event that causes a secondary and extended inflammation characterized by infiltration of immune cells, including T lymphocytes, release of pro-inflammatory mediators in the lesion site, and tissue degeneration. Current therapeutic approaches for SCI are limited to glucocorticoids (GC) due to their potent anti-inflammatory activity. GC efficacy resides, in part, in the capability to inhibit NF-κB, T lymphocyte activation, and the consequent cytokine production. In this study, we performed experiments aimed to test the susceptibility of glucocorticoid-induced leucine zipper (GILZ) transgenic (GILZ(TG)) mice, in which GILZ is selectively over-expressed in T lymphocytes, to SCI induction. Consistent with a decreased inflammatory response, GILZ(TG) were less susceptible to SCI as compared to wild-type littermates. Notably, inhibition of NF-κB activation and nuclear translocation, diminished T lymphocytes activation and tissue infiltration, as well as decreased release of cytokines were evident in GILZ(TG) as compared to wild-type mice. Moreover, GILZ(TG) showed a reduced tumor necrosis factor-α, IL-1β, Inductible nitric oxide synthase (iNOS) and nytrotyrosine production, apoptosis, and neuronal tissue damage. Together these results indicate that GILZ mimics the anti-inflammatory effect of GC and represents a potential pharmacological target for modulation of T lymphocyte-mediated immune response in inflammatory disorders, such as SCI.

Topics: Animals; Apoptosis; bcl-2-Associated X Protein; Cyclin D1; Cytokines; Dinoprostone; Disease Models, Animal; Enzyme-Linked Immunosorbent Assay; Gene Expression Regulation; Glial Fibrillary Acidic Protein; In Situ Nick-End Labeling; Inflammation; Mice; Mice, Transgenic; Nitric Oxide Synthase Type II; Peroxidase; Signal Transduction; Spinal Cord Injuries; T-Lymphocytes; Transcription Factors

Mutation of tumor suppressor gene, adenomatous polyposis coli (APC), is the primary molecular event in the development of most intestinal carcinomas. Animal model with APC gene mutation is an effective tool for study of preventive approaches against intestinal carcinomas. We aimed to evaluate the effect of Riccardin D, a macrocyclic bisbibenzyl compound, as a chemopreventive agent against intestinal adenoma formation in APC(Min/+) mice.. APC(Min/+) mice were given Riccardin D by p.o. gavage for 7 weeks. Mice were sacrificed, and the number, size and histopathology of intestinal polyps were examined under a microscope. We performed immunohistochemical staining, western blotting, reverse transcriptase-polymerase chain reaction (RT-PCR) and enzyme-linked immunosorbent assay (ELISA) in intestinal polyps to investigate the mechanism of chemopreventive effect of Riccardin D.. Riccardin D treatment resulted in a significant inhibition of intestinal adenoma formation, showing a reduction of polyp number by 41.7%, 31.1% and 44.4%, respectively, in proximal, middle and distal portions of small intestine. The activity of Riccardin D against polyp formation was more profound in colon, wherein Riccardin D decreased polyp number by 79.3%. Size distribution analysis revealed a significant reduction in large-size polyps (2-3 mm) by 40.0%, 42.5% and 33.3%, respectively, in proximal, middle and distal portions of small intestine, and 77.8% in colon. Histopathological analysis of the intestinal polyps revealed mostly hyperplastic morphology without obvious dysplasia in Riccardin D-treated mice. Molecular analyses of the polyps suggested that the inhibitory effect of Riccardin D on intestinal adenoma formation was associated with its abilities of reduction in cell proliferation, induction of apoptosis, antiangiogenesis, inhibition of the Wnt signaling pathway and suppression of inflammatory mediators in polyps.. Our results suggested that Riccardin D exerts its chemopreventive effect against intestinal adenoma formation through multiple mechanisms including anti-proliferative, apoptotic, anti-angiogenic and anti-inflammatory activity.

Topics: Adenoma; Adenomatous Polyposis Coli Protein; Animals; Apoptosis; beta Catenin; Biological Products; Caspases; Cell Proliferation; Colon; Cyclin D1; Cyclooxygenase 2; Hepatophyta; Inflammation; Intestinal Neoplasms; Intestinal Polyps; Intestine, Small; Mice; Neovascularization, Pathologic; NF-kappa B; Phenyl Ethers; Precancerous Conditions; Signal Transduction; Stilbenes

