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Hypoxia enhances protective effect of placental-derived mesenchymal stem cells on damaged intestinal epithelial cells by promoting secretion of insulin-like growth factor-1.

ABSTRACT: Apoptosis and necrosis of intestinal epithelial cells (IECs), induced by ischemia-reperfusion (I/R) injury, can lead to dysfunction of the intestinal barrier, which could cause multiple organ dysfunction syndromes. Mesenchymal stem cells (MSCs) have the potential of providing protective effects on damaged IECs via paracrine action. This study investigated whether hypoxia can enhance the protective effect of placental-derived MSCs (pMSCs) on H2O2-treated-caco2 cells, and explored the possible mechanism. The pMSCs isolated by tissue culture were fibroblast-like, positive for CD73, CD90 and CD105 and can differentiate into chondrocytes and endothelial cells. Five days after treatment with H2O2, the numbers of living caco2 cells significantly decreased. More live H2O2-treated-caco2 cells were observed in pMSCs hypoxia culture medium (pMSCs-HCM) than pMSCs normoxia culture medium (pMSCs-NCM), and the application of a specific antibody that blocked insulin-like growth factor-1 (IGF-1) leads to a significant decrease of the protective effect of pMSCs-HCM. Hypoxia can promote IGF-1 expression of pMSCs at mRNA and protein levels, and caco2 stably expressed IGF-1 receptor. Knocking down IGF-1 expression in pMSCs by siRNA resulted in a significant attenuation of the increase in apoptosis of H2O2-treated-caco2 cultured in pMSCs-HCM. In conclusion, hypoxia can increase the protective effect of pMSCs on H2O2-treated-caco2 cells via a promotion of their paracrine actions, and the key cytokine involved is IGF-1.

SUBMITTER: Du L

PROVIDER: S-EPMC3958833 | biostudies-other | 2014 Jan

REPOSITORIES: biostudies-other

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Hypoxia enhances protective effect of placental-derived mesenchymal stem cells on damaged intestinal epithelial cells by promoting secretion of insulin-like growth factor-1.

Du Lili L Yu Yanqiu Y Ma Haiying H Lu Xiaomei X Ma Ling L Jin Yunan Y Zhang Haipeng H

International journal of molecular sciences 20140127 2

Apoptosis and necrosis of intestinal epithelial cells (IECs), induced by ischemia-reperfusion (I/R) injury, can lead to dysfunction of the intestinal barrier, which could cause multiple organ dysfunction syndromes. Mesenchymal stem cells (MSCs) have the potential of providing protective effects on damaged IECs via paracrine action. This study investigated whether hypoxia can enhance the protective effect of placental-derived MSCs (pMSCs) on H2O2-treated-caco2 cells, and explored the possible mec ...[more]

PMID: 24473145

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Project description:BackgroundIschemia reperfusion injury (IRI) is the major cause of intestinal damage in clinic. Although either mesenchymal stromal cells (MSCs) or interleukin 37 (IL-37) shows some beneficial roles to ameliorate IRI, their effects are limited. In this study, the preventative effects of IL-37 gene-modified MSCs (IL-37-MSCs) on intestinal IRI are investigated.MethodsIntestinal IRI model was established by occluding the superior mesenteric artery for 30 minutes and then reperfused for 72 hours in rats. Forty adult male Sprague-Dawley rats were randomly divided into the sham control, IL-37-MSC-treated, MSC-treated, recombinant IL-37- (rIL-37-) treated, and untreated groups. Intestinal damage was assessed by H&E staining. The levels of gut barrier function factors (diamine oxidase and D-Lactate) and inflammation cytokine IL-1β were assayed using ELISA. The synthesis of tissue damage-related NLRP3 inflammasome and downstream cascade reactions including cleaved caspase-1, IL-1β, and IL-18 was detected by western blot. The mRNA levels of proinflammatory mediators IL-6 and TNF-α, which are downstream of IL-1β and IL-18, were determined by qPCR. Data were analyzed by one-way analysis of variance (ANOVA) after the normality test and followed by post hoc analysis with the least significant difference (LSD) test.ResultsIL-37-MSCs were able to migrate to the damaged tissue and significantly inhibit intestinal IRI. As compared with MSCs or the rIL-37 monotherapy group, IL-37-MSC treatment both improved gut barrier function and decreased local and systemic inflammation cytokine IL-1β level in IRI rats. In addition, tissue damage-related NLRP3 and downstream targets (cleaved caspase-1, IL-1β, and IL-18) were significantly decreased in IRI rats treated with IL-37-MSCs. Furthermore, IL-1β- and IL-18-related proinflammatory mediator IL-6 and TNF-α mRNA expressions were all significantly decreased in IRI rats treated with IL-37-MSCs.ConclusionThe results suggest that IL-37 gene modification significantly enhances the protective effects of MSCs against intestinal IRI. In addition, NLRP3-related signaling pathways could be associated with IL-37-MSC-mediated protection.

