Marrow-derived Clonal Mesenchymal Stem Cells Research Articles

50997 A phase II trial of human bone marrow-derived clonal mesenchymal stem cells for treatment of adults with moderate-to-severe atopic dermatitis

50997 A phase II trial of human bone marrow-derived clonal mesenchymal stem cells for treatment of adults with moderate-to-severe atopic dermatitis

Transplantation of human bone marrow-derived clonal mesenchymal stem cells reduces fibrotic scar formation in a rat spinal cord injury model.

This study aimed to evaluate the therapeutic effect on tissue repair and scar formation of human bone marrow-derived clonal mesenchymal stem cells (hcMSCs) homogeneously isolated by using a subfractionation culturing method, in comparison with the non-clonal MSCs (hMSCs), in a rat spinal cord injury (SCI) model. The SCI was made using a vascular clip at the T9 level. Cells were transplanted into the lesion site 3days after injury. A functional test was performed over 4weeks employing a BBB score. Rats were killed for histological analysis at 3days, 1week and 4weeks after injury. The transplantation of hMSCs and hcMSCs significantly reduced lesion size and the fluid-filled cavity at 4weeks in comparison with the control group injected with phosphate buffered saline (PBS) (p<0.01). Transplantation of hcMSCs showed more axons reserved than that of hMSCs in the lesion epicentre filled with non-neuronal tissues. In addition, hMSCs and hcMSCs clearly reduced the inflammatory reaction and intraparenchymal hemorrhaging, compared with the PBS group. Interestingly, hcMSCs largely decreased Col IV expression, one of the markers of fibrotic scars. hcMSCs yielded therapeutic effects more than equal to those of hMSCs on the SCI. Both hMSCs and hcMSCs created an increase in axon regeneration and reduced scar formation around the SCI lesion. Copyright © 2017 John Wiley & Sons, Ltd.

Bone marrow-derived clonal mesenchymal stem cells inhibit NK- and T-cell infiltration and reduce alopecia areata in mouse model

Bone marrow-derived clonal mesenchymal stem cells inhibit NK- and T-cell infiltration and reduce alopecia areata in mouse model

Hypoxia Suppresses Spontaneous Mineralization and Osteogenic Differentiation of Mesenchymal Stem Cells via IGFBP3 Up-Regulation.

Hypoxia has diverse stimulatory effects on human adipose-derived stem cells (ASCs). In the present study, we investigated whether hypoxic culture conditions (2% O2) suppress spontaneous mineralization and osteogenic differentiation of ASCs. We also investigated signaling pathways and molecular mechanisms involved in this process. We found that hypoxia suppressed spontaneous mineralization and osteogenic differentiation of ASCs, and up-regulated mRNA and protein expression of Insulin-like growth factor binding proteins (IGFBPs) in ASCs. Although treatment with recombinant IGFBPs did not affect osteogenic differentiation of ASCs, siRNA-mediated inhibition of IGFBP3 attenuated hypoxia-suppressed osteogenic differentiation of ASCs. In contrast, overexpression of IGFBP3 via lentiviral vectors inhibited ASC osteogenic differentiation. These results indicate that hypoxia suppresses spontaneous mineralization and osteogenic differentiation of ASCs via intracellular IGFBP3 up-regulation. We determined that reactive oxygen species (ROS) generation followed by activation of the MAPK and PI3K/Akt pathways play pivotal roles in IGFBP3 expression under hypoxia. For example, ROS scavengers and inhibitors for MAPK and PI3K/Akt pathways attenuated the hypoxia-induced IGFBP3 expression. Inhibition of Elk1 and NF-κB through siRNA transfection also led to down-regulation of IGFBP3 mRNA expression. We next addressed the proliferative potential of ASCs with overexpressed IGFBP3, but IGFBP3 overexpression reduced the proliferation of ASCs. In addition, hypoxia reduced the osteogenic differentiation of bone marrow-derived clonal mesenchymal stem cells. Collectively, our results indicate that hypoxia suppresses the osteogenic differentiation of mesenchymal stem cells via IGFBP3 up-regulation.

