transforming-growth-factor-beta and Vulvar-Neoplasms

To explore the inhibition of the proliferation of vulvar squamous cell carcinoma (VSCC) by astragaloside IV.. MTT examined the cell proliferation of VSCC. Flow cytometry analyzed cell cycle and apoptosis. Western blot assay detected the expression of some relevant proteins.. AS-IV reduced the proliferation of SW962 cells in a concentration- and time-dependent manner, induced cell-cycle arresting in G0/G1 phase, as demonstrated by the up-regulation of P53 and P21 expression, and the down-regulation of cyclin D1 expression. AS-IV enhanced the expression of Bax and cleaved-caspase 3, and suppressed Bcl-2 and Bcl-xl expression, which resulted in apoptosis increased. Furthermore, the expression of Beclin-1 and LC3-B was upregulated and that of P62 was downregulated, which suggested that AS-IV could increase the incidence of autophagy in SW962 cells. After inhibiting autophagy by 3-methyladenine (3-MA), cell apoptosis decreased upon AS-IV treatment. Similarly, TGF-β1 stimulated SW962 cells, cell proliferation enhanced, and the expression of TGF-βRII and Smad4 was decreased. Furthermore, the expression of proteins that promote apoptosis and autophagy decreased. After AS-IV treatment, the expression levels of the above proteins exhibited the opposite effect.. AS-IV inhibits cell proliferation and induces apoptosis and autophagy through the TGF-β/Smad signaling pathway in VSCC.

Topics: Apoptosis; Autophagy; Carcinoma, Squamous Cell; Cell Line, Tumor; Cell Proliferation; Female; G1 Phase Cell Cycle Checkpoints; Humans; Saponins; Signal Transduction; Smad Proteins; Transforming Growth Factor beta; Triterpenes; Vulvar Neoplasms

MicroRNAs (miRNAs), a class of small non-coding RNA molecules, are associated with a variety of human cancers. Currently, little data are available regarding miRNA expression in vulvar squamous cell carcinoma (VSCC); the mechanism of action of miRNAs in VSCC still requires investigation. The aim of the present study was to investigate the miRNA expression profile in VSCC using a miRCURY™ LNA array. The expression levels of selected miRNAs were quantified by RT-qPCR. The relationship between miR-590-5p expression and clinical pathology was assessed. The expression levels of crucial transforming growth factor-β (TGF-β) and Smad pathway factors were detected. We further investigated the role of miR-590-5p via in vitro studies in the A431 human VSCC cell line. A total of 157 miRNAs showed significantly altered expression in this type of carcinoma. Of particular interest, miR-590-5p, miR-182-5p and miR-183-5p were upregulated, and miR-603, miR-103a-3p and miR-107 were downregulated. A positive relationship was found between miR-590-5p expression and lymph node metastasis. In VSCC, TGFβ1 and TGFβ2 were significantly overexpressed and TGFβRII and Smad4 were significantly underexpressed at both the RNA and protein levels. In A431 cells, overexpression of miR-590-5p promoted proliferation, migration and G1-S phase transition and downregulated TGFβRII. The knockdown of TGFβRII by siRNA promoted malignant behaviours in the A431 cells. In conclusion, we present the miRNA expression profile in VSCC, and our findings suggest that the upregulation of miR-590-5p promotes cellular malignant behaviours via the target gene TGFβRII.

Topics: Aged, 80 and over; Carcinoma, Squamous Cell; Cell Line, Tumor; Female; Gene Expression Profiling; Gene Expression Regulation, Neoplastic; Humans; MicroRNAs; Middle Aged; Neoplasm Metastasis; Protein Serine-Threonine Kinases; Receptor, Transforming Growth Factor-beta Type II; Receptors, Transforming Growth Factor beta; Smad Proteins; Transforming Growth Factor beta; Up-Regulation; Vulvar Neoplasms

