基于p38 MAPK/NF-κB/MLCK信号通路探讨半夏泻心汤修复肠黏膜屏障的作用机制

Exploring the Mechanism of Banxia Xiexin Decoction in Repairing Intestinal Mucosal Barrier Based on p38 MAPK/NF-κB/MLCK Signaling Pathway

目的:探寻半夏泻心汤(BXD)在体内抑制肠道炎症反应及修复肠黏膜屏障的作用机制。方法:36只雌性SPF级SD大鼠随机分为空白组、模型组、西药组(美沙拉嗪组)以及半夏泻心汤低剂量组(BXD-低剂量组)、中剂量组(BXD-中剂量组)、高剂量组(BXD-高剂量组)。适应性喂养7 d后,除空白组外,其余各组大鼠自由饮用5%DSS 7 d构建溃疡性结肠炎大鼠模型,造模成功后,空白组与模型组灌胃生理盐水,西药组灌胃美沙拉嗪,0.054 g/d),半夏泻心汤低剂量组(2.205 g/kg)、中剂量组(4.41 g/kg)、高剂量组(8.82 g/kg)灌胃给予半夏泻心汤提取物,均给药15 d。采用疾病活动指数评估造模及药物治疗后大鼠的一般变化情况;苏木精-伊红(HE)染色法,确认结肠组织病变损伤程度;ELISA检测血清中炎性因子含量表达变化情况;qRT-PCR法、Western blot法检测p38 MAPK/NF-κB/MLCK/信号通路关键蛋白表达。结果:5%葡聚糖硫酸钠(DSS)诱导的溃疡性结肠炎模型大鼠出现体质量减轻、腹泻、脓血便及结肠缩短,造模成功。BXD及美沙拉嗪给药后,大鼠的DAI评分出现不同程度的降低。HE结果显示美沙拉嗪及高剂量BXD给药后明显改善了DSS导致的大鼠结肠组织的病理情况。酶联免疫吸附实验结果提示,高剂量BXD及美沙拉嗪给药后可以显著抑制大鼠促炎因子的表达水平,从而达到抑制肠道炎症的治疗效果。Western blot和qPCR结果表明,BXD可通过调控MAPK、MLCK、NF-κB、Occludin的表达量修复肠黏膜屏障。结论:半夏泻心汤可降低肠黏膜通透性以及修复受损的肠黏膜,其机制可能是通过抑制p38 MAPK/NF-κB/MLCK信号通路实现的。

Objective:To explore the mechanism by which Banxia Xiexin Decoction(BXD)inhibits intestinal inflammatory response and repairs the intestinal mucosal barrier in vivo.Methods:Thirty-six female SPFgrade SD rats were randomly divided into a blank group,a model group,a Western medicine group(mesalazine group),and low,medium,and high-dose Banxia Xiexin Decoction groups(BXD-low,BXD-medium,and BXD-high dose groups).After 7 days of adaptive feeding,all groups except the blank group were freely given 5%DSS for 7 days to induce an ulcerative colitis rat model.Following successful modeling,the blank and model groups were gavaged with physiological saline,the Western medicine group was gavaged with mesalazine(0.054 g/d),and the BXD groups were gavaged with Banxia Xiexin Decoction extract at doses of 2.205 g/kg,4.41 g/kg,and 8.82 g/kg respectively,for 15 days.The Disease Activity Index(DAI)was used to assess general changes after modeling and drug treatment.Hematoxylin-eosin(HE)staining confirmed the extent of colonic tissue damage.ELISA detected changes in the expression of inflammatory factors in serum,and qRT-PCR and Western blot were used to detect the expression of key proteins in the p38 MAPK/NF-κB/MLCK signaling pathway.Results:The 5%DSS-induced ulcerative colitis model rats showed weight loss,diarrhea,bloody stools,and shortened colons,confirming successful modeling.After administration of BXD and mesalazine,DAI scores decreased to varying degrees.HE results indicated that mesalazine and high-dose BXD significantly improved DSS-induced pathological changes in rat colonic tissue.ELISA results suggested that high-dose BXD and mesalazine significantly inhibited the expression levels of pro-inflammatory factors,thereby achieving an anti-inflammatory effect in the intestines.Western blot and qPCR results showed that BXD could repair the intestinal mucosal barrier by regulating the expression of MAPK,MLCK,NF-κB,and Occludin.Conclusion:Banxia Xiexin Decoction can reduce intestinal mucosal permeability and repair damaged intestinal mucosa,potentially through the inhibition of the p38 MAPK/NF-κB/MLCK signaling pathway.