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Sample records for marrow-derived mesenchymal stem

  1. Enhanced adipogenic differentiation of bovine bone marrow-derived mesenchymal stem cells

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Until now, the isolation and characterization of bovine bone marrow-derived mesenchymal stem cells (bBM-MSCs) have not been established, which prompted us to optimize the differentiation protocol for bBM-MSCs. In this study, bBM-MSCs were freshly isolated from three 6-month-old cattle and used for p...

  2. Immunoregulatory effects of bone marrow-derived mesenchymal stem cells in the nasal polyp microenvironment.

    PubMed

    Pezato, Rogério; de Almeida, Danilo Cândido; Bezerra, Thiago Freire; Silva, Fernando de Sá; Perez-Novo, Claudina; Gregório, Luís Carlos; Voegels, Richard Louis; Câmara, Niels Olsen; Bachert, Claus

    2014-01-01

    Nasal polyposis is a severe, chronic inflammatory condition of the paranasal sinuses and is frequently associated with asthma and aspirin sensitivity. Mesenchymal stem cells exhibit a potent immunosuppressive effect in several inflammatory conditions, and their role in nasal polyposis remains little explored. Hence, we investigated whether bone marrow-derived mesenchymal stem cells could modulate cell phenotype in the nasal polyp milieu. After coculture with mesenchymal stem cells, the frequency of these inflammatory cells was found to decrease. Furthermore, mesenchymal stem cells promoted strong inhibition of CD4+ and CD8+ T cell proliferation, increased the frequency of CD4+CD25+Foxp3 T cells, and changed the global cytokine profile from an inflammatory to an anti-inflammatory response. We believe that mesenchymal stem cells may be a very useful adjunct for investigation of the inflammatory process in nasal polyposis, contributing to better understanding of the inflammatory course of this condition. PMID:24707116

  3. Suppression of ornithine decarboxylase promotes osteogenic differentiation of human bone marrow-derived mesenchymal stem cells.

    PubMed

    Tsai, Yo-Hsian; Lin, Kuan-Lian; Huang, Yuan-Pin; Hsu, Yi-Chiang; Chen, Chung-Hwan; Chen, Yuhsin; Sie, Min-Hua; Wang, Gwo-Jaw; Lee, Mon-Juan

    2015-07-22

    Ornithine decarboxylase (ODC) is the rate-limiting enzyme for polyamine biosynthesis. Suppression of ODC by its irreversible inhibitor, α-difluoromethylornithine (DFMO), or by RNA interference through siRNA, enhanced osteogenic gene expression and alkaline phosphatase activity, and accelerated matrix mineralization of human bone marrow-derived mesenchymal stem cells (hBMSCs). Besides, adipogenic gene expression and lipid accumulation was attenuated, indicating that the enhanced osteogenesis was accompanied by down-regulation of adipogenesis when ODC was suppressed. A decrease in the intracellular polyamine content of hBMSCs during osteogenic induction was observed, suggesting that the level of endogenous polyamines is regulated during differentiation of hBMSCs. This study elucidates the role of polyamine metabolism in the lineage commitment of stem cells and provides a potential new indication for DFMO as bone-stimulating drug. PMID:26140984

  4. Bone-marrow-derived mesenchymal stem cells inhibit gastric aspiration lung injury and inflammation in rats.

    PubMed

    Zhou, Jing; Jiang, Liyan; Long, Xuan; Fu, Cuiping; Wang, Xiangdong; Wu, Xiaodan; Liu, Zilong; Zhu, Fen; Shi, Jindong; Li, Shanqun

    2016-09-01

    Gastric aspiration lung injury is one of the most common clinical events. This study investigated the effects of bone-marrow-derived mesenchymal stem cells (BMSCs) on combined acid plus small non-acidified particle (CASP)-induced aspiration lung injury. Enhanced green fluorescent protein (EGFP(+) ) or EGFP(-) BMSCs or 15d-PGJ2 were injected via the tail vein into rats immediately after CASP-induced aspiration lung injury. Pathological changes in lung tissues, blood gas analysis, the wet/dry weight ratio (W/D) of the lung, levels of total proteins and number of total cells and neutrophils in bronchoalveolar lavage fluid (BALF) were determined. The cytokine levels were measured using ELISA. Protein expression was determined by Western blot. Bone-marrow-derived mesenchymal stem cells treatment significantly reduced alveolar oedema, exudation and lung inflammation; increased the arterial partial pressure of oxygen; and decreased the W/D of the lung, the levels of total proteins and the number of total cells and neutrophils in BALF in the rats with CASP-induced lung injury. Bone-marrow-derived mesenchymal stem cells treatment decreased the levels of tumour necrosis factor-α and Cytokine-induced neutrophil chemoattractant (CINC)-1 and the expression of p-p65 and increased the levels of interleukin-10 and 15d-PGJ2 and the expression of peroxisome proliferator-activated receptor (PPAR)-γ in the lung tissue in CASP-induced rats. Tumour necrosis factor-α stimulated BMSCs to secrete 15d-PGJ2 . A tracking experiment showed that EGFP(+) BMSCs were able to migrate to local lung tissues. Treatment with 15d-PGJ2 also significantly inhibited CASP-induced lung inflammation and the production of pro-inflammatory cytokines. Our results show that BMSCs can protect lung tissues from gastric aspiration injury and inhibit lung inflammation in rats. A beneficial effect might be achieved through BMSC-derived 15d-PGJ2 activation of the PPAR-γ receptor, reducing the production of

  5. Therapeutic potential of bone marrow-derived mesenchymal stem cells for cutaneous wound healing.

    PubMed

    Chen, Jerry S; Wong, Victor W; Gurtner, Geoffrey C

    2012-01-01

    Despite advances in wound care, many wounds never heal and become chronic problems that result in significant morbidity and mortality to the patient. Cellular therapy for cutaneous wounds has recently come under investigation as a potential treatment modality for impaired wound healing. Bone marrow-derived mesenchymal stem cells (MSCs) are a promising source of adult progenitor cells for cytotherapy as they are easy to isolate and expand and have been shown to differentiate into various cell lineages. Early studies have demonstrated that MSCs may enhance epithelialization, granulation tissue formation, and neovascularization resulting in accelerated wound closure. It is currently unclear if these effects are mediated through cellular differentiation or by secretion of cytokines and growth factors. This review discusses the proposed biological contributions of MSCs to cutaneous repair and their clinical potential in cell-based therapies. PMID:22787462

  6. Epigenetic Plasticity Drives Adipogenic and Osteogenic Differentiation of Marrow-derived Mesenchymal Stem Cells.

    PubMed

    Meyer, Mark B; Benkusky, Nancy A; Sen, Buer; Rubin, Janet; Pike, J Wesley

    2016-08-19

    Terminal differentiation of multipotent stem cells is achieved through a coordinated cascade of activated transcription factors and epigenetic modifications that drive gene transcription responsible for unique cell fate. Within the mesenchymal lineage, factors such as RUNX2 and PPARγ are indispensable for osteogenesis and adipogenesis, respectively. We therefore investigated genomic binding of transcription factors and accompanying epigenetic modifications that occur during osteogenic and adipogenic differentiation of mouse bone marrow-derived mesenchymal stem cells (MSCs). As assessed by ChIP-sequencing and RNA-sequencing analyses, we found that genes vital for osteogenic identity were linked to RUNX2, C/EBPβ, retinoid X receptor, and vitamin D receptor binding sites, whereas adipocyte differentiation favored PPARγ, retinoid X receptor, C/EBPα, and C/EBPβ binding sites. Epigenetic marks were clear predictors of active differentiation loci as well as enhancer activities and selective gene expression. These marrow-derived MSCs displayed an epigenetic pattern that suggested a default preference for the osteogenic pathway; however, these patterns were rapidly altered near the Adipoq, Cidec, Fabp4, Lipe, Plin1, Pparg, and Cebpa genes during adipogenic differentiation. Surprisingly, we found that these cells also exhibited an epigenetic plasticity that enabled them to trans-differentiate from adipocytes to osteoblasts (and vice versa) after commitment, as assessed by staining, gene expression, and ChIP-quantitative PCR analysis. The osteogenic default pathway may be subverted during pathological conditions, leading to skeletal fragility and increased marrow adiposity during aging, estrogen deficiency, and skeletal unloading. Taken together, our data provide an increased mechanistic understanding of the epigenetic programs necessary for multipotent differentiation of MSCs that may prove beneficial in the development of therapeutic strategies. PMID:27402842

  7. Sertoli cells promote proliferation of bone marrow-derived mesenchymal stem cells in co-culture.

    PubMed

    Zhang, Fenxi; Lu, Ming; Liu, Hengxing; Ren, Tongming; Miao, Yingying; Wang, Jingjing

    2016-05-01

    Bone marrow-derived mesenchymal stem cells (BMSCs) are a major source for cell transplantation. The proliferative ability of BMSCs is an important determinant of the efficiency of transplant therapy. Sertoli cells are "nurse" cells for development of sperm cells. Our recent study showed that Sertoli cells promoted proliferation of human umbilical cord mesenchymal stem cells (hUCMSCs) in co-culture. Studies by other groups also showed that Sertoli cells promoted growth of endothelial cells and neural stem cells. In this study, we investigated the effect of Sertoli cells on proliferation of BMSCs. Our results showed that Sertoli cells in co-culture significantly enhanced proliferation of BMSCs (P < 0.01). Moreover, co-culture with Sertoli cells also markedly increased mRNA and/or protein expressions of Mdm2, p-Akt and Cyclin D1, and decreased p53 expression in BMSCs (P < 0.01 or < 0.05). These findings indicate that Sertoli cells have the potential to enhance proliferation of BMSCs. PMID:27319049

  8. A Modified Method of Insulin Producing Cells' Generation from Bone Marrow-Derived Mesenchymal Stem Cells

    PubMed Central

    Czubak, Paweł; Putowski, Lechosław

    2014-01-01

    Type 1 diabetes mellitus is a result of autoimmune destruction of pancreatic insulin producing β-cells and so far it can be cured only by insulin injection, by pancreas transplantation, or by pancreatic islet cells' transplantation. The methods are, however, imperfect and have a lot of disadvantages. Therefore new solutions are needed. The best one would be the use of differentiated mesenchymal stem cells (MSCs). In the present study, we investigated the potential of the bone marrow-derived MSCs line for in vitro differentiation into insulin producing cells (IPSs). We applied an 18-day protocol to differentiate MSCs. Differentiating cells formed cell clusters some of which resembled pancreatic islet-like cells. Using dithizone we confirmed the presence of insulin in the cells. What is more, the expression of proinsulin C-peptide in differentiated IPCs was analyzed by flow cytometry. For the first time, we investigated the influence of growth factors' concentration on IPCs differentiation efficiency. We have found that an increase in the concentration of growth factors up to 60 ng/mL of β-FGF/EGF and 30 ng/mL of activin A/β-cellulin increases the percentage of IPCs. Further increase of growth factors does not show any increase of the percentage of differentiated cells. Our findings suggest that the presented protocol can be adapted for differentiation of insulin producing cells from stem cells. PMID:25405207

  9. A modified method of insulin producing cells' generation from bone marrow-derived mesenchymal stem cells.

    PubMed

    Czubak, Paweł; Bojarska-Junak, Agnieszka; Tabarkiewicz, Jacek; Putowski, Lechosław

    2014-01-01

    Type 1 diabetes mellitus is a result of autoimmune destruction of pancreatic insulin producing β-cells and so far it can be cured only by insulin injection, by pancreas transplantation, or by pancreatic islet cells' transplantation. The methods are, however, imperfect and have a lot of disadvantages. Therefore new solutions are needed. The best one would be the use of differentiated mesenchymal stem cells (MSCs). In the present study, we investigated the potential of the bone marrow-derived MSCs line for in vitro differentiation into insulin producing cells (IPSs). We applied an 18-day protocol to differentiate MSCs. Differentiating cells formed cell clusters some of which resembled pancreatic islet-like cells. Using dithizone we confirmed the presence of insulin in the cells. What is more, the expression of proinsulin C-peptide in differentiated IPCs was analyzed by flow cytometry. For the first time, we investigated the influence of growth factors' concentration on IPCs differentiation efficiency. We have found that an increase in the concentration of growth factors up to 60 ng/mL of β-FGF/EGF and 30 ng/mL of activin A/β-cellulin increases the percentage of IPCs. Further increase of growth factors does not show any increase of the percentage of differentiated cells. Our findings suggest that the presented protocol can be adapted for differentiation of insulin producing cells from stem cells. PMID:25405207

  10. Therapeutic effects of bone marrow-derived mesenchymal stem cells on radiation-induced lung injury.

    PubMed

    Xia, Chengcheng; Chang, Pengyu; Zhang, Yuyu; Shi, Weiyan; Liu, Bin; Ding, Lijuan; Liu, Min; Gao, Ling; Dong, Lihua

    2016-02-01

    Radiation-induced lung injury (RILI) is a fatal condition featured by interstitial pneumonitis and fibrosis. Mesenchymal stem cells (MSCs) have been widely used for treating RILI in rodent models. In the present study, we aimed to investigate whether the therapeutic effects of human bone marrow-derived mesenchymal stem cells (hBM-MSCs) on RILI were in a dose-dependent manner. A total of 100 mice were randomly divided into: a control group (n=25), subject to lung irradiation and injection of phosphate-buffered solution (PBS) via the tail vein; and the hBM-MSC group, subject to lung irradiation followed by injection of a low dose (1x103 hBM-MSCs/g), medium dose (5x103 hBM-MSCs/g) and high dose (1x104 hBM-MSCs/g) of hBM-MSCs in PBS through the tail vein, respectively. After sacrifice, the pulmonary tissues were subject to hematoxylin and eosin (H&E) staining, Masson's trichrome staining and immunohistochemical staining to investigate the pathological changes. Immunofluorescent staining was performed to evaluate the differentiation capacity of hBM-MSCs in vivo by analyzing the expression of SPC and PECAM. hBM-MSCs improved the survival rate and histopathological features in the irradiated mice, especially in the low-dose group. Marked decrease in collagen deposition was noted in the irradiated mice treated using a low dose of hBM-MSCs. In addition, hBM-MSCs attenuated secretion and expression of IL-10 and increased the expression of TNF-α. Furthermore, hBM-MSCs had the potential to differentiate into functional cells upon lung injury. Low-dose hBM-MSCs contributed to functional recovery in mice with RILI. PMID:26717975

  11. Chemically Functionalized Silk for Human Bone Marrow-Derived Mesenchymal Stem Cells Proliferation and Differentiation.

    PubMed

    Zheng, Ke; Chen, Ying; Huang, Wenwen; Lin, Yinan; Kaplan, David L; Fan, Yimin

    2016-06-15

    To produce biocompatible, mechanically robust, and conductive materials for bone tissue engineering, chemical oxidation using sodium hyprochlorite (NaClO) was utilized to introduce carboxyl groups onto silk fibroin (SF). A final carboxyl content of 1.09 mM/g SF was obtained, corresponding to ∼47% of the primary hydroxymethyl groups on the silk. Interestingly, both infrared (IR) spectroscopy and circular dichroism (CD) spectra demonstrated that the resulting oxidized silk (OxSF) self-assembled into β-sheet structures under aqueous conditions and this contributed to the mechanical properties of the as-prepared silk-based scaffolds and the mineralized OxSF scaffolds (M-OxSF). The OxSF scaffolds had a compressive modulus of 211 ± 75 KPa in the hydrated state, 10 times higher than that of the SF scaffolds, and the modulus of the M-OxSF scaffolds was increased to 758 ± 189 KPa. Human bone marrow-derived mesenchymal stem cells (hMSCs) grown on the scaffolds during osteogenesis showed that the OxSF scaffolds supported the proliferation and differentiation of hMSCs in vitro. PMID:27177120

  12. Vanadate inhibits dexamethasone-induced apoptosis of rat bone marrow-derived mesenchymal stem cells.

    PubMed

    Fan, Qie; Zhan, Xinli; Li, Xiaofeng; Zhao, Jinmin; Chen, Yueping

    2015-01-01

    Apoptosis of bone marrow-derived mesenchymal stem cells (BM-MSCs) has been shown to contribute to the development of osteoporosis, which is often the result of long-term use of glucocorticoid drugs such as dexamethasone (Dex). However, it remains unknown whether Dex induces apoptosis of BM-MSCs, and whether a chemical agent like vanadate can block such effects. To investigate these two issues, we isolated BM-MSCs from SD rats and treated the cells with different doses of Dex. We found that Dex induced apoptosis in dose- and time-dependent manners. Pretreating BM-MSCs with vanadate prevented Dex-induced apoptosis. Furthermore, we found that expression of caspases (3, 8, and 9) increased in Dex-treated BM-MSC and was attenuated by vanadate pretreatment. These results not only demonstrate the role of vanadate in the inhibition of Dex-induced apoptosis of BM-MSCs, but also reveal the therapeutic potential of vanadate in glucocorticoid-mediated osteoporosis. PMID:25887871

  13. Increased Differentiation Capacity of Bone Marrow-Derived Mesenchymal Stem Cells in Aquaporin-5 Deficiency

    PubMed Central

    Yi, Fei; Khan, Muhammad; Gao, Hongwen; Hao, Feng; Sun, Meiyan; Zhong, Lili; Lu, Changzheng; Feng, Xuechao

    2012-01-01

    Mesenchymal stem cells (MSCs) are adult stem cells with a self-renewal and multipotent capability and express extensively in multitudinous tissues. We found that water channel aquaporin-5 (AQP5) is expressed in bone marrow-derived MSCs (BMMSCs) in the plasma membrane pattern. BMMSCs from AQP5−/− mice showed significantly lower plasma membrane water permeability than those from AQP5+/+ mice. In characterizing the cultured BMMSCs from AQP5−/− and AQP5+/+ mice, we found no obvious differences in morphology and proliferation between the 2 genotypes. However, the multiple differentiation capacity was significantly higher in AQP5−/− than AQP5+/+ BMMSCs as revealed by representative staining by Oil Red O (adipogenesis); Alizarin Red S and alkaline phosphatase (ALP; osteogenesis); and type II collagen and Safranin O (chondrogenesis) after directional induction. Relative mRNA expression levels of 3 lineage differentiation markers, including PPARγ2, C/EBPα, adipsin, collagen 1a, osteopontin, ALP, collagen 11a, collagen 2a, and aggrecan, were significantly higher in AQP5−/− -differentiating BMMSCs, supporting an increased differentiation capacity of AQP5−/− BMMSCs. Furthermore, a bone-healing process was accelerated in AQP5−/− mice in a drill-hole injury model. Mechanistic studies indicated a significantly lower apoptosis rate in AQP5−/− than AQP5+/+ BMMSCs. Apoptosis inhibitor Z-VAD-FMK increased the differentiation capacity to a greater extent in AQP5+/+ than AQP5−/− BMMSCs. We conclude that AQP5-mediated high plasma membrane water permeability enhances the apoptosis rate of differentiating BMMSCs, thus decreasing their differentiation capacity. These data implicate AQP5 as a novel determinant of differentiation of BMMSCs and therefore a new molecular target for regulating differentiation of BMMSCs during tissue repair and regeneration. PMID:22420587

  14. The effect of rat bone marrow derived mesenchymal stem cells transplantation for restoration of olfactory disorder.

    PubMed

    Jo, Hyogyeong; Jung, Minyoung; Seo, Dong Jin; Park, Dong Joon

    2015-11-13

    The purpose of the study was to investigate the effect of bone marrow-derived mesenchymal stem cells (BMSCs) transplantation on olfactory epithelium (OE) of morphologic and functional restoration following neural Sensorineural Disorder in rats. Except the Normal group, twenty-one rats underwent Triton X-100 (TX-100) irrigation to induce degeneration of OE, and then BMSCs and PBS were treated from the both medial canthus to the rear part of the both nasal cavity into the experimental group and then were observed for restoration according to time point. At two and four weeks after transplantation with BMSCs, restoration of OE was observed with olfactory marker protein (OMP) and behavioral test. And we observed the expression of OMP, nerve growth factor (NGF) and brain-derived neurotrophic factor (BDNF). After TX-100 irrigation, the OE almost disappeared in 3 days. At four weeks after transplantation with BMSCs, the thickness and cellular composition of OE was considerably restored to normal group and expression of OMP was markedly increased when compared with PBS group and reduced the searching time in the behavioral test. Furthermore at two weeks after treatment with BMSCs, expression of NGF and BDNF was greatly increased when compared with PBS group. However at four weeks after treatment with BMSCs, expression of NGF and BDNF was slightly decreased. Our results suggest the BMSCs transplantation affect restoration of OE and olfaction, most likely via regulation of the neurotrophic factor expression, especially the expression of NGF and BDNF and has a possibility of a new therapeutic strategy for the treatment of olfactory disorder caused by the degeneration of OE. PMID:26427869

  15. CD44 promotes the migration of bone marrow-derived mesenchymal stem cells toward glioma

    PubMed Central

    YIN, QIANG; ZHOU, YANG-YANG; WANG, PENG; MA, LI; LI, PENG; WANG, XIAO-GUANG; SHE, CHUN-HUA; LI, WEN-LIANG

    2016-01-01

    Previous in vivo and in vitro studies have shown that human mesenchymal stem cells (MSCs) exhibit tropism for gliomas. However, the mechanism underlying this directed migration remains unclear. The aim of the present study was to investigate the possible mechanism underlying platelet-derived growth factor-BB (PDGF-BB)-induced chemotactic migration of bone marrow-derived MSCs (BMSCs) toward glioma. Rat glioma C6 cell-conditioned medium was utilized to evaluate the chemotactic response of BMSCs toward glioma using an in vitro migration assay. Recombinant rat PDGF-BB was added to C6 cell-conditioned medium to assess its effect on the tropism of BMSCs. The effect of PDGF-BB on the expression levels of cluster of differentiation (CD)44 in BMSCs was evaluated by reverse transcription-polymerase chain reaction (RT-PCR) and immunofluorescence assays. The results revealed that chemotactic migration was induced in BMSCs by rat glioma C6 cell-conditioned medium, which was enhanced by PDGF-BB treatment in a dose-dependent manner. Furthermore, RT-PCR and immunofluorescence assays showed that CD44 expression was upregulated in BMSCs following treatment with 40 ng/ml PDGF-BB for 12 h. Additionally, 3-h pretreatment with the anti-CD44 neutralizing antibody OX-50 was observed to attenuate the tropism of BMSCs toward glioma in the presence or absence of PDGF-BB. The results of the present study indicate that CD44 mediates the tropism of BMSCs toward glioma, and PDGF-BB promotes the migration of BMSCs toward glioma via the upregulation of CD44 expression in BMSCs. These findings suggest CD44 inhibition may be a potential therapeutic target for the treatment of glioma. PMID:27073479

  16. Treatment of radiation-induced acute intestinal injury with bone marrow-derived mesenchymal stem cells

    PubMed Central

    ZHENG, KAI; WU, WEIZHEN; YANG, SHUNLIANG; HUANG, LIANGHU; CHEN, JIN; GONG, CHUNGUI; FU, ZHICHAO; LIN, RUOFEI; TAN, JIANMING

    2016-01-01

    The aim of the present study was to investigate the ability of bone marrow-derived mesenchymal stem cells (BMSCs) to repair radiation-induced acute intestinal injury, and to elucidate the underlying repair mechanism. Male Sprague-Dawley rats were subjected to whole abdominal irradiation using a single medical linear accelerator (12 Gy) and randomly assigned to two groups. Rats in the BMSC-treated group were injected with 1 ml BMSC suspension (2×106 cells/ml) via the tail vein, while the control group rats were injected with normal saline. BMSCs were identified by detecting the expression of CD29, CD90, CD34 and CD45 using flow cytometry. The expression of the cytokines stromal cell-derived factor 1 (SDF-1), prostaglandin E2 (PGE2) and interleukin (IL)-2 was detected using immunohistochemical techniques. Plasma citrulline concentrations were evaluated using an ELISA kit. Rat general conditions, including body weight, and changes in cellular morphology were also recorded. The results suggested that BMSCs exerted a protective effect on radiation-induced acute intestinal injury in rats. The histological damage was rapidly repaired in the BMSC-treated group. In addition, the BMSC-treated group showed significantly reduced radiation injury scores (P<0.01), mildly reduced body weight and plasma citrulline levels, significantly more rapid recovery (P<0.01), significantly reduced expression of the cytokines PGE2 and IL-2 (P<0.05) and significantly increased SDF-1 expression (P<0.01) compared with the control group. In summary, the present results indicate that BMSCs are able to effectively reduce inflammation and promote repair of the structure and function of intestinal tissues damaged by radiation exposure, suggesting that they may provide a promising therapeutic agent. PMID:27284330

  17. Beneficial Effects of Autologous Bone Marrow-Derived Mesenchymal Stem Cells in Naturally Occurring Tendinopathy

    PubMed Central

    Smith, Roger Kenneth Whealands; Werling, Natalie Jayne; Dakin, Stephanie Georgina; Alam, Rafiqul; Goodship, Allen E.; Dudhia, Jayesh

    2013-01-01

    Tendon injuries are a common age-related degenerative condition where current treatment strategies fail to restore functionality and normal quality of life. This disease also occurs naturally in horses, with many similarities to human tendinopathy making it an ideal large animal model for human disease. Regenerative approaches are increasingly used to improve outcome involving mesenchymal stem cells (MSCs), supported by clinical data where injection of autologous bone marrow derived MSCs (BM-MSCs) suspended in marrow supernatant into injured tendons has halved the re-injury rate in racehorses. We hypothesized that stem cell therapy induces a matrix more closely resembling normal tendon than the fibrous scar tissue formed by natural repair. Twelve horses with career-ending naturally-occurring superficial digital flexor tendon injury were allocated randomly to treatment and control groups. 1X107 autologous BM-MSCs suspended in 2 ml of marrow supernatant were implanted into the damaged tendon of the treated group. The control group received the same volume of saline. Following a 6 month exercise programme horses were euthanized and tendons assessed for structural stiffness by non-destructive mechanical testing and for morphological and molecular composition. BM-MSC treated tendons exhibited statistically significant improvements in key parameters compared to saline-injected control tendons towards that of normal tendons and those in the contralateral limbs. Specifically, treated tendons had lower structural stiffness (p<0.05) although no significant difference in calculated modulus of elasticity, lower (improved) histological scoring of organisation (p<0.003) and crimp pattern (p<0.05), lower cellularity (p<0.007), DNA content (p<0.05), vascularity (p<0.03), water content (p<0.05), GAG content (p<0.05), and MMP-13 activity (p<0.02). Treatment with autologous MSCs in marrow supernatant therefore provides significant benefits compared to untreated tendon repair in

  18. Beneficial effects of autologous bone marrow-derived mesenchymal stem cells in naturally occurring tendinopathy.

    PubMed

    Smith, Roger Kenneth Whealands; Werling, Natalie Jayne; Dakin, Stephanie Georgina; Alam, Rafiqul; Goodship, Allen E; Dudhia, Jayesh

    2013-01-01

    Tendon injuries are a common age-related degenerative condition where current treatment strategies fail to restore functionality and normal quality of life. This disease also occurs naturally in horses, with many similarities to human tendinopathy making it an ideal large animal model for human disease. Regenerative approaches are increasingly used to improve outcome involving mesenchymal stem cells (MSCs), supported by clinical data where injection of autologous bone marrow derived MSCs (BM-MSCs) suspended in marrow supernatant into injured tendons has halved the re-injury rate in racehorses. We hypothesized that stem cell therapy induces a matrix more closely resembling normal tendon than the fibrous scar tissue formed by natural repair. Twelve horses with career-ending naturally-occurring superficial digital flexor tendon injury were allocated randomly to treatment and control groups. 1X10(7) autologous BM-MSCs suspended in 2 ml of marrow supernatant were implanted into the damaged tendon of the treated group. The control group received the same volume of saline. Following a 6 month exercise programme horses were euthanized and tendons assessed for structural stiffness by non-destructive mechanical testing and for morphological and molecular composition. BM-MSC treated tendons exhibited statistically significant improvements in key parameters compared to saline-injected control tendons towards that of normal tendons and those in the contralateral limbs. Specifically, treated tendons had lower structural stiffness (p<0.05) although no significant difference in calculated modulus of elasticity, lower (improved) histological scoring of organisation (p<0.003) and crimp pattern (p<0.05), lower cellularity (p<0.007), DNA content (p<0.05), vascularity (p<0.03), water content (p<0.05), GAG content (p<0.05), and MMP-13 activity (p<0.02). Treatment with autologous MSCs in marrow supernatant therefore provides significant benefits compared to untreated tendon repair in

  19. Bone marrow-derived mesenchymal stem cells promote growth and angiogenesis of breast and prostate tumors

    PubMed Central

    2013-01-01

    Introduction Mesenchymal stem cells (MSCs) are known to migrate to tumor tissues. This behavior of MSCs has been exploited as a tumor-targeting strategy for cell-based cancer therapy. However, the effects of MSCs on tumor growth are controversial. This study was designed to determine the effect of MSCs on the growth of breast and prostate tumors. Methods Bone marrow-derived MSCs (BM-MSCs) were isolated and characterized. Effects of BM-MSCs on tumor cell proliferation were analyzed in a co-culture system with mouse breast cancer cell 4T1 or human prostate cancer cell DU145. Tumor cells were injected into nude mice subcutaneously either alone or coupled with BM-MSCs. The expression of cell proliferation and angiogenesis-related proteins in tumor tissues were immunofluorescence analyzed. The angiogenic effect of BM-MSCs was detected using a tube formation assay. The effects of the crosstalk between tumor cells and BM-MSCs on expression of angiogenesis related markers were examined by immunofluorescence and real-time PCR. Results Both co-culturing with mice BM-MSCs (mBM-MSCs) and treatment with mBM-MSC-conditioned medium enhanced the growth of 4T1 cells. Co-injection of 4T1 cells and mBM-MSCs into nude mice led to increased tumor size compared with injection of 4T1 cells alone. Similar experiments using DU145 cells and human BM-MSCs (hBM-MSCs) instead of 4T1 cells and mBM-MSCs obtained consistent results. Compared with tumors induced by injection of tumor cells alone, the blood vessel area was greater in tumors from co-injection of tumor cells with BM-MSCs, which correlated with decreased central tumor necrosis and increased tumor cell proliferation. Furthermore, both conditioned medium from hBM-MSCs alone and co-cultures of hBM-MSCs with DU145 cells were able to promote tube formation ability of human umbilical vein endothelial cells. When hBM-MSCs are exposed to the DU145 cell environment, the expression of markers associated with neovascularization (macrophage

  20. Small-diameter human vessel wall engineered from bone marrow-derived mesenchymal stem cells (hMSCs)

    PubMed Central

    Gong, Zhaodi; Niklason, Laura E.

    2008-01-01

    Using biodegradable scaffold and a biomimetic perfusion system, our lab has successfully engineered small-diameter vessel grafts using endothelial cells (ECs) and smooth muscle cells (SMCs) obtained from vessels in various species. However, translating this technique into humans has presented tremendous obstacles due to species and age differences. SMCs from elderly persons have limited proliferative capacity and a reduction in collagen production, which impair the mechanical strength of engineered vessels. As an alternative cell source, adult human bone marrow-derived mesenchymal stem cells (hMSCs) were studied for their ability to differentiate into SMCs in culture plates as well as in a bioreactor system. In the former setting, immunofluorescence staining showed that MSCs, after induction for 14 days, expressed smooth muscle α-actin (SMA) and calponin, early and mid-SMC phenotypic markers, respectively. In the latter setting, vessel walls were constructed with MSC-derived SMCs. Various factors (i.e., matrix proteins, soluble factors, and cyclic strain) in the engineering system were further investigated for their effects on hMSC cell proliferation and differentiation into SMCs. Based on a screening of multiple factors, the engineering system was optimized by dividing the vessel culture into proliferation and differentiation phases. The vessel walls engineered under the optimized conditions were examined histologically and molecularly, and found to be substantially similar to native vessels. In conclusion, bone marrow-derived hMSCs can serve as a new cell source of SMCs in vessel engineering. Optimization of the culture conditions to drive SMC differentiation and matrix production significantly improved the quality of the hMSC-derived engineered vessel wall.—Gong, Z., Niklason, L. E. Small-diameter human vessel wall engineered from bone marrow-derived mesenchymal stem cells (hMSCs). PMID:18199698

  1. Detection of BCR/ABL Translocation in Bone Marrow Derived Mesenchymal Stem Cells in Egyptian CML Patients

    PubMed Central

    Gaafar, Taghrid Mohamed; Raafat, Inas Ismail; Aly, Azza Ahmed; Mohamed, Nagwa Abd EL-Ghaffar; Farid, Reem Jan; Saad, Neveen Ezzat; EL-Hawary, Rabab; Mostafaa, Naglaa; Ahmed, Mirhan Mohamed

    2015-01-01

    BACKGROUND: Chronic myeloid leukemia (CML) is a clonal myeloproliferative disorder of hematopoietic stem cells. It is characterized at the cytogenetic level by Philadelphia (ph) chromosome and at the molecular level by the BCR/ABL gene rearrangement. Bone marrow derived mesenchymal stem cells (MSCs) are pluripotent stem cells that can differentiate into several mesenchymal tissues. AIM: To observe the biological characteristics of MSCS from CML patients and to determine whether MSCs harbor the abnormal BCR/ABL translocation similar to CML bone marrow cells. SUBJECTS AND METHODS: Characterized MSCs were isolated from 12 newly diagnosed Philadelphia positive untreated CML patients. RESULTS: MSCs can be readily isolated from CML marrow and exhibit major expansion. Flow cytometry analysis revealed the typical MSC phenotype. Moreover; MSCs do not harbor the BCR/ABL translocation confirmed by karyotype and real time PCR. CONCLUSION: MSCs from CML patients express the typical MSC phenotype; and do not express the BCR/ABL gene. Since; MSCs are able to support engraftment of hematopoietic stem cells in stem cell transplantation(SCT) as well as suppress alloreactive T cells causing graft versus –host disease, this current study provides evidence that in a SCT setting of CML patients, autologous MSCs could be a source of stem cell support in future cell therapy applications.

  2. Retinal Electrophysiological Effects of Intravitreal Bone Marrow Derived Mesenchymal Stem Cells in Streptozotocin Induced Diabetic Rats

    PubMed Central

    Akkoç, Tolga; Eraslan, Muhsin; Şahin, Özlem; Özkara, Selvinaz; Vardar Aker, Fugen; Subaşı, Cansu; Karaöz, Erdal; Akkoç, Tunç

    2016-01-01

    Diabetic retinopathy is the most common cause of legal blindness in developed countries at middle age adults. In this study diabetes was induced by streptozotocin (STZ) in male Wistar albino rats. After 3 months of diabetes, rights eye were injected intravitreally with green fluorescein protein (GFP) labelled bone marrow derived stem cells (BMSC) and left eyes with balanced salt solution (Sham). Animals were grouped as Baseline (n = 51), Diabetic (n = 45), Diabetic+BMSC (n = 45 eyes), Diabetic+Sham (n = 45 eyes), Healthy+BMSC (n = 6 eyes), Healthy+Sham (n = 6 eyes). Immunohistology analysis showed an increased retinal gliosis in the Diabetic group, compared to Baseline group, which was assessed with GFAP and vimentin expression. In the immunofluorescence analysis BMSC were observed to integrate mostly into the inner retina and expressing GFP. Diabetic group had prominently lower oscillatory potential wave amplitudes than the Baseline group. Three weeks after intravitreal injection Diabetic+BMSC group had significantly better amplitudes than the Diabetic+Sham group. Taken together intravitreal BMSC were thought to improve visual function. PMID:27300133

  3. The role of growth factors in maintenance of stemness in bone marrow-derived mesenchymal stem cells

    SciTech Connect

    Eom, Young Woo; Oh, Ji-Eun; Lee, Jong In; Baik, Soon Koo; Rhee, Ki-Jong; Shin, Ha Cheol; Kim, Yong Man; Ahn, Chan Mug; Kong, Jee Hyun; Kim, Hyun Soo; Shim, Kwang Yong

    2014-02-28

    Highlights: • Expression of FGF-2, FGF-4, EGF, and HGF decreased during long-term culture of BMSCs. • Loss of growth factors induced autophagy, senescence and decrease of stemness. • FGF-2 increased proliferation potential via AKT and ERK activation in BMSCs. • FGF-2 suppressed LC3-II expression and down-regulated senescence of BMSCs. • HGF was important in maintenance of the differentiation potential of BMSCs. - Abstract: Mesenchymal stem cells (MSCs) are an active topic of research in regenerative medicine due to their ability to secrete a variety of growth factors and cytokines that promote healing of damaged tissues and organs. In addition, these secreted growth factors and cytokines have been shown to exert an autocrine effect by regulating MSC proliferation and differentiation. We found that expression of EGF, FGF-4 and HGF were down-regulated during serial passage of bone marrow-derived mesenchymal stem cells (BMSCs). Proliferation and differentiation potentials of BMSCs treated with these growth factors for 2 months were evaluated and compared to BMSCs treated with FGF-2, which increased proliferation of BMSCs. FGF-2 and -4 increased proliferation potentials at high levels, about 76- and 26-fold, respectively, for 2 months, while EGF and HGF increased proliferation of BMSCs by less than 2.8-fold. Interestingly, differentiation potential, especially adipogenesis, was maintained only by HGF treatment. Treatment with FGF-2 rapidly induced activation of AKT and later induced ERK activation. The basal level of phosphorylated ERK increased during serial passage of BMSCs treated with FGF-2. The expression of LC3-II, an autophagy marker, was gradually increased and the population of senescent cells was increased dramatically at passage 7 in non-treated controls. But FGF-2 and FGF-4 suppressed LC3-II expression and down-regulated senescent cells during long-term (i.e. 2 month) cultures. Taken together, depletion of growth factors during serial passage

  4. [Present status of research in bone marrow-derived mesenchymal stem cells for promoting the healing of diabetic ulcer].

    PubMed

    Zheng, Shu-Juan; Jia, Chi-Yu

    2012-08-01

    The delayed healing of diabetic ulcer has been haunting the surgeons and researchers for a long time. Although we have been researching and exploring the effective therapies for many years, the progress has been limited. Bone marrow-derived mesenchymal stem cells (BMSCs) have gradually won worldwide attention for their characteristics of differentiating into tissue repair cells and secreting multiple cytokines as well as growth factors. In recent years, the role of BMSCs in the treatment of diabetic ulcer has been drawing more and more attention. This article reviewed the advancement in the research of BMSCs in promoting the healing of diabetic ulcer. Through a discussion of the treatment of diabetic ulcer, the related research in BMSCs, as well as its role in diabetic ulcer treatment, the mechanism of BMSCs in promoting healing of diabetic ulcers is discussed. We expect through further research, unified criteria for the quality of BMSCs, application approach and dosage of BMSCs could be established. PMID:23248965

  5. Low level light promotes the proliferation and differentiation of bone marrow derived mesenchymal stem cells

    NASA Astrophysics Data System (ADS)

    Ahn, Jin-Chul; Rhee, Yun-Hee; Choi, Sun-Hyang; Kim, Dae Yu; Chung, Phil-Sang

    2015-03-01

    Low-level light irradiation (LLLI) reported to stimulate the proliferation or differentiation of a variety of cell types. However, very little is known about the effect of light therapy on stem cells. The aim of the present study was to evaluate the effect of LLLI on the molecular physiological change of human bone marrow derived stem cells (hBMSC) by wavelength (470, 630, 660, 740 and 850, 50mW). The laser diode was performed with different time interval (0, 7.5, 15, 30J/cm2, 50mW) on hBMSC. To determine the molecular physiological changes of cellular level of hBMSC, the clonogenic assay, ATP assay, reactive oxygen species (ROS) detection, mitochondria membrane potential (MMPΦ) staining and calcium efflux assay were assessed after irradiation. There was a difference between with and without irradiation on hBMSCs. An energy density up to 30 J/cm² improved the cell proliferation in comparison to the control group. Among these irradiated group, 630 and 660nm were significantly increased the cell proliferation. The cellular level of ATP and calcium influx was increased with energy dose-dependent in all LLLI groups. Meanwhile, ROS and MMPΦ were also increased after irradiation except 470nm. It can be concluded that LLLI using infrared light and an energy density up to 30 J/cm² has a positive stimulatory effect on the proliferation or differentiation of hBMSCs. Our results suggest that LLLI may influence to the mitochondrial membrane potential activity through ATP synthesis and increased cell metabolism which leads to cell proliferation and differentiation.

