目的

探讨罗乐胃蜜膏对酸刺激诱导人食管上皮细胞（HEEC）氧化应激和炎症因子表达的影响及作用。

方法

HEEC 细胞进行传代培养，实验进一步分为5 组：空白对照组（Control/n=6）、酸刺激组（n=6）、罗乐维蜜膏预处理组（LL 组，n=6）、平溃加瓦日西麦尔瓦衣特蜜膏预处理组（PK 组，n=6）以及艾司奥美拉唑预处理组（AS 组，n=6）。酸刺激采用pH 值酸性培养基（pH=4）每日刺激3 次，每次15 min，培养48 h。LL 组、PK 组和AS 组分别加入浓度为1 mmol/L 的LL、PK 和AS 孵育4 h 之后，进行酸刺激48 h。每组分别提取总核糖核酸（RNA），进行实时荧光定量PCR（RT-PCR）实验检测烟酰胺腺嘌呤二核苷酸磷酸-4（Nox-4）、抗氧化酶［锰超氧化物歧化酶（Mn SOD）、谷胱甘肽过氧化物酶（GSH-Px）、过氧化氢酶（CAT）］以及炎症因子指标［单核细胞趋化蛋白-1（MCP-1）、白细胞介素-6（L-6）及肿瘤坏死因子α（TNF-α）］的mRNA 表达水平。

结果

经pH=4 酸刺激培养48 h HEEC 细胞显著增加Nox-4 的mRNA 表达水平（P＜0.0001）；LL组、PK 组以及AS 组均显著抑制Nox-4 mRNA 高表达（P＜0.0001）；LL 组抑制Nox-4 mRNA 高表达效果优于PK 组（P＜0.01）。pH=4 酸刺激培养显著降低HEEC 细胞抗氧化酶（Mn SOD、GSH-Px、CAT）mRNA 的表达水平（P＜0.0001）。LL 组、PK 组和AS 组均明显增加上述抗氧化酶的mRNA表达水平（P＜0.0001）；与PK 组比较，LL 组明显上升抗氧化酶的mRNA 表达水平（P＜0.05）。酸刺激培养显著增加HEEC 细胞炎症因子（MCP-1、IL-6、TNF-α）的mRNA 表达水平（P＜0.0001）。LL 组、PK 组和AS 组均明显降低上述炎症因子的mRNA 表达水平（P＜0.0001）；与PK 组比较，LL组明显降低炎症因子的mRNA 表达水平（P＜0.01）。

结论

罗乐胃蜜膏能显著抑制酸刺激诱导HEEC细胞氧化应激和炎症因子的高表达，其深入的机制有待进一步研究。

Objective

To investigate the effects of luole weimi ointment on oxidative stress and inflammatory cytokines expression in human esophageal epithelial cells （HEEC） induced by acid stimulation.

Methods

HEEC cells were cultured and the experiment was further divided into 5 groups：Control group（n=6）, acid stimulation group（n=6）, luoliwei pretreatment group （LL group, n=6）, pingkui jiawarixi maierwayite honey ointment pretreatment group （PK group, n=6） and esomeprazole pretreatment group（AS group, n=6）. The acid was stimulated with pH value acid medium （pH=4） 3 times a day, 15min each time, for 48 h； LL group, PK group and AS group were added with 1 mmol/L LL, PK and AS for 4 h, and then were stimulated with acid for 48 h. Total ribonucleic acid （RNA） was extracted from each group, and quantitative real-time polymerase chain reaction （RT-PCR） was performed to detect the mRNA expression levels of nicotinamide adenine dinucleotide phosphate oxidase-4 （Nox-4）, antioxidant enzymes ［manganese superoxide dismutase （Mn SOD）, glutathione peroxidase （GSH-Px）, catalase （CAT）］ and inflammatory factor indicators ［monocyte chemotactic protein 1 （MCP-1）, interleukin-6 （IL-6） and tumor necrosis factor α （TNF-α）］.

Results

After pH=4 acid stimulation for 48 h, the mRNA expression level of Nox-4 in HEEC cells was significantly increased （P＜0.0001）； the high expression of Nox-4 mRNA was inhibited in LL, PK and AS group （P＜0.0001）； the effect of LL group on inhibiting Nox-4 mRNA was better than that of PK group （P＜0.01）. pH=4 acid stimulation significantly reduced the expression of antioxidant enzymes （MnSOD,GSH-Px, CAT） mRNA in HEEC cells （P＜0.0001）. The mRNA expression levels of the above antioxidant enzymes were significantly increased in LL group, PK group and AS group （P＜0.0001）； compared with the PK group, the mRNA expression level of the antioxidant enzymes in LL group was significantly increased（P＜0.05）. Acid stimulation significantly increased the mRNA expression levels of inflammatory factors（MCP-1, IL-6, TNF-α） in HEEC cells （P＜0.0001）. The mRNA expression levels of the above inflammatory factors were significantly decreased in LL, PK and AS group （P＜0.0001）； it is noteworthy that compared with the PK group, the mRNA expression level of the inflammatory factors in LL group was significantly decreased （P＜0.01）.

Conclusion

Luole weimi ointment can significantly inhibit the oxidative stress and the inflammatory cytokines in HEEC cells induced by acid stimulation, and its mechanism needs further study.