论文部分内容阅读
目的探讨12-脂氧化酶(12-LO)对系膜细胞血管紧张素(Ang)Ⅱ1型受体(ATlR)表达的影响。方法用AngⅡ刺激正常和12-LO基因敲除小鼠肾系膜细胞后,观察p38 MAPK活性和细胞外基质(ECM)蛋白的变化。用12-LO的作用产物12(S)-HETE刺激的系膜细胞、转染12-LO基因的系膜细胞和采用显微切割法从正常和12-LO基因敲除小鼠肾脏提取的肾小球来观察AT1R的表达。采用RT-PCR和Western印迹分别检测目标基因mRNA和蛋白的表达。结果AngⅡ的刺激可诱导正常系膜细胞p38 MAPK活性和ECM蛋白表达增高。然而,AngⅡ的刺激不能诱导12-LO基因敲除小鼠系膜细胞p38 MAPK活性和ECM蛋白表达升高。剂量依赖性和时间依赖性实验结果表明12(S)-HETE刺激可引起系膜细胞AT1蛋白水平增高,且AT1R mRNA水平升高有统计学意义(P＜0．01)。敲除肾小球内12-LO基因可有效地降低AT1R mRNA的表达(P＜0．01),转染12-LO基因至系膜细胞使AT1R蛋白和mRNA的表达明显增多(P＜0．01)。结论12-LO可上调系膜细胞AT1R的表达。
Objective To investigate the effect of 12-lipoxygenase (12-LO) on the expression of angiotensin Ⅱ type 1 receptor (AT1R) in mesangial cells. Methods After stimulating the mesangial cells of normal and 12-LO knockout mice with AngⅡ, the changes of p38 MAPK activity and extracellular matrix (ECM) protein were observed. Mesangial cells stimulated with the 12 (S) -HETE, an effector of 12-LO, mesangial cells transfected with the 12-LO gene, and kidney extracted from the kidneys of normal and 12-LO knockout mice using microdissection Pellet to observe AT1R expression. RT-PCR and Western blot were used to detect the expression of target gene mRNA and protein, respectively. Results AngⅡ stimulated p38 MAPK activity and ECM protein expression in normal mesangial cells. However, stimulation with Ang II failed to induce p38 MAPK activity and ECM protein expression in mesangial cells of 12-LO knockout mice. The results of dose-dependent and time-dependent experiments indicated that 12 (S) -HETE stimulation could induce the increase of AT1 protein and the increase of AT1R mRNA in mesangial cells (P <0.01). The knockdown of 12-LO gene in glomerulus could effectively reduce the expression of AT1R mRNA (P <0.01). The transfection of 12-LO gene into mesangial cells increased the expression of AT1R protein and mRNA (P <0.01). 01). Conclusion 12-LO can up-regulate the expression of AT1R in mesangial cells.