Zhang Xin, Wang Qian, Li Fagen, Li Suna, Lin Hepu, Huo Yanhong
Department of Nephrology, The Seventh Medical Center of PLA General Hospital, Beijing, China.
Nephron. 2023;147(8):496-509. doi: 10.1159/000529212. Epub 2023 Jan 30.
We here evaluated the efficacy of piceatannol (PIC) in high glucose (HG)-induced injury of renal tubular epithelial cells HK-2.
After the establishment of an HG-induced cell injury model and the treatment with PIC at both high and low concentrations and/or acetazolamide (ACZ, the inhibitor of carbonic anhydrase 2 [CA2]), MTT and flow cytometry assays were carried out to confirm the viability and apoptosis of HK-2 cells. The levels of oxidative stress markers lactate dehydrogenase (LDH), malondialdehyde (MDA), and reactive oxygen species (ROS), the ratio of glutathione/oxidized glutathione (GSH/GSSG), and the CA2 activity were determined. Both quantitative reverse-transcription polymerase chain reaction and Western blot were used to calculate the expressions of CA2 (the predicted target gene of PIC via intersecting the data from bioinformatic analyses) and AKT pathway-related (phosphatase and tensin homolog [PTEN], phosphorylated [p]-AKT, AKT) and apoptosis-related proteins (Bcl-2 and cleaved caspase-3).
HG suppressed cell viability and the levels of GSH/GSSG ratio, CA2, pThr308-AKT/AKT, pSer473-AKT/AKT, and Bcl-2, while promoting cell apoptosis, the levels of LDH, MDA, and ROS, and the expressions of PTEN and cleaved caspase-3. All effects of HG were reversed by PIC at a high concentration. CA2 was predicted and identified as the target of PIC. In HG-treated HK-2 cells, additionally, ACZ reversed the effects of PIC on the viability, apoptosis, and levels of both oxidative stress markers and AKT pathway- and apoptosis-related factors.
PIC protects against HG-induced injury of HK-2 cells via regulating CA2.
我们在此评估了白皮杉醇（PIC）对高糖（HG）诱导的肾小管上皮细胞HK-2损伤的疗效。
建立HG诱导的细胞损伤模型后，用高、低浓度的PIC和/或乙酰唑胺（ACZ，碳酸酐酶2 [CA2] 的抑制剂）进行处理，然后进行MTT和流式细胞术检测，以确认HK-2细胞的活力和凋亡情况。测定氧化应激标志物乳酸脱氢酶（LDH）、丙二醛（MDA）和活性氧（ROS）的水平、谷胱甘肽/氧化型谷胱甘肽（GSH/GSSG）的比值以及CA2活性。采用定量逆转录聚合酶链反应和蛋白质印迹法来计算CA2（通过生物信息学分析数据交叉确定为PIC的预测靶基因）、AKT通路相关蛋白（磷酸酶和张力蛋白同源物 [PTEN]、磷酸化 [p]-AKT、AKT）和凋亡相关蛋白（Bcl-2和裂解的半胱天冬酶-3）的表达。
HG抑制细胞活力、GSH/GSSG比值、CA2、pThr308-AKT/AKT、pSer473-AKT/AKT和Bcl-2的水平，同时促进细胞凋亡、LDH、MDA和ROS的水平以及PTEN和裂解的半胱天冬酶-3的表达。高浓度的PIC可逆转HG的所有作用。CA2被预测并鉴定为PIC的靶点。此外，在HG处理的HK-2细胞中，ACZ可逆转PIC对细胞活力、凋亡以及氧化应激标志物和AKT通路及凋亡相关因子水平的影响。
PIC通过调节CA2来保护HK-2细胞免受HG诱导的损伤。
