山药总蛋白对高糖诱导的人脐静脉内皮细胞氧化应激的保护作用及机制研究

姜蕾蕾; 冯佳宝; 刘颖; 金辰蓉; 刘美辰; 王思明; 赵大庆; 于士婷

doi:10.13386/j.issn1002-0306.2023060254

山药总蛋白对高糖诱导的人脐静脉内皮细胞氧化应激的保护作用及机制研究

Protective Effect and Mechanism of Chinese Yam Total Protein on High D-Glucose Induced Oxidative Stress in Human Umbilical Vein Endothelial Cells

摘要

摘要: 探究山药总蛋白对高浓度D-葡萄糖（High concentrations of D-glucose，HG）诱导的人脐静脉内皮细胞（Human umbilical vein endothelial cells，HUVECs）氧化应激损伤的保护作用及其机制。利用50 mmol/L HG建立HUVECs损伤模型。将HUVECs随机分为对照组、模型组、山药总蛋白低剂量组（0.5 mg/mL）以及山药总蛋白高剂量组（1 mg/mL），评估山药总蛋白对HG处理的HUVECs的细胞活力、细胞形态及血管生成能力的影响；检测细胞中活性氧（Reactive oxygen species，ROS）的水平和超氧化物歧化酶（Superoxide dismutase，SOD）、过氧化氢酶（Catalase，CAT）的活力；以及培养基上清液中一氧化氮（Nitric oxide，NO）、8-羟基脱氧鸟苷（8-Hydroxy deoxyguanosine，8-OHdG）、丙二醛（Malondialdehyde，MDA）、白介素-1β（Interleukin-1β，IL-1β）、白介素-6（Interleukin-6，IL-6）的水平；测定B细胞淋巴瘤2（Bcl-2）、Bcl-2相关X蛋白（Bax）、硫氧还蛋白相互作用蛋白（Txnip）、NOD样受体蛋白3（NLRP3）、半胱氨酸天冬氨酸蛋白水解酶剪切体-1（Cleaved caspase-1）和IL-1β蛋白的表达水平。结果显示，山药总蛋白能显著提高HG处理后的HUVECs的细胞活力（P<0.01），改善细胞凋亡形态，并显著升高SOD和CAT的活力（P<0.05或P<0.01）以及凋亡相关蛋白（Bcl-2/Bax）的比值、血管形成能力、NO分泌量。此外，山药总蛋白还显著降低了HUVECs的ROS、MDA、8-OHdG、IL-1β和IL-6的水平（P<0.05或P<0.01）及炎症相关蛋白（Txnip、NLRP3、cleaved Caspase-1、IL-1β）的表达水平（P<0.05或P<0.01），并且上述指标的变化均呈现明显的浓度依赖性。本研究结果表明，山药总蛋白可能通过抑制Txnip/NLRP3信号通路从而保护HUVECs拮抗HG诱导的氧化应激，恢复HUVECs内的氧化平衡，进一步减轻细胞的炎症反应。

Abstract: To explored the protective effect and mechanism of Chinese yam total protein in human umbilical vein endothelial cells (HUVECs) with high-concentration D-glucose (HG)-induced oxidative stress injury. The HUVECs injury model was established by 50 mmol/L HG. HUVECs were randomly divided into control, model, low-dose Chinese yam total protein (0.5 mg/mL), and high-dose Chinese yam total protein (1 mg/mL) groups. The effects of Chinese yam total protein on the viability, morphology, and angiogenetic ability of HUVECs treated with HG were evaluated. The reactive oxygen species (ROS) level, superoxide dismutase (SOD) and catalase (CAT) activities in cells were determined. The nitric oxide (NO), 8-hydroxy deoxyguanosine (8-OHdG), malondialdehyde (MDA), interleukin-1β (IL-1β), and interleukin-6 (IL-6) levels in culture medium supernatant were measured. Protein expression levels of B-cell lymphoma 2 (Bcl-2), Bcl-2-associated X (Bax), thioredoxin-interacting protein (Txnip), NOD-like receptor protein 3 (NLRP3), cleaved Caspase-1, and IL-1β were determined. The results showed that Chinese yam total protein could significantly increased the cell viability of HUVECs treated with HG (P<0.01), improved cell apoptotic morphology, and significantly increased the activities of SOD and CAT (P<0.05, P<0.01) and ratio of apoptosis-related proteins (Bcl-2/Bax), angiogenetic ability and nitric oxide (NO) secretion. Moreover, Chinese yam total protein significantly decreased the levels of ROS, MDA, 8-OHdG, IL-1β, and IL-6 (P<0.05, P<0.01) and the expression levels of inflammation-related proteins (Txnip, NLRP3, cleaved Caspase-1, and IL-1β) (P<0.05, P<0.01). Changes in these indicators showed a significant concentration-dependent manner. The results indicate that Chinese yam total protein may protect against HG-induced oxidative stress by inhibiting the Txnip/NLRP3 signaling pathway, restoring the oxidative balance, and reducing the inflammatory response of HUVECs.