Apoptosis Of Mesenchymal Stem Cells Research Articles

The delivery of hsa-miR-11401 by extracellular vesicles can relieve doxorubicin-induced mesenchymal stem cell apoptosis

BackgroundChemotherapy is an effective anti-tumor treatment. Mesenchymal stem cells (MSCs), exerting therapy effect on injured tissues during chemotherapy, may be damaged in the process. The possibility of self-healing through long-range paracrine and the mechanisms are unclear.MethodsDoxorubicin, a commonly used chemotherapy drug, was to treat human umbilical cord-derived mesenchymal stem cells (hUC-MSCs) for 6 h as an in vitro cell model of chemotherapy-induced damage. Then we use extracellular vesicles derived from placental mesenchymal stem cells (hP-MSCs) to investigate the therapeutic potential of MSCs-EVs for chemotherapy injury. The mechanism was explored using microRNA sequencing.ResultsMSC-derived extracellular vesicles significantly alleviated the chemotherapy-induced apoptosis. Using microRNA sequencing, we identified hsa-miR-11401, which was downregulated in the Dox group but upregulated in the EV group. The upregulation of hsa-miR-11401 reduced the expression of SCOTIN, thereby inhibiting p53-dependent cell apoptosis.ConclusionsHsa-miR-11401 expressed by MSCs can be transported to chemotherapy-damaged cells by EVs, reducing the high expression of SCOTIN in damaged cells, thereby inhibiting SCOTIN-mediated apoptosis.

Probucol enhances the therapeutic efficiency of mesenchymal stem cells in the treatment of erectile dysfunction in diabetic rats by prolonging their survival time via Nrf2 pathway

BackgroundIntracavernous injection of mesenchymal stem cells (MSCs) is a promising method for diabetic mellitus-induced erectile dysfunction (DMED), but short survival time of MSCs in cavernous is a fatal defect for therapy. This study investigated therapeutic efficiency and potential mechanism of probucol combined with MSCs.MethodsIn vivo study, a total of forty-eight 10-week-old male Sprague-Dawley (SD) rats were used. Twelve rats received intraperitoneal injection of PBS as the sham group; the rest received intraperitoneal injection of 60 mg/kg streptozotocin to establish DM models. DM rats were randomly divided into three groups: received intracavernosal (IC) injection of either PBS (DM group), MSCs (M group), or administrated probucol after intracavernosal injection of MSCs (P + M group). Erectile function was assessed by electrical stimulation of the cavernous nerves with real-time intracavernous pressure measurement. After euthanasia, penile tissue was investigated for histologic examination and Western blotting. In in vitro experiment, H2O2 was used to create oxidative stress environment to detect changes in cell viability. CCK8 was used to measure cell viability of MSCs treated with or without probucol. Intracellular ROS changes were detected by flow cytometry. Autophagy and apoptosis were detected by Western blotting and confocal microscopy.ResultsRecovery of erectile function was observed in the P + M group. The combination therapy decreased fibrosis and increased endothelial function compared with MSC therapy alone. Western blotting results confirmed the increased expression of Nrf2 and HO-1 in cavernous body. H2O2 induced high oxidative stress and reduced cell viability in vitro, which was gradually reversed with increased concentration of probucol. H2O2 reduced Nrf2 expression, which was reversed by probucol’s intervention. Furthermore, the expression of Bax, Caspase3, and Cleaved-Caspase3 decreased, and the expression of Bcl-2 increased in a dose-dependent manner because of probucol’s intervention. In addition, Beclin1 and LC3II both increased in a dose-dependent manner. Meanwhile, the expression of P62 decreased. In the study of autophagy flux, we found probucol did not block it.ConclusionProbucol enhanced therapeutic efficiency of MSCs in DMED by prolonging their survival time, which mediated through improving the transplanted microenvironment of MSCs, increasing self-antioxidant ability of MSCs, strengthening protective autophagy, and inhibiting apoptosis of MSCs via Nrf2 pathway.Graphical abstractSchematic model showing combined probucol and MSCs to improve DMED. Probucol increases self-antioxidant ability of MSCs, strengthening protective autophagy and inhibiting apoptosis via Nrf2/HO-1 and Nrf2/autophagy pathways.

