黄芩汤通过抑制 TNF-α 诱导的 NF-κB 激活减轻 DSS 诱导的黏膜损伤并促进上皮修复。

Huangqin Decoction Attenuates DSS-Induced Mucosal Damage and Promotes Epithelial Repair via Inhibiting TNF-α-Induced NF-κB Activation.

机构信息

Gastroenterology Department, Affiliated Hospital of Integrated Traditional Chinese and Western Medicine, Nanjing University of Chinese Medicine, Nanjing, 210028, China.

Gastroenterology Department, Jiangsu Province Hospital on Integration of Chinese and Western Medicine, Nanjing, 210028, China.

出版信息

Chin J Integr Med. 2022 Feb;28(2):124-129. doi: 10.1007/s11655-021-3343-4. Epub 2021 Dec 7.

DOI:10.1007/s11655-021-3343-4

PMID:34874516

Abstract

OBJECTIVE

To investigate the protective effect of Chinese herbal formula Huangqin Decoction (HQD) on ulcerative colitis mouse model induced by dextran sulphate sodium (DSS) and human intestinal epithelial cell injury induced by tumour necrosis factor-α (TNF-α).

METHODS

In vivo, 30 male C57BL/6 mice were divided into 5 groups using a random number table (n=6 per group), including control, DSS, 5-aminosalicylic acid (5-ASA), HQD low- (HQD-L) and high-dose (HQD-H) groups. The colitis mouse model was established by 3% (w/v) DSS water for 5 days. Meanwhile, mice in the HQD-L, HQD-H and 5-ASA groups were administrated with 100, 200 mg/kg HQD or 100 mg/kg 5-ASA, respectively, once daily by gavage. After 9 days of administration, the body weight, disease activity index (DAI) score and colon length of mice were measured, the pathological changes of colons were analyzed by hematoxylin-eosin staining (HE) staining, and the levels of serum interleukin (IL)-6, IL-1β and TNF-α were measured by enzyme linked immunosorbent assay. In vitro, the human colon epithelial normal cells (FHC cells) were exposed to HQD (0.6 mg/mL) for 12 h and then treated with TNF-α (10 ng/mL) for 24 h. The tight junction (TJ) protein expression levels of Claudin-4 and Occludin, and the protein phosphorylation levels of p65 and inhibitor of nuclear factor kappaB (NF-κB)-α (IκBα) were measured by Western blot.

RESULTS

In vivo, compared with the DSS group, HQD-H treatment attenuated the weight loss and reduced DAI score of mice on the 8th day (P<0.05). Moreover, HQD-H treatment ameliorated the colon shortening in the DSS-induced colitis mice (P<0.05). HE staining showed HQD attenuated the pathological changes of colitis mice, and the histological scores of HQD-H and 5-ASA groups were significantly decreased compared with the DSS group (P<0.05). Meanwhile, HQD-H and 5-ASA significantly decreased the serum IL-1β, IL-6 and TNF-α levels of mice (P<0.05). In vitro experiments showed that HQD up-regulated Occludin and Claudin-4 protein expressions and inhibited p-p65 and p-IκBα levels in FHC cells compared with the TNF-α group (P<0.05).

CONCLUSION

HQD significantly relieved the symptoms in DSS-induced colitis mice by inhibiting pro-inflammatory cytokines expression and maintained the homeostasis of TJ protein in FHC cells by suppressing TNF-α-induced NF-κB activation.

摘要

目的

探讨黄芩汤（HQD）对葡聚糖硫酸钠（DSS）诱导的溃疡性结肠炎（UC）小鼠模型和肿瘤坏死因子-α（TNF-α）诱导的人肠上皮细胞损伤的保护作用。

方法

体内实验将 30 只雄性 C57BL/6 小鼠采用随机数字表法分为 5 组（n=6），分别为对照组、DSS 组、5-氨基水杨酸（5-ASA）组、HQD 低剂量（HQD-L）组和 HQD 高剂量（HQD-H）组。采用 3%（w/v）DSS 水诱导 5 天建立结肠炎小鼠模型。同时，HQD-L、HQD-H 和 5-ASA 组分别给予 100、200mg/kg HQD 或 100mg/kg 5-ASA 灌胃，每天 1 次。给药 9 天后，测量小鼠的体重、疾病活动指数（DAI）评分和结肠长度，苏木精-伊红（HE）染色分析结肠病理变化，酶联免疫吸附试验（ELISA）法检测血清白细胞介素（IL）-6、IL-1β和 TNF-α水平。体外实验将人结肠上皮正常细胞（FHC 细胞）暴露于 HQD（0.6mg/mL）12h 后，再用 TNF-α（10ng/mL）处理 24h。采用 Western blot 法检测 Claudin-4 和 Occludin 紧密连接（TJ）蛋白表达水平以及 p65 和核因子κB（NF-κB）-α 抑制剂（IκBα）磷酸化水平。

结果

体内实验结果显示，与 DSS 组相比，HQD-H 治疗可减轻第 8 天小鼠的体重减轻和 DAI 评分（P<0.05）。此外，HQD-H 治疗可改善 DSS 诱导的结肠炎小鼠的结肠缩短（P<0.05）。HE 染色显示 HQD 可减轻结肠炎小鼠的病理变化，HQD-H 和 5-ASA 组的组织学评分明显低于 DSS 组（P<0.05）。同时，HQD-H 和 5-ASA 可显著降低小鼠血清中 IL-1β、IL-6 和 TNF-α水平（P<0.05）。体外实验结果表明，与 TNF-α 组相比，HQD 可上调 FHC 细胞中 Occludin 和 Claudin-4 蛋白表达，并抑制 p-p65 和 p-IκBα 水平（P<0.05）。