NLRP-3抑制剂MCC950对胃酸刺激诱导HEECs氧化应激、抗氧化蛋白及炎症因子表达的作用

Effects of the NLRP-3 inhibitor MCC950 on oxidative stress and expressions of antioxidant proteins and inflammatory cytokines in HEECs induced by gastric acid stimulation

摘要:

目的 探讨NOD样受体蛋白3（NLRP-3）抑制剂MCC950对胃酸刺激诱导人食管上皮细胞（HEECs）氧化应激、抗氧化蛋白及炎症因子表达的作用。方法 HEECs细胞经传代培养，将细胞分为空白对照组、酸刺激组、胆盐刺激组、脂多糖（LPS）处理组、MCC950预处理+酸刺激组、MCC950预处理+胆盐刺激组、MCC950预处理+LPS刺激组、N-乙酰半胱氨酸（NAC）预处理+酸刺激组、NAC预处理+胆盐刺激组、NAC预处理+LPS组等，每组细胞孵育48 h；并提取总RNA，后进行逆转录聚合酶链反应检测并分析氧化应激、抗氧化蛋白和炎症因子指标mRNA表达水平。结果 酸刺激组、胆盐刺激组和LPS处理组Nox-4 mRNA相对表达量较空白对照组高（P <0.05），酸刺激组、胆盐刺激组和LPS处理组Nrf-2、HO-1 mRNA相对表达量较空白对照组低（P <0.05）。MCC950+酸刺激组、MCC950+胆盐刺激组、MCC950+胆盐刺激组Nox-4 mRNA相对表达量分别较酸刺激组、胆盐刺激组、LPS处理组低（P <0.05），MCC950+酸刺激组、NAC+酸刺激组、MCC950+胆盐刺激组、NAC+胆盐刺激组、MCC950+LPS处理组、NAC+LPS处理组Nrf-2、HO-1 mRNA相对表达量分别较酸刺激组、胆盐刺激组、LPS处理组高（P <0.05）。酸刺激组、胆盐刺激组和LPS处理组MCP-1、白细胞介素-6（IL-6）、肿瘤坏死因子-α（TNF-α） mRNA相对表达量较空白对照组高（P <0.05），CC950+酸刺激组、NAC+酸刺激组、MCC950+胆盐刺激组、NAC+胆盐刺激组、MCC950+LPS处理组、NAC+LPS处理组MCP-1、IL-6、TNF-α mRNA相对表达量分别较酸刺激组、胆盐刺激组、LPS处理组低（P <0.05）。结论 胃酸诱导HEECs细胞Nox-4高表达和抗氧化蛋白低表达，促进炎症因子的过量表达，MCC950能有效抑制炎症损伤的发生。

Abstract:

Objective To explore the effects of the NOD-like receptor protein 3 (NLRP3) inhibitor on oxidative stress and expressions of antioxidant proteins and inflammatory cytokines in human esophageal epithelial cells (HEECs) induced by gastric acid stimulation.Methods HEECs were cultured and divided into the blank control group, acid group, bile salt group, lipopolysaccharide (LPS) group, MCC950 pretreatment + acid group, MCC950 pretreatment + bile salt group, MCC950 pretreatment + LPS group, N-acetylcysteine (NAC) pretreatment + acid group, NAC pretreatment + bile salt group, and NAC pretreatment + LPS group. Cells were subjected to various treatment conditions for 48 hours. The total RNA was extracted, and quantitative real-time polymerase chain reaction was performed to detect the mRNA expression levels of oxidative stress indicators, antioxidant proteins, and inflammatory cytokines.Results As compared to the blank control group, the relative mRNA expression of Nox-4 was higher (P < 0.05) and that of Nrf-2 and HO-1 was lower (P < 0.05) in the acid group, bile salt group, and LPS group. The relative mRNA expression of Nox-4 in the MCC950 pretreatment + acid group, MCC950 pretreatment + bile salt group, and MCC950 pretreatment + LPS group was lower than that in the acid group, bile salt group, and LPS group, respectively (P < 0.05). The relative mRNA expressions of Nrf-2 and HO-1 in the MCC950 pretreatment + acid group, NAC pretreatment + acid group, MCC950 pretreatment + bile salt group, NAC pretreatment + bile salt group, MCC950 pretreatment + LPS group, and NAC pretreatment + LPS group were higher compared to those in the acid group, bile salt group, and LPS group, respectively (P < 0.05). The relative mRNA expressions of MCP-1, interleukin-6 (IL-6), and tumor necrosis factor-α (TNF-α) in the acid group, bile salt group, and LPS group were higher than those in the blank control group (P < 0.05), while they were lower in the MCC950 pretreatment + acid group, NAC pretreatment + acid group, MCC950 pretreatment + bile salt group, NAC pretreatment + bile salt group, MCC950 pretreatment + LPS group, and NAC pretreatment + LPS group compared to those in the acid group, bile salt group, and LPS group, respectively (P < 0.05).Conclusion Gastric acid induces high expression of Nox-4 and low expression of antioxidant proteins in HEECs cells, resulting in excessive production of inflammatory cytokines. MCC950 effectively attenuates the inflammation-mediated cellular injury.