Combining Mesenchymal Stem Cell from Stromal Vascular Fraction with Scaffold (in silico, Biocompatibility, and Attachment Study)

AIM: The aim of this study is to evaluate the in silico, biocompatibility, and attachment of mesenchymal stem cell (MSC) from stromal vascular fraction (SVF) combined with scaffolds.
 METHODS: This research is true experimental study. In silico study using HEX version 8.0. Attachment of MSC from SVF to scaffolds was evaluated using electron microscope and measured. Biocompatibility was evaluated using viability and apoptosis of MSC from SVF after combined with MSC with scaffold.
 RESULTS: From in silico study, the strongest bound to scaffold is between MSC from SVF to hydroxyapatite (HA) in both receptor which was integrin alpha V to HA which need a total energy of −89.24 (J/Mol) and integrin beta 2 to HA which need a total energy of −177.8 (J/Mol). MSC from SVF cells is capable to combine with three types of bone substitution material. At HA-calcium sulfate administration, SVF cells had apoptosis of 30.27%, and viable cells were 69.73%. At the administration of bovine bone cancellous, SVF cells had apoptosis of 22.20% and viable cells of 77.80%. From microscope electron study, the best result of attachment was obtained from MSC to HA-calcium phosphate with an average value of 12.66 cluster cells counting per 100 μm2 scaffold material.
 CONCLUSION: From in silico study showed that MSC from SVF could attach to scaffold with stronger binding between integrin alpha V and integrin beta 2 to HA scaffold. Bovine bone cancellous has the best biocompatibility and attachment among other scaffold with the highest viability and lowest cell apoptosis. From microscope electron study, we can prove that MSC can make a cluster cell in those scaffolds with HA-calcium sulfate having the biggest cluster cell counting.

A novel role of vitronectin in promoting survival of mesenchymal stem cells under serum deprivation stress

BackgroundDue to their immunomodulatory and trophic support functions, mesenchymal stem cells (MSCs) are promising in the field of cell-based regenerative medicine. However, MSC survival post-transplantation is challenged by various microenvironment stress factors. Here, we investigated the role of vitronectin (VTN) in the survival strategy of MSCs under serum deprivation stress condition.MethodsProliferation kinetics and cell adhesion of MSCs under serum deprivation were determined from population doublings and cell-matrix de-adhesion studies, respectively. mRNA and protein expression levels of VTN were confirmed by qRT-PCR and Western blotting, respectively. Immunofluorescence technique revealed distribution of VTN under serum deprivation stress. siRNA and inhibitor-based studies were performed to confirm the role and regulation of VTN. Apoptosis and cell cycle status of MSCs were assessed using flow cytometric analysis.ResultsSubjecting MSCs to serum deprivation led to significant increase in cell spread area and cell-matrix adhesion. An upregulation of VTN expression was noted with an arrest in G0/G1 phase of cell cycle and no appreciable apoptotic change. Pro-survival PI3kinase pathway inhibition led to further increase in VTN expression with no apoptotic change. siRNA-mediated inhibition of VTN resulted in reversal in G0/G1 cell cycle arrest and a marked increase in apoptosis, suggesting a role of VTN in preventing serum deprivation-induced apoptotic cell death. In addition, p65 knockdown resulted in downregulation of VTN establishing an association between NF-κβ pathway and VTN.ConclusionsVTN was identified as a survival factor in providing protection from serum deprivation-induced apoptosis in MSCs.

Long non-coding RNA MEG3 is involved in osteogenic differentiation and bone diseases (Review).