P576. Therapeutic effects of mouse bone marrow-derived clonal mesenchymal stem cells in a mouse model of inflammatory bowel disease

P576. Therapeutic effects of mouse bone marrow-derived clonal mesenchymal stem cells in a mouse model of inflammatory bowel disease

Pharmacokinetics and In Vivo Fate of Intra-Articularly Transplanted Human Bone Marrow-Derived Clonal Mesenchymal Stem Cells

In this study, we report the pharmacokinetics and in vivo fate of intra-articularly transplanted human mesenchymal stem cells (MSCs) in comparison with those of intravenously administered cells. Bone marrow-derived human clonal mesenchymal stem cells (hcMSCs) were transplanted to nude mice through intravenous or intra-articular routes. The numbers of hcMSCs in blood and tissue samples were measured by the quantitative real-time-polymerase chain reaction (qPCR) with human Alu (hAlu) as a detection marker. Following intra-articular transplantation, the blood levels of hcMSCs peaked 8 h postdose and gradually diminished, showing a 95-fold higher mean residence time than hcMSCs delivered through the intravenous route. Unlike intravenously administered hcMSCs, intra-articularly injected hcMSCs were mainly retained at injection joint sites where their levels 8 h postdose were 116-fold higher than those in muscle tissues. Regardless of injection routes, biodistribution patterns did not significantly differ between normal and osteoarthritis-induced mice. Quantitative analysis using hAlu-specific qPCR revealed that hcMSC levels in joint tissues were significantly higher than those in muscle tissues 120 days postdose. These dramatic differences in kinetic behavior and fate of intra-articularly transplanted hcMSCs compared with intravenously administered hcMSCs may provide insights useful for the development of human MSCs for arthritis therapeutics.

Therapeutic effects of mouse bone marrow-derived clonal mesenchymal stem cells in a mouse model of inflammatory bowel disease

Mouse bone marrow-derived clonal mesenchymal stem cells (mcMSCs), which were originated from a single cell by a subfractionation culturing method, are recognized as new paradigm for stem cell therapy featured with its homogenous cell population. Next to proven therapeutic effects against pancreatitis, in the current study we demonstrated that mcMSCs showed significant therapeutic effects in dextran sulfate sodium (DSS)-induced experimental colitis model supported with anti-inflammatory and restorative activities. mcMSCs significantly reduced the disease activity index (DAI) score, including weight loss, stool consistency, and intestinal bleeding and significantly increased survival rates. The pathological scores were also significantly improved with mcMSC. We have demonstrated that especial mucosal regeneration activity accompanied with significantly lowered level of apoptosis as beneficiary actions of mcMSCs in UC models. The levels of inflammatory cytokines including TNF-α, IFN-γ, IL-1β, IL-6, and IL-17 were all significantly concurrent with significantly repressed NF-κB activation compared to the control group and significantly decreased infiltrations of responsible macrophage and neutrophil. Conclusively, our findings provide the rationale that mcMSCs are applicable as a potential source of cell-based therapy in inflammatory bowel diseases, especially contributing either to prevent relapse or to accelerate healing as solution to unmet medical needs in IBD therapy.

Galectin-9 is Involved in Immunosuppression Mediated by Human Bone Marrow-derived Clonal Mesenchymal Stem Cells.

Bone marrow-derived mesenchymal stem cells (MSCs) have immunomodulatory properties and can suppress exaggerated pro-inflammatory immune responses. Although the exact mechanisms remain unclear, a variety of soluble factors are known to contribute to MSC-mediated immunosuppression. However, functional redundancy in the immunosuppressive properties of MSCs indicates that other uncharacterized factors could be involved. Galectin-9, a member of the β-galactoside binding galectin family, has emerged as an important regulator of innate and adaptive immunity. We examined whether galectin-9 contributes to MSC-mediated immunosuppression. Galectin-9 was strongly induced and secreted from human MSCs upon stimulation with pro-inflammatory cytokines. An in vitro immunosuppression assay using a knockdown approach revealed that galectin-9-deficient MSCs do not exert immunosuppressive activity. We also provided evidence that galectin-9 may contribute to MSC-mediated immunosuppression by binding to its receptor, TIM-3, expressed on activated lymphocytes, leading to apoptotic cell death of activated lymphocytes. Taken together, our findings demonstrate that galectin-9 is involved in MSC-mediated immunosuppression and represents a potential therapeutic factor for the treatment of inflammatory diseases.

Therapeutic effect of human clonal bone marrow-derived mesenchymal stem cells in severe acute pancreatitis.

