scholarly journals Mesenchymal stem cells–microvesicle-miR-451a ameliorate early diabetic kidney injury by negative regulation of P15 and P19

2018 ◽  
Vol 243 (15-16) ◽  
pp. 1233-1242 ◽  
Author(s):  
Ling Zhong ◽  
Guangneng Liao ◽  
Xiaojiao Wang ◽  
Lan Li ◽  
Jie Zhang ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Diabetic Nephropathy ◽  
Surface Markers ◽  
Cell Cycle Inhibitors ◽  
Injured Kidney ◽  
E Cadherin

Microvesicles (MVs) from mesenchymal stem cells (MSCs) have been reported as a new communicated way between cells. This study evaluated the influence and underlying mechanism of MVs-shuttled miR-451a on renal fibrosis and epithelial mesenchymal transformation (EMT) in diabetic nephropathy (DN) with hyperuricemia. MVs were isolated from MSCs-cultured medium by gradient ultracentrifugation. The level of miR-451a in MSCs and MVs was analyzed by qPCR. The changes of miR-451a, E-cadherin, α-SMA, P15INK4b (P15), and P19INK4d (P19) were measured in hyperglycosis and hyperuricemia-induced cell (HK-2) and mouse models. The changes of cell cycle were analyzed by flow cytometry. The ability of proliferation and viability was measured by BrdU and CCK8, respectively. Dual-luciferase reporter assays were conducted to determine the target binding sites. The renal function and histological changes of mice were analyzed. MVs showed the same surface markers as MSCs but much higher miR-451a expression (4.87 ± 2.03 fold higher than MSCs). miR-451a was decreased to 26% ± 11% and 6.7% ± 0.82% in injured HK-2 cells and kidney, respectively. MV-miR-451a enhanced the HK2 cells proliferation and viability in vitro, and decreased the morphologic and functional injury of kidney in vivo. Moreover, infusion of MV-miR-451a reduced the level of α-SMA and raised E-cadherin expression. These effects were responsible for the improved arrested cell cycle and down-regulation of P15 and P19 via miR-451a targeting their 3′-UTR sites. This study demonstrated that MSC–MV-miR-451a could inhibit cell cycle inhibitors P15 and P19 to restart the blocked cell cycle and reverse EMT in vivo and in vitro, and thus miR-451a is potentially a new target for DN therapy. Impact statement The mechanism of MSCs repairing the injured kidney in diabetic nephropathy is not yet clear. In the research, MVs showed the same surface markers as MSCs but much higher MiR-451a expression. miR-451a was decreased in both injured HK-2 cells and kidneys. MV-miR-451a stimulated the cell proliferation and viability in vitro and promoted structural and functional improvements of injured kidney in vivo. Infusion of MV-miR-451a ameliorated EMT by reducing α-SMA and increasing E-cadherin. These effects relied on the improved cell cycle arrest and the down-regulation of P15 and P19 via miR-451a binding to their 3′-UTR region. This study demonstrated that MSC–MV-miR-451a could specifically inhibit cell cycle inhibitors to restart the blocked cell cycle and reverse EMT in vivo and in vitro. Therefore, miR-451a may be a new target for DN therapy.

scholarly journals PTPRU, a quiescence-induced receptor tyrosine phosphatase negatively regulates osteogenic differentiation of human mesenchymal stem cells

2021 ◽  
Author(s):  
Mohammad Rumman ◽  
Jyotsna Dhawan
Keyword(s):  
Cell Cycle ◽  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Osteogenic Differentiation ◽  
Tyrosine Phosphatase ◽  
Quiescent State ◽  
Differentiation Potential

Bone marrow mesenchymal stem cells (MSCs) are heterogeneous osteo-progenitors that are mainly responsible for bone regeneration and homeostasis. In vivo, a subpopulation of bone marrow MSCs persists in a quiescent state, providing a source of new cells for repair. Previously, we reported that induction of quiescence in hMSCs in vitro skews their differentiation potential in favour of osteogenesis while suppressing adipogenesis. Here, we uncover a new role for a protein tyrosine phosphatase, receptor type U (PTPRU) in repressing osteogenesis during quiescence. A 75 kD PTPRU protein isoform was found to be specifically induced during quiescence and down-regulated during cell cycle reactivation. Using siRNA-mediated knockdown, we report that in proliferating hMSC, PTPRU preserves self-renewal, while in quiescent hMSC, PTPRU not only maintains reversibility of cell cycle arrest but also suppresses expression of osteogenic lineage genes. Knockdown of PTPRU in proliferating or quiescent hMSC de-represses osteogenic markers, and enhances induced osteogenic differentiation. We also show that PTPRU positively regulates a β-catenin-TCF transcriptional reporter. Taken together, our study suggests a role for a quiescence-induced 75kD PTPRU isoform in modulating bone differentiation in hMSC, potentially involving the Wnt pathway.

