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Exosomes Derived From Mesenchymal Stem Cells Ameliorate Renal Ischemic-Reperfusion Injury Through Inhibiting Inflammation and Cell Apoptosis.

ABSTRACT: This study aimed to investigate the underlying mechanism of mesenchymal stem cells (MSCs) on protection of renal ischemia reperfusion injury (IRI). Exosomes originated from MSCs (MSC-ex) were extracted according to the instructions of Total Exosome Isolation Reagent. Rats were divided into five groups: sham-operated, IRI, MSC, MSC-ex, and MSC-ex + RNAase group. MSCs or MSC-ex were injected via carotid artery. The renal function test and pathological detection were applied to determine the renoprotection of MSC-ex on IRI. Western blotting and quantitative reverse transcription polymerase chain reaction (RT-qPCR) were conducted to examine the levels of apoptosis-related proteins and inflammatory cytokines. Our results revealed that MSC-derived exosomes attenuated renal dysfunction, histologic damage, and decreased apoptosis. The expression levels of inflammatory cytokines, such as interleukin 6 (IL-6), tumor necrosis factor-? (TNF-?), nuclear factor kappa B (NF-?B), and interferon gamma (IFN-?), were decreased by the MSC-ex treatment. The expression levels of caspase-9, cleaved caspase-3, Bax, and Bcl-2 caused by IR were also inhibited by MSC-ex. MSC-ex + RNAase group shared the similar pattern of changes with IRI group, likely due to the ability of RNA hydrolase to eliminate the function of exosomes. Our results demonstrated that exosomes originating from MSCs have protective effects on IRI via inhibiting cell apoptosis and inflammatory responses. Out findings may provide a new insight into therapeutic mechanism of MSCs on renal IRI.

SUBMITTER: Li L

PROVIDER: S-EPMC6907421 | biostudies-literature | 2019

REPOSITORIES: biostudies-literature

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Exosomes Derived From Mesenchymal Stem Cells Ameliorate Renal Ischemic-Reperfusion Injury Through Inhibiting Inflammation and Cell Apoptosis.

Li Long L Wang Rulin R Jia Yichen Y Rong Ruiming R Xu Ming M Zhu Tongyu T

Frontiers in medicine 20191119

This study aimed to investigate the underlying mechanism of mesenchymal stem cells (MSCs) on protection of renal ischemia reperfusion injury (IRI). Exosomes originated from MSCs (MSC-ex) were extracted according to the instructions of Total Exosome Isolation Reagent. Rats were divided into five groups: sham-operated, IRI, MSC, MSC-ex, and MSC-ex + RNAase group. MSCs or MSC-ex were injected via carotid artery. The renal function test and pathological detection were applied to determine the renopr ...[more]

PMID: 31867333

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Project description:IntroductionClinically, a good deal of injury from stroke results from ischemic-reperfusion. There is a loss of cerebral parenchyma and its associated cells, disruption of neuronal connections, compromise of the blood-brain barrier, and inflammation. We tested whether exogenously engrafted human neural stem cells could migrate rapidly and extensively to damaged regions, following transplantation into a neurogenic site where migration cues are already underway during stroke onset, then counteract a number of these pathological processes.MethodsOne day post-injury, we injected human neural stem cells (hNSCs) into the ipsilesional hippocampus of a mouse model of stroke with middle cerebral artery occlusion to induce focal ischemia followed by reperfusion (MCAO/R). The time frame for hNSC transplantation corresponded to upregulation of endogenous proinflammatory cytokines. We examined the effect of hNSC transplantation on pathological processes and behavioral dysfunction 48 hours post-injury.ResultsTwenty-four hours after transplantation, engrafted hNSCs had migrated extensively to the lesion, and infarct volume was reduced relative to MCAO/R controls. The behavioral deficits seen in MCAO/R controls were also significantly improved. Given this rapid response, we hypothesized that the mechanisms underlying therapeutic activity were anti-inflammatory rather than due to cell replacement. In support of this idea, in hNSC-transplanted mice we observed reduced microglial activation, decreased expression of proinflammatory factors (tumor necrosis factor-α, interleukin (IL)-6, IL-1β, monocyte chemotactic protein-1, macrophage inflammatory protein-1α) and adhesion molecules (intercellular adhesion molecule-1, vascular cell adhesion molecule-1), and amelioration of blood-brain barrier damage.ConclusionsWhile long-term effects of engrafted hNSCs on the amelioration of ischemic stroke-induced behavioral dysfunction in a rodent model have been reported, our study is the first to show rapid, beneficial impacts on behavioral function (within 24 hours) upon early delivery of hNSCs into the hippocampus.

