Miaomiao Fan

Ghrelin improves functional survival of engrafted adipose-derived mesenchymal stem cells in ischemic heart through PI3K/Akt signaling pathway.

Mesenchymal stem cells (MSCs) have been proposed as a promising cell population for cell therapy and regenerative medicine applications. However, the low retention and poor survival of engrafted cells hampered the therapeutic efficacy of engrafted MSCs. Ghrelin is a 28-amino-acid peptide hormone and is proved to exert a protective effect on the cardiovascular system. This study is designed to investigate the protective effects of ghrelin on engrafted adipose-derived mesenchymal stem cells (ADMSCs) and its beneficial effects with cellular therapy in mice myocardial infarction (MI). Results showed that intramyocardial injection of ADMSCs combining with ghrelin administration inhibited host cardiomyocyte apoptosis, reduced fibrosis, and improved cardiac function. To reveal possible mechanisms, ADMSCs were subjected to hypoxia/serum deprivation (H/SD) injury to simulate ischemic conditions in vivo. Ghrelin (10−8 M, 33712 pg/ml) improved ADMSCs survival under H/SD condition. Western blot assay revealed that ghrelin increased AKT phosphorylation both in vivo and in vitro, decreased the proapoptotic protein Bax, and increased the antiapoptotic protein Bcl-2 in vitro, while these effects were abolished by PI3K inhibitor LY294002. These revealed that ghrelin may serve as a promising candidate for hormone-driven approaches to improve the efficacy of mesenchymal stem cell-based therapy for cardiac ischemic disease via PI3K/AKT pathway.

Dihydromyricetin Protects against Diabetic Cardiomyopathy in Streptozotocin-Induced Diabetic Mice

Diabetic cardiomyopathy (DCM) is an important cause of heart failure in diabetic patients. The present study sought to explore the potential effects of dihydromyricetin (DHM) on DCM and its possible mechanism. A diabetic model was induced by intraperitoneal injection of streptozotocin (STZ) in C57BL/6J mice. Two weeks after the STZ injection, mice were randomly allocated into the following 4 groups for treatment: the control group (CON), the control treated with DHM group (CON + DHM), the diabetes group (DM), and the diabetes treated with DHM group (DM + DHM). DHM was dissolved in distilled water and administered daily by gavage. For 14 weeks, the CON + DHM group and DM + DHM group were given a dose of 100 mg/kg/day DHM (Sigma-Aldrich), while the CON and DM groups were intragastrically given equivalent volumes of distilled water. Assessments and comparisons were made among the groups based on cardiac function and structural changes, inflammation factors, markers of oxidative stress, mitochondria function, apoptosis, and autophagy. The DHM treatment normalized body weight, preserved cardiac function, attenuated oxidative stress (MDA, SOD, and GSH-Px), reduced the levels of inflammation factors (IL-6, TNF-α), alleviated pathological changes, improved mitochondrial function (ATP content, CS activity, and complex Ι/ΙΙ/ΙΙΙ/ΙV/V activities), inhibited cardiac apoptosis, and restored autophagy in diabetic mice. DHM may have a great therapeutic potential in the treatment of DCM.

SIRT1 Activation by Resveratrol Alleviates Cardiac Dysfunction via Mitochondrial Regulation in Diabetic Cardiomyopathy Mice

