晚期糖基化终末产物与其受体相互作用对足细胞凋亡的影响

Impact of interaction of advanced glycation end product and its receptor on podocytes apoptosis

目的探讨可溶性与复合型晚期糖基化终末产物（AGE）与晚期糖基化终末产物受体（RAGE）的相互作用对足细胞凋亡的影响。方法以可溶性（CML—BSA、AGE—BSA）和复合型（AGE修正胶原1V）AGE刺激小鼠足细胞，并用浓度分别为10、50、100mg／L的AGE刺激细胞，应用TUNEL染色和荧光激活细胞分类（FACS）法来计数凋亡和坏死的足细胞。用RAGEiRNA转染足细胞后，以同样剂量的可溶性和复合型AGE刺激足细胞，观察凋亡情况的改变。结果可溶性和复合型AGE均可诱导小鼠足细胞凋亡，复合型AGE引起的足细胞凋亡是可溶性AGE的2～3倍（均P〈0．01）。AGE呈剂量依赖性引起足细胞凋亡。用RAGEiRNA转染足细胞，降低60％～70％RAGE基因活性后，可溶性AGE引起的凋亡率明显下降，复合型AGE诱导的凋亡有下降趋势，但不明显。只有在AGE100mg／L刺激后才发生细胞坏死。结论可溶性AGE主要通过与RAGE相互作用引起足细胞凋亡，复合型AGE部分通过与RAGE相互作用诱导足细胞凋亡。减少AGE生成和RAGE表达可能是预防肾脏病进展的重要途径。

Objective To study the effects of the interaction of advanced glycation end products （AGEs） and the receptor of AGEs （RAGE） on apoptosis of mice podocytes. Methods Podocytes were exposed to soluble AGEs such as bovine serum albmnin （BSA）, carboxymethyl-lysin （CML）-BSA, AGE-BSA and matrix-bound AGEs （AGE-modified collagen 1V ）, and to different concentrations of AGE, such as 10 mg/L, 50 rag/L, 100 mg/L. Apoptosis was assessed by TUNEL staining. Fluorescence-activated cell sorting （FACS） was used for the quantification of apoptotic and necrotic podocytes ＂after Annexin V-tluorescein isothiocyanate （FITC） and propidium iodide （PI） labeling. Apoptosis was described as the ratio of apoptotic cells to the total number cells under the high-power field, siRNA was transfected into podocytes through combining Dharmacon on Targetplus SMART pool siRNA reagents and Amaxa RNAi nucleofection kit. Results The apoptosis rate was higher in podocytes exposed to either CML-BSA or AGE-BSA than that exposed to BSA. There was a two- to three-fold increase in apoptosis collagen Ⅳ as compared with native collagen Ⅳ exposure occurred in a dose-dependent manner when podocytes were cultured in AGE-modified The apoptotic response of podocytes to AGE-BSA Podocyte necrosis occurred only at the highest concentration of AGE-BSA（100 mg/L）. AGE-BSA failed to induce apoptosis in podocytes transfected with RAGE siRNA. RAGE-specific gene knockdown did not significantly reduce the apoptosis of podocytes cultured in AGE-modified collagen IV. Conclusions The AGE-RAGE plays a major role in the apoptosis of podocytes triggered by soluble AGEs, but not bound AGEs. Reduction of AGE burden and RAGE expression may be important approaches to prevent the progression of kidney disease.