  6. Neural stem cells induce bone-marrow-derived mesenchymal stem cells to generate neural stem-like cells via juxtacrine and paracrine interactions

    SciTech Connect

    Alexanian, Arshak R. . E-mail: aalexan@mcw.edu

    2005-11-01

    Several recent reports suggest that there is far more plasticity that previously believed in the developmental potential of bone-marrow-derived cells (BMCs) that can be induced by extracellular developmental signals of other lineages whose nature is still largely unknown. In this study, we demonstrate that bone-marrow-derived mesenchymal stem cells (MSCs) co-cultured with mouse proliferating or fixed (by paraformaldehyde or methanol) neural stem cells (NSCs) generate neural stem cell-like cells with a higher expression of Sox-2 and nestin when grown in NS-A medium supplemented with N2, NSC conditioned medium (NSCcm) and bFGF. These neurally induced MSCs eventually differentiate into {beta}-III-tubulin and GFAP expressing cells with neuronal and glial morphology when grown an additional week in Neurobasal/B27 without bFGF. We conclude that juxtacrine interaction between NSCs and MSCs combined with soluble factors released from NSCs are important for generation of neural-like cells from bone-marrow-derived adherent MSCs.

  7. Repeated autologous bone marrow-derived mesenchymal stem cell injections improve radiation-induced proctitis in pigs.

    PubMed

    Linard, Christine; Busson, Elodie; Holler, Valerie; Strup-Perrot, Carine; Lacave-Lapalun, Jean-Victor; Lhomme, Bruno; Prat, Marie; Devauchelle, Patrick; Sabourin, Jean-Christophe; Simon, Jean-Marc; Bonneau, Michel; Lataillade, Jean-Jacques; Benderitter, Marc

    2013-11-01

    The management of proctitis in patients who have undergone very-high-dose conformal radiotherapy is extremely challenging. The fibrosis-necrosis, fistulae, and hemorrhage induced by pelvic overirradiation have an impact on morbidity. Augmenting tissue repair by the use of mesenchymal stem cells (MSCs) may be an important advance in treating radiation-induced toxicity. Using a preclinical pig model, we investigated the effect of autologous bone marrow-derived MSCs on high-dose radiation-induced proctitis. Irradiated pigs received repeated intravenous administrations of autologous bone marrow-derived MSCs. Immunostaining and real-time polymerase chain reaction analysis were used to assess the MSCs' effect on inflammation, extracellular matrix remodeling, and angiogenesis, in radiation-induced anorectal and colon damages. In humans, as in pigs, rectal overexposure induces mucosal damage (crypt depletion, macrophage infiltration, and fibrosis). In a pig model, repeated administrations of MSCs controlled systemic inflammation, reduced in situ both expression of inflammatory cytokines and macrophage recruitment, and augmented interleukin-10 expression in rectal mucosa. MSC injections limited radiation-induced fibrosis by reducing collagen deposition and expression of col1a2/col3a1 and transforming growth factor-β/connective tissue growth factor, and by modifying the matrix metalloproteinase/TIMP balance. In a pig model of proctitis, repeated injections of MSCs effectively reduced inflammation and fibrosis. This treatment represents a promising therapy for radiation-induced severe rectal damage. PMID:24068742

  8. Osteoanagenesis after transplantation of bone marrow-derived mesenchymal stem cells using polyvinylidene chloride film as a scaffold.

    PubMed

    Hamajima, Soichiro; Hayashi, Tatsuhide; Sato, Yamato; Sasaki, Keisuke; Kawai, Tatsushi

    2011-01-01

    The aim of this study was to develop a new cell transplantation technique for osteoanagenesis at bone defect sites. Polyvinylidene chloride (PVDC) film was evaluated because of its good biocompatibility and flexibility. We used this film as both a cell scaffold and a barrier membrane. Initially, the cell compatibility of the PVDC film for fibroblast-like cells and osteoblast-like cells was confirmed. Subsequently, bone marrow cells were obtained from rats and cultured on PVDC films in two kinds of medium. The PVDC films with bone marrow-derived mesenchymal stem cells (MSCs) were then applied to critical-sized bone defects in the calvarial bone of rats. After the transplantation, the surgical sites were dissected out and evaluated by soft X-ray radiography, micro-CT analysis and histological examinations. The bone marrow-derived MSC-transplanted rats showed greater bone regeneration than the control rats. Therefore, PVDC film is considered to be useful as a scaffold for bone regeneration. PMID:21946492

  9. Platelet-rich Plasma and Bone Marrow-derived Mesenchymal Stem Cells in Sports Medicine.

    PubMed

    Gobbi, Alberto; Fishman, Matthew

    2016-06-01

    Regenerative medicine is a fast-growing field in orthopedic sports medicine. Platelet-rich plasma contains multiple factors that have been shown to augment healing, thereby stimulating its use in multiple areas of acute and chronic injuries. Mesenchymal stem cells have pluripotent potential to form into tissues pertinent to orthopedics, such as cartilage and bone. As such, there is been a surge in the research directed toward steering those stem cells into a particular lineage as part of treatment for a variety of soft-tissue, cartilage, and bone pathologies. Overall, there are promising reports of their potential success, but there is a need for continued investigation into the efficacy of platelet-rich plasma and stem cells in sports medicine. PMID:27135289

  10. Characterization of Nestin, a Selective Marker for Bone Marrow Derived Mesenchymal Stem Cells

    PubMed Central

    Xie, Liang; Zeng, Xin; Hu, Jing; Chen, Qianming

    2015-01-01

    Mesenchymal stem cells (MSCs) are multipotent cells capable of differentiating into multiple cell lineages and contributing to tissue repair and regeneration. Characterization of the physiological function of MSCs has been largely hampered by lack of unique markers. Nestin, originally found in neuroepithelial stem cells, is an intermediate filament protein expressed in the early stages of development. Increasing studies have shown a particular association between Nestin and MSCs. Nestin could characterize a subset of bone marrow perivascular MSCs which contributed to bone development and closely contacted with hematopoietic stem cells (HSCs). Nestin expressing (Nes+) MSCs also play a role in the progression of various diseases. However, Nes+ cells were reported to participate in angiogenesis as MSCs or endothelial progenitor cells (EPCs) in several tissues and be a heterogeneous population comprising mesenchymal cells and endothelial cells in the developing bone marrow. In this review article, we will summarize the progress of the research on Nestin, particularly the function of Nes+ cells in bone marrow, and discuss the feasibility of using Nestin as a specific marker for MSCs. PMID:26236348

  11. Immortalization of bone marrow-derived porcine mesenchymal stem cells and their differentiation into cells expressing cardiac phenotypic markers.

    PubMed

    Moscoso, Isabel; Rodriguez-Barbosa, Jose-Ignacio; Barallobre-Barreiro, Javier; Anon, Patricia; Domenech, Nieves

    2012-08-01

    Mesenchymal stem cells (MSCs) may be among the first stem cell types to be utilized in the clinic for cell therapy, because of their ease of isolation and extensive differentiation potential. Using a porcine model, we have established several cell lines from MSCs to facilitate in vitro and in vivo studies of their potential use for cellular therapy. Bone marrow-derived primary MSCs were immortalized using the pRNS-1 plasmid. We obtained four stable immortalized cell lines that exhibited higher proliferative capacities than the parental cells. All four cell lines displayed a common phenotype similar to that of primary mesenchymal cells, characterized by constitutively high expressions of CD90, CD29, CD44, SLA I and CD46, while CD172a, CD106 and CD56 were less expressed. Remarkably, treatment with 5-azacytidine-stimulated porcine MSCs lines to differentiate into cells that were positive for cardiac phenotypic markers, such as α-actin, connexin-43, sarcomeric actin, serca-2 and, to a lesser extent, desmin and troponin-T. These porcine MSC lines will be valuable biological tools for developing strategies for ex vivo expansion and differentiation of MSCs into a specific lineage. PMID:22162515

  12. Development of a rapid culture method to induce adipocyte differentiation of human bone marrow-derived mesenchymal stem cells

    SciTech Connect

    Ninomiya, Yuichi; Sugahara-Yamashita, Yzumi; Nakachi, Yutaka; Tokuzawa, Yoshimi; Okazaki, Yasushi; Nishiyama, Masahiko

    2010-04-02

    Human mesenchymal stem cells (hMSCs) derived from bone marrow are multipotent stem cells that can regenerate mesenchymal tissues such as adipose, bone or muscle. It is thought that hMSCs can be utilized as a cell resource for tissue engineering and as human models to study cell differentiation mechanisms, such as adipogenesis, osteoblastogenesis and so on. Since it takes 2-3 weeks for hMSCs to differentiate into adipocytes using conventional culture methods, the development of methods to induce faster differentiation into adipocytes is required. In this study we optimized the culture conditions for adipocyte induction to achieve a shorter cultivation time for the induction of adipocyte differentiation in bone marrow-derived hMSCs. Briefly, we used a cocktail of dexamethasone, insulin, methylisobutylxanthine (DIM) plus a peroxisome proliferator-activated receptor {gamma} agonist, rosiglitazone (DIMRo) as a new adipogenic differentiation medium. We successfully shortened the period of cultivation to 7-8 days from 2-3 weeks. We also found that rosiglitazone alone was unable to induce adipocyte differentiation from hMSCs in vitro. However, rosiglitazone appears to enhance hMSC adipogenesis in the presence of other hormones and/or compounds, such as DIM. Furthermore, the inhibitory activity of TGF-{beta}1 on adipogenesis could be investigated using DIMRo-treated hMSCs. We conclude that our rapid new culture method is very useful in measuring the effect of molecules that affect adipogenesis in hMSCs.

  13. Human gingiva-derived mesenchymal stem cells are superior to bone marrow-derived mesenchymal stem cells for cell therapy in regenerative medicine

    SciTech Connect

    Tomar, Geetanjali B.; Srivastava, Rupesh K.; Gupta, Navita; Barhanpurkar, Amruta P.; Pote, Satish T.; Jhaveri, Hiral M.; Mishra, Gyan C.; Wani, Mohan R.

    2010-03-12

    Mesenchymal stem cells (MSCs) are capable of self-renewal and differentiation into multiple cell lineages. Presently, bone marrow is considered as a prime source of MSCs; however, there are some drawbacks and limitations in use of these MSCs for cell therapy. In this study, we demonstrate that human gingival tissue-derived MSCs have several advantages over bone marrow-derived MSCs. Gingival MSCs are easy to isolate, homogenous and proliferate faster than bone marrow MSCs without any growth factor. Importantly, gingival MSCs display stable morphology and do not loose MSC characteristic at higher passages. In addition, gingival MSCs maintain normal karyotype and telomerase activity in long-term cultures, and are not tumorigenic. Thus, we reveal that human gingiva is a better source of MSCs than bone marrow, and large number of functionally competent clinical grade MSCs can be generated in short duration for cell therapy in regenerative medicine and tissue engineering.

  14. Overexpression of FABP3 inhibits human bone marrow derived mesenchymal stem cell proliferation but enhances their survival in hypoxia

    SciTech Connect

    Wang, Suna Zhou, Yifu; Andreyev, Oleg; Hoyt, Robert F.; Singh, Avneesh; Hunt, Timothy; Horvath, Keith A.

    2014-04-15

    Studying the proliferative ability of human bone marrow derived mesenchymal stem cells in hypoxic conditions can help us achieve the effective regeneration of ischemic injured myocardium. Cardiac-type fatty acid binding protein (FABP3) is a specific biomarker of muscle and heart tissue injury. This protein is purported to be involved in early myocardial development, adult myocardial tissue repair and responsible for the modulation of cell growth and proliferation. We have investigated the role of FABP3 in human bone marrow derived mesenchymal stem cells under ischemic conditions. MSCs from 12 donors were cultured either in standard normoxic or modified hypoxic conditions, and the differential expression of FABP3 was tested by quantitative {sup RT}PCR and western blot. We also established stable FABP3 expression in MSCs and searched for variation in cellular proliferation and differentiation bioprocesses affected by hypoxic conditions. We identified: (1) the FABP3 differential expression pattern in the MSCs under hypoxic conditions; (2) over-expression of FABP3 inhibited the growth and proliferation of the MSCs; however, improved their survival in low oxygen environments; (3) the cell growth factors and positive cell cycle regulation genes, such as PCNA, APC, CCNB1, CCNB2 and CDC6 were all down-regulated; while the key negative cell cycle regulation genes TP53, BRCA1, CASP3 and CDKN1A were significantly up-regulated in the cells with FABP3 overexpression. Our data suggested that FABP3 was up-regulated under hypoxia; also negatively regulated the cell metabolic process and the mitotic cell cycle. Overexpression of FABP3 inhibited cell growth and proliferation via negative regulation of the cell cycle and down-regulation of cell growth factors, but enhances cell survival in hypoxic or ischemic conditions. - Highlights: • FABP3 expression pattern was studied in 12 human hypoxic-MSCs. • FABP3 mRNA and proteins are upregulated in the MSCs under hypoxic conditions.

  15. Micro-/Nano- sized hydroxyapatite directs differentiation of rat bone marrow derived mesenchymal stem cells towards an osteoblast lineage

    NASA Astrophysics Data System (ADS)

    Huang, Yan; Zhou, Gang; Zheng, Lisha; Liu, Haifeng; Niu, Xufeng; Fan, Yubo

    2012-03-01

    Regenerative medicine consisting of cells and materials provides a new way for the repair and regeneration of tissues and organs. Nano-biomaterials are highlighted due to their advantageous features compared with conventional micro-materials. The aim of this study is to investigate the effects of micro-/nano- sized hydroxyapatite (μ/n-HA) on the osteogenic differentiation of rat bone marrow derived mesenchymal stem cells (rBMSCs). μ/n-HA were prepared by a microwave synthesizer and precipitation method, respectively. Different sizes of μ/n-HA were characterized by IR, XRD, SEM, TEM and co-cultured with rBMSCs. It was shown that rBMSCs expressed higher levels of osteoblast-related markers by n-HA than μ-HA stimulation. The size of HA is an important factor for affecting the osteogenic differentiation of rBMSCs. This provides a new avenue for mechanistic studies of stem cell differentiation and a new approach to obtain more committed differentiated cells.

  16. Micro-/nano- sized hydroxyapatite directs differentiation of rat bone marrow derived mesenchymal stem cells towards an osteoblast lineage.

    PubMed

    Huang, Yan; Zhou, Gang; Zheng, Lisha; Liu, Haifeng; Niu, Xufeng; Fan, Yubo

    2012-04-01

    Regenerative medicine consisting of cells and materials provides a new way for the repair and regeneration of tissues and organs. Nano-biomaterials are highlighted due to their advantageous features compared with conventional micro-materials. The aim of this study is to investigate the effects of micro-/nano- sized hydroxyapatite (μ/n-HA) on the osteogenic differentiation of rat bone marrow derived mesenchymal stem cells (rBMSCs). μ/n-HA were prepared by a microwave synthesizer and precipitation method, respectively. Different sizes of μ/n-HA were characterized by IR, XRD, SEM, TEM and co-cultured with rBMSCs. It was shown that rBMSCs expressed higher levels of osteoblast-related markers by n-HA than μ-HA stimulation. The size of HA is an important factor for affecting the osteogenic differentiation of rBMSCs. This provides a new avenue for mechanistic studies of stem cell differentiation and a new approach to obtain more committed differentiated cells. PMID:22371072

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

    PubMed

    Park, Jin Seok; Yi, Tac-Ghee; Park, Jong-Min; Han, Young Min; Kim, Jun-Hyung; Shin, Dong-Hee; Tak, Seon Ji; Lee, Kyuheon; Lee, Youn Sook; Jeon, Myung-Shin; Hahm, Ki-Baik; Song, Sun U; Park, Seok Hee

    2015-11-01

    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. PMID:26566304

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

    PubMed Central

    Na, K; Yoo, H S; Zhang, Y X; Choi, M-S; Lee, K; Yi, T G; Song, S U; Jeon, M-S

    2014-01-01

    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. PMID:25032868

  19. Differentiation of Bone Marrow: Derived Mesenchymal Stem Cells into Hepatocyte-like Cells.

    PubMed

    Al Ghrbawy, Nesrien M; Afify, Reham Abdel Aleem Mohamed; Dyaa, Nehal; El Sayed, Asmaa A

    2016-09-01

    Cirrhosis is the end-stage liver fibrosis, whereby normal liver architecture is disrupted by fibrotic bands, parenchymal nodules and vascular distortion. Portal hypertension and hepatocyte dysfunction are the end results and give rise to major systemic complications and premature death. Mesenchymal stem cells (MSC) have the capacity of self-renew and to give rise to cells of various lineages, so MSC can be isolated from bone marrow (BM) and induced to differentiate into hepatocyte-like cells. MSC were induced to differentiate into hepatocyte-like cells by hepatotic growth factor (HGF) and fibroblast growth factor-4 (FGF-4). Differentiated cells were examined for the expression of hepatocyte-specific markers and hepatocyte functions. MSC were isolated. Flow cytometry analysis showed that they expressed the MSC-specific markers, reverse transcriptase-polymerase chain reaction (RT-PCR) demonstrated that MSC expressed the hepatocyte-specific marker cytokeratin 18 (CK-18) following hepatocyte induction. This study demonstrates that BM-derived-MSC can differentiate into functional hepatocyte-like cells following the induction of HGF and FGF-4. MSC can serve as a favorable cell source for tissue engineering in the treatment of liver disease. PMID:27429519

  20. Comparison of the osteogenic capacity of minipig and human bone marrow-derived mesenchymal stem cells.

    PubMed

    Heino, Terhi J; Alm, Jessica J; Moritz, Niko; Aro, Hannu T

    2012-07-01

    Minipigs are a recommended large animal model for preclinical testing of human orthopedic implants. Mesenchymal stem cells (MSCs) are the key repair cells in bone healing and implant osseointegration, but the osteogenic capacity of minipig MSCs is incompletely known. The aim of this study was to isolate and characterize minipig bone marrow (BM) and peripheral blood (PB) MSCs in comparison to human BM-MSCs. BM sample was aspirated from posterior iliac crest of five male Göttingen minipigs (age 15 ± 1 months). PB sample was drawn for isolation of circulating MSCs. MSCs were selected by plastic-adherence as originally described by Friedenstein. Cell morphology, colony formation, proliferation, surface marker expression, and differentiation were examined. Human BM-MSCs were isolated and cultured from adult fracture patients (n = 13, age 19-60 years) using identical techniques. MSCs were found in all minipig BM samples, but no circulating MSCs could be detected. Minipig BM-MSCs had similar morphology, proliferation, and colony formation capacities as human BM-MSCs. Unexpectedly, minipig BM-MSCs had a significantly lower ability than human BM-MSCs to form differentiated and functional osteoblasts. This observation emphasizes the need for species-specific optimization of MSC culture protocol before direct systematic comparison of MSCs between human and various preclinical large animal models can be made. PMID:22570220

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

    PubMed

    Na, K; Yoo, H S; Zhang, Y X; Choi, M-S; Lee, K; Yi, T G; Song, S U; Jeon, M-S

    2014-01-01

    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. PMID:25032868

  2. The canine epiphyseal-derived mesenchymal stem cells are comparable to bone marrow derived-mesenchymal stem cells

    PubMed Central

    CHANG, Ya-Pei; HONG, Hsuan-Ping; LEE, Yen-Hua; LIU, I-Hsuan

    2014-01-01

    Mesenchymal stem cells (MSCs) hold great potential in cell therapy and have attracted increasing interests in a wide range of biomedical sciences. However, the scarcity of MSCs and the prolonged isolation procedure limited the clinical application. To address these 2 issues, we developed a method to isolate MSCs from bone biopsy tissues of euthanized canine body donors. Compared to the traditional method to isolate MSCs from aspirated bone marrow (BMSCs), the isolation procedure for MSCs from harvested epiphyseal cancellous bone (EMSCs) was less time-consuming. The isolated EMSCs had similar plastic-adherence, tri-lineage differentiation and consistent surface marker profiles compared to BMSCs. We harvested BMSCs and EMSCs from 24 euthanized cases from clinics and 42 euthanized donors from a local shelter. The successful rate for EMSC isolation is significantly higher compared to BMSC isolation, while the other properties of the isolated MSCs including the clonogenicity, proliferative potentials and molecular phenotypes were not discernibly different between the MSCs established by the two methods. In conclusion, we demonstrated a new procedure to harvest MSCs by bone biopsy at the epiphyseal region. This method is less time consuming and more reliable, and the resulting MSCs are comparable to those harvested by bone marrow aspiration. The combination of the two methods can greatly improve the efficiency to harvest MSCs. PMID:25391394

  3. Bone Marrow-Derived Mesenchymal Stem Cells Enhance Angiogenesis via their α6β1 Integrin Receptor

    PubMed Central

    Carrion, Bita; Kong, Yen P.; Kaigler, Darnell; Putnam, Andrew J

    2013-01-01

    Bone marrow-derived mesenchymal stem cells (BMSCs) facilitate the angiogenic response of endothelial cells (ECs) within three-dimensional (3D) matrices in vivo and in engineered tissues in vitro in part through paracrine mediators and by acting as stabilizing pericytes. However, the molecular interactions between BMSCs and nascent tubules during the process of angiogenesis are not fully understood. In this study, we have used a tractable 3D co-culture model to explore the functional role of the α6β1 integrin adhesion receptor on BMSCs in sprouting angiogenesis. We report that knockdown of the α6 integrin subunit in BMSCs significantly reduces capillary sprouting, and causes their failure to associate with the nascent vessels. Furthermore, we demonstrate that the BMSCs with attenuated α6 integrin proliferate at a significantly lower rate relative to either control cells expressing non-targeting shRNA or wild type BMSCs; however, despite adding more cells to compensate for this deficit in proliferation, deficient sprouting persists. Collectively, our findings demonstrate that the α6 integrin subunit in BMSCs is important for their ability to stimulate vessel morphogenesis. This conclusion may have important implications in the optimization of cell-based strategies to promote angiogenesis. PMID:24056178

  4. The Effect of Quercetin on the Osteogenesic Differentiation and Angiogenic Factor Expression of Bone Marrow-Derived Mesenchymal Stem Cells

    PubMed Central

    Zhou, Yuning; Wu, Yuqiong; Jiang, Xinquan; Zhang, Xiuli; Xia, Lunguo; Lin, Kaili; Xu, Yuanjin

    2015-01-01

    Bone marrow-derived mesenchymal stem cells (BMSCs) are widely used in regenerative medicine in light of their ability to differentiate along the chondrogenic and osteogenic lineages. As a type of traditional Chinese medicine, quercetin has been preliminarily reported to promote osteogenic differentiation in osteoblasts. In the present study, the effects of quercetin on the proliferation, viability, cellular morphology, osteogenic differentiation and angiogenic factor secretion of rat BMSCs (rBMSCs) were examined by MTT assay, fluorescence activated cell sorter (FACS) analysis, real-time quantitative PCR (RT-PCR) analysis, alkaline phosphatase (ALP) activity and calcium deposition assays, and Enzyme-linked immunosorbent assay (ELISA). Moreover, whether mitogen-activated protein kinase (MAPK) signaling pathways were involved in these processes was also explored. The results showed that quercetin significantly enhanced the cell proliferation, osteogenic differentiation and angiogenic factor secretion of rBMSCs in a dose-dependent manner, with a concentration of 2 μM achieving the greatest stimulatory effect. Moreover, the activation of the extracellular signal-regulated protein kinases (ERK) and p38 pathways was observed in quercetin-treated rBMSCs. Furthermore, these induction effects could be repressed by either the ERK inhibitor PD98059 or the p38 inhibitor SB202190, respectively. These data indicated that quercetin could promote the proliferation, osteogenic differentiation and angiogenic factor secretion of rBMSCs in vitro, partially through the ERK and p38 signaling pathways. PMID:26053266

  5. Altered MicroRNA Expression Profile in Exosomes during Osteogenic Differentiation of Human Bone Marrow-Derived Mesenchymal Stem Cells

    PubMed Central

    Zhang, Shui-Jun; Zhao, Chen; Qiu, Bin-Song; Gu, Hai-Feng; Hong, Jian-Fei; Cao, Li; Chen, Yu; Xia, Bing; Bi, Qin; Wang, Ya-Ping

    2014-01-01

    The physiological role of microRNAs (miRNAs) in osteoblast differentiation remains elusive. Exosomal miRNAs isolated from human bone marrow-derived mesenchymal stem cells (BMSCs) culture were profiled using miRNA arrays containing probes for 894 human matured miRNAs. Seventy-nine miRNAs (∼8.84%) could be detected in exosomes isolated from BMSC culture supernatants when normalized to endogenous control genes RNU44. Among them, nine exosomal miRNAs were up regulated and 4 miRNAs were under regulated significantly (Relative fold>2, p<0.05) when compared with the values at 0 day with maximum changes at 1 to 7 days. Five miRNAs (miR-199b, miR-218, miR-148a, miR-135b, and miR-221) were further validated and differentially expressed in the individual exosomal samples from hBMSCs cultured at different time points. Bioinformatic analysis by DIANA-mirPath demonstrated that RNA degradation, mRNA surveillance pathway, Wnt signaling pathway, RNA transport were the most prominent pathways enriched in quantiles with differential exosomal miRNA patterns related to osteogenic differentiation. These data demonstrated exosomal miRNA is a regulator of osteoblast differentiation. PMID:25503309

  6. Effects of matrix metalloproteinase-1 on the myogenic differentiation of bone marrow-derived mesenchymal stem cells in vitro

    SciTech Connect

    Zheng, Zhenyang; Leng, Yan; Zhou, Chen; Ma, Zhenyu; Zhong, Zhigang; Shi, Xing-Ming; Zhang, Weixi

    2012-11-16

    Highlights: Black-Right-Pointing-Pointer MMP-1 is a member of the zinc-dependent endopeptidase family. Black-Right-Pointing-Pointer MMP-1 has no cytotoxic effects on BMSCs. Black-Right-Pointing-Pointer MMP-1 can promote the myogenic differentiation of BMSCs. Black-Right-Pointing-Pointer MyoD and desmin were chosen as myogenic markers in this study. -- Abstract: Matrix metalloproteinase-1 (MMP-1) is a member of the family of zinc-dependent endopeptidases that are capable of degrading extracellular matrix (ECM) and certain non-matrix proteins. It has been shown that MMP-1 can enhance muscle regeneration by improving the differentiation and migration of myoblasts. However, it is still not known whether MMP-1 can promote the myogenesis of bone marrow-derived mesenchymal stem cells (BMSCs). To address this question, we isolated BMSCs from C57BL/6J mice and investigated the effects of MMP-1 on their proliferation and myogenic differentiation. Our results showed that MMP-1 treatment, which had no cytotoxic effects on BMSCs, increased the mRNA and protein levels of MyoD and desmin in a dose-dependent manner, indicating that MMP-1 promoted myogenic differentiation of BMSCs in vitro. These results suggest that BMSCs may have a therapeutic potential for treating muscular disorders.

  7. Effect of bone marrow derived mesenchymal stem cells on lung pathology and inflammation in ovalbumin-induced asthma in mouse

    PubMed Central

    Mohammadian, Maryam; Boskabady, Mohammad Hosein; Kashani, Iraj Ragerdi; Jahromi, Gila Pirzad; Omidi, Amene; Nejad, Amir Kavian; Khamse, Safoura; Sadeghipour, Hamid Reza

    2016-01-01

    Objective(s): Bone marrow-derived mesenchymal stem cells (BMSCs) have attracted significant interest to treat asthma and its complication. In this study, the effects of BMSCs on lung pathology and inflammation in an ovalbumin-induced asthma model in mouse were examined. Materials and Methods: BALB/c mice were divided into three groups: control group (animals were not sensitized), asthma group (animals were sensitized by ovalbumin), asthma+BMSC group (animals were sensitized by ovalbumin and treated with BMSCs). BMSCs were isolated and characterized and then labeled with Bromodeoxyuridine (BrdU). After that the cells transferred into asthmatic mice. Histopathological changes of the airways, BMSCs migration and total and differential white blood cell (WBC) count in bronchoalveolar lavage (BAL) fluid were evaluated. Results: A large number of BrdU-BMSCs were found in the lungs of mice treated with BMSCs. The histopathological changes, BAL total WBC counts and the percentage of neutrophils and eosinophils were increased in asthma group compared to the control group. Treatment with BMSCs significantly decreased airway pathological indices, inflammatory cell infiltration, and also goblet cell hyperplasia. Conclusion: The results of this study revealed that BMSCs therapy significantly suppressed the lung pathology and inflammation in the ovalbumin induced asthma model in mouse. PMID:27096065

  8. Regulation of Tenomodulin Expression Via Wnt/β-catenin Signaling in Equine Bone Marrow-derived Mesenchymal Stem Cells.

    PubMed

    Miyabara, Shihori; Yuda, Yohei; Kasashima, Yoshinori; Kuwano, Atsutoshi; Arai, Katsuhiko

    2014-01-01

    Tenomodulin has been recognized as a biomarker for tendon differentiation, and its gene expression is regulated by several transcription factors including Scleraxis and Mohawk. In this study, we found a novel regulatory mechanism of tenomodulin expression. Equine bone marrow-derived mesenchymal stem cells (BMSCs) in monolayer culture showed a low mRNA level of tenomodulin in comparison with the level in the tendon. When cultured in collagen gel containing a glycogen synthase kinase-3 (GSK-3) inhibitor (BIO), expression of tenomodulin in BMSCs increased up to the level in the tendon. Participation of GSK-3 in its gene expression was further demonstrated by a gene silencing experiment with small interference RNA corresponding to GSK-3, suggesting that Wnt/β-catenin signaling mediated expression of tenomodulin. These results were confirmed by nuclear translocation of β-catenin in BIO-treated BMSCs cultured in collagen gel. Under this culture condition, expression of tenomodulin-related transcription factors including Scleraxis and Mohawk was not affected, suggesting that Wnt/β-catenin signaling was independent from these transcription factors. Additionally, BIO strongly enhanced expression of type XIV collagen in collagen-embedded BMSCs up to the level in the tendon, and other tendon-related extracellular matrix components such as decorin and fibromodulin were also upregulated. Taken together, these results indicated that activation of Wnt/β-catenin signaling could induce differentiation of BMSCs into tenomodulin-expressing tendon cells in collagen gel. PMID:24834008

  9. Inhibition of AQP1 Hampers Osteosarcoma and Hepatocellular Carcinoma Progression Mediated by Bone Marrow-Derived Mesenchymal Stem Cells.

    PubMed

    Pelagalli, Alessandra; Nardelli, Anna; Fontanella, Raffaela; Zannetti, Antonella

    2016-01-01

    The complex cross-talk between tumor cells and their surrounding stromal environment plays a key role in the pathogenesis of cancer. Among several cell types that constitute the tumor stroma, bone marrow-derived mesenchymal stem cells (BM-MSCs) selectively migrate toward the tumor microenvironment and contribute to the active formation of tumor-associated stroma. Therefore, here we elucidate the involvement of BM-MSCs to promote osteosarcoma (OS) and hepatocellular carcinoma (HCC) cells migration and invasion and deepening the role of specific pathways. We analyzed the function of aquaporin 1 (AQP1), a water channel known to promote metastasis and neoangiogenes. AQP1 protein levels were analyzed in OS (U2OS) and HCC (SNU-398) cells exposed to conditioned medium from BM-MSCs. Tumor cell migration and invasion in response to BM-MSC conditioned medium were evaluated through a wound healing assay and Boyden chamber, respectively. The results showed that the AQP1 level was increased in both tumor cell lines after treatment with BM-MSC conditioned medium. Moreover, BM-MSCs-mediated tumor cell migration and invasion were hampered after treatment with AQP1 inhibitor. These data suggest that the recruitment of human BM-MSCs into the tumor microenvironment might cause OS and HCC cell migration and invasion through involvement of AQP1. PMID:27409610

  10. Inhibition of AQP1 Hampers Osteosarcoma and Hepatocellular Carcinoma Progression Mediated by Bone Marrow-Derived Mesenchymal Stem Cells

    PubMed Central

    Pelagalli, Alessandra; Nardelli, Anna; Fontanella, Raffaela; Zannetti, Antonella

    2016-01-01

    The complex cross-talk between tumor cells and their surrounding stromal environment plays a key role in the pathogenesis of cancer. Among several cell types that constitute the tumor stroma, bone marrow-derived mesenchymal stem cells (BM-MSCs) selectively migrate toward the tumor microenvironment and contribute to the active formation of tumor-associated stroma. Therefore, here we elucidate the involvement of BM-MSCs to promote osteosarcoma (OS) and hepatocellular carcinoma (HCC) cells migration and invasion and deepening the role of specific pathways. We analyzed the function of aquaporin 1 (AQP1), a water channel known to promote metastasis and neoangiogenes. AQP1 protein levels were analyzed in OS (U2OS) and HCC (SNU-398) cells exposed to conditioned medium from BM-MSCs. Tumor cell migration and invasion in response to BM-MSC conditioned medium were evaluated through a wound healing assay and Boyden chamber, respectively. The results showed that the AQP1 level was increased in both tumor cell lines after treatment with BM-MSC conditioned medium. Moreover, BM-MSCs-mediated tumor cell migration and invasion were hampered after treatment with AQP1 inhibitor. These data suggest that the recruitment of human BM-MSCs into the tumor microenvironment might cause OS and HCC cell migration and invasion through involvement of AQP1. PMID:27409610

  11. The Effect of Quercetin on the Osteogenesic Differentiation and Angiogenic Factor Expression of Bone Marrow-Derived Mesenchymal Stem Cells.

    PubMed

    Zhou, Yuning; Wu, Yuqiong; Jiang, Xinquan; Zhang, Xiuli; Xia, Lunguo; Lin, Kaili; Xu, Yuanjin

    2015-01-01

    Bone marrow-derived mesenchymal stem cells (BMSCs) are widely used in regenerative medicine in light of their ability to differentiate along the chondrogenic and osteogenic lineages. As a type of traditional Chinese medicine, quercetin has been preliminarily reported to promote osteogenic differentiation in osteoblasts. In the present study, the effects of quercetin on the proliferation, viability, cellular morphology, osteogenic differentiation and angiogenic factor secretion of rat BMSCs (rBMSCs) were examined by MTT assay, fluorescence activated cell sorter (FACS) analysis, real-time quantitative PCR (RT-PCR) analysis, alkaline phosphatase (ALP) activity and calcium deposition assays, and Enzyme-linked immunosorbent assay (ELISA). Moreover, whether mitogen-activated protein kinase (MAPK) signaling pathways were involved in these processes was also explored. The results showed that quercetin significantly enhanced the cell proliferation, osteogenic differentiation and angiogenic factor secretion of rBMSCs in a dose-dependent manner, with a concentration of 2 μM achieving the greatest stimulatory effect. Moreover, the activation of the extracellular signal-regulated protein kinases (ERK) and p38 pathways was observed in quercetin-treated rBMSCs. Furthermore, these induction effects could be repressed by either the ERK inhibitor PD98059 or the p38 inhibitor SB202190, respectively. These data indicated that quercetin could promote the proliferation, osteogenic differentiation and angiogenic factor secretion of rBMSCs in vitro, partially through the ERK and p38 signaling pathways. PMID:26053266

  12. Homing and restorative effects of bone marrow-derived mesenchymal stem cells on cisplatin injured ovaries in rats.

    PubMed

    Liu, Jiabin; Zhang, Haiying; Zhang, Yun; Li, Nan; Wen, Yuku; Cao, Fanglei; Ai, Hao; Xue, Xiaoou

    2014-12-31

    Premature ovarian failure (POF) is a long-term adverse effect of chemotherapy treatment. However, current available treatment regimens are not optimal. Emerging evidence suggests that bone marrow-derived mesenchymal stem cells (BMSCs) could restore the structure and function of injured tissues, but the homing and restorative effects of BMSCs on chemotherapy injured ovaries are still not clear. In this study, we found that granulosa cell (GC) apoptosis induced by cisplatin was reduced when BMSCs were migrated to granulosa cells (GCs) in vitro. Chemotherapy-induced POF was induced by intraperitoneal injection of cisplatin in rats. BMSCs labeled with enhanced green fluorescent protein (EGFP) were injected into the rats via the tail vein to investigate the homing and distribution of BMSCs in vivo. The number of BMSCs in the ovarian hilum and medulla was greater than in the cortex, but no BMSCs were found in the follicles and corpus lutea. In addition, the BMSCs treatment group's antral follicle count and estradiol levels increased after 30 days, compared with the POF group. Hence, our study demonstrates that intravenously delivered BMSCs can home to the ovaries, and restore its structure and function in POF model rats. PMID:25410907

  13. Attachment and growth of human bone marrow derived mesenchymal stem cells on regenerated antheraea pernyi silk fibroin films.

    PubMed

    Luan, Xi-Ying; Wang, Yong; Duan, Xiang; Duan, Qiao-Yan; Li, Ming-Zhong; Lu, Shen-Zhou; Zhang, Huan-Xiang; Zhang, Xue-Guang

    2006-12-01

    Silk fibroin of the silkworm Bombyx mori has been studied extensively, while the research on Antheraea pernyi silk fibroin (A. pernyi SF) in biomaterials is only at an early stage. In this study, the attachment, morphology, growth and phenotype of human bone marrow derived mesenchymal stem cells (hBMSCs) cultured on the regenerated A. pernyi SF films were studied in vitro. The results indicated that the attachment of hBMSCs on the regenerated A. pernyi SF films was almost the same as that on the collagen films. MTT and cell counting analyses demonstrated that the growth of hBMSCs on the regenerated A. pernyi SF films was better than that on controls. Moreover, electron scanning microscopy and fluorescence-activated cell sorting assays showed that the regenerated A. pernyi SF supported hBMSCs growth and functional maintenance compared with the controls. These data suggest that the regenerated A. pernyi SF, like Bombyx mori silk fibroin (B. mori SF) and collagen, can support hBMSCs attachment, growth and phenotypic maintenance, and has better biocompatibilities for hBMSCs in vitro culture. PMID:18458403

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

    PubMed

    Kim, Si-Na; Lee, Hyun-Joo; Jeon, Myung-Shin; Yi, TacGhee; Song, Sun U

    2015-10-01

    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. PMID:26557808

  15. NR2F2 regulates bone marrow-derived mesenchymal stem cell-promoted proliferation of Reh cells.

    PubMed

    Zhu, Ni; Wang, Huafang; Wei, Jieping; Wang, Binsheng; Shan, Wei; Lai, Xiaoyu; Zhao, Yanmin; Yu, Jian; Huang, He

    2016-08-01

    Bone marrow-derived mesenchymal stem cells (BM-MSCs) are pivotal components of the leukemic microenvironment. BM-MSCs have been previously reported to promote the proliferation of leukemic cells. To further understand the molecular mechanisms of BM-MSC-induced proliferation of leukemic cells, the present study co-cultured acute lymphoblastic leukemia (ALL) Reh cells with BM-MSCs. The current study used methods including shRNA, flow cytometry, MTT, reverse transcription-quantitative polymerase chain reaction, ELISA and western blotting. The data of the present study demonstrated that BM‑MSCs promote the proliferation of Reh cells and the NR2F2 mRNA and protein levels were elevated in BM‑MSCs following co‑culture. Additionally, it was demonstrated that shRNA knockdown of NR2F2 inhibited BM‑MSC‑induced proliferation of Reh cells. Furthermore, following downregulation of NR2F2, vascular endothelial growth factor A (VEGFA) secretion by BM‑MSCs was reduced. The present study demonstrated that NR2F2 mediates BM‑MSC‑induced proliferation of Reh cells, partially via regulation of VEGFA. Disrupting microenvironmental support by targeting NR2F2 may be a potential therapeutic strategy for ALL. PMID:27314877

  16. Induction of Spermatogenesis by Bone Marrow-derived Mesenchymal Stem Cells in Busulfan-induced Azoospermia in Hamster

    PubMed Central

    Tamadon, Amin; Mehrabani, Davood; Rahmanifar, Farhad; Jahromi, Alireza Raayat; Panahi, Mohadeseh; Zare, Shahrokh; Khodabandeh, Zahra; Jahromi, Iman Razeghian; Tanideh, Nader; Dianatpour, Mehdi; Ramzi, Mani; Koohi-Hoseinabadi, Omid

    2015-01-01

    Background Bone marrow-derived mesenchymal stem cells (BM-MSCs) have potential of differentiation and they secrete anti-inflammatory cytokines and growth factors which make them appropriate for cell therapy. Aim of the Work Were to evaluate the healing effect of BM-MSCs transplantation on germinal cells of busulfan-induced azoospermic hamsters. Material and Methods In the present experimental case control study, BM-MSCs were isolated from bone marrow of donor albino hamsters. Five mature male recipient hamsters received two doses of 10 mg/kg of busulfan with 21 days interval to stop endogenous spermatogenesis. After induction of azoospermia, right testis of hamsters was injected with 106 BM-MSCs via efferent duct and the left one remained as azoospermia control testis. Five normal mature hamsters were selected as normal intact control. After 35 days, testes and epididymis of three groups were removed for histological evaluation. Results Histomorphological analyses of BM-MSCs treated testes and epididymis showed the epithelial tissue of seminiferous tubules had normal morphology and spermatozoa were present in epididymis tubes. Spermatogenesis was observed in most cell-treated seminiferous tubules. The untreated seminiferous tubules were empty. Conclusion Transplanted BM-MSCs could successfully induce spermatogenesis in seminiferous tubules of azoospermic hamster. Therefore, BM-MSCs can be an attractive candidate in cell transplantation of azoospermia. PMID:26634062

  17. Effect of inflammatory environment on equine bone marrow derived mesenchymal stem cells immunogenicity and immunomodulatory properties.