Background Diabetic cardiomyopathy (DCM) is a major threat for diabetic patients. Silent information regulator 1 (SIRT1) has a regulatory effect on mitochondrial dynamics, which is associated with DCM pathological changes. Our study aims to investigate whether resveratrol, a SRIT1 activator, could exert a protective effect against DCM. Methods and Results Cardiac-specific SIRT1 knockout (SIRT1KO) mice were generated using Cre-loxP system. SIRT1KO mice displayed symptoms of DCM, including cardiac hypertrophy and dysfunction, insulin resistance, and abnormal glucose metabolism. DCM and SIRT1KO hearts showed impaired mitochondrial biogenesis and function, while SIRT1 activation by resveratrol reversed this in DCM mice. High glucose caused increased apoptosis, impaired mitochondrial biogenesis, and function in cardiomyocytes, which was alleviated by resveratrol. SIRT1 deletion by both SIRT1KO and shRNA abolished the beneficial effects of resveratrol. Furthermore, the function of SIRT1 is mediated via the deacetylation effect on peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α), thus inducing increased expression of nuclear respiratory factor 1 (NRF-1), NRF-2, estrogen-related receptor-α (ERR-α), and mitochondrial transcription factor A (TFAM). Conclusions Cardiac deletion of SIRT1 caused phenotypes resembling DCM. Activation of SIRT1 by resveratrol ameliorated cardiac injuries in DCM through PGC-1α-mediated mitochondrial regulation. Collectively, SIRT1 may serve as a potential therapeutic target for DCM.

Therapeutic efficacy of apelin on transplanted mesenchymal stem cells in hindlimb ischemic mice via regulation of autophagy

Mesenchymal stem cells (MSCs)-based therapy provides a promising avenue for the management of peripheral arterial disease (PAD). However, engrafted MSCs are subjected to acute cell death in the ischemic microenvironment. Apelin has been shown to protect bone marrow MSCs against apoptosis although the mechanism of action remains elusive. Here we demonstrated that apelin promoted functional survival of AD-MSCs in ischemic hindlimbs and provoked a synergetic effect with AD-MSCs to restore hindlimb blood perfusion and limb functions. Further in vitro studies revealed that a biphasic response in autophagy was induced by apelin in AD-MSCs during hypoxia and hypoxia/reoxygenation (H/R) stages to exert cytoprotective effects against H/R injury. Mechanistically, apelin increased the viability of AD-MSCs via promoting protective autophagy during hypoxia, which was accompanied with activation of AMPK and inhibition of mammalian target of rapamycin (mTOR). To the contrary, apelin suppressed autophagic cell death during reoxygenation, which was accompanied with activation of Akt and inhibition of Beclin1. Our findings indicated that apelin facilitated AD-MSCs-based therapy in PAD, possibly through promoting survival of AD-MSCs by way of autophagy regulation. Our data support the promises of apelin as a novel strategy to improve MSC-based therapy for PAD, possibly through autophagy modulation in MSCs.

Endothelial deletion of mTORC1 protects against hindlimb ischemia in diabetic mice via activation of autophagy, attenuation of oxidative stress and alleviation of inflammation

Abstract Peripheral arterial disease (PAD) complicated with diabetes mellitus (DM) still remains a thorny issue due to lack of effective strategies. Our previous study has demonstrated that inhibition of mTORC1 protected adipose‐derived stromal cells from hindlimb ischemic injury in PAD mice. However, whether inhibition of mTORC1 could protect against PAD in diabetes mellitus and the underlying mechanisms remained elusive. In this study, we employed endothelial‐specific raptor (an essential component of the mTORC1 signaling complex) knockout (KO) mice (Tie2‐mTORC1ko) to investigate whether and how mTORC1 downregulation could alleviate hindlimb ischemic injury in diabetic mice. Tie2‐mTORC1ko mice and their wild‐type littermates were intraperitoneally injected with streptozocin to induce type 1 diabetic model, after which the hyperglycemic mice were randomly allocated to sham operation or PAD operation (femoral artery ligation). The restoration of hindlimb blood perfusion and recovery of limb functions were improved in diabetic Tie2‐mTORC1ko PAD mice with significant improvements of autophagy, angiogenesis and vascular integrity as well as attenuation of apoptosis, inflammation and oxidative stress. In vitro, high glucose combining with hypoxia/serum deprivation treatment (HG+H/SD) significantly triggered apoptosis, reactive oxygen species generation and inflammation while inhibited autophagy and tube formation in HUVECs. The effect could be accentuated and attenuated by mTORC1 over‐expression (TSC2 siRNA) and mTORC1 silencing (raptor siRNA), respectively. Moreover, autophagy inhibitor 3‐MA could simulate the effects of TSC2 siRNA while autophagy inducer rapamycin could mimic the effects of raptor siRNA, suggesting that the beneficial effects of mTORC1 deletion were associated with autophagy induction. In conclusion, our present study demonstrates that endothelial mTORC1 deletion protects against hindlimb ischemic injury in diabetic mice possibly via activation of autophagy, attenuation of oxidative stress and alleviation of inflammation. Therapeutics targeting mTORC1 may therefore represents a promising strategy to rescue limb ischemia in diabetes mellitus. Graphical abstract Figure. No Caption available. HighlightsEndothelial deletion of mTORC1 protects agaisnst limb ischemia in diabetic mice.mTORC1 silencing preserved endothelial viability and function.mTORC1 silencing restored impaired autophagy to scavenge damaged mitochondria.mTORC1 deletion attenuated oxidative stress in ischemsic gastrocnemius muscle.mTORC1‐p70S6K axis exhibited pro‐inflammatory potential.