Vulvar cancer accounts for about 3-5% of all female genital carcinomas. TGF-P protein is a member of a superfamily of cytokines that regulate cell functions. A correlation between this protein and many neoplastic processes was reported.. In our study we analyzed TGF-β expression in vulvar tumor among patients with diagnosed squamous cell carcinoma (with and without inguinal nodes metastases).. Paraffin embedded blocks obtained from vulvar tissues and inguinal nodes (from 31 patients with vulvar carcinoma FIGO ll-IV) were prepared. Next, the hematoxylin and eosin staining was performed. Monoclonal antibody NCL-TGF-beta was used for immunohistochemical tests.. Higher expression of TGF-beta in cancer cells corresponds to more advanced cancer stages (FIGO). A positive correlation between TGF-beta and metastases, as well as a number of inguinal nodes metastases was observed. The ratio between the number of stained cells in vulvar tumor and of inflammatory cells proved to be higher in FIGO stage III than IV Possibly TGF-beta increase in vulvar tumor contributes to the breakdown of immunological processes limiting cancer progression. Higher TGF-beta expression leads to metastasis in regional lymphatic nodes.. TGF-beta overproduction is observed in vulvar neoplastic processes. In early stages of carcinogenesis TGF-beta inhibits cancer cell proliferation, but in more advanced stages it accelerates cancer progression by inhibiting the immunological response.

Topics: Adult; Aged; Carcinoma, Squamous Cell; Disease Progression; Female; Humans; Immunohistochemistry; Lymphatic Metastasis; Middle Aged; Neoplasm Staging; Transforming Growth Factor beta; Vulva; Vulvar Neoplasms

Numerous investigations show that cytokines have a significant role in the regulation of cell growth. There is also increasing evidence for the role of transcription factors in cytokine-mediated growth-regulation of cancer cells. Our previous data demonstrate that several cytokines are able to inhibit DNA synthesis of vulvar carcinoma cells. The aim of this study was to investigate the effect of growth-inhibitory cytokines on the binding activity of transcription factor AP-1 and NF-kappaB in two vulvar carcinoma cell lines UM-SCV-6 and UM-SCV-1A in vitro. The effects of interferon gamma (IFN-gamma), interleukins 10 (IL-10) and 13 (IL-13), transforming growth factor beta(1) (TGF-beta(1)) and tumor necrosis factor alpha (TNF-alpha) on the DNA binding proteins were studied by electrophoretic mobility shift assay (EMSA). Our results showed that NF-kappaB and AP-1 were constitutively activated in both cell lines. The binding activity of NF-kappaB was found to be stimulated by TNF-alpha in both vulvar carcinoma cell lines while no effect on AP-1 was found by any of the cytokines. The binding activity of NF-kappaB was decreased by IL-10 and IL-13 in UM-SCV-1A cells suggesting that the pathway by which TNF-alpha activates NF-kappaB differs from that activated by interleukins.

Topics: Carcinoma, Squamous Cell; Cytokines; Electrophoresis; Female; Humans; Interferon-gamma; Interleukin-10; Interleukin-13; NF-kappa B; Recombinant Proteins; Signal Transduction; Transcription Factor AP-1; Transforming Growth Factor beta; Transforming Growth Factor beta1; Tumor Cells, Cultured; Tumor Necrosis Factor-alpha; Vulvar Neoplasms

Resistance to the growth inhibitory effects of transforming growth factor beta (TGFbeta) has been associated with decreased levels of the TGFbeta type II receptor (TbetaR-II) and has been correlated with tumorigenicity. Previously, we reported an A --> G mutation at position -364 in the TbetaR-II promoter in A431 tumor cells which results in reduced TbetaR-II promoter activity. In this study, we show that the CDP/Cut (CCAAT displacement protein) transcription factor, a transcriptional repressor, binds both the wild type and the mutant TbetaR-II promoter. We also demonstrate that the A --> G mutation increases CDP/Cut binding affinity, and that overexpression of CDP/Cut reduces transcription from TbetaR-II promoter reporter constructs. Increased binding of the CDP/Cut repressor protein, as a result of a mutation at position -364, represents a novel mechanism of regulation in a neoplastic cell of the promoter of a tumor suppressor gene, TbetaR-II.

Topics: Binding Sites; DNA Mutational Analysis; Female; Gene Expression Regulation, Neoplastic; Homeodomain Proteins; Humans; Neoplasms; Nuclear Proteins; Promoter Regions, Genetic; Protein Binding; Protein Serine-Threonine Kinases; Receptor, Transforming Growth Factor-beta Type II; Receptors, Transforming Growth Factor beta; Repressor Proteins; Transcription Factors; Transforming Growth Factor beta; Tumor Cells, Cultured; Vulvar Neoplasms