Coordinated Regulation of Hematopoietic and Mesenchymal Stem Cells in a Bone Marrow Niche.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 2-2 ◽  
Author(s):  
Simón Méndez-Ferrer ◽  
Tatyana V. Michurina ◽  
Francesca Ferraro ◽  
Amin Mazloom ◽  
Ben MacArthur ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Stem Cell ◽  
Osteoblastic Differentiation ◽  
Cell Depletion ◽  
Self Renewal

Abstract Abstract 2 Despite their therapeutic potential, mesenchymal stem cells (MSCs) remain poorly defined owing to their heterogeneity, the inability to assess in vivo self-renewal and the scarcity of markers allowing their identification, isolation and genetic manipulation. In the bone marrow (BM) of Nestin (Nes)-Gfp transgenic mice, CD31− CD45− GFP+ peri-vascular cells expressing endogenous nestin are associated with hematopoietic stem cells (HSCs) and innervated by fibers from the sympathetic nervous system (SNS). Flow cytometry sorting of BM CD45− Nes:GFP+ and CD45− Nes:GFP− cells has revealed that Nes:GFP+ cells, despite their rarity (4.0 ± 0.6% CD45− cells), contain all the colony-forming unit-fibroblastic (CFU-F) activity and have the exclusive capacity of forming self-renewing, multipotent clonal spheres that differentiate robustly along osteoblastic, chondrocytic and adipocytic lineages. To test in vivo self-renewal, single spheres derived from Nes-Gfp / Col2.3-Cre / R26R triple-transgenic animals were allowed to attach to phosphocalcic ceramic ossicles that were subcutaneously implanted into littermate mice that did not carry the transgenes. Histological analyses after 2 months revealed the presence of β-galactosidase+ osteoblasts (OBs) derived from Nes:GFP+ cells and not from 30,000 control CD45− Nes:GFP− cells. Hematopoietic areas were associated with Nes:GFP+ cells, that yielded per ossicle 310 ± 32 GFP+ secondary spheres (n=6), 38.6 ± 1.9% of which showed spontaneous multilineage differentiation into Col2.3+ OBs and Oil Red O+ adipocytes. Single secondary spheres subjected to a subsequent round of transplantation yielded after 8 months 8,557 ± 537 GFP+ spheres per ossicle (n = 7), which also generated Col2.3+ OBs, as a further proof of their self-renewal, osteoblastic differentiation potential and donor origin. Lineage-tracing studies in Nes-Cre / R26R mice have revealed the contribution of nestin-expressing cells in endochondral and membranous ossification. Administration of tamoxifen to adult Nes-CreERT2 mice bred to different reporter lines revealed that adult nestin-expressing BM cells could generate OBs, chondrocytes and osteocytes after 8-month chasing, suggesting an active role for adult nestin+ MSCs in physiological bone turnover. Genome-wide comparison analyses have shown that BM CD45− Nes:GFP+ cells are distinct from other stem cells but closest to in vitro expanded MSCs. Applying gene ontology analyses, metabolic and cell cycle genes were up- and down-regulated, respectively, in BM CD45− Nes:GFP+ cells. We have studied gene regulation, cell cycle and fate in response to granulocyte-colony stimulating factor (G-CSF), parathormone (PTH) and signals from the SNS, stimuli that regulate both hematopoietic and mesenchymal lineages in the BM. Cell cycle studies from FACS-sorted, flushed BM samples have confirmed that CD45− Nes:GFP+ cells are much more quiescent (90% G0/G1) than CD45− Nes:GFP− cells (58% G0/G1) but are selectively induced to proliferate after chemical sympathectomy (61% G0/G1) or PTH (70% G0/G1) administration in mice (n = 4–5). The inhibitory effects of the SNS and G-CSF (95% G0/G1) on BM CD45− Nes:GFP+ cells were not limited to cell cycle but also involved osteoblastic differentiation and expression of HSC maintenance genes. By contrast, in vivo or in vitro treatment with PTH selectively induced proliferation and osteoblastic differentiation of CD45− Nes:GFP+ cells, which express PTH receptor 1. We generated selective cell depletion models by intercrossing Nes-Cre and Nes-CreERT2 mice with a Cre-inducible diphtheria toxin receptor line (iDTR). In both models, HSC numbers decreased by ∼ 50% in the BM and increased in the spleen, an effect directly caused by selective BM cell depletion, as per in vitro experiments. In the more specific Nes-CreERT2 model, this effect was specific for HSCs and not for more mature progenitors. Cell depletion in Nes-Cre / iDTR and Nes-CreERT2 / iDTR mice reduced homing of hematopoietic progenitors by 73 and 90%, respectively. Finally, combined two-photon and confocal microscopy of the calvarial BM has demonstrated that highly purified, labeled HSCs rapidly (≤ 2h) home near Nes:GFP+ cells. Thus, cytokines, hormones, and the SNS regulate both HSC maintenance and bone formation in the BM stem cell niche through direct control of nestin-expressing MSCs. These results uncover an unprecedented partnership between two distinct somatic stem cell types and argue for a unique peri-vascular niche in the BM formed by MSC-HSC pairs. Disclosures: Scadden: Fate Therapeutics: Consultancy. Frenette:Glycomimetic: Research Funding.