    PubMed

    Barrachina, L; Remacha, A R; Romero, A; Vázquez, F J; Albareda, J; Prades, M; Ranera, B; Zaragoza, P; Martín-Burriel, I; Rodellar, C

    2016-03-01

    Mesenchymal stem cells (MSCs) are being investigated for the treatment of equine joint diseases because of their regenerative potential. Recently, the focus mainly has addressed to their immunomodulatory capacities. Inflammation plays a central role in joint pathologies, since the release of proinflammatory mediators to the synovial fluid (SF) leads to the activation of enzymatic degradation of the cartilage. MSCs can modulate the local immune environment through direct or paracrine interaction with immune cells, suppressing their proliferation and re-addressing their functions. Proinflammatory molecules can induce MSC immunoregulatory potential, but they could also increase the expression of immunogenic molecules. Studying the effect of inflammatory environment on MSC immunomodulation and immunogenicity profiles is mandatory to improve cellular therapies. The aim of this study was to analyse the response of equine bone marrow MSCs (eBM-MSCs) to three inflammatory conditions. Equine BM-MSCs from three animals were exposed to: (a) 20% allogeneic inflammatory SF (SF); (b) 50 ng/ml of TNFα and IFNγ (CK50) and (c) 20 ng/ml of TNFα and IFNγ (CK20). After 72 h of exposure, expression of immunogenic and immunomodulation-related molecules, including cell-to-cell contact and paracrine signalling molecules, were analysed by RT-qPCR and flow cytometry. The gene expression of adhesion molecules was upregulated whereas MSC migration-related genes were downregulated by all inflammatory conditions tested. CK culture conditions significantly upregulated the expression of COX-2, iNOS, IDO and IL-6. MHC-I gene expression was upregulated by all conditions, whereas MHC-II was upregulated only after CK priming. The expression of CD40 did not significantly change, whereas the ligand, CD40L, was downregulated in CK conditions. Flow cytometry showed an increase in the percentage of positive cells and mean fluorescence intensity (MFI) of the MHC-I and MHC-II molecules at CK50

  18. Oxygen tension regulates the osteogenic, chondrogenic and endochondral phenotype of bone marrow derived mesenchymal stem cells

    SciTech Connect

    Sheehy, Eamon J.; Buckley, Conor T.; Kelly, Daniel J.

    2012-01-06

    Highlights: Black-Right-Pointing-Pointer Expansion in low oxygen enhances MSC proliferation and osteogenesis. Black-Right-Pointing-Pointer Differentiation in low oxygen enhances chondrogenesis and suppresses hypertrophy. Black-Right-Pointing-Pointer Oxygen can regulate the MSC phenotype for use in tissue engineering applications. -- Abstract: The local oxygen tension is a key regulator of the fate of mesenchymal stem cells (MSCs). The objective of this study was to investigate the effect of a low oxygen tension during expansion and differentiation on the proliferation kinetics as well as the subsequent osteogenic and chondrogenic potential of MSCs. We first hypothesised that expansion in a low oxygen tension (5% pO{sub 2}) would improve both the subsequent osteogenic and chondrogenic potential of MSCs compared to expansion in a normoxic environment (20% pO{sub 2}). Furthermore, we hypothesised that chondrogenic differentiation in a low oxygen environment would suppress hypertrophy of MSCs cultured in both pellets and hydrogels used in tissue engineering strategies. MSCs expanded at 5% pO{sub 2} proliferated faster forming larger colonies, resulting in higher cell yields. Expansion at 5% pO{sub 2} also enhanced subsequent osteogenesis of MSCs, whereas differentiation at 5% pO{sub 2} was found to be a more potent promoter of chondrogenesis than expansion at 5% pO{sub 2}. Greater collagen accumulation, and more intense staining for collagen types I and X, was observed in pellets maintained at 20% pO{sub 2} compared to 5% pO{sub 2}. Both pellets and hydrogels stained more intensely for type II collagen when undergoing chondrogenesis in a low oxygen environment. Differentiation at 5% pO{sub 2} also appeared to inhibit hypertrophy in both pellets and hydrogels, as demonstrated by reduced collagen type X and Alizarin Red staining and alkaline phosphatase activity. This study demonstrates that the local oxygen environment can be manipulated in vitro to either stabilise a

  19. Promoting effect of small molecules in cardiomyogenic and neurogenic differentiation of rat bone marrow-derived mesenchymal stem cells

    PubMed Central

    Khanabdali, Ramin; Saadat, Anbarieh; Fazilah, Maizatul; Bazli, Khairul Fidaa’ Khairul; Qazi, Rida-e-Maria; Khalid, Ramla Sana; Hasan Adli, Durriyyah Sharifah; Moghadamtousi, Soheil Zorofchian; Naeem, Nadia; Khan, Irfan; Salim, Asmat; Shamsuddin, ShamsulAzlin Ahmad; Mohan, Gokula

    2016-01-01

    Small molecules, growth factors, and cytokines have been used to induce differentiation of stem cells into different lineages. Similarly, demethylating agents can trigger differentiation in adult stem cells. Here, we investigated the in vitro differentiation of rat bone marrow mesenchymal stem cells (MSCs) into cardiomyocytes by a demethylating agent, zebularine, as well as neuronal-like cells by β-mercaptoethanol in a growth factor or cytokines-free media. Isolated bone marrow-derived MSCs cultured in Dulbecco’s Modified Eagle’s Medium exhibited a fibroblast-like morphology. These cells expressed positive markers for CD29, CD44, and CD117 and were negative for CD34 and CD45. After treatment with 1 μM zebularine for 24 hours, the MSCs formed myotube-like structures after 10 days in culture. Expression of cardiac-specific genes showed that treated MSCs expressed significantly higher levels of cardiac troponin-T, Nkx2.5, and GATA-4 compared with untreated cells. Immunocytochemical analysis showed that differentiated cells also expressed cardiac proteins, GATA-4, Nkx 2.5, and cardiac troponin-T. For neuronal differentiation, MSCs were treated with 1 and 10 mM β-mercaptoethanol overnight for 3 hours in complete and serum-free Dulbecco’s Modified Eagle’s Medium, respectively. Following overnight treatment, neuron-like cells with axonal and dendritic-like projections originating from the cell body toward the neighboring cells were observed in the culture. The mRNA expression of neuronal-specific markers, Map2, Nefl, Tau, and Nestin, was significantly higher, indicating that the treated cells differentiated into neuronal-like cells. Immunostaining showed that differentiated cells were positive for the neuronal markers Flk, Nef, Nestin, and β-tubulin. PMID:26766903

  20. Population doubling level-dependent change of secreted glycosaminoglycan in equine bone marrow-derived mesenchymal stem cells

    PubMed Central

    SASAO, Takafumi; FUKUDA, Yuki; YOSHIDA, Sayako; MIYABARA, Shihori; KASASHIMA, Yoshinori; KUWANO, Atsutoshi; ARAI, Katsuhiko

    2015-01-01

    ABSTRACT In regenerative medicine using transplantation of mesenchymal stem cells (MSCs), the importance of regulating the quality of MSCs has been well recognized; however, there is little information concerning the relationship between the population doubling level (PDL) and the stemness of MSCs in equine medicine. In this study, we showed that the amount of glycosaminoglycan (GAG) secreted by bone marrow-derived MSCs (BMSCs) decreases with increase of PDL. Enzymatic digestion and two-dimensional electrophoresis revealed that a main component of GAG produced by BMSCs was hyaluronan with a small amount of chondroitin sulfate. Increase of PDL downregulated the expression of MSC CD markers, including CD44, CD73, CD90, CD105, and CD146, along with loss of differentiation capacity. Thus, the effect of hyaluronan supplement to the growth medium on both expression of CD markers and the tri-lineage potential of BMSCs was evaluated. Expression of CD73 and CD90 was preserved by continuous addition of hyaluronan to the growth medium, whereas mRNA levels corresponding to CD44, CD105 and CD146 were not preserved by supplementation of hyaluronan. BMSCs subcultured with hyaluronan-supplemented growth medium to PDL-12 showed osteogenic capacity, however adipogenic and chondrogenic activities at PDL-12 were not preserved by exogenous hyaluronan. These results suggest that downregulation of CD44, CD105 and CD146 might not affect the osteogenic capacity. Taken together, the results suggested that supplementation of hyaluronan to the growth medium might be effective at maintaining the osteogenic capacity of equine BMSCs. PMID:26435680

  1. [Differentiation of rat bone marrow-derived mesenchymal stem cells into cardiomyocyte-like cells induced by cyclic stretching strain].

    PubMed

    Kuang, Wei; Tang, Min; He, Xueling; Wu, Wenchao; Liu, Xiaojing; Li, Liang

    2014-06-01

    Bone marrow-derived mesenchymal stem cells (BMSCs) are multipotent stem cells that differentiate into a variety of cell types and widely used in tissue regeneration engineering. The purpose of this study is to investigate whether the cyclic biaxial stretching strain could promote the rat BMSCs (rBMSCs) to differentiate into cardiomyocyte-like cells in vitro. The second or third generation of rBMSCs were randomly divided into the cyclic stretching stain group, the control group and the blank group. Those rBMSCs in the cyclic stretching strain group were seeded on a silicone membrane with complete medium were exposed to biaxial stretching strain of 10% of membrane at a frequency of 1 Hz lasting for 6 h, 12 h and 24 h. Those in the control group were seeded on silicone membrane with complete medium. Those in the blank group were seeded in the 6-wells plates with complete medium. The mRNA expression of GATA4 and myocyte-specific enhancer factor 2C (MEF-2C) were detected by the real-time fluorescent quantification PCR and the protein expression of connexin 43 (Cx43) was detected by using the Western blot method. The results showed that the mRNA expression level of the GATA4 and MEF-2C, and the protein expression level of Cx43 were significantly higher in the cyclic stretching strain groups, compared with those in the relative control groups (P < 0.05). It suggests that cyclic biaxial stretching strain could play a part in the induction of rBMSCs to differentiate into cardiomyocyte-like cells in vitro, but the differentiation mechanism is still unclear. PMID:25219242

  2. Bone marrow-derived mesenchymal stem cells in three-dimensional culture promote neuronal regeneration by neurotrophic protection and immunomodulation.

    PubMed

    Han, Sufang; Wang, Bin; Li, Xing; Xiao, Zhifeng; Han, Jin; Zhao, Yannan; Fang, Yongxiang; Yin, Yanyun; Chen, Bing; Dai, Jianwu

    2016-07-01

    Accumulating evidence has revealed three-dimensional (3D) culture could better mimic the stem cell niche in vivo in comparison with conventional two-dimensional (2D) culture. In this study, we found that bone marrow derived mesenchymal stem cells (BMSCs) cultured in 3D collagen scaffold (3D BMSCs) exhibited distinctive features including significantly enhancing neurotrophic factor secretions and reducing macrophage activations challenged by lipopolysaccharide (LPS) in vitro. To further evaluate 3D BMSCs' potential benefits to the regeneration of spinal cord injury (SCI), the 3D and 2D BMSCs were respectively implanted in rat hemisected SCI. Compared with 2D cohort, 3D BMSCs transplantation significantly reduced the expressions of inflammatory cytokines such as TNF-α, IL-1β, and IL-6 at 5 days after transplantation, markedly enhanced axonal regeneration, and promoted motor functional recovery during 8 weeks of observation. When Nocodazole was used to depolymerize the cytoskeleton of 3D BMSCs, the changed expressions of neurotrophic factors and inflammatory cytokines were blunted, at least partially. Thus synergistic effects of neuronal protection and immunomodulation of 3D BMSCs may lead to a better functional recovery of SCI and the underlying mechanism may involve the alteration of their cellular morphology because of 3D culture. This study contributes to a better understanding of the cellular characteristics of 3D BMSCs and provides a novel strategy to promote the repair of the injured spinal cord. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 104A: 1759-1769, 2016. PMID:26990583

  3. Fibroblast Growth Factor 2 Regulates High Mobility Group A2 Expression in Human Bone Marrow-Derived Mesenchymal Stem Cells.

    PubMed

    Kalomoiris, Stefanos; Cicchetto, Andrew C; Lakatos, Kinga; Nolta, Jan A; Fierro, Fernando A

    2016-09-01

    Mesenchymal stem cells (MSCs) are an excellent source for numerous cellular therapies due to their simple isolation, low immunogenicity, multipotent differentiation potential and regenerative secretion profile. However, over-expanded MSCs show decreased therapeutic efficacy. This shortcoming may be circumvented by identifying methods that promote self-renewal of MSCs in culture. HMGA2 is a DNA-binding protein that regulates self-renewal in multiple types of stem cells through chromatin remodeling, but its impact on human bone marrow-derived MSCs is not known. Using an isolation method to obtain pure MSCs within 9 days in culture, we show that expression of HMGA2 quickly decreases during early expansion of MSCs, while let-7 microRNAs (which repress HMGA2) are simultaneously increased. Remarkably, we demonstrate that FGF-2, a growth factor commonly used to promote self-renewal in MSCs, rapidly induces HMGA2 expression in a time- and concentration-dependent manner. The signaling pathway involves FGF-2 receptor 1 (FGFR1) and ERK1/2, but acts independent from let-7. By silencing HMGA2 using shRNAs, we demonstrate that HMGA2 is necessary for MSC proliferation. However, we also show that over-expression of HMGA2 does not increase cell proliferation, but rather abrogates the mitogenic effect of FGF-2, possibly through inhibition of FGFR1. In addition, using different methods to assess in vitro differentiation, we show that modulation of HMGA2 inhibits adipogenesis, but does not affect osteogenesis of MSCs. Altogether, our results show that HMGA2 expression is associated with highly proliferating MSCs, is tightly regulated by FGF-2, and is involved in both proliferation and adipogenesis of MSCs. J. Cell. Biochem. 117: 2128-2137, 2016. © 2016 Wiley Periodicals, Inc. PMID:26888666

  4. An in vitro expansion score for tissue-engineering applications with human bone marrow-derived mesenchymal stem cells.

    PubMed

    Bertolo, Alessandro; Mehr, Marco; Janner-Jametti, Tiziana; Graumann, Ursula; Aebli, Niklaus; Baur, Martin; Ferguson, Stephen J; Stoyanov, Jivko V

    2016-02-01

    Human bone marrow-derived mesenchymal stem cells (MSCs) have limited growth potential in vitro and cease to divide due to replicative senescence, which from a tissue-engineering perspective has practical implications, such as defining the correct starting points for differentiation and transplantation. Time spent in culture before the loss of required differentiation potential is different and reflects patient variability, which is a problem for cell expansion. This study aimed to develop a score set which can be used to quantify the senescent state of MSCs and predict whether cells preserve their ability to differentiate to osteogenic, adipogenic and chondrogenic phenotypes, based on colony-forming unit (CFU) assay, population doubling time (PDT), senescence-associated β-galactosidase (SA-β-Gal) activity, cell size, telomere length and gene expression of MSCs cultured in vitro over 11 passages. This set of morphological, physiological and genetic senescence markers was correlated to the ability of MSCs to differentiate. Differentiation efficiency was assessed by marker genes and protein expression. CFUs decreased with increasing passage number, whereas SA-β-Gal activity and PDT increased; however, the correlation with MSCs' differentiation potential was sometimes unexpected. The expression of genes related to senescence was higher in late-passage cells than in early-passage cells. Early-passage cells underwent efficient osteogenic differentiation, with mid-passage cells performing best in chondrogenic differentiation. Late-passage cells preserve only adipogenic differentiation potential. Based on this marker set, we propose a senescence score in which combined markers give a reliable quality control of MSCs, not depending only on mechanistic passage number. PMID:23576360

  5. Bone marrow-derived mesenchymal stem cells enhance angiogenesis via their α6β1 integrin receptor

    SciTech Connect

    Carrion, Bita; Kong, Yen P.; Kaigler, Darnell; Putnam, Andrew J.

    2013-11-15

    Bone marrow-derived mesenchymal stem cells (BMSCs) facilitate the angiogenic response of endothelial cells (ECs) within three-dimensional (3D) matrices in vivo and in engineered tissues in vitro in part through paracrine mediators and by acting as stabilizing pericytes. However, the molecular interactions between BMSCs and nascent tubules during the process of angiogenesis are not fully understood. In this study, we have used a tractable 3D co-culture model to explore the functional role of the α6β1 integrin adhesion receptor on BMSCs in sprouting angiogenesis. We report that knockdown of the α6 integrin subunit in BMSCs significantly reduces capillary sprouting, and causes their failure to associate with the nascent vessels. Furthermore, we demonstrate that the BMSCs with attenuated α6 integrin proliferate at a significantly lower rate relative to either control cells expressing non-targeting shRNA or wild type BMSCs; however, despite adding more cells to compensate for this deficit in proliferation, deficient sprouting persists. Collectively, our findings demonstrate that the α6 integrin subunit in BMSCs is important for their ability to stimulate vessel morphogenesis. This conclusion may have important implications in the optimization of cell-based strategies to promote angiogenesis. Highlights: • BMSCs stimulate angiogenesis, but the mechanisms remain unclear. • We silenced the expression of the α6 integrin subunit in BMSCs. • Silencing this receptor subunit significantly inhibited angiogenic sprouting. • Knocking down α6 integrin affected laminin and αSMA expression. • Silencing α6 integrin expression also reduced BMSC proliferation.

  6. Intervertebral disc regeneration using platelet-rich plasma-containing bone marrow-derived mesenchymal stem cells: A preliminary investigation

    PubMed Central

    WANG, SHAN-ZHENG; JIN, JI-YANG; GUO, YU-DONG; MA, LIANG-YU; CHANG, QING; PENG, XIN-GUI; GUO, FANG-FANG; ZHANG, HAI-XIANG; HU, XIN-FENG; WANG, CHEN

    2016-01-01

    Platelet-rich plasma (PRP) is a promising strategy for intervertebral disc degeneration (IDD). However, the short half-life of growth factors released from PRP cannot continuously stimulate the degenerated discs. Thus, the present study hypothesized that the combined use of PRP and bone marrow-derived mesenchymal stem cells (BMSCs) may repair the early degenerated discs in the long term for their synergistic reparative effect. In the present study, following the induction of early IDD by annular puncture in rabbits, PRP was prepared and mixed with BMSCs (PRP-BMSC group) for injection into the early degenerated discs. As controls, phosphate-buffered saline (PBS; PBS group) and PRP (PRP group) were similarly injected. Rabbits without any intervention served as a control group. At 8 weeks following treatment, histological changes of the injected discs were assessed. Magnetic resonance imaging (MRI) was used to detect the T2-weighted signal intensity of the targeted discs at weeks 1, 2 and 8 following treatment. Annular puncture resulted in disc narrowing and decreased T2-weighted signal intensity. At weeks 1 and 3, MRI examinations showed regenerative changes in the PRP-BMSC group and PRP group, whereas the PBS group exhibited a continuous degenerative process of the discs. At 8 weeks post-injection, the PRP-BMSCs induced a statistically significant restoration of discs, as shown by MRI (PRP-BMSCs, vs.PRP and PBS; P<0.05), which was also confirmed by histological evaluations. Thus, compared with PRP, the administration of PRP-containing BMSCs resulted in a superior regenerative effect on the early degenerated discs, which may be a promising therapeutic strategy for the restoration of early degenerated discs. PMID:26956080

  7. Effects of Tricalcium Silicate Cements on Osteogenic Differentiation of Human Bone Marrow-Derived Mesenchymal Stem Cells In Vitro

    PubMed Central

    Eid, Ashraf A.; Hussein, Khalid A.; Niu, Li-na; Li, Guo-hua; Watanabe, Ikuya; Al-Shabrawey, Mohamed; Pashley, David H.; Tay, Franklin R.

    2014-01-01

    Tricalcium silicate cements have been successfully employed in the biomedical field as bioactive bone and dentin substitutes, with widely acclaimed osteoactive properties. This research analyzed the effects of different tricalcium silicate cement formulations on the temporal osteoactivity profile of human bone marrow-derived mesenchymal stem cells (hMW-MSCs). These cells were exposed to 4 commercially-available tricalcium silicate cement formulations in osteogenic differentiation medium. After 1, 3, 7 and 10 days, quantitative real time-polymerase chain reaction and Western blotting were performed to detect the expression of target osteogenic markers ALP, RUNX2, OSX, OPN, MSX2, and OCN. After 3, 7, 14 and 21 days, alkaline phosphatase assay was performed to detect changes in intracellular enzyme level. Alizarin Red S assay was performed after 28 days to detect extracellular matrix mineralization. In the presence of tricalcium silicate cements, target osteogenic markers were downregulated at the mRNA and protein levels at all time-points. Intracellular alkaline phosphatase enzyme levels and extracellular mineralization of the experimental groups were not significantly different from the untreated control. Quantitative polymerase chain reaction results showed increases in downregulation of RUNX2, OSX, MSX2 and OCN with increase in time of exposure to the tricalcium silicate cements, while ALP showed peak downregulation at day 7. For Western blotting, OSX, OPN, MSX2 and OCN showed increased downregulation with increased exposure time to the tested cements. Alkaline phosphatase enzyme levels generally declined after day 7. Based on these results, it is concluded that tricalcium silicate cements do not induce osteogenic differentiation of hBM-MSCs in vitro. PMID:24726977

  8. Chemically primed bone-marrow derived mesenchymal stem cells show enhanced expression of chemokine receptors contributed to their migration capability

    PubMed Central

    Bidkhori, Hamid Reza; Ahmadiankia, Naghmeh; Matin, Maryam Moghaddam; Heirani-tabasi, Asieh; Farshchian, Moein; Naderi-meshkin, Hojjat; Shahriyari, Mina; Dastpak, Mahtab; Bahrami, Ahmad Reza

    2016-01-01

    Objective(s): The limited homing potential of bone-marrow-derived mesenchymal stem cells (BM-MSC) is the key obstacle in MSC-based therapy. It is believed that chemokines and chemokine receptor interactions play key roles in cellular processes associated with migration. Meanwhile, MSCs express a low level of distinct chemokine receptors and they even lose these receptors on their surface after a few passages which influence their therapeutic applications negatively. This study investigated whether treatment of BM-MSCs with hypoxia-mimicking agents would increase expression of some chemokine receptors and cell migration. Materials and Methods: BM-MSCs were treated at passage 2 for our gene expression profiling. All qPCR experiments were performed by SYBR Green method in CFX-96 Bio-Rad Real-Time PCR. The Boyden chamber assay was utilized to investigate BM-MSC homing. Results: Possible approaches to increasing the expression level of chemokine receptors by different hypoxia-mimicking agents such as valproic acid (VPA), CoCl2, and desferrioxamine (DFX) are described. Results show DFX efficiently up-regulate the CXCR7 and CXCR4 gene expression while VPA increase only the CXCR7 gene expression and no significant change in expression level of CXCR4 and the CXCR7 gene was detectable by CoCl2 treatment. Chemotaxis assay results show that pre-treatment with DFX, VPA, and Cocl2 enhances significantly the migration ability of BM-MSCs compared with the untreated control group and DFX treatment accelerates MSCs homing significantly with a higher rate than VPA and Cocl2 treatments. Conclusion: Our data supports the notion that pretreatment of MSC with VPA and DFX improves the efficiency of MSC therapy by triggering homing regulatory signaling pathways. PMID:27096059

  9. Comparison of the Treatment Efficiency of Bone Marrow-Derived Mesenchymal Stem Cell Transplantation via Tail and Portal Veins in CCl4-Induced Mouse Liver Fibrosis

    PubMed Central

    Truong, Nhung Hai; Nguyen, Nam Hai; Le, Trinh Van; Vu, Ngoc Bich; Huynh, Nghia; Nguyen, Thanh Van; Le, Huy Minh; Phan, Ngoc Kim

    2016-01-01

    Because of self-renewal, strong proliferation in vitro, abundant sources for isolation, and a high differentiation capacity, mesenchymal stem cells are suggested to be potentially therapeutic for liver fibrosis/cirrhosis. In this study, we evaluated the treatment effects of mouse bone marrow-derived mesenchymal stem cells (BM-MSCs) on mouse liver cirrhosis induced by carbon tetrachloride. Portal and tail vein transplantations were examined to evaluate the effects of different injection routes on the liver cirrhosis model at 21 days after transplantation. BM-MSCs transplantation reduced aspartate aminotransferase/alanine aminotransferase levels at 21 days after injection. Furthermore, BM-MSCs induced positive changes in serum bilirubin and albumin and downregulated expression of integrins (600- to 7000-fold), transforming growth factor, and procollagen-α1 compared with the control group. Interestingly, both injection routes ameliorated inflammation and liver cirrhosis scores. All mice in treatment groups had reduced inflammation scores and no cirrhosis. In conclusion, transplantation of BM-MSCs via tail or portal veins ameliorates liver cirrhosis in mice. Notably, there were no differences in treatment effects between tail and portal vein administrations. In consideration of safety, we suggest transfusion of bone marrow-derived mesenchymal stem cells via a peripheral vein as a potential method for liver fibrosis treatment. PMID:26839564

  10. Autologous Bone Marrow-Derived Mesenchymal Stem Cells Modulate Molecular Markers of Inflammation in Dogs with Cruciate Ligament Rupture

    PubMed Central

    Muir, Peter; Hans, Eric C.; Racette, Molly; Volstad, Nicola; Sample, Susannah J.; Heaton, Caitlin; Holzman, Gerianne; Schaefer, Susan L.; Bloom, Debra D.; Bleedorn, Jason A.; Hao, Zhengling; Amene, Ermias; Suresh, M.; Hematti, Peiman

    2016-01-01

    Mid-substance rupture of the canine cranial cruciate ligament rupture (CR) and associated stifle osteoarthritis (OA) is an important veterinary health problem. CR causes stifle joint instability and contralateral CR often develops. The dog is an important model for human anterior cruciate ligament (ACL) rupture, where rupture of graft repair or the contralateral ACL is also common. This suggests that both genetic and environmental factors may increase ligament rupture risk. We investigated use of bone marrow-derived mesenchymal stem cells (BM-MSCs) to reduce systemic and stifle joint inflammatory responses in dogs with CR. Twelve dogs with unilateral CR and contralateral stable partial CR were enrolled prospectively. BM-MSCs were collected during surgical treatment of the unstable CR stifle and culture-expanded. BM-MSCs were subsequently injected at a dose of 2x106 BM-MSCs/kg intravenously and 5x106 BM-MSCs by intra-articular injection of the partial CR stifle. Blood (entry, 4 and 8 weeks) and stifle synovial fluid (entry and 8 weeks) were obtained after BM-MSC injection. No adverse events after BM-MSC treatment were detected. Circulating CD8+ T lymphocytes were lower after BM-MSC injection. Serum C-reactive protein (CRP) was decreased at 4 weeks and serum CXCL8 was increased at 8 weeks. Synovial CRP in the complete CR stifle was decreased at 8 weeks. Synovial IFNγ was also lower in both stifles after BM-MSC injection. Synovial/serum CRP ratio at diagnosis in the partial CR stifle was significantly correlated with development of a second CR. Systemic and intra-articular injection of autologous BM-MSCs in dogs with partial CR suppresses systemic and stifle joint inflammation, including CRP concentrations. Intra-articular injection of autologous BM-MSCs had profound effects on the correlation and conditional dependencies of cytokines using causal networks. Such treatment effects could ameliorate risk of a second CR by modifying the stifle joint inflammatory response

  11. Autologous Bone Marrow-Derived Mesenchymal Stem Cells Modulate Molecular Markers of Inflammation in Dogs with Cruciate Ligament Rupture.

    PubMed

    Muir, Peter; Hans, Eric C; Racette, Molly; Volstad, Nicola; Sample, Susannah J; Heaton, Caitlin; Holzman, Gerianne; Schaefer, Susan L; Bloom, Debra D; Bleedorn, Jason A; Hao, Zhengling; Amene, Ermias; Suresh, M; Hematti, Peiman

    2016-01-01

    Mid-substance rupture of the canine cranial cruciate ligament rupture (CR) and associated stifle osteoarthritis (OA) is an important veterinary health problem. CR causes stifle joint instability and contralateral CR often develops. The dog is an important model for human anterior cruciate ligament (ACL) rupture, where rupture of graft repair or the contralateral ACL is also common. This suggests that both genetic and environmental factors may increase ligament rupture risk. We investigated use of bone marrow-derived mesenchymal stem cells (BM-MSCs) to reduce systemic and stifle joint inflammatory responses in dogs with CR. Twelve dogs with unilateral CR and contralateral stable partial CR were enrolled prospectively. BM-MSCs were collected during surgical treatment of the unstable CR stifle and culture-expanded. BM-MSCs were subsequently injected at a dose of 2x106 BM-MSCs/kg intravenously and 5x106 BM-MSCs by intra-articular injection of the partial CR stifle. Blood (entry, 4 and 8 weeks) and stifle synovial fluid (entry and 8 weeks) were obtained after BM-MSC injection. No adverse events after BM-MSC treatment were detected. Circulating CD8+ T lymphocytes were lower after BM-MSC injection. Serum C-reactive protein (CRP) was decreased at 4 weeks and serum CXCL8 was increased at 8 weeks. Synovial CRP in the complete CR stifle was decreased at 8 weeks. Synovial IFNγ was also lower in both stifles after BM-MSC injection. Synovial/serum CRP ratio at diagnosis in the partial CR stifle was significantly correlated with development of a second CR. Systemic and intra-articular injection of autologous BM-MSCs in dogs with partial CR suppresses systemic and stifle joint inflammation, including CRP concentrations. Intra-articular injection of autologous BM-MSCs had profound effects on the correlation and conditional dependencies of cytokines using causal networks. Such treatment effects could ameliorate risk of a second CR by modifying the stifle joint inflammatory response

  12. Isolation, characterization and the multi-lineage differentiation potential of rabbit bone marrow-derived mesenchymal stem cells

    PubMed Central

    Tan, Sik-Loo; Ahmad, Tunku Sara; Selvaratnam, Lakshmi; Kamarul, Tunku

    2013-01-01

    Mesenchymal stem cells (MSCs) are recognized by their plastic adherent ability, fibroblastic-like appearance, expression of specific surface protein markers, and are defined by their ability to undergo multi-lineage differentiation. Although rabbit bone marrow-derived MSCs (rbMSCs) have been used extensively in previous studies especially in translational research, these cells have neither been defined morphologically and ultrastructurally, nor been compared with their counterparts in humans in their multi-lineage differentiation ability. A study was therefore conducted to define the morphology, surface marker proteins, ultrastructure and multi-lineage differentiation ability of rbMSCs. Herein, the primary rbMSC cultures of three adult New Zealand white rabbits (at least 4 months old) were used for three independent experiments. rbMSCs were isolated using the gradient-centrifugation method, an established technique for human MSCs (hMSCs) isolation. Cells were characterized by phase contrast microscopy observation, transmission electron microscopy analysis, reverse transcriptase-polymerase chain reaction (PCR) analysis, immunocytochemistry staining, flow cytometry, alamarBlue® assay, histological staining and quantitative (q)PCR analysis. The isolated plastic adherent cells were in fibroblastic spindle-shape and possessed eccentric, irregular-shaped nuclei as well as rich inner cytoplasmic zones similar to that of hMSCs. The rbMSCs expressed CD29, CD44, CD73, CD81, CD90 and CD166, but were negative (or dim positive) for CD34, CD45, CD117 and HLD-DR. Despite having similar morphology and phenotypic expression, rbMSCs possessed significantly larger cell size but had a lower proliferation rate as compared with hMSCs. Using established protocols to differentiate hMSCs, rbMSCs underwent osteogenic, adipogenic and chondrogenic differentiation. Interestingly, differentiated rbMSCs demonstrated higher levels of osteogenic (Runx2) and chondrogenic (Sox9) gene expressions

  13. Bone marrow-derived mesenchymal stem cells migrate to healthy and damaged salivary glands following stem cell infusion.

    PubMed

    Schwarz, Silke; Huss, Ralf; Schulz-Siegmund, Michaela; Vogel, Breda; Brandau, Sven; Lang, Stephan; Rotter, Nicole

    2014-09-01

    Xerostomia is a severe side effect of radiation therapy in head and neck cancer patients. To date, no satisfactory treatment option has been established. Because mesenchymal stem cells (MSCs) have been identified as a potential treatment modality, we aimed to evaluate stem cell distribution following intravenous and intraglandular injections using a surgical model of salivary gland damage and to analyse the effects of MSC injections on the recruitment of immune cells. The submandibular gland ducts of rats were surgically ligated. Syngeneic adult MSCs were isolated, immortalised by simian virus 40 (SV40) large T antigen and characterized by flow cytometry. MSCs were injected intravenously and intraglandularly. After 1, 3 and 7 days, the organs of interest were analysed for stem cell recruitment. Inflammation was analysed by immunohistochemical staining. We were able to demonstrate that, after intravenous injection, MSCs were recruited to normal and damaged submandibular glands on days 1, 3 and 7. Unexpectedly, stem cells were recruited to ligated and non-ligated glands in a comparable manner. After intraglandular injection of MSCs into ligated glands, the presence of MSCs, leucocytes and macrophages was enhanced, compared to intravenous injection of stem cells. Our data suggest that injected MSCs were retained within the inflamed glands, could become activated and subsequently recruited leucocytes to the sites of tissue damage. PMID:24810808

  14. Toxicological effects of pet food ingredients on canine bone marrow-derived mesenchymal stem cells and enterocyte-like cells.

    PubMed

    Ortega, M T; Jeffery, B; Riviere, J E; Monteiro-Riviere, N A

    2016-02-01

    We developed an in vitro method to assess pet food ingredients safety. Canine bone marrow-derived mesenchymal stem cells (BMSC) were differentiated into enterocyte-like cells (ELC) to assess toxicity in cells representing similar patterns of exposure in vivo. The toxicological profile of clove leave oil, eugenol, guanosine monophosphate (GMP), GMP + inosine monophosphate, sorbose, ginger root extract, cinnamon bark oil, cinnamaldehyde, thyme oil, thymol and citric acid was assessed in BMSC and ELC. The LC50 for GMP + inosine monophosphate was 59.42 ± 0.90 and 56.7 ± 3.5 mg ml(-1) for BMSC and ELC; 56.84 ± 0.95 and 53.66 ± 1.36 mg ml(-1) for GMP; 0.02 ± 0.001 and 1.25 ± 0.47 mg ml(-1) for citric acid; 0.077 ± 0.002 and 0.037 ± 0.01 mg ml(-1) for cinnamaldehyde; 0.002 ± 0.0001 and 0.002 ± 0.0008 mg ml(-1) for thymol; 0.080 ± 0.003 and 0.059 ± 0.001 mg ml(-1) for thyme oil; 0.111 ± 0.002 and 0.054 ± 0.01 mg ml(-1) for cinnamon bark oil; 0.119 ± 0.0004 and 0.099 ± 0.011 mg ml(-1) for clove leave oil; 0.04 ± 0.001 and 0.028 ± 0.002 mg ml(-1) for eugenol; 2.80 ± 0.11 and 1.75 ± 0.51 mg ml(-1) for ginger root extract; > 200 and 116.78 ± 7.35 mg ml(-1) for sorbose. Lemon grass oil was evaluated at 0.003-0.9 in BMSC and .03-0.9 mg ml(-1) in ELC and its mechanistic effect was investigated. The gene toxicology studies showed regulation of 61% genes in CYP450 pathway, 37% in cholestasis and 33% in immunotoxicity pathways for BMSC. For ELC, 80% for heat shock response, 69% for beta-oxidation and 65% for mitochondrial energy metabolism. In conclusion, these studies provide a baseline against which differential toxicity of dietary feed ingredients can be assessed in vitro for direct effects on canine cells and demonstrate differential toxicity in differentiated cells that represent gastrointestinal epithelial cells. PMID

  15. Donor-matched comparison of dental pulp stem cells and bone marrow-derived mesenchymal stem cells in a rat model

    PubMed Central

    Alge, Daniel L.; Zhou, Dan; Adams, Lyndsey L.; Wyss, Brandon K.; Shadday, Matthew D.; Woods, Erik J.; Chu, T.M. Gabriel; Goebel, W. Scott

    2010-01-01

    Dental pulp stem cells (DPSC) have drawn much interest for the regeneration of mineralized tissues, and several studies have compared DPSC to bone marrow-derived mesenchymal stem cells (BMMSC). However, conflicting results, possibly due to donor-associated variability, have been published and the regenerative potential of DPSC is currently unclear. In the present study we have sought to address this problem using a donor-matched experimental design to robustly compare the biological properties of DPSC and BMMSC. All experiments were performed using cells isolated from a single adult Sprague-Dawley rat. Our results show that DPSC and BMMSC had similar morphologies and flow cytometry profiles, were capable of forming colonies in vitro, and were capable of osteogenic, chondrogenic, and adipogenic differentiation. However, quantitative comparisons revealed that DPSC had a faster population doubling time and a higher percentage of stem/progenitor cells in the population as determined by clonogenic assays. Furthermore, while both cell populations formed mineral in vitro, DPSC had significantly higher alkaline phosphatase activity than BMMSC after three weeks in osteogenic medium. These data show several key differences between DPSC and BMMSC and support the possibility of using DPSC for mineralized tissue regeneration. PMID:19842108

  16. Comparing the immunoregulatory effects of stem cells from human exfoliated deciduous teeth and bone marrow-derived mesenchymal stem cells.