Project description:Transmissible gastroenteritis virus (TGEV) is a coronavirus characterized by diarrhea and high morbidity rates, and the mortality rate is 100% in piglets less than 2 weeks old. Pigs infected with TGEV often suffer secondary infection by other pathogens, which aggravates the severity of diarrhea, but the mechanisms remain unknown. Here, we hypothesized that persistent TGEV infection stimulates the epithelial-mesenchymal transition (EMT), and thus enterotoxigenic Escherichia coli (ETEC) can more easily adhere to generating cells. Intestinal epithelial cells are the primary targets of TGEV and ETEC infections. We found that TGEV can persistently infect porcine intestinal columnar epithelial cells (IPEC-J2) and cause EMT, consistent with multiple changes in key cell characteristics. Infected cells display fibroblast-like shapes; exhibit increases in levels of mesenchymal markers with a corresponding loss of epithelial markers; have enhanced expression levels of interleukin-1? (IL-1?), IL-6, IL-8, transforming growth factor ? (TGF-?), and tumor necrosis factor alpha (TNF-?) mRNAs; and demonstrate increases in migratory and invasive behaviors. Additional experiments showed that the activation of the phosphatidylinositol 3-kinase (PI3K)/Akt and extracellular signal-regulated kinase (ERK) signaling pathways via TGF-? is critical for the TGEV-mediated EMT process. Cellular uptake is also modified in cells that have undergone EMT. TGEV-infected cells have higher levels of integrin ?5 and fibronectin and exhibit enhanced ETEC K88 adhesion. Reversal of EMT reduces ETEC K88 adhesion and inhibits the expression of integrin ?5 and fibronectin. Overall, these results suggest that TGEV infection induces EMT in IPEC-J2 cells, increasing the adhesion of ETEC K88 in the intestine and facilitating dual infection.IMPORTANCE Transmissible gastroenteritis virus (TGEV) causes pig diarrhea and is often followed by secondary infection by other pathogens. In this study, we showed that persistent TGEV infection induces an EMT in porcine intestinal columnar epithelial cells (IPEC-J2) and enhances the adhesion of the secondary pathogen ETEC K88. Additional experiments suggest that integrin ?5 and fibronectin play an important role in TGEV-enhanced ETEC K88 adhesion. Reversal of EMT reduces the expression of integrin ?5 and fibronectin and also reduces ETEC K88 adhesion. We conclude that TGEV infection triggers EMT and facilitates dual infection. Our results provide new insights into secondary infection and suggest that targeted anti-EMT therapy may have implications for the prevention and treatment of secondary infection.

Project description: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.

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Hypoxia enhances protective effect of placental-derived mesenchymal stem cells on damaged intestinal epithelial cells by promoting secretion of insulin-like growth factor-1.

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Hypoxia enhances protective effect of placental-derived mesenchymal stem cells on damaged intestinal epithelial cells by promoting secretion of insulin-like growth factor-1.

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