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Expression and function of NET-1 in human skin squamous cell carcinoma.

ABSTRACT: To evaluate the clinicopathological significance of NET-1 in human skin squamous cell carcinoma (SSCC). The expression of NET-1 and Ki67 protein was detected using immunostaining from 60 SSCC cases, 50 SIN samples and ten normal skin tissues. The vectors expressing NET-1, siRNA NET-1 and shRNA NET-1 were constructed, as well as negative controls (target-off). In transfected A431 cells, the expression of NET-1 was detected by qRT-PCR, Western blot and immunofluorescence staining; the proliferation and migration of cells was evaluated by MTT, flow cytometry, wound healing and transwell chamber assays. The stable cell lines transfected with shRNANET-1 was inoculated in nude mice for in vivo study. (1) The levels of NET-1 were significantly higher in SSCC (96.67 %) and SIN III (93.75 %) than that in SIN I and II (41.18 %), (P < 0.05). NET-1 expression was significantly enhanced in spindle-cell SSCC (75 %) versus other histological types (P < 0.05). (2) The expression of NET-1 in A431 cells transfected with siRNANET-1 or shRNANET-1 was significantly decreased; the proliferation and migration of these cells were obviously inhibited as compared to controls (P < 0.05). (3) The growth of subcutaneous tumors was significantly inhibited associated with reduction in the expression of NET-1 vs. the negative control or untreated group (P < 0.05). The overexpression of NET-1 in tumor cells may be closely related to the malignant phenotype of SSCC. NET-1 RNAi used in this study can specifically and effectively downregulate NET-1 gene expression; thus SSCC proliferation, invasion and tumor growth were attenuated. NET-1 might be one of the potential targets for SSCC therapy.

SUBMITTER: zhang J

PROVIDER: S-EPMC4000423 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:ObjectiveMiR-21 is an oncomir expressed by malignant cells and/or tumor microenvironment components. In this study we focused on understanding the effects of stromal miR-21 on esophageal malignant cells.DesignMiR-21 expression was evaluated in formalin-fixed paraffin-embedded samples from patients with esophageal squamous-cell carcinoma (SCC) by quantitative RT-PCR. MiR-21 tissue distribution was visualized with in situ hybridization. A co-culture system of normal fibroblasts and esophageal cancer cells was used to determine the effects of fibroblasts on miR-21 expression levels, and on SCC cell migration and invasion.ResultsMiR-21 was overexpressed in SCCs, when compared to the adjacent non-tumor tissues (P = 0.0007), and was mainly localized in the cytoplasm of stromal cells adjacent to malignant cells. Accordingly, miR-21 expression was increased in tumors with high versus low stromal content (P = 0.04). When co-cultured with normal fibroblasts, miR-21 expression was elevated in SCC cells (KYSE-30), while its expression was restricted to fibroblasts when co-cultured with adenocarcinoma cells (OE-33 and FLO-1). MiR-21 was detected in conditioned media of cancer cell lines, illustrating the release of this miRNA into the environment. Co-culturing with normal fibroblasts or addition of fibroblast conditioned media caused a significant increase in cell migration and invasion potency of KYSE-30 cells (P<0.0001). In addition, co-culturing cancer cells with fibroblasts and expression of miR-21 induced the expression of the cancer associated fibroblast (CAF) marker S100A4.ConclusionsMiR-21 expression is mostly confined to the SCC stroma and its release from fibroblasts influences the migration and invasion capacity of SCC cells. Moreover, miR-21 may be an important factor in "activating" fibroblasts to CAFs. These findings provide new insights into the role of CAFs and the extracellular matrix in tumor microenvironment formation and in tumor cell maintenance, and suggest miR-21 may contribute to cellular crosstalk in the tumor microenvironment.

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Expression and function of NET-1 in human skin squamous cell carcinoma.

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