    PubMed

    Alipour, Razieh; Adib, Minoo; Masoumi Karimi, Masoumeh; Hashemi-Beni, Batool; Sereshki, Nasrin

    2013-12-01

    Stem cells from human exfoliated deciduous teeth (SHED) have been introduced recently and possess characteristics similar to mesenchymal stem cells (MSCs). Because of their convenient accessibility and safety of harvest, SHED can be a preferable source for the ever-increasing MSCs' applications  While they are new, their immunoproperties have not been adequately studied. In this study, we aimed to explore the effect of SHED on T lymphocytes and compare it to conventional MSCs (BMMSCs).At first the isolated T lymphocytes were activated specifically/nonspecifically in vitro and cocultured with SHED or BMMSCs under the same conditions, subsequently their proliferation and cytokine secretion (IL-2 and IFN-γ) were measured.In our experiment, BMMSCs and SHED inhibit the proliferation and cytokine production of both PHA and alloantigen stimulated T lymphocytes in a dose-dependent manner. In direct and indirect contact to T lymphocytes, the inhibition of BMMSCs (but not of SHED) was significantly different The cytokine production from activated T cells was affected differently by two types of MSCs. The inhibition decreased by the separation of lymphocytes and MSCs by a semipermeable membrane, but it was not abolished.This study showed that SHED suppress the activation of human T lymphocytes in vitro like other MSCs. Compared to BMMSCs, this suppression was alleviated. In the equal conditions, the pattern of immune-modulation of BMMSCs and SHED was different, suggesting that SHED do not exert the exact mechanisms of BMMSCs' immunosuppression., This finding should be verified by further studies focused on the detailed mechanisms  of the immunomodulation of SHED and also BMMSCs. PMID:23996709

  17. Up-regulation of immunomodulatory effects of mouse bone-marrow derived mesenchymal stem cells by tetrahydrocannabinol pre-treatment involving cannabinoid receptor CB2

    PubMed Central

    Xie, Junran; Xiao, Dongju; Xu, Yun; Zhao, Jinning; Jiang, Li; Hu, Xuming; Zhang, Yaping; Yu, Lina

    2016-01-01

    Chronic pain is commonly and closely correlated with inflammation. Both cannabinoid signaling and mesenchymal stem cells (MSCs) have been demonstrated to reduce inflammatory pain. Although cannabinoid signaling is essential for mesenchymal stem cell survival and differentiation, little is known about its role in modulatory effect of MSCs on inflammation and pain sensitivity. Here we showed that mouse bone-marrow derived MSCs (BM-MSCs) expressed both cannabinoid receptor type 1 and 2 (CB1 and CB2). CB2 expression level in BM-MSCs increased with their maturation. In addition, we found that tetrahydrocannabinol (THC) activated CB2 receptor and ERK signaling, consequently enhancing the modulation of MSCs on inflammation-associated cytokine release from lipopolysaccharides-stimulated microglia. Consistent with in vitro data, THC pretreatment enhanced the immunomodulatory effects of BM-MSC on thermal hyperalgesia and mechanical allodynia in chronic constriction injury model, by decreasing the release of pro-inflammation cytokines. Our study revealed the crucial role of THC in promoting the immunomodulatory effects of MSCs and proposed a new strategy to alleviate pain based on stem cells therapy. PMID:26824325

  18. Effects of magnetic nanoparticle-incorporated human bone marrow-derived mesenchymal stem cells exposed to pulsed electromagnetic fields on injured rat spinal cord.

    PubMed

    Cho, Hyunjin; Choi, Yun-Kyong; Lee, Dong Heon; Park, Hee Jung; Seo, Young-Kwon; Jung, Hyun; Kim, Soo-Chan; Kim, Sung-Min; Park, Jung-Keug

    2013-01-01

    Transplanting mesenchymal stem cells into injured lesions is currently under study as a therapeutic approach for spinal cord injury. In this study, the effects of a pulsed electromagnetic field (PEMF) on injured rat spinal cord were investigated in magnetic nanoparticle (MNP)-incorporated human bone marrow-derived mesenchymal stem cells (hBM-MSCs). A histological analysis revealed significant differences in MNP-incorporated cell distribution near the injured site under the PEMF in comparison with that in the control group. We confirmed that MNP-incorporated cells were widely distributed in the lesions under PEMF. The results suggest that MNP-incorporated hBM-MSCs were guided by the PEMF near the injured site, and that PEMF exposure for 8 H per day over 4 weeks promoted behavioral recovery in spinal cord injured rats. The results show that rats with MNP-incorporated hBM-MSCs under a PEMF were more effective on the Basso, Beattie, and Bresnahan behavioral test and suggest that the PEMF enhanced the action of transplanted cells for recovery of the injured lesion. PMID:24033637

  19. Characterization and Expression of Senescence Marker in Prolonged Passages of Rat Bone Marrow-Derived Mesenchymal Stem Cells

    PubMed Central

    Ridzuan, Noridzzaida; Al Abbar, Akram; Yip, Wai Kien; Maqbool, Maryam

    2016-01-01

    The present study is aimed at optimizing the in vitro culture protocol for generation of rat bone marrow- (BM-) derived mesenchymal stem cells (MSCs) and characterizing the culture-mediated cellular senescence. The initial phase of generation and characterization was conducted using the adherent cells from Sprague Dawley (SD) rat's BM via morphological analysis, growth kinetics, colony forming unit capacity, immunophenotyping, and mesodermal lineage differentiation. Mesenchymal stem cells were successfully generated and characterized as delineated by the expressions of CD90.1, CD44H, CD29, and CD71 and lack of CD11b/c and CD45 markers. Upon induction, rBM-MSCs differentiated into osteocytes and adipocytes and expressed osteocytes and adipocytes genes. However, a decline in cell growth was observed at passage 4 onwards and it was further deciphered through apoptosis, cell cycle, and senescence assays. Despite the enhanced cell viability at later passages (P4-5), the expression of senescence marker, β-galactosidase, was significantly increased at passage 5. Furthermore, the cell cycle analysis has confirmed the in vitro culture-mediated cellular senescence where cells were arrested at the G0/G1 phase of cell cycle. Although the currently optimized protocols had successfully yielded rBM-MSCs, the culture-mediated cellular senescence limits the growth of rBM-MSCs and its potential use in rat-based MSC research. PMID:27579045

  20. Characterization and Expression of Senescence Marker in Prolonged Passages of Rat Bone Marrow-Derived Mesenchymal Stem Cells.

    PubMed

    Ridzuan, Noridzzaida; Al Abbar, Akram; Yip, Wai Kien; Maqbool, Maryam; Ramasamy, Rajesh

    2016-01-01

    The present study is aimed at optimizing the in vitro culture protocol for generation of rat bone marrow- (BM-) derived mesenchymal stem cells (MSCs) and characterizing the culture-mediated cellular senescence. The initial phase of generation and characterization was conducted using the adherent cells from Sprague Dawley (SD) rat's BM via morphological analysis, growth kinetics, colony forming unit capacity, immunophenotyping, and mesodermal lineage differentiation. Mesenchymal stem cells were successfully generated and characterized as delineated by the expressions of CD90.1, CD44H, CD29, and CD71 and lack of CD11b/c and CD45 markers. Upon induction, rBM-MSCs differentiated into osteocytes and adipocytes and expressed osteocytes and adipocytes genes. However, a decline in cell growth was observed at passage 4 onwards and it was further deciphered through apoptosis, cell cycle, and senescence assays. Despite the enhanced cell viability at later passages (P4-5), the expression of senescence marker, β-galactosidase, was significantly increased at passage 5. Furthermore, the cell cycle analysis has confirmed the in vitro culture-mediated cellular senescence where cells were arrested at the G0/G1 phase of cell cycle. Although the currently optimized protocols had successfully yielded rBM-MSCs, the culture-mediated cellular senescence limits the growth of rBM-MSCs and its potential use in rat-based MSC research. PMID:27579045

  1. Differential alkaline phosphatase responses of rat and human bone marrow derived mesenchymal stem cells to 45S5 bioactive glass

    PubMed Central

    Reilly, Gwendolen C.; Radin, Shula; Chen, Andrew T.; Ducheyne, Paul

    2009-01-01

    Bioactive glass is used as both a bone filler and as a coating on implants, and has been advocated as a potential osteogenic scaffold for tissue engineering. Rat derived mesenchymal stem cells (MSCs) show elevated levels of levels of alkaline phosphatase activity when grown on 45S5 bioactive glass as compared to standard tissue culture plastic. Similarly, exposure to the dissolution products of 45S5 elevates alkaline phosphatase activity and other osteogenic markers in these cells. We investigated whether human MSCs grown under the same laboratory conditions as rat MSCs would exhibit similar responses. In general, human MSCs produce markedly less alkaline phosphatase activity than rat MSCs, regardless of cell culture conditions, and do not respond to the growth factor BMP-2 in the same way as rat MSCs. In our experiments there was no difference in alkaline phosphatase activity between human MSCs grown on 45S5 bioactive glass or tissue culture plastic, in samples from five different orthopaedic patients, regardless of culture media composition. Neither was there any consistent effect of 45S5 dissolution products on human MSCs from three different donors. These results suggest that the positive effects of bioactive glass on bone growth in human patients are not mediated by accelerated differentiation of mesenchymal stem cells. PMID:17586040

  2. Pelleted Bone Marrow Derived Mesenchymal Stem Cells Are Better Protected from the Deleterious Effects of Arthroscopic Heat Shock

    PubMed Central

    Kalamegam, Gauthaman; Abbas, Mohammed; Gari, Mamdooh; Alsehli, Haneen; Kadam, Roaa; Alkaff, Mohammed; Chaudhary, Adeel; Al-Qahtani, Mohammed; Abuzenadah, Adel; Kafienah, Wael; Mobasheri, Ali

    2016-01-01

    Introduction: The impact of arthroscopic temperature on joint tissues is poorly understood and it is not known how mesenchymal stem cells (MSCs) respond to the effects of heat generated by the device during the process of arthroscopy assisted experimental cell-based therapy. In the present study, we isolated and phenotypically characterized human bone marrow mesenchymal stem cells (hBMMSCs) from osteoarthritis (OA) patients, and evaluated the effect of arthroscopic heat on cells in suspension and pellet cultures. Methods: Primary cultures of hBMMSCs were isolated from bone marrow aspirates of OA patients and cultured using DMEM supplemented with 10% FBS and characterized for their stemness. hBMMSCs (1 × 106 cells) cultured as single cell suspensions or cell pellets were exposed to an illuminated arthroscope for 10, 20, or 30 min. This was followed by analysis of cellular proliferation and heat shock related gene expression. Results: hBMMSCs were viable and exhibited population doubling, short spindle morphology, MSC related CD surface markers expression and tri-lineage differentiation into adipocytes, chondrocytes and osteoblasts. Chondrogenic and osteogenic differentiation increased collagen production and alkaline phosphatase activity. Exposure of hBMMSCs to an illuminated arthroscope for 10, 20, or 30 min for 72 h decreased metabolic activity of the cells in suspensions (63.27% at 30 min) and increased metabolic activity in cell pellets (62.86% at 10 min and 68.57% at 20 min). hBMMSCs exposed to 37, 45, and 55°C for 120 s demonstrated significant upregulation of BAX, P53, Cyclin A2, Cyclin E1, TNF-α, and HSP70 in cell suspensions compared to cell pellets. Conclusions: hBMMSC cell pellets are better protected from temperature alterations compared to cell suspensions. Transplantation of hBMMSCs as pellets rather than as cell suspensions to the cartilage defect site would therefore support their viability and may aid enhanced cartilage regeneration. PMID

  3. Different Forms of Tenascin-C with Tenascin-R Regulate Neural Differentiation in Bone Marrow-Derived Human Mesenchymal Stem Cells

    PubMed Central

    Tsai, Hung-Li; Chiu, Wen-Ta; Fang, Chia-Lang; Hwang, Shiaw-Min; Renshaw, Perry F.

    2014-01-01

    Mesenchymal stem cells (MSCs) are currently thought to transdifferentiate into neural lineages under specific microenvironments. Studies have reported that the tenascin family members, tenascin-C (TnC) and tenascin-R (TnR), regulate differentiation and migration, in addition to neurite outgrowth and survival in numerous types of neurons and mesenchymal progenitor cells. However, the mechanisms by which TnC and TnR affect neuronal differentiation are not well understood. In this study, we hypothesized that different forms of tenascin might regulate the neural transdifferentiation of human bone marrow-derived mesenchymal stem cells. Human MSCs were cultured in media incorporated with soluble tenascins, or on precoated tenascins. In a qualitative polymerase chain reaction analysis, adding a soluble TnC and TnR mixture to the medium significantly enhanced the expression of neuronal and glial markers, whereas no synaptic markers were expressed. Conversely, in groups of cells treated with coated TnC, hMSCs showed neurite outgrowth and synaptic marker expression. After being treated with coated TnR, hMSCs exhibited neuronal differentiation; however, it inhibited neurite outgrowth and synaptic marker expression. A combination of TnC and TnR significantly promoted hMSC differentiation in neurons or oligodendrocytes, induced neurite formation, and inhibited differentiation into astrocytes. Furthermore, the effect of the tenascin mixture showed dose-dependent effects, and a mixture ratio of 1:1 to 1:2 (TnC:TnR) provided the most obvious differentiation of neurons and oligodendrocytes. In a functional blocking study, integrin α7 and α9β1-blocking antibodies inhibited, respectively, 80% and 20% of mRNA expression by hMSCs in the coated tenascin mixture. In summary, the coated combination of TnC and TnR appeared to regulate neural differentiation signaling through integrin α7 and α9β1 in bone marrow-derived hMSCs. Our findings demonstrate novel mechanisms by which tenascin

  4. Characterization of bone marrow-derived mesenchymal stem cells from dimethyloxallyl glycine-preconditioned mice: Evaluation of the feasibility of dimethyloxallyl glycine as a mobilization agent

    PubMed Central

    GE, TINGTING; YU, QIN; LIU, WEI; CONG, LI; LIU, LIZHEN; WANG, YAN; ZHOU, LIPING; LIN, DEJU

    2016-01-01

    The prolyl hydroxylase inhibitor dimethyloxallyl glycine (DMOG) has been increasingly studied with regards to stem cell therapy. Previous studies have demonstrated that endogenous mesenchymal stem cells (MSCs) may be mobilized into peripheral circulation by pharmaceutical preconditioning. In addition, our previous study confirmed that DMOG, as a novel mobilization agent, could induce mouse/rat MSC migration into peripheral blood circulation. Therefore, the present study conducted studies to characterize bone marrow-derived MSCs (BM-MSCs) collected from mice following DMOG intraperitoneal injection. The surface antigen immune phenotype, differentiation capability, proliferative ability, migratory capacity and paracrine capacity of the BM-MSCs collected from DMOG-preconditioned mice (DBM-MSCs) or normal saline-treated mice (NBM-MSCs) were evaluated by means of flow cytometry, differentiation induction, Cell Counting kit-8, Transwell assay and enzyme-linked immunosorbent assay, respectively. Compared with NBM-MSCs, DBM-MSCs displayed a similar immune phenotype and multilineage differentiation capability, reduced proliferative ability and migratory capacity, and similar transforming growth factor and platelet-derived growth factor secretion capacity. These results provide a novel insight into the biological properties of BM-MSCs from mice preconditioned with DMOG. DBM-MSCs exhibited slightly distinct characteristics to NBM-MSCs; however, they may have therapeutic potential for future stem cell therapy. In addition, the present study suggested that DMOG may be used as a novel mobilization agent in future clinical trials as no adverse effects were observed. PMID:26935134

  5. Inducible Lentivirus-Mediated Expression of the Oct4 Gene Affects Multilineage Differentiation of Adult Human Bone Marrow-Derived Mesenchymal Stem Cells.

    PubMed

    Hao, Qiang; An, Jia-Qiang; Hao, Fei; Yang, Chun; Lu, Tao; Qu, Ting-Yu; Zhao, Li-Ru; Duan, Wei-Ming

    2015-10-01

    The octamer-binding transcription factor 4 (Oct4) gene plays an important role in maintaining the undifferentiated state of embryonic stem cells (ESCs) and reprogramming adult somatic cells into induced pluripotent stem cells (iPSCs). In the present study, we transduced human bone marrow-derived mesenchymal stem cells (hMSCs) using tetracycline-on (Tet-On) lentiviruses carrying human Oct4 to examine the effects of regulated expression of human Oct4 on the proliferation and differentiation of hMSCs. hMSCs were efficiently transduced by Tet-On lentiviruses to express regulated levels of human Oct4 with doxycycline (Dox), as examined by immunofluorescent staining, flow cytometry, and quantitative real time-PCR (qRT-PCR) assays. Ectopic expression of Oct4 in transduced hMSCs increased the ability of colony formation. Continued expression of Oct4 further enhanced adipogenic differentiation of hMSCs, and transient expression of Oct4 sufficiently enhanced osteogenic differentiation of hMSCs. qRT-PCR analysis showed that ectopic expression of Oct4 in transduced hMSCs temporally increased the expression of Sox2 and c-Myc. Interestingly, ectopic expression of Oct4 reduced neuronal differentiation of hMSCs when incubated under neuronal differentiation conditions. Our results suggest that ectopic expression of human Oct4 leads to temporal changes in multilineage differentiation of hMSCs and may inhibit neuronal differentiation of hMSCs. PMID:26230571

  6. Crosstalk between bone marrow-derived mesenchymal stem cells and regulatory T cells through a glucocorticoid-induced leucine zipper/developmental endothelial locus-1-dependent mechanism.

    PubMed

    Yang, Nianlan; Baban, Babak; Isales, Carlos M; Shi, Xing-Ming

    2015-09-01

    Bone marrow is a reservoir for regulatory T (T(reg)) cells, but how T(reg) cells are regulated in that environment remains poorly understood. We show that expression of glucocorticoid (GC)-induced leucine zipper (GILZ) in bone marrow mesenchymal lineage cells or bone marrow-derived mesenchymal stem cells (BMSCs) increases the production of T(reg) cells via a mechanism involving the up-regulation of developmental endothelial locus-1 (Del-1), an endogenous leukocyte-endothelial adhesion inhibitor. We found that the expression of Del-1 is increased ∼4-fold in the bone tissues of GILZ transgenic (Tg) mice, and this increase is coupled with a significant increase in the production of IL-10 (2.80 vs. 0.83) and decrease in the production of IL-6 (0.80 vs. 2.33) and IL-12 (0.25 vs. 1.67). We also show that GILZ-expressing BMSCs present antigen in a way that favors T(reg) cells. These results indicate that GILZ plays a critical role mediating the crosstalk between BMSCs and T(reg) in the bone marrow microenvironment. These data, together with our previous findings that overexpression of GILZ in BMSCs antagonizes TNF-α-elicited inflammatory responses, suggest that GILZ plays important roles in bone-immune cell communication and BMSC immune suppressive functions. PMID:26038125

  7. Physical Activity Increases the Total Number of Bone-Marrow-Derived Mesenchymal Stem Cells, Enhances Their Osteogenic Potential, and Inhibits Their Adipogenic Properties

    PubMed Central

    Marędziak, Monika; Śmieszek, Agnieszka; Chrząstek, Klaudia; Basinska, Katarzyna; Marycz, Krzysztof

    2015-01-01

    Aging and sedentary lifestyle are common nowadays and are associated with the increasing number of chronic diseases. Thus, physical activity is recommended as one of three healthy behavior factors that play a crucial role in health prophylaxis. In the present study, we were interested whether physical activity influences the number and potential of bone-marrow-derived mesenchymal stem cells BMMSCs. In this study, four-week-old male C57Bl/6 mice were trained on a treadmill at progressive speeds over a 5-week period. Comparisons made between exercised (EX) and sedentary animal groups revealed (i) significantly higher number of MSCs in EX animals, (ii) elevated alkaline phosphatase (ALP) activity, (iii) increased level of osteopontin (OPN) and osteocalcin (OCL), and (iv) reduced marrow cavity fat. The results obtained support the thesis that EX may play a substantial role in the regeneration of mesenchymal tissues. Therefore, EX may represent a novel, nonpharmacological strategy of slowing down age-related decline of the musculoskeletal functions. PMID:26167185

  8. BKCa and hEag1 channels regulate cell proliferation and differentiation in human bone marrow-derived mesenchymal stem cells.

    PubMed

    Zhang, Ying-Ying; Yue, Jianbo; Che, Hui; Sun, Hai-Ying; Tse, Hung-Fat; Li, Gui-Rong

    2014-02-01

    Human bone marrow-derived mesenchymal stem cells (MSCs) serve as a reservoir for the continuous renewal of various mesenchymal tissues; however, cellular physiology of ion channels is not fully understood. The present study investigated potential roles of large-conductance Ca(2+) -activated potassium (BKCa ) channels and ether-à-go-go potassium (hEag1 or Kv10.1) channels in regulating cell proliferation and differentiation in human MSCs. We found that inhibition of BKCa with paxilline or hEag1 with astemizole, or knockdown of BKCa with shRNAs targeting KCa1.1 or hEag1 channels with shRNAs targeting KCNH1 arrested the cells at G0/G1 phase. In addition, silencing BKCa or hEag1 channels significantly reduced adipogenic differentiation with decrease of lipid accumulation and expression of the adipocyte marker PPARγ, and decreased osteogenic differentiation with reduction of mineral precipitation and osteocalcin. These effects were accompanied with a reduced cyclin D1, cyclin E, p-ERK1/2, and p-Akt. Our results demonstrate that BKCa and hEag1 channels not only regulate cell proliferation, but also participate in the adipogenic and osteogenic differentiations in human MSCs, which indicates that BKCa and hEag1 channels may be essential in maintaining bone marrow physiological function and bone regeneration. PMID:23881642

  9. Atherogenic Cytokines Regulate VEGF-A-Induced Differentiation of Bone Marrow-Derived Mesenchymal Stem Cells into Endothelial Cells.

    PubMed

    Ikhapoh, Izuagie Attairu; Pelham, Christopher J; Agrawal, Devendra K

    2015-01-01

    Coronary artery stenting or angioplasty procedures frequently result in long-term endothelial dysfunction or loss and complications including arterial thrombosis and myocardial infarction. Stem cell-based therapies have been proposed to support endothelial regeneration. Mesenchymal stem cells (MSCs) differentiate into endothelial cells (ECs) in the presence of VEGF-A in vitro. Application of VEGF-A and MSC-derived ECs at the interventional site is a complex clinical challenge. In this study, we examined the effect of atherogenic cytokines (IL-6, TNFα, and Ang II) on EC differentiation and function. MSCs (CD44(+), CD73(+), CD90(+), CD14(-), and CD45(-)) were isolated from the bone marrow of Yucatan microswine. Naïve MSCs cultured in differentiation media containing VEGF-A (50 ng/mL) demonstrated increased expression of EC-specific markers (vWF, PECAM-1, and VE-cadherin), VEGFR-2 and Sox18, and enhanced endothelial tube formation. IL-6 or TNFα caused a dose-dependent attenuation of EC marker expression in VEGF-A-stimulated MSCs. In contrast, Ang II enhanced EC marker expression in VEGF-A-stimulated MSCs. Addition of Ang II to VEGF-A and IL-6 or TNFα was sufficient to rescue the EC phenotype. Thus, Ang II promotes but IL-6 and TNFα inhibit VEGF-A-induced differentiation of MSCs into ECs. These findings have important clinical implications for therapies intended to increase cardiac vascularity and reendothelialize coronary arteries following intervention. PMID:26106428

  10. Transient Receptor Potential Melastatin Type 7 Channel Is Critical for the Survival of Bone Marrow Derived Mesenchymal Stem Cells

    PubMed Central

    Feng, Ji-Ming; Figueiredo, Marxa L.; Zhang, Hanjie; Nelson, Piper L.; Marigo, Vanessa; Beck, Andreas

    2010-01-01

    The transient receptor potential melastatin type 7 channel (TRPM7) is a member of the TRP family of ion channels that is essential for cell proliferation and viability. Mesenchymal stem cells (MSCs) from bone marrow are a potential source for tissue repair due to their ability to differentiate into specialized cells. However, the role of TRPM7 in stem cells is unknown. In this study, we characterized TRPM7 in mouse MSCs using molecular biology, immunocytochemistry, and patch clamp. We also investigated TRPM7 function using a lentiviral vector and specific shRNA to knockdown gene expression. By RT-PCR and immunocytochemistry, we identified TRPM7, but not TRPM6, a close family member with similar function. Electrophysiological recordings during depletion of intracellular Mg2+ or Mg2+-ATP resulted in the development of currents typical for the channel. Furthermore, 2-aminoethoxydiphenyl borate (1 pM–100 μM) inhibited TRPM7 in a concentration-dependent manner. The molecular suppression of TRPM7 significantly decreased MSC proliferation and viability as determined by MTT assay. In addition, TRPM7 gene expression was up-regulated during osteogenesis. These findings demonstrate that TRPM7 is required for MSC survival and perhaps involved in the differentiation process. PMID:19929312

  11. A Member of the Nuclear Receptor Superfamily, Designated as NR2F2, Supports the Self-Renewal Capacity and Pluripotency of Human Bone Marrow-Derived Mesenchymal Stem Cells

    PubMed Central

    Zhu, Ni; Wang, Huafang; Wang, Binsheng; Wei, Jieping; Shan, Wei; Feng, Jingjing; Huang, He

    2016-01-01

    Mesenchymal stem cells are characterized with self-renewal capacity and pluripotency. NR2F2 is a nuclear receptor that has been detected in the mesenchymal compartment of developing organs. However, whether NR2F2 plays a role in the stemness maintenance of mesenchymal stem cells has not been explored yet. In this study, we investigated the function of NR2F2 in bone marrow-derived mesenchymal stem cells via shRNA-mediated knock-down of NR2F2. The suppression of NR2F2 impaired the colony-forming efficacy of mesenchymal stem cells. The inhibition of colony-forming capacity may be attributed to the acceleration of senescence through upregulation of P21 and P16. The downregulation of NR2F2 also suppressed both osteogenic and adipogenic differentiation processes. In conclusion, NR2F2 plays an important role in the stemness maintenance of bone marrow-derived mesenchymal stem cells. PMID:26783404

  12. Epigallocatechin-3-gallate Protects against Hydrogen Peroxide-Induced Inhibition of Osteogenic Differentiation of Human Bone Marrow-Derived Mesenchymal Stem Cells

    PubMed Central

    Wang, Dawei; Wang, Yonghui; Xu, Shihong; Wang, Fu; Wang, Bomin; Han, Ke; Sun, Daqing; Li, Lianxin

    2016-01-01

    Oxidative stress induces bone loss and osteoporosis, and epigallocatechin-3-gallate (EGCG) may be used to combat these diseases due to its antioxidative property. Herein, oxidative stress in human bone marrow-derived mesenchymal stem cells (BM-MSCs) was induced by H2O2, resulting in an adverse effect on their osteogenic differentiation. However, this H2O2-induced adverse effect was nullified when the cells were treated with EGCG. In addition, treatment of BM-MSCs with EGCG alone also resulted in the enhancement of osteogenic differentiation of BM-MSCs. After EGCG treatment, expressions of β-catenin and cyclin D1 were upregulated, suggesting that the Wnt pathway was involved in the effects of EGCG on the osteogenic differentiation of BM-MSCs. This was also confirmed by the fact that the Wnt pathway inhibitor, Dickkopf-1 (DKK-1), can nullify the EGCG-induced enhancement effect on BM-MSC's osteogenic differentiation. Hence, our results suggested that EGCG can reduce the effects of oxidative stress on Wnt pathway in osteogenic cells, which supported a potentially promising therapy of bone disorders induced by oxidative stress. Considering its positive effects on BM-MSCs, EGCG may also be beneficial for stem cell-based bone repair. PMID:26977159

  13. Effects of Line and Pillar Array Microengineered SiO2 Thin Films on the Osteogenic Differentiation of Human Bone Marrow-Derived Mesenchymal Stem Cells.

    PubMed

    Carvalho, Angela; Pelaez-Vargas, Alejandro; Hansford, Derek J; Fernandes, Maria H; Monteiro, Fernando J

    2016-02-01

    A primary goal in bone tissue engineering is the design of implants that induce controlled, guided, and rapid healing. The events that normally lead to the integration of an implant into bone and determine the performance of the device occur mainly at the tissue-implant interface. Topographical surface modification of a biomaterial might be an efficient tool for inducing stem cell osteogenic differentiation and replace the use of biochemical stimuli. The main goal of this work was to develop micropatterned bioactive silica thin films to induce the osteogenic differentiation of human bone marrow-derived mesenchymal stem cells (hMSCs) only through topographical stimuli. Line and pillar micropatterns were developed by a combination of sol-gel/soft lithography and characterized by scanning electron microscopy, atomic force microscopy, and contact angle measurements. hMSCs were cultured onto the microfabricated thin films and flat control for up to 21 days under basal conditions. The micropatterned groups induced levels of osteogenic differentiation and expression of osteoblast-associated markers higher than those of the flat controls. Via comparison of the micropatterns, the pillars caused a stronger response of the osteogenic differentiation of hMSCs with a higher level of expression of osteoblast-associated markers, ALP activity, and extracellular matrix mineralization after the cells had been cultured for 21 days. These findings suggest that specific microtopographic cues can direct hMSCs toward osteogenic differentiation. PMID:26771563

  14. Enhanced hepatic differentiation of rat bone marrow-derived mesenchymal stem cells in spheroidal aggregate culture on a decellularized liver scaffold

    PubMed Central

    Bao, Ji; Wu, Qiong; Wang, Yujia; Li, Yi; Li, Li; Chen, Fei; Wu, Xiujuan; Xie, Mingjun; Bu, Hong

    2016-01-01

    In the present study, we aimed to determine whether the combination of aggregate culture and decellularized liver scaffolds (DLSs) promoted the hepatic differentiation of murine bone marrow-derived mesenchymal stem cells (BM-MSCs) into high yields of mature hepatocytes in vitro. Four culturing methods for differentiation [single cell (2D), spheroids (3D), 2D + DLS and 3D + DLS] were studied. To determine the differentiation stages of the MSCs, RT-qPCR of the hepatocyte genes, immunostaining of hepatocyte markers, and functional analyses were all performed. Compared with the other groups, hepatocyte-like cells which differentiated from BM-MSC spheroids on extracellular matrix (ECM) exhibited more intensive staining of stored glycogen, an elevated level of urea biosynthesis and albumin secretion as well as the higher expression of hepatocyte-specific genes. Our results indicated that DLSs combined with spheroidal aggregate culture may be used as an effective method to facilitate the hepatic maturation of BM-MSCs and may have future applications in stem cell-based liver regenerative medicine. PMID:27314916

  15. Enhanced hepatic differentiation of rat bone marrow-derived mesenchymal stem cells in spheroidal aggregate culture on a decellularized liver scaffold.

    PubMed

    Bao, Ji; Wu, Qiong; Wang, Yujia; Li, Yi; Li, Li; Chen, Fei; Wu, Xiujuan; Xie, Mingjun; Bu, Hong

    2016-08-01

    In the present study, we aimed to determine whether the combination of aggregate culture and decellularized liver scaffolds (DLSs) promoted the hepatic differentiation of murine bone marrow-derived mesenchymal stem cells (BM-MSCs) into high yields of mature hepatocytes in vitro. Four culturing methods for differentiation [single cell (2D), spheroids (3D), 2D + DLS and 3D + DLS] were studied. To determine the differentiation stages of the MSCs, RT-qPCR of the hepatocyte genes, immunostaining of hepatocyte markers, and functional analyses were all performed. Compared with the other groups, hepatocyte-like cells which differentiated from BM‑MSC spheroids on extracellular matrix (ECM) exhibited more intensive staining of stored glycogen, an elevated level of urea biosynthesis and albumin secretion as well as the higher expression of hepatocyte-specific genes. Our results indicated that DLSs combined with spheroidal aggregate culture may be used as an effective method to facilitate the hepatic maturation of BM-MSCs and may have future applications in stem cell-based liver regenerative medicine. PMID:27314916

  16. Protective effect of 17β-estradiol on serum deprivation-induced apoptosis and oxidative stress in bone marrow-derived mesenchymal stem cells.

    PubMed

    Mirzamohammadi, S; Mehrabani, M; Tekiyehmaroof, N; Sharifi, A M

    2016-03-01

    Stem cell transplantation has indicated great promise for cell therapy in a wide range of diseases, but poor and insufficient viability of cells within damaged tissues has limited its potential therapeutic effects. 17 β-Estradiol (E2) is a steroid hormone that plays an important role in expression of many genes and regulating proliferation, viability, and intracellular redox status in different cell types. In this study, we aimed to assess the effect of E2 on bone marrow-derived mesenchymal stem cells (BM-MSCs). Apoptosis was induced by serum deprivation (SD), and cells were exposed to E2 in the presence or absence of serum for varying periods of time, after which cell viability was measured by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Expression of proapoptotic and antiapoptotic proteins after exposure to E2 was examined by Western blotting. The ability of E2 to prevent reactive oxygen species (ROS) production was also measured. The results indicated that E2 significantly enhanced the viability of the cells and protected BM-MSCs against SD-induced overproduction of ROS. It could reduce lipid peroxidation, total antioxidant power, and also Bax/Bcl-2 ratio as well as expression of caspase-3. Taken together, our data support that E2 treatment protects BM-MSCs against SD-induced damage by regulating ROS production and upregulation of antiapoptotic/proapoptotic proteins ratio. PMID:25964380

  17. Bone marrow-derived mesenchymal stem cell-secreted IL-8 promotes the angiogenesis and growth of colorectal cancer

    PubMed Central

    Cai, Jianye; Shi, Jianqiang; Sui, Xin; Cao, Yong; Huang, Weijun; Chen, Xiaoyong; Cai, Zijie; Li, Hongyu; Bardeesi, Adham Sameer A.; Zhang, Bin; Liu, Muyun; Song, Wu; Wang, Maosheng; Xiang, Andy Peng

    2015-01-01

    Mesenchymal stem cells (MSCs) have recently been shown to home to tumors and contribute to the formation of the tumor-associated stroma. In addition, MSCs can secrete paracrine factors to facilitate tumor progression. However, the involvement of MSC-derived cytokines in colorectal cancer (CRC) angiogenesis and growth has not been clearly addressed. In this study, we report that interleukin-8 (IL-8) was the most highly upregulated pro-angiogenic factor in MSCs co-cultured with CRC cells and was expressed at substantially higher levels in MSCs than CRC cells. To evaluate the effect of MSC-derived IL-8 on CRC angiogenesis and growth, we used MSCs that expressed small hairpin (interfering) RNAs (shRNA) targeting IL-8 (shIL-8-MSCs). We found that MSC-secreted IL-8 promoted human umbilical vein endothelial cell (HUVEC) proliferation and migration, tube-formation ability and CRC cell proliferation. Additionally, in vivo studies showed that MSCs promoted tumor angiogenesis partially through IL-8. Taken together, these findings suggest that IL-8 secreted by MSCs promotes CRC angiogenesis and growth and can therefore serve as a potential novel therapeutic target. PMID:26517517

  18. Nanoparticle Labeling of Bone Marrow-Derived Rat Mesenchymal Stem Cells: Their Use in Differentiation and Tracking

    PubMed Central

    Akhan, Ece; Tuncel, Donus; Akcali, Kamil C.

    2015-01-01

    Mesenchymal stem cells (MSCs) are promising candidates for cellular therapies due to their ability to migrate to damaged tissue without inducing immune reaction. Many techniques have been developed to trace MSCs and their differentiation efficacy; however, all of these methods have limitations. Conjugated polymer based water-dispersible nanoparticles (CPN) represent a new class of probes because they offer high brightness, improved photostability, high fluorescent quantum yield, and noncytotoxicity comparing to conventional dyes and quantum dots. We aimed to use this tool for tracing MSCs' fate in vitro and in vivo. MSC marker expression, survival, and differentiation capacity were assessed upon CPN treatment. Our results showed that after CPN labeling, MSC markers did not change and significant number of cells were found to be viable as revealed by MTT. Fluorescent signals were retained for 3 weeks after they were differentiated into osteocytes, adipocytes, and chondrocytes in vitro. We also showed that the labeled MSCs migrated to the site of injury and retained their labels in an in vivo liver regeneration model. The utilization of nanoparticle could be a promising tool for the tracking of MSCs in vivo and in vitro and therefore can be a useful tool to understand differentiation and homing mechanisms of MSCs. PMID:25654092

  19. Endothelial Progenitor Cell Fraction Contained in Bone Marrow-Derived Mesenchymal Stem Cell Populations Impairs Osteogenic Differentiation

    PubMed Central

    Duttenhoefer, Fabian; Lara de Freitas, Rafael; Loibl, Markus; Bittermann, Gido; Geoff Richards, R.; Alini, Mauro; Verrier, Sophie

    2015-01-01

    In bone tissue engineering (TE) endothelial cell-osteoblast cocultures are known to induce synergies of cell differentiation and activity. Bone marrow mononucleated cells (BMCs) are a rich source of mesenchymal stem cells (MSCs) able to develop an osteogenic phenotype. Endothelial progenitor cells (EPCs) are also present within BMC. In this study we investigate the effect of EPCs present in the BMC population on MSCs osteogenic differentiation. Human BMCs were isolated and separated into two populations. The MSC population was selected through plastic adhesion capacity. EPCs (CD34+ and CD133+) were removed from the BMC population and the resulting population was named depleted MSCs. Both populations were cultured over 28 days in osteogenic medium (Dex+) or medium containing platelet lysate (PL). MSC population grew faster than depleted MSCs in both media, and PL containing medium accelerated the proliferation for both populations. Cell differentiation was much higher in Dex+ medium in both cases. Real-time RT-PCR revealed upregulation of osteogenic marker genes in depleted MSCs. Higher values of ALP activity and matrix mineralization analyses confirmed these results. Our study advocates that absence of EPCs in the MSC population enables higher osteogenic gene expression and matrix mineralization and therefore may lead to advanced bone neoformation necessary for TE constructs. PMID:26491682

  20. Characteristics and response of mouse bone marrow derived novel low adherent mesenchymal stem cells acquired by quantification of extracellular matrix

    PubMed Central

    Zheng, Ri-Cheng; Heo, Seong-Joo; Koak, Jai-Young; Lee, Joo-Hee; Park, Ji-Man

    2014-01-01

    PURPOSE The aim of present study was to identify characteristic and response of mouse bone marrow (BM) derived low-adherent bone marrow mesenchymal stem cells (BMMSCs) obtained by quantification of extracellular matrix (ECM). MATERIALS AND METHODS Non-adherent cells acquired by ECM coated dishes were termed low-adherent BMMSCs and these cells were analyzed by in vitro and in vivo methods, including colony forming unit fibroblast (CFU-f), bromodeoxyuridine (BrdU), multi-potential differentiation, flow cytometry and transplantation into nude mouse to measure the bone formation ability of these low-adherent BMMSCs. Titanium (Ti) discs with machined and anodized surfaces were prepared. Adherent and low-adherent BMMSCs were cultured on the Ti discs for testing their proliferation. RESULTS The amount of CFU-f cells was significantly higher when non-adherent cells were cultured on ECM coated dishes, which was made by 7 days culturing of adherent BMMSCs. Low-adherent BMMSCs had proliferation and differentiation potential as adherent BMMSCs in vitro. The mean amount bone formation of adherent and low-adherent BMMSCs was also investigated in vivo. There was higher cell proliferation appearance in adherent and low-adherent BMMSCs seeded on anodized Ti discs than machined Ti discs by time. CONCLUSION Low-adherent BMMSCs acquired by ECM from non-adherent cell populations maintained potential characteristic similar to those of the adherent BMMSCs and therefore could be used effectively as adherent BMMSCs in clinic. PMID:25352957

  1. Expression of OCT-4 and SOX-2 in Bone Marrow-Derived Human Mesenchymal Stem Cells during Osteogenic Differentiation

    PubMed Central

    Matic, Igor; Antunovic, Maja; Brkic, Sime; Josipovic, Pavle; Mihalic, Katarina Caput; Karlak, Ivan; Ivkovic, Alan; Marijanovic, Inga

    2016-01-01

    AIM: Determine the levels of expression of pluripotency genes OCT-4 and SOX-2 before and after osteogenic differentiation of human mesenchymal stem cells (hMSCs). METHODS: Human MSCs were derived from the bone marrow and differentiated into osteoblasts. The analyses were performed on days 0 and 14 of the cell culture. In vitro differentiation was evaluated due to bone markers – alkaline phosphatase (AP) activity and the messenger RNA (mRNA) expression of AP and bone sialoprotein (BSP). The OCT-4 and SOX-2 expression was evaluated at mRNA level by real-time qPCR and at protein level by immunocytochemistry. RESULTS: In vitro cultures on day 14 showed an increase in AP activity and upregulation of AP and BSP gene expression. OCT-4 and SOX-2 in undifferentiated hMSCs on day 0 is detectable and very low compared to tumor cell lines as a positive control. Immunocytochemistry detected OCT-4 in the cell nuclei prior (day 0) and post differentiation (day 14). On the same time points, cultures were negative for SOX-2 protein. CONCLUSION: Messenger RNA for pluripotency markers OCT-4 and SOX-2 isolated from hMSCs was less present, while OCT-4 protein was detected in cell nuclei prior and post differentiation into osteoblast lineage. PMID:27275321

  2. Assessment of the role of flavonoids for inducing osteoblast differentiation in isolated mouse bone marrow derived mesenchymal stem cells.

