摘要
目的探讨幼鼠单侧输尿管完全梗阻（completeunilateralureteralobstruction，CUUO）肾脏miRNA-101表达与肾间质纤维化进展的关系。方法选取30只SD雄性幼鼠，年龄（8±1）周，体重（180±20）g，数字随机分为sham组、CUUO7d组和14d组。分别于梗阻后7d和14d处死取其梗阻侧。肾脏，将其切开分为均等的两部分，一部分用于RT-PCR，采用相对定量法检测不同梗阻时间段梗阻侧肾脏中miRNA-101表达。另一部分用于Westernblot及免疫组织化学检测rSMA、E-cadherin的相对定量及定性表达。结果免疫组织化学测定结果显示a-SMA及E-cadherin主要表达于肾间质，呈棕黄色，其中mSMA和E-cadherin在CUUO14d组、UUO7d组及sham组显色依次递减和增强；组织学上可见CUUO7d组及14d组肾小管管腔增大，明显不同于sham组。RT-PCR结果显示miRNA-101相对表达量（folddifference）sham组是CUUO14d组的（14．50±5．12）倍，CU—UO7d组是14d组的（3．45±2．27）倍，三组间差异有统计学意义（P〈O．05）；Westernblot显示梗阻侧肾脏rSMA在CUUO14d组、CUUO7d和sham表达分别为1．22±0．48、0．50±0．06和0．21±0．03，与相应时间段miRNA-101表达成负相关（r=-0．74，P〈0．05）。相反，E-cadherin在CUUO14d组、CUUO7d和sham表达分别为0．24±0．22、0．50±0．44和0．76±0．20。与相应时间段miRNA-101表达成正相关（r=0．88，P％0．05）。结论幼鼠完全梗阻肾脏miRNA-101表达随着梗阻时间的延长而下降，与反映肾脏间质纤维化严重程度的蛋白表达相关，提示miRNA-101的变化可能参与梗阻肾脏纤维化过程的进展。
Objective To explore the correlation of miRNA-101 expression with the progression of renal interstitial fibrosis in young rats with complete unilateral ureteral obstruction （CUUO）. Methods Thirty male Sprague Dawley rats （age 8 ± 1 weeks, weight 180 ± 20 g） were randomly divided into 3 groups of sham, CUUO 7 days （D7） and 14 days （I）14）. The kidney samples were collected at Days 7 and 14 respectively and used for detecting the expression of miRNA-101 by real- time polymerase chain reaction （RT-PCR） and α-SMA and E-cadherin protein by Western blot and immunohistochemistry. The correlations between their expressions were analyzed. Results Immunohistochemistry showed that α-SMA and E-cadherin were predominantly expressed in renal interstitiurn. The expression of α-SMA gradually declined in all groups. However there was a gradually elevated expression of E-cadherin. The tubular lumen of D7 and D14 groups expanded more dramatically than that of sham group. RT-PCR results showed that the expression magnitude of MiRNA-101 in sham and D7 groups was （14. 50 ± 5. 12） and （3. 45 ± 2. 27） folds of D14 group respectively. Significant difference existed among 3 groups （P 〈 0. 05）. The expression level of α-SMA protein was 1, 22 ± O. 48, 0. 50 ± 0. 06 and 0. 21 ± 0. 03 in D14, D7 and sham respectively and there were negative correlations with miRNA-101 （r = -0. 74, P〈0. 05）. In contrast, the expression level of E-cadherin was 0. 24 ± 0. 22, 0. 50 ± 0. 44 and 0. 76 ± 0. 20 in D14, D7 and sham respectively and there were positive correlations with miRNA-101 （r = 0. 88, P〈0. 05）. Conclusions The renal expression of miRNA-101 decreases with obstructive duration and it is related with renal interstitial fibrosis proteins. And miRNA-101 may be significantly correlated with the progression of renal interstitial fibrosis.
出处
《中华小儿外科杂志》
CSCD
北大核心
2014年第4期290-295,共6页
Chinese Journal of Pediatric Surgery
基金
基金项目：国家自然基金（81370869）
NSFC-河南人才培养联合基金（U1304803）
