亚硝酸钠诱导的草鱼肝细胞凋亡中内质网应激IRE1通路的作用研究

为探究亚硝酸钠诱导草鱼肝细凋亡中内质网应激IRE1通路的作用, 草鱼(Ctenopharyngodon idellus)肝细胞L8824分别置于亚硝酸钠浓度为0、5、20和50 mg/L暴露12h和24h。同时采用三磷酸肌醇受体拮抗剂2-APB和IRE1α抑制剂STF-083010分别和20 mg/L亚硝酸钠共同孵育L8824细胞24h。检测细胞凋亡以及c-Jun氨基末端激酶(c-Jun N-terminal kinase, jnk)、B淋巴细胞瘤-2(B-cell lymphoma-2, bcl-2)、bcl-2相关X蛋白(bcl-2 associated X protein, bax)、caspase9、caspase3、肌醇酶1α (inositol requiring enzyme 1α, ire1α)、X盒结合蛋白 1s (X-box binding protein 1s, xbp1s)和葡萄糖调节蛋白78 (glucose-regulated protein78, grp78)基因的表达和细胞质内钙离子浓度。结果表明: 与对照组相比, 亚硝酸钠处理组细胞凋亡率显著上升, jnk、bax、caspase9、caspase3、ire1α、xbp1s和grp78 mRNA的表达显著升高, bcl-2 mRNA的表达量显著下降, 细胞质内钙离子浓度显著上升。与亚硝酸钠单处理组相比, STF-083010处理组细胞凋亡率显著下降, ire1α、xbp1s、grp78、jnk、bax、caspase3 mRNA表达量显著下降, bcl-2 mRNA表达量显著上升, 2-APB和STF-083010两个处理组细胞质内钙离子浓度均显著下降。结果显示, 高浓度的亚硝酸钠可诱导草鱼肝细胞凋亡和钙离子紊乱, 内质网应激IRE1通路发挥了作用。

ROLE OF ENDOPLASMIC RETICULUM STRESS IRE1 PATHWAY IN HEPATOCYTE APOPTOSIS OF GRASS CARP CTENOPHARYNGODON IDELLA INDUCED BY SODIUM NITRITE

Nitrite, a common pollutant in aquaculture, is an intermediate product of nitrogen cycle in ecosystem. To explore the mechanisms of sodium nitrite-induced cell apoptosis, grass carp liver cell (L8824) were exposed to four concentrations of sodium nitrite (0, 5 mg/L, 20 mg/L and 50 mg/L) with or without treatments of phosphoinositide receptor antagonist 2-APB and IRE1 inhibitors STF-083010. Cell apoptosis related gene expression of jnk, bcl-2, bax, caspase9, caspase3, ire1α, xbp1s and grp78 and the cytoplasmic calcium ion concentration were assessed. The results showed that nitrite significantly increased the apoptosis rate, cytoplasm calcium ion concentration and mRNA levels of jnk, bax, caspase9, caspase3, ire1α, xbp1s and grp78 and significantly decreased bcl-2 mRNA level, which were reversed by the STF-083010 treatment. Besides, both 2-APB and STF-083010 reduced the sodium nitrite-induced cytoplasmic calcium ion. These results indicate that endoplasmic reticulum stress-related IRE1 pathway plays pivotal role nitrite-mediated L8824 cell apoptosis and calcium dyshomeostasis.