Stem cell therapy for nerve injury

Peripheral nerve injury has remained a substantial clinical complication with no satisfactory treatment options.Despite the great development in the field ofmicrosurgery,some severe types of neural injuries cannot be treated without causing tension to the injured nerve.Thus current studies have focused on the new approaches for the treatment of peripheral nerve injuries.Stem cells with the ability to differentiate into a variety of cell types have brought a new perspective to this matter.In this review,we will discuss the use of three main sources of mesenchymal stem cells in the treatment of peripheral nerve injuries.

Utility of co-transplanting mesenchymal stem cells in islet transplantation

Islet transplantation is characterized by the transplantation of isolated islets from donor pancreata into a diabetic recipient. Although it is a viable choice in the treatment of insulin dependent diabetes mellitus, most patients (approximately 90%) require insulin five years after transplantation. Recently, the co-transplantation of mesenchymal stem cells (MSCs) and islets in animal studies has revealed the effectiveness of MSCs co-transplantation for improving islet function. Themechanisms underlying the beneficial impact of MSCs include immunomodulation and the promotion of angiogenesis. In this review, we discuss MSCs and how they support improved graft survival and function.

Effect of CXCR3/HO-1 genes modified bone marrow mesenchymal stem cells on small bowel transplant rejection

AIM To investigate whether bone marrow mesenchymal stem cells(BMMSCs) modified with the HO-1 and CXCR3 genes can augment the inhibitory effect of BMMSCs on small bowel transplant rejection.METHODS Lewis rat BMMSCs were cultured in vitro. Thirdpassage BMMSCs were transduced with the CXCR3/HO-1 genes or the HO-1 gene alone. The rats were divided into six groups and rats in the experimental group were pretreated with BMMSCs 7 d prior to smallbowel transplant. Six time points(instant, 1 d, 3 d, 7 d, 10 d, and 14 d)(n = 6) were chosen for each group. Hematoxylin-eosin staining was used to observe pathologic rejection, while immunohistochemistry and Western blot were used to detect protein expression. Flow cytometry was used to detect T lymphocytes and enzyme linked immunosorbent assay was used to detect cytokines.RESULTS The median survival time of BMMSCs from the CXCR3/HO-1 modified group(53 d) was significantly longer than that of the HO-1 modified BMMSCs group(39 d), the BMMSCs group(26 d), and the NS group(control group)(16 d)(P < 0.05). Compared with BMMSCs from the HO-1 modified BMMSCs, BMMSCs, and NS groups, rejection of the small bowel in the CXCR3/HO-1 modified group was significantly reduced, while the weight of transplant recipients was also significantly decreased(P < 0.05). Furthermore, IL-2, IL-6, IL-17, IFN-γ, and TNF-α levels were significantly decreased and the levels of IL-10 and TGF-β were significantly increased(P < 0.05). CONCLUSION BMMSCs modified with the CXCR3 and HO-1 genes can abrogate the rejection of transplanted small bowel more effectively and significantly increase the survival time of rats that receive a small bowel transplant.

Mesenchymal stem cells rescue acute hepatic failure by polarizing M2 macrophages

AIM To investigate whether M1 or M2 polarization contributes to the therapeutic effects of mesenchymal stem cells(MSCs) in acute hepatic failure(AHF).METHODS MSCs were transfused into rats with AHF induced by D-galactosamine(DGal N). The therapeutic effects of MSCs were evaluated based on survival rate and hepatocyte proliferation and apoptosis. Hepatocyte regeneration capacity was evaluated by the expression of the hepatic progenitor surface marker epithelial cell adhesion molecule(Ep CAM). Macrophage polarization was analyzed by M1 markers [CD68,tumor necrosis factor alpha(TNF-α),interferon-γ(IFN-γ),inducible nitric oxide synthase(INOS)] and M2 markers [CD163,interleukin(IL)-4,IL-10,arginase-1(Arg-1)] in the survival and death groups after MSC transplantation.RESULTS The survival rate in the MSC-treated group was increased compared with the DPBS-treated control group(37.5% vs 10%). MSC treatment protected rats with AHF by reducing apoptotic hepatocytes and promoting hepatocyte regeneration. Immunohistochemical analysis showed that MSC treatment significantly increased the expression of Ep CAM compared with the control groups(P < 0.001). Expression of Ep CAM in the survival group was significantly up-regulated compared with the death group after MSC transplantation(P = 0.003). Transplantation of MSCs significantly improved the expression of CD163 and increased the gene expression of IL-10 and Arg-1 in the survival group. IL-4 concentrations were significantly increased compared to the death group after MSC transplantation(88.51 ± 24.51 pg/m L vs 34.61 ± 6.6 pg/m L,P < 0.001). In contrast,macrophages showed strong expression of CD68,TNF-α,and INOS in the death group. The concentration of IFN-γ was significantly increased compared to the survival group after MSC transplantation(542.11 ± 51.59 pg/m L vs 104.07 ± 42.80 pg/m L,P < 0.001).CONCLUSION M2 polarization contributes to the therapeutic effects of MSCs in AHF by altering levels of anti-inflammatory and pro-inflammatory factors.

