低氧诱导因子短期活化后诱导 miR-29c表达上调可延缓5／6肾切除大鼠的肾病进展

摘要

目的：探讨适度活化低氧诱导因子（hypoxia‐inducible factor ，HIF）对延缓残肾慢性肾脏病进展的作用及可能机制。方法：雄性SD大鼠采用二步法5／6肾大部切除术建立残肾模型，随机分为L‐mimosine （L‐Mim）治疗组（术后5～12周短期给予脯氨酸羟化酶抑制剂，隔日50 mg／kg腹腔给药）和未治疗残肾组，同时设立假手术对照组。术后12周末处死大鼠留取标本。结果：L‐Mim治疗组大鼠血肌酐水平[（82．4±6．3）比（130．1±24．1）μmol／L ， P＜0．05]、24 h尿蛋白水平[（0．7±0．1）比（1．7±0．5） g／d ，P＜0．05]以及残肾病理改变较未治疗残肾组大鼠有显著改善。miRNA芯片分析结果提示：L‐Mim治疗组肾皮质miR‐29c丰度高于未治疗残肾组，伴 HIF‐1α和 HIF‐2α表达增强。经荧光素酶报告检测系统和体外突变实验明确原肌球蛋白1（TPM1）为miR‐29c靶基因之一。HK2细胞转染pre‐miR‐29c寡核苷酸后可以抑制 TGF‐β1（3 ng／mL ，24 h）诱导的原肌球蛋白水平上调（P＜0．05或0．01）。结论：大鼠残肾肾间质纤维化病变明显并伴miR‐29c水平下调，适度活化 HIF水平可通过上调miR‐29c表达延缓残肾功能恶化。%Objective:To investigate the role and probable mechanism of moderate activation of hypoxia‐inducible factor(HIF) in slowing chronic kidney disease progression of remnant kidney .Methods :Rat models of remnant kidney were established by 5/6 subtotal nephrectomy in male Sprague‐Dawley rats . And then they were randomly allocated to L‐mimosine (L‐Mim ) treatment group ,in which the rats were treated with intraperitoneal injections of L‐Mim during 5‐12 week after operation ,and untreated remnant kidney group .Meanwhile ,sham operated rats were set as control group .All rats were sacrificed at the end of week 12 ,and the specimens were collected .Results:The serum creatinine level in L‐Mim treatment group was lower than that in untreated remnant kidney group(82 .4 ± 6 .3 vs .130 .1 ± 24 .1 μmol/L ,P<0 .05) ,as well as the 24 h Ualb level (0 .7 ± 0 .1 vs .1 .7 ± 0 .5 g/d , P< 0 .05) .And the pathological changes in in L‐Mim treatment group was slightly improved while compared to untreated remnant kidney group .The result of miRNA microarray analysis showed that miR‐29c in renal cortex was up‐regulated in L‐Mim group compared with untreated remnant group and meanwhile the expressions of HIF‐1αand HIF‐2αincreased .Tropomyosin 1 (TPM1) met the sequence criteria for microRNA‐target interaction ,which was later confirmed by luciferase reporter system and mutation test in vitro .HK2 cell transfected with pre‐miT‐29c oligonucleotide could inhibit the tropomyosin up‐regulation induced by TGF‐β1 treatment (3 ng/mL ,24 h) , P<0 .05 or 0 .01 .Conclusions :Renal interstitial fibrosis in rat remnant kidney was significant ,and it was accompanied by the miR‐29c down‐regulation .Moderate activation of HIF level may attenuate the deterioration of renal function by up‐regulating miR‐29c expression .