Activation of Cannabinoid Receptor Type II by AM1241 Ameliorates Myocardial Fibrosis via Nrf2-Mediated Inhibition of TGF-β1/Smad3 Pathway in Myocardial Infarction Mice.

Aims: Myocardial interstitial fibrosis is a major histologic landmark resulting in cardiac dysfunction after myocardial infarction (MI). Activation of cannabinoid receptor type II (CB2 receptor) have been demonstrated to reduce fibrosis in hepatic cirrhotic rat. However, the anti-fibrotic effect of CB2 receptor activation in infarcted hearts was still unclear. In this study, we aimed to investigate the effects of a CB2 receptor selective agonist AM1241 on myocardial fibrosis post MI in mice. Methods: Echocardiograph was conducted to assess cardiac function. Fibrosis markers such as type I and type III collagen, fibronectin, Plasminogen activator inhibitor(PAI)-1 and tissue inhibitor of metalloprotease(TIMP)-1 were examined by Western blot, while collagens were directly observed by Sirius-red staining. Primary cultured cardiac fibroblasts(CFs) were subjected to hypoxia/serum deprivation (H/SD) injury to simulate ischemic conditions in vivo. Nrf2 siRNA were applied to explore the role of Nrf2 and TGF-β1/Smad3 pathway in this process. Results: Echocardiography showed that AM1241 significantly improved cardiac function, suppressed the expression of fibrosis markers such as collagen I and collagen III, fibronectin, PAI-1 and TIMP-1 in mice with MI. In cardiac fibroblasts subjected to H/SD injury, AM1241 reduced the elevated levels of α-SMA, collagen I and collagen III, which were partially abrogated by the Nrf2 siRNA transfection. Furthermore, AM1241 not only activated and accelerated the translocation of Nrf2 to nucleus, but also inhibited TGF-β1/ Smad3 pathway in an Nrf2 dependent manner. Conclusion: CB2 receptor agonist AM1241 alleviated myocardial interstitial fibrosis via Nrf2 -mediated down-regulation of TGF-β1/Smad3 pathway, which suggested that CB2 receptor activation might represent a promising target for retarding cardiac fibrosis after MI.

Cardiotrophin-1 promotes cardiomyocyte differentiation from mouse induced pluripotent stem cells via JAK2/STAT3/Pim-1 signaling pathway

