Apoptosis Of Mesenchymal Stem Cells Research Articles

Blocking Nox2 improves mesenchymal stem cells therapy in myocardial infarction via antagonizing oxidant and promoting survival.

An increase in reactive oxygen species (ROS) plays a key role in aging and apoptosis in mesenchymal stem cells derived from bone marrow (BMSCs). NADPH oxidase Nox2 serves as an important source of intracellular ROS formation. This study is designed to determine if blocking Nox2 enhances anti-apoptotic and anti-aging ability of BMSCs to oxidant stress, and thus improves therapeutic efficacy in myocardial infarction (MI). Nox2 inhibitor (Acetovanillone) and Nox2 siRNA were used to block Nox2 in BMSCs, and the cell viability, apoptosis, senescence and survival of BMSCs were determined by CCK-8, Edu staining, TUNEL staining, β-galactosidase (β-gal) assay and DAPI labeling. Here we found that both Nox2 inhibitor and Nox2 knockdown remarkably countered the decrease of viability, and the increase of aging and apoptosis of BMSCs by H2 O2 . Whereas, Nox2 overexpression exacerbated the viability reduction, senescence and apoptosis of BMSCs. The ROS accumulation in BMSCs was also suppressed by Nox2 blocking. Further study uncovered that Nox2 inhibitor caused the downregulation of p-p53, p21, p-FoxO1 and Bax, and the upregulation of anti-apoptotic protein Bcl-2. In vivo, Nox2 knockdown in grafted BMSCs led to the improvement of EF and FS in infarcted myocardium than BMSCs without Nox2 knockdown. Consistently, more retention and survival of BMSCs were found after Nox2 knockdown. Taken together, Nox2 inhibition enhances anti-aging and anti-apoptotic ability of BMSCs, and thus promotes survival and retention of BMSCs, which provides a new strategy for improving BMSCs-based therapy.

In Vitro Survival of Human Mesenchymal Stem Cells is Enhanced in Artificial Endolymph with Moderately High Concentrations of Potassium.

While mesenchymal stem cells (MSCs) are promising candidates for inner ear hair cell regeneration, to date, there have been no convincing reports indicating whether MSCs can survive in the cochlea for more than a few weeks, as the high levels of potassium (K+) in the endolymph (EL) are thought to be toxic to transplanted stem cells. For conditioning the EL for MSC transplantation, we conducted this in vitro study to examine the effects of artificial EL with altered K+ concentration levels, in the range of 5-153.8 mM, on proliferation, apoptosis, and morphological changes in MSCs derived from various human tissues. Our findings demonstrate that altering the K+ concentration in artificial EL could significantly influence the survival of MSCs in vitro. We discovered that K+ concentrations of 55-130 mM in artificial EL could enhance the survival of MSCs in vitro. However, MSCs exhibited reduced proliferation regardless of K+ concentration.

Effects of HSYA on the proliferation and apoptosis of MSCs exposed to hypoxic and serum deprivation conditions.

As a primary active ingredient of safflor yellow, hydroxysafflor yellow A (HSYA) exhibits notable antioxidative and neuroprotective effects. The aim of the present study was to investigate the protective effects of HSYA in mesenchymal stem cells (MSCs) exposed to hypoxia (5% O2) and serum deprivation (H/SD), and to explore the mechanisms underlying HSYA-mediated protection. Under H/SD conditions, HSYA was applied to protect MSCs against injury. Cell viability, proliferation, apoptosis and reactive oxygen species (ROS) levels were determined using an 5-ethynyl-2′-deoxyuridine assay, MTT assay, Hoechst 33342/propidium iodide and 2′,7′-dichlorodihydrofluorescein diacetate staining, respectively. The results revealed that 160 mg/l HSYA significantly reduced apoptosis and ROS levels compared with the H/SD group; however, HSYA demonstrated minimal effects on cell proliferation. A western blot assay demonstrated that HSYA reduced cleaved caspase-3 expression and cytC release from the mitochondria to the cytoplasm when compared with the H/SD group. In addition, western blotting and RT-qPCR analyses revealed that HSYA treatment significantly increased the expression of hypoxia inducible factor-1α (HIF-1α) and vascular endothelial growth factor (VEGF). In conclusion, the results of the current study demonstrated that HSYA exerts protective effects against H/SD-induced apoptosis in MSCs potentially via activation of the HIF-1α/VEGF signaling pathway and stabilization of the mitochondrial membrane.

