血管紧张素Ⅱ对胰岛凋亡及Fas/Fas-L表达影响的实验研究

Experiment of study on the effects of angiotensin Ⅱ on apoptosis and Fas/Fas-L expression in islet

目的观察血管紧张素Ⅱ(angiotensinⅡ,AngⅡ)对体外培养犬胰岛细胞凋亡的诱导作用及氯沙坦(losartan)抑制胰岛凋亡和对凋亡相关基因Fas和Fas-L表达的影响。方法将获得的纯化犬胰岛经不同处理培养48h后,采用光镜和透射电镜观察胰岛细胞凋亡情况,原位末端标记法检测胰岛细胞凋亡百分率,免疫组化法和流式细胞术检测胰岛细胞Fas和Fas-L表达。结果体外培养犬胰岛经1.0μmol/L AngⅡ处理48h后,胰岛细胞出现调亡(16.41±3.01)%现象,电镜见细胞皱缩、胞核裂解,电泳显示核酸断裂片呈"梯形"结构,Fas(11.46±1.97)%和Fas-L(14.68±3.51)%表达亦显著增强。氯沙坦组凋亡细胞数减少至(9.25±1.58)%,Fas(6.85±1.89)%和Fas-L(8.16±2.13)%表达减弱。两两比较差异有统计学意义(P<0.05)。结论AngⅡ诱导胰岛细胞凋亡。凋亡由AT1受体介导,Fas和Fas-L参与其凋亡过程。氯沙坦具有胰岛保护作用。

Objective To investigate the effect of angiotensin Ⅱ on canine islet apoptosis in vitro, and observe the influence of losartan on islet apoptosis inhibition and the expressions of apoptosis-relevant genes Fas and Fas-L. Methods Purified canine islets were cultured with different methods for 48 h, then the apoptosis of islets were observed with light microscope, transmission electronic microscope （ TEM ）, and terminal deoxynucleotidyl transferase biotin-dUTP nicv labeling （TUNEL）. The expressions of Fas and Fas-L were examined by immunohistochemistry and flow cytometry. Results After in vitro culture with 1.0 μmol/L angiotensin Ⅱ ,apoptosis appeared in （16.41 ± 3. 01 ）% islets. Cell shrinkage and karyorrhexis were also found in the islets. Electrophoresis analysis showed that nucleic acid fragmentation took on trapezoid-like shape. The expressions of Fas （ 11.46 ± 1.97 ） % and Fas-L （ 14.68 ± 3.51 ） % were improved significantly. In the group of islets treated with losartan, the apoptotic islets reduced to （9.25 ± 1.58 ） %, the expressions of Fas / Fas-L reduced to （6.85 ± 1.89） % and （ 8.16 ± 2.13） %. There is a significant difference between each two groups（P 〈 0.05 ）. Conclusion Angiotensin Ⅱ could induce islet apoptosis. The apoptosis is mediated by AT1 and Fas and Fas-L take part in the apoptotic procedure. Losartan shows protective effect on islets.