scholarly journals Prospective Isolation of Murine and Human Bone Marrow Mesenchymal Stem Cells Based on Surface Markers

2013 ◽  
Vol 2013 ◽  
pp. 1-7 ◽  
Author(s):  
Yo Mabuchi ◽  
Diarmaid D. Houlihan ◽  
Chihiro Akazawa ◽  
Hideyuki Okano ◽  
Yumi Matsuzaki
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Therapeutic Potential ◽  
Research Field ◽  
Specific Reference ◽  
Human Mscs ◽  
Surface Markers ◽  
Prospective Isolation

Mesenchymal stem cells (MSCs) are currently defined as multipotent stromal cells that undergo sustainedin vitrogrowth and can give rise to cells of multiple mesenchymal lineages, such as adipocytes, chondrocytes, and osteoblasts. The regenerative and immunosuppressive properties of MSCs have led to numerous clinical trials exploring their utility for the treatment of a variety of diseases (e.g., acute graft-versus-host disease, Crohn’s disease, multiple sclerosis, osteoarthritis, and cardiovascular diseases including heart failure and myocardial infarction). On the other hand, conventionally cultured MSCs reflect heterogeneous populations that often contain contaminating cells due to the significant variability in isolation methods and the lack of specific MSC markers. This review article focuses on recent developments in the MSC research field, with a special emphasis on the identification of novel surface markers for thein vivolocalization and prospective isolation of murine and human MSCs. Furthermore, we discuss the physiological importance of MSC subtypesin vivowith specific reference to data supporting their contribution to HSC niche homeostasis. The isolation of MSCs using selective markers (combination of PDGFRαand Sca-1) is crucial to address the many unanswered questions pertaining to these cells and has the potential to enhance their therapeutic potential enormously.

scholarly journals Simultaneous impact of atorvastatin and mesenchymal stem cells for glioblastoma multiform suppression in rat glioblastoma multiform model

2020 ◽  
Vol 47 (10) ◽  
pp. 7783-7795
Author(s):  
Arash Goodarzi ◽  
Mehdi Khanmohammadi ◽  
Arman Ai ◽  
Hamid Khodayari ◽  
Armin Ai ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Cell Cycle Progression ◽  
C6 Cells ◽  
Glioblastoma Multiform ◽  
Apoptotic Activity ◽  
And Migration