    PubMed

    Srivastava, Swati; Bankar, Rohini; Roy, Partha

    2013-06-15

    Quercetin and rutin are common flavonoids in fruit and vegetables, and have been reported to affect bone development. However, the effect of flavonoids on osteoblast differentiation remains a matter of controversy. In the present study, mouse bone marrow mesenchymal stem cells (BMMSCs) were isolated and characterized for their use in osteoblast differentiation using two flavonoids, quercetin and rutin. BMMSCs were cultured in various concentrations of quercetin and rutin during the osteoblast differentiation period of 10 days. Both quercetin and rutin were found to up regulate the osteoblast differentiation in dose dependent manner, albeit to lesser extent in case of former than that of latter. Quercetin and rutin also increased alkaline phosphatase activity by about 150 and 240% and demonstrated mineralization up to 110 and 200% respectively as compared to control (which was considered as 100%). Further, both the flavonoids were also found to increase the expression of some of the prominent markers for differentiation of osteoblast like osteopontin, osterix, RunX2, osteoprotegerin and osteocalcin. The current data suggests that certain classes of flavonoids like rutin and quercetin can be used in the cure and management of osteodegenerative disorders due to their osteoblast specific differentiation activities. PMID:23570998

  3. The effect of magnetic field during freezing and thawing of rat bone marrow-derived mesenchymal stem cells.

    PubMed

    Shikata, H; Kaku, M; Kojima, S-I; Sumi, H; Kojima, S-T; Yamamoto, T; Yashima, Y; Kawata, T; Tanne, K; Tanimoto, K

    2016-08-01

    Previous studies showed that a programmed freezer with magnetic field can maintain a high survival rate of mesenchymal stem cells (MSCs). The purpose of this study was to evaluate the influences of magnetic field during freezing and thawing on the survival of MSCs isolated from rat bone marrow. The cells were frozen by a normal programmed freezer or a programmed freezer with magnetic field (CAS-LAB1) and cryopreserved for 7 days at -150 °C. Then, the cells were thawed in the presence or absence of magnetic field. Immediately after thawing, the number of surviving or viable cells was counted. The cell proliferation was examined after 1-week culture. Cryopreserved MSCs which were frozen by a normal freezer or a CAS freezer were transplanted into bone defects artificially made in calvaria of 4-week-old rats. Non-cryopreserved MSCs were used as a control. The rats were sacrificed at 8, 16, or 24 weeks after transplantation and the bone regeneration area was measured. Proliferation rates of MSCs after 1 week were significantly higher in the CAS-freezing-thawing group than in the CAS-freezing group. The extent of new bone formation in the CAS-freezing-thawing group tended to be larger than in CAS-freezing group 24 weeks after transplantation. These results suggest that a magnetic field enhances cell survival during thawing as well as freezing. PMID:27346603

  4. The synergistic effect of VEGF and biomorphic silicon carbides topography on in vivo angiogenesis and human bone marrow derived mesenchymal stem cell differentiation.

    PubMed

    Díaz-Rodríguez, P; Gómez-Amoza, J L; Landin, M

    2015-08-01

    Topographical features of biomaterials are able to modulate cell attachment, spreading and differentiation. The addition of growth factors to implantable biomaterials can modify these cellular responses, enhancing their therapeutic potential. The aim of this research is to establish the influence of biomorphic silicon carbide ceramics (bioSiCs) surface topography on the proliferation and osteoblastic differentiation of mesenchymal stem cells and the potential synergistic effect of the ceramic porous structure together with vascular endothelial growth factor loading (VEGF) on the surface mediated osteoblastic differentiation. Three porous bioSiCs with important differences in their microstructure were obtained from different natural precursors. Samples loaded with or without VEGF through ionic interactions were cultured with human umbilical vein endothelial cells (HUVEC) or bone marrow derived mesenchymal stem cells (hMSCs). Cell behaviour and protein activity with regard to bioSiC porous structure and surface properties were analysed. An in vivo model (Chick Chorioallantoic Membrane; CAM) was used to assess the capability of the VEGF loaded systems to promote angiogenesis. Experimental data show that loaded systems were able to control the release of VEGF for up to 15 d ensuring the activity of the protein, increasing the proliferation of HUVECs and the formation of new blood vessels in the CAM. It was found that the selection of bioSiCs with a higher pore size promoted a higher concentration of osteoblastic differentiation markers of MSCs cultured on the surface of bioSiCs. Furthermore, the addition of VEGF to the systems was able to promote a faster osteoblastic differentiation according to the qPCR results, suggesting a synergy between both the surface properties and the controlled release of the growth factor. The VEGF loaded sapelli bioSiC was found to be the most promising material for bone tissue engineering applications. PMID:26238485

  5. Contact-dependent abrogation of bone marrow-derived plasmacytoid dendritic cell differentiation by murine mesenchymal stem cells.

    PubMed

    Hackstein, Holger; Tschipakow, Inna; Bein, Gregor; Nold, Philipp; Brendel, Cornelia; Baal, Nelli

    2016-07-15

    Plasmacytoid dendritic cells (pDCs) are rare central regulators of antiviral immunity and unsurpassed producers of interferon-α (IFN-α). Despite their crucial role as a link between innate and adaptive immunity, little is known about the modulation of pDC differentiation by other bone marrow (BM) cells. In this study, we investigated the modulation of pDC differentiation in Flt-3 ligand (Flt3L)-supplemented BM cultures, using highly purified mesenchymal stem cells (MSCs) that were FACS-isolated from murine BM based on surface marker expression and used after in vitro expansion. Initial analysis revealed an almost complete inhibition of BM-derived pDC expansion in the presence of >2% MSC. This inhibition was cell contact-dependent and soluble factor-independent, as indicated by trans-well experiments. The abrogation of functional pDC development by MSCs was confirmed after TLR9 stimulation, revealing a complete, contact-dependent suppression of the IFN-a producing capacity of pDCs in Flt3L MSC BM co-cultures. MSC selectively inhibited pDC development in contrast to myeloid DC development, as indicated by the significantly increased numbers of myeloid DC in Flt3L-supplemented BM cultures. The absence of significant MSC-mediated inhibitory effects on myeloid DC differentiation was confirmed by additional experiments in GM-CSF/IL-4-supplemented BM cultures. In summary, we describe a novel contact-dependent immunomodulatory mechanism of MSC that targets the BM-derived expansion of functional pDCs. PMID:27233615

  6. Growth and osteogenic differentiation of alveolar human bone marrow-derived mesenchymal stem cells on chitosan/hydroxyapatite composite fabric.

    PubMed

    Kim, Beom-Su; Kim, Jin Seong; Chung, Yong Sik; Sin, Yeon-Woo; Ryu, Kyung-Ho; Lee, Jun; You, Hyung-Keun

    2013-06-01

    Scaffolds can be used for tissue engineering because they can serve as templates for cell adhesion and proliferation for tissue repair. In this study, chitosan/hydroxyapatite (CS/HAp) composites were prepared by coprecipitation synthesis. Then, CS and CS/HAp fabrics were prepared by wet spinning. CS fibers with a diameter of 15 ± 1.3 μm and CS/HAp fibers with a diameter of 22 ± 1.2 μm were successfully produced; incorporation of HAp into the CS/HAp fibers was confirmed by X-ray diffraction analysis. Biological in vitro evaluations showed that human mesenchymal stem cells (hMSCs) cultured on CS/HAp fabric showed increased proliferation compared to those cultured on pure CS fabric, which was observed using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay, DNA content assay, and [(3) H] thymidine incorporation assay. Neither the CS nor CS/HAp scaffold exhibited any cytotoxicity to hMSCs, as shown by viability staining and cytotoxicity fluorescence image assays. After 10 days of culturing, the attachment of cells onto the scaffold was observed by scanning electron microscopy. Furthermore, under osteogenic differentiation conditions, alkaline phosphatase (ALP) activity and calcium accumulation was higher in cells cultured on the CS/HAp scaffold than in cells cultured on the CS scaffold. The mRNA expression of osteoblast markers, including ALP, osteocalcin, Co1Ia1, and runt-related transcription factor 2, was higher in cells cultured on CS/HAp than in cells cultured on the CS fabric. The results of this study indicate that the CS/HAp composite fabric may serve as a good scaffold for bone tissue engineering applications. PMID:23135904

  7. Hepatocyte Growth Factor Modification Enhances the Anti-Arrhythmic Properties of Human Bone Marrow-Derived Mesenchymal Stem Cells

    PubMed Central

    Du, Wei; Yu, Yi-Chao; Ju, Wei-Zhu; Man, Yi-Long; Li, Xiao-Rong; Chen, Yan; Wang, Zi-Dun; Gu, Wei-Juan; Zhang, Feng-Xiang; Wang, Hua; Wu, Chu-Tse; Cao, Ke-Jiang

    2014-01-01

    Background/Aims Chronic myocardial infarction (MI) results in the formation of arrhythmogenic substrates, causing lethal ventricular arrhythmia (VA). We aimed to determine whether mesenchymal stem cells (MSCs) carrying a hepatocyte growth factor (HGF) gene modification (HGF-MSCs) decrease the levels of arrhythmogenic substrates and reduce the susceptibility to developing VA compared with unmodified MSCs and PBS in a swine infarction model. Methods The left descending anterior artery was balloon-occluded to establish an MI model. Four weeks later, the randomly grouped pigs were administered MSCs, PBS or HGF-MSCs via thoracotomy. After an additional four weeks, dynamic electrocardiography was performed to assess heart rate variability, and programmed electrical stimulation was conducted to evaluate the risk for VA. Then, the pigs were euthanized for morphometric, immunofluorescence and western blot analyses. Results: The HGF-MSC group displayed the highest vessel density and Cx43 expression levels, and the lowest levels of apoptosis, and tyrosine hydroxylase (TH) and growth associated protein 43 (GAP43) expression. Moreover, the HGF-MSC group exhibited a decrease in the number of sympathetic nerve fibers, substantial decreases in the low frequency and the low-/high- frequency ratio and increases in the root mean square of successive differences (rMSSD) and the percentage of successive normal sinus R-R intervals longer than 50 ms (pNN50), compared with the other two groups. Finally, the HGF-MSC group displayed the lowest susceptibility to developing VA. Conclusion HGF-MSCs displayed potent antiarrhythmic effects, reducing the risk for VA. PMID:25360679

  8. Mitochondrial Function and Energy Metabolism in Umbilical Cord Blood- and Bone Marrow-Derived Mesenchymal Stem Cells

    PubMed Central

    Palomäki, Sami; Lehtonen, Siri; Ritamo, Ilja; Valmu, Leena; Nystedt, Johanna; Laitinen, Saara; Leskelä, Hannnu-Ville; Sormunen, Raija; Pesälä, Juha; Nordström, Katrina; Vepsäläinen, Ari; Lehenkari, Petri

    2012-01-01

    Human mesenchymal stem cells (hMSCs) are an attractive choice for a variety of cellular therapies. hMSCs can be isolated from many different tissues and possess unique mitochondrial properties that can be used to determine their differentiation potential. Mitochondrial properties may possibly be used as a quality measure of hMSC-based products. Accordingly, the present work focuses on the mitochondrial function of hMSCs from umbilical cord blood (UCBMSC) cells and bone marrow cells from donors younger than 18 years of age (BMMSC <18) and those more than 50 years of age (BMMSC >50). Changes of ultrastructure and energy metabolism during osteogenic differentiation in all hMSC types were studied in detail. Results show that despite similar surface antigen characteristics, the UCBMSCs had smaller cell surface area and possessed more abundant rough endoplasmic reticulum than BMMSC >50. BMMSC <18 were morphologically more UCBMSC-like. UCBMSC showed dramatically higher mitochondrial-to-cytoplasm area ratio and elevated superoxide and manganese superoxide dismutase (MnSOD) levels as compared with BMMSC >50 and BMMSC <18. All hMSCs types showed changes indicative of mitochondrial activation after 2 weeks of osteogenic differentiation, and the increase in mitochondrial-to-cytoplasm area ratio appears to be one of the first steps in the differentiation process. However, BMMSC >50 showed a lower level of mitochondrial maturation and differentiation capacity. UCBMSCs and BMMSCs also showed a different pattern of exocytosed proteins and glycoproteoglycansins. These results indicate that hMSCs with similar cell surface antigen expression have different mitochondrial and functional properties, suggesting different maturation levels and other significant biological variations of the hMSCs. Therefore, it appears that mitochondrial analysis presents useful characterization criteria for hMSCs intended for clinical use. PMID:21615273

  9. Low Molecular Weight Fraction of Commercial Human Serum Albumin Induces Morphologic and Transcriptional Changes of Bone Marrow-Derived Mesenchymal Stem Cells.

    PubMed

    Bar-Or, David; Thomas, Gregory W; Rael, Leonard T; Gersch, Elizabeth D; Rubinstein, Pablo; Brody, Edward

    2015-08-01

    Osteoarthritis (OA) is the most common chronic disease of the joint; however, the therapeutic options for severe OA are limited. The low molecular weight fraction of commercial 5% human serum albumin (LMWF5A) has been shown to have anti-inflammatory properties that are mediated, in part, by a diketopiperazine that is present in the albumin preparation and that was demonstrated to be safe and effective in reducing pain and improving function when administered intra-articularly in a phase III clinical trial. In the present study, bone marrow-derived mesenchymal stem cells (BMMSCs) exposed to LMWF5A exhibited an elongated phenotype with diffuse intracellular F-actin, pronounced migratory leading edges, and filopodia-like projections. In addition, LMWF5A promoted chondrogenic condensation in "micromass" culture, concurrent with the upregulation of collagen 2α1 mRNA. Furthermore, the transcription of the CXCR4-CXCL12 axis was significantly regulated in a manner conducive to migration and homing. Several transcription factors involved in stem cell differentiation were also found to bind oligonucleotide response element probes following exposure to LMWF5A. Finally, a rapid increase in PRAS40 phosphorylation was observed following treatment, potentially resulting in the activation mTORC1. Proteomic analysis of synovial fluid taken from a preliminary set of patients indicated that at 12 weeks following administration of LMWF5A, a microenvironment exists in the knee conducive to stem cell infiltration, self-renewal, and differentiation, in addition to indications of remodeling with a reduction in inflammation. Taken together, these findings imply that LMWF5A treatment may prime stem cells for both mobilization and chondrogenic differentiation, potentially explaining some of the beneficial effects achieved in clinical trials. PMID:26041739

  10. Suppression of Evi1 promotes the osteogenic differentiation and inhibits the adipogenic differentiation of bone marrow-derived mesenchymal stem cells in vitro.

    PubMed

    An, Qijun; Wu, Dou; Ma, Yuehong; Zhou, Biao; Liu, Qiang

    2015-12-01

    Osteoporosis (OP) is considered a complex disease with a strong genetic impact, mainly affecting post-menopausal women and is also a common cause of fracture. Elucidating the molecular mechanisms that regulate the osteogenic differentiation of bone marrow-derived mesenchymal stem cells (BMSCs) is crucial to developing treatment strategies to combat OP. In the present study, we found that ectopic viral integration site‑1 (Evi1) was highly expressed during the process of adipogenesis of rat BMSCs. Notably, Evi1 levels markedly increased on day 3 of adipogenic differentiation following the addition of adipogenic induction supplements. In addition, we interfered with the expression of the Evi1 gene in the adipogenesis of BMSCs by supplementing adenoviral plasmids and measured the expression levels of bone sialoprotein (BSP), osteocalcin (OCN), osteopontin (OPN), peroxisome proliferator‑activated receptor γ2 (PPARγ2) and lipoprotein lipase (LPL) by RT-qPCR and western blot analysis. The mRNA and protein levels of osteogenic and adipogenic markers in the BMSCs were up‑ and downregulated, respectively following the silencing of siEvi1. Our experimental results substantiate that the suppression of Evi1 in BMSCs by RNA interference inhibits adipogenic differentiation, while it promotes osteogenic differentiation. The results from our study demonstrated that the Evi1 gene may be targeted as a therapeutic strategy for promoting bone formation. PMID:26497332

  11. Enhancement of tendon-to-bone healing after anterior cruciate ligament reconstruction using bone marrow-derived mesenchymal stem cells genetically modified with bFGF/BMP2

    PubMed Central

    Chen, Biao; Li, Bin; Qi, Yong-Jian; Ni, Qu-Bo; Pan, Zheng-Qi; Wang, Hui; Chen, Liao-Bin

    2016-01-01

    Many strategies, including various growth factors and gene transfer, have been used to augment healing after anterior cruciate ligament (ACL) reconstruction. The biological environment regulated by the growth factors during the stage of tendon-bone healing was considered important in controlling the integrating process. The purpose of this study was to evaluate the effects of bone marrow-derived mesenchymal stem cells (BMSCs) genetically modified with bone morphogenetic protein 2 (BMP2) and basic fibroblast growth factor (bFGF) on healing after ACL reconstruction. BMSCs were infected with an adenoviral vector encoding BMP2 (AdBMP2) or bFGF (AdbFGF). Then, the infected BMSCs were surgically implanted into the tendon-bone interface. At 12 weeks postoperatively, the formation of abundant cartilage-like cells, smaller tibial bone tunnel and significantly higher ultimate load and stiffness levels, through histological analysis, micro-computed tomography and biomechanical testing, were observed. In addition, the AdBMP2-plus-AdbFGF group had the smallest bone tunnel and the best mechanical properties among all the groups. The addition of BMP2 or bFGF by gene transfer resulted in better cellularity, new bone formation and higher mechanical property, which contributed to the healing process after ACL reconstruction. Furthermore, the co-application of these two genes was more powerful and efficient than either single gene therapy. PMID:27173013

  12. Eicosapentaenoic acid attenuates dexamethasome-induced apoptosis by inducing adaptive autophagy via GPR120 in murine bone marrow-derived mesenchymal stem cells

    PubMed Central

    Gao, B; Han, Y-H; Wang, L; Lin, Y-J; Sun, Z; Lu, W-G; Hu, Y-Q; Li, J-Q; Lin, X-S; Liu, B-H; Jie, Q; Yang, L; Luo, Z-J

    2016-01-01

    Long-term use of glucocorticoids is a widespread clinical problem, which currently has no effective solution other than discontinuing the use. Eicosapentaenoic acid (EPA), an omega-3 long chain polyunsaturated fatty acid (n-3 PUFA), which is largely contained in fish or fish oil, has been reported to promote cell viability and improve bone metabolism. However, little is known about the effects of EPA on dexamethasome (Dex)-induced cell apoptosis. In this study, we showed that EPA-induced autophagy of murine bone marrow-derived mesenchymal stem cells (mBMMSCs). Meanwhile, EPA, but not arachidonic acid (AA), markedly inhibited Dex-induced apoptosis and promoted the viability of mBMMSCs. We also observed that EPA-induced autophagy was modulated by GPR120, but not GPR40. Further experiments showed that the mechanism of EPA-induced autophagy associated with GPR120 modulation involved an increase in the active form of AMP-activated protein kinase and a decrease in the activity of mammalian target of RAPA. The protective effect of EPA on Dex-induced apoptosis via GPR120-meditated induction of adaptive autophagy was supported by in vivo experiments. In summary, our findings may have important implications in developing future strategies to use EPA in the prevention and therapy of the side effects induced by long-term Dex-abuse. PMID:27228350

  13. Biological Response of Human Bone Marrow-Derived Mesenchymal Stem Cells to Commercial Tantalum Coatings with Microscale and Nanoscale Surface Topographies

    NASA Astrophysics Data System (ADS)

    Skoog, Shelby A.; Kumar, Girish; Goering, Peter L.; Williams, Brian; Stiglich, Jack; Narayan, Roger J.

    2016-06-01

    Tantalum is a promising orthopaedic implant coating material due to its robust mechanical properties, corrosion resistance, and excellent biocompatibility. Previous studies have demonstrated improved biocompatibility and tissue integration of surface-treated tantalum coatings compared to untreated tantalum. Surface modification of tantalum coatings with biologically inspired microscale and nanoscale features may be used to evoke optimal tissue responses. The goal of this study was to evaluate commercial tantalum coatings with nanoscale, sub-microscale, and microscale surface topographies for orthopaedic and dental applications using human bone marrow-derived mesenchymal stem cells (hBMSCs). Tantalum coatings with different microscale and nanoscale surface topographies were fabricated using a diffusion process or chemical vapor deposition. Biological evaluation of the tantalum coatings using hBMSCs showed that tantalum coatings promote cellular adhesion and growth. Furthermore, hBMSC adhesion to the tantalum coatings was dependent on surface feature characteristics, with enhanced cell adhesion on sub-micrometer- and micrometer-sized surface topographies compared to hybrid nano-/microstructures. Nanostructured and microstructured tantalum coatings should be further evaluated to optimize the surface coating features to promote osteogenesis and enhance osseointegration of tantalum-based orthopaedic implants.

  14. Bone marrow derived mesenchymal stem cells alleviated brain injury via down-regulation of interleukin-1β in focal cerebral ischemic rats

    PubMed Central

    Zhao, Yansong; Wang, Xiaoli; Dong, Peng; Xu, Qinyan; Ma, Ze; Mu, Qingjie; Sun, Xihe; Jiang, Zhengchen; Wang, Xin

    2016-01-01

    Interleukin-1β (IL-1β) plays an important role in brain injury after focal ischemia, and bone marrow-derived mesenchymal stem cells (BMSCs) are capable of reducing the expression of IL-1β, we investigated the effects of BMSCs transplantation on brain edema and cerebral infarction as well as the underlying mechanisms via IL-1β. Male Sprague-Dawley rats were randomly divided into five groups: Normal + phosphate-buffered saline (PBS), middle cerebral artery occlusion (MCAO) + PBS, Normal + BMSCs, MCAO + BMSCs and MCAO + IL-1ra (an antagonist of IL-1β). BMSCs were transplanted 24 hours after MCAO, and brain edema was evaluated by Magnetic Resonance Imaging (MRI) and brain water content method after BMSCs transplantation. The expression of NeuN and AQP4 was analyzed by immunofluorescence staining. Protein level of AQP4 and IL-1β was detected by western blot analysis 48 hours after transplantation. The results showed that BMSCs transplantation reduced brain edema by measurement of brain water content and ADC Value of MRI, as well as the expression of AQP4 and IL-1β. It was also found that BMSCs transplantation could alleviate the cerebral infarction volume and neuronal damage. Both the brain edema and the cerebral infarction were associated with IL-1β expression. In conclusion, BMSCs transplantation was capable of alleviating brain edema as well as reducing cerebral infarction via down-regulation of IL-1β expression, thus repair the injured brain in focal cerebral ischemic rats.

  15. Novel biomimetic tripolymer scaffolds consisting of chitosan, collagen type 1, and hyaluronic acid for bone marrow-derived human mesenchymal stem cells-based bone tissue engineering.

    PubMed

    Mathews, Smitha; Bhonde, Ramesh; Gupta, Pawan Kumar; Totey, Satish

    2014-11-01

    Human bone marrow-derived mesenchymal stem cells (hMSCs) are an ideal osteogenic cell source for bone tissue engineering (BTE). A scaffold, in the context of BTE, is the extracellular matrix (ECM) that provides the unique microenvironment and play significant role in regulating cell behavior, differentiation, and development in an in vitro culture system. In this study, we have developed novel biomimetic tripolymer scaffolds for BTE using an ECM protein, collagen type 1; an ECM glycosaminoglycan, hyaluronic acid; and a natural osteoconductive polymer, chitosan. The scaffolds were characterized by scanning electron microscopy (SEM) and swelling ratio. The scaffolds were seeded with hMSCs and tested for cytocompatibility and osteogenic potential. The scaffolds supported cell adhesion, enhanced cell proliferation, promoted cell migration, showed good cell viability, and osteogenic potential. The cells were able to migrate out from the scaffolds in favorable conditions. SEM, alkaline phosphatase assay, and immunofluorescent staining confirmed the differentiation of hMSCs to osteogenic lineage in the scaffolds. In conclusion, we have successfully developed biomimetic scaffolds that supported the proliferation and differentiation of hMSCs. These scaffolds hold great promise as a cell-delivery vehicle for regenerative therapies and as a support system for enhancing bone regeneration. PMID:24723571

  16. The Effect of Bone Marrow-Derived Mesenchymal Stem Cells and Their Conditioned Media Topically Delivered in Fibrin Glue on Chronic Wound Healing in Rats.

    PubMed

    Mehanna, Radwa A; Nabil, Iman; Attia, Noha; Bary, Amany A; Razek, Khalid A; Ahmed, Tamer A E; Elsayed, Fatma

    2015-01-01

    Bone marrow-derived mesenchymal stem cells (BM-MSCs) represent a modern approach for management of chronic skin injuries. In this work, we describe BM-MSCs application versus their conditioned media (CM) when delivered topically admixed with fibrin glue to enhance the healing of chronic excisional wounds in rats. Fifty-two adult male rats were classified into four groups after induction of large-sized full-thickness skin wound: control group (CG), fibrin only group (FG), fibrin + MSCs group (FG + SCs), and fibrin + CM group (FG + CM). Healing wounds were evaluated functionally and microscopically. Eight days after injury, number of CD68+ macrophages infiltrating granulation tissue was considerably higher in the latter two groups. Although--later--none of the groups depicted a substantially different healing rate, the quality of regenerated skin was significantly boosted by the application of either BM-MSCs or their CM both (1) structurally as demonstrated by the obviously increased mean area percent of collagen fibers in Masson's trichrome-stained skin biopsies and (2) functionally as supported by the interestingly improved epidermal barrier as well as dermal tensile strength. Thus, we conclude that topically applied BM-MSCs and their CM-via fibrin vehicle--could effectively improve the quality of healed skin in chronic excisional wounds in rats, albeit without true acceleration of wound closure. PMID:26236740

  17. A novel antagonist of CXCR4 prevents bone marrow-derived mesenchymal stem cell-mediated osteosarcoma and hepatocellular carcinoma cell migration and invasion.

    PubMed

    Fontanella, Raffaela; Pelagalli, Alessandra; Nardelli, Anna; D'Alterio, Crescenzo; Ieranò, Caterina; Cerchia, Laura; Lucarelli, Enrico; Scala, Stefania; Zannetti, Antonella

    2016-01-01

    Recent findings suggest that bone marrow-derived mesenchymal stem cells (BM-MSCs) are recruited into the microenvironment of developing tumors, where they contribute to metastatic processes. The aim of this study was to investigate the role of BM-MSCs in promoting osteosarcoma and hepatocellular carcinoma cell progression in vitro and the possible mechanisms involved in these processes. U2OS and SNU-398 are osteosarcoma and hepatocellular carcinoma cell lines, respectively, that can be induced to proliferate when cultured in the presence of BM-MSCs. To determine the effect of BM-MSCs on U2OS and SNU-398 cells, the AKT and ERK signaling pathways were investigated, and increases were observed in active P-Akt and P-Erk forms. Moreover, BM-MSCs caused an increase in tumor cell migration and invasion that was derived from the enhancement of CXCR4 levels. Thus, when tumor cells were treated with the CXCR4 antagonist AMD3100, a reduction in their migration and invasion was observed. Furthermore, a new CXCR4 inhibitor, Peptide R, which was recently developed as an anticancer agent, was used to inhibit BM-MSC-mediated tumor invasion and to overcome AMD3100 toxicity. Taken together, these results suggest that inhibiting CXCR4 impairs the cross-talk between tumor cells and BM-MSCs, resulting in reduced metastatic potential in osteosarcoma and hepatocellular carcinoma cells. PMID:26517945

  18. The Effect of Bone Marrow-Derived Mesenchymal Stem Cells and Their Conditioned Media Topically Delivered in Fibrin Glue on Chronic Wound Healing in Rats

    PubMed Central

    Mehanna, Radwa A.; Nabil, Iman; Attia, Noha; Bary, Amany A.; Razek, Khalid A.; Ahmed, Tamer A. E.; Elsayed, Fatma

    2015-01-01

    Bone marrow-derived mesenchymal stem cells (BM-MSCs) represent a modern approach for management of chronic skin injuries. In this work, we describe BM-MSCs application versus their conditioned media (CM) when delivered topically admixed with fibrin glue to enhance the healing of chronic excisional wounds in rats. Fifty-two adult male rats were classified into four groups after induction of large-sized full-thickness skin wound: control group (CG), fibrin only group (FG), fibrin + MSCs group (FG + SCs), and fibrin + CM group (FG + CM). Healing wounds were evaluated functionally and microscopically. Eight days after injury, number of CD68+ macrophages infiltrating granulation tissue was considerably higher in the latter two groups. Although—later—none of the groups depicted a substantially different healing rate, the quality of regenerated skin was significantly boosted by the application of either BM-MSCs or their CM both (1) structurally as demonstrated by the obviously increased mean area percent of collagen fibers in Masson's trichrome-stained skin biopsies and (2) functionally as supported by the interestingly improved epidermal barrier as well as dermal tensile strength. Thus, we conclude that topically applied BM-MSCs and their CM—via fibrin vehicle—could effectively improve the quality of healed skin in chronic excisional wounds in rats, albeit without true acceleration of wound closure. PMID:26236740

  19. Long-Term Effects of Bone Marrow-Derived Mesenchymal Stem Cells in Dextran Sulfate Sodium-Induced Murine Chronic Colitis

    PubMed Central

    Lee, Hyun Jung; Oh, Sun-Hee; Jang, Hui Won; Kwon, Ji-Hee; Lee, Kyoung Jin; Kim, Chung Hee; Park, Soo Jung; Hong, Sung Pil; Cheon, Jae Hee; Kim, Tae Il; Kim, Won Ho

    2016-01-01

    Background/Aims Bone marrow-derived mesenchymal stem cells (BM-MSCs) have shown beneficial effects in experimental colitis models, but the underlying mechanisms are not fully understood. We investigated the long-term effects of BM-MSCs, particularly in mice with chronic colitis. Methods Chronic colitis was induced by administering 3% dextran sulfate sodium (DSS) in a series of three cycles. BM-MSCs were injected intravenously into DSS-treated mice three times during the first cycle. On day 33, the therapeutic effects were evaluated with clinicopathologic profiles and histological scoring. Inflammatory mediators were measured with real-time polymerase chain reaction. Results Systemic infusion of BM-MSCs ameliorated the severity of colitis, and body weight restoration was significantly promoted in the BM-MSC-treated mice. In addition, BM-MSC treatment showed a sustained beneficial effect throughout the three cycles. Microscopic examination revealed that the mice treated with BM-MSCs had fewer inflammatory infiltrates, a lesser extent of inflammation, and less crypt structure damage compared with mice with DSS-induced colitis. Anti-inflammatory cytokine levels of interleukin-10 were significantly increased in the inflamed colons of BM-MSC-treated mice compared with DSS-induced colitis mice. Conclusions Systemic infusion of BM-MSCs at the onset of disease exerted preventive and rapid recovery effects, with long-term immunosuppressive action in mice with repeated DSS-induced chronic colitis. PMID:27114436

  20. Promoting Effects on Proliferation and Chondrogenic Differentiation of Bone Marrow-Derived Mesenchymal Stem Cells by Four “Kidney-Tonifying” Traditional Chinese Herbs

    PubMed Central

    Cai, Bin; Zhang, Ai-guo; Zhang, Xian; Ge, Wen-jie; Dai, Guo-da; Tan, Xiang-ling; Roodrajeetsing, Gopaul; Cai, Jian-ping

    2015-01-01

    Traditional Chinese medicine can promote the proliferation of bone marrow-derived mesenchymal stem cells (BMSCs). We chose four “Kidney-tonifying” Chinese herbal medicines, Radix Astragali, Salvia, Herba Epimedii, and Saussurea Involucrata, to evaluate whether they had positive effects on the proliferation of BMSCs and TGF-β1-induced chondrogenic differentiation of BMSCs. The four Chinese herbal medicines were intragastrically administered to Sprague-Dawley rats, respectively, to prepare drug-containing serums of corresponding Chinese herbs. BMSCs were isolated, cultured, and exposed to culture solution containing 1%, 5%, 10%, and 15% (v/v) Radix Astragali-, Salvia-, Herba Epimedii-, and Saussurea Involucrata-containing serum, respectively. TGF-β1-induced BMSCs were addressed in the same manner. Collagen type II protein was assessed by immunofluorescence methods. To assess whether the drug-containing serums had positive effects on the proliferation of BMSCs and TGF-β1-induced BMSCs, MTT method was assessed. The proliferation of BMSCs was significantly enhanced when exposed to culture solutions containing 1% and 5% Radix Astragali-, 1% and 5% Salvia-, 5% Herba Epimedii-, and 1%, 5%, and 10% Saussurea Involucrata-containing serum. The proliferation of TGF-β1-induced BMSCs was significantly enhanced when exposed to 1%, 5%, and 15% Radix Astragali-, 10% and 15% Salvia-, 5%, and 15% Herba Epimedii-, and 1%, 5%, and 10% Saussurea Involucrata-containing serum. PMID:26137494

  1. Glucose-Dependent Insulinotropic Peptide Prevents Serum Deprivation-Induced Apoptosis in Human Bone Marrow-Derived Mesenchymal Stem Cells and Osteoblastic Cells.

    PubMed

    Berlier, J L; Kharroubi, I; Zhang, J; Dalla Valle, A; Rigutto, S; Mathieu, M; Gangji, V; Rasschaert, J

    2015-12-01

    Human bone marrow-derived mesenchymal stem cells (hBMSC) are able to differentiate into cells of connective tissue lineages, including bone and cartilage. They are therefore considered as a promising tool for the treatment of bone degenerative diseases. One of the major issues in regenerative cell therapy is the biosafety of fetal bovine serum used for cell culture. Therefore, the development of a culture medium devoid of serum but preserving hBMSC viability will be of clinical value. The glucose-dependent insulinotropic peptide (GIP) has an anti-apoptotic action in insulin-producing cells. Interestingly, GIP also exerts beneficial effects on bone turnover by acting on osteoblasts and osteoclasts. We therefore evaluated the ability of GIP to prevent cell death in osteoblastic cells cultured in serum-free conditions. In hBMSC and SaOS-2 cells, activation of the GIP receptor increased intracellular cAMP levels. Serum deprivation induced apoptosis in SaOS-2 and hBMSC that was reduced by 30 and 50 %, respectively, in the presence of GIP. The protective effect of GIP involves activation of the adenylate cyclase pathway and inhibition of caspases 3/7 activation. These findings demonstrate that GIP exerts a protective action against apoptosis in hBMSC and suggest a novel approach to preserve viability of hBMSC cultured in the absence of serum. PMID:26254594

  2. Eicosapentaenoic acid attenuates dexamethasome-induced apoptosis by inducing adaptive autophagy via GPR120 in murine bone marrow-derived mesenchymal stem cells.

    PubMed

    Gao, B; Han, Y-H; Wang, L; Lin, Y-J; Sun, Z; Lu, W-G; Hu, Y-Q; Li, J-Q; Lin, X-S; Liu, B-H; Jie, Q; Yang, L; Luo, Z-J

    2016-01-01

    Long-term use of glucocorticoids is a widespread clinical problem, which currently has no effective solution other than discontinuing the use. Eicosapentaenoic acid (EPA), an omega-3 long chain polyunsaturated fatty acid (n-3 PUFA), which is largely contained in fish or fish oil, has been reported to promote cell viability and improve bone metabolism. However, little is known about the effects of EPA on dexamethasome (Dex)-induced cell apoptosis. In this study, we showed that EPA-induced autophagy of murine bone marrow-derived mesenchymal stem cells (mBMMSCs). Meanwhile, EPA, but not arachidonic acid (AA), markedly inhibited Dex-induced apoptosis and promoted the viability of mBMMSCs. We also observed that EPA-induced autophagy was modulated by GPR120, but not GPR40. Further experiments showed that the mechanism of EPA-induced autophagy associated with GPR120 modulation involved an increase in the active form of AMP-activated protein kinase and a decrease in the activity of mammalian target of RAPA. The protective effect of EPA on Dex-induced apoptosis via GPR120-meditated induction of adaptive autophagy was supported by in vivo experiments. In summary, our findings may have important implications in developing future strategies to use EPA in the prevention and therapy of the side effects induced by long-term Dex-abuse. PMID:27228350

  3. 15-Deoxy-Δ12,14-Prostaglandin J2 Inhibits Homing of Bone Marrow-Derived Mesenchymal Stem Cells Triggered by Chronic Liver Injury via Redox Pathway

    PubMed Central

    Liu, Xin; Jia, Shuangshuang; Li, Weiyang; Yang, Le; Yang, Lin; Wang, Lin; Li, Liying

    2015-01-01

    It has been reported that bone marrow-derived mesenchymal stem cells (BMSCs) have capacity to migrate to the damaged liver and contribute to fibrogenesis in chronic liver diseases. 15-Deoxy-Δ12,14-prostaglandin J2 (15d-PGJ2), an endogenous ligand for peroxisome proliferator-activated receptor gamma (PPARγ), is considered a new inhibitor of cell migration. However, the actions of 15d-PGJ2 on BMSC migration remain unknown. In this study, we investigated the effects of 15d-PGJ2 on the migration of BMSCs using a mouse model of chronic liver fibrosis and primary mouse BMSCs. Our results demonstrated that in vivo, 15d-PGJ2 administration inhibited the homing of BMSCs to injured liver by flow cytometric analysis and, in vitro, 15d-PGJ2 suppressed primary BMSC migration in a dose-dependent manner determined by Boyden chamber assay. Furthermore, the repressive effect of 15d-PGJ2 was blocked by reactive oxygen species (ROS) inhibitor, but not PPARγ antagonist, and action of 15d-PGJ2 was not reproduced by PPARγ synthetic ligands. In addition, 15d-PGJ2 triggered a significant ROS production and cytoskeletal remodeling in BMSCs. In conclusion, our results suggest that 15d-PGJ2 plays a crucial role in homing of BMSCs to the injured liver dependent on ROS production, independently of PPARγ, which may represent a new strategy in the treatment of liver fibrosis. PMID:26457076

  4. Differentiation of Human Bone Marrow-Derived Mesenchymal Stem Cells into Insulin-Producing Cells: Evidence for Further Maturation In Vivo

    PubMed Central

    Gabr, Mahmoud M.; Zakaria, Mahmoud M.; Refaie, Ayman F.; Khater, Sherry M.; Ashamallah, Sylvia A.; Ismail, Amani M.; El-Halawani, Sawsan M.; Ghoneim, Mohamed A.

    2015-01-01

    The aim of this study was to provide evidence for further in vivo maturation of insulin-producing cells (IPCs) derived from human bone marrow-derived mesenchymal stem cells (HBM-MSCs). HBM-MSCs were obtained from three insulin-dependent type 2 diabetic volunteers. Following expansion, cells were differentiated according to a trichostatin-A/GLP protocol. One million cells were transplanted under the renal capsule of 29 diabetic nude mice. Blood glucose, serum human insulin and c-peptide levels, and glucose tolerance curves were determined. Mice were euthanized 1, 2, 4, or 12 weeks after transplantation. IPC-bearing kidneys were immunolabeled, number of IPCs was counted, and expression of relevant genes was determined. At the end of in vitro differentiation, all pancreatic endocrine genes were expressed, albeit at very low values. The percentage of IPCs among transplanted cells was small (≤3%). Diabetic animals became euglycemic 8 ± 3 days after transplantation. Thereafter, the percentage of IPCs reached a mean of ~18% at 4 weeks. Relative gene expression of insulin, glucagon, and somatostatin showed a parallel increase. The ability of the transplanted cells to induce euglycemia was due to their further maturation in the favorable in vivo microenvironment. Elucidation of the exact mechanism(s) involved requires further investigation. PMID:26064925

  5. Simulated intervertebral disc-like assembly using bone marrow-derived mesenchymal stem cell sheets and silk scaffolds for annulus fibrosus regeneration.

