Chang'an Ⅱ Decoction(肠安Ⅱ号方)-Containing Serum Ameliorates Tumor Necrosis Factor-α-Induced Intestinal Epithelial Barrier Dysfunction via MLCK-MLC Signaling Pathway in Rats

作者：CHEN Ting; YIN Xiao-lan; KANG Nan; WANG Xiao-ge; LI Bao-shuang; JI Hai-jie; ZHANG Yin-qiang; BIAN Li-qun; ZHANG Bei-hua; WANG Feng-yun; TANG Xu-dong

来源：Chinese Journal of Integrative Medicine, 2020, 26(10): 745-753.

Objective: To investigate the effect of Chang’an Ⅱ Decoction(肠安Ⅱ号方)-containing serum on intestinal epithelial barrier dysfunction in rats. Methods: Tumor necrosis factor(TNF)-α-induced injury of Caco-2 monolayers were established as an inflammatory model of human intestinal epithelium. Caco-2 monolayers were treated with blank serum and Chang’an Ⅱ Decoction-containing serum that obtained from the rats which were treated with distilled water and Chang’an Ⅱ Decoction intragastrically at doses of 0.49, 0.98, 1.96 g/(kg·d) for 1 week, respectively. After preparation of containing serum, cells were divided into the normal group, the model group, the Chang’an Ⅱ-H, M, and L groups(treated with 30 ng/mL TNF-α and medium plus 10% high, middle-, and low-doses Chang’an Ⅱ serum, respectively). Epithelial barrier function was assessed by transepithelial electrical resistance(TER) and permeability of fluorescein isothiocyanate(FITC)-labeled dextran. Transmission electron microscopy was used to observe the ultrastructure of tight junctions(TJs). Immunofluorescence of zonula occludens-1(ZO-1), claudin-1 and nuclear transcription factor-kappa p65(NF-κBp65) were measured to determine the protein distribution. The mRNA expression of myosin light chain kinase(MLCK) was measured by real-time polymerase chain reaction. The expression levels of MLCK, myosin light chain(MLC) and p-MLC were determined by Western blot. Results: Chang’an Ⅱ Decoction-containing serum significantly attenuated the TER and paracellular permeability induced by TNF-α. It alleviated TNF-α-induced morphological alterations in TJ proteins. The increases in MLCK mRNA and MLCK, MLC and p-MLC protein expressions induced by TNF-α were significantly inhibited in the Chang’an Ⅱ-H group. Additionally, Chang’an Ⅱ Decoction significantly attenuated translocation of NF-κBp65 into the nucleus. Conclusion: High-dose Chang’an Ⅱ-containing serum attenuates TNF-α-induced intestinal barrier dysfunction. The underlying mechanism may be involved in inhibiting the MLCK-MLC phosphorylation signaling pathway mediated by NF-κBp65.

• 单位
  中医学科学院; 河南大学