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目的探讨胆碱能通路对急性缺糖缺氧性肾小管细胞损伤的影响。方法分离培养大鼠肾内巨噬细胞,构建巨噬细胞与肾小管上皮细胞共培养体系及缺糖缺氧(OGD)细胞模型,据处理不同将细胞分为OGD组、乙酰胆碱(ACh 100μmol/L)+OGD组和ACh+加兰他敏(Gal 10μmol/L)+OGD组。采用ELISA法检测各组上清液肿瘤坏死因子α(TNF-α)、白细胞介素-1β(IL-1β)和IL-10的表达;MTT法检测肾小管细胞活力;RT-qPCR及Western blotting检测胆碱酯酶(AChE)m RNA和蛋白表达;比色法检测AChE活性。结果 ACh+OGD组TNF-α和IL-1β水平均低于OGD组,加入Gal之后,TNF-α和IL-1β水平进一步下降;ACh+OGD组肾小管活力高于OGD组,加入Gal之后,肾小管活力进一步增强,差异有统计学意义(P<0.05或P<0.01)。3组之间巨噬细胞ACh E m RNA和蛋白表达差异均无统计学意义(P>0.05)。与OGD组比较,ACh+OGD组与ACh+Gal+OGD组肾小管细胞活力减弱,但ACh+OGD组与ACh+Gal+OGD组间差异无统计学意义(P=0.368)。结论 ACh和Gal可抑制肾脏巨噬细胞分泌炎性介质并抑制胆碱酯酶活性,减轻急性缺氧性肾小管细胞损伤。调控巨噬细胞的胆碱能通路可能是未来急性缺氧性肾损伤的治疗方向。
Objective To investigate the effects of cholinergic pathway on renal tubular cell injury induced by acute hypoglycemia and hypoxia. Methods Rat renal macrophages were isolated and cultured to construct macrophages and renal tubular epithelial cells co-culture system and hypoglycemic hypoxia (OGD) cell model. The cells were divided into OGD group, ACh 100μmol / L ) + OGD group and ACh + galantamine (Gal 10μmol / L) + OGD group. The expressions of TNF-α, IL-1β and IL-10 in supernatants were detected by ELISA. The viability of renal tubular cells was detected by MTT assay. The expressions of TNF-α, IL- The AChE mRNA and protein were detected by colorimetric assay. AChE activity was detected by colorimetric assay. Results The levels of TNF-α and IL-1β in ACh + OGD group were lower than those in OGD group. The levels of TNF-α and IL-1β in ACh + OGD group were lower than those in OGD group. Tubular viability further increased, the difference was statistically significant (P <0.05 or P <0.01). There were no significant differences in ACh E m RNA and protein expression between macrophages in three groups (P> 0.05). Compared with OGD group, the viability of renal tubular cells in ACh + OGD group and ACh + Gal + OGD group decreased, but there was no significant difference between ACh + OGD group and ACh + Gal + OGD group (P = 0.368). Conclusion ACh and Gal can inhibit the secretion of inflammatory mediators and inhibit the activity of cholinesterase in renal macrophages and reduce the damage of acute hypoxic renal tubular cells. Cholinergic pathways regulating macrophages may be the treatment of acute hypoxic renal injury in the future.