Oleuropein attenuates hydrogen peroxide-induced autophagic cell death in human adipose-derived stem cells

Mesenchymal stem cells (MSCs) are multipotent progenitor cells with self-renewing properties; thus, transplanting functionally enhanced MSCs might be a promising strategy for cell therapy against ischemic diseases. However, extensive oxidative damage in ischemic tissue affects the cell fate of transplanted MSCs, eventually resulting in cell damage and autophagic cell death. Oleuropein (OLP) is a bioactive compound isolated from olives and olive oil that harbors antioxidant properties. This study aimed to investigate the potential cytoprotective effects of OLP against oxidative stress and autophagic cell death in MSCs. We found that short-term priming with OLP attenuated H2O2-induced apoptosis by regulating the pro-apoptotic marker Bax and the anti-apoptotic markers Bcl-2 and Mcl-1. Notably, OLP inhibits H2O2 -induced autophagic cell death by modulating autophagy-related death signals, including mTOR (mammalian target of rapamycin), ULK1 (unc-51 like autophagy activating kinase 1), Beclin-1, AMPK (AMP-activated protein kinase), and LC3 (microtubule-associated protein 1a/1b-light chain 3). Our data suggest that OLP might reduce H2O2-induced autophagy and cell apoptosis in MSCs by regulating both the AMPK-ULK axis and the Bcl-2-Mcl-1 axis. Consequently, short-term cell priming with OLP might enhance the therapeutic effect of MSCs against ischemic vascular diseases, which provides an important potential improvement for emerging therapeutic strategies.

Novel Role of ER Stress and Mitochondria Stress in Serum-deprivation Induced Apoptosis of Rat Mesenchymal Stem Cells.

The poor survival of mesenchymal stem cells (MSCs) compromises the efficacy of stem cell therapy. Growth factor deprivation is one of the important factors that have challenged the survival of donor MSCs in cell therapy. In this study, the aim was to evaluate the effect of serum deprivation on the cell death of MSCs and to investigate the underlying mechanisms. Apoptosis of MSCs was evaluated with Hoechst 33342/PI staining. Signaling pathways involved in serum-deprivation induced apoptosis were analyzed using Western blotting. The results revealed that serum deprivation induced apoptosis in MSCs within 72 h of treatment. Serum deprivation was shown to lead to protein expression alterations in Bax, Bcl-2, casepase-3, casepase-8, GRP78, and CHOP during experiments. The data suggested that the mitochondria death pathway, the extrinsic apoptotic pathway and the endoplastic reticulum(ER) stress pathway were all involved in MSCs apoptosis. The increase in expression of CHOP and the simultaneous decrease in Bcl-2 expression suggest a synergistic effect in apoptosis induction in both the mitochondrion and the ER.

Radial shockwave treatment promotes human mesenchymal stem cell self-renewal and enhances cartilage healing

BackgroundShockwaves and mesenchymal stem cells (MSCs) have been widely accepted as useful tools for many orthopedic applications. However, the modulatory effects of shockwaves on MSCs remain controversial. In this study, we explored the influence of radial shockwaves on human bone marrow MSCs using a floating model in vitro and evaluated the healing effects of these cells on cartilage defects in vivo using a rabbit model.MethodsMSCs were cultured in vitro, harvested, resuspended, and treated with various doses of radial shockwaves in a floating system. Cell proliferation was evaluated by growth kinetics and Cell Counting Kit-8 (CCK-8) assay. In addition, the cell cycle and apoptotic activity were analyzed by fluorescence activated cell sorting. To explore the “stemness” of MSCs, cell colony-forming tests and multidifferentiation assays were performed. We also examined the MSC subcellular structure using transmission electron microscopy and examined the healing effects of these cells on cartilage defects by pathological analyses.ResultsThe results of growth kinetics and CCK-8 assays showed that radial shockwave treatment significantly promoted MSC proliferation. Enhanced cell growth was also reflected by an increase in the numbers of cells in the S phase and a decrease in the numbers of cells arrested in the G0/G1 phase in shockwave-treated MSCs. Unexpectedly, shockwaves caused a slight increase in MSC apoptosis rates. Furthermore, radial shockwaves promoted self-replicating activity of MSCs. Transmission electron microscopy revealed that MSCs were metabolically activated by shockwave treatment. In addition, radial shockwaves favored MSC osteogenic differentiation but inhibited adipogenic activity. Most importantly, MSCs pretreated by radial shockwaves exhibited an enhanced healing effect on cartilage defects in vivo. Compared with control groups, shockwave-treated MSCs combined with bio-scaffolds significantly improved histological scores of injured rabbit knees.ConclusionsIn the present study, we found that radial shockwaves significantly promoted the proliferation and self-renewal of MSCs in vitro and safely accelerated the cartilage repair process in vivo, indicating favorable clinical outcomes.