    PubMed

    See, Eugene Yong-Shun; Toh, Siew Lok; Goh, James Cho Hong

    2012-07-01

    Most studies on the intervertebral disc (IVD) focus on the regeneration of the nucleus pulposus (NP). However, without a proper strategy to regenerate the damaged annulus fibrosus (AF), the NP replacements are bound to fail. Therefore the objective of this study was to investigate whether the use of bone marrow-derived mesenchymal stem cells (BMSCs) to form cell sheets, and incorporating them onto silk scaffolds, has the potential to regenerate the annulus fibrosus. The BMSC cell sheets and silk scaffolds were wrapped around a silicone NP substitute to form a simulated IVD-like assembly. The simulated IVD-like assembly was cultured for 4 weeks in static conditions and it was shown that the BMSC cell sheets remained viable, with no significant change in cell numbers. Histological analysis showed that the BMSC cell sheets adhered well onto the silk scaffolds and glycosaminoglycans (GAGs) were detected within the extracellular matrix (ECM). The ratio of collagen type I to collagen type II within the ECM of the BMSC cell sheets also decreased significantly over the period of culture. The results suggested that extensive remodelling of the ECM occurred within the simulated IVD-like assembly, and it is suitable for the regeneration of the inner AF. PMID:21800436

  6. Histomorphometric evaluation of treatment of rat azoosper-mic seminiferous tubules by allotransplantation of bone marrow-derived mesenchymal stem cells

    PubMed Central

    Rahmanifar, Farhad; Tamadon, Amin; Mehrabani, Davood; Zare, Shahrokh; Abasi, Sorush; Keshavarz, Saeideh; Dianatpour, Mehdi; Khodabandeh, Zahra; Jahromi, Iman Raze ghian; Koohi-Hoseinabadi, Omid

    2016-01-01

    Objective(s): Bone marrow-derived mesenchymal stem cells (BM-MSCs) potentials make them appropriate for cell therapy including ability of differentiation and release of anti-inflammatory cytokines and growth factors secreta. For treatment of azoospermia to induce proliferation and differentiation of germ cells, MSCs transplantation has been introduced. The aim of the present experimental case-control study was to histomorphometric evaluation of the germinal cells in seminiferous tubules of azoospermic rats before and after BM-MSCs allotransplantation. Materials and Methods: In the present study, BM-MSCs were isolated from six male rats and confirmed. Their testes also served as intact negative controls. The recipient rats (n=6) were received two doses of 10 mg/kg of busulfan with 21 days interval to induce azoospermia. After cessation of spermatogenesis, the rats were allotransplanted with the BM-MSCs into efferent duct of right testes. Thirty-five days later, the right cell-treated testes were compared to left azoospermic ones. Results: Histomorphometric analyses showed that the seminiferous tubules treated with BM-MSCs had normal morphology in comparison with azoospermic testes, which were without germinal layer. In most BM-MSCs-treated seminiferous tubules, spermatogenesis was observed. Conclusion: The allotransplanted BM-MSCs could induce spermatogenesis in seminiferous tubules of azoospermic rats. PMID:27482347

  7. Acoustic-frequency vibratory stimulation regulates the balance between osteogenesis and adipogenesis of human bone marrow-derived mesenchymal stem cells.

    PubMed

    Chen, Xi; He, Fan; Zhong, Dong-Yan; Luo, Zong-Ping

    2015-01-01

    Osteoporosis can be associated with the disordered balance between osteogenesis and adipogenesis of bone marrow-derived mesenchymal stem cells (BM-MSCs). Although low-frequency mechanical vibration has been demonstrated to promote osteogenesis, little is known about the influence of acoustic-frequency vibratory stimulation (AFVS). BM-MSCs were subjected to AFVS at frequencies of 0, 30, 400, and 800 Hz and induced toward osteogenic or adipogenic-specific lineage. Extracellular matrix mineralization was determined by Alizarin Red S staining and lipid accumulation was assessed by Oil Red O staining. Transcript levels of osteogenic and adipogenic marker genes were evaluated by real-time reverse transcription-polymerase chain reaction. Cell proliferation of BM-MSCs was promoted following exposure to AFVS at 800 Hz. Vibration at 800 Hz induced the highest level of calcium deposition and significantly increased mRNA expression of COL1A1, ALP, RUNX2, and SPP1. The 800 Hz group downregulated lipid accumulation and levels of adipogenic genes, including FABP4, CEBPA, PPARG, and LEP, while vibration at 30 Hz supported adipogenesis. BM-MSCs showed a frequency-dependent response to acoustic vibration. AFVS at 800 Hz was the most favorable for osteogenic differentiation and simultaneously suppressed adipogenesis. Thus, acoustic vibration could potentially become a novel means to prevent and treat osteoporosis. PMID:25738155

  8. A novel strontium(II)-modified calcium phosphate bone cement stimulates human-bone-marrow-derived mesenchymal stem cell proliferation and osteogenic differentiation in vitro.

    PubMed

    Schumacher, M; Lode, A; Helth, A; Gelinsky, M

    2013-12-01

    In the present study, the in vitro effects of novel strontium-modified calcium phosphate bone cements (SrCPCs), prepared using two different approaches on human-bone-marrow-derived mesenchymal stem cells (hMSCs), were evaluated. Strontium ions, known to stimulate bone formation and therefore already used in systemic osteoporosis therapy, were incorporated into a hydroxyapatite-forming calcium phosphate bone cement via two simple approaches: incorporation of strontium carbonate crystals and substitution of Ca(2+) by Sr(2+) ions during cement setting. All modified cements released 0.03-0.07 mM Sr(2+) under in vitro conditions, concentrations that were shown not to impair the proliferation or osteogenic differentiation of hMSCs. Furthermore, strontium modification led to a reduced medium acidification and Ca(2+) depletion in comparison to the standard calcium phosphate cement. In indirect and direct cell culture experiments with the novel SrCPCs significantly enhanced cell proliferation and differentiation were observed. In conclusion, the SrCPCs described here could be beneficial for the local treatment of defects, especially in the osteoporotic bone. PMID:23917042

  9. Cellular behaviour of hepatocyte-like cells from nude mouse bone marrow-derived mesenchymal stem cells on galactosylated poly(D,L-lactic-co-glycolic acid).

    PubMed

    Roh, Hyun; Yang, Dae Hyeok; Chun, Heung Jae; Khang, Gilson

    2015-07-01

    Previously, the galactosylation of poly(d,l-lactic-co-glycolic acid) (PLGA) surface was accomplished by grafting allylamine (AA), using inductively coupled plasma-assisted chemical vapour deposition (ICP-CVD) and conjugating lactobionic acid (LA) with AA via 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide and N-hydroxysuccinimide (EDC/NHS) activation for hepatic tissue-engineering purposes. As a continuation study, the cellular behaviour of hepatocyte-like cells (HLCs) on the surface of the galactosylated PLGA were investigated. Nude mouse bone marrow-derived mesenchymal stem cells (MSCs) were cultured under hepatogenic conditions and the differentiated cells were characterized by reverse-transcription polymerase chain reaction (RT-PCR), immunofluorescence and periodic acid-Schiff (PAS) staining. Galactosylated PLGA enhanced the proliferation rate of HLCs compared to the control; HLCs on the surface of the sample became aggregated and formed spheroids after 3 days of culture. A large number of cells on the surface of the sample exhibited increased liver-specific functional activities, such as albumin and urea secretions. In addition, multicellular spheroids in the sample strongly expressed phospholyated focal adhesion kinase (pFAK) (cell-matrix interactions), E-cadherin (cell-cell interactions) and connexin 32 (Cox32; gap junction). PMID:23784953

  10. Enhancement of tendon-to-bone healing after anterior cruciate ligament reconstruction using bone marrow-derived mesenchymal stem cells genetically modified with bFGF/BMP2.

    PubMed

    Chen, Biao; Li, Bin; Qi, Yong-Jian; Ni, Qu-Bo; Pan, Zheng-Qi; Wang, Hui; Chen, Liao-Bin

    2016-01-01

    Many strategies, including various growth factors and gene transfer, have been used to augment healing after anterior cruciate ligament (ACL) reconstruction. The biological environment regulated by the growth factors during the stage of tendon-bone healing was considered important in controlling the integrating process. The purpose of this study was to evaluate the effects of bone marrow-derived mesenchymal stem cells (BMSCs) genetically modified with bone morphogenetic protein 2 (BMP2) and basic fibroblast growth factor (bFGF) on healing after ACL reconstruction. BMSCs were infected with an adenoviral vector encoding BMP2 (AdBMP2) or bFGF (AdbFGF). Then, the infected BMSCs were surgically implanted into the tendon-bone interface. At 12 weeks postoperatively, the formation of abundant cartilage-like cells, smaller tibial bone tunnel and significantly higher ultimate load and stiffness levels, through histological analysis, micro-computed tomography and biomechanical testing, were observed. In addition, the AdBMP2-plus-AdbFGF group had the smallest bone tunnel and the best mechanical properties among all the groups. The addition of BMP2 or bFGF by gene transfer resulted in better cellularity, new bone formation and higher mechanical property, which contributed to the healing process after ACL reconstruction. Furthermore, the co-application of these two genes was more powerful and efficient than either single gene therapy. PMID:27173013

  11. Bone Marrow-Derived Mesenchymal Stem Cells Improve Diabetic Neuropathy by Direct Modulation of Both Angiogenesis and Myelination in Peripheral Nerves

    PubMed Central

    Han, Ji Woong; Choi, Dabin; Lee, Min Young; Huh, Yang Hoon; Yoon, Young-sup

    2016-01-01

    Recent evidence has suggested that diabetic neuropathy (DN) is pathophysiologically related to both impaired angiogenesis and a deficiency of neurotrophic factors in the nerves. It is widely known that vascular and neural growths are intimately associated. Mesenchymal stem cells (MSCs) promote angiogenesis in ischemic diseases and have neuroprotective effects, particularly on Schwann cells. Accordingly, we investigated whether DN could be improved by local transplantation of MSCs by augmenting angiogenesis and neural regeneration such as remyelination. In sciatic nerves of streptozotocin (STZ)-induced diabetic rats, motor and sensory nerve conduction velocities (NCVs) and capillary density were reduced, and axonal atrophy and demyelination were observed. After injection of bone marrow-derived MSCs (BM-MSCs) into hindlimb muscles, NCVs were restored to near-normal levels. Histological examination demonstrated that injected MSCs were preferentially and durably engrafted in the sciatic nerves, and a portion of the engrafted MSCs were distinctively localized close to vasa nervora of sciatic nerves. Furthermore, vasa nervora increased in density, and the ultrastructure of myelinated fibers in nerves was observed to be restored. Real-time RT-PCR experiments showed that gene expression of multiple factors involved in angiogenesis, neural function, and myelination were increased in the MSC-injected nerves. These findings suggest that MSC transplantation improved DN through direct peripheral nerve angiogenesis, neurotrophic effects, and restoration of myelination. PMID:25975801

  12. Hypoxia pretreatment of bone marrow-derived mesenchymal stem cells seeded in a collagen-chitosan sponge scaffold promotes skin wound healing in diabetic rats with hindlimb ischemia.

    PubMed

    Tong, Chuan; Hao, Haojie; Xia, Lei; Liu, Jiejie; Ti, Dongdong; Dong, Liang; Hou, Qian; Song, Haijing; Liu, Huiling; Zhao, Yali; Fu, Xiaobing; Han, Weidong

    2016-01-01

    Bone marrow-derived mesenchymal stem cells (BM-MSCs) have properties that make them promising for the treatment of chronic nonhealing wounds. The major challenge is ensuring an efficient, safe, and painless delivery of BM-MSCs. Tissue-engineered skin substitutes have considerable benefits in skin damage resulting from chronic nonhealing wounds. Here, we have constructed a three-dimensional biomimetic scaffold known as collagen-chitosan sponge scaffolds (CCSS) using the cross-linking and freeze-drying method. Scanning electron microscopy images showed that CCSS had an interconnected network pore configuration about 100 μm and exhibited a suitable swelling ratio for maintaining morphological stability and appropriate biodegradability to improve biostability using swelling and degradation assays. Furthermore, BM-MSCs were seeded in CCSS using the two-step seeding method to construct tissue-engineered skin substitutes. In addition, in this three-dimensional biomimetic CCSS, BM-MSCs secreted their own collagen and maintain favorable survival ability and viability. Importantly, BM-MSCs exhibited a significant upregulated expression of proangiogenesis factors, including HIF-1α, VEGF, and PDGF following hypoxia pretreatment. In vivo, hypoxia pretreatment of the skin substitute observably accelerated wound closure via the reduction of inflammation and enhanced angiogenesis in diabetic rats with hindlimb ischemia. Thus, hypoxia pretreatment of the skin substitutes can serve as ideal bioengineering skin substitutes to promote optimal diabetic skin wound healing. PMID:26463737

  13. Autophagy regulates the apoptosis of bone marrow-derived mesenchymal stem cells under hypoxic condition via AMP-activated protein kinase/mammalian target of rapamycin pathway.

    PubMed

    Zhang, Zheng; Yang, Ming; Wang, Yabin; Wang, Le; Jin, Zhitao; Ding, Liping; Zhang, Lijuan; Zhang, Lina; Jiang, Wei; Gao, Guojie; Yang, Junke; Lu, Bingwei; Cao, Feng; Hu, Taohong

    2016-06-01

    Bone marrow-derived mesenchymal stem cells (BM-MSCs) have been demonstrated as an ideal autologous stem cells source for cell-based therapy for myocardial infarction (MI). However, poor viability of donor stem cells after transplantation limits their therapeutic efficiency, whereas the underlying mechanism is still poorly understood. Autophagy, a highly conserved process of cellular degradation, is required for maintaining homeostasis and normal function. Here, we investigated the potential role of autophagy on apoptosis in BM-MSCs induced by hypoxic injury. BM-MSCs, isolated from male C57BL/6 mice, were subjected to hypoxia and serum deprivation (H/SD) injury for 6, 12, and 24 h, respectively. The autophagy state was regulated by 3-methyladenine (3MA) and rapamycin administration. Furthermore, compound C was administrated to inhibit AMPK. The apoptosis induced by H/SD was determined by TUNEL assays. Meanwhile, autophagy was measured by GFP-LC3 plasmids transfection and transmission electron microscope. Moreover, protein expressions were evaluated by Western blot assay. In the present study, we found that hypoxic stress increased autophagy and apoptosis in BM-MSCs time dependently. Meanwhile, hypoxia increased the activity of AMPK/mTOR signal pathway. Moreover, increased apoptosis in BM-MSCs under hypoxia was abolished by 3-MA, whereas was aggravated by rapamycin. Furthermore, the increased autophagy and apoptosis in BM-MSCs induced by hypoxia were abolished by AMPK inhibitor compound C. These data provide evidence that hypoxia induced AMPK/mTOR signal pathway activation which regulated the apoptosis and autophagy in BM-MSCs. Furthermore, the apoptosis of BM-MSCs under hypoxic condition was regulated by autophagy via AMPK/mTOR pathway. PMID:27005844

  14. Therapeutic Potential of Bone Marrow-Derived Mesenchymal Stem Cells on Experimental Liver Injury Induced by Schistosoma mansoni: A Histological Study

    PubMed Central

    Fikry, Heba; Gawad, Sara Abdel; Baher, Walaa

    2016-01-01

    Background and Objectives Bone marrow derived mesenchymal stem cells (BM-MSCs) have been proposed as effective treatment of many diseases owing to their unique ability to differentiate into other cell types in vivo. Schistosoma mansoni (S. mansoni) infection is characterized by hepatic granuloma formation around schistosome eggs at acute stage of infection, followed by hepatic fibrosis at chronic and advanced stages. Whether BM-MSCs have an ameliorative effect on hepatic tissue injury caused by S. mansoni infection or not, was inspected in the current study. Materials and Results Female Swiss Albino mice were divided into a control group and an experimental group. Half of control animals served as donors for bone marrow stem cells, and the other half was used to collect liver samples. Experimental group was injected with circariae of S. mansoni, and then subdivided into three subgroups; Subgroup B1, sacrificed after eight weeks of infection without treatment, subgroup B2, received BM-MSCs at the eighth week and sacrificed four weeks later, and subgroup B3, was untreated till the twelfth week of infection. Histological examination of liver samples showed the formation of granulomas and liver fibrosis which were extensive in subgroup B3. However, treated subgroup illustrated improvement of liver histology, signs of hepatocytes regeneration, and possible contribution of oval cell in the process of hepatic and biliary regeneration. Conclusion BM-MSCs decreased liver fibrosis and contributed to an increase in oval cells, generation of new hepatocytes and/or to the improvement of resident hepatocytes in S. mansoni infected mice. PMID:27426091

  15. TNF-α Inhibits FoxO1 by Upregulating miR-705 to Aggravate Oxidative Damage in Bone Marrow-Derived Mesenchymal Stem Cells during Osteoporosis.

    PubMed

    Liao, Li; Su, Xiaoxia; Yang, Xiaohong; Hu, Chenghu; Li, Bei; Lv, Yajie; Shuai, Yi; Jing, Huan; Deng, Zhihong; Jin, Yan

    2016-04-01

    Decline of antioxidant defense after estrogen deficiency leads to oxidative damage in bone marrow-derived mesenchymal stem cells (BMMSCs), resulting a defect of bone formation in osteoporosis. Forkhead box O1 (FoxO1) protein is crucial for defending physiological oxidative damage in bone. But whether FoxO1 is involved in the oxidative damage during osteoporosis is largely unknown. In this study, we found that FoxO1 protein accumulation was decreased in BMMSCs of ovariectomized mice. The decrease of FoxO1 resulted in the suppression of manganese superoxide dismutase (Sod2) and catalase (Cat) expression and accumulation of reactive oxygen species (ROS), inhibiting the osteogenic differentiation of BMMSCs. The decline of FoxO1 protein was caused by tumor necrosis factor-alpha (TNF-α) accumulated after estrogen deficiency. Mechanistically, TNF-α activated NF-κB pathway to promote microRNA-705 expression, which function as a repressor of FoxO1 through post-transcriptional regulation. Inhibition of NF-κB pathway or knockdown of miR-705 largely prevented the decline of FoxO1-mediated antioxidant defense caused by TNF-α and ameliorated the oxidative damage in osteoporotic BMMSCs. Moreover, the accumulated ROS further activated NF-κB pathway with TNF-α, which formed a feed-forward loop to persistently inhibiting FoxO1 protein accumulation in BMMSCs. In conclusion, our study revealed that the decline of FoxO1 is an important etiology factor of osteoporosis and unclosed a novel mechanism of FoxO1 regulation by TNF-α. These findings suggested a close correlation between inflammation and oxidative stress in stem cell dysfunction during degenerative bone diseases. PMID:26700816

  16. Effects of TGF-β1 and alginate on the differentiation of rabbit bone marrow-derived mesenchymal stem cells into a chondrocyte cell lineage

    PubMed Central

    WANG, WAN-ZONG; YAO, XIAO-DONG; HUANG, XIAO-JIN; LI, JIN-QUAN; XU, HAO

    2015-01-01

    The aim of the present study was to investigate the effect of a three-dimensional (3D) culture system of sodium alginate gel on the directional differentiation induction of bone marrow-derived mesenchymal stem cells (BMSCs) into chondrocytes, as well as the in vitro gene transfection technique. The biological characteristics of the passage and proliferation of rabbit BMSCs were investigated under conditions of in vitro monolayer and 3D culture of sodium alginate gel. Transforming growth factor (TGF)-β1 gene recombinant adenoviral cosmid vectors and the recombinant adenoviral vector Ad.TGF-β1 were constructed, and the effect of Ad.TGF-β1 transfection on the differentiation of BMSCs into chondrocytes was investigated. The whole bone marrow rinsing method was used to obtain, separate and purify the rabbit BMSCs, and the in vitro monolayer and 3D culture of sodium alginate gel were thus successfully and stably established. A safe, stable and efficient method of constructing Ad.TGF-β1 TGF-β1 gene recombinant adenoviral vectors was established. Following TGF-β1 transfection, BMSCs were able to continuously secrete significantly increased amounts of specific extracellular matrix components of chondrocytes, such as collagen II and proteoglycans. Furthermore, the effects in the post-gene transfection 3D culture group were found to be enhanced compared with those in the monolayer culture group. In conclusion, the 3D culture system of sodium alginate gel and in vitro gene transfection exhibited significant inductive effects on differentiation, which could be used to promote BMSCs to differentiate into chondrocytes. PMID:26622428

  17. Periostin: A Downstream Mediator of EphB4-Induced Osteogenic Differentiation of Human Bone Marrow-Derived Mesenchymal Stem Cells

    PubMed Central

    Zhang, Fei; Zhang, Zehua; Sun, Dong; Dong, Shiwu; Xu, Jianzhong; Dai, Fei

    2016-01-01

    Erythropoietin-producing hepatocyte B4 (EphB4) has been reported to be a key molecular switch in the regulation of bone homeostasis, but the underlying mechanism remains poorly understood. In this study, we investigated the role of EphB4 in regulating the expression of periostin (POSTN) within bone marrow-derived mesenchymal stem cells (MSCs) and assessed its effect and molecular mechanism of osteogenic induction in vitro. Treatment with ephrinB2-FC significantly increased the expression of POSTN in MSCs, and the inhibition of EphB4 could abrogate this effect. In addition, osteogenic markers were upregulated especially in MSCs overexpressing EphB4. To elucidate the underlying mechanism of cross talk between EphB4 and the Wnt pathway, we detected the change in protein expression of phosphorylated-glycogen synthase kinase 3β-serine 9 (p-GSK-3β-Ser9) and β-catenin, as well as the osteogenic markers Runx2 and COL1. The results showed that GSK-3β activation and osteogenic marker expression levels were downregulated by ephrinB2-FC treatment, but these effects were inhibited by blocking integrin αvβ3 in MSCs. Our findings demonstrate that EphB4 can promote osteogenic differentiation of MSCs via upregulation of POSTN expression. It not only helps to reveal the interaction mechanism between EphB4 and Wnt pathway but also brings a better understanding of EphB4/ephrinB2 signaling in bone homeostasis. PMID:26788070

  18. Effect of Chromatin-Remodeling Agents in Hepatic Differentiation of Rat Bone Marrow-Derived Mesenchymal Stem Cells In Vitro and In Vivo

    PubMed Central

    Ye, Danna; Li, Tong; Heraud, Philip; Parnpai, Rangsun

    2016-01-01

    Epigenetic events, including covalent histone modifications and DNA methylation, play fundamental roles in the determination of lineage-specific gene expression and cell fates. The aim of this study was to determine whether the DNA methyltransferase inhibitor (DNMTi) 5-aza-2′-deoxycytidine (5-aza-dC) and the histone deacetylase inhibitor (HDACi) trichostatin A (TSA) promote the hepatic differentiation of rat bone marrow-derived mesenchymal stem cells (rBM-MSCs) and their therapeutic effect on liver damage. 1 μM TSA and 20 μM 5-aza-dC were added to standard hepatogenic medium especially at differentiation and maturation steps and their potential function on hepatic differentiation in vitro and in vivo was determined. Exposure of rBM-MSCs to 1 μM TSA at both the differentiation and maturation steps considerably improved hepatic differentiation. TSA enhanced the development of the hepatocyte shape, promoted the chronological expression of hepatocyte-specific markers, and improved hepatic functions. In contrast, treatment of rBM-MSCs with 20 μM 5-aza-dC alone or in combination with TSA was ineffective in improving hepatic differentiation in vitro. TSA and/or 5-aza-dC derived hepatocytes-like cells failed to improve the therapeutic potential in liver damage. We conclude that HDACis enhance hepatic differentiation in a time-dependent manner, while DNMTis do not induce the hepatic differentiation of rBM-MSCs in vitro. Their in vivo function needs further investigation. PMID:27242905

  19. Autophagy Plays a Protective Role in Tumor Necrosis Factor-α-Induced Apoptosis of Bone Marrow-Derived Mesenchymal Stem Cells.

    PubMed

    Yang, Rui; Ouyang, Yi; Li, Weiping; Wang, Peng; Deng, Haiquan; Song, Bin; Hou, Jingyi; Chen, Zhong; Xie, Zhongyu; Liu, Zhenhua; Li, Jinteng; Cen, Shuizhong; Wu, Yanfeng; Shen, Huiyong

    2016-05-15

    Bone marrow-derived mesenchymal stem cells (BMSCs) are being broadly investigated for treating numerous inflammatory diseases. However, the low survival rate of BMSCs during the transplantation process has limited their application. Autophagy can maintain cellular homeostasis and protect cells against environmental stresses. Tumor necrosis factor-α (TNF-α) is an important inflammatory cytokine that can induce both autophagy and apoptosis of BMSCs. However, the actual role of autophagy in TNF-α-induced apoptosis of BMSCs remains poorly understood. In the current study, BMSCs were treated with TNF-α/cycloheximide (CHX), and cell death was examined by the Cell Counting Kit-8, Hoechst 33342 staining, and flow cytometric analysis as well as by the level of caspase-3 and caspase-8. Meanwhile, autophagic flux was examined by analyzing the level of microtubule-associated protein light chain 3 B (LC3B)-II and SQSTEM1/p62 and by examining the amount of green fluorescent protein-LC3B by fluorescence microscopy. Then, the cell death and autophagic flux of BMSCs were examined after pretreatment and cotreatment with 3-methyladenine (3-MA, autophagy inhibitor) or rapamycin (Rap, autophagy activator) together with TNF-α/CHX. Moreover, BMSCs pretreated with lentiviruses encoding short hairpin RNA of beclin-1 (BECN1) were treated with TNF-α/CHX, and then cell death and autophagic flux were detected. We showed that BMSCs treated with TNF-α/CHX presented dramatically elevated autophagic flux and cell death. Furthermore, we showed that 3-MA and shBECN1 treatment accelerated TNF-α/CHX-induced apoptosis, but that Rap treatment ameliorated cell death. Our results demonstrate that autophagy protects BMSCs against TNF-α-induced apoptosis. Enhancing the autophagy of BMSCs may elevate cellular survival in an inflammatory microenvironment. PMID:26985709

  20. Probable impact of age and hypoxia on proliferation and microRNA expression profile of bone marrow-derived human mesenchymal stem cells

    PubMed Central

    Mohd Ali, Norlaily; Boo, Lily; Yeap, Swee Keong; Ky, Huynh; Satharasinghe, Dilan A.; Liew, Woan Charn; Cheong, Soon Keng; Kamarul, Tunku

    2016-01-01

    Decline in the therapeutic potential of bone marrow-derived mesenchymal stem cells (MSC) is often seen with older donors as compared to young. Although hypoxia is known as an approach to improve the therapeutic potential of MSC in term of cell proliferation and differentiation capacity, its effects on MSC from aged donors have not been well studied. To evaluate the influence of hypoxia on different age groups, MSC from young (<30 years) and aged (>60 years) donors were expanded under hypoxic (5% O2) and normal (20% O2) culture conditions. MSC from old donors exhibited a reduction in proliferation rate and differentiation potential together with the accumulation of senescence features compared to that of young donors. However, MSC cultured under hypoxic condition showed enhanced self-renewing and proliferation capacity in both age groups as compared to normal condition. Bioinformatic analysis of the gene ontology (GO) and KEGG pathway under hypoxic culture condition identified hypoxia-inducible miRNAs that were found to target transcriptional activity leading to enhanced cell proliferation, migration as well as decrease in growth arrest and apoptosis through the activation of multiple signaling pathways. Overall, differentially expressed miRNA provided additional information to describe the biological changes of young and aged MSCs expansion under hypoxic culture condition at the molecular level. Based on our findings, the therapeutic potential hierarchy of MSC according to donor’s age group and culture conditions can be categorized in the following order: young (hypoxia) > young (normoxia) > old aged (hypoxia) > old aged (normoxia). PMID:26788424

  1. Determining the minimum number of detectable cardiac-transplanted 111In-tropolone-labelled bone-marrow-derived mesenchymal stem cells by SPECT

    NASA Astrophysics Data System (ADS)

    Jin, Yuan; Kong, Huafu; Stodilka, Rob Z.; Wells, R. Glenn; Zabel, Pamela; Merrifield, Peter A.; Sykes, Jane; Prato, Frank S.

    2005-10-01

    In this work, we determined the minimum number of detectable 111In-tropolone-labelled bone-marrow-derived stem cells from the maximum activity per cell which did not affect viability, proliferation and differentiation, and the minimum detectable activity (MDA) of 111In by SPECT. Canine bone marrow mesenchymal cells were isolated, cultured and expanded. A number of samples, each containing 5 × 106 cells, were labelled with 111In-tropolone from 0.1 to 18 MBq, and cell viability was measured afterwards for each sample for 2 weeks. To determine the MDA, the anthropomorphic torso phantom (DataSpectrum Corporation, Hillsborough, NC) was used. A point source of 202 kBq 111In was placed on the surface of the heart compartment, and the phantom and all compartments were then filled with water. Three 111In SPECT scans (duration: 16, 32 and 64 min; parameters: 128 × 128 matrix with 128 projections over 360°) were acquired every three days until the 111In radioactivity decayed to undetectable quantities. 111In SPECT images were reconstructed using OSEM with and without background, scatter or attenuation corrections. Contrast-to-noise ratio (CNR) in the reconstructed image was calculated, and MDA was set equal to the 111In activity corresponding to a CNR of 4. The cells had 100% viability when incubated with no more than 0.9 MBq of 111In (80% labelling efficiency), which corresponded to 0.14 Bq per cell. Background correction improved the detection limits for 111In-tropolone-labelled cells. The MDAs for 16, 32 and 64 min scans with background correction were observed to be 1.4 kBq, 700 Bq and 400 Bq, which implies that, in the case where the location of the transplantation is known and fixed, as few as 10 000, 5000 and 2900 cells respectively can be detected.

  2. Compensatory cellular reactions to nonsteroidal anti-inflammatory drugs on osteogenic differentiation in canine bone marrow-derived mesenchymal stem cells.

    PubMed

    Oh, Namgil; Kim, Sangho; Hosoya, Kenji; Okumura, Masahiro

    2014-05-01

    The suppressive effects of nonsteroidal anti-inflammatory drugs (NSAIDs) on the bone healing process have remained controversial, since no clinical data have clearly shown the relationship between NSAIDs and bone healing. The aim of this study was to assess the compensatory response of canine bone marrow-derived mesenchymal stem cells (BMSCs) to several classes of NSAIDs, including carprofen, meloxicam, indomethacin and robenacoxib, on osteogenic differentiation. Each of the NSAIDs (10 µM) was administered during 20 days of the osteogenic process with human recombinant IL-1β (1 ng/ml) as an inflammatory stimulator. Gene expression of osteoblast differentiation markers (alkaline phosphatase and osteocalcin), receptors of PGE2 (EP2 and EP4) and enzymes for prostaglandin (PG) E2 synthesis (COX-1, COX-2, cPGES and mPGES-1) was measured by using quantitative reverse transcription-polymerase chain reaction. Protein production levels of alkaline phosphatase, osteocalcin and PGE2 were quantified using an alkaline phosphatase activity assay, osteocalcin immunoassay and PGE2 immunoassay, respectively. Histologic analysis was performed using alkaline phosphatase staining, von Kossa staining and alizarin red staining. Alkaline phosphatase and calcium deposition were suppressed by all NSAIDs. However, osteocalcin production showed no significant suppression by NSAIDs. Gene expression levels of PGE2-related receptors and enzymes were upregulated during continuous treatment with NSAIDs, while certain channels for PGE2 synthesis were utilized differently depending on the kind of NSAIDs. These data suggest that canine BMSCs have a compensatory mechanism to restore PGE2 synthesis, which would be an intrinsic regulator to maintain differentiation of osteoblasts under NSAID treatment. PMID:24419976

  3. Lithium stimulates human bone marrow derived mesenchymal stem cell proliferation through GSK-3β-dependent β-catenin/Wnt pathway activation.

    PubMed

    Zhu, Zhenzhong; Yin, Junhui; Guan, Junjie; Hu, Bin; Niu, Xin; Jin, Dongxu; Wang, Yang; Zhang, Changqing

    2014-12-01

    Mesenchymal stem cells (MSCs) are multipotent cells that have been widely used in cell based transplantation therapy. The use of MSCs requires in vitro expansion in order to fulfill their regenerative capacity. Therefore the proliferative ability of MSCs is one of the key factors which determine MSC therapeutic efficacy. In the present study, we showed for the first time that lithium, a well-known antidepressant, reversibly promoted the proliferation of human bone marrow derived MSCs in vitro. MSCs treated with 5 mm lithium proliferated more rapidly than untreated cells without undergoing apoptosis. Lithium increased the proportion of cells in S phase as well as cyclin D1 expression. Mechanistic studies revealed that these effects were dependent upon the activation of the glycogen synthase kinase 3β (GSK-3β) mediated canonical Wnt pathway. Lithium induced Ser9 phosphorylation, which results in the inhibition of GSK-3β activity, β-catenin accumulation and Wnt pathway activation. Utilizing a specific GSK-3β inhibitor SB216763 or siRNA-mediated inhibition of GSK-3β produced effects similar to those induced by lithium. In contrast, either quercetin, an inhibitor of the β-catenin/TCF pathway, or siRNA-mediated knockdown of β-catenin abolished the proliferative effect of lithium, suggesting that lithium stimulates MSC proliferation via the GSK-3β-dependent β-catenin/Wnt pathway. Collectively, these studies elucidate a novel role of lithium, which may not only provide a simple and effective way to strengthen MSC transplantation therapy efficacy but also shed light on lithium's clinical application for the treatment of certain disorders resulting from β-catenin/Wnt pathway suppression. PMID:25265417

  4. A CXCL5- and bFGF-Dependent Effect of PDGF-B-Activated Fibroblasts in Promoting Trafficking and Differentiation of Bone Marrow-Derived Mesenchymal Stem Cells

    PubMed Central

    Nedeau, April E.; Bauer, Richard J.; Gallagher, Katherine; Chen, Haiying; Liu, Zhao-Jun

    2008-01-01

    Adult bone marrow-derived mesenchymal stem cells (MSCs) are able to differentiate into myofibroblasts and be recruited into wound lesions and contribute to wound healing. The cellular and molecular mechanism responsible for MSC trafficking and differentiation, however, are poorly understood. Local resting resident fibroblasts are activated after injury and play a critical role in recruiting MSCs. We investigated the role of platelet derived growth factor-B-activated fibroblasts (PDGF-B-aFBs) in regulating recruitment, migration and differentiation of MSCs from GFP transgenic mice in an in vitro wound healing assay and a novel three-dimensional (3D) model. PDGF-B-aFBs caused significant increases in MSCs migration velocity compared to control as demonstrated by time-lapse photography in an in vitro wound healing assay. Consistently, invasion/migration of MSCs into 3D collagen gels was enhanced in the presence of PDGF-B-aFbs. In addition, PDGF-B-aFBs induced differentiation of MSCs into myofibroblast. The regulatory effects of PDGF-B-aFBs are likely to be mediated by basic fibroblast growth factor (bFGF) and epithelial neutrophil activating peptide-78 (ENA-78 or CXCL5) as protein array analysis indicated an elevated levels of these two soluble factors in culture supernatant of PDGF-B-aFBs. Blocking antibodies against bFGF and CXCL5 were able to inhibit both trafficking and differentiation of MSCs into 3D collagen gels while supplement of exogenous bFGF and/or CXCL5 promoted invasion/migration of MSCs into 3D collagen gels. Our results reveal that PDGF-B-aFBs play a key role in recruitment/migration and differentiation of MSCs and implicate a bFGF- and CXCL5-dependent mechanism in mediating these effects. PMID:18570917

  5. Bone marrow-derived mesenchymal stem cells enhance autophagy via PI3K/AKT signalling to reduce the severity of ischaemia/reperfusion-induced lung injury

    PubMed Central

    Li, Jing; Zhou, Jian; Zhang, Dan; Song, Yuanlin; She, Jun; Bai, Chunxue

    2015-01-01

    Autophagy, a type II programmed cell death, is essential for cell survival under stress, e.g. lung injury, and bone marrow-derived mesenchymal stem cells (BM-MSCs) have great potential for cell therapy. However, the mechanisms underlying the BM-MSC activation of autophagy to provide a therapeutic effect in ischaemia/reperfusion-induced lung injury (IRI) remain unclear. Thus, we investigate the activation of autophagy in IRI following transplantation with BM-MSCs. Seventy mice were pre-treated with BM-MSCs before they underwent lung IRI surgery in vivo. Human pulmonary micro-vascular endothelial cells (HPMVECs) were pre-conditioned with BM-MSCs by oxygen-glucose deprivation/reoxygenation (OGD) in vitro. Expression markers for autophagy and the phosphoinositide 3-kinase/protein kinase B (PI3K/Akt) signalling pathway were analysed. In IRI-treated mice, administration of BM-MSCs significantly attenuated lung injury and inflammation, and increased the level of autophagy. In OGD-treated HPMVECs, co-culture with BM-MSCs attenuated endothelial permeability by decreasing the level of cell death and enhanced autophagic activation. Moreover, administration of BM-MSCs decreased the level of PI3K class I and p-Akt while the expression of PI3K class III was increased. Finally, BM-MSCs-induced autophagic activity was prevented using the inhibitor LY294002. Administration of BM-MSCs attenuated lung injury by improving the autophagy level via the PI3K/Akt signalling pathway. These findings provide further understanding of the mechanisms related to BM-MSCs and will help to develop new cell-based therapeutic strategies in lung injury. PMID:26177266

  6. Evaluation of autologous bone marrow-derived mesenchymal stem cells on renal regeneration after experimentally induced acute kidney injury in dogs.

    PubMed

    Lim, Chae-Young; Han, Jae-Ik; Kim, Seung-Gon; Lee, Chang-Min; Park, Hee-Myung

    2016-02-01

    OBJECTIVE To evaluate the usefulness of autologous bone marrow-derived mesenchymal stem cell (BM-MSC) therapy for the treatment of dogs with experimentally induced acute kidney injury. ANIMALS 6 healthy dogs. PROCEDURES After induction of kidney injury (day 0) with cisplatin (5 mg/kg, IV), dogs immediately received saline (0.9% NaCl) solution (10 mL; n = 3) or BM-MSCs (1 × 10(6) cells/kg in 10 mL of saline solution; 3) IV. A CBC, serum biochemical analysis, and urinalysis were performed for each dog before administration of cisplatin and on days 1 through 4. Glomerular filtration rate was determined for all dogs on days -7 and 2; BM-MSC tracking by MRI was performed on BM-MSC-treated dogs on days -14 and 4. After sample collection and BM-MSC tracking on day 4, all dogs were euthanized; kidney tissue samples underwent histologic evaluation, immunohistochemical analysis, and cytokine profiling via reverse transcriptase PCR assays. RESULTS Kidney tissue from both groups had mononuclear inflammatory cell infiltration, tubular necrosis, dilated tubules, and glomerular damage. However, there was less fibrotic change and increased proliferation of renal tubular epithelial cells in the BM-MSC-treated dogs, compared with findings for the control dogs. Expressions of tumor necrosis factor-α and transforming growth factor-β were lower in the BM-MSC-treated group, compared with findings for the control group. Laboratory data revealed no improvement in the renal function in BM-MSC-treated dogs. CONCLUSIONS AND CLINICAL RELEVANCE Results of this study suggested that autologous BM-MSCs may accelerate renal regeneration after experimentally induced acute kidney injury in dogs. (Am J Vet Res 2016;77:208-217). PMID:27027716

  7. Targeted delivery of CYP2E1 recombinant adenovirus to malignant melanoma by bone marrow-derived mesenchymal stem cells as vehicles.