Osteogenic differentiation originating from mesenchymal stem cells (MSCs) requires tight co-ordination of transcriptional factors, signaling pathways and biomechanical cues. Dysregulation of such reciprocal networks may influence the proliferation and apoptosis of MSCs and osteoblasts, thereby impairing bone metabolism and homeostasis. An increasing number of studies have shown that long non-coding (lnc)RNAs are involved in osteogenic differentiation and thus serve an important role in the initiation, development, and progression of bone diseases such as tumors, osteoarthritis and osteoporosis. It has been reported that the lncRNA, maternally expressed gene 3 (MEG3), regulates osteogenic differentiation of multiple MSCs and also acts as a critical mediator in the development of bone formation and associated diseases. In the present review, the proposed mechanisms underlying the roles of MEG3 in osteogenic differentiation and its potential effects on bone diseases are discussed. These discussions may help elucidate the roles of MEG3 in osteogenic differentiation and highlight potential biomarkers and therapeutic targets for the treatment of bone diseases.

MiRNA-98-5p Targeting IGF2BP1 Induces Mesenchymal Stem Cell Apoptosis by Modulating PI3K/Akt and p53 in Immune Thrombocytopenia

Immune thrombocytopenia (ITP) is a common hematological autoimmune disease, in which defective mesenchymal stem cells (MSCs) are potentially involved. Our previous study suggested that MSCs in ITP patients displayed enhanced apoptosis. MicroRNAs (miRNAs) play important roles in ITP by affecting megakaryopoiesis, platelet production and immunoregulation, whereas the roles of miRNAs in ITP-MSCs remain unknown. In a previous study, we performed microarray analysis to obtain mRNA and miRNA profiles of ITP-MSCs. In the present study, we reanalyze the data and identify miR-98-5p as a candidate miRNA contributing to MSC deficiency in ITP. miR-98-5p acts through targeting insulin-like growth factor 2 mRNA-binding protein 1 (IGF2BP1), and the subsequent downregulation of insulin-like growth factor 2 (IGF-2) causes inhibition of the phosphatidylinositol 3-kinase (PI3K)/Akt pathway, which is involved in the process of MSC deficiency. Furthermore, miR-98-5p upregulates p53 by inhibiting β-transducin repeat-containing protein (β-TrCP)-dependent p53 ubiquitination. Moreover, miR-98-5p overexpression impairs the therapeutic effect of MSCs in ITP mice. All-trans retinoic acid (ATRA) protects MSCs from apoptosis by downregulating miR-98-5p, thus providing a potential therapeutic approach for ITP. Our findings demonstrate that miR-98-5p is a critical regulator of ITP-MSCs, which will help us thoroughly understand the pathogenesis of ITP.

The Therapeutic Potential of Apoptotic Mesenchymal Stem Cells in Muscle Regeneration

Macrophages are critical for muscle regeneration and have an unusually high level of responsiveness to their environment. Because of this sensitivity, their beneficial effects may be realized only upon certain activation conditions. Previous studies in our lab have shown that transplantation of classically activated macrophages and anti‐inflammatory macrophages sorted from injured muscle enhanced functional regeneration in ischemia‐reperfusion muscles. The phagocytosis of apoptotic cells that are present in injured muscle or from transplantation serves as a well‐known activation signal for macrophages to initiate a regenerative and anti‐inflammatory profile leading to enhanced muscle regeneration. The aim of this study is to investigate if the phagocytosis of apoptotic mesenchymal stem cells (MSCs) can polarize bone marrow‐derived macrophages (BMDMs) towards a regenerative and anti‐inflammatory phenotype and subsequently if apoptotic MSCs in different stages of apoptosis have differential effects. We induced apoptosis in MSCs and incubated them at different time intervals to obtain differential stages of apoptosis, which were characterized by the amount of externalized phosphatidylserine (PS) on their cell membrane. Apoptotic MSCs with different concentrations of PS were then co‐cultured with BMDMs for 48 hours. Cell surface markers and gene expression of characteristic of pro‐ and anti‐inflammatory polarized BMDMs were measured. Our results showed that apoptotic MSCs without incubation significantly increased the expression of F4/80 and CD206 on the cell surface of BMDMs. They also increased the gene expression of MRC‐1 and IL‐10 as well as decreased the expression of iNOS and IL‐1β in co‐cultured BMDMs. The extent of these effects exhibited a decreasing trend when incubation time of apoptotic MSCs increased. In conclusion, the extent of MSC apoptosis as characterized by PS expression resulted in differential anti‐inflammatory and regenerative effects on BMDMs, wherein the apoptotic MSCs in the earlier stage of apoptosis result in the greatest increase in anti‐inflammatory phenotype, which reveals their therapeutic potential in muscle regeneration.Support or Funding InformationThis project is funded by National Institutes of Health Grant 2R01EB015007‐05A1 (to Laura J. Suggs and Roger P. Farrar)