Project description:BackgroundAcute kidney injury (AKI) is an emerging global healthcare issue without effective therapy yet. Autophagy recycles damaged organelles and helps maintain tissue homeostasis in acute renal ischemia-reperfusion (I/R) injury. Hypoxic mesenchymal stem cells (HMSCs) represent an innovative cell-based therapy in AKI. Moreover, the conditioned medium of HMSCs (HMSC-CM) rich in beneficial trophic factors may serve as a cell-free alternative therapy. Nonetheless, whether HMSCs or HMSC-CM mitigate renal I/R injury via modulating tubular autophagy remains unclear.MethodsRenal I/R injury was induced by clamping of the left renal artery with right nephrectomy in male Sprague-Dawley rats. The rats were injected with either PBS, HMSCs, or HMSC-CM immediately after the surgery and sacrificed 48 h later. Renal tubular NRK-52E cells subjected to hypoxia-reoxygenation (H/R) injury were co-cultured with HMSCs or treated with HMSC-CM to assess the regulatory effects of HSMCs on tubular autophagy and apoptosis. The association of tubular autophagy gene expression and renal recovery was also investigated in patients with ischemic AKI.ResultHMSCs had a superior anti-oxidative effect in I/R-injured rat kidneys as compared to normoxia-cultured mesenchymal stem cells. HMSCs further attenuated renal macrophage infiltration and inflammation, reduced tubular apoptosis, enhanced tubular proliferation, and improved kidney function decline in rats with renal I/R injury. Moreover, HMSCs suppressed superoxide formation, reduced DNA damage and lipid peroxidation, and increased anti-oxidants expression in renal tubular epithelial cells during I/R injury. Co-culture of HMSCs with H/R-injured NRK-52E cells also lessened tubular cell death. Mechanistically, HMSCs downregulated the expression of pro-inflammatory interleukin-1β, proapoptotic Bax, and caspase 3. Notably, HMSCs also upregulated the expression of autophagy-related LC3B, Atg5 and Beclin 1 in renal tubular cells both in vivo and in vitro. Addition of 3-methyladenine suppressed the activity of autophagy and abrogated the renoprotective effects of HMSCs. The renoprotective effect of tubular autophagy was further validated in patients with ischemic AKI. AKI patients with higher renal LC3B expression were associated with better renal recovery.ConclusionThe present study describes that the enhancing effect of MSCs, and especially of HMCSs, on tissue autophagy can be applied to suppress renal tubular apoptosis and attenuate renal impairment during renal I/R injury in the rat. Our findings provide further mechanistic support to HMSCs therapy and its investigation in clinical trials of ischemic AKI.