Severe acute pancreatitis (SAP), a common necroinflammatory disease initiated by the premature activation of digestive enzymes within the pancreatic acinar cells, is associated with significant morbidity and mortality. In this study, we investigated whether human bone marrow-derived clonal mesenchymal stem cells (hcMSCs), isolated from human bone marrow aspirate according to our newly established isolation protocol, have potential therapeutic effects in SAP. SAP was induced by three intraperitoneal (i.p.) injections of cerulein (100μg/kg) and sequential LPS (10mg/kg) in Sprague-Dawley (SD) rats. hcMSCs (1×10(6)/head) were infused on 24h after LPS injection via the tail vein. The rats were sacrificed 3days after infusion of hcMSCs. We observed that infused hcMSCs reduced the levels of serum amylase and lipase. Infused hcMSCs ameliorated acinar cell necrosis, pancreatic edema, and inflammatory infiltration. Also, infused hcMSCs decreased the level of malondialdehyde, and increased the levels of glutathione peroxidase and superoxide dismutase. The number of TUNEL positive acinar cells was reduced after hcMSCs infusion. In addition, hcMSCs reduced the expression levels of pro-inflammation mediators and cytokines, and increased the expression of SOX9 in SAP. Taken together, hcMSCs could effectively relieve injury of pancreatitis as a promising therapeutics for SAP.

Bone marrow-derived clonal mesenchymal stem cells inhibit ovalbumin-induced atopic dermatitis.

Mesenchymal stem cells (MSCs) possess immunomodulatory activities, including suppression of T- and B-cell activation. However, their effects on atopic dermatitis (AD) have not yet been studied. Using an ovalbumin-induced AD mouse model, we investigated whether MSCs can be used as therapeutics in AD. We isolated both allogeneic and syngeneic clonal MSCs (cMSCs) from mouse bone marrow according to the subfractionation culturing method. Our cMSCs suppressed both T- and B-cell activation. T-cell proliferation and cytokine production, including interferon (IFN)-γ and interleukin (IL)-4, were suppressed by inhibition of transcription factors, such as T-bet, GATA-3, and c-Maf. Those transcription factors were nitric oxide dependent. Immunoglobulin E (IgE) suppression occurred through downregulation of AID and BLIMP-1, important regulators for isotype class switch and B-cell differentiation. The cMSCs were injected intravenously into ovalbumin-induced AD mouse model, and the therapeutic effects were analyzed. Injection of both allogeneic and syngeneic cMSCs in an AD mouse model inhibited cell infiltration in skin lesions and decreased the serum level of IgE. IL-4 expression was also suppressed by cMSCs in both the lymph node and skin. The cMSCs migrated to skin lesions and draining lymph nodes. Taken together, these data demonstrated that cMSCs, which suppressed T- and B-cell functions, can be used for the treatment of AD in mice.

Molecular Characterization of Neurally Differentiated Human Bone Marrow-derived Clonal Mesenchymal Stem Cells

Bone marrow-derived mesenchymal stem cells (MSCs) are multipotent, with the ability to differentiate into different cell types. Additionally, the immunomodulatory activity of MSCs can downregulate inflammatory responses. The use of MSCs to repair injured tissues and treat inflammation, including in neuroimmune diseases, has been extensively explored. Although MSCs have emerged as a promising resource for the treatment of neuroimmune diseases, attempts to define the molecular properties of MSCs have been limited by the heterogeneity of MSC populations. We recently developed a new method, the subfractionation culturing method, to isolate homogeneous human clonal MSCs (hcMSCs). The hcMSCs were able to differentiate into fat, cartilage, bone, neuroglia, and liver cell types. In this study, to better understand the properties of neurally differentiated MSCs, gene expression in highly homogeneous hcMSCs was analyzed. Neural differentiation of hcMSCs was induced for 14 days. Thereafter, RNA and genomic DNA was isolated and subjected to microarray analysis and DNA methylation array analysis, respectively. We correlated the transcriptome of hcMSCs during neural differentiation with the DNA methylation status. Here, we describe and discuss the gene expression profile of neurally differentiated hcMSCs. These findings will expand our understanding of the molecular properties of MSCs and contribute to the development of cell therapy for neuroimmune diseases.