Tauroursodeoxycholic Acid Protects against the Effects of P-Cresol-Induced Reactive Oxygen Species via the Expression of Cellular Prion Protein.

Mesenchymal stem cells (MSCs) could be a promising solution in the treatment of various diseases including chronic kidney disease (CKD). However, endoplasmic reticulum (ER) stress induced by ischemia in the area of application limits the integration and survival of MSCs in patients. In our study, we generated ER stress-induced conditions in MSCs using P-cresol. As P-cresol is a toxic compound accumulated in the body of CKD patients and induces apoptosis and inflammation through reactive oxygen species (ROS), we observed ER stress-induced MSC apoptosis activated by oxidative stress, which in turn resulted from ROS generation. To overcome stress-induced apoptosis, we investigated the protective effects of tauroursodeoxycholic acid (TUDCA), a bile acid, on ER stress in MSCs. In ER stress, TUDCA treatment of MSCs reduced ER stress-associated protein activation, including GRP78, PERK, eIF2α, ATF4, IRE1α, and CHOP. Next, to explore the protective mechanism adopted by TUDCA, TUDCA-mediated cellular prion protein (PrPC) activation was assessed. We confirmed that PrPC expression significantly increased ROS, which was eliminated by superoxide dismutase and catalase in MSCs. These findings suggest that TUDCA protects from inflammation and apoptosis in ER stress via PrPC expression. Our study demonstrates that TUDCA protects MSCs against inflammation and apoptosis in ER stress by PrPC expression in response to P-cresol exposure.

Effect of neurotrophic factor -3 on the proliferation and apoptosis of bone marrow mesenchymal stem cells and mechanisms

Objective To investigate the effect of neurotrophin-3 (NT-3) on the proliferation and apoptosis of bone marrow mesenchymal stem cells (BMSCs) in rats and possible mechanisms. Methods The NT-3 overexpression and lentiviral transfection of BMSCs were co-cultured with neuronal cells respectively and then they were divided into overexpression control group, NT-3 transfection group and shRNA-NT-3 transfection group (NT-3 silencing group). MTT assay was used to detect the cell culture for 24 h, 48 h and 72 h. Cell cycle and apoptosis were detected by flow cytometry for 48 h. Real-time quantitative PCR was used to detect the expression of C/EBPβmRNA.The expression of C/EBPβprotein was detected by Western blot. Results MTT results showed that the proliferation ability of BMSCs in the NT-3 overexpression group was significantly higher than that in the control group (0.650±0.042, 0.826±0.074) at 48 h and 72 h (P<0.05). Compared with the control group (P<0.05), the cell cycle and apoptosis of BMSCs in NT-3 silencing group were significantly decreased at 48 h and 72 h (P<0.05). The results of 48 h cell cycle and apoptosis showed that the percentage of G1 phase in BMSCs was decreased, G2 and S were increased and the apoptosis was decreased. The percentage of G1 phase in G2-S phase and the increase of apoptosis were increased in NT-3 silencing group. The results of Western Blot showed that C/EBPβ mRNA and protein levels were significantly up-regulated in BMSCs of NT-3 overexpression group and significantly decreased in NT-3 silencing group (P<0.05). Conclusion NT-3 may promote the expression of C/EBP beta and affect the expression of its downstream target genes, which can inhibit the apoptosis of BMSCs cells. Key words: Bone marrow mesenchymal stem cells; Neurotrophin-3; CCAAT/enhancer binding proteins β; Neuronal; Apoptosis

2,5-hexanedione induces bone marrow mesenchymal stem cell apoptosis via inhibition of Akt/Bad signal pathway.

