Indoleamine 2,3-dioxgenase-transfected mesenchymal stem cells suppress heart allograft rejection by increasing the production and activity of dendritic cells and regulatory T cells.

Expression of indoleamine 2,3-dioxygenase (IDO) in mesenchymal stem cells (MSC) is thought to contribute to MSC-mediated immunosuppression. A lentiviral-based transgenic system was used to generate bone marrow stem cells (BMSC) which stably expressed IDO (IDO-BMSCs). Coculture of IDO-BMSCs with dendritic cells (DC) or T cells was used to evaluate the immunomodulatory effect of IDO-BMSCs. A heterotopic heart transplant model in rats was used to evaluate allograft rejection after IDO-BMSC treatment. Mechanisms of IDO-BMSC-mediated immunosuppression were investigated by evaluating levels of proinflammatory and anti-inflammatory cytokines, and production of Tregs. A significant decrease in DC marker-positive cells and a significant increase in Tregs were observed in IDO-BMSC cocultured. Treatment of transplanted rats with IDO-BMSCs was associated with significantly prolonged graft survival. Compared with the control groups, transplanted animals treated with IDO-BMSCs had a (1) significantly higher ejection fraction and fractional shortening, (2) significantly lower expression of CD86, CD80, and MHCII, and significantly higher expression in CD274, and Tregs, and (3) significantly higher levels of interleukin-10 (IL-10), transforming growth factor beta-1 (TGF-ß1), TGF-ß2, and TGF-ß3, and significantly lower levels of IL-2 and interferon gamma. Our results expand our understanding of the molecular mechanisms underlying suppression of heart allograft rejection via IDO-expressing BMSCs.

Bone marrow mesenchymal stem cells overexpressing GATA-4 improve cardiac function following myocardial infarction.

INTRODUCTION: The present study aimed to examine whether GATA-4 overexpressing bone marrow mesenchymal stem cells can improve cardiac function in a murine myocardial infarction model compared with bone marrow mesenchymal stem cells alone. METHODS: A lentiviral-based transgenic system was used to generate bone mesenchymal stem cells which stably expressed GATA-4 (GATA-4-bone marrow mesenchymal stem cells). Apoptosis and the myogenic phenotype of the bone marrow mesenchymal stem cells were measured using Western blot and immunofluorescence assays co-cultured with cardiomyocytes. Cardiac function, bone marrow mesenchymal stem cell homing, cardiac cell apoptosis, and vessel number following transplantation were assessed, as well as the expression of c-Kit. RESULTS: In GATA-4-bone marrow mesenchymal stem cells-cardiomyocyte co-cultures, expression of myocardial-specific antigens, cTnT, connexin-43, desmin, and α-actin was increased compared with bone marrow mesenchymal stem cells alone. Caspase 8 and cytochrome C expression was lower, and the apoptotic rate was significantly lower in GATA-4 bone marrow mesenchymal stem cells. Cardiac function following myocardial infarction was also increased in the GATA-4 bone marrow mesenchymal stem cell group as demonstrated by enhanced ejection fraction and left ventricular fractional shortening. Analysis of the cardiac tissue revealed that the GATA-4 bone marrow mesenchymal stem cell group had a greater number of DiR-positive cells suggestive of increased homing and/or survival. Transplantation with GATA-4-bone marrow mesenchymal stem cells significantly increased the number of blood vessels, decreased the proportion of apoptotic cells, and increased the mean number of cardiac c-kit-positive cells. CONCLUSION: GATA-4 overexpression in bone marrow mesenchymal stem cells exerts anti-apoptotic effects by targeting cytochrome C and Fas pathways, promotes the aggregation of bone marrow mesenchymal stem cells in cardiac tissue, facilitates angiogenesis, and effectively mobilizes c-kit-positive cells following myocardial infarction, leading to the improvement of cardiac function after MI.

Exosomes Derived from IDO1-Overexpressing Rat Bone Marrow Mesenchymal Stem Cells Promote Immunotolerance of Cardiac Allografts.