3,3'Diindolylmethane (DIM), a natural phytochemical, has shown inhibitory effects on the growth and migration of a variety of cancer cells; however, whether DIM has similar effects on vascular smooth muscle cells (VSMCs) remains unknown. The purpose of this study was to assess the effects of DIM on the proliferation and migration of cultured VSMCs and neointima formation in a carotid injury model, as well as the related cell signaling mechanisms.. DIM dose-dependently inhibited the platelet-derived growth factor (PDGF)-BB-induced proliferation of VSMCs without cell cytotoxicity. This inhibition was caused by a G0/G1 phase cell cycle arrest demonstrated by fluorescence-activated cell-sorting analysis. We also showed that DIM-induced growth inhibition was associated with the inhibition of the expression of cyclin D1 and cyclin-dependent kinase (CDK) 4/6 as well as an increase in p27(Kip1) levels in PDGF-stimulated VSMCs. Moreover, DIM was also found to modulate migration of VSMCs and smooth muscle-specific contractile marker expression. Mechanistically, DIM negatively modulated PDGF-BB-induced phosphorylation of PDGF-recptorβ (PDGF-Rβ) and the activities of downstream signaling molecules including Akt/glycogen synthase kinase(GSK)3β, extracellular signal-regulated kinase1/2 (ERK1/2), and signal transducers and activators of transcription 3 (STAT3). Our in vivo studies using a mouse carotid arterial injury model revealed that treatment with 150 mg/kg DIM resulted in significant reduction of the neointima/media ratio and proliferating cell nuclear antigen (PCNA)-positive cells, without affecting apoptosis of vascular cells and reendothelialization. Infiltration of inflammatory cells was also inhibited by DIM administration.. These results demonstrate that DIM can suppress the phenotypic modulation of VSMCs and neointima hyperplasia after vascular injury. These beneficial effects on VSMCs were at least partly mediated by the inhibition of PDGF-Rβ and the activities of downstream signaling pathways. The results suggest that DIM has the potential to be a candidate for the prevention of restenosis.

Topics: Animals; Apoptosis; Becaplermin; Carotid Artery Diseases; Cell Cycle Checkpoints; Cell Movement; Cell Proliferation; Cells, Cultured; Cyclin D1; Cyclin-Dependent Kinases; G1 Phase; Human Umbilical Vein Endothelial Cells; Humans; Indoles; Inflammation; Male; Mice; Muscle, Smooth, Vascular; Myocytes, Smooth Muscle; Neointima; Phosphorylation; Proliferating Cell Nuclear Antigen; Proto-Oncogene Proteins c-sis; Rats, Sprague-Dawley; Resting Phase, Cell Cycle; Signal Transduction

Radiation-induced brain injury occurs in many patients receiving cranial radiation therapy, and these deleterious effects are most profound in younger patients. Impaired neurocognitive functions in both humans and rodents are associated with inflammation, demyelination, and neural stem cell dysfunction. Here we evaluated the utility of lithium and a synthetic retinoid receptor agonist in reducing damage in a model of brain-focused irradiation in juvenile mice. We found that lithium stimulated brain progenitor cell proliferation and differentiation following cranial irradiation while also preventing oligodendrocyte loss in the dentate gyrus of juvenile mice. In response to inflammation induced by radiation, which may have encumbered the optimal reparative action of lithium, we used the anti-inflammatory synthetic retinoid Am80 that is in clinical use in the treatment of acute promyelocytic leukemia. Although Am80 reduced the number of cyclooxygenase-2-positive microglial cells following radiation treatment, it did not enhance lithium-induced neurogenesis recovery, and this alone was not significantly different from the effect of lithium on this proinflammatory response. Similarly, lithium was superior to Am80 in supporting the restoration of new doublecortin-positive neurons following irradiation. These data suggest that lithium is superior in its restorative effects to blocking inflammation alone, at least in the case of Am80. Because lithium has been in routine clinical practice for 60 years, these preclinical studies indicate that this drug might be beneficial in reducing post-therapy late effects in patients receiving cranial radiotherapy and that blocking inflammation in this context may not be as advantageous as previously suggested.

Topics: Animals; Behavior, Animal; Benzoates; Brain; Cell Differentiation; Cell Proliferation; Cognition Disorders; Cranial Irradiation; Cyclin D1; Cyclooxygenase 2; Dentate Gyrus; Immunohistochemistry; Inflammation; Lithium; Male; Mice; Mice, Inbred C57BL; Neural Stem Cells; Neurogenesis; Neurons; Oligodendroglia; Radiation Injuries, Experimental; Stem Cells; Tetrahydronaphthalenes

1. Inflammation-induced proliferation of cardiac fibroblasts plays an important role in cardiac remodelling. Pharmacological doses of exogenous glucocorticoids (GC) are the most effective therapy for inflammatory diseases. Similarly, physiological concentrations of endogenous GC have recently been shown to have anti-inflammatory effects. Therefore, the aim of the present study was to determine whether a physiological concentration of GC could inhibit pro-inflammatory cytokine-stimulated proliferation of cardiac fibroblasts and to explore the mechanisms involved. 2. Cardiac fibroblasts were isolated from adult male Sprague-Dawley rats and cell proliferation was measured using a CCK-8 kit. Western blotting was used to detect protein expression of extracellular-regulated kinase (ERK) 1/2 and nuclear factor (NF)-κB. 3. Cardiac fibroblast proliferation was significantly increased by tumour necrosis factor-α, interleukin (IL)-1β and angiotensin II and was accompanied by upregulated protein expression of ERK1/2 and NF-κB. A physiological concentration of hydrocortisone (127 ng/mL) not only inhibited the proliferation of cardiac fibroblasts, but also suppressed activation of ERK1/2 and NF-κB. These effects of hydrocortisone were abrogated by the glucocorticoid receptor (GR) antagonist RU-486 (100 nmol/L). Furthermore, inflammation-induced cardiac fibroblast proliferation was also blocked by the mitogen-activated protein kinase kinase 1/2 inhibitor U0126 (100 nmol/L) and the NF-κB inhibitor pyrrolidine dithiocarbamate (1 μmol/L). Cytokine-induced ERK1/2 phosphorylation and cyclin D1 expression were attenuated by U0126, suggesting that the ERK1/2 and NF-κB signalling pathways were involved in cardiac fibroblast proliferation. 4. In conclusion, the results of the present study indicate that a physiological concentration of hydrocortisone can inhibit inflammation-induced proliferation of cardiac fibroblasts by preventing the activation of ERK1/2 and NF-κB.

