The Roles And Mechanism Of CSE/H₂S System In Breast Cancer Cells

Objective: Endogenous hydrogen sulfide（H2S） may be involved in many physiological and pathological processes, and has important roles in some kinds of tumors. But H₂S is rarely reported in breast cancer,a woman tumor with high incidence. Therefore, this paper explore the roles and mechanism of endogenous H₂S in breast cancer cells.Methods:Western blot was used to detect the expression of CSE in normal breast cells MCF-10 A and breast cancer cells MCF-7, MDA-MB-231 cells;Methylene blue method was used to detect the effect of CSE siRNA and CSE inhibitor（PAG）on endogenous hydrogen sulfide production in MCF-7 cells;the effect of CSE siRNA and PAG on cell viability in MCF-7 cells was determined by MTT assay;EdU assay was used to detect the effect of CSE siRNA and PAG on proliferation of MCF-7 cells;flow cytometry was used to test the roles of CSE siRNA and PAG on cell cycle and apoptosis in MCF-7 cells;scratch test was used to detect the migration of MCF-7 cells;RT-PCR and Western blot was used to detect the expression of CSE in MCF-7 cells; Methylene blue method was used to detect the effect of STAT3 siRNA on endogenous H₂S production in MCF-7 cells;Luciferase assay was used to detect the activation of STAT3 on CSE promoter.Results:1.Western blot results showed that CSE was overexpressed in breast cancer MCF-7 cells,compared with normal breast cells MCF-10 A.2.Methylene blue results found that CSE siRNA and PAG inhibited the production of endogenous H₂S in MCF-7 cells.3.MTT results showed that cell viability was inhibited in MCF-7 cells treated with CSE siRNA and PAG.4.EdU assay results showed that CSE siRNA and PAG inhibited proliferation of MCF-7 cells.5.Flow cytometry results showed that MCF-7 cells exposed to CSE siRNA and PAG was arrested in S phase.6.Scratch assay results showed that PAG inhibited MCF-7 cells migration at a time-dependent and concentration-dependent way.7.Flow cytometry results showed that induction by CSE siRNA and PAG didn’t induce apoptosis of MCF-7 cells.8.RT-PCR and Western blot showed that STAT3 siRNA decreased CSE mRNA and CSE protein levels in MCF-7 cells.9.Methylene blue results found that STAT3 siRNA decreased endogenous H₂S levels in MCF-7 cells.10.Luciferase assay showed that STAT3 can activate CSE promoter.Conclusion:1.Downregulation of CSE/H₂S system inhibited growth and proliferation of MCF-7 cells,and arrested cell cycle in S phase.2.Downregulation of CSE / H₂S system inhibited migration of MCF-7 cells.3.Inhibition of CSE/H₂S system had no significant effect on apoptosis of MCF-7 cells.4.STAT3 might regulate the expression of CSE by activating CSE promoter, and consequently affect the production of endogenous H₂S.