Mesenchymal stromal cell-based therapy: Regulatory and translational aspects in gastroenterology

The past decade has witnessed an outstanding scientific production focused towards the possible clinical applications of mesenchymal stromal cells(MSCs) in autoimmune and chronic inflammatory diseases. This raised the need of novel standards to adequately address quality, efficacy and safety issues of this advanced therapy. The development of a streamlined regulation is currently hampered by the complexity of analyzing dynamic biological entities rather than chemicals. Although numerous pieces of evidence show efficacy in reducing intestinal inflammation, some inconsistencies between the mechanisms of action of rodent vs human MSCs suggest caution before assigning translational value to preclinical studies. Preliminary evidence from clinical trials showed efficacy of MSCs in the treatment of fistulizing Crohn's disease(CD), and preparations of heterologous MSCs for CD treatment are currently tested in ongoing clinical trials. However, safety issues, especially in longterm treatment, still require solid clinical data. In this regard, standardized guidelines for appropriate dosing and methods of infusion could enhance the likelihood to predict more accurately the number of responders and the duration of remission periods. In addition, elucidating MSC mechanisms of action could lead to novel and more reliable formulations such as those derived from the MSCs themselves(e.g., supernatants).

Regenerative medicine using dental pulp stem cells for liver diseases

Acute liver failure is a refractory disease and its pro-gnosis, if not treated using liver transplantation, is extremely poor. It is a good candidate for regenerative medicine, where stem cell-based therapies play a central role. Mesenchymal stem cells(MSCs) are known to differentiate into multiple cell lineages including hepatocytes. Autologous cell transplant without any foreign gene induction is feasible using MSCs, thereby avoiding possible risks of tumorigenesis and immune rejection. Dental pulp also contains an MSC population that differentiates into hepatocytes. A point worthy of special mention is that dental pulp can be obtained from deciduous teeth during childhood and can be subsequently harvested when necessary after deposition in a tooth bank. MSCs have not only a regenerative capacity but also act in an anti--inflammatory manner via paracrine mechanisms. Promising efficacies and difficulties with the use of MSC derived from teeth are summarized in this review.

The Experimental Study on Constructing the Tissue Engineered Myocardium-like Tissue in vitro with Bone Mesenchymal Stem Cells

Objective:Unlike other tissues,myocardium has not substitute whick can be used to repair damaged cardiac tissue.This paper proposes engineering 3-D myocardium-like tissue constructs in vitro with bone mesenchymal stem cells(BMSCs) of infant and poly-lactic-co-glycolic acid(PLGA)in vitro.Methods:Bone marrow was obtained from the sternal marrow cavum outflow of infant with congenital heart disease (CHD)undergoing cardiac operation.BMSCs were obtained by density gradient centrifugation.The cells in passages two were induced in DMED with 10 umol/L 5- Azacytidine(5-Aza)for 24 h.When the induced BMSCS were cultured nearly into filled,the cells were planted in the scaffold of PLGA in 5.5×106 cells/cm2.The cell- scaffold complex has been cultured in the shake cultivation for 1 week,then the complex has been planted in the dorse of the nude mouse.When the experiment had been finished,the histology,immunology,real time PCR and so on were done.Results: The BMSCs of infant with congenital heart disease have the properties of the stable growth and the rapid proliferation.The immunohistochemistry showed that tissue engineered myocardium constructed in vitro expressed some cardiac related proteins such asα-actin,Cx-43,Desmine,cTNI and so on.The transparent myofilaments,gap junctions and intercalated disk-like structure formation could be observed in the 3D tissue-like constructs by transmission electron microscope(TEM).The engineered myocardium-like tissue had the auto-myocardial property as assessed by real time- PCR and so on.Conclusion:The engineered myocardial tissue-like constructs could be built with infant BMSCs and PLGA in vitro.Our results may provide the first step on the long road toward engineering myocardial material for repairing the defect or augmenting the tract in CHD,such as ventricular septal defect,tetralogy of Fallot and so on.