Topics: Angiotensin II; Animals; Cell Growth Processes; Cells, Cultured; Cyclin D1; Hydrocortisone; Inflammation; Inflammation Mediators; Interleukin-1beta; Male; MAP Kinase Signaling System; Mitogen-Activated Protein Kinase 1; Mitogen-Activated Protein Kinase 3; Myocardium; Myofibroblasts; NF-kappa B; Rats; Rats, Sprague-Dawley; Receptors, Glucocorticoid; Signal Transduction; Tumor Necrosis Factor-alpha; Up-Regulation

Signals generated by the dynamic mechanical strain critically regulate endothelial cell proliferation and angiogenesis; however, the molecular basis remains unclear. We investigated the mechanisms by which human dermal microvascular endothelial cells (HDMECs) perceive mechanical signals and relay them intracellularly to regulate gene expression and endothelial cell proliferation. HDMECs were exposed to low/physiologic levels of dynamic strain and probed for the differential activation/inhibition of kinases in the mechanosignaling cascade associated with endothelial cell gene activation. Because angiogenesis is important at inflammatory sites, we also assessed the mechanisms of mechanosignaling in the presence of an proinflammatory cytokine IL-1beta. In this article, we demonstrate that the mechanosignaling cascade is initiated by vascular endothelial growth receptor-2 (VEGFR2) activation. Mechanoactivation of VEGFR2 results in its nuclear translocation and elevation of PI3K-dependent Ser473-Akt phosphorylation. Subsequently, activated Akt inactivates the kinase activity of the serine/threonine kinase, glycogen synthase kinase-3beta (GSK3beta), via its Ser9 phosphorylation. Thus, inactive GSK3beta fails to phosphorylate cyclin D1 and prevents its proteosomal degradation and, consequently, promotes endothelial cell survival and proliferation. In the presence of IL-1beta, cyclin D1 is phosphorylated and degraded, leading to inhibition of cell proliferation. However, mechanical signals repress cyclin D1 phosphorylation and upregulate cell proliferation, despite the presence of IL-1beta. The data indicate that the VEGFR2/Akt/GSK3beta signaling cascade plays a critical role in sensing and phospho-relaying mechanical stimuli in endothelial cells. Furthermore, mechanical forces control highly interconnected networks of proinflammatory and Akt signaling cascades to upregulate endothelial cell proliferation.

Topics: Blotting, Western; Cell Proliferation; Cell Survival; Cells, Cultured; Cyclin D1; Endothelial Cells; Gene Expression; Glycogen Synthase Kinase 3; Glycogen Synthase Kinase 3 beta; Humans; Inflammation; Interleukin-1beta; Phosphorylation; Proto-Oncogene Proteins c-akt; Reverse Transcriptase Polymerase Chain Reaction; Signal Transduction; Stress, Mechanical; Tumor Necrosis Factor-alpha; Tyrosine; Up-Regulation; Vascular Endothelial Growth Factor Receptor-2

Inflammation and fibrogenesis are directly related to chronic liver disease progression, including hepatocellular carcinoma (HCC) development. Currently there are few therapeutic options available to inhibit liver fibrosis. We have evaluated the hepatoprotective and anti-fibrotic potential of orally-administered 5'-methylthioadenosine (MTA) in Mdr2(-/-) mice, a clinically relevant model of sclerosing cholangitis and spontaneous biliary fibrosis, followed at later stages by HCC development.. MTA was administered daily by gavage to wild type and Mdr2(-/-) mice for three weeks. MTA anti-inflammatory and anti-fibrotic effects and potential mechanisms of action were examined in the liver of Mdr2(-/-) mice with ongoing fibrogenesis and in cultured liver fibrogenic cells (myofibroblasts).. MTA treatment reduced hepatomegaly and liver injury. α-Smooth muscle actin immunoreactivity and collagen deposition were also significantly decreased. Inflammatory infiltrate, the expression of the cytokines IL6 and Mcp-1, pro-fibrogenic factors like TGFβ2 and tenascin-C, as well as pro-fibrogenic intracellular signalling pathways were reduced by MTA in vivo. MTA inhibited the activation and proliferation of isolated myofibroblasts and down-regulated cyclin D1 gene expression at the transcriptional level. The expression of JunD, a key transcription factor in liver fibrogenesis, was also reduced by MTA in activated myofibroblasts.. Oral MTA administration was well tolerated and proved its efficacy in reducing liver inflammation and fibrosis. MTA may have multiple molecular and cellular targets. These include the inhibition of inflammatory and pro-fibrogenic cytokines, as well as the attenuation of myofibroblast activation and proliferation. Downregulation of JunD and cyclin D1 expression in myofibroblasts may be important regarding the mechanism of action of MTA. This compound could be a good candidate to be tested for the treatment of (biliary) liver fibrosis.

