RARs受体在缺氧性RTEC损伤中的作用及分子机制

Injury to renal tubular epithelial cell(RTEC) is the key factor and central link of renal interstitial fibrosis(RIF). Hypoxia is the most common factor of RTEC damage. The abnormal expression of retinoic acid receptors(RARs) is involved in the development and progression of renal disease. Our previous studies had found that the expressions of RARα and RARβ were decreased obviously and may be implicated in the progression of RIF in rats. However, the expression and mechanism of action of RARs in hypoxia-induced RTEC injury remain unknown at present. Our study will be first conducted to explore the function and mechanism, including: 1.the model of hypoxia-induced RTEC injury was established to study the subcellular localization and expressions of RARα、RARβ and RARγ in RTEC. 2.The effect of RARs on hypoxia-induced RTEC injury was studied by detecting the expressions of the three subtype of RARs、transforming growth factor-β1(TGF-β1) and α-smooth muscle actin(α-SMA), and the changes of mitochodria membrane potential、superoxide anioncontent、Ca2+ concentration and superoxide dismutase activity. 3.Construction of virus vector, siRNA silencing endogenous and transfecting exogenous genes of RARα、RARβ and RARγ, and their expressions were intervened by agonist/antagonist, to further explore the role and molecular mechanisms of RARs in hypoxia-induced RTEC injury and provide new ideas for clinical prevention and treatment of RIF.

肾小管上皮细胞（RTEC）损伤是肾间质纤维化（RIF）的关键因素。缺氧是RTEC损伤最常见原因。维甲酸受体（RARs）表达异常参与了肾脏疾病的发生发展。我们前期研究发现，在RIF大鼠中，肾脏RARα、RARβ表达降低可能参与RIF发生发展。但RARs受体在缺氧性RTEC损伤中表达及作用机制目前尚不清楚。本项目首次探讨该作用及机制，内容包括：1.建立缺氧致RTEC损伤模型，检测RARα、RARβ和RARγ等在RTEC中的亚细胞定位及表达。2.检测RARs受体三种亚型、TGF-β1、α-SMA等表达及线粒体膜电位、超氧阴离子、Ca2+浓度和超氧化物歧化酶活力变化，研究RARs受体对RTEC损伤的影响。3.构建病毒载体，siRNA沉默内源性和转染外源性RARα、RARβ和RARγ基因，激动剂/阻断剂干预其表达，探讨RARs受体在缺氧性RTEC损伤中的作用及分子机制，为临床防治RIF提供新思路。

肾小管上皮细胞（RTEC）损伤是肾间质纤维化（RIF）的关键因素。缺氧是RTEC损伤最常见原因之一。维甲酸受体（RARs）表达异常参与了肾脏疾病的发生发展。我们前期研究发现，在RIF大鼠中，肾脏组织RARα和RARβ表达降低可能参与RIF发生发展。但RARs受体在缺氧性RTEC损伤中的表达及作用机制目前尚不清楚。本项目通过建立缺氧致RTEC损伤模型，构建病毒载体，siRNA沉默内源性和转染外源性RARα、RARβ和RARγ基因，检测RARα、RARβ和RARγ、TGF-β1等表达，MMP、O2−、细胞活力、SOD和GSH酶活力变化。结果显示：⑴ 缺氧损伤48 h和72 h的RTEC中，RARα和RARγ的mRNA和蛋白表达均显著降低，而RARβ的mRNA和蛋白表达未见明显变化，TGF-β1、Col-IV及FN的蛋白表达均显著增多，O2−产生显著增多，MMP明显下降，细胞活力、SOD和GSH酶活力明显降低。⑵ 下调RARα或RARγ表达后，O2−红色荧光密度和MMP荧光密度明显增大，TGF-β1、Col-IV和FN表达显著增多，细胞活力、SOD和GSH酶活力明显降低；而上调RARα或RARγ表达后，O2−红色荧光密度和MMP荧光密度均明显减少，TGF-β1、Col-IV和FN表达显著减少，细胞活力、SOD和GSH酶活力明显增加；而上调或下调RARβ表达后上述指标均未出现明显变化。结论：在缺氧性损伤的RTEC中，上调RARα或RARγ的表达具有减轻氧化损伤和纤维化的作用。其分子机制可能与RARα或RARγ减少O2−产生，增加SOD和GSH酶活力，减少TGF-β1产生，进而减少ECM积聚有关。本研究共发表了论文4篇（其中SCI论文2篇，中文核心期刊2篇）。培养了研究生毕业3名（其中博士研究生1名，硕士研究生2名）。