Project description:Acute kidney injury (AKI) is associated with an abrupt loss of kidney function that results in significant morbidity and mortality. Considerable effort has focused around the identification of diagnostic biomarkers and the analysis of molecular events. Most studies have adopted organ-wide approaches that do not fully capture the interplay among different cell types in the pathophysiology of AKI. To extend our understanding of molecular and cellular events in AKI, we developed a mouse line that enables the identification of translational profiles in specific cell types by CRE recombinase-dependent activation of an eGFP-tagged L10a ribosomal protein subunit, and consequently, translating ribosome affinity purification (TRAP) of mRNA populations. By utilizing cell-type specific CRE-driver lines, in this study we identify distinct cellular responses in an ischemia reperfusion injury (IRI) model of AKI. Cell-specific translational expression profiles were uncovered 24 hours after IRI from four populations enriched for distinct anatomical and cellular subgroups: nephron, interstitial cell populations, vascular endothelium, and macrophages/monocytes by Affymetrix microarray. A construct containing the CAGGS promoter driving eGFP-L10a, with a loxP-site flanked triple SV40 polyA cassette between promoter and eGFP-L10a cassette was targeted into the ubiquitously active Rosa26 locus. The upstream polyA cassette is designed to block activity of the downstream eGFP-L10a cassette. CRE-dependent removal of this transcriptional block activates eGFP::L10a production within the CRE-producing cell, and all of its descendants. Mice carrying the conditional eGFP-L10a allele, referred to as L10a, were maintained in a homozygous state. L10a mice were crossed to four CRE strains to activate eGFP::L10a expression in four predominantly non-overlapping cellular compartments in the kidney. A Six2-Tet-GFP::CRE allele is active exclusively within nephron progenitors; consequently, historical labeling results in eGFP::L10a expression throughout the main body of the nephron. A Foxd1-GFP::CRE allele is active in the progenitors of many of the interstitial cell lineages including those generating mesangial and non-glomerular pericytes. In addition, Foxd1 is normally expressed in podocytes. Cdh5-CRE is reported to be active throughout the vascular endothelium, and finally, Lyz2-CRE specifically labels cells of the myeloid lineage, notably macrophages, monocytes and dendritic cells. Mice carrying any CRE allele and the L10a allele are designated generically CRE-L10a. six2-L10a, foxd1-L10a, cdh5-L10a and lyz2-L10a denote specifically mice that are compound heterozygotes for the indicated CRE driver and L10a. CRE-L10a, L10a heterozygous littermates without CRE allele, C57BL/6 wild type mice were subjected to renal bilateral warm ischemia 28 minutes followed by 24-hour reperfusion when the kidney TRAP RNA and total RNA were isolated and subjected to Affymetrix microarray. Biological triplicates for each CRE-L10a line underwent no Surgery; sham Surgery and IRI treatment.

Project description:BackgroundHuman umbilical cord mesenchymal stem cells (UC-MSCs) are multipotent progenitor cells representing an attractive therapeutic tool for tissue damage and inflammation owing to their unique immunomodulatory properties. This study was designed to determine the protective effects and underlying mechanisms of UC-MSCs on acute liver failure (ALF).MethodsALF was induced in mice by intraperitoneal injection of D-galactosamine (D-GalN) and lipopolysaccharide (LPS). Mice were intravenously injected with 1×106 UC-MSCs one hour before or six hours after D-GalN/LPS injection. Liver function was valued by serum biochemical parameters and hematoxylin-eosin staining. Inflammatory cytokine and chemokine levels were measured by real-time PCR, and inflammatory cells infiltration was observed by immunofluorescence staining. Hepatocyte apoptosis and pyroptosis related proteins were detected by western blot. Murine macrophage Raw264.7 in the presentation of LPS was treated with the UC-MSCs condition medium (UC-MSCs-CM), and then the levels of inflammatory cytokines and NOD-like receptor family pyrin domain containing 3 (NLRP3) inflammasome in Raw264.7 were measured.ResultsUC-MSCs significantly reduced the mortality, decreased serum alanine aminotransferase and aspartate aminotransferase levels, and improved the pathological damage. Moreover, UC-MSCs inhibited inflammatory cytokine and chemokine levels, especially TNF-α, interleukins-6 (IL-6), IL-1β, monocyte chemoattractant protein (MCP-1), CC-chemokines ligand 2 (CCL2), C-X-C motif ligand 2 (CXCL2), and reduced macrophage, neutrophil and T lymphocyte infiltration into the liver tissue. UC-MSCs also attenuated hepatocyte apoptosis, as evidenced by decreased TUNEL positive cells, increased Bcl-xl/Bax protein ratio and downregulated cleaved caspase 3 levels. NLRP3 inflammasome activation, IL-1β maturation and cleaved caspase1 were suppressed by UC-MSC administration. Furthermore, the UC-MSCs-CM reduced the levels of inflammatory cytokines and the activation of NLRP3 inflammasome in Raw264.7.ConclusionsOur results demonstrated that UC-MSCs exerted therapeutic effects on ALF by inhibiting apoptosis, inflammation, and pyroptosis.