2,5-Hexanedione (HD) is an important bioactive metabolite of n-hexane and mediates the neurotoxicity of parent compound. Studies show that HD induces apoptotic death of neural progenitor cells. However, its underlying mechanism remains unknown. Mesenchymal stem cells (MSCs) are multipotential stem cells with the ability to differentiate into various cell types and have been used as cell model for studying the toxic effects of chemicals on stem cells. In this study, we exposed rat bone marrow MSCs to 0, 10, 20, and 40 mM HD in vitro. Apoptosis and disruption of mitochondrial transmembrane potential were estimated by immunochemistry staining. The expression of Akt, Bad, phosphorylated Akt (p-Akt), and Bad (p-Bad) as well as cytochrome c in mitochondria and cytosol were examined by Western blot. Moreover, caspase 3 activity, viability, and death of cells were measured by spectrophotometry. Our results showed that HD induced cell apoptosis and increased caspase 3 activity. HD down-regulated the expression levels of p-Akt, p-Bad and induced MMP depolarization, followed by cytochrome c release. Moreover, HD led to a concentration-dependent increase in the MSCs death, which was relative to MSCs apoptosis. However, these toxic effects of HD on the MSCs were significantly mitigated in the presence of IGF, which could activate PI3 K/Akt pathway. These results indicated that HD induced mitochondria-mediated apoptosis in the MSCs via inhibiting Akt/Bad signaling pathway and apoptotic death of MSCs via the signaling pathway. These results might provide some clues for studying further the mechanisms of HD-induced stem cell apoptosis and adverse effect on neurogenesis.

Human mesenchymal stem cells lose their functional properties after paclitaxel treatment

Mesenchymal stem cells (MSCs) are an integral part of the bone marrow niche and aid in the protection, regeneration and proliferation of hematopoietic stem cells after exposure to myelotoxic taxane anti-cancer agents, but the influence of taxane compounds on MSCs themselves remains incompletely understood. Here, we show that bone marrow-derived MSCs are highly sensitive even to low concentrations of the prototypical taxane compound paclitaxel. While MSCs remained metabolically viable, they were strongly impaired regarding both their proliferation and their functional capabilities after exposure to paclitaxel. Paclitaxel treatment resulted in reduced cell migration, delays in cellular adhesion and significant dose-dependent inhibition of the stem cells’ characteristic multi-lineage differentiation potential. Cellular morphology and expression of the defining surface markers remained largely unaltered. Paclitaxel only marginally increased apoptosis in MSCs, but strongly induced premature senescence in these stem cells, thereby explaining the preservation of the metabolic activity of functionally inactivated MSCs. The reported sensitivity of MSC function to paclitaxel treatment may help to explain the severe bone marrow toxicities commonly caused by taxane-based anti-cancer treatments.

Pigment epithelium-derived factor from ARPE19 promotes proliferation and inhibits apoptosis of human umbilical mesenchymal stem cells in serum-free medium

Clinical expansion of mesenchymal stem cells (MSCs) is hampered by the lack of knowledge regarding how to prevent MSC apoptosis and promote their proliferation in serum-free medium. Our in vitro studies demonstrated that human umbilical cord MSCs (HUCMSCs) underwent apoptosis in the serum-free medium. When HUCMSCs were co-cultured with retinal pigment epithelial cells (ARPE19), however, HUCMSCs exhibited normal growth and morphology in serum-free medium. Their colony formation was promoted by the conditioned medium (CM) of ARPE19 cells on Matrigel. Proteomics analysis showed that pigment epithelium-derived factor (PEDF) was one of the most abundant extracellular proteins in the ARPE19 CM, whereas enzyme-linked immunosorbent assay confirmed that large amounts of PEDF was secreted from ARPE19 cells. Adding anti-PEDF-blocking antibodies to the co-culture of HUCMSCs with ARPE19 cells increased apoptosis of HUCMSCs. Conversely, treatment with PEDF significantly reduced apoptosis and increased proliferation of HUCMSCs in serum-free medium. PEDF was further demonstrated to exert this anti-apoptotic effect by inhibiting P53 expression to suppress caspase activation. In vivo studies demonstrated that co-injection of HUCMSCs with ARPE19 cells in immunocompromised NOD-SCID mice also increased survival and decreased apoptosis of HUCMSCs. PEDF also showed no negative effect on the mesoderm differentiation capability of HUCMSCs. In conclusion, this study is the first to demonstrate that PEDF promotes HUCMSC proliferation and protects them from apoptosis by reducing p53 expression in the serum-free medium. This study provides crucial information for clinical-scale expansion of HUCMSCs.