Background The induced pluripotent stem cell (iPSC) has shown great potential in cellular therapy of myocardial infarction (MI), while its application is hampered by the low efficiency of cardiomyocyte differentiation. The present study was designed to investigate the effects of cardiotrophin-1 (CT-1) on cardiomyocyte differentiation from mouse induced pluripotent stem cells (miPSCs) and the underlying mechanisms involved. Methods The optimal treatment condition for cardiomyocyte differentiation from miPSCs was established with ideal concentration (10 ng/mL) and duration (from day 3 to day 14) of CT-1 administration. Up-regulated expression of cardiac specific genes that accounted for embryonic cardiogenesis was observed by quantitative RT-PCR. Elevated amount of α-myosin heavy chain (α-MHC) and cardiac troponin I (cTn I) positive cells were detected by immunofluorescence staining and flow cytometry analysis in CT-1 group. Results Transmission electron microscopic analysis revealed that cells treated with CT-1 showed better organized sacromeric structure and more mitochondria, which are morphological characteristic of matured cardiomyocytes. Western blot demonstrated that CT-1 promotes cardiomyocyte differentiation from miPSCs partly via JAK2/STAT3/Pim-1 pathway as compared with control group. Conclusions These findings suggested that CT-1 could enhance the cardiomyocyte differentiation as well as the maturation of mouse induced pluripotent stem cell derived cardiomyocytes by regulating JAK2/STAT3/Pim-1signaling pathway.

Reduced silent information regulator 1 signaling exacerbates sepsis-induced myocardial injury and mitigates the protective effect of a liver X receptor agonist

Abstract Myocardial injury and dysfunction are critical manifestations of sepsis. Previous studies have reported that liver X receptor (LXR) activation is protective during sepsis. However, whether LXR activation protects against septic heart injury and its underlying mechanisms remain elusive. This study was designed to determine the role of LXR activation in the septic heart with a focus on SIRT1 (silent information regulator 1) signaling. Male cardiac‐specific SIRT1 knockout mice (SIRT1‐/‐) and their wild‐type littermates were subjected to sepsis by cecal ligation and puncture (CLP) in the presence or absence of LXR agonist T0901317. The survival rate of mice was recorded during the 7‐day period post CLP. Our results demonstrated that SIRT1‐/‐ mice suffered from exacerbated mortality and myocardial injury in comparison with their wild‐type littermates. Meanwhile, T0901317 treatment improved mice survival, accompanied by significant ameliorations of myocardial injury and dysfunction in wild‐type mice but not in SIRT1‐/‐ mice. Furthermore, the levels of myocardial inflammatory cytokines (TNF‐&agr;, IL‐6, IL‐1&bgr;, MCP‐1, MPO and HMGB1), oxidative stress (ROS generation, MDA), endoplasmic‐reticulum (ER) stress (protein levels of CHOP, GRP78, GRP94, IRE1&agr;, and ATF6), and cardiac apoptosis following CLP were inhibited by T0901317 treatment in wild‐type mice but not in SIRT1‐/‐ mice. Mechanistically, T0901317 enhanced SIRT1 signaling and the subsequent deacetylation and activation of antioxidative FoxO1 and anti‐ER stress HSF1, as well as the deacetylation and inhibition of pro‐inflammatory NF‐KB and pro‐apoptotic P53, thereby alleviating sepsis‐induced myocardial injury and dysfunction. Our data support the promise of LXR activation as an effective strategy for relieving heart septic injury. Graphical abstract No caption available. HighlightsReduced SIRT1 signaling exacerbates sepsis‐induced myocardial injury.LXR agonist T0901317 mitigates sepsis‐induced myocardial injury.T0901317 attenuates oxidative stress, ER stress, inflammation, and apoptosis.SIRT1 signaling is strongly involved in the protective effects of LXR agonist.

Activation of cannabinoid receptor type II by AM1241 protects adipose-derived mesenchymal stem cells from oxidative damage and enhances their therapeutic efficacy in myocardial infarction mice via Stat3 activation

