Résumé
AIM: To investigate the effects of transient receptor potential channel 6 (TRPC6) on the proliferation of rheumatoid arthritis fibroblast-like synoviocytes (RA-FLS) induced by IL-1β.METHODS: The mRNA expression of TRPC6 in synovial tissues from RA or OA patients was studied by RT-qPCR.RA-FLS were cultured by enzyme digestion and tissue adhesion methods.The method of flow cytometry was applied to identify the RA-FLS.RA-FLS were treated with different concentrations (0, 0.25, 0.5, 1, 2, 4, 8 and 16 μg/L) of IL-1β for 36 h.The cell viability was examined by CCK-8 assay.RA-FLS were incubated with IL-1β (16 μg/L) for different time (12, 24, 36, 48, 60 and 72 h), and the cell viability was measured by CCK-8 assay.The interference efficiency of TRPC6-siRNA was determined by RT-qPCR and Western blotting.After incubation in the presence or absence of IL-1β medium, the cell viability, the percentage of EdU-positive cells and the percentage of (G2/M+S) phase were measured by CCK-8 assay, EdU labeling assay and flow cytometry, respectively.RESULTS: The mRNA expression of TRPC6 was found in synovial tissue with higher levels in RA patients than that in OA patients.TRPC6-siRNA significantly decreased the mRNA and protein expression of TRPC6 (P<0.05).When RA-FLS were treated with IL-1β, the proliferation of RA-FLS was increased (P<0.05).The differences of the cell viability, the percentage of EdU-positive cells and the (G2/M+S) phase percentage between TRPC6-siRNA group and blank control group or NC-siRNA group were significant, in the presence of IL-1β (P<0.05).However, they were not significant in the absence of IL-1β.CONCLUSION: TRPC6 is involved in the proliferation of RA-FLS induced by IL-1β.Silencing of TRPC6 gene inhibits the growth of RA-FLS induced by IL-1β.
Résumé
Objective To explore the expression of transient receptor potential channel 6(TRPC6)in human breast cancer cells, and to clarify the correlation of TRPC6 with the invasion potential of breast cancer cells. Methods The human breast cancer cell strains MCF-7 (hypo-invasion group)and MDA-MB-231 (hyper-invasion group)were cultured.The expressions of TRPC6 mRNA and protein in in two groups were detected by RT-PCR and Western blotting methods.Then the MDA-MB-231 cells were divided into control group and SKF96365 group, the effects of SKF96365 on the invasion ability of MDA-MB-231 cells invitro were explored by wound healing assay and Transwell experiment.Results The results of Western blotting and RT-PCR showed that the expression levels of TRPC6 mRNA and protein in MDA-MB-231 cells were higher than that in MCF-7 cells(P<0.05).The wound healing assay showed the numbers of migrating cells in 5,25 and 40μmol·L-1 SKF96365 groups (76.24±7.54, 45.33±4.50,25.12±1.57)were lower than those in control group (130.48±9.55)(P<0.05).The Transwell experiment results indicated that the invasiveness of MDA-MB-231 cells were inhibited significantly by SKF96365 compared with control group (P<0.05).Conclusion The invasion ability of human breast cancer MDA-MB-231 cells is promoted by upregulating the TRPC6 expression, which indicates that the TRPC6 may play role in the metastasis of human breast cancer.