All-trans retinoic acid promotes smooth muscle cell differentiation of rabbit bone marrow-derived mesenchymal stem cells

Bone marrow-derived mesenchymal stem cells are multipotent stem cells, an attractive resource for regenerative medicine. Accumulating evidence suggests that all-trans retinoic acid plays a key role in the devel- opment and differentiation of smooth muscle cells. In the present study, we demonstrate, for the first time, that rabbit bone marrow-derived mesenchymal stem cells differentiate into smooth muscle cells upon the treatment with all-trans retinoic acid. All-trans retinoic acid increased the expression of myocardin, caldesmon, 22-kDa smooth muscle cellspecific protein (SM22α), and SM-myosin heavy chains in rabbit bone marrow-derived mesenchymal stem cells, as detected by reverse transcription polymerase chain reaction (PCR). Immunostaining of SM22α and SM-myosin heavy chains using monoclonal antibodies also indicated smooth muscle cell differentiation of rabbit bone marrowderived mesenchymal stem cells following the treatment with all-trans retinoic acid. In addition, more than 47% of bone marrow-derived mesenchymal stem cells demonstrated the contractile phenotype of smooth muscle cells. Western blot results showed that SM-1 and SM-2 were highly expressed in the differentiated cells. These results suggest that all-trans retinoic acid may serve as a potent agent for functional smooth muscle cell differentiation in tissue engineering.

Angiopoietin-1 preconditioning enhances survival and functional recovery of mesenchymal stem cell transplantation

Objective:Mesenchymal stem cell (MSC) transplantation is a promising therapy for ischemic heart diseases. However, poor cell survival after transplantation greatly limits the therapeutic efficacy of MSCs. The purpose of this study was to investigate the protective effect of angiopoietin-1 (Ang1) preconditioning on MSC survival and sub-sequent heart function improvement after transplantation. Methods: MSCs were cultured with or without 50 ng/ml Ang1 in complete medium for 24 h prior to experiments on cell survival and transplantation. 3-(4,5- Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and Hoechst staining were applied to evaluate MSC survival after serum deprivation in vitro, while cell survival in vivo was detected by terminal deoxynucleotidyl trans-ferase biotin-dUPT nick end labeling (TUNEL) assay 24 and 72 h after transplantation. Heart function and infarct size were measured four weeks later by small animal echocardiography and Masson's trichrome staining, respectively. Results: Ang1 preconditioning induced Akt phosphorylation and increased expression of Bcl-2 and the ratio of Bcl-2/Bax. In comparison with non-preconditioned MSCs, Ang1-preconditioned cell survival was significantly increased while the apoptotic rate decreased in vitro. However, the PI3K/Akt pathway inhibitor, LY294002, abrogated the protective effect of Ang1 preconditioning. After transplantation, the Ang1-preconditioned-MSC group showed a lower death rate, smaller infarct size, and better heart functional recovery compared to the non-preconditioned-MSC group. Conclusions:Ang1 preconditioning enhances MSC survival, contributing to further improvement of heart function.