Effect of High Hydrostatic Pressure Applied Before Cryopreservation on the Survival Rate and Quality of Porcine Mesenchymal Stem Cells After Thawing

ABSTRACTThe aim of the present study was to examine the effects of varied high hydrostatic pressure (HHP) values on survival rate, proliferation rate, cell multipotency (transcript expression of SOX2, C-MYC, and REX1) and apoptosis (expression of phosphatidylserine (PS), SURVIVIN at the RNA level and BAX at the protein level) of porcine mesenchymal stem cells (MSCs). MSCs were isolated from porcine bone marrow and cultured in vitro. Before cryopreservation and storage in liquid nitrogen, MSCs were subjected to HHP at the varied pressures of 20, 30, 40, 50, or 60 MPa for 1 h at 24°C. Immediately after thawing and after 8 days of in vitro culture, cells were subjected to trypan blue staining, cell counting, real-time Polymerase Chain Reaction (PCR), western blotting, and fluorescence microscopy. BAX protein expression was only estimated immediately after HHP to exclusively examine the impact of HHP on apoptosis of MSCs. The viability of MSC subjected to 40, 50, and 60 MPa and estimated immediately after thawing increased significantly (P < 0.001 for 60 MPa and P < 0.05 for 40 and 50 MPa) in comparison to control. The proliferation rate of MSCs subjected to 40 MPa HHP was significantly higher than in the control group (P < 0.02) after 8 days of in vitro culture. After 8 days of in vitro culture, no significant differences were noted in the survival rates, PS exposure, or levels of SOX2, C-MYC, REX1, and SURVIVIN gene expression in all analyzed groups compared to control. In conclusion: 40–60 MPa HHP has a positive impact by improving cell viability in short term. 20–60 MPa HHP does not induce nor decrease apoptosis in MSCs. Fortunately, HHP does not induce differentiation of MSC. Our results calls for further analysis using HHP values higher than 60 MPa.

Sox9 augments BMP2-induced chondrogenic differentiation by downregulating Smad7 in mesenchymal stem cells (MSCs)

Cartilage injuries caused by arthritis or trauma pose formidable challenges for effective clinical management due to the limited intrinsic proliferative capability of chondrocytes. Autologous stem cell-based therapies and transgene-enhanced cartilage tissue engineering may open new avenues for the treatment of cartilage injuries. Bone morphogenetic protein 2 (BMP2) induces effective chondrogenesis of mesenchymal stem cells (MSCs) and can thus be explored as a potential therapeutic agent for cartilage defect repair. However, BMP2 also induces robust endochondral ossification. Although the precise mechanisms through which BMP2 governs the divergence of chondrogenesis and osteogenesis remain to be fully understood, blocking endochondral ossification during BMP2-induced cartilage formation may have practical significance for cartilage tissue engineering. Here, we investigate the role of Sox9-donwregulated Smad7 in BMP2-induced chondrogenic differentiation of MSCs. We find that overexpression of Sox9 leads to a decrease in BMP2-induced Smad7 expression in MSCs. Sox9 inhibits BMP2-induced expression of osteopontin while enhancing the expression of chondrogenic marker Col2a1 in MSCs. Forced expression of Sox9 in MSCs promotes BMP2-induced chondrogenesis and suppresses BMP2-induced endochondral ossification. Constitutive Smad7 expression inhibits BMP2-induced chondrogenesis in stem cell implantation assay. Mouse limb explant assay reveals that Sox9 expands BMP2-stimulated chondrocyte proliferating zone while Smad7 promotes BMP2-intitated hypertrophic zone of the growth plate. Cell cycle analysis indicates that Smad7 induces significant early apoptosis in BMP2-stimulated MSCs. Taken together, our results strongly suggest that Sox9 may facilitate BMP2-induced chondrogenesis by downregulating Smad7, which can be exploited for effective cartilage tissue engineering.

Lactoferrin Protects Human Mesenchymal Stem Cells from Oxidative Stress-Induced Senescence and Apoptosis.