The poor survival of cells in ischemic sites diminishes the therapeutic efficacy of stem cell therapy. Previously we and others have reported that Cannabinoid receptor type II (CB2) is protective during heart ischemic injury for its anti-oxidative activity. However, whether CB2 activation could improve the survival and therapeutic efficacy of stem cells in ischemic myocardium and the underlying mechanisms remain elusive. Here, we showed evidence that CB2 agonist AM1241 treatment could improve the functional survival of adipose-derived mesenchymal stem cells (AD-MSCs) in vitro as well as in vivo. Moreover, AD-MSCs adjuvant with AM1241 improved cardiac function, and inhibited cardiac oxidative stress, apoptosis and fibrosis. To unveil possible mechanisms, AD-MSCs were exposed to hydrogen peroxide/serum deprivation to simulate the ischemic environment in myocardium. Results delineated that AM1241 blocked the apoptosis, oxidative damage and promoted the paracrine effects of AD-MSCs. Mechanistically, AM1241 activated signal transducers and activators of transcription 3 (Stat3) through the phosphorylation of Akt and ERK1/2. Moreover, the administration of AM630, LY294002, U0126 and AG490 (inhibitors for CB2, Akt, ERK1/2 and Stat3, respectively) could abolish the beneficial actions of AM1241. Our result support the promise of CB2 activation as an effective strategy to optimize stem cell-based therapy possibly through Stat3 activation.The poor survival of cells in ischemic sites diminishes the therapeutic efficacy of stem cell therapy. Previously we and others have reported that Cannabinoid receptor type II (CB2) is protective during heart ischemic injury for its anti-oxidative activity. However, whether CB2 activation could improve the survival and therapeutic efficacy of stem cells in ischemic myocardium and the underlying mechanisms remain elusive. Here, we showed evidence that CB2 agonist AM1241 treatment could improve the functional survival of adipose-derived mesenchymal stem cells (AD-MSCs) in vitro as well as in vivo. Moreover, AD-MSCs adjuvant with AM1241 improved cardiac function, and inhibited cardiac oxidative stress, apoptosis and fibrosis. To unveil possible mechanisms, AD-MSCs were exposed to hydrogen peroxide/serum deprivation to simulate the ischemic environmentin myocardium. Results delineated that AM1241 blocked the apoptosis, oxidative damage and promoted the paracrine effects of AD-MSCs. Mechanistically, AM1241 activated signal transducers and activators of transcription 3 (Stat3) through the phosphorylation of Akt and ERK1/2. Moreover, the administration of AM630, LY294002, U0126 and AG490 (inhibitors for CB2, Akt, ERK1/2 and Stat3, respectively) could abolish the beneficial actions of AM1241. Our result support the promise of CB2 activation as an effective strategy to optimize stem cell-based therapy possibly through Stat3 activation.

A Novel Mechanism of Mesenchymal Stromal Cell-Mediated Protection against Sepsis: Restricting Inflammasome Activation in Macrophages by Increasing Mitophagy and Decreasing Mitochondrial ROS

Sepsis, a systemic inflammatory response to infection, is the leading cause of death in the intensive care unit (ICU). Previous studies indicated that mesenchymal stromal cells (MSCs) might have therapeutic potential against sepsis. The current study was designed to investigate the effects of MSCs on sepsis and the underlying mechanisms focusing on inflammasome activation in macrophages. The results demonstrated that the bone marrow-derived mesenchymal stem cells (BMSCs) significantly increased the survival rate and organ function in cecal ligation and puncture (CLP) mice compared with the control-grouped mice. BMSCs significantly restricted NLRP3 inflammasome activation, suppressed the generation of mitochondrial ROS, and decreased caspase-1 and IL-1β activation when cocultured with bone marrow-derived macrophages (BMDMs), the effects of which could be abolished by Mito-TEMPO. Furthermore, the expression levels of caspase-1, IL-1β, and IL-18 in BMDMs were elevated after treatment with mitophagy inhibitor 3-MA. Thus, BMSCs exert beneficial effects on inhibiting NLRP3 inflammasome activation in macrophages primarily via both enhancing mitophagy and decreasing mitochondrial ROS. These findings suggest that restricting inflammasome activation in macrophages by increasing mitophagy and decreasing mitochondrial ROS might be a crucial mechanism for MSCs to combat sepsis.