Tongxinluo promotes mesenchymal stem cell tube formation in vitro

Objective:To study whether Tongxinluo (TXL) can induce angiogenesis in bone marrow mesenchymal stem cells (MSCs),and to investigate the underlying mechanism.Methods:Bone marrow MSCs were obtained from male Sprague-Dawley rats.We established an angiogenesis model in vitro via matrigel experiment.MSCs were seeded on matrigel coated 24-well plates,and treated by TXL 50 and 100 mg/L.After 24 h,we observed the tube formations of MSCs in the matrigel.Cell migration ability was examined by wound scratch test and transwell assay.Expressions of vascular endothelial growth factor (VEGF),fetal liver kinase-1 (Flk-1),matrix metalloproteinase-2 (MMP-2),MMP-9,and tissue inhibitor of metalloproteinase-2 (TIMP-2) were analyzed at the protein level by Western blot.Gelatin zymography assay was applied to investigating the MSC paracrine abilities of pro-MMP-2 and activated MMP-2.Results:TXL promoted MSC tube formation in matrigel.The ratio of TXL 100 mg/L treated-MSC tubular length was increased 3.04-fold compared to the control group (P<0.05).Scratch test and transwell assay showed that TXL could improve the cell migration ability of MSCs.Western blot experiments showed that TXL promoted MSC synthesis of MMP-2,but it had no influence on the expressions of MMP-9 and TIMP-2.This effect was confirmed by gelatin zymography assay,which showed that TXL increased MSC secretion of pro-MMP-2 and activated MMP-2.VEGF expression of TXL treated-MSCs was increased compared to the control group.The expression of Flk-1 was not different among the groups.Conclusions:This study demonstrates that TXL can promote the tube formation of MSCs,and the underlying mechanisms are associated with increased migration ability of MSCs and the up-regulation of MMP-2 and VEGF expressions.

Immunomodulative effects of mesenchymal stem cells derived from human embryonic stem cells in vivo and in vitro

Objective: Human embryonic stem cells (hESCs) have recently been reported as an unlimited source of mesenchymal stem cells (MSCs).The present study not only provides an identical and clinically compliant MSC source derived from hESCs (hESC-MSCs),but also describes the immunomodulative effects of hESC-MSCs in vitro and in vivo for a carbon tetrachloride (CCl4)-induced liver inflammation model.Methods: Undifferentiated hESCs were treated with Rho-associated kinase (ROCK) inhibitor and induced to fibroblast-looking cells.These cells were tested for their surface markers and multilineage differentiation capability.Further more,we analyzed their immune characteristics by mixed lymphocyte reactions (MLRs) and animal experiments.Results: hESC-MSCs show a homogenous fibroblastic morphology that resembles bone marrow-derived MSCs (BM-MSCs).The cell markers and differentiation potential of hESC-MSCs are also similar to those of BM-MSCs.Unlike their original cells,hESC-MSCs possess poor immunogenicity and can survive and be engrafted into a xenogenic immunocompetent environment.Conclusions: The hESC-MSCs demonstrate strong inhibitory effects on lymphocyte proliferation in vitro and anti-inflammatory infiltration properties in vivo.This study offers information essential to the applications of hESC-MSC-based therapies and evidence for the therapeutic mechanisms of action.

Growth factors induce the improved cardiac remodeling in autologous mesenchymal stem cell-implanted failing rat hearts

Therapeutically delivered mesenchymal stem cells (MSCs) improve ventricular remodeling. However,the mechanism underlying MSC cardiac remodeling has not been clearly determined. Congestive heart failure (CHF) was induced in rats by cauterization of the left ventricular free wall. MSCs were cultured from autologous bone marrow and injected into the border zone and the remote myocardium 5 d after injury. Ten weeks later,when compared with sham operation,CHF significantly increased nucleus mitotic index,capillary density,and expression of insulin-like growth factor 1,hepatocyte growth factor and vascular endothelial growth factor in the border zone (P<0.01) and decreased each of them in the remote myocardium (P<0.05 or P<0.01). MSC implantation in CHF dramatically elevated ex-pression of these growth factors in the remote myocardium and further elevated their expression in the border zone when compared with CHF without MSC addition (P<0.05 or P<0.01). This was paralleled by a higher nucleus mitotic index and a significantly increased capillary density both in the remote myocardium and in the border zone,and by a lower percentage of area of collagen and a higher percentage of area of myocardium in the border zone (P<0.05 or P<0.01),and cardiac remodeling markedly improved. Autologous MSC implantation promoted expression of growth factors in rat failing myocardium,which might enhance cardiomyogenesis and angiogenesis,and improved cardiac remodeling.