    PubMed

    Wang, Jishi; Ma, Dan; Li, Yan; Yang, Yuan; Hu, Xiaoyan; Zhang, Wei; Fang, Qin

    2014-03-01

    The aim of this study was to explore the effects of bone marrow-derived mesenchymal stem cells (BMSCs) as intermediate carriers on targeting of P450 gene recombinant adenovirus to malignant melanoma in vitro and in vivo. BMSCs were transduced with pAd5-CMV-CYP2E1 recombinant adenovirus. BMSC migration was detected by Transwell plates in vitro and by superparamagnetic iron oxide particles in vivo. Growth-inhibitory effect and apoptosis were determined by MTT and immunity fluorescence staining. Anticancer effects were examined by a human melanoma nude mouse model in vivo. BMSCs moved toward A375 cells in Transwell plates. Numerous superparamagnetic MSCs labeled with iron oxide were identified in the peripheral areas of the tumor, but were detected in primary organs by Prussian blue staining. BMSC-CYP2E1 cells mediated a bystander killing effect on CYP2E1-negative A375 cells during coculture (IC50 values for A375 cells cocultured with BMSC-EGFP and BMSC-CYP2E1 were 4.08 and 2.68 mmol/l, respectively). Intravenously injecting CYP2E1 recombinant adenovirus-loaded BMSCs in mice with established human melanoma managed to target the tumor site, and BMSCs with forced expression of CYP2E1 inhibited the growth of malignant cells in vivo by activating 5-(3,3-dimethyl-1-triazeno)imidazole-4-carboxamide. BMSCs may serve as a platform of P450 gene-directed enzyme prodrug therapy for the delivery of chemotherapeutic prodrugs to tumors. PMID:24413391

  8. A double blind randomized placebo controlled phase I/II study assessing the safety and efficacy of allogeneic bone marrow derived mesenchymal stem cell in critical limb ischemia

    PubMed Central

    2013-01-01

    Background Peripheral vascular disease of the lower extremities comprises a clinical spectrum that extends from no symptoms to presentation with critical limb ischemia (CLI). Bone marrow derived Mesenchymal Stem Cells (BM- MSCs) may ameliorate the consequences of CLI due to their combinatorial potential for inducing angiogenesis and immunomodulatory environment in situ. The primary objective was to determine the safety of BM- MSCs in patients with CLI. Methods Prospective, double blind randomized placebo controlled multi-center study was conducted in patients with established CLI as per Rutherford classification in category II-4, III-5, or III-6 with infra-inguinal arterial occlusive disease and were not suitable for or had failed revascularization treatment. The primary end point was incidence of treatment – related adverse events (AE). Exploratory efficacy end points were improvement in rest pain, increase in Ankle Brachial Pressure Index (ABPI), ankle pressure, healing of ulcers, and amputation rates. Twenty patients (BM-MSC: Placebo = 1:1) were administered with allogeneic BM-MSCs at a dose of 2 million cells/kg or placebo (PlasmaLyte A) at the gastrocnemius muscle of the ischemic limb. Results Improvement was observed in the rest pain scores in both the arms. Significant increase in ABPI and ankle pressure was seen in BM-MSC arm compared to the placebo group. Incidence of AEs in the BM-MSC arm was 13 vs. 45 in the placebo arm where as serious adverse events (SAE) were similar in both the arms (5 in BM-MSC and 4 in the placebo group). SAEs resulted in death, infected gangrene, amputations in these patients. It was observed that the SAEs were related to disease progression and not related to stem cells. Conclusion BM-MSCs are safe when injected IM at a dose of 2 million cells/kg body weight. Few efficacy parameters such as ABPI and ankle pressure showed positive trend warranting further studies. Trial registration NIH website (http

  9. Controlled Dual Growth Factor Delivery From Microparticles Incorporated Within Human Bone Marrow-Derived Mesenchymal Stem Cell Aggregates for Enhanced Bone Tissue Engineering via Endochondral Ossification

    PubMed Central

    Dang, Phuong N.; Dwivedi, Neha; Phillips, Lauren M.; Yu, Xiaohua; Herberg, Samuel; Bowerman, Caitlin; Solorio, Loran D.; Murphy, William L.

    2016-01-01

    Bone tissue engineering via endochondral ossification has been explored by chondrogenically priming cells using soluble mediators for at least 3 weeks to produce a hypertrophic cartilage template. Although recapitulation of endochondral ossification has been achieved, long-term in vitro culture is required for priming cells through repeated supplementation of inductive factors in the media. To address this challenge, a microparticle-based growth factor delivery system was engineered to drive endochondral ossification within human bone marrow-derived mesenchymal stem cell (hMSC) aggregates. Sequential exogenous presentation of soluble transforming growth factor-β1 (TGF-β1) and bone morphogenetic protein-2 (BMP-2) at various defined time courses resulted in varying degrees of chondrogenesis and osteogenesis as demonstrated by glycosaminoglycan and calcium content. The time course that best induced endochondral ossification was used to guide the development of the microparticle-based controlled delivery system for TGF-β1 and BMP-2. Gelatin microparticles capable of relatively rapid release of TGF-β1 and mineral-coated hydroxyapatite microparticles permitting more sustained release of BMP-2 were then incorporated within hMSC aggregates and cultured for 5 weeks following the predetermined time course for sequential presentation of bioactive signals. Compared with cell-only aggregates treated with exogenous growth factors, aggregates with incorporated TGF-β1- and BMP-2-loaded microparticles exhibited enhanced chondrogenesis and alkaline phosphatase activity at week 2 and a greater degree of mineralization by week 5. Staining for types I and II collagen, osteopontin, and osteocalcin revealed the presence of cartilage and bone. This microparticle-incorporated system has potential as a readily implantable therapy for healing bone defects without the need for long-term in vitro chondrogenic priming. Significance This study demonstrates the regulation of chondrogenesis

  10. hTERT- and hCTLA4Ig-expressing human bone marrow-derived mesenchymal stem cells: in vitro and in vivo characterization and osteogenic differentiation.

    PubMed

    Dai, Fei; Yang, Sisi; Zhang, Fei; Shi, Dongwen; Zhang, Zehua; Wu, Jun; Xu, Jianzhong

    2014-07-22

    Multipotent mesenchymal stem cells (MSCs) are commonly used as seed cells in studies of tissue engineering and regenerative medicine but their clinical application is limited, due to insufficient numbers of autogeneic MSCs, immune rejection of allogeneic MSCs and replicative senescence. We constructed two gene expression vectors for transfection of the human telomerase reverse transcriptase (hTERT) and cytotoxic T lymphocyte-associated antigen 4-Ig (CTLA4Ig) genes into human bone marrow-derived stem cells (hBMSCs). Successful transfection of both genes generated hTERT-CTLA4Ig hBMSCs that expressed both telomerase (shown by immunohistochemistry and a TRAPeze assay) and CTLA4Ig (demonstrated by immunocytochemistry and western blotting) without apparent mutual interference. Both hTERT BMSCs (92 population doublings) and hTERT-CTLA4Ig hBMSCs (60 population doublings) had an extended lifespan compared with hBMSCs (18 population doublings). Cell cycle analysis revealed that, compared with hBMSCs, a lower proportion of hTERT hBMSCs were in G0 /G1 phase but a higher proportion were in S phase; compared with hTERT hBMSCs, a higher proportion of hTERT-CTLA4Ig hBMSCs were in G0 /G1 phase, while a lower proportion were in S and G2 /M phases. hTERT-CTLA4Ig hBMSCs retained their capacity for osteogenic differentiation in vitro, shown by the detection of hydroxyapatite mineral deposition (labelled tetracycline fluorescence staining), calcareous nodules (alizarin red S staining), alkaline phosphatase (calcium-cobalt method) and osteocalcin (immunocytochemistry). Furthermore, subcutaneous transplantation of hTERT-CTLA4Ig hBMSCs in a rat xenotransplantation model resulted in the successful generation of bone-like tissue, confirmed using radiography and histological assessment. We propose that allogeneic hTERT-CTLA4Ig hBMSCs may be ideal seed cells for bone tissue engineering. Copyright © 2014 John Wiley & Sons, Ltd. PMID:25047146

  11. Lentiviral-mediated multiple gene transfer to chondrocytes promotes chondrocyte differentiation and bone formation in rabbit bone marrow-derived mesenchymal stem cells.

    PubMed

    Liu, Ping; Sun, Liang; Chen, Hui; Sun, Shui; Zhou, Dongsheng; Pang, Bo; Wang, Jian

    2015-11-01

    The aim of the present study was to provide a theoretical and experimental foundation on the differentiation of stem cells through the induction of multiple genes. The lentiviral vector carrying TGF-β1 and IL-10 genes was transfected to bone marrow-derived mesenchymal stem cells (BMSCs) which differentiated into chondrogenesis. Healthy New Zealand white rabbits, 2-3 months of age were used in the present study. A 6-8 ml of bone marrow was isolated from the iliac and tibial shaft of each rabbit. The BMSCs suspension was aspired following centrifugation of the bone marrow by percoll separating medium. The BMSCs were primarily cultured and subcultured in vitro, then divided into four groups according to the difference of lentivirus vectors: group A, receiving transforming growth factor β1 (TGF‑β1); group B, receiving TGF-β1 and Interleukin-10 (IL-10); group C, empty vector transfection; and group D, receiving no cell growth factor. Fluorescence expression was detected 12 h after transfecting the lentiviral vector carrying the TGF-β1 and IL-10 gene to BMSCs. The transfection efficiency was approximately 70% with a MOI=100 after 96 h. Expression of SOX-9 aggrecan and Type Ⅱ collagen in groups A-E on day 7 and 14 was detected by RT-PCR and western blot analysis. The expression level of three genes expressed in groups A and C were higher compared to the expression in groups B, D and E. The expression level of the three genes expressed in group B was higher compared to the expression in group D. The expression level of three genes expressed in group A and C showed no statistical difference. Cytokines therefore play an important role in cell proliferation and chondrogenic differentiation. TGF-β1 has a synergistic effect in the differentiation. In addition, IL-10 may have a protective role in the restoration of cartilaginous tissue. PMID:26328747

  12. Controlled Dual Growth Factor Delivery From Microparticles Incorporated Within Human Bone Marrow-Derived Mesenchymal Stem Cell Aggregates for Enhanced Bone Tissue Engineering via Endochondral Ossification.

    PubMed

    Dang, Phuong N; Dwivedi, Neha; Phillips, Lauren M; Yu, Xiaohua; Herberg, Samuel; Bowerman, Caitlin; Solorio, Loran D; Murphy, William L; Alsberg, Eben

    2016-02-01

    Bone tissue engineering via endochondral ossification has been explored by chondrogenically priming cells using soluble mediators for at least 3 weeks to produce a hypertrophic cartilage template. Although recapitulation of endochondral ossification has been achieved, long-term in vitro culture is required for priming cells through repeated supplementation of inductive factors in the media. To address this challenge, a microparticle-based growth factor delivery system was engineered to drive endochondral ossification within human bone marrow-derived mesenchymal stem cell (hMSC) aggregates. Sequential exogenous presentation of soluble transforming growth factor-β1 (TGF-β1) and bone morphogenetic protein-2 (BMP-2) at various defined time courses resulted in varying degrees of chondrogenesis and osteogenesis as demonstrated by glycosaminoglycan and calcium content. The time course that best induced endochondral ossification was used to guide the development of the microparticle-based controlled delivery system for TGF-β1 and BMP-2. Gelatin microparticles capable of relatively rapid release of TGF-β1 and mineral-coated hydroxyapatite microparticles permitting more sustained release of BMP-2 were then incorporated within hMSC aggregates and cultured for 5 weeks following the predetermined time course for sequential presentation of bioactive signals. Compared with cell-only aggregates treated with exogenous growth factors, aggregates with incorporated TGF-β1- and BMP-2-loaded microparticles exhibited enhanced chondrogenesis and alkaline phosphatase activity at week 2 and a greater degree of mineralization by week 5. Staining for types I and II collagen, osteopontin, and osteocalcin revealed the presence of cartilage and bone. This microparticle-incorporated system has potential as a readily implantable therapy for healing bone defects without the need for long-term in vitro chondrogenic priming. Significance: This study demonstrates the regulation of chondrogenesis

  13. Comparison of human adipose-derived stem cells and bone marrow-derived stem cells in a myocardial infarction model.

    PubMed

    Rasmussen, Jeppe Grøndahl; Frøbert, Ole; Holst-Hansen, Claus; Kastrup, Jens; Baandrup, Ulrik; Zachar, Vladimir; Fink, Trine; Simonsen, Ulf

    2014-02-01

    Treatment of myocardial infarction (MI) with bone marrow-derived mesenchymal stem cells and recently also adipose-derived stem cells has shown promising results. In contrast to clinical trials and their use of autologous bone marrow-derived cells from the ischemic patient, the animal MI models are often using young donors and young, often immune-compromised, recipient animals. Our objective was to compare bone marrow-derived mesenchymal stem cells with adipose-derived stem cells from an elderly ischemic patient in the treatment of MI using a fully grown non-immune-compromised rat model. Mesenchymal stem cells were isolated from adipose tissue and bone marrow and compared with respect to surface markers and proliferative capability. To compare the regenerative potential of the two stem cell populations, male Sprague-Dawley rats were randomized to receive intramyocardial injections of adipose-derived stem cells, bone marrow-derived mesenchymal stem cells, or phosphate-buffered saline 1 week following induction of MI. After 4 weeks, left ventricular ejection fraction (LVEF) was improved in the adipose-derived stem cell group, and scar wall thickness was greater compared with the saline group. Adipose-derived as well as bone marrow-derived mesenchymal stem cells prevented left ventricular end diastolic dilation. Neither of the cell groups displayed increased angiogenesis in the myocardium compared with the saline group. Adipose-derived stem cells from a human ischemic patient preserved cardiac function following MI, whereas this could not be demonstrated for bone marrow-derived mesenchymal stem cells, with only adipose-derived stem cells leading to an improvement in LVEF. Neither of the stem cell types induced myocardial angiogenesis, raising the question whether donor age and health have an effect on the efficacy of stem cells used in the treatment of MI. PMID:23211469

  14. Reinforced chitosan-based heart valve scaffold and utility of bone marrow-derived mesenchymal stem cells for cardiovascular tissue engineering

    NASA Astrophysics Data System (ADS)

    Albanna, Mohammad Zaki

    Recent research has demonstrated a strong correlation between the differentiation profile of mesenchymal stem cells (MSCs) and scaffold stiffness. Chitosan is being widely studied for tissue engineering applications due to its biocompatibility and biodegradability. However, its use in load-bearing applications is limited due to moderate to low mechanical properties. In this study, we investigated the effectiveness of a fiber reinforcement method for enhancing the mechanical properties of chitosan scaffolds. Chitosan fibers were fabricated using a solution extrusion and neutralization method and incorporated into porous chitosan scaffolds. The effects of different fiber/scaffold mass ratios, fiber mechanical properties and fiber lengths on scaffold mechanical properties were studied. The results showed that incorporating fibers improved scaffold strength and stiffness in proportion to the fiber/scaffold mass ratio. A fiber-reinforced heart valve leaflet scaffold achieved strength values comparable to the radial values of human pulmonary and aortic valves. Additionally, the effects of shorter fibers (2 mm) were found to be up to 3-fold greater than longer fibers (10 mm). Despite this reduction in fiber mechanical properties caused by heparin crosslinking, the heparin-modified fibers still improved the mechanical properties of the reinforced scaffolds, but to a lesser extent than the unmodified fibers. The results demonstrate that chitosan fiber-reinforcement can be used to generate tissue-matching mechanical properties in porous chitosan scaffolds and that fiber length and mechanical properties are important parameters in defining the degree of mechanical improvement. We further studied various chemical and physical treatments to improve the mechanical properties of chitosan fibers. With combination of chemical and physical treatments, fiber stiffness improved 40fold compared to unmodified fibers. We also isolated ovine bone marrow-derived MSCs and evaluated their

  15. Lectin-like oxidized LDL receptor-1 expresses in mouse bone marrow-derived mesenchymal stem cells and stimulates their proliferation

    SciTech Connect

    Zhang, Fenxi; Wang, Congrui; Jing, Suhua; Ren, Tongming; Li, Yonghai; Cao, Yulin; Lin, Juntang

    2013-04-15

    The bone marrow-derived mesenchymal stem cells (bmMSCs) have been widely used in cell transplant therapy, and the proliferative ability of bmMSCs is one of the determinants of the therapy efficiency. Lectin-like oxidized low density lipoprotein receptor-1 (LOX-1) as a transmembrane protein is responsible for binding, internalizing and degrading oxidized low density lipoprotein (ox-LDL). It has been identified that LOX-1 is expressed in endothelial cells, vascular smooth muscle cells, cardiomyocytes, fibroblasts and monocytes. In these cells, low concentration of ox-LDL (<40 μg/mL) stimulates their proliferation via LOX-1 activation. However, it is poor understood that whether LOX-1 is expressed in bmMSCs and which role it plays. In this study, we investigated the status of LOX-1 expression in bmMSCs and its function on bmMSC proliferation. Our results showed that primary bmMSCs exhibiting a typical fibroblast-like morphology are positive for CD44 and CD90, but negative for CD34 and CD45. LOX-1 in both mRNA and protein levels is highly expressed in bmMSCs. Meanwhile, bmMSCs exhibit a strong potential to take up ox-LDL. Moreover, LOX-1 expression in bmMSCs is upregulated by ox-LDL with a dose- and time-dependent manner. Presence of ox-LDL also enhances the proliferation of bmMSCs. Knockdown of LOX-1 expression significantly inhibits ox-LDL-induced bmMSC proliferation. These findings indicate that LOX-1 plays a role in bmMSC proliferation. - Highlights: ► LOX-1 expresses in bmMSCs and mediates uptake of ox-LDL. ► Ox-LDL stimulates upregulation of LOX-1 in bmMSCs. ► Ox-LDL promotes bmMSC proliferation and expression of Mdm2, phosphor-Akt, phosphor-ERK1/2 and phosphor-NF-κB. ► LOX-1 siRNA inhibits ox-LDL-induced bmMSC proliferation and expression cell survival signals.

  16. Effects of Acellular Amniotic Membrane Matrix and Bone Marrow-Derived Mesenchymal Stem Cells in Improving Random Skin Flap Survival in Rats

    PubMed Central

    Chehelcheraghi, Farzaneh; Eimani, Hossein; Homayoonsadraie, Seyed; Torkaman, Giti; Amini, Abdollah; Alavi Majd, Hamid; Shemshadi, Hashem

    2016-01-01

    Background The necrotic skin flap represents a great challenge in plastic and reconstructive surgery. In this study, we evaluated the effect of bioscaffolds, acellular amniotic membranes (AAMs), and bone marrow-derived mesenchymal stem cells (BM-MSCs) on random skin flap (RSF) survival in rats by applying a cell-free extracellular matrix scaffold as a supportive component for the growth and proliferation of BM-MSCs on RSFs. AAM matrix scaffolds were created by incubating AMs in ethylenediaminetetraacetic acid 0.05% at 37°C, and cell scrapers were used. Objectives The aim of the present study was to assess the effect of AAM as a scaffold in TE, and combined with transplanted BM-MSCs, on the survival of RSFs and on the biomechanical parameters of the incision-wound flap margins 7 days after flap elevation. Materials and Methods BM-MSCs and AAMs were transplanted into subcutaneous tissue in the flap area. On the 7th postoperative day, the surviving flap areas were measured using digital imaging software, and the flap tissue was collected for evaluation. Forty rats were randomly divided into four groups of 10 each: group 1 received an AAM injection; group 2 underwent BM-MSC transplantation; group 3 received both AAM injection + BM-MSC transplantation; and group 4 was the control group, receiving only saline. Results The survival area in the AAM/BM-MSC group was significantly higher than in the control group (18.49 ± 1.58 versus 7.51 ± 2.42, P < 0.05). The biomechanical assessment showed no significant differences between the experimental groups and the control group (P > 0.05), and there was no correlation with flap survival. Conclusions Our findings showed that the treatment of flaps with BM-MSC and AAM transplantations significantly promoted flap survival compared to a control group. The viability of the flap was improved by combining BM-MSCs with AAM matrix scaffolds. PMID:27621924

  17. Effects of bone marrow-derived mesenchymal stem cells on the autophagic activity of alveolar macrophages in a rat model of silicosis

    PubMed Central

    ZHU, HUI-XING; GAO, JUN-LING; ZHAO, MAN-MAN; LI, RAN; TIAN, YAN-XIA; WANG, XIN; ZHANG, JUAN; YUAN, JU-XIANG; CUI, JIAN-ZHONG

    2016-01-01

    The aim of the present study was to evaluate the effects of bone marrow-derived mesenchymal stem cells (BMSCs) on the expression of the autophagy-associated proteins, microtubule-associated protein light chain 3 (LC-3) and autophagy-related gene Beclin-1 (Beclin-1), in alveolar macrophages (AMs) in a rat model of silicosis. Furthermore, the study investigated the molecular mechanisms underlying the effects of BMSC treatment. A population of 60 adult female Sprague-Dawley (SD) rats were allocated at random into three groups, namely the control, model and BMSC treatment groups (n=20 per group). BMSCs were isolated from five male SD rats (age, 6–8 weeks) and cultured in vitro. The silicosis model was established using a single 1.0-ml infusion of silicon dioxide suspension administered via non-exposed tracheal intubation. Rats in the BMSC treatment group received a 1.0-ml transplantation of BMSCs (1×106/ml). The rats were sacrificed on days 1, 7, 14 and 28 after modeling, and AMs were extracted from the rats using bronchoalveolar lavage. Third-generation BMSCs were identified using flow cytometry with fluorescein isothiocyanate staining, and the morphological characteristics of the AMs were observed using hematoxylin and eosin staining. The expression levels of LC-3 and Beclin-1 were determined using immunocytochemistry sand western blot analysis. The expression levels of LC-3 and Beclin-1 were found to be increased at all the time points in the model group. LC-3 and Beclin-1 levels began to increase at day 1, peaked at day 14 and decreased after day 28; however, the levels remained elevated compared with the basal expression levels. The AMs of the BMSC treatment group exhibited significantly alleviated pathological symptoms compared with the model group AMs, as indicated by significantly decreased expression levels of LC-3 and Beclin-1 at each time point. Therefore, the results indicated that autophagy was promoted in the AMs of the silicosis model rats

  18. Intranasal delivery of bone marrow-derived mesenchymal stem cells, macrophages, and microglia to the brain in mouse models of Alzheimer's and Parkinson's disease.

    PubMed

    Danielyan, Lusine; Beer-Hammer, Sandra; Stolzing, Alexandra; Schäfer, Richard; Siegel, Georg; Fabian, Claire; Kahle, Philipp; Biedermann, Tilo; Lourhmati, Ali; Buadze, Marine; Novakovic, Ana; Proksch, Barbara; Gleiter, Christoph H; Frey, William H; Schwab, Matthias

    2014-01-01

    In view of the rapid preclinical development of cell-based therapies for neurodegenerative disorders, traumatic brain injury, and tumors, the safe and efficient delivery and targeting of therapeutic cells to the central nervous system is critical for maintaining therapeutic efficacy and safety in the respective disease models. Our previous data demonstrated therapeutically efficacious and targeted delivery of mesenchymal stem cells (MSCs) to the brain in the rat 6-hydroxydopamine model of Parkinson's disease (PD). The present study examined delivery of bone marrow-derived MSCs, macrophages, and microglia to the brain in a transgenic model of PD [(Thy1)-h[A30P] αS] and an APP/PS1 model of Alzheimer's disease (AD) via intranasal application (INA). INA of microglia in naive BL/6 mice led to targeted and effective delivery of cells to the brain. Quantitative PCR analysis of eGFP DNA showed that the brain contained the highest amount of eGFP-microglia (up to 2.1 × 10(4)) after INA of 1 × 10(6) cells, while the total amount of cells detected in peripheral organs did not exceed 3.4 × 10(3). Seven days after INA, MSCs expressing eGFP were detected in the olfactory bulb (OB), cortex, amygdala, striatum, hippocampus, cerebellum, and brainstem of (Thy1)-h[A30P] αS transgenic mice, showing predominant distribution within the OB and brainstem. INA of eGFP-expressing macrophages in 13-month-old APP/PS1 mice led to delivery of cells to the OB, hippocampus, cortex, and cerebellum. Both MSCs and macrophages contained Iba-1-positive population of small microglia-like cells and Iba-1-negative large rounded cells showing either intracellular amyloid β (macrophages in APP/PS1 model) or α-synuclein [MSCs in (Thy1)-h[A30P] αS model] immunoreactivity. Here, we show, for the first time, intranasal delivery of cells to the brain of transgenic PD and AD mouse models. Additional work is needed to determine the optimal dosage (single treatment regimen or repeated

  19. The Effect of Purified Multi-potent Human Bone-marrow Derived Mesenchymal Stem Cells on Rotator Cuff Tendon Healing in an Athymic Rat

    PubMed Central

    Degen, Ryan; Carbone, Andrew; Carballo, Camilia; Zong, Jianchun; Chen, Tony; Ying, Lilly; Lebaschi, Amir; Deng, Xiang-Hua; Rodeo, Scott A.

    2016-01-01

    Objectives: Bone marrow concentrates are being used to augment soft tissue healing. However, only 0.01% of these cells meet the criteria of a mesenchymal stem cell (MSC), which likely accounts for the variability in reported results. Previous studies using an established rat rotator cuff repair model have demonstrated that bone marrow-derived MSCs had no effect on healing. In this study we evaluated the effect of purified human MSCs on rotator cuff healing in an athymic rat model. Our hypothesis is that purified human MSCs added to the repair site will improve biomechanical strength and fibrocartilage formation of the healing tendon. Methods: Fifty-two athymic rats underwent unilateral detachment and repair of the supraspinatus tendon with either fibrin glue (control) or fibrin glue with 106 hMSCs (experimental) applied at the repair site. Flow cytometry verified the stem cell phenotype of the cells as CD73+, CD90+, CD105+, CD 14-, CD34- and CD45-. Rats were sacrificed at 2 and 4 weeks, with 10 used for biomechanical testing and 3 for histologic analysis from each group. Results: Biomechanical testing revealed a significant increase in failure load (11.5±2.4 N vs. 8.5±2.4 N, p=0.002) and stiffness (7.1±1.2 N/mm vs. 5.7±2.1 N/mm, p>0.17).Biomechanical testing revealed a significant increase in failure load (11.5±2.4 N vs. 8.5±2.4 N, p=0.002) and stiffness (7.1±1.2 N/mm vs. 5.7±2.1 N/mm, p<0.001) in the experimental group compared with the control group at 2 weeks. Similarly, safranin-O staining identified increased fibrocartilage formation at the repair site at 2 weeks in the hMSC group (18.6±2.9% vs. 9.1±1.6%, p=0.026). These effects dissipated by 4 weeks, with no significant biomechanical or histologic differences between groups (p>0.17). Conclusion: These data demonstrate the potential for stem cells to augment tendon healing. This is the first study to use purified stem cells, rather than simple bone marrow concentrate. In the future, cell

  20. Functional Comparison of Human-Induced Pluripotent Stem Cell-Derived Mesenchymal Cells and Bone Marrow-Derived Mesenchymal Stromal Cells from the Same Donor

    PubMed Central

    Diederichs, Solvig

    2014-01-01

    Mesenchymal stem cells (MSCs) have a high potential for therapeutic efficacy in treating diverse musculoskeletal injuries and cardiovascular diseases, and for ameliorating the severity of graft-versus-host and autoimmune diseases. While most of these clinical applications require substantial cell quantities, the number of MSCs that can be obtained initially from a single donor is limited. Reports on the derivation of MSC-like cells from pluripotent stem cells (PSCs) are, thus, of interest, as the infinite proliferative capacity of PSCs opens the possibility to generate large amounts of uniform batches of MSCs. However, characterization of such MSC-like cells is currently inadequate, especially with regard to the question of whether these cells are equivalent or identical to MSCs. In this study, we have derived MSC-like cells [induced PSC-derived MSC-like progenitor cells (iMPCs)] using four different methodologies from a newly established induced PSC line reprogrammed from human bone marrow stromal cells (BMSCs), and compared the iMPCs directly with the originating parental BMSCs. The iMPCs exhibited typical MSC/fibroblastic morphology and MSC-typical surface marker profile, and they were capable of differentiation in vitro along the osteogenic, chondrogenic, and adipogenic lineages. However, compared with the parental BMSCs, iMPCs displayed a unique expression pattern of mesenchymal and pluripotency genes and were less responsive to traditional BMSC differentiation protocols. We, therefore, conclude that iMPCs generated from PSCs via spontaneous differentiation represent a distinct population of cells which exhibit MSC-like characteristics. PMID:24625206

  1. Co-infusion of autologous adipose tissue derived neuronal differentiated mesenchymal stem cells and bone marrow derived hematopoietic stem cells, a viable therapy for post-traumatic brachial plexus injury: a case report.

    PubMed

    Thakkar, Umang G; Vanikar, Aruna V; Trivedi, Hargovind L

    2014-01-01

    Stem cell therapy is emerging as a viable approach in regenerative medicine. A 31-year-old male with brachial plexus injury had complete sensory-motor loss since 16 years with right pseudo-meningocele at C5-D1 levels and extra-spinal extension up to C7-D1, with avulsion on magnetic resonance imaging and irreversible damage. We generated adipose tissue derived neuronal differentiated mesenchymal stem cells (N-AD-MSC) and bone marrow derived hematopoietic stem cells (HSC-BM). Neuronal stem cells expressed β-3 tubulin and glial fibrillary acid protein which was confirmed on immunofluorescence. On day 14, 2.8 ml stem cell inoculum was infused under local anesthesia in right brachial plexus sheath by brachial block technique under ultrasonography guidance with a 1.5-inch-long 23 gauge needle. Nucleated cell count was 2 × 10 4 /μl, CD34+ was 0.06%, and CD45-/90+ and CD45-/73+ were 41.63% and 20.36%, respectively. No untoward effects were noted. He has sustained recovery with re-innervation over a follow-up of 4 years documented on electromyography-nerve conduction velocity study. PMID:25116721

  2. Novel daidzein analogs enhance osteogenic activity of bone marrow-derived mesenchymal stem cells and adipose-derived stromal/stem cells through estrogen receptor dependent and independent mechanisms

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Osteoporosis is a disease characterized by low bone mineral density (BMD) and increased risk of fractures. Studies have demonstrated the use of phytoestrogens, or plant-derived estrogens, such as genistein anddaidzein, to effectively increase osteogenic activity of bone marrow-derived mesenchymal s...

  3. Bone marrow-derived mesenchymal stem cells induce both polyclonal expansion and differentiation of B cells isolated from healthy donors and systemic lupus erythematosus patients.

    PubMed

    Traggiai, Elisabetta; Volpi, Stefano; Schena, Francesca; Gattorno, Marco; Ferlito, Francesca; Moretta, Lorenzo; Martini, Alberto

    2008-02-01

    Human bone marrow multipotent mesenchymal stromal cells are progenitor cells that can be expanded in vitro and differentiate into various cells of mesodermal origin. They contribute to the bone marrow reticular niche, where mature B cells and long-lived plasma cells are maintained. Multipotent mesenchymal stromal cells were recently shown to modulate T- and B-cell proliferation and differentiation, dendritic cell maturation, and natural killer activity. These immunoregulatory properties encouraged a possible use of these cells to modulate autoimmune responses in humans. We studied the influence of bone marrow mesenchymal stem cells on highly purified B-cell subsets isolated from healthy donors and total B cells from pediatric systemic lupus erythematosus patients. Bone marrow mesenchymal stem cells promoted proliferation and differentiation into immunoglobulin-secreting cells of transitional and naive B cells stimulated with an agonist of Toll-like receptor 9, in the absence of B cell receptor triggering. They strongly enhanced proliferation and differentiation into plasma cells of memory B-cell populations. A similar effect was observed in response to polyclonal stimulation of B cells isolated from pediatric patients with systemic lupus erythematosus. This study casts important questions on bone marrow mesenchymal stem cells as a therapeutic tool in autoimmune diseases in which B-cell activation is crucially implicated in the pathogenesis of the disease. PMID:18024418

  4. Co-infusion of autologous adipose tissue derived insulin-secreting mesenchymal stem cells and bone marrow derived hematopoietic stem cells: viable therapy for type III.C. a diabetes mellitus.

    PubMed

    Thakkar, Umang G; Vanikar, Aruna V; Trivedi, Hargovind L

    2013-01-01

    Transition from acute pancreatitis to insulin-dependent diabetes mellitus (IDDM) is a rare manifestation of primary hyperparathyroidism caused by parathyroid adenoma because of impaired glucose tolerance and suppresses insulin secretion. We report the case of a 26-year-old male with pancreatic diabetes caused by parathyroid adenoma induced chronic pancreatitis. He had serum C-peptide 0.12 ng/ml, glutamic acid decarboxylase antibody 5.0 IU/ml, and glycosylated hemoglobin (HbA1C) 8.9%, and required 72 IU/day of biphasic-isophane insulin injection for uncontrolled hyperglycemia. We treated him with his own adipose tissue derived insulin-secreting mesenchymal stem-cells (IS-ADMSC) along with his bone marrow derived hematopoietic stem cells (BM-HSC). Autologous IS-ADMSC + BM-HSC were infused into subcutaneous tissue, portal and thymic circulation without any conditioning. Over a follow-up of 27 months, the patient is maintaining fasting and postprandial blood sugar levels of 132 and 165 mg/dl, respectively, with HbA1C 6.8% and requiring 36 IU/day of biphasic-isophane insulin. Co-infusion of IS-ADMSC + BM-HSC offers a safe and viable therapy for type III.C.a Diabetes Mellitus. PMID:24385073

  5. Cigarette smoke inhibits recruitment of bone-marrow-derived stem cells to the uterus.

    PubMed

    Zhou, Yuping; Gan, Ye; Taylor, Hugh S

    2011-02-01

    Cigarette smoking leads to female infertility and a decreased incidence of endometriosis. Bone marrow derived stem cells are recruited to uterine endometrium and endometriosis. The effect of cigarette smoking on stem cell recruitment to any organ is uncharacterized. We hypothesized that bone marrow-derived mesenchymal stem cell recruitment to the uterus and differentiation would be diminished by cigarette smoke. We used human mesenchymal stem cells (hMSC) in vitro and a mouse model of cigarette smoke exposure. After myeloablation female C57BL/6J received bone marrow cells from males. Mice were exposed to room air or smoke from unfiltered cigarettes. Immunofluorescence and Y-FISH was performed on uterine sections. In vitro hMSCs were treated with 8-Br-cAMP to induce endometrial cell differentiation with or without cigarette smoke extract (CSE) and decidualization assessed morphologically and by prolactin expression. After 4 weeks the total number of Y-chromosome cells in the uterus was reduced by 68% in the smoke exposed mice. Both leukocytes and bone marrow derived endometrial cells were reduced by 60% and 73%, respectively. Differentiation of bone marrow derived cell to endometrial epithelial cells was reduced by 84%. hMSC treated with CSE failed to show cytological characteristics of decidualization. mRNA levels of the decidualization marker prolactin were decreased by 90% in CSE treated cells. Smoking inhibits both recruitment of bone marrow derived stem cells to uterus and stem cell differentiation. Inhibition of stem cells recruitment may be a general mechanism by which smoking leads to long term organ damage through inability to repair or regenerate multiple tissues. PMID:20955787

  6. Brazilian minipig as a large-animal model for basic research and stem cell-based tissue engineering. Characterization and in vitro differentiation of bone marrow-derived mesenchymal stem cells

    PubMed Central

    STRAMANDINOLI-ZANICOTTI, Roberta Targa; CARVALHO, André Lopes; REBELATTO, Carmen Lúcia Kuniyoshi; SASSI, Laurindo Moacir; TORRES, Maria Fernanda; SENEGAGLIA, Alexandra Cristina; BOLDRINILEITE, Lidiane Maria; CORREA-DOMINGUEZ, Alejandro; KULIGOVSKY, Crisciele; BROFMAN, Paulo Roberto Slud

    2014-01-01

    Stem cell-based regenerative medicine is one of the most intensively researched medical issues. Pre-clinical studies in a large-animal model, especially in swine or miniature pigs, are highly relevant to human applications. Mesenchymal stem cells (MSCs) have been isolated and expanded from different sources. Objective This study aimed at isolating and characterizing, for the first time, bone marrow-derived MSCs (BM-MSCs) from a Brazilian minipig (BR1). Also, this aimed to validate a new large-animal model for stem cell-based tissue engineering. Material and Methods Bone marrow (BM) was aspirated from the posterior iliac crest of twelve adult male BR1 under general anesthesia. MSCs were selected by plastic-adherence as originally described by Friedenstein. Cell morphology, surface marker expression, and cellular differentiation were examined. The immunophenotypic profile was determined by flow cytometry. The differentiation potential was assessed by cytological staining and by RT-PCR. Results MSCs were present in all minipig BM samples. These cells showed fibroblastic morphology and were positive for the surface markers CD90 (88.6%), CD29 (89.8%), CD44 (86.9%) and negative for CD34 (1.61%), CD45 (1.83%), CD14 (1.77%) and MHC-II (2.69%). MSCs were differentiated into adipocytes, osteoblasts, and chondroblasts as demonstrated by the presence of lipidic-rich vacuoles, the mineralized extracellular matrix, and the great presence of glycosaminoglycans, respectively. The higher gene expression of adipocyte fatty-acid binding protein (AP2), alkaline phosphatase (ALP) and collagen type 2 (COLII) also confirmed the trilineage differentiation (p<0.001, p<0.001, p=0.031; respectively). Conclusions The isolation, cultivation, and differentiation of BM-MSCs from BR1 makes this animal eligible as a useful large-animal model for stem cell-based studies in Brazil. PMID:25025563

  7. Forced expression of Sox2 or Nanog in human bone marrow derived mesenchymal stem cells maintains their expansion and differentiation capabilities

    SciTech Connect

    Go, Masahiro J. Takenaka, Chiemi; Ohgushi, Hajime

    2008-03-10

    Mesenchymal stem cells (MSCs) derived from human bone marrow have capability to differentiate into cells of mesenchymal lineage. The cells have already been applied in various clinical situations because of their expansion and differentiation capabilities. The cells lose their capabilities after several passages, however. With the aim of conferring higher capability on human bone marrow MSCs, we introduced the Sox2 or Nanog gene into the cells. Sox2 and Nanog are not only essential for pluripotency and self-renewal of embryonic stem cells, but also expressed in somatic stem cells that have superior expansion and differentiation potentials. We found that Sox2-expressing MSCs showed consistent proliferation and osteogenic capability in culture media containing basic fibroblast growth factor (bFGF) compared to control cells. Significantly, in the presence of bFGF in culture media, most of the Sox2-expressing cells were small, whereas the control cells were elongated in shape. We also found that Nanog-expressing cells even in the absence of bFGF had much higher capabilities for expansion and osteogenesis than control cells. These results demonstrate not only an effective way to maintain proliferation and differentiation potentials of MSCs but also an important implication about the function of bFGF for self-renewal of stem cells including MSCs.

  8. The Effect of Recombinant Human Interleukin-6 on Osteogenic Differentiation and YKL-40 Expression in Human, Bone Marrow-Derived Mesenchymal Stem Cells.