Topics: Adenosine; Animals; ATP Binding Cassette Transporter, Subfamily B; ATP-Binding Cassette Sub-Family B Member 4; Cyclin D1; Cytokines; Disease Progression; Fibroblasts; Fibrosis; Inflammation; Liver; Liver Diseases; Mice; Mice, Transgenic; Proto-Oncogene Proteins c-jun; Thionucleosides

Consumption of green tea polyphenols (GTPs) in drinking water prevents photocarcinogenesis in mice; however, the molecular mechanisms underlying this effect have not been fully elucidated. Using IL-12p40 knockout (KO) mice and their wild-type counterparts and an established photocarcinogenesis protocol, we found that although administration of GTPs (0.2%, w/v) in drinking water significantly reduced UVB-induced tumor development in wild-type mice, this treatment had a nonsignificant effect in IL-12-KO mice. GTPs resulted in reduction in the levels of markers of inflammation (cyclooxygenase-2, prostaglandin E(2), proliferating cell nuclear antigen, and cyclin D1) and proinflammatory cytokines (tumor necrosis factor-alpha, IL-6, and IL-1beta) in chronically UVB-exposed skin and skin tumors of wild-type mice but less effective in IL-12p40-KO mice. UVB-induced DNA damage (cyclobutane pyrimidine dimers) was resolved rapidly in GTPs-treated wild-type mice than untreated wild-type mice and this resolution followed the same time course as the GTPs-induced reduction in the levels of inflammatory responses. This effect of GTPs was less pronounced in IL-12-KO mice. The above results were confirmed by treatment of IL-12-KO mice with murine recombinant IL-12 and treatment of wild-type mice with neutralizing anti-IL-12 antibody. To our knowledge, it is previously unreported that prevention of photocarcinogenesis by GTPs is mediated through IL-12-dependent DNA repair and a subsequent reduction in skin inflammation.

Topics: Administration, Oral; Animals; Beverages; Cyclin D1; Cyclooxygenase 2; Cytokines; Dinoprostone; Disease Models, Animal; DNA Repair; Female; Flavonoids; Inflammation; Interleukin-12; Male; Mice; Mice, Inbred C3H; Mice, Inbred C57BL; Mice, Knockout; Neoplasms, Radiation-Induced; Phenols; Plant Extracts; Polyphenols; Proliferating Cell Nuclear Antigen; Skin Neoplasms; Ultraviolet Rays

Grape seed proanthocyanidins (GSPs) possess anticarcinogenic activities. Here, we assessed the effects of dietary GSPs on 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced skin tumor promotion in 7,12-dimethylbenz[a]anthracene (DMBA)-initiated mouse skin. Administration of dietary GSPs (0.2 and 0.5%, wt/wt) supplemented with control AIN76A diet resulted in significant inhibition of TPA-induced skin tumor promotion in C3H/HeN mice. The mice treated with GSPs developed a significantly lower tumor burden in terms of the percentage of mice with tumors (P < 0.05), total number of tumors per group (P < 0.01, n = 20) and total tumor volume per tumor-bearing mouse (P < 0.01-0.001) as compared with the mice that received the control diet. GSPs also delayed the malignant progression of papillomas into carcinomas. As TPA-induced inflammatory responses are used routinely as markers of skin tumor promotion, we assessed the effect of GSPs on biomarkers of TPA-induced inflammation. Immunohistochemical analysis and western blotting revealed that GSPs significantly inhibited expression of cyclooxygenase-2 (COX-2), prostaglandin E(2) (PGE(2)) and markers of proliferation (proliferating cell nuclear antigen and cyclin D1) in both the DMBA-initiated/TPA-promoted mouse skin and skin tumors. In short-term experiments in which the mouse skin was treated with acute or multiple TPA applications, we found that dietary GSPs inhibited TPA-induced edema, hyperplasia, leukocytes infiltration, myeloperoxidase, COX-2 expression and PGE(2) production in the mouse skin. The inhibitory effect of GSPs was also observed against other structurally different skin tumor promoter-induced inflammation in the skin. Together, our results show that dietary GSPs inhibit chemical carcinogenesis in mouse skin and that the inhibition of skin tumorigenesis by GSPs is associated with the inhibition of inflammatory responses caused by tumor promoters.

