Yajieshaba prevents lipopolysaccharide-induced intestinal barrier injury via anti-inflammatory and anti-apoptosis

doi:10.19852/j.cnki.jtcm.2022.05.005

Abstract

Abstract:

OBJECTIVE: To investigate the protective effect of Yajieshaba (YJSB) on the intestinal barrier dysfunction induced by lipopolysaccharide (LPS).

METHODS: C57BL/6 mice and rat intestinal epithelial cells were treated with LPS. Thiazolyl Blue Tetrazolium Bromide assay were used to detect cell viability. D-Lactate, diamine oxidase and myeloperoxidase and cytokines were determined by enzyme-linked immunosorbent assay. Western blot was used to detect apoptosis-related proteins and tight junction (TJ) proteins. Real-time quantitative polymerase chain reaction was used to quantify the levels of mRNA expression of cytokines. Histological analysis was performed by hematoxylin and eosin staining. An immunofluorescence staining assay was performed to determine the expression level of TJ protein.

RESULTS: YJSB increased cell viability and decreased apoptosis, maintained intestinal permeability after LPS-induced. YJSB inhibited LPS-induced decrease of TJ protein expression, pro-inflammatory cytokine levels and neutrophil infiltration.

CONCLUSION: YJSB protect against LPS-induced intestinal barrier dysfunction via anti-inflammatory and anti-apoptosis, suggesting its therapeutic potential against intestinal barrier injury-related diseases.

Key words: inflammation, apoptosis, lipopolysaccharides, Yajieshaba, intestinal barrier function

YANG Liping, YU Xinglin, ZHANG Chao, CHEN Pu, DUAN Xiaohua. Yajieshaba prevents lipopolysaccharide-induced intestinal barrier injury via anti-inflammatory and anti-apoptosis[J]. Journal of Traditional Chinese Medicine, 2022, 42(5): 707-714.

Figures/Tables 6

Figure 1YJSB attenuates LPS-mediated IEC-6 cell cytotoxicity A: IEC-6 cells were treated with LPS at various concentrations (25, 50, 100, 200 μg/mL) for 24 h; B: IEC-6 cells were incubated with YJSB (30, 60, 120, 240, 480 μg/mL) for 24 h. C: IEC-6 cells were treated with YJSB (30, 60, 120, 240, 480 μg/mL) and LPS (l00 μg/mL) for 24 h. YJSB: Yajieshaba; LPS: lipopolysaccharide; aP < 0.05, bP < 0.01vs control group; cP < 0.05, dP < 0.01 vs LPS group.

Figure 2 Effect of YJSB on the expression of TJ proteins A: the TJ integrity of IEC-6 cells was observed by fluorescence microscope using Hoechst staining, suggested that LPS altered the distribution of TJ protein at cell boundaries, and this effect was significantly inhibited by YJSB (×200 magnification), A1, B1, C1: control groups; A2, B2, C2: LPS groups; A3, B3, C3: YJSB groups; B: Western blot detection of the expression of TJ proteins in mice intestines. TJ: tight junction; YJSB: Yajieshaba; LPS: lipopolysaccharide; ZO-1: zonulaoccludens-1. aP < 0.01 vs control group; bP < 0.05 vs LPS group.

Figure 3 Effect of YJSB on intestinal histological appearance and permeability A: D-Lactate; B: DAO; C: effect of YJSB on body weight of the mice; D: intestinal histopathological injury was observed by hematoxylin and eosin (HE) staining (×100), D1: control group; D2,: LPS group; D3, D4: YJSB groups; E: villus heights; F: crypt depth. G: V/C. YJSB: Yajieshaba; LPS: lipopolysaccharide. V/C: villus height /crypt depth ratio; DAO: diamine oxidase. aP < 0.01 vs control group; bP < 0.05 vs LPS group.

Figure 4 Effect of YJSB on LPS-induced apoptosis Western blot detection of the expression levels of Bcl-2 and Bax in mice intestines. A: Bcl-2 and Bax protein levels were determined by Western bloting; B: relative density of Bcl-2 and Bax. YJSB: Yajieshaba; LPS: lipopolysaccharide; Bcl-2: B-cell lymphoma-2; Bax: Bcl-2 assaciated X protein. aP < 0.01vs control group; bP < 0.01 vs LPS group.

Figure 5 YJSB ameliorates LPS-induced inflammatory response TNF-α (A), IL-1β (B) and IL-6 (C) in IEC-6 cells. mRNA levels of TNF-α (D), IL-1β (E) and IL-6 (F) in jejunum homogenates of mice. MPO level in mice plasma (F). YJSB: Yajieshaba; LPS: lipopolysaccharide; TNF-α: tumor necrosis factor-α; IL-1β: interleukin-1β; IL-6: interleukin-6; MPO: myeloperoxidase; aP < 0.01 vs vehicle control group. bP < 0.05, cP < 0.01vs LPS-treated group.

Figure 6 Mechanism of YJSB in protecting the intestinal barrier function YJSB: Yajieshaba; LPS: lipopolysaccharide; ZO-1: zonulaoccludens-1; DAO: diamine oxidase; Bcl-2: B-cell lymphoma-2; Bax: Bcl-2 assaciated X protein; TNF-α: tumor necrosis factor-α; IL-1β: interleukin-1β; IL-6: interleukin-6; MPO: myeloperoxidase.

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