RXRα and MRTF-A have a synergistic effect in the retinoic acid-induced neural-like differentiation of adult bone marrow-derived mesenchymal stem cells.

Mesenchymal stem cells (MSCs) have multilineage differentiation potential and can transform into neuron cells under an appropriate environment. Retinoic acid (RA) facilitates the neuronal differentiation of MSCs. We found that RXRα, a RA receptor, was significantly upregulated in RA-induced process. Here, we show that RXRα collaborated with myocardin-related transcription factor-A (MRTF-A) to strongly promote the RA-induced process as evidenced by the increase in NF-H expression and NF-H promoter transcription activity. Our studies reveal that RXRα and MRTF-A exhibit protein interactions and synergistically inhibit the MSCs apoptosis by enhancing the P21 expression. Furthermore, RXRα and MRTF-A can activate P21 transcription by affecting the formation of the MRTF-A/RXRα/RARE complex. These findings reveal the important roles of RXRα and MRTF-A signaling in RA-induced neural-like differentiation of MSCs and describe a new mechanism underlying the synergistic interaction of RXRα and MRTF-A.

Fine-tuning of Rat Mesenchymal Stem Cell Senescence via Microtopography of Polymeric Substrates

ABSTRACTCellular senescence, a driver of aging and age-related diseases, is a stable state found in metabolically active cells characterized by irreversible cell growth arrest and dramatic changes in metabolism, gene expression and secretome profile. Endogenous regeneration efficacy of mesenchymal stem cells (MSCs) could be attenuated due to senescence. MSCs can be modulated by not only biochemical signals but also by physical cues such as substrate topography. To provide a cell culture substrate that can prevent MSC senescence over an extended period of in vitro cultivation, here, the cell- and immunocompatible poly(ether imide) (PEI) substrate was used. Two distinct levels of roughness were created on the bottom surfaces of PEI inserts via injection molding: Low-R (similar to the thickness of attached single MSC, Rq: 3.9 ± 0.2 µm) and High-R (larger than single MSC thickness. Rq: 22.7 ± 0.8 µm). Cell expansion, lysosomal enzymatic activity, apoptosis and paracrine effects of senescent MSCs were examined by cell counting, detection of senescence-associated β-galactosidase (SA β-gal), Caspase 3/7, and CFSE labeling. MSCs showed high cell viability and similar spindle-shaped morphology on all investigated surfaces. Cells on Low-R presented the highest expansion (80000 ± 1805 cells), as compared to cells on smooth PEI and High-R. The low apoptosis level (0.08 vs 0.12 from smooth PEI) and senescence ratio (35% vs. 54% from smooth PEI) were observed in MSCs cultured on Low-R. The secretome from Low-R effectively prevents senescence and supports the proliferation of neighboring cells (1.5-fold faster) as compared to the smooth PEI secretome. In summary, the Low-R PEI provided a superior surface environment for MSCs, which promoted proliferation, inhibited apoptosis and senescence, and effectively influenced the proliferation of neighboring cells via their paracrine effect. Such microroughness can be considered as a key parameter for improving the therapeutic potential of endogenous regeneration, anti-organismal aging and anti-age-related pathologies via directly promoting cell growth and modulating paracrine effects of the senescence associated secretome.

Ginsenoside Rg1 as an Effective Regulator of Mesenchymal Stem Cells.