Bone Marrow-Derived Clonal Mesenchymal Stem Cells as a Source of Cell Therapy for Promoting Vocal Fold Wound Healing

We investigated whether mouse bone marrow-derived clonal mesenchymal stem cells (BM-cMSCs) could promote vocal fold (VF) wound healing by using a xenograft animal model. Homogeneous BM-cMSCs isolated by a subfractionation culturing method from the bone marrow aspirates of green fluorescent protein transgenic mice were injected into the VFs of rabbits immediately after direct mechanical injury. Macroscopic, biomechanical (rheometric), histologic, immunohistochemical, and transcriptional evaluations were performed on the scarred VFs 1 to 3 months after injury. Engraftment of the implanted BM-cMSCs was determined by detection of green fluorescent protein cells in the recipient VF by confocal microscopy. The BM-cMSC-treated VFs showed improved morphological properties and viscoelasticity as compared to control VFs injected with phosphate-buffered saline solution. Histologic and immunohistochemical evaluations showed less excessive collagen deposition and increased density of glycosaminoglycans in the BM-cMSC-treated VFs as compared to the control VFs at 3 months after injury (p = 0.003 and p = 0.037, respectively). BM-cMSC transplantation led to a significant attenuation of fibronectin (p = 0.036) and transforming growth factor beta1 (p = 0.042) messenger RNA expression at 1 month after injury. Green fluorescent protein-expressing BM-cMSCs engrafted in recipient VFs were found at 1 month after implantation. BM-cMSCs appeared to survive in the injured xenogeneic VFs after transplantation for up to 1 month and favorably enhanced the wound healing of VFs after injury. We conclude that BM-cMSCs are a possible source of cell therapy for vocal fold regeneration.

Intraglandular transplantation of bone marrow-derived clonal mesenchymal stem cells for amelioration of post-irradiation salivary gland damage

External irradiation in head and neck cancers may induce irreversible hyposalivation and consequent xerostomia, stemming from radiation damage to salivary glands (SGs). As cell-based therapy has been reported to be able to repair or restore damaged SG tissues, we attempted to determine whether bone marrow-derived clonal mesenchymal stem cells (BM-cMSCs) can ameliorate irradiation-induced salivary gland damage via a murine model. External irradiation at a dose of 15Gy was delivered to the neck fields of C57BL/6 mice. We directly administered either homologous mouse BM-cMSCs labeled with PKH26 (treatment group) or PBS (control group) into SGs 24h after irradiation. Salivary flow rate (SFR) and lag time of salivation were measured at 12weeks after transplantation. At 4 and 12weeks post-transplantation, we performed morphological, histological, and immunofluorescent examinations. Transdifferentiation of administered BM-cMSCs into salivary epithelial cells was observed by confocal microscopy. SFR was significantly increased in BM-cMSCs-transplanted mice compared with PBS-injected mice at 12weeks after transplantation. Administration of BM-cMSCs preserved the microscopic morphologies of SGs, with more functional acini in BM-cMSC-transplanted SGs than in PBS-injected SGs. Immunofluorescent staining revealed less apoptotic cells and increased microvessel density in BM-cMSC-transplanted SGs compared with PBS-injected SGs. PKH-26 labeled BM-cMSCs were detected in transplanted SGs at 4weeks after transplantation and in vivo transdifferentiation of BM-cMSCs into acinar cells was also observed. This study suggests that BM-cMSCs can ameliorate salivary damage following irradiation and can be used as a source of cell-based therapy for restoration of irradiation-induced salivary hypofunction.

Treatment of Atopic Dermatitis by Bone Marrow-derived Clonal Mesenchymal Stem Cells (119.3)

Abstract Mesenchymal stem cells (MSCs) can perform immunomodulatory activities, including suppression of T- and B-cell proliferation. MSCs have been primarily studied for their suppression of pro-inflammatory Th1 responses, but their therapeutic effects on atopic dermatitis (AD), which is a Th2-dominant disease, have not been reported. In this study, using an ovalbumin-induced mouse AD model, we investigated whether MSCs reduce the allergic inflammation in AD. We used mouse bone marrow-derived clonal mesenchymal stem cells (mcMSCs) isolated by our newly established isolation protocol termed the subfractionation culturing method (SCM). When injected intravenously into animals between antigen challenges, mcMSCs were found to inhibit the majority of AD-specific pathological changes by decreasing cell infiltration in skin lesions and reducing serum level of IgE. To explore the mechanism of this effect, we investigated the effects of mcMSCs on IgE production in B cells. We found that mcMSCs significantly inhibited IgE production when B cells were stimulated either by lipopolysaccharide (LPS) or anti-CD40 antibodies in the presence of interleukin (IL)-4. In addition, allogeneic mcMSCs can inhibit both isotype class switching and terminal differentiation of B cells probably by inhibiting NF-κB activation. These results suggest that mcMSC-based cell therapy is a promising therapeutic modality for AD treatment.