Intermedin及其受体系统在单侧输尿管梗阻大鼠肾组织的表达

Expression of intermedin and its receptors in renal tissues of rats with unilateral ureteral obstruction( UUO)

目的观察大鼠单侧输尿管梗阻(UUO)模型中肾组织intermedin(IMD)及其受体系统-降钙素受体样受体(CRLR)/受体活性修饰蛋白(RAMP)表达的变化。方法健康雄性Wistar大鼠48只,随机分为假手术组和UUO组,分别于术后1、3、7、14 d随机选取6只,留取梗阻侧及假手术侧肾组织。HE、Masson染色观察肾组织病理变化;Real-Time RT-PCR检测肾组织中IMD、CRLR、RAMP1、RAMP2、RAMP3的mRNA表达;免疫组织化学法检测肾组织中IMD、CRLR、RAMP1、RAMP2、RAMP3的蛋白表达。采用析因设计的方差分析进行统计学分析。结果肾脏组织病理染色结果显示,假手术组术后各时间点肾小球、肾小管均未见明显损伤;与假手术组相比,UUO组肾脏病理改变明显,肾间质纤维化程度随梗阻时间延长逐渐加重(P<0.05);Real-Time RT-PCR及免疫组织化学结果显示,IMD、CRLR和RAMP1、RAMP2、RAMP3的mRNA及其蛋白质表达从UUO术后1 d开始升高(与假手术组比较P<0.05),分别于术后3d(RAMP2和RAMP3)和术后7 d(IMD、CRLR和RAMP1)达到高峰,以后逐渐下降。结论 IMD及其受体系统在UUO大鼠肾组织表达上调,提示IMD可能在肾脏间质纤维化中发挥重要作用。

Objective To investigate the expression changes of intermedin （IMD） and its receptor system, namely calcitonin receptor-like receptor （CRLR） and receptor activity-modifying proteins （RAMP1/ 2/3）, in renal tissues of rats with unilateral ureteral obstruction （UUO）. Methods Forty-eight male Wistar rats were randomly divided into UUO group （n = 24 ） and sham-operated group （n = 24 ）. UUO model was made by ligation of left ureter. For sham-operated group, rats were subjected to left flank incisions, and the kidney was exposed, but the ureter was not ligated. Groups of six animals each were killed at 1, 3, 7, and 14 d after operation, respectively. Kidneys were harvested for further analysis. Paraffin-embedded transverse kidney slices were sectioned at 4 μm, and stained with hematoxylin and eosin. For analyzing the degree of tubulointerstitial collagen deposition, sections were stained with Masson trichrome, mRNA expression levels of IMD, CRLR, and RAMPs were detected by real-time reverse transcription-polymerase chanin reaction （RT- PCR）. Protein expression levels of IMD, CRLR, and RAMPs were detected by immunohistochemical staining. The variance analysis of factorial design was used for data analysis. Results Renal morphology showed that no changes were detected in the sham-operated group. In the UUO group, there were significant pathological changes, and the interstitial fibrosis gradually increased with time （P 〈 0.05）. Real-time RT-PCR and immunohistochemical staining revealed that, after UUO operation, the mRNA and protein expression levels of IMD, CRLR, RAMP1, RAMP2, and RAMP3 increased from day 1 （P 〈 0.05, compared with sham group）, and reached peak value at day 3 （RAMP2 and RAMP3 ） or at day 7 （IMD, CRLR and RAMP1 ）, respectively, and then gradually declined. Conclusion Our results indicated that IMD and its receptors were upregulated after UUO, which might play a role in the renal fibrosis.