Project description:BackgroundHypoxia preconditioning has been proven to be an effective method to enhance the therapeutic action of mesenchymal stem cells (MSCs). However, the beneficial effects of hypoxic MSCs in ischemia/reperfusion (I/R) lung injury have yet to be investigated. In this study, we hypothesized that the administration of hypoxic MSCs would have a positive therapeutic impact on I/R lung injury at molecular, cellular, and functional levels.MethodsI/R lung injury was induced in isolated and perfused rat lungs. Hypoxic MSCs were administered in perfusate at a low (2.5×105 cells) and high (1×106 cells) dose. Rats ventilated with a low tidal volume of 6 ml/kg served as controls. Hemodynamics, lung injury indices, inflammatory responses and activation of apoptotic pathways were determined.ResultsI/R induced permeability pulmonary edema with capillary leakage and increased levels of reactive oxygen species (ROS), pro-inflammatory cytokines, adhesion molecules, cytosolic cytochrome C, and activated MAPK, NF-κB, and apoptotic pathways. The administration of a low dose of hypoxic MSCs effectively attenuated I/R pathologic lung injury score by inhibiting inflammatory responses associated with the generation of ROS and anti-apoptosis effect, however this effect was not observed with a high dose of hypoxic MSCs. Mechanistically, a low dose of hypoxic MSCs down-regulated P38 MAPK and NF-κB signaling but upregulated glutathione, prostaglandin E2, IL-10, mitochondrial cytochrome C and Bcl-2. MSCs infused at a low dose migrated into interstitial and alveolar spaces and bronchial trees, while MSCs infused at a high dose aggregated in the microcirculation and induced pulmonary embolism.ConclusionsHypoxic MSCs can quickly migrate into extravascular lung tissue and adhere to other inflammatory or structure cells and attenuate I/R lung injury through anti-oxidant, anti-inflammatory and anti-apoptotic mechanisms. However, the dose of MSCs needs to be optimized to prevent pulmonary embolism and thrombosis.

Project description:INTRODUCTION: Studies have demonstrated that mesenchymal stromal cells (MSCs) could reverse acute and chronic kidney injury by a paracrine or endocrine mechanism, and microvesicles (MVs) have been regarded as a crucial means of intercellular communication. In the current study, we focused on the therapeutic effects of human Wharton-Jelly MSCs derived microvesicles (hWJMSC-MVs) in renal ischemia/reperfusion injury and its potential mechanisms. METHODS: MVs isolated from conditioned medium were injected intravenously in rats immediately after ischemia of the left kidney for 60 minutes. The animals were sacrificed at 24 hours, 48 hours and 2 weeks after reperfusion. The infiltration of inflammatory cells was identified by the immunostaining of CD68+ cells. ELISA was employed to determine the inflammatory factors in the kidney and serum von Willebrand Factor (VWF). Tubular cell proliferation and apoptosis were identified by immunostaining. Renal fibrosis was assessed by Masson's tri-chrome straining and alpha-smooth muscle actin (?-SMA) staining. The CX3CL1 expression in the kidney was measured by immunostaining and Western blot, respectively. In vitro, human umbilical vein endothelial cells were treated with or without MVs for 24 or 48 hours under hypoxia injury to test the CX3CL1 by immunostaining and Western blot. RESULTS: After administration of hWJMSC-MVs in acute kidney injury (AKI) rats, renal cell apoptosis was mitigated and proliferation was enhanced, inflammation was also alleviated in the first 48 hours. MVs also could suppress the expression of CX3CL1 and decrease the number of CD68+ macrophages in the kidney. In the late period, improvement of renal function and abrogation of renal fibrosis were observed. In vitro, MVs could down-regulate the expression of CX3CL1 in human umbilical vein endothelial cells under hypoxia injury at 24 or 48 hours. CONCLUSIONS: A single administration of MVs immediately after ischemic AKI could ameliorate renal injury in both the acute and chronic stage, and the anti-inflammatory property of MVs through suppression of CX3CL1 may be a potential mechanism. This establishes a substantial foundation for future research and treatment.

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Exosomes Derived From Mesenchymal Stem Cells Ameliorate Renal Ischemic-Reperfusion Injury Through Inhibiting Inflammation and Cell Apoptosis.

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Exosomes Derived From Mesenchymal Stem Cells Ameliorate Renal Ischemic-Reperfusion Injury Through Inhibiting Inflammation and Cell Apoptosis.

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