Abstract Glioblastoma multiform (GBM) is known as an aggressive glial neoplasm. Recently incorporation of mesenchymal stem cells with anti-tumor drugs have been used due to lack of immunological responses and their easy accessibility. In this study, we have investigated the anti-proliferative and apoptotic activity of atorvastatin (Ator) in combination of mesenchymal stem cells (MSCs) on GBM cells in vitro and in vivo. The MSCs isolated from rats and characterized for their multi-potency features. The anti-proliferative and migration inhibition of Ator and MSCs were evaluated by MTT and scratch migration assays. The annexin/PI percentage and cell cycle arrest of treated C6 cells were evaluated until 72 h incubation. The animal model was established via injection of C6 cells in the brain of rats and subsequent injection of Ator each 3 days and single injection of MSCs until 12 days. The growth rate, migrational phenotype and cell cycle progression of C6 cells decreased and inhibited by the interplay of different factors in the presence of Ator and MSCs. The effect of Ator and MSCs on animal models displayed a significant reduction in tumor size and weight. Furthermore, histopathology evaluation proved low hypercellularity and mitosis index as well as mild invasive tumor cells for perivascular cuffing without pseudopalisading necrosis and small delicate vessels in Ator + MSCs condition. In summary, Ator and MSCs delivery to GBM model provides an effective strategy for targeted therapy of brain tumor.

Autologous transplantation of mesenchymal stem cells into the portal vein of the miniature pig; a preliminary experiment for NOTES approach

2019 ◽  
Vol 98 (9) ◽  
pp. 350-355
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Cell Therapy ◽  
Portal Vein ◽  
Liver Parenchyma ◽  
Miniature Pig ◽  
Hepatic Diseases ◽  
Study Results

Introduction: There is evidence that mesenchymal stem cells (MSCs) could trans-differentiate into the liver cells in vitro and in vivo and thus may be used as an unfailing source for stem cell therapy of liver disease. Combination of MSCs (with or without their differentiation in vitro) and minimally invasive procedures as laparoscopy or Natural Orifice Transluminal Endoscopic Surgery (NOTES) represents a chance for many patients waiting for liver transplantation in vain. Methods: Over 30 millions of autologous MSCs at passage 3 were transplanted via the portal vein in an eight months old miniature pig. The deposition of transplanted cells in liver parenchyma was evaluated histologically and the trans-differential potential of CM-DiI labeled cells was assessed by expression of pig albumin using immunofluorescence. Results: Three weeks after transplantation we detected the labeled cells (solitary, small clusters) in all 10 samples (2 samples from each lobe) but no diffuse distribution in the samples. The localization of CM-DiI+ cells was predominantly observed around the portal triads. We also detected the localization of albumin signal in CM-DiI labeled cells. Conclusion: The study results showed that the autologous MSCs (without additional hepatic differentiation in vitro) transplantation through the portal vein led to successful infiltration of intact miniature pig liver parenchyma with detectable in vivo trans-differentiation. NOTES as well as other newly developed surgical approaches in combination with cell therapy seem to be very promising for the treatment of hepatic diseases in near future.

Differentiation of mesenchymal stem cells from humans and animals into insulin-producing cells: An overview in vitro induction forms

2020 ◽  
Vol 16 ◽  
Author(s):  
Bruna O. S. Câmara ◽  
Bruno M. Bertassoli ◽  
Natália M. Ocarino ◽  
Rogéria Serakides
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Culture Medium ◽  
Adipogenic Differentiation ◽  
Medium Composition ◽  
Cell Therapies ◽  
Insulin Producing Cells ◽  
Msc Differentiation

The use of stem cells in cell therapies has shown promising results in the treatment of several diseases, including diabetes mellitus, in both humans and animals. Mesenchymal stem cells (MSCs) can be isolated from various locations, including bone marrow, adipose tissues, synovia, muscles, dental pulp, umbilical cords, and the placenta. In vitro, by manipulating the composition of the culture medium or transfection, MSCs can differentiate into several cell lineages, including insulin-producing cells (IPCs). Unlike osteogenic, chondrogenic, and adipogenic differentiation, for which the culture medium and time are similar between studies, studies involving the induction of MSC differentiation in IPCs differ greatly. This divergence is usually evident in relation to the differentiation technique used, the composition of the culture medium, the cultivation time, which can vary from a few hours to several months, and the number of steps to complete differentiation. However, although there is no “gold standard” differentiation medium composition, most prominent studies mention the use of nicotinamide, exedin-4, ß-mercaptoethanol, fibroblast growth factor b (FGFb), and glucose in the culture medium to promote the differentiation of MSCs into IPCs. Therefore, the purpose of this review is to investigate the stages of MSC differentiation into IPCs both in vivo and in vitro, as well as address differentiation techniques and molecular actions and mechanisms by which some substances, such as nicotinamide, exedin-4, ßmercaptoethanol, FGFb, and glucose, participate in the differentiation process.