Recently, breakthroughs have been made in the use of mesenchymal stem cells (MSCs) to treat various diseases. Several stem cell types have been authorized as drugs by the European Medicines Agency and the U.S. Food and Drug Administration. The Chinese official document “Notification of the management of stem cell clinical research (trial)” was also published in August 2015. Currently, China has approved 106 official stem cell clinical research filing agencies and 62 clinical research projects, which are mostly focused on MSC therapy. Hence, the optimization and development of stem cell drugs is imperative. During this process, maximizing MSC expansion, minimizing cell loss during MSC transplantation, improving the homing rate, precisely regulating the differentiation of MSCs, and reducing MSC senescence and apoptosis are major issues in MSC preclinical research. Similar to artemisinin extracted from the stems and leaves of Artemisia annua, ginsenoside Rg1 (Rg1) is purified from the root or stem of ginseng. In the human body, Rg1 regulates organ function, which is inseparable from its regulation of adult stem cells. Rg1 treatment may effectively regulate the proliferation, differentiation, senescence, and apoptosis of MSCs in different microenvironments in vitro or in vivo. In this review, we discuss recent advances in understanding the effect of Rg1 on MSCs and describe the issues that must be addressed and prospects regarding Rg1 regulation of MSCs in preclinical or clinical studies.

Dexamethasone promotes mesenchymal stem cell apoptosis and inhibits osteogenesis by disrupting mitochondrial dynamics.

Long‐term or heavy use of glucocorticoids can cause severe necrosis of the femoral head, but the underlying mechanisms are still unclear. Recent studies have found that mitochondrial dynamics play an important role in femoral head necrosis. Here, we investigated the effect of dexamethasone on the mitochondrial function of mesenchymal stem cells. We observed that high concentrations of dexamethasone (10−6 mol·L−1) decreased cell activity, promoted apoptosis, elevated levels of reactive oxygen species and disrupted mitochondrial dynamics. Furthermore, dexamethasone (10−6 mol·L−1) inhibited osteogenesis of stem cells and promoted adipogenesis. These findings may facilitate greater understanding of the adverse effects of dexamethasone on the femoral head.

Effects of VEGF-Notch Signaling Pathway on Proliferation and Apoptosis of Bone Marrow MSC in Patients with Aplastic Anemia

To evaluate the regulation of VEGF-Notch signaling pathway on proliferation and apoptosis of mesenchymal stem cells (MSC) in the patients with aplastic anemia (AA). The bone marrow specimens of AA patients were collected for isolation and identification of BM MSC. Westen blot was used to detect the expression of VEGF-Notch signaling pathway-related proteins (VEGF, VEGFR, Notch 1, Jagged 1, Delta-like 1 and Hes1). The VEGF (100 ng/ml) and DAPT (γ-secretase inhibitor, 10 μmol/L) were respectively added into MSC culture system in oder to activate and inhibit the signaling transduction of VEGF-Notch in BM MSC. The proliferation, apoptosis and cell cycle of MSC in AA patients were detected by CCK8 assay and flow cyfometry. The adipogenic differentiation of BM MSC was detected by oil red O staining. The VEGF-Notch signaling pathway was significantly inhibited in AA BM tissues and AA MSC (P＜0.05) detected by Western blot. The intervention of VEGF and DAPT significantly activated and inhibited VEGF-Notch signaling in AA MSC, respectively. CCK8 assay showed that VEGF intervention significantly promoted the proliferation of MSC in AA patients (P＜0.05). Flow cytometry showed that VEGF significantly inhibited apoptosis of MSCs by blocking S phase cells (P＜0.05). The activation of VEGF-Notch can restore the proliferation function of MSC in AA patients.

Effects of VEGF-Notch Signaling Pathway on Proliferation and Apoptosis of Bone Marrow MSC in Patients with Aplastic Anemia