Topics: 9,10-Dimethyl-1,2-benzanthracene; Animals; Carcinogens; Cyclin D1; Cyclooxygenase 2; Dinoprostone; Female; Grape Seed Extract; Inflammation; Mice; Papilloma; Plant Extracts; Proanthocyanidins; Proliferating Cell Nuclear Antigen; Skin Neoplasms; Tetradecanoylphorbol Acetate

Topics: Administration, Oral; Animals; Beverages; Cyclin D1; Cyclooxygenase 2; Disease Models, Animal; DNA Repair; Flavonoids; Inflammation; Interleukin-12; Mice; Mice, Knockout; Neoplasms, Radiation-Induced; Phenols; Plant Extracts; Polyphenols; Proliferating Cell Nuclear Antigen; Prostaglandins; Skin Neoplasms; Ultraviolet Rays

Interleukin (IL)-12 deficiency exacerbates tumorigenesis in ultraviolet (UV) radiation-induced skin. Here, we assessed the effects of IL-12 deficiency on 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced tumor promotion in 7,12-dimethylbenz(a)anthracene (DMBA)-initiated mouse skin. Using this two-stage chemical carcinogenesis protocol, we found that the development of DMBA/TPA-induced skin tumors was diminished in IL-12p40-knockout mice than in their wild-type counterparts. At the termination of the experiment (at 24 weeks), the skin tumor incidence and tumor multiplicity were significantly lower (P < 0.005) in interleukin-12-knockout (IL-12 KO) mice than in their wild-type counterparts, as was the malignant transformation of DMBA/TPA-induced papillomas to carcinomas (P < 0.01). Analysis of samples collected at the termination of the experiments for biomarkers of inflammation by immunohistochemical analysis, western blotting, enzyme-linked immunosorbent assay and real-time polymerase chain reaction revealed significantly lower levels of cyclooxygenase-2 (COX-2), prostaglandin (PG) E(2), proliferating cell nuclear antigen, cyclin D1 and the proinflammatory cytokines (tumor necrosis factor-alpha, IL-1beta and IL-6) in the DMBA/TPA-treated tumors and tumor-uninvolved skin of IL-12 KO mice than the skin and tumors of DMBA/TPA-treated wild-type mice. Analysis of the skin 6 h after TPA treatment showed that the TPA-induced promotion of skin edema, inflammatory leukocyte infiltration, COX-2 expression and PGE(2) production was significantly lower in the skin of the IL-12-KO mice than their wild-type counterparts. These results indicate that DMBA/TPA-induced skin tumor development differs from UVB-induced skin tumor development in that endogenous IL-12 acts to inhibit UVB-induced skin tumor development and malignant progression of the skin tumors to carcinoma. In the case of DMBA/TPA-induced skin tumor development, the endogenous IL-12 modulates the tumor promoter stimulation of inflammatory responses.

Topics: 9,10-Dimethyl-1,2-benzanthracene; Animals; Carcinogens; Cyclin D1; Cyclooxygenase 2; Cytokines; Female; Inflammation; Interleukin-12 Subunit p40; Interleukin-6; Mice; Mice, Knockout; Proliferating Cell Nuclear Antigen; Skin Neoplasms; Tetradecanoylphorbol Acetate; Tumor Necrosis Factor-alpha

NF-kappaB activity is tightly regulated by IkappaB class of proteins. IkappaB proteins possess ankyrin repeats for binding to and inhibiting NF-kappaB. The regulatory protein, NPR1 from Brassica juncea possesses ankyrin repeats with sequence similarity to IkappaBalpha subgroup. Therefore, we examined whether stably expressed BjNPR1 could function as IkappaB in inhibiting NF-kappaB in human glioblastoma cell lines. We observed that BjNPR1 bound to NF-kappaB and inhibited its nuclear translocation. Further, BjNPR1 expression down-regulated the NF-kappaB target genes iNOS, Cox-2, c-Myc and cyclin D1 and reduced the proliferation rate of U373 cells. Finally, BjNPR1 decreased the levels of pERK, pJNK and PKCalpha and increased the Caspase-3 and Caspase-8 activities. These results suggested that inhibition of NF-kappaB activation by BjNPR1 can be a promising therapy in NF-kappaB dependent pathologies.

Topics: Active Transport, Cell Nucleus; Amino Acid Sequence; Ankyrin Repeat; Cell Line, Tumor; Cell Proliferation; Cyclin D1; Cyclooxygenase 2; Down-Regulation; eIF-2 Kinase; Gene Expression; Gene Expression Regulation; Genes, Reporter; Humans; I-kappa B Proteins; Inflammation; Molecular Sequence Data; Mustard Plant; Neoplasms; NF-kappa B; Nitric Oxide Synthase Type II; Plant Proteins; Protein Kinase C-alpha; Proto-Oncogene Proteins c-myc

Apurinic/apyrimidinic endonuclease 1/redox factor-1 (APE1) functions in both DNA repair and redox signaling, making it an attractive emerging therapeutic target. However, the role of APE1 in cutaneous inflammatory responses is largely unknown. In this study, we report that APE1 is a key upstream regulator in TLR2-dependent keratinocyte inflammatory responses. We found that nuclear expression of APE1 in epidermal layers was markedly up-regulated in psoriatic skin. APE1 was essential for the transcriptional activation and nuclear translocation of hypoxia-inducible factor-1alpha and NF-kappaB, both of which are crucial for inflammatory signaling in keratinocytes. Moreover, APE1 played a crucial role in the expression of TLR2-mediated inflammatory mediators, including TNF-alpha, CXCL8, and LL-37, in HaCaT cells and human primary keratinocytes. Silencing of APE1 attenuated cyclin D1/cyclin-dependent kinase 4 expression and phosphorylation of ERK1/2 and Akt, thereby affecting keratinocyte proliferation. Importantly, TLR2-induced generation of reactive oxygen species contributed to the nuclear translocation and expression of APE1, suggesting an autoregulatory circuit in which the subcellular localization of APE1 is associated with the production of APE1 per se through reactive oxygen species-dependent signaling. Taken together, these findings establish a role for APE1 as a master regulator of TLR2-dependent inflammatory responses in human keratinocytes.