BACKGROUND: The immunosuppressive activity of mesenchymal stem cells (MSCs) has been exploited to induce tolerance after organ transplantation. The indoleamine 2,3-dioxygenase (IDO) may have beneficial effects in the immunoregulatory properties of MSCs. It was recently revealed that exosomes derived from MSCs play important roles in mediating the biological functions of MSCs. This study aimed to explore the roles of exosomes derived from MSCs in the induction of immune tolerance. METHODS: Dendritic cells (DCs) and T-cells were cultured with exosomes derived from rat bone marrow MSCs (BMSCs) overexpressing IDO1 or controls. For the in-vivo study, rats received heart transplants and were treated with exosomes from IDO-BMSCs and heart function was evaluated. Flow cytometry was used to detect expression of cell surface markers. Cytokine levels were detected in culture supernatants or serum samples. Protein and microRNA expressions in exosomes were investigated by chips. RESULTS: Exosomes from IDO-BMSCs cultured with DCs and T-cells (1) downregulated CD40, CD86, CD80, MHC-II, CD45RA, CD45RA+CD45RB, OX62, and upregulated CD274 expression, (2) increased the number of regulatory T-cells (Tregs) and decreased the number of CD8+ T-cells, and (3) decreased the levels of pro-inflammatory cytokines, but increased the levels of anti-inflammatory cytokines compared with the other groups. Transplanted rats, which were injected with exosomes from IDO-BMSCs, had reduced allograft-targeting immune responses and improved cardiac allograft function. Exosomes secreted by IDO-BMSCs exhibited significant upregulations of the immunoregulatory protein FHL-1, miR-540-3p, and a downregulation of miR-338-5p. CONCLUSION: Exosomes derived from IDO-BMSCs can be used to promote immunotolerance and prolong the survival of cardiac allografts.

ETHYLENE RESPONSE FACTOR 74 (ERF74) plays an essential role in controlling a respiratory burst oxidase homolog D (RbohD)-dependent mechanism in response to different stresses in Arabidopsis.

Recent studies indicate that the ETHYLENE RESPONSE FACTOR VII (ERF-VII) transcription factor is an important regulator of osmotic and hypoxic stress responses in plants. However, the molecular mechanism of ERF-VII-mediated transcriptional regulation remains unclear. Here, we investigated the role of ERF74 (a member of the ERF-VII protein family) by examining the abiotic stress tolerance of an ERF74 overexpression line and a T-DNA insertion mutant using flow cytometry, transactivation and electrophoretic mobility shift assays. 35S::ERF74 showed enhanced tolerance to drought, high light, heat and aluminum stresses, whereas the T-DNA insertion mutant erf74 and the erf74;erf75 double mutant displayed higher sensitivity. Using flow cytometry analysis, we found that erf74 and erf74;erf75 lines lack the reactive oxygen species (ROS) burst in the early stages of various stresses, as a result of the lower expression level of RESPIRATORY BURST OXIDASE HOMOLOG D (RbohD). Furthermore, ERF74 directly binds to the promoter of RbohD and activates its expression under different abiotic stresses. Moreover, induction of stress marker genes and ROS-scavenging enzyme genes under various stress conditions is dependent on the ERF74-RbohD-ROS signal pathway. We propose a pathway that involves ERF74 acting as an on-off switch controlling an RbohD-dependent mechanism in response to different stresses, subsequently maintaining hydrogen peroxide (H2 O2 ) homeostasis in Arabidopsis.

[The crystal behavior of calcium carbonate in water-soluable chitin].

Based on the basic principles of biominerlization, the paper analyses calcium carbonate crystallization in waterable chitin solution under the control of chitin, using chitin as the matrix; and analyses the effect on crystals by varying temperature or pH of the system. The obtained calcium carbonate was characterized by Fourier transform infrared spectroscopy (FTIR), scanning electronic microscopy(SEM) and X ray powder diffraction (XRD). As a result, it was found that crystals were different formed in purity water; and the obtained crystals are different in different concentration chitin solution. Calcium carbonate has effect on chitin during the calcium carbonate formation process, so there is the interaction between chitin and calcium carbonate.