To evaluate the regulation of VEGF-Notch signaling pathway on proliferation and apoptosis of mesenchymal stem cells (MSC) in the patients with aplastic anemia (AA). The bone marrow specimens of AA patients were collected for isolation and identification of BM MSC. Westen blot was used to detect the expression of VEGF-Notch signaling pathway-related proteins (VEGF, VEGFR, Notch 1, Jagged 1, Delta-like 1 and Hes1). The VEGF (100 ng/ml) and DAPT (γ-secretase inhibitor, 10 μmol/L) were respectively added into MSC culture system in oder to activate and inhibit the signaling transduction of VEGF-Notch in BM MSC. The proliferation, apoptosis and cell cycle of MSC in AA patients were detected by CCK8 assay and flow cyfometry. The adipogenic differentiation of BM MSC was detected by oil red O staining. The VEGF-Notch signaling pathway was significantly inhibited in AA BM tissues and AA MSC (P＜0.05) detected by Western blot. The intervention of VEGF and DAPT significantly activated and inhibited VEGF-Notch signaling in AA MSC, respectively. CCK8 assay showed that VEGF intervention significantly promoted the proliferation of MSC in AA patients (P＜0.05). Flow cytometry showed that VEGF significantly inhibited apoptosis of MSCs by blocking S phase cells (P＜0.05). The activation of VEGF-Notch can restore the proliferation function of MSC in AA patients.

Full title: High glucose protects mesenchymal stem cells from metformin-induced apoptosis through the AMPK-mediated mTOR pathway

Micro- and macro-vascular events are directly associated with hyperglycemia in patients with type 2 diabetes mellitus (T2DM), but whether intensive glucose control decreases the risk of diabetic cardiovascular complications remains uncertain. Many studies have confirmed that impaired quality and quantity of mesenchymal stem cells (MSCs) plays a pathogenic role in diabetes. Our previous study found that the abundance of circulating MSCs was significantly decreased in patients with T2DM, which was correlated with the progression of diabetic complications. In addition, metformin-induced MSC apoptosis is one of the reasons for the decreased quantity of endogenous or exogenous MSCs during intensive glucose control. However, the role of glucose in metformin-induced MSC apoptosis during intensive glucose control in T2DM remains unknown. In this study, we found that metformin induces MSC apoptosis during intensive glucose control, while high glucose (standard glucose control) could significantly reverse its adverse effect in an AMPK-mTOR pathway dependent manner. Thus, our results indicate that the poorer clinical benefit of the intensive glucose control strategy may be related to an adverse effect due to metformin-induced MSC apoptosis during intensive glucose control therapy in patients with T2DM.

In a Rat Model of Acute Liver Failure, Icaritin Improved the Therapeutic Effect of Mesenchymal Stem Cells by Activation of the Hepatocyte Growth Factor/c-Met Pathway

Acute liver failure (ALF) is a serious life-threatening condition. Mesenchymal stem cells (MSCs) may be an effective treatment for this condition and a good alternative to liver transplantation. Icaritin (ICT) is an active ingredient of the genus Epimedium, a traditional Chinese medicine, with the potential to enhance the proliferation of MSCs. The purpose of this study was to explore whether ICT increased the therapeutic effects of MSCs and explore its underlying mechanisms. For in vivo experiments, a rat ALF model was established by intraperitoneal injection of D(+)-galactosamine/ lipopolysaccharide. MSCs cocultured with ICT were used to treat ALF rats and the protective effects assessed as survival rate, levels of serum AST and ALT, and histological changes in liver tissue. For in vitro experiments, MSCs were treated in serum-free culture for 72 h to simulate the disruption of intrahepatic microcirculation. MSCs apoptosis was examined to determine whether ICT rescued impaired MSCs. The role of the hepatocyte growth factor (HGF)/c-Met pathway in MSCs was assessed by constructing genetically modified MSCs overexpressing c-Met and by using the c-Met receptor inhibitor (crizotinib). The results showed that MSCs increased the survival rate of ALF rats and reduced liver damage. MSCs cocultured with ICT exerted a greater therapeutic effect than MSCs alone. Further, the HGF/c-Met pathway played a key role in the antiapoptotic activity of MSCs, which was associated with the optimized efficacy of ICT. In conclusion, this study demonstrated that ICT enhances the therapeutic effect of MSCs in a model of ALF, improving the antiapoptotic potential of MSCs by upregulation of the HGF/c-Met pathway. The combination of stem cell therapy with traditional herbal extracts may improve MSC-based clinical applications.