Topics: Antimicrobial Cationic Peptides; Apoptosis; Cathelicidins; Cell Line; Cyclin D1; Cyclin-Dependent Kinase 4; DNA-(Apurinic or Apyrimidinic Site) Lyase; Epidermis; Extracellular Signal-Regulated MAP Kinases; Humans; Hypoxia-Inducible Factor 1; Inflammation; Interferon Inducers; Interleukin-8; Keratinocytes; NF-kappa B; Poly I-C; Proto-Oncogene Proteins c-akt; Psoriasis; Reactive Oxygen Species; RNA, Small Interfering; Signal Transduction; Toll-Like Receptor 2; Transcriptional Activation; Transfection; Tumor Necrosis Factor-alpha; Up-Regulation; Zymosan

IL-12 deficiency has been shown to promote photocarcinogenesis in mice. As UVB-induced inflammation is an important tumor-promoting event in the development of skin tumors, we determined the effects of IL-12-deficiency on UVB-induced inflammatory responses in mice. For this purpose, IL-12-knockout (IL-12 KO) and their wild-type counterparts were subjected to a photocarcinogenesis protocol; skin and tumor samples were collected at the termination of the experiment, and analyzed for biomarkers of inflammation and their mediators. We found that the levels of infiltrating leukocytes, myeloperoxidase, proliferating cell-nuclear antigen (PCNA), COX-2, PGE2, and the proinflammatory cytokines IL-1beta, TNF-alpha, and IL-6 were higher in the UVB-exposed skin of IL-12 KO than in that of wild-type mice. In a short-term experiment, pretreatment of IL-12 KO mice with rIL-12 (50 ng per mouse) before each exposure to UVB increased the repair rate of UVB-induced cyclobutane pyrimidine dimers, while inhibiting UVB-induced increases in myeloperoxidase, COX-2, PGE2, PCNA, TNF-alpha, and IL-1beta in the skin as compared with non-rIL-12-treated IL-12 KO mice. Similarly, tumors of IL-12 KO mice expressed higher levels of inflammatory responses than those of wild-type mice. Together, our data suggest that IL-12 KO mice are more susceptible to both UVB-induced inflammation and photocarcinogenesis because of the deficiency in the repair of UVB-induced DNA damage.

Topics: Animals; Cell Movement; Cyclin D1; Cyclooxygenase 2; Dinoprostone; DNA Damage; Female; Genetic Predisposition to Disease; Inflammation; Interleukin-12; Leukocytes; Mice; Mice, Knockout; Neoplasms, Radiation-Induced; Proliferating Cell Nuclear Antigen; Skin; Skin Neoplasms

Topics: Animals; Chemistry, Pharmaceutical; Cyclin D1; Drug Delivery Systems; Inflammation; Leukocytes; Mice; Nanoparticles; RNA, Small Interfering

Obesity is considered a chronic low-grade inflammatory state. The white adipose tissue produces a variety of inflammation-related proteins whose expression is increased in obese subjects. The nonadipose cell fraction, which includes infiltrated macrophages, is a determinant source of inflammation-related molecules within the adipose tissue. Our working hypothesis is that macrophage infiltration affects fat expansion through a paracrine action on adipocyte differentiation. Human primary preadipocytes were then differentiated in the presence of conditioned media obtained from macrophages differentiated from blood monocytes. Preadipocytes treated by macrophage-conditioned medium displayed marked reduction of adipogenesis as assessed by decreased cellular lipid accumulation and reduced gene expression of adipogenic and lipogenic markers. In addition to this effect, the activation of macrophages by lipopolysaccharides stimulated nuclear factor kappaB signaling, increased gene expression and release of proinflammatory cytokines and chemokines, and induced preadipocyte proliferation. This phenomenon was associated with increased cyclin D1 gene expression and maintenance of the fibronectin-rich matrix. Anti-TNFalpha neutralizing antibody inhibits the inflammatory state of preadipocytes positioning TNFalpha as an important mediator of inflammation in preadipocytes. Strikingly, conditioned media produced by macrophages isolated from human adipose tissue exerted comparable effects with activated macrophages, i.e. decreased adipogenesis and increased inflammatory state in the preadipocytes. These data show that macrophage-secreted factors inhibit the formation of mature adipocytes, suggesting possible role in limiting adipose tissue expansion in humans.

