胱硫醚-γ-裂解酶在三种微血管内皮细胞上的表达

Expression of Cystathionine γ-lyase in Three Kinds of Microvascular Endothelial Cells

目的鉴定微血管内皮细胞上是否表达生成硫化氢(H2S)的酶,为深入研究硫化氢的心血管作用提供新线索。方法原代培养大鼠心脏微血管内皮细胞(CMEC),设计相应引物后用RT-PCR的方法检测胱硫醚-γ-裂解酶(CSE)和胱硫醚-β-合成酶(CBS)基因转录情况,进一步用兔抗大鼠CSE和CBS多克隆抗体对CMEC进行蛋白水平的免疫荧光染色。此外对另两种微血管内皮细胞株RF/6A和bEnd.3也检测了H2S生成酶的表达。结果三种微血管内皮细胞都扩增出CSE基因的特异条带,而CBS基因都未扩增出相应产物。CMEC的免疫荧光染色显示CSE抗体染色为阳性,CBS抗体染色为阴性。结论首次发现了微血管内皮细胞上存在CSE的表达,提示内皮细胞可以通过生成硫化氢调节心血管活动。

Objective To identify whether microvascular endothelial cells express H2 S-generating enzymes for further study of H2 S on cardiovascular effects. Methods Primary rat cardiac microvascular endothelial cells （CMEC） were cultured. Appropriate primers were designed to analyze cystathionine-γ-lyase （CSE） and cystathioninc-β-synthase （CBS） mRNA expression using RT-PCR. For immunofluorescence studies, cells were incubated with rabbit polyclonal antibodies against CBS or CSE. Another two microvascular endothelial cell lines RF/6A and bEnd. 3 were also examined for mRNA expression of H2S-generating enzymes. Results Using RT-PCR, the fragments of CSE, but not CBS, were detected in all three kinds of endothelial cells. For immunofluorescence staining, CMEC was positively stained with antibody against CSE while CBS staining was negative. Conclusion The present study provides the first evidence of expression of CSE but not CBS in mi- crovascular endothelial cells. Endothelial cells may participate in cardiovascular regulation by generating H2S.