| 摘要: |
| 目的 考察黄连化浊胶囊对分泌及Ras相关GTP酶1A（sar-1A）敲除诱导的外被蛋白Ⅱ（COP-Ⅱ）功能障碍胰岛β细胞内质网应激的干预效果。方法 将小鼠胰岛β细胞Min6分为空白组（Min6细胞无任何干预）、阴性对照（NC）组（Min6细胞转染sar-1A-NC-siRNA）及sar-1A siRNA组（Min6细胞转染sar-1A siRNA）。收集sar-1A siRNA转染的Min6细胞，分为空白血清组（用含10%大鼠空白血清的培养基培养），5%、10%、20%黄连化浊胶囊组（分别用含5%、10%、20%黄连化浊胶囊含药血清的培养基培养），以及罗格列酮组（用含10%罗格列酮含药血清的培养基培养）。采用qPCR检测Min6细胞中sar-1A miRNA的表达、免疫荧光法检测sec31蛋白的表达，蛋白质印迹法检测胰岛素原、磷酸化真核翻译起始因子2α（p-eIF2α）、C/EBP同源蛋白（CHOP）、X盒结合蛋白1（XBP1）、肌醇需求酶1（IRE1）、X盒（X-box）蛋白的表达。结果 空白组与NC组Min6细胞中sar-1A mRNA、sec31蛋白及胰岛素原、p-eIF2α、CHOP、XBP1、IRE1、X-box蛋白表达差异均无统计学意义（P均>0.05）；与NC组相比，sar-1A siRNA组Min6细胞中sar-1A mRNA、sec31蛋白及胰岛素原和X-box蛋白表达均降低（P均< 0.05），p-eIF2α、CHOP、XBP1、IRE1蛋白表达均升高（P均<0.05）。与空白血清组相比，5%、10%、20%黄连化浊胶囊组及罗格列酮组Min6细胞中sar-1A mRNA表达均升高（P均<0.05），5%、10%、20%黄连化浊胶囊组Min6细胞中sar-1A mRNA表达逐渐升高，各组间差异均有统计学意义（P均<0.05）。空白血清组与5%黄连化浊胶囊组Min6细胞中sec31蛋白及胰岛素原、p-eIF2α、CHOP、XBP1、IRE1、X-box蛋白表达差异均无统计学意义（P均>0.05）；与5%黄连化浊胶囊组相比，10%、20%黄连化浊胶囊组及罗格列酮组Min6细胞中sec31蛋白和胰岛素原表达均增加（P均<0.05），p-eIF2α、CHOP、XBP1、IRE1、X-box蛋白表达均降低（P均<0.05）。结论 sar-1A敲除后胰岛β细胞存在COP-Ⅱ功能障碍，黄连化浊胶囊能够降低sar-1A敲除后胰岛β细胞的内质网应激水平。 |
| 关键词: 胰岛β细胞 黄连化浊胶囊 内质网应激 分泌及Ras相关GTP酶1A |
| DOI：10.16781/j.CN31-2187/R.20220623 |
| 投稿时间：2022-07-25修订日期：2022-09-02 |
| 基金项目:兰州市科技发展计划项目（2022-ZD-89）. |
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| Intervention effects of Huanglian Huazhuo capsule on endoplasmic reticulum stress of islet β cells with coat protein Ⅱ dysfunction induced by secretion associated Ras related GTPase 1A knockdown |
| ZHOU Chunnan,WANG Yuanming* |
(Department of Endocrinology, Affiliated Hospital of Gansu University of Chinese Medicine, Lanzhou 730200, Gansu, China
*Corresponding author) |
| Abstract: |
| Objective To investigate the intervention effects of Huanglian Huazhuo capsule on endoplasmic reticulum stress (ERS) of islet β cells with coat proteinⅡ (COP-Ⅱ) dysfunction induced by secretion associated Ras related GTPase 1A (sar-1A) knockout. Methods Mouse islet β cells Min6 were divided into 3 groups:blank group (Min6 cells without any intervention), negative control (NC) group (Min6 cells were transfected with sar-1A-NC-small interfering RNA[siRNA]), and sar-1A siRNA group (Min6 cells were transfected with sar-1A siRNA). Min6 cells transfected with sar-1A siRNA were collected and divided into blank serum group (cells cultured in medium with 10% blank serum), 5%, 10%, and 20% Huanglian Huazhuo capsule groups (cells cultured in medium with 5%, 10%, and 20% serum containing Huanglian Huazhuo capsule), and rosiglitazone group (cells cultured in medium with 10% serum containing rosiglitazone). The expression of sar-1A mRNA in Min6 cells was detected by quantitative polymerase chain reaction, the expression of sec31 protein was detected by immunofluorescence, and the expression of proinsulin, phosphorylated eukaryotic translation initiation factor 2α (p-eIF2α), C/EBP homologous protein (CHOP), X-box binding protein 1 (XBP1), inosital-requiring enzyme 1 (IRE1), and X-box proteins was detected by Western blotting. Results There were no significant differences in sar-1A mRNA, sec31, proinsulin, p-eIF2α, CHOP, XBP1, IRE1, or X-box protein between the blank group and the NC group (all P>0.05). Compared with the NC group, the sar-1A siRNA group showed decreased expression of sar-1A mRNA, sec31 protein, proinsulin, and X-box protein (all P<0.05), and increased expression of p-eIF2α, CHOP, XBP1, and IRE1 proteins (all P<0.05). Compared with the blank serum group, the sar-1A mRNA expression in the 5%, 10%, and 20% Huanglian Huazhuo capsule groups and the rosiglitazone group was significantly increased (all P<0.05). The expression of sar-1A mRNA was gradually increased in the 5%, 10%, and 20% Huanglian Huazhuo capsule groups (all P<0.05). There was no significant difference in the expression of sec31, proinsulin, p-eIF2α, CHOP, XBP1, IRE1, or X-box protein between the blank serum group and the 5% Huanglian Huazhuo capsule group (all P>0.05). Compared with the 5% Huanglian Huazhuo capsule group, sec31 protein and proinsulin in Min6 cells of 10%, 20% Huanglian Huazhuo capsule groups and rosiglitazone group were significantly increased (all P<0.05), and p-eIF2α, CHOP, XBP1, IRE1, and X-box proteins were significantly decreased (all P<0.05). Conclusion COP-Ⅱ dysfunction is found in the islet β cells after sar-1A knockdown, and Huanglian Huazhuo capsule can reduce ERS of islet β cells after sar-1A knockdown. |
| Key words: islet β cell Huanglian Huazhuo capsule endoplasmic reticulum stress secretion associated Ras related GTPase 1A |
