诱导HEK293细胞增殖中钙离子及TRPC6的变化及意义

目的建立AngⅡ诱导人胚肾细胞增殖模型,探讨增殖效果与Ca2+变化及TRPC6表达的关系。方法用不同浓度AngⅡ诱导细胞,在不同时间(0,6,12,24,48 h),予LOS干预24 h后用激光扫描共聚焦显微镜测Ca2+变化,RT-PCR、Western Blot检测TRPC6表达情况。采用MTT法及台盼蓝法检测细胞增殖及活性,确定最适浓度及时间构建增殖模型。结果 AngⅡ10-6mol/L诱导HEK293 24 h后细胞增殖及Ca2+升高明显,LOS干预后细胞增殖及Ca2+明显下降。TRPC6表达(+/-),胞膜、胞质定位不清。结论 HEK293细胞自身无明显表达TRPC6,可利用此表达系统进一步研究外源性TRPC6与Ca2+变化的关系。AngⅡ诱导HEK293细胞增殖与Ca2+升高有关,具有一定的时间和剂量依赖性。

The changes and its significance of Ca~(2+) and TRPC 6 expression in the proliferation of HEK 293 cells

Objective It is to establish HEK293 cell mode induced by Ang Ⅱ and to explore the relationship between cell proliferation and the changes of Ca2+ and TRPC6 expression.Methods The cells were induced by different concentration of Ang Ⅱ and intervened by LOS for 24 h at different time point(0,6,12,24,28 h).Then the changes of Ca2+ were detected by LSCM,the expression of TRPC6 was detected by RT-PCR,Western Blot method,the cell proliferation and activity was detected by MTT method and trypan-blue-stain method to confirm the best concentration and time for model construction.Results The cell proliferation and Ca2+ increase were obvious after inducing by Ang Ⅱ in HEK293 cells,and the changes were inhibit after LOS's intervention.TRPC6 expression was(+/-),the positions of cytoplasm and cytomembrane were not clear.Conclusion HEK293 cells don't have obvious expression of TRPC6,and this expression system can be used for further study on exogenous TRPC6 and Ca2+ change.The induction of Ang Ⅱ may be related with increase of Ca2+ with a time and dosage dependent.