AT1R-Mediated Apoptosis of Bone Marrow Mesenchymal Stem Cells Is Associated with mtROS Production and mtDNA Reduction.

Angiotensin II (Ang II) is used as an inducer for the differentiation of mesenchymal stem cells (MSCs). Whether the commonly used doses of Ang II for MSC differentiation affect cell apoptosis has not been elucidated. In this study, we investigated the effect of Ang II on the apoptosis of bone marrow MSCs (BMMSCs), and its relations to the activation of Ang II receptor-1- (AT1R-) signaling, mitochondrial ROS (mtROS) generation, and mitochondrial DNA (mtDNA) leakage. AT1R expression in BMMSCs was identified by immunostaining and Western-blotting assays. BMMSC viability was measured by MTT assay following exposure to 1 nM~1 mM Ang II for 12 hours. Cell apoptosis, mtROS, and mtDNA levels were detected by FAM-FLICA® Poly Caspase, MitoSOX™ superoxide, and PicoGreen staining, respectively. The expressions of Bcl2 and Bax were measured by Western-blotting assays. Next, we used losartan to block AT1R-signaling and subsequently measured apoptosis, mtROS, and mtDNA levels, again. The maximum viability of BMMSCs was in response to 100 nM Ang II, after that it began to decrease with the increase of Ang II doses, indicating that Ang II (≧1 μM) may cause apoptosis of BMMSCs. As expected, 1 μM and 10 μM Ang II both caused BMMSC apoptosis. Furthermore, 1 μM and 10 μM Ang II could also induce mtROS generation and cause a marked mtDNA leakage. The application of losartan markedly inhibited Ang II-induced mtROS production, mtDNA leakage, and BMMSC apoptosis. In conclusion, the activation of AT1R-signaling stimulates apoptosis of BMMSCs, which is associated mtROS production and mtDNA reduction.

Spreading Shape and Area Regulate the Osteogenesis of Mesenchymal Stem Cells.

Mesenchymal stem cells (MSCs) have strong self-renewal ability and multiple differentiation potential. Some studies confirmed that spreading shape and area of single MSCs influence cell differentiation, but few studies focused on the effect of the circularity of cell shape on the osteogenic differentiation of MSCs with a confined area during osteogenic process. In the present study, MSCs were seeded on a micropatterned island with a spreading area lower than that of a freely spreading area. The patterns had circularities of 1.0 or 0.4, respectively, and areas of 314, 628, or 1256μm2. After the cells were grown on a micropatterned surface for 1 or 3days, cell apoptosis and F-actin were stained and analyzed. In addition, the expression of β-catenin and three osteogenic differentiation markers were immunofluorescently stained and analyzed, respectively. Of these MSCs, the ones with star-like shapes and large areas promoted the expression of osteogenic differentiation markers and the survival of cells. The expression of F-actin and its cytosolic distribution or orientation also correlated with the spreading shape and area. When actin polymerization was inhibited by cytochalasin D, the shape-regulated differentiation and apoptosis of MSCs with the confined spreading area were abolished. This study demonstrated that a spreading shape of low circularity and a larger spreading area are beneficial to the survival and osteogenic differentiation of individual MSCs, which may be regulated through the cytosolic expression and distribution of F-actin.

Canagliflozin Impairs Blood Reperfusion of the Ischemic Lower Limb by Inhibiting the Retention and Paracrine Function of Mesenchymal Stem Cells