Mesenchymal stem cells (MSCs) have been suggested as a primary candidate for cell therapy applications because they have self-renewal and differentiation capabilities. Although they can be expanded in ex vivo system, clinical application of these cells is still limited because they survive poorly and undergo senescence or apoptosis when transplanted and exposed to environmental factors such as oxidative stress. Thus, reducing oxidative stress is expected to improve the efficacy of MSC therapy. The milk protein lactoferrin is a multifunctional iron-binding glycoprotein that plays various roles, including reduction of oxidative stress. Thus, we explored the effect of lactoferrin on oxidative stress-induced senescence and apoptosis of human MSCs (hMSCs). Measurement of reactive oxygen species (ROS) revealed that lactoferrin inhibited the production of hydrogen peroxide-induced intracellular ROS, suggesting lactoferrin as a good candidate as an antioxidant in hMSCs. Pretreatment of lactoferrin suppressed hydrogen peroxide-induced senescence of hMSCs. In addition, lactoferrin reduced hydrogen peroxide-induced apoptosis via inhibition of caspase-3 and Akt activation. These results demonstrate that lactoferrin can be a promising factor to protect hMSCs from oxidative stress-induced senescence and apoptosis, thus increasing the efficacy of MSC therapy.

HDAC6 deficiency induces apoptosis in mesenchymal stem cells through p53 K120 acetylation

The acetylation of p53 is critical in modulating its pro-apoptotic roles. However, its regulatory mechanism and physiological significance are unclear. Here, we show HDAC6 negatively regulates pro-apoptotic acetylation of p53 at lysine residue 120 (K120) in mesenchymal stem cells (MSCs). The loss of HDAC6 expression in MSCs increases K120 acetylation of p53, which is successfully reversed by the wild-type but not by catalytically dead HDAC6. Deletion of HDAC6 induces caspase-dependent apoptosis by promoting transactivation of Bax and suppression of Bcl-2. Moreover, HDAC6 deficiency leads to mitochondrial dysfunction characterized by aberrant reactive oxygen species production and defective oxidative phosphorylation, which is reversed by ectopic expression of wild-type or acetylation mimetic p53. This study demonstrates that HDAC6 is a critical regulator of a pro-apoptotic p53 K120 acetylation and mitochondrial function in MSCs, suggesting that the modulation of HDAC6 activity could be a novel approach to improve MSC- based therapies.

Injectable co-gels of collagen and decellularized vascular matrix improve MSC-based therapy for acute kidney injury

Transplantation of mesenchymal stem cells (MSCs) is promising for treatment of acute kidney injury (AKI), but their therapeutic effects are often limited under normal conditions. In this study, we prepared the co-gels of decellularized vascular matrix and collagen, and investigated whether the co-gels increase the therapeutic potentials of MSCs on AKI. In vitro studies indicated that the co-gels enhanced the paracrine effects of MSCs, and significantly reduced the apoptosis of MSCs under oxidative environments. When the co-gels were co-transplanted with MSCs into the kidney of model rats with ischemia-reperfusion (I/R)-induced AKI, the survival and paracrine effects of MSCs were enhanced in the injured kidney. More importantly, the co-gels increased the therapeutic effects of MSCs for AKI, as indicated by cell apoptosis, tissue damage, vascularization and renal function. Therefore, the co-gels of decellularized vascular matrix and collagen improved the therapeutic effects of MSCs, and might be promising for AKI treatment.

Iron accumulation leads to bone loss by inducing mesenchymal stem cells apoptosis

Iron accumulation leads to bone loss by inducing mesenchymal stem cells apoptosis

Abstract 24: Graphene Oxide Containing Thermosensitive Hydrogel Improves the Survival of Transplanted Stem Cells in Ischemic Myocardium