scholarly journals Lithium-Doped Biological-Derived Hydroxyapatite Coatings Sustain In Vitro Differentiation of Human Primary Mesenchymal Stem Cells to Osteoblasts

Coatings ◽  
2019 ◽  
Vol 9 (12) ◽  
pp. 781 ◽  
Author(s):  
Paula E. Florian ◽  
Liviu Duta ◽  
Valentina Grumezescu ◽  
Gianina Popescu-Pelin ◽  
Andrei C. Popescu ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Doping Agent ◽  
Hydroxyapatite Coatings ◽  
3D Network ◽  
Immunofluorescence Labelling ◽  
Good Biocompatibility ◽  
Adhesion Process

This study is focused on the adhesion and differentiation of the human primary mesenchymal stem cells (hMSC) to osteoblasts lineage on biological-derived hydroxyapatite (BHA) and lithium-doped BHA (BHA:LiP) coatings synthesized by Pulsed Laser Deposition. An optimum adhesion of the cells on the surface of BHA:LiP coatings compared to control (uncoated Ti) was demonstrated using immunofluorescence labelling of actin and vinculin, two proteins involved in the initiation of the cell adhesion process. BHA:LiP coatings were also found to favor the differentiation of the hMSC towards an osteoblastic phenotype in the presence of osteoinductive medium, as revealed by the evaluation of osteoblast-specific markers, osteocalcin and alkaline phosphatase. Numerous nodules of mineralization secreted from osteoblast cells grown on the surface of BHA:LiP coatings and a 3D network-like organization of cells interconnected into the extracellular matrix were evidenced. These findings highlight the good biocompatibility of the BHA coatings and demonstrate that the use of lithium as a doping agent results in an enhanced osteointegration potential of the synthesized biomaterials, which might therefore represent viable candidates for future in vivo applications.

scholarly journals Comparative analysis of mouse bone marrow and adipose tissue mesenchymal stem cells for critical limb ischemia cell therapy

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Pegah Nammian ◽  
Seyedeh-Leili Asadi-Yousefabad ◽  
Sajad Daneshi ◽  
Mohammad Hasan Sheikhha ◽  
Seyed Mohammad Bagher Tabei ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Adipose Tissue ◽  
Cell Therapy ◽  
Femoral Artery ◽  
Critical Limb Ischemia ◽  
Limb Ischemia

Abstract Introduction Critical limb ischemia (CLI) is the most advanced form of peripheral arterial disease (PAD) characterized by ischemic rest pain and non-healing ulcers. Currently, the standard therapy for CLI is the surgical reconstruction and endovascular therapy or limb amputation for patients with no treatment options. Neovasculogenesis induced by mesenchymal stem cells (MSCs) therapy is a promising approach to improve CLI. Owing to their angiogenic and immunomodulatory potential, MSCs are perfect candidates for the treatment of CLI. The purpose of this study was to determine and compare the in vitro and in vivo effects of allogeneic bone marrow mesenchymal stem cells (BM-MSCs) and adipose tissue mesenchymal stem cells (AT-MSCs) on CLI treatment. Methods For the first step, BM-MSCs and AT-MSCs were isolated and characterized for the characteristic MSC phenotypes. Then, femoral artery ligation and total excision of the femoral artery were performed on C57BL/6 mice to create a CLI model. The cells were evaluated for their in vitro and in vivo biological characteristics for CLI cell therapy. In order to determine these characteristics, the following tests were performed: morphology, flow cytometry, differentiation to osteocyte and adipocyte, wound healing assay, and behavioral tests including Tarlov, Ischemia, Modified ischemia, Function and the grade of limb necrosis scores, donor cell survival assay, and histological analysis. Results Our cellular and functional tests indicated that during 28 days after cell transplantation, BM-MSCs had a great effect on endothelial cell migration, muscle restructure, functional improvements, and neovascularization in ischemic tissues compared with AT-MSCs and control groups. Conclusions Allogeneic BM-MSC transplantation resulted in a more effective recovery from critical limb ischemia compared to AT-MSCs transplantation. In fact, BM-MSC transplantation could be considered as a promising therapy for diseases with insufficient angiogenesis including hindlimb ischemia.

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