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Protective effects of CXCR3/HO?1 gene?modified BMMSCs on damaged intestinal epithelial cells: Role of the p38?MAPK signaling pathway.

ABSTRACT: The purpose of the present study was to investigate whether bone marrow mesenchymal stem cells (BMMSCs) modified by CXC?chemokine receptor type 3 (CXCR3) and heme oxygenase?1 (HO?1) genes can repair damaged intestinal epithelial cells in vitro, and the role of the p38 mitogen?activated protein kinase (p38?MAPK) pathway in this process. A model of intestinal epithelial crypt cell line?6 (IEC?6) damage was created, and BMMSCs were transfected with either the CXCR3 and/or HO?1 gene in vitro. There were nine experimental groups in which the damaged IEC?6 cells were co?cultured with differentially?treated BMMSCs and lymphocytes for 24 h. Reverse transcription?quantitative polymerase chain reaction analysis, immunohistochemistry and a western blot analysis were performed to detect stem cell transfection, the repair of damaged intestinal epithelial cells and the expression of related molecules in the P38?MAPK pathway, respectively. Crystal violet staining and live cell imaging were used to detect the chemotaxis of BMMSCs. Flow cytometry was used to detect T lymphocyte activity and the surface markers expressed on BMMSCs. An ELISA was used to quantify cytokine production. The adenovirus (Ad)?CXCR3/MSCs exhibited the characteristics of stem cells and exhibited chemotaxis. The Ad?CXCR3/MSCs and Ad?(CXCR3 + HO)/MSCs exhibited increased expression of tight junction protein zonula occludens?1 (ZO?1) and anti?proliferating cell nuclear antigen in the damaged IEC?6 cells, and apoptosis of the damaged IEC?6 cells was decreased. BMMSCs inhibited the phosphorylation of p38, in addition to downstream molecules of the p38MAPK signaling pathway. The Ad?CXCR3/MSCs and Ad?(CXCR3 + HO)/MSCs exhibited significantly decreased expression levels of downstream molecules, including phosphorylated (p)?p38, p?activated transcription factor 2, p?C/EBP homologous protein?10, and p?myocyte enhancer factor 2C, and target molecules (e.g., apoptotic bodies). The effects of Ad?(CXCR3 + HO)/MSCs on the repair of the damaged intestinal tract and inhibition of the p38?MAPK pathway was more marked than those in other groups on day 7 post?surgery in the rejection model for small bowel transplantation. BMMSCs modified by the CXCR3 and HO?1 genes exhibited superior ability to repair damaged intestinal epithelial cells and served this role via the p38?MAPK pathway.

SUBMITTER: Yin M

PROVIDER: S-EPMC6445595 | biostudies-literature | 2019 May

REPOSITORIES: biostudies-literature

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Project description:The present study aimed to explore whether heme oxygenase-1 (HO-1)-modified bone marrow mesenchymal stem cells (BMMSCs) have a protective effect on liver transplantation with steatotic liver grafts in rats, and to determine the role of the intestinal microbiota in such protection. HO-1/BMMSCs were obtained by transduction of Hmox1 gene [encoding heme oxygenase (HO-1)]-encoding adenoviruses into primary rat BMMSCs. Steatotic livers were obtained by feeding rats a high-fat diet, and a model of liver transplantation with steatotic liver grafts was established. The recipients were treated with BMMSCs, HO-1/BMMSCs, or neither, via the portal vein. Two time points were used: postoperative day 1 (POD 1) and POD 7. The results showed that under the effect of HO-1/BMMSCs, the degree of steatosis in the liver grafts was significantly reduced, and the level of liver enzymes and the levels of pro-inflammatory cytokines in plasma were reduced. The effect of HO-1/BMMSCs was better than that of pure BMMSCs in the prolongation of the rats' postoperative time. In addition, HO-1/BMMSCs promoted the recovery of recipients' intestinal structure and function, especially on POD 7. The intestinal villi returned to normal, the expression of tight junction proteins was restored, and intestinal permeability was reduced on POD 7. The intestinal bacterial of the LT group showed significantly weakened energy metabolism and overgrowth. On POD 1, the abundance of Akkermansiaceae was higher. On POD 7, the abundance of Clostridiaceae increased, the level of lipopolysaccharide increased, the intestinal mucosal barrier function was destroyed, and the levels of several invasive bacteria increased. When treated with HO-1/BMMSCs, the energy metabolism of intestinal bacteria was enhanced, and on POD 1, levels bacteria that protect the intestinal mucosa, such as Desulfovibrionaceae, increased significantly. On POD 7, the changed intestinal microbiota improved lipid metabolism and increased the levels of butyrate-producing bacteria, such as Lachnospiraceae. In conclusion, HO-1/BMMSCs have protective effects on steatotic liver grafts and the intestinal barrier function of the recipients. By improving lipid metabolism and increasing the abundance of butyrate-producing bacteria, the changed intestinal microbiota has a protective effect and prolongs the recipients' survival time.

Project description:p38 MAPK inhibition may have additive and synergistic anti-inflammatory effects when used with corticosteroids. We investigated crosstalk between p38 MAPK inhibitors and corticosteroids in bronchial epithelial cells to investigate synergistic effects on cytokine production and the molecular mechanisms involved. Effects of the p38 MAPK inhibitor BIRB-796 and dexamethasone alone and in combination on LPS, polyI:C or TNFα -induced IL-6, CXCL8 and RANTES were assessed in 16HBEs (human epithelial cell line) and on TNFα-induced IL-6 and CXCL8 in primary human epithelial cells from asthma patients and healthy controls. 16HBEs were used to assess effects of BIRB-796 alone and in combination with dexamethasone on glucocorticoid receptor (GR) activity by reporter gene assay, expression of GR target genes and nuclear localisation using Western blot. The effects of BIRB-796 on TNFα stimulated phosphorylation of p38 MAPK and GR at serine (S) 226 by Western blot. Epithelial levels of phosphorylated p38 MAPK and GR S226 were determined by immunohistochemistry in bronchial biopsies from asthma patients and healthy controls. BIRB-796 in combination with dexamethasone increased inhibition of cytokine production in a synergistic manner. Combination treatment significantly increased GR nuclear localisation compared to dexamethasone alone. BIRB-796 inhibited TNFα-induced p38 MAPK and GR S226 phosphorylation. Phosphorylated GR S226 and p38 MAPK levels were increased in bronchial epithelium of more severe asthma patients. Molecular crosstalk exists between p38 MAPK activation and GR function in human bronchial epithelial cells, which alters GR activity. Combining a p38 MAPK inhibitor and a corticosteroid may demonstrate therapeutic potential in severe asthma. KEY MESSAGES: • Combination of corticosteroid and p38 inhibitor in human bronchial epithelial cells • Combination increased cytokine inhibition synergistically and nuclear GR • p38 MAPK inhibition reduced TNFα-induced phosphorylation of GR at S226 but not S211 • Phosphorylated GRS226 and p38 is increased in bronchial epithelium in severe asthma • Combining a p38 inhibitor and a corticosteroid may be effective in asthma treatment.

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Protective effects of CXCR3/HO?1 gene?modified BMMSCs on damaged intestinal epithelial cells: Role of the p38?MAPK signaling pathway.

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