Bone marrow derived mesenchymal stem cell (MSC) therapy has the potential to preserve cardiac function and prevent heart failure following a myocardial infarction (MI). However, in our recent studies, long term survival of transplanted stem cells was not detected, which was associated with deterioration of cardiac function. Therefore, strategies to enhance survival of transplanted cells would preserve the benefits of MSCs therapy for heart repair. Recently, graphene oxide (GO) nanosheets are being widely investigated for their ability to promote adherence and homing of transplanted MSCs to the extra cellular matrix in the heart. However, the presence of reactive oxygen functional groups (-OH, -COOH) in the structure of GO increases oxidative stress and activate apoptotic pathways in MSCs, which impairs the benefits of GO based biomaterials. Therefore, to nullify these side effects of GO, we synthesized a novel thermosensitive hydrogel by conjugating GO with chitosan (CS). Our Fourier transformed infrared spectrum (FTIR) analysis revealed an interaction between the oxygen functional groups of GO and amino (-NH2) groups of chitosan. Our data also demonstrate that CS masked the reactive functional groups of GO and prevented GO mediated ROS induction. This novel chitosan-graphene oxide (CS-GO) composite exhibited optimal porosity for cell conjugation and retention. Furthermore, this novel biomaterial was cyto-compatible as it exerted negligible toxicity to encapsulated MSCs. Upon exposure to hypoxic/ischemic environment, we observed a significant increase in apoptosis in MSCs as evidenced by an increase in the Bax/Bcl-xl ratio and caspase 3 activation. However, conjugation of MSCs to CS-GO polymer prevented hypoxia induced apoptosis. Therefore, coupling of GO with chitosan prevented GO mediated oxidative stress and downregulated hypoxia induced apoptosis of MSCs. Therefore, the outcome of this study will provide a novel delivery system for MSCs to the heart and improve their long term survival in the infarcted heart.

Hydrogel elasticity and microarchitecture regulate dental-derived mesenchymal stem cell-host immune system cross-talk.

In this study, we demonstrate that alginate hydrogel regulates the viability and the fate of the encapsulated dental-derived MSCs through modulation of NF-kB pathway. Alginate hydrogels with smaller pores and higher elasticity prevent pro-inflammatory cytokine-induced MSC apoptosis by down-regulating the Caspase-3- and 8- associated proapoptotic cascade, leading to higher amounts of ectopic bone regeneration. MSCs encapsulated in hydrogel with higher elasticity exhibited lower expression levels of NF-kB p65 and Cox-2 in vivo. These findings confirm that the fate of encapsulated MSCs are affected by the stiffness and microarchitecture of the encapsulating hydrogel biomaterial, as well as presence of T-lymphocytes/pro-inflammatory cytokines providing evidence concerning material science, stem cell biology, the molecular mechanism of dental-derived MSC-associated therapies, and the potential clinical therapeutic impact of MSCs.

Complement C5a induces mesenchymal stem cell apoptosis during the progression of chronic diabetic complications.

Regeneration and repair mediated by mesenchymal stem cells (MSCs) are key self-protection mechanisms against diabetic complications, a reflection of diabetes-related cell/tissue damage and dysfunction. MSC abnormalities have been reported during the progression of diabetic complications, but little is known about whether a deficiency in these cells plays a role in the pathogenesis of this disease. In addition to MSC resident sites, peripheral circulation is a major source of MSCs that participate in the regeneration and repair of damaged tissue. Therefore, we investigated whether there is a deficiency of circulating MSC-like cells in people with diabetes and explored the underlying mechanisms. The abundance of MSC-like cells in peripheral blood was evaluated by FACS. Selected diabetic and non-diabetic serum (DS and NDS, respectively) samples were used to mimic diabetic and non-diabetic microenvironments, respectively. The proliferation and survival of MSCs under different serum conditions were analysed using several detection methods. The survival of MSCs in diabetic microenvironments was also investigated in vivo using leptin receptor mutant (Lepr db/db ) mice. Our data showed a significant decrease in the abundance of circulating MSC-like cells, which was correlated with complications in individuals with type 2 diabetes. DS strongly impaired the proliferation and survival of culture-expanded MSCs through the complement system but not through exposure to high glucose levels. DS-induced MSC apoptosis was mediated, at least in part, by the complement C5a-dependent upregulation of Fas-associated protein with death domain (FADD) and the Bcl-2-associated X protein (BAX)/B cell lymphoma 2 (Bcl-2) ratio, which was significantly inhibited by neutralising C5a or by the pharmacological or genetic inhibition of the C5a receptor (C5aR) on MSCs. Moreover, blockade of the C5a/C5aR pathway significantly inhibited the apoptosis of transplanted MSCs in Lepr db/db recipient mice. C5a-dependent apoptotic death is probably involved in MSC deficiency and in the progression of complications in individuals with type 2 diabetes. Therefore, anticomplement therapy may be a novel intervention for diabetic complications.