In a previous report, we observed by light microscopy the extracellular matrix in 51 vulvar squamous carcinomas and found that some tumors has a prominent stromal response in the form of a regional or diffuse zone of extracellular myxoid matrix containing immature collagen and fibroblasts at the tumor-stromal junction. These tumors were associated with clitoral involvement, ulcerative nonexophytic growth pattern, older age groups, poorer survival rate, and more extensive lymph node metastases than when prominent fibromyxoid stromal response (PFSR) was absent. This behavior was demonstrated despite the fact that these tumors were not larger, more deeply invasive, or of higher grade than when PFSR was absent. In the current immunohistochemical study, we examined cytokine, cell adhesion receptor, and tumor suppressor gene expression in 50 vulvar squamous carcinomas using a panel of antibodies to identify any potential role of these proteins in the development of a PFSR. Semiquantification of expression into none, focal (< 25% of cells showing expression), regional (25-50%), and diffuse (> 50%) patterns revealed PFSR to be statistically associated with high CD44, transforming growth factor (TGF) beta 3, and p53 protein expression, but not with fibroblast growth factor, epidermal growth factor, epidermal growth factor receptor, or E-cadherin expression. When expression of CD44 and either stromal or tumor TGF-beta 3 expression was high, i.e., regional or diffuse in distribution, 15 (50%) of 30 cases were associated with PFSR. In contrast, only 1 (7%) of 14 cases was associated with PFSR when expression was high for only one of these two proteins and none of 3 cases was associated with response when expression was low for both proteins (p = 0.005). Furthermore, in cases showing high expression for both TGF-beta 3 and CD44, PFSR was found in 13 (72%) of 18 cases when p53 expression was diffuse compared with 2 (17%) of 12 cases when expression was less (p = 0.01). Since TGF-beta acts mitogenically for fibroblasts and has been shown to be an inhibitor of epithelial cell growth, its high expression in a carcinoma with PFSR would suggest loss of effect on the epithelial component but an intact effect on the stroma. Since CD44 is known to act as a receptor for hyaluronic acid, which is a prominent stromal component and known to play an important role in cell mobility and tumor aggressiveness, its high expression in association with PFSR would suggest a role of CD44 ov

Topics: Carcinoma, Squamous Cell; Collagen; Cytokines; Female; Fibroblasts; Gene Expression; Genes, Tumor Suppressor; Humans; Hyaluronan Receptors; Immunohistochemistry; Integrins; Prognosis; Stromal Cells; Transforming Growth Factor beta; Tumor Suppressor Protein p53; Vulvar Neoplasms

The regulation of parathyroid hormone-related protein (PTH-rP) mRNA levels and immunoreactive (ir)PTH-rP formation by peptide growth factors, particularly transforming growth factor-beta (TGF-beta) and epidermal growth factor (EGF), in squamous cell carcinomas of gynecologic origin is largely unknown.. PTH-rP mRNA levels were evaluated by Northern analysis in A431 cells (derived from a human vulvar epidermoid carcinoma) and ME-180 cells (derived from a human papillomavirus-infected squamous cell carcinoma of the cervix). PTH-rP protein levels in cell culture media were evaluated using both radioimmunoassay and immunoradiometric assay techniques. These results were compared with those from a lung carcinoid cell line known to produce PTH-rP, namely, NCI-H727 cells.. TGF-beta 1 or EGF treatment caused an increase in the levels of PTH-rP mRNA in A431 cells; these increases in PTH-rP mRNA were detectable after 60 minutes of treatment, were maximal at approximately 4-8 hours, and were approximately additive. Immunoreactive PTH-rP was not detectable (using two different PTH-rP immunoassays) in the culture medium or cell sonicates of A431 cells before or after treatment with TGF-beta 1, EGF, or TGF-beta 1 plus EGF. ME-180 cells responded to EGF (but not to TGF-beta 1) with an increase in the level of PTH-rP mRNA as early as 2 hours; irPTH-rP was present (by use of either immunoassay) in the medium of these cells at 8 and 24 hours. In NCI-H727 (human lung carcinoid) cells, TGF-beta 1 and EGF acted alone and synergistically to effect increases in PTH-rP mRNA and the accumulation of irPTH-rP.. TGF-beta 1 and EGF regulation of PTH-rP gene expression in squamous cell carcinomas of gynecologic origin is unique for each cell line studied and different from that in human lung carcinoid cells.

Topics: Base Sequence; Carcinoid Tumor; Carcinoma, Squamous Cell; Epidermal Growth Factor; Female; Gene Expression Regulation, Neoplastic; Genital Neoplasms, Female; Humans; Lung Neoplasms; Molecular Sequence Data; Neoplasm Proteins; Parathyroid Hormone; Parathyroid Hormone-Related Protein; Proteins; RNA, Messenger; Transforming Growth Factor beta; Tumor Cells, Cultured; Uterine Cervical Neoplasms; Vulvar Neoplasms