Background: Canagliflozin (CANA) administration increases amputation risk of the lower limb in the clinic. The present study aimed to investigate whether and how CANA interferer with intracellular physiological processes of mesenchymal stem cells (MSCs), and its contribution to ischemic lower limb. Methods: The in vivo blood flow recovery of ischemic lower limb following CANA treatment was evaluated. The cellular function of MSCs after CANA treatment were also assessed in vitro. In silico docking analysis and mutant substitution assay were conducted to confirm the interaction of CANA and glutamate dehydrogenase 1 (GDH1). Findings: Following CANA treatment, attenuated angiogenesis and hampered recovery of blood flow were observed in ischemic region, and the proliferation inhibition, migration inhibition, and apoptosis of MSCs was evidenced. CANA was involved in MSC mitochondrial respiratory malfunction, ATP production inhibition, cytochrome c release and vessel endothelial growth factor A (VEGFA) secretion inhibition, which contribute to reduced tissue repair capacity of MSCs, resulting in compromised angiogenesis and blood reperfusion in ischemic region. The detrimental effects of CANA on MSCs result from the inhibition of GDH1 by CANA (evidenced by in silico docking analysis and H199A-GDH1/ N392A-GDH1 mutant substitution). Interpretation: Our work highlights that GDH1 activity inhibition by CANA interferes with metabolic activity of mitochondria, which deteriorates retention and VEGFA secretion of MSCs, hampering angiogenesis and the recovery of blood flow following lower limb ischemia. Funding Statement: This study was financially supported by National Natural Science Foundation of China (grant # 81800314 for J. N.), Natural Science Foundation of Zhejiang Province (grant # LQ18H090007 for Z. L.) and Wenzhou Science and Technology Bureau Foundation (grant # Y20190163 for Y. L.). Declaration of Interests: The authors declare no competing financial interests. Ethics Approval Statement: All experimental protocols were approved by the Animal Care and Use Committee of Wenzhou Medical University.

O-cyclic phytosphingosine-1-phosphate stimulates HIF1\u03b1-dependent glycolytic reprogramming to enhance the therapeutic potential of mesenchymal stem cells

O-cyclic phytosphingosine-1-phosphate (cP1P) is a novel chemically synthesized sphingosine metabolite derived from phytosphingosine-1-phosphate. Although structurally similar to sphingosine-1-phosphate (S1P), its biological properties in stem cells remain to be reported. We investigated the effect of cP1P on the therapeutic potential of mesenchymal stem cells (MSCs) and their regulatory mechanism. We found that, under hypoxia, cP1P suppressed MSC mitochondrial dysfunction and apoptosis. Metabolic data revealed that cP1P stimulated glycolysis via the upregulation of glycolysis-related genes. cP1P-induced hypoxia-inducible factor 1 alpha (HIF1α) plays a key role for MSC glycolytic reprogramming and transplantation efficacy. The intracellular calcium-dependent PKCα/mammalian target of the rapamycin (mTOR) signaling pathway triggered by cP1P regulated HIF1α translation via S6K1, which is critical for HIF1 activation. Furthermore, the cP1P-activated mTOR pathway induced bicaudal D homolog 1 expression, leading to HIF1α nuclear translocation. In conclusion, cP1P enhances the therapeutic potential of MSC through mTOR-dependent HIF1α translation and nuclear translocation.

Photobiomodulation therapy compensate the impairments of diabetic bone marrow mesenchymal stem cells.

Pathophysiologic conditions associated with diabetes mellitus affect mesenchymal stem cells (MSCs), and this phenomenon may lead to some diabetic secondary complications. The present study was conducted to evaluate the impact of photobiomodulation (PBM) on rat diabetic MSC (DMSC) behavior in vitro. For the purpose of PBM, we used helium-neon laser with a wavelength of 632.8nm at three different energy densities (0.5, 1, 2J/cm2) and radiation periodicity of once, twice, and thrice. The survival, proliferation, and apoptosis in the normal MSCs (NMSCs), DMSCs, and diabetic MSCs, which were laser irradiated (DMSCs+L), were assessed using MTT assay, Ki67 immunofluorescence staining, and TUNEL assay, respectively. Our results demonstrated that DMSCs have significantly lower survival (P < 0.05) and proliferation rates (P < 0.001), and dramatically higher population doubling time (PDT, P < 0.001) and apoptosis rates (P < 0.001) as compared to NMSCs. Moreover, PBM with energy density of 1J/cm2 and the periodicity of 1 or 2 times could improve diabetic MSC capabilities in the term of survival, proliferation, and apoptosis. Considering these findings, it is suggested that PBM could improve the ability of diabetic MSCs in vitro prior to transplantation or may rise their capabilities in their native niche in vivo.