Topics: Adipocytes; Adipogenesis; Adipose Tissue; Biomarkers; Cell Differentiation; Cell Proliferation; Cells, Cultured; Chemokines; Culture Media; Cyclin D1; Cytokines; Female; Gene Expression; Humans; Inflammation; Inflammation Mediators; Lipid Metabolism; Lipopolysaccharides; Macrophages; NF-kappa B; Signal Transduction; Stem Cells

Tyrosine kinase receptors such as members of the epidermal growth factor receptor family and their respective ligands are frequently overexpressed in pancreatic cancer as well as in chronic pancreatitis. In this study, the role of ErbB2 in the exocrine pancreas was examined by ectopic overexpression under the control of the proximal rat elastase promoter. Three independent transgenic mouse lines overexpressing ErbB2 were established by pronuclear injection. Pancreatic mRNA and protein levels were analyzed by real time PCR, immunohistochemistry and immunoblot analysis, RAS activity by using a specific immunoprecipitation assay and various kinase activities by phosphospecific antibodies. Overexpression of ErbB2 in the exocrine pancreas resulted in increased RAS activity and downstream activation of ERK1/2, but not in transgenic increased proliferation of acinar and ductal cells. At later timepoints, some mice showed focal areas of acinar cell damage with upregulated mRNA levels for Cyclin D1 and p16(INK4a). Despite the increased mRNA level, cyclin D1 protein levels were downregulated. We observed areas of focal infiltrations with inflammatory cells interspersed in the exocrine pancreas. NF-kappaB activity was induced in transgenic acinar cells compared with controls contributing to high levels of chemokine and cytokine gene expression such as CCR-1 and CCL3. These data suggest that overexpression of ErbB2 in acinar cells leads to increased RAS activity without cell cycle progression and mediates inflammation via NF-kappaB. We conclude that the biological response of ErbB2/RAS signaling depends highly on cellular context.

Topics: Animals; Blotting, Western; Cell Proliferation; Cyclin D1; Cyclin-Dependent Kinase Inhibitor p16; Gene Expression Profiling; Gene Expression Regulation; Humans; Immunohistochemistry; Inflammation; Mice; Mice, Inbred C57BL; Mice, Inbred DBA; Mice, Transgenic; Mitogen-Activated Protein Kinase 1; Mitogen-Activated Protein Kinase 3; NF-kappa B; Oncogene Proteins v-rel; Pancreas, Exocrine; ras Proteins; Rats; Receptor, ErbB-2; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger

Silibinin, a flavanone from milk thistle, inhibits the growth of tumors in several rodent models. We examined the effects of dietary silibinin on the growth, progression, and angiogenesis of urethane-induced lung tumors in mice.. A/J mice (15 per group) were injected with urethane (1 mg/g body weight) or saline alone and fed normal diets for 2 weeks, after which they were fed diets containing different doses of silibinin (0%-1% [wt/wt] silibinin) for 18 or 27 weeks. Immunohistochemistry and Western blot analysis were used to examine angiogenesis and enzymatic markers of inflammation, proliferation, and apoptosis. All statistical tests were two-sided.. Urethane-injected mice exposed to silibinin had statistically significantly lower lung tumor multiplicities than urethane-injected mice fed the control diet lacking silibinin (i.e., control mice). Mice that received urethane and 1% (wt/wt) dietary silibinin for 18 weeks had 93% fewer large (i.e., 1.5-2.5-mm-diameter) lung tumors than control mice (mean number of tumors/mouse: 27 in the urethane group versus 2 in the urethane + 1% silibinin group, difference = 25 tumors/mouse, 95% confidence interval [CI] = 13 to 37 tumors/mouse, P = .005). Lung tumors of silibinin-fed mice had 41%-74% fewer cells positive for the cell proliferation markers proliferating cell nuclear antigen and cyclin D1 than lung tumors of control mice. Tumor microvessel density was reduced by up to 89% with silibinin treatment (e.g., 56 microvessels/400x field in tumors from control mice versus 6 microvessels/400x field in tumors from urethane + 1% silibinin-treated mice [difference = 50 microvessels/400x field, 95% CI = 46 to 54 microvessels/400x field; P<.001]). Silibinin decreased lung tumor expression of vascular endothelial growth factor (VEGF) and of inducible nitric oxide synthase and cyclooxygenase-2, two enzymes that promote lung tumor growth and progression by inducing VEGF expression.. Silibinin inhibits lung tumor angiogenesis in an animal model and merits investigation as a chemopreventive agent for suppressing lung cancer progression.

