R.S. Mehta Jain Department of Biochemistry and Cell Biology, KBIRVO, Vision Research Foundation, Chennai 600006, India; School of Chemical and Biotechnology, SASTRA Deemed to be University, Thanjavur, India.
R.S. Mehta Jain Department of Biochemistry and Cell Biology, KBIRVO, Vision Research Foundation, Chennai 600006, India.
Life Sci. 2020 Apr 1;246:117397. doi: 10.1016/j.lfs.2020.117397. Epub 2020 Feb 4.
Paraoxonase (PON), an antioxidant enzyme, comprises of three members (PON1, 2 and 3). Hyperglycemia accelerates formation of AGE in diabetes which mediates endothelial dysfunction. PON1 is studied in diabetes due to its association with HDL, lipid peroxidation and vascular complications but PON2 is not explored much. Recently decreased PON2 activity is reported in monocytes of Type 2 diabetes mellitus (T2DM) patients but its regulation by factors like high glucose and AGE has not been studied.
The aim of the current study is to identify the effect of AGEs on PON2 expression and activity and its implications on endothelial dysfunction in HUVECs.
HUVECs were exposed to Glycated albumin (GA)/Carboxymethyl lysine (CML) for 24 h to check for PON2 expression. The ER stress markers GRP78 and IRE1α, pro-inflammatory genes MCP-1, IL-6, IL-8, ICAM1, VCAM1 were assessed by qPCR, western blotting/ELISA. Endothelial-leukocyte adhesion and ROS were assessed using Calcein AM and DCFDA method. One-way ANOVA and student's t-test was done using Graphpad Prism.
AGE exposure significantly decreased PON2 expression and activity with increased oxidative stress, ER stress and inflammation. PON2 overexpression significantly reduced ROS, ER stress and inflammation as well as inhibited NFκB, and ERK1/2, phosphorylation induced by GA/CML treatment. Concomitantly, silencing of PON2 accelerated AGEs induced effects.
This is the first study to show that both GA and CML down regulates the PON2 expression and activity in HUVECs and over expression of PON2 alleviates AGEs induced endothelial dysfunction.
过氧化物酶（PON）是一种抗氧化酶，由三个成员（PON1、2 和 3）组成。高血糖加速糖尿病中 AGE 的形成，从而介导内皮功能障碍。PON1 因其与 HDL、脂质过氧化和血管并发症的关联而在糖尿病中进行研究，但 PON2 研究较少。最近有报道称，2 型糖尿病（T2DM）患者的单核细胞中 PON2 活性降低，但尚未研究高血糖和 AGE 等因素对其的调节作用。
本研究旨在确定 AGE 对 PON2 表达和活性的影响及其对 HUVECs 内皮功能障碍的影响。
将 HUVECs 暴露于糖化白蛋白（GA）/羧甲基赖氨酸（CML）24 小时，以检查 PON2 的表达。通过 qPCR、Western blot/ELISA 评估内质网应激标志物 GRP78 和 IRE1α、促炎基因 MCP-1、IL-6、IL-8、ICAM1、VCAM1。使用 Calcein AM 和 DCFDA 法评估内皮-白细胞黏附和 ROS。使用 Graphpad Prism 进行单因素方差分析和学生 t 检验。
AGE 暴露显著降低 PON2 的表达和活性，同时增加氧化应激、内质网应激和炎症。PON2 过表达显著降低了 ROS、内质网应激和炎症，抑制了 NFκB 和 ERK1/2 的磷酸化，从而减轻了 GA/CML 处理引起的炎症。同时，PON2 的沉默加速了 AGE 诱导的作用。
这是第一项表明 GA 和 CML 均可下调 HUVECs 中 PON2 的表达和活性，而过表达 PON2 可减轻 AGE 诱导的内皮功能障碍的研究。