Topics: Animals; Antineoplastic Agents; Apoptosis; Biomarkers, Tumor; Blotting, Western; Carcinogens; Cell Proliferation; Cyclin D1; Cyclooxygenase 2; Disease Models, Animal; Fibroblast Growth Factor 2; Immunohistochemistry; Inflammation; Lung Neoplasms; Mice; Microcirculation; Neovascularization, Pathologic; Nitric Oxide Synthase Type II; Platelet Endothelial Cell Adhesion Molecule-1; Proliferating Cell Nuclear Antigen; Silybin; Silymarin; Urethane; Vascular Endothelial Growth Factor A

Rho family GTPases and STAT3 act as mediators of cytokine and growth factor signaling in a variety of cellular functions involved in inflammation, tumorigenesis, and development. In the course of searching for their functional connections, we found by using STAT3 knock-out mouse embryonic fibroblasts that RhoA, Rac1, and Cdc42 could cause nonspecific activation of STAT3 promoter-driven luciferase reporter in the absence of STAT3, raising concerns to a body of literature where STAT3 was associated with Rho GTPases based on the reporter system. We also found that although active RhoA, Rac1, and Cdc42 could all mediate Ser-727 and Tyr-705 phosphorylation and nuclear translocation of STAT3, the Rho GTPases were able to induce STAT3 activation independently of the interleukin-6 autocrine pathway, and active RhoA, Rac1, or Cdc42 could not form a stable complex with STAT3 as previously suggested, indicating an unappreciated mechanism of STAT3 activation by the Rho GTPases. The RhoA-induced STAT3 activation partly depended on Rho-associated kinase (ROK) and involved multiple effector signals as revealed by the examination of effector domain mutants of RhoA. Genetic deletion of STAT3 led to a loss of response to RhoA in myosin light chain phosphorylation and actin stress fiber induction but sensitized the cells to RhoA or ROK-stimulated cell migration. STAT3 was required for the RhoA-induced NF-kappaB and cyclin D1 transcription and was involved in NF-kappaB nuclear translocation. Furthermore, loss of STAT3 expression inhibited RhoA-promoted cell proliferation and blocked RhoA or ROK induced anchorage-independent growth. These phenotypic changes in STAT3-/- cells could be rescued by reconstituting STAT3 gene. Our studies carried out in STAT3 null cells demonstrate unambiguously that STAT3 represents an essential effector pathway of Rho GTPases in regulating multiple cellular functions including actin cytoskeleton reorganization, cell migration, gene activation, and proliferation.

Topics: Actins; Active Transport, Cell Nucleus; Animals; cdc42 GTP-Binding Protein; Cell Movement; Cell Nucleus; Cell Proliferation; Cells, Cultured; Cyclin D1; Cytoskeleton; DNA-Binding Proteins; DNA, Complementary; Fibroblasts; Gene Deletion; Genes, Reporter; Immunoblotting; Inflammation; Interleukin-6; Luciferases; Mice; Mice, Knockout; Microscopy, Fluorescence; Models, Biological; Mutation; Myosin Light Chains; NF-kappa B; Phosphorylation; Promoter Regions, Genetic; Protein Structure, Tertiary; rac1 GTP-Binding Protein; Retroviridae; rho GTP-Binding Proteins; rhoA GTP-Binding Protein; Serine; Signal Transduction; STAT3 Transcription Factor; Trans-Activators; Tyrosine; Wound Healing

While cell-cycle markers have been used to differentiate benign vs. malignant lesions and to classify malignant lesions, benign keratoses have not been well studied using such markers. We hypothesized that inflammation or irritation of benign keratoses may be related to a shift in the cell cycle. We compared the immunohistochemical staining patterns of 10 seborrheic keratoses (SKs), 10 inflamed seborrheic keratoses (iSKs), and 10 inverted follicular keratoses (IFKs) using antibodies to p53, bcl-1, and bcl-2. Staining with antibodies to p53 was slightly increased in IFKs compared with iSKs or non-inflamed seborrheic keratoses. Bcl-1 staining was similar in all lesions. A population of bcl-2-positive dendritic cells was seen within the epidermal portion of IFKs. Keratinocyte bcl-2 staining was significantly higher in SKs compared with the other two keratoses. Bcl-2 may be increased in SKs as an anti-apoptotic mechanism.

Topics: Biomarkers; Cell Cycle; Cyclin D1; Darier Disease; Humans; Immunohistochemistry; Inflammation; Keratosis; Keratosis, Seborrheic; Proto-Oncogene Proteins c-bcl-2; Staining and Labeling; Tissue Distribution; Tumor Suppressor Protein p53

Inflammatory breast cancer (IBC) is a rare but particularly aggressive form of primary breast cancer. In contrast to noninflammatory breast cancer (non IBC), the molecular alterations underlying IBC are poorly known. We postulated that the kind and frequency of these alterations might differ between IBC and non IBC and account for its particular aggressiveness. We investigated allelic losses associated with primary breast cancer (on chromosome arms 1p, 3p, 6p, 6q, 7q, 8p, 9p, 11p, 11q, 16q, 17p and 17q) by analyzing 71 microsatellite markers in 66 cases of IBC. Loss of heterozygosity (LOH) was frequent, with a mean fractional allelic loss (FAL) index of 52%. Relative to published data on non IBC, allelic loss was particularly frequent at 3p21-p14, 6p, 8p22, 11q, 13q14 and 17q21, suggesting the presence of genes that are markedly altered in IBC. In contrast, the DNA amplification levels of ERBB2, MYC and CCND1, as measured by real-time quantitative PCR, did not differ between IBC and non IBC.

Topics: Breast Neoplasms; Cyclin D1; Female; Genes, erbB-2; Genes, myc; Humans; Inflammation; Loss of Heterozygosity