    PubMed

    Lieder, Ramona; Sigurjonsson, Olafur E

    2014-02-01

    Human mesenchymal stem cells are an attractive cell source for tissue engineering and regenerative medicine applications, especially because of their differentiation potential toward the mesenchymal lineage. Furthermore, this cell type participates in the regeneration of tissue damage and plays an important role in immunity. Similarly, chitinase-like proteins have been proposed to aid in tissue remodeling, inflammation, and differentiation processes. The chitinase-like protein YKL-40 in particular is indicated in preventing damage to the extracellular matrix in response to proinflammatory cytokines, even though its biological function remains speculative. Finally, interleukin (IL)-6, a pleiotropic acute phase protein, participates in the regulation of bone turnover and immunoregulation. The physiological role of IL-6 in bone homeostasis is complex, exerting different effects on osteoblasts and osteoclasts depending on their differentiation stage. The aim of this study was to determine the effect of recombinant human IL-6 (5 ng/mL) on YKL-40 expression and osteogenic differentiation of human mesenchymal stem cells. Recombinant human IL-6 induced a donor-dependent change in mineralization and significantly promoted YKL-40 protein secretion. However, YKL-40 gene expression remained unaffected, and no statistically significant differences in the expression of osteogenic marker genes could be observed. PMID:24570843

  9. Bone-marrow-derived mesenchymal stem cells as a target for cytomegalovirus infection: Implications for hematopoiesis, self-renewal and differentiation potential

    SciTech Connect

    Smirnov, Sergey V.; Harbacheuski, Ryhor; Lewis-Antes, Anita; Zhu Hua; Rameshwar, Pranela; Kotenko, Sergei V. . E-mail: kotenkse@umdnj.edu

    2007-03-30

    Mesenchymal stem cells (MSCs) in bone marrow (BM) regulate the differentiation and proliferation of adjacent hematopoietic precursor cells and contribute to the regeneration of mesenchymal tissues, including bone, cartilage, fat and connective tissue. BM is an important site for the pathogenesis of human cytomegalovirus (HCMV) where the virus establishes latency in hematopoietic progenitors and can transmit after reactivation to neighboring cells. Here we demonstrate that BM-MSCs are permissive to productive HCMV infection, and that HCMV alters the function of MSCs: (i) by changing the repertoire of cell surface molecules in BM-MSCs, HCMV modifies the pattern of interaction between BM-MSCs and hematopoietic cells; (ii) HCMV infection of BM-MSCs undergoing adipogenic or osteogenic differentiation impaired the process of differentiation. Our results suggest that by altering BM-MSC biology, HCMV may contribute to the development of various diseases.

  10. Unexpected complication in a rat stroke model: exacerbation of secondary pathology in the thalamus by subacute intraarterial administration of human bone marrow-derived mesenchymal stem cells

    PubMed Central

    Mitkari, Bhimashankar; Kerkelä, Erja; Nystedt, Johanna; Korhonen, Matti; Jolkkonen, Jukka

    2015-01-01

    This study examined whether human bone marrow mesenchymal stromal/stem cells (BMMSCs) could alleviate the secondary pathology in the thalamus after middle cerebral artery occlusion (MCAO) in rats. Atypical accumulation of both amyloid-β (Aβ) and calcium in the thalamus was significantly higher in rats receiving the BMMSCs infusion 48 hours after MCAO as compared with the vehicle MCAO group. The elevated Aβ/calcium accumulation correlated with the level of impaired sensorimotor function. Although secondary pathology in the thalamus seems to be rodent specific, it needs to be taken into account because it may impair long-term behavioral recovery and negate therapeutic treatment effects. PMID:25564231

  11. Intratracheal transplantation of bone marrow-derived mesenchymal stem cells reduced airway inflammation and up-regulated CD4⁺CD25⁺ regulatory T cells in asthmatic mouse.

    PubMed

    Ge, Xiahui; Bai, Chong; Yang, Jianming; Lou, Guoliang; Li, Qiang; Chen, Ruohua

    2013-07-01

    Mesenchymal stem cells attenuate the severity of lung injury due to their immunomodulatory properties. The effect of bone marrow-derived mesenchymal stem cells on asthma is seldom reported. We have examined the effect of BMSCs on airway inflammation in asthma. Forty female BALB/c mice were equally randomised into PBS group, BMSCs treatment group, BMSCs control group and asthmatic group. Reactivity of the airway to acetylcholine was measured by barometric plethysmography. Cytokine profiles of bronchoalveolar lavage fluid and serum were determined by enzyme-linked immunosorbent assay. Morphometric analysis was done with haematoxylin and periodic-acid Schiff staining. Engraftment of BMSCs in asthmatic mice significantly decreased the number of eosinophils and mononuclear cells in bronchoalveolar lavage fluid and the airway (P < 0.05). Both goblet cell hyperplasia and responsiveness to acetylcholine were significantly reduced in BMSCs treatment groups. Moreover, BMSCs engraftment caused significant increases the ratio of Treg in pulmonary lymph node and interleukin-10 (IL-10) and interleukin-12 levels in BALF and serum. We conclude that BMSCs engraftment ameliorated airway inflammation and improved lung function in asthmatic mouse and the protective effect might be mediated by upregulating Treg and partly involved with increasing IL-10. PMID:23483727

  12. Bone-Marrow-Derived Mesenchymal Stem Cells Promote Proliferation and Neuronal Differentiation of Niemann–Pick Type C Mouse Neural Stem Cells by Upregulation and Secretion of CCL2

    PubMed Central

    Lee, Hyun; Kang, Ji Eun; Lee, Jong Kil; Bae, Jae-sung

    2013-01-01

    Abstract Niemann–Pick type C (NP-C) disease is a neurodegenerative disorder characterized neuropathologically by ballooned neurons distended with lipid storage and widespread neuronal loss. Neural stem cells (NSC) derived from NP-C disease models have decreased ability for self-renewal and neuronal differentiation. Investigation of neurogenesis in the adult brain has suggested that NP-C disease can be overcome, or at least ameliorated, by the generation of new neurons. Bone-marrow-derived mesenchymal stem cells (BM-MSCs) are regarded as potential candidates for use in the treatment of neurodegenerative disorders because of their ability to promote neurogenesis. The underlying mechanisms of BM-MSC-induced promotion of neurogenesis, however, have not been resolved. The aim of the present study was to examine the mechanism of neurogenesis by BM-MSCs in NP-C disease. Coculture of embryonic NSCs from NP-C mice that exhibit impaired ability for self-renewal and decreased rates of neuronal differentiation with BM-MSCs resulted in an enhanced capacity for self-renewal and an increased ability for differentiation into neurons or oligodendrocytes. In addition, results of in vivo studies have demonstrated that transplantation of intracerebral BM-MSCs resulted in stimulated proliferation and neuronal differentiation of NSCs within the subventricular zone. Of particular interest, enhanced proliferation and neuronal differentiation of endogenous NP-C mouse NSCs showed an association with elevated release of the chemokine (C-C motif) ligand 2 (CCL2) from BM-MSCs. These effects suggest that soluble CCL2 derived from BM-MSCs can modulate endogenous NP-C NSCs, resulting in their improved proliferation and neuronal differentiation in mice. PMID:23659480

  13. Bone Marrow-Derived Mesenchymal Stem Cell Therapy as a Candidate Disease-Modifying Strategy in Parkinson's Disease and Multiple System Atrophy

    PubMed Central

    Park, Hyun Jung

    2009-01-01

    Parkinson's disease (PD) and multiple system atrophy (MSA) are neurodegenerative diseases representative of α-synucleinopathies characterized pathologically by α-synuclein-abundant Lewy bodies and glial cytoplasmic inclusions, respectively. Embryonic stem cells, fetal mesencephalic neurons, and neural stem cells have been introduced as restorative strategies in PD animals and patients, but ethical and immunological problems as well as the serious side effects of tumorigenesis and disabling dyskinesia have limited clinical application of these stem cells. Meanwhile, cell therapy using mesenchymal stem cells (MSCs) is attractive clinically because these cells are free from ethical and immunological problems. MSCs are present in adult bone marrow and represent <0.01% of all nucleated bone marrow cells. MSCs are themselves capable of multipotency, differentiating under appropriate conditions into chondrocytes, skeletal myocytes, and neurons. According to recent studies, the neuroprotective effect of MSCs is mediated by their ability to produce various trophic factors that contribute to functional recovery, neuronal cell survival, and stimulation of endogenous regeneration and by immunoregulatory properties that not only inhibit nearly all cells participating in the immune response cell-cell-contact-dependent mechanism, but also release various soluble factors associated with immunosuppressive activity. However, the use of MSCs as neuroprotectives in PD and MSA has seldom been studied. Here we comprehensively review recent advances in the therapeutic roles of MSCs in PD and MSA, especially focusing on their neuroprotective properties and use in disease-modifying therapeutic strategies. PMID:19513327

  14. Monocyte Chemoattractant Protein-Induced Protein 1 (MCPIP1) Enhances Angiogenic and Cardiomyogenic Potential of Murine Bone Marrow-Derived Mesenchymal Stem Cells

    PubMed Central

    Labedz-Maslowska, Anna; Lipert, Barbara; Berdecka, Dominika; Kedracka-Krok, Sylwia; Jankowska, Urszula; Kamycka, Elzbieta; Sekula, Malgorzata; Madeja, Zbigniew; Dawn, Buddhadeb; Jura, Jolanta; Zuba-Surma, Ewa K.

    2015-01-01

    The current evidence suggests that beneficial effects of mesenchymal stem cells (MSCs) toward myocardial repair are largely due to paracrine actions of several factors. Although Monocyte chemoattractant protein-induced protein 1 (MCPIP1) is involved in the regulation of inflammatory response, apoptosis and angiogenesis, whether MCPIP1 plays any role in stem cell-induced cardiac repair has never been examined. By employing retroviral (RV)-transduced overexpression of MCPIP1, we investigated the impact of MCPIP1 on viability, apoptosis, proliferation, metabolic activity, proteome, secretome and differentiation capacity of murine bone marrow (BM) - derived MSCs. MCPIP1 overexpression enhanced angiogenic and cardiac differentiation of MSCs compared with controls as indicated by elevated expression of genes accompanying angiogenesis and cardiomyogenesis in vitro. The proangiogenic activity of MCPIP1-overexpressing MSCs (MCPIP1-MSCs) was also confirmed by increased capillary-like structure formation under several culture conditions. This increase in differentiation capacity was associated with decreased proliferation of MCPIP1-MSCs when compared with controls. MCPIP1-MSCs also expressed increased levels of proteins involved in angiogenesis, autophagy, and induction of differentiation, but not adverse inflammatory agents. We conclude that MCPIP1 enhances endothelial and cardiac differentiation of MSCs. Thus, modulating MCPIP1 expression may be a novel approach useful for enhancing the immune-regulatory, anti-apoptotic, anti-inflammatory and regenerative capacity of BM-derived MSCs for myocardial repair and regeneration of ischemic tissues. PMID:26214508

  15. The Effect of Bone Marrow-Derived Mesenchymal Stem Cell Transplantation on Allodynia and Hyperalgesia in Neuropathic Animals: A Systematic Review with Meta-Analysis.

    PubMed

    Hosseini, Mostafa; Yousefifard, Mahmoud; Aziznejad, Heidar; Nasirinezhad, Farinaz

    2015-09-01

    Stem cell transplantation has been considered a possible therapeutic method for neuropathic pain. However, no quantitative data synthesis of stem cell therapy for neuropathic pain exists. Therefore, the present systematic review and meta-analysis assessed the efficacy of bone marrow mesenchymal stem cell (BMMSC) transplantation on alleviating pain symptoms in animal models of neuropathic pain. In the present meta-analysis, controlled animal studies assessing the effect of administrating BMMSC on neuropathic pain were included through an extensive literature search of online databases. After collecting data, effect sizes were computed and the standardized mean difference (SMD) with 95% confidence interval (CI) was entered in all analyses. Random-effects models were used for data analysis. Sensitivity and subgroup analyses were performed to investigate expected or measured heterogeneity. Finally, 14 study were included. The analyses showed that BMMSC transplantation lead to significant improvement on allodynia (SMD = 2.06; 95% CI, 1.09 to 3.03; I(2) = 99.7%; P < .001). The type of neuropathy (P = .036), time between injury and intervention (P = .02), and the number of transplanted cells (P = .023) influence the improvement of allodynia after BMMSC transplantation. BMMSC transplantation has no effect on hyperalgesia (SMD = .3; 95% CI, -1.09 to 1.68; I(2) = 100%; P < .001) unless it occurs during the first 4 days after injury (P = .02). The present systematic review with meta-analysis suggests that BMMSC transplantation improves allodynia but does not have any significant effect on hyperalgesia unless it is given during the first 4 days after injury. PMID:25985918

  16. Trichostatin A-Mediated Epigenetic Transformation of Adult Bone Marrow-Derived Mesenchymal Stem Cells Biases the In Vitro Developmental Capability, Quality, and Pluripotency Extent of Porcine Cloned Embryos

    PubMed Central

    Samiec, Marcin; Opiela, Jolanta; Lipiński, Daniel

    2015-01-01

    The current research was conducted to explore the in vitro developmental outcome and cytological/molecular quality of porcine nuclear-transferred (NT) embryos reconstituted with adult bone marrow-derived mesenchymal stem cells (ABM-MSCs) that were epigenetically transformed by treatment with nonspecific inhibitor of histone deacetylases, known as trichostatin A (TSA). The cytological quality of cloned blastocysts was assessed by estimation of the total cells number (TCN) and apoptotic index. Their molecular quality was evaluated by real-time PCR-mediated quantification of gene transcripts for pluripotency- and multipotent stemness-related markers (Oct4, Nanog, and Nestin). The morula and blastocyst formation rates of NT embryos derived from ABM-MSCs undergoing TSA treatment were significantly higher than in the TSA-unexposed group. Moreover, the NT blastocysts generated using TSA-treated ABM-MSCs exhibited significantly higher TCN and increased pluripotency extent measured with relative abundance of Oct4 and Nanog mRNAs as compared to the TSA-untreated group. Altogether, the improvements in morula/blastocyst yields and quality of cloned pig embryos seem to arise from enhanced abilities for promotion of correct epigenetic reprogramming of TSA-exposed ABM-MSC nuclei in a cytoplasm of reconstructed oocytes. To our knowledge, we are the first to report the successful production of mammalian high-quality NT blastocysts using TSA-dependent epigenomic modulation of ABM-MSCs. PMID:25866813

  17. Dynamics of bone marrow-derived endothelial progenitor cell/mesenchymal stem cell interaction in co-culture and its implications in angiogenesis

    SciTech Connect

    Aguirre, A.; Planell, J.A.; Engel, E.

    2010-09-17

    Research highlights: {yields} BM-EPCs and MSCs establish complex, self-organizing structures in co-culture. {yields} Co-culture decreases proliferation by cellular self-regulatory mechanisms. {yields} Co-cultured cells present an activated proangiogenic phenotype. {yields} qRT-PCR and cluster analysis identify new target genes playing important roles. -- Abstract: Tissue engineering aims to regenerate tissues and organs by using cell and biomaterial-based approaches. One of the current challenges in the field is to promote proper vascularization in the implant to prevent cell death and promote host integration. Bone marrow endothelial progenitor cells (BM-EPCs) and mesenchymal stem cells (MSCs) are bone marrow resident stem cells widely employed for proangiogenic applications. In vivo, they are likely to interact frequently both in the bone marrow and at sites of injury. In this study, the physical and biochemical interactions between BM-EPCs and MSCs in an in vitro co-culture system were investigated to further clarify their roles in vascularization. BM-EPC/MSC co-cultures established close cell-cell contacts soon after seeding and self-assembled to form elongated structures at 3 days. Besides direct contact, cells also exhibited vesicle transport phenomena. When co-cultured in Matrigel, tube formation was greatly enhanced even in serum-starved, growth factor free medium. Both MSCs and BM-EPCs contributed to these tubes. However, cell proliferation was greatly reduced in co-culture and morphological differences were observed. Gene expression and cluster analysis for wide panel of angiogenesis-related transcripts demonstrated up-regulation of angiogenic markers but down-regulation of many other cytokines. These data suggest that cross-talk occurs in between BM-EPCs and MSCs through paracrine and direct cell contact mechanisms leading to modulation of the angiogenic response.

  18. In Vivo Implanted Bone Marrow-Derived Mesenchymal Stem Cells Trigger a Cascade of Cellular Events Leading to the Formation of an Ectopic Bone Regenerative Niche

    PubMed Central

    Tasso, Roberta; Ulivi, Valentina; Reverberi, Daniele; Lo Sicco, Claudia; Descalzi, Fiorella

    2013-01-01

    We recently reported that mouse bone marrow stromal cells, also known as bone marrow (BM)-derived mesenchymal stem cells (MSCs), seeded onto a scaffold and implanted in vivo, led to an ectopic bone deposition by host cells. This MSCs capacity was critically dependent on their commitment level, being present only in MSCs cultured in presence of fibroblast growth factor-2. Taking advantage of a chimeric mouse model, in this study we show that seeded MSCs trigger a cascade of events resulting in the mobilization of macrophages, the induction of their functional switch from a proinflammatory to a proresolving phenotype, and the subsequent formation of a bone regenerative niche through the recruitment, within the first 2 weeks of implantation, of endothelial progenitors and of cells with an osteogenic potential (CD146+CD105+), both of them derived from the BM. Moreover, we demonstrated that, in an inflammatory environment, MSCs secrete a large amount of prostaglandin E2 playing a key role in the macrophage phenotype switch. PMID:23924051

  19. In Vitro Expansion of Bone Marrow Derived Mesenchymal Stem Cells Alters DNA Double Strand Break Repair of Etoposide Induced DNA Damage

    PubMed Central

    Hare, Ian; Gencheva, Marieta; Evans, Rebecca; Fortney, James; Piktel, Debbie; Vos, Jeffrey A.; Howell, David; Gibson, Laura F.

    2016-01-01

    Mesenchymal stem cells (MSCs) are of interest for use in diverse cellular therapies. Ex vivo expansion of MSCs intended for transplantation must result in generation of cells that maintain fidelity of critical functions. Previous investigations have identified genetic and phenotypic alterations of MSCs with in vitro passage, but little is known regarding how culturing influences the ability of MSCs to repair double strand DNA breaks (DSBs), the most severe of DNA lesions. To investigate the response to DSB stress with passage in vitro, primary human MSCs were exposed to etoposide (VP16) at various passages with subsequent evaluation of cellular damage responses and DNA repair. Passage number did not affect susceptibility to VP16 or the incidence and repair kinetics of DSBs. Nonhomologous end joining (NHEJ) transcripts showed little alteration with VP16 exposure or passage; however, homologous recombination (HR) transcripts were reduced following VP16 exposure with this decrease amplified as MSCs were passaged in vitro. Functional evaluations of NHEJ and HR showed that MSCs were unable to activate NHEJ repair following VP16 stress in cells after successive passage. These results indicate that ex vivo expansion of MSCs alters their ability to perform DSB repair, a necessary function for cells intended for transplantation. PMID:26880992

  20. Shp2 regulates chlorogenic acid-induced proliferation and adipogenic differentiation of bone marrow-derived mesenchymal stem cells in adipogenesis.

    PubMed

    Zhou, Rong-Ping; Deng, Ming-Tao; Chen, Lan-Ying; Fang, Ning; Du, Chuan; Chen, Lin-Pan; Zou, Ye-Qing; Dai, Jiang-Hua; Zhu, Mei-Lan; Wang, Wei; Lin, Si-Jian; Liu, Rong-Hua; Luo, Jun

    2015-06-01

    Chlorogenic acid (CGA) exhibits various biological properties, including the inhibition of oxidation, obesity, apoptosis and tumorigenesis. CGA is also able to promote cell survival and proliferation. The aim of the present study was to determine the effects and underlying molecular mechanisms of CGA on the adipogenesis of bone marrow‑derived mesenchymal stem cells (BMSCs). Treatment with CGA had a marginal effect on cell proliferation, by promoting the expression levels of phosphorylated Akt and cyclin D1. Furthermore, treatment with CGA also upregulated the phosphorylation of extracellular signal‑regulated kinase (Erk) and inhibited the adipocyte differentiation of BMSCs by inhibiting the expression of peroxisome proliferator‑activated receptor (PPAR)γ and CCAAT/enhancer binding protein (C/EBP)α. However, knockdown of the expression of Shp2 attenuated CGA‑induced proliferation and inhibited the phosphorylation of Akt and expression of cyclin D1. Furthermore, CGA treatment upregulated Erk phosphorylation and decreased the expression levels of PPARγ and CEBPα, which was inhibited by treatment with the Shp2 PTPase activity inhibitor, NSC‑87877. The results of the present study suggested that CGA‑induced Akt and Erk pathways regulate proliferation and differentiation and that Shp2 is important in the proliferation and differentiation of BMSCs. PMID:25634525

  1. Intravenous Administration of Bone Marrow-Derived Mesenchymal Stem Cells Induces a Switch from Classical to Atypical Symptoms in Experimental Autoimmune Encephalomyelitis

    PubMed Central

    Kurte, Mónica; Bravo-Alegría, Javiera; Torres, Alexander; Carrasco, Vania; Ibáñez, Cristina; Vega-Letter, Ana María; Fernández-O'Ryan, Catalina; Irarrázabal, Carlos E.; Figueroa, Fernando E.; Fuentealba, Rodrigo A.; Riedel, Claudia; Carrión, Flavio

    2015-01-01

    Potent immunosuppressive and regenerative properties of mesenchymal stem cells (MSCs) position them as a novel therapy for autoimmune diseases. This research examines the therapeutic effect of MSCs administration at different disease stages in experimental autoimmune encephalomyelitis (EAE). Classical and atypical scores of EAE, associated with Th1 and Th17 response, respectively, and also Treg lymphocytes, were evaluated. MSCs administration at the onset (EAE+MSConset) induced an important amelioration of the clinical signs and less lasting effect at the peak of EAE (EAE+MSCpeak). No effect was observed when MSCs were applied after EAE stabilization (EAE+MSClate). Surprisingly, EAE atypical signs were detected in EAE+MSCpeak and EAE+MSClate mice. However, no correlation was found in Th17/Th1 ratio. Interestingly, regardless of time administration, MSCs significantly reduced IL-6 and also T-bet, RORγT, and Foxp3 mRNA levels in brain samples of EAE mice. The downregulation of IL-6 could restore the well-functioning of the blood-brain barrier of EAE mice, correlated with a decreased number of brain infiltrating leukocytes. These results suggest that the inflammatory status is important to be considered for administering MSCs in autoimmune pathologies, leading to a further research to clarify the effect of MSCs for multiple sclerosis. PMID:25838828

  2. Bone marrow-derived mesenchymal stem cells improve diabetes-induced cognitive impairment by exosome transfer into damaged neurons and astrocytes

    PubMed Central

    Nakano, Masako; Nagaishi, Kanna; Konari, Naoto; Saito, Yuki; Chikenji, Takako; Mizue, Yuka; Fujimiya, Mineko

    2016-01-01

    The incidence of dementia is higher in diabetic patients, but no effective treatment has been developed. This study showed that rat bone marrow mesenchymal stem cells (BM-MSCs) can improve the cognitive impairments of STZ-diabetic mice by repairing damaged neurons and astrocytes. The Morris water maze test demonstrated that cognitive impairments induced by diabetes were significantly improved by intravenous injection of BM-MSCs. In the CA1 region of the hippocampus, degeneration of neurons and astrocytes, as well as synaptic loss, were prominent in diabetes, and BM-MSC treatment successfully normalized them. Since a limited number of donor BM-MSCs was observed in the brain parenchyma, we hypothesized that humoral factors, especially exosomes released from BM-MSCs, act on damaged neurons and astrocytes. To investigate the effectiveness of exosomes for treatment of diabetes-induced cognitive impairment, exosomes were purified from the culture media and injected intracerebroventricularly into diabetic mice. Recovery of cognitive impairment and histological abnormalities similar to that seen with BM-MSC injection was found following exosome treatment. Use of fluorescence-labeled exosomes demonstrated that injected exosomes were internalized into astrocytes and neurons; these subsequently reversed the dysfunction. The present results indicate that exosomes derived from BM-MSCs might be a promising therapeutic tool for diabetes-induced cognitive impairment. PMID:27102354

  3. Bone marrow-derived mesenchymal stem cells improve diabetes-induced cognitive impairment by exosome transfer into damaged neurons and astrocytes.

    PubMed

    Nakano, Masako; Nagaishi, Kanna; Konari, Naoto; Saito, Yuki; Chikenji, Takako; Mizue, Yuka; Fujimiya, Mineko

    2016-01-01

    The incidence of dementia is higher in diabetic patients, but no effective treatment has been developed. This study showed that rat bone marrow mesenchymal stem cells (BM-MSCs) can improve the cognitive impairments of STZ-diabetic mice by repairing damaged neurons and astrocytes. The Morris water maze test demonstrated that cognitive impairments induced by diabetes were significantly improved by intravenous injection of BM-MSCs. In the CA1 region of the hippocampus, degeneration of neurons and astrocytes, as well as synaptic loss, were prominent in diabetes, and BM-MSC treatment successfully normalized them. Since a limited number of donor BM-MSCs was observed in the brain parenchyma, we hypothesized that humoral factors, especially exosomes released from BM-MSCs, act on damaged neurons and astrocytes. To investigate the effectiveness of exosomes for treatment of diabetes-induced cognitive impairment, exosomes were purified from the culture media and injected intracerebroventricularly into diabetic mice. Recovery of cognitive impairment and histological abnormalities similar to that seen with BM-MSC injection was found following exosome treatment. Use of fluorescence-labeled exosomes demonstrated that injected exosomes were internalized into astrocytes and neurons; these subsequently reversed the dysfunction. The present results indicate that exosomes derived from BM-MSCs might be a promising therapeutic tool for diabetes-induced cognitive impairment. PMID:27102354

  4. In vitro osteogenic induction of human marrow-derived mesenchymal stem cells by PCL fibrous scaffolds containing dexamethazone-loaded chitosan microspheres.

    PubMed

    Omidvar, Noushin; Ganji, Fariba; Eslaminejad, Mohamadreza Baghaban

    2016-07-01

    This research reports the encapsulation of dexamethasone (Dex) within the chitosan microspheres (CSMs) embedded in a fibrous structure of poly(ɛ-caprolactone) (PCL) to provide a platform for osteogenic differentiation of human mesenchymal stem cells (hMSCs). Dex loaded CSMs were prepared by spray drying a mixture of chitosan and Dex. Then, they were electrospun with PCL solution to create a bilayer fibrous scaffold (PCL/CSMs-Dex). The CSMs act as good depots for sustained release of Dex over a period of 14 days, without noticeable burst release. This is mainly attributed to the core-shell structure of the final PCL/CSMs-Dex-matrix, which could prolong the release and eliminate the initial burst. The water contact angle of PCL scaffolds decreased from 141.4 ± 3.8 to 118.4 ± 7.6 in the presence of CSMs. Improved proliferation of hMSCs cultured on PCL/CSMs-Dex scaffolds was also evidenced. Furthermore, osteogenic assays showed an increase in alkaline phosphatase activity and mineral deposits. The expression of bone-specific genes also confirmed the osteogenic differentiation of cells cultured on these Dex-loaded core-shell structures. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 104A: 1657-1667, 2016. PMID:26916786

  5. A novel bispecific immunotoxin delivered by human bone marrow-derived mesenchymal stem cells to target blood vessels and vasculogenic mimicry of malignant gliomas

    PubMed Central

    Zhang, Yonghong; Sun, Xinlin; Huang, Min; Ke, Yiquan; Wang, Jihui; Liu, Xiao

    2015-01-01

    Background In previous years, immunotoxins have been shown to be a greatly promising therapeutic tool for brain malignancies, such as gliomas. Human mesenchymal stem cells (hMSCs) exhibit tropism to tumor tissue. However, the effect of bispecific immunotoxins in malignant gliomas is still unknown. The aim of this study was to investigate the function of bispecific immunotoxins in human malignant gliomas. Materials and methods In the present study, the bispecific immunotoxin VEGF165-ephrin A1-PE38KDEL was established using deoxyribonucleic acid shuffling and cloning techniques. The VEGF165-ephrin A1-PE38KDEL was delivered by hMSCs to mouse malignant gliomas. The effects of the bispecific immunotoxins on glioma-derived blood vessels and vasculogenic mimicry to elucidate the molecular mechanisms underlying the antitumorigenic effects of immunotoxins were examined in vivo. Results In vitro, transfected hMSCs significantly inhibited the cell viability of gliomas cell lines U87 and U251 in a dose-dependent manner compared with untransfected hMSCs (P<0.01). In vivo, the intratumoral injection of engineered hMSCs was effective at inhibiting tumor growth in a malignant glioma tumor model. Conclusion The bispecific immunotoxin secreted from hMSCs acts as a novel strategy for improving treatment options for malignant gliomas in the clinic. PMID:26089644

  6. Autophagy-Modulated Human Bone Marrow-Derived Mesenchymal Stem Cells Accelerate Liver Restoration in Mouse Models of Acute Liver Failure

    PubMed Central

    Amiri, Fatemeh; Molaei, Sedigheh; Bahadori, Marzie; Nasiri, Fatemeh; Deyhim, Mohammad Reza; Jalili, Mohammad Ali; Nourani, Mohammad Reza; Habibi Roudkenar, Mehryar

    2016-01-01

    Background: Mesenchymal stem cells (MSCs) have been recently received increasing attention for cell-based therapy, especially in regenerative medicine. However, the low survival rate of these cells restricts their therapeutic applications. It is hypothesized that autophagy might play an important role in cellular homeostasis and survival. This study aims to investigate the regenerative potentials of autophagy-modulated MSCs for the treatment of acute liver failure (ALF) in mice. Methods: ALF was induced in mice by intraperitoneal injection of 1.5 ml/kg carbon tetrachloride. Mice were intravenously infused with MSCs, which were suppressed in their autophagy pathway. Blood and liver samples were collected at different intervals (24, 48 and 72 h) after the transplantation of MSCs. Both the liver enzymes and tissue necrosis levels were evaluated using biochemical and histopathological assessments. The survival rate of the transplanted mice was also recorded during one week. Results: Biochemical and pathological results indicated that 1.5 ml/kg carbon tetrachloride induces ALF in mice. A significant reduction of liver enzymes and necrosis score were observed in autophagy-modulated MSC-transplanted mice compared to sham (with no cell therapy) after 24 h. After 72 h, liver enzymes reached their normal levels in mice transplanted with autophagy-suppressed MSCs. Interestingly, normal histology without necrosis was also observed. Conclusion: Autophagy suppression in MSCs ameliorates their liver regeneration potentials due to paracrine effects and might be suggested as a new strategy for the improvement of cell therapy in ALF. PMID:26899739

  7. Oleate Abrogates Palmitate-Induced Lipotoxicity and Proinflammatory Response in Human Bone Marrow-Derived Mesenchymal Stem Cells and Osteoblastic Cells.

    PubMed

    Gillet, C; Spruyt, D; Rigutto, S; Dalla Valle, A; Berlier, J; Louis, C; Debier, C; Gaspard, N; Malaisse, W J; Gangji, V; Rasschaert, J

    2015-11-01

    Osteoporosis is a metabolic bone disease associated with unequilibrated bone remodeling resulting from decreased bone formation and/or increased bone resorption, leading to progressive bone loss. In osteoporotic patients, low bone mass is associated with an increase of bone marrow fat resulting from accumulation of adipocytes within the bone marrow. Marrow adipocytes are active secretory cells, releasing cytokines, adipokines and free fatty acids (FA) that influence the bone marrow microenvironment and alter the biology of neighboring cells. Therefore, we examined the effect of palmitate (Palm) and oleate (Ole), 2 highly prevalent FA in human organism and diet, on the function and survival of human mesenchymal stem cells (MSC) and MSC-derived osteoblastic cells. The saturated FA Palm exerted a cytotoxic action via initiation of endoplasmic reticulum stress and activation of the nuclear factor κB (NF-κB) and ERK pathways. In addition, Palm induced a proinflammatory response, as determined by the up-regulation of Toll-like receptor 4 expression as well as the increase of IL-6 and IL-8 expression and secretion. Moreover, we showed that MSC-derived osteoblastic cells were more sensitive to lipotoxicity than undifferentiated MSC. The monounsaturated FA Ole fully neutralized Palm-induced lipotoxicity by impairing activation of the pathways triggered by the saturated FA. Moreover, Ole promoted Palm detoxification by fostering its esterification into triglycerides and storage in lipid droplets. Altogether, our data showed that physiological concentrations of Palm and Ole differently modulated cell death and function in bone cells. We therefore propose that FA could influence skeletal health. PMID:26327577

  8. Bone Marrow-Derived Mesenchymal Stem Cells Have Innate Procoagulant Activity and Cause Microvascular Obstruction Following Intracoronary Delivery: Amelioration by Antithrombin Therapy.

    PubMed

    Gleeson, Birgitta M; Martin, Kenneth; Ali, Mohammed T; Kumar, Arun H S; Pillai, M Gopala-Krishnan; Kumar, Sujith P G; O'Sullivan, John F; Whelan, Derek; Stocca, Alessia; Khider, Wisam; Barry, Frank P; O'Brien, Timothy; Caplice, Noel M

    2015-09-01

    Mesenchymal stem cells (MSCs) are currently under investigation as tools to preserve cardiac structure and function following acute myocardial infarction (AMI). However, concerns have emerged regarding safety of acute intracoronary (IC) MSC delivery. This study aimed to characterize innate prothrombotic activity of MSC and identify means of its mitigation toward safe and efficacious therapeutic IC MSC delivery post-AMI. Expression of the initiator of the coagulation cascade tissue factor (TF) on MSC was detected and quantified by immunofluorescence, FACS, and immunoblotting. MSC-derived TF antigen was catalytically active and capable of supporting thrombin generation in vitro. Addition of MSCs to whole citrated blood enhanced platelet thrombus deposition on collagen at arterial shear, an effect abolished by heparin coadministration. In a porcine AMI model, IC infusion of 25 × 10(6) MSC during reperfusion was associated with a decrease in coronary flow reserve but not when coadministered with an antithrombin agent (heparin). Heparin reduced MSC-associated thrombosis incorporating platelets and VWF within the microvasculature. Heparin-assisted therapeutic MSC delivery also reduced apoptosis in the infarct border zone at 24 hours, significantly improved infarct size, left ventricular (LV) ejection fraction, LV volumes, wall motion, and attenuated histologic evidence of scar formation at 6 weeks post-AMI. Heparin alone or heparin-assisted fibroblast control cell delivery had no such effect. Procoagulant TF activity of therapeutic MSCs is associated with reductions in myocardial perfusion when delivered IC may be successfully managed by heparin coadministration. This study highlights an important mechanistic insight into safety concerns associated with therapeutic IC MSC delivery for AMI. PMID:25969127

  9. Human bone marrow derived mesenchymal stem cells do not undergo transformation after long-term in vitro culture and do not exhibit telomere maintenance mechanisms.

    PubMed

    Bernardo, Maria Ester; Zaffaroni, Nadia; Novara, Francesca; Cometa, Angela Maria; Avanzini, Maria Antonietta; Moretta, Antonia; Montagna, Daniela; Maccario, Rita; Villa, Raffaella; Daidone, Maria Grazia; Zuffardi, Orsetta; Locatelli, Franco

    2007-10-01

    Significant improvement in the understanding of mesenchymal stem cell (MSC) biology has opened the way to their clinical use. However, concerns regarding the possibility that MSCs undergo malignant transformation have been raised. We investigated the susceptibility to transformation of human bone marrow (BM)-derived MSCs at different in vitro culture time points. MSCs were isolated from BM of 10 healthy donors and propagated in vitro until reaching either senescence or passage (P) 25. MSCs in the senescence phase were closely monitored for 8 to 12 weeks before interrupting the cultures. The genetic characterization of MSCs was investigated through array-comparative genomic hybridization (array-CGH), conventional karyotyping, and subtelomeric fluorescent in situ hybridization analysis both before and after prolonged culture. MSCs were tested for the expression of telomerase activity, human telomerase reverse transcriptase (hTERT) transcripts, and alternative lengthening of telomere (ALT) mechanism at different passages. A huge variability in terms of proliferative capacity and MSCs life span was noted between donors. In eight of 10 donors, MSCs displayed a progressive decrease in proliferative capacity until reaching senescence. In the remaining two MSC samples, the cultures were interrupted at P25 to pursue data analysis. Array-CGH and cytogenetic analyses showed that MSCs expanded in vitro did not show chromosomal abnormalities. Telomerase activity and hTERT transcripts were not expressed in any of the examined cultures and telomeres shortened during the culture period. ALT was not evidenced in the MSCs tested. BM-derived MSCs can be safely expanded in vitro and are not susceptible to malignant transformation, thus rendering these cells suitable for cell therapy approaches. PMID:17909019

  10. Demonstration of the presence of independent pre-osteoblastic and pre-adipocytic cell populations in bone marrow-derived mesenchymal stem cells.

    PubMed

    Post, S; Abdallah, B M; Bentzon, J F; Kassem, M

    2008-07-01

    Mesenchymal stem cells (MSC) are defined as plastic-adherent, clonal cells that are common progenitors for osteoblasts and adipocytes. An inverse relationship between bone and fat has been observed in several clinical conditions and has been suggested to be caused by re-directing MSC differentiation into one particular lineage. However, this inverse relationship between bone and fat is not consistent and under certain in vivo conditions, bone and fat can change independently suggesting separate precursor cell populations. In order to test for this hypothesis, we extensively characterized two plastic-adherent clonal MSC lines (mMSC1 and mMSC2) derived from murine bone marrow. The two cell lines grew readily in culture and have undergone more than 100 population doublings with no apparent differences in their growth rates. Both cell lines were positive for the murine MSC marker Sca-1 and mMSC1 was also positive for CD13. Both cell lines were exposed to in vitro culture induction of osteogenesis and adipogenesis. mMSC1 and not mMSC2 were only able to differentiate to adipocytes evidenced by the expression of adipocyte markers (aP2, adiponectin, adipsin, PPARgamma2 and C/EBPa) and the presence of mature adipocytes visualized by Oil Red O staining. On the other hand, mMSC2 and not mMSC1 differentiated to osteoblast lineage as demonstrated by up-regulation of osteoblastic makers (CBFA1/RUNX2, Osterix, alkaline phosphatase, bone sialoprotein and osteopontin) and formation of alizarin red stained mineralized matrix in vitro. Consistent with the in vitro results, mMSC2 and not mMSC1, were able to form bone in vivo after subcutaneous implantation in immune-deficient (NOD/SCID) mice. Our data suggest that contrary to the current belief, bone marrow contains clonal subpopulations of cells that are committed to either osteoblast or adipocyte lineage. These cell populations may undergo independent changes during aging and in bone diseases and thus represent important targets for

  11. Effects of the dichloromethane fraction of Dipsaci Radix on the osteoblastic differentiation of human alveolar bone marrow-derived mesenchymal stem cells.

    PubMed

    Kim, Beom-Su; Kim, Yoon-Chul; Zadeh, Homa; Park, Yoon-Jeong; Pi, Sung-Hee; Shin, Hyung-Shik; You, Hyung-Keun

    2011-01-01

    Dipsaci Radix is the dried root of Dipsacus asper Wall. It has been used in Korean herbal medicine to treat bone fractures. In this study, we examined the effect of the dichloromethane fraction of Dipsaci Radix (DR(DM)) on the osteoblastic differentiation of human alveolar bone marrow-derived MSCs (ABM-MSCs). The ABM-MSCs were isolated from healthy subjects and cultured in vitro, followed by phenotypic characterization. They showed a fibroblast-like morphology and expressed CD29, CD44, CD73, and CD105, but not CD34. Calcified nodules were generated in response to both dexamethasone (DEX) and DR(DM). There was a significant increase in the alkaline phosphatase (ALP) activity and protein expression of bone sialoprotein (BSP) and osteocalcin (OC) in response to DEX and DR(DM) as compared to control. These results provide evidence for the osteogenic potential of cultured ABM-MSCs in response to DR(DM). Also, an active single compound was additionally isolated from DR(DM). The single compound (hederagenin 3-O-(2-O-acetyl)-α-L-arabinopyranoside) also significantly increased ALP activity and the level of protein expression of BSP and OC. These results highlight the possible clinical applications of DR(DM) and hederagenin 3-O-(2-O-acetyl)-α-L-arabinopyranoside in bone regeneration. PMID:21228489

  12. Activin Receptor-Like Kinase Receptors ALK5 and ALK1 Are Both Required for TGFβ-Induced Chondrogenic Differentiation of Human Bone Marrow-Derived Mesenchymal Stem Cells

    PubMed Central

    de Kroon, Laurie M. G.; Narcisi, Roberto; Blaney Davidson, Esmeralda N.; Cleary, Mairéad A.; van Beuningen, Henk M.; Koevoet, Wendy J. L. M.; van Osch, Gerjo J. V. M.; van der Kraan, Peter M.

    2015-01-01

    Introduction Bone marrow-derived mesenchymal stem cells (BMSCs) are promising for cartilage regeneration because BMSCs can differentiate into cartilage tissue-producing chondrocytes. Transforming Growth Factor β (TGFβ) is crucial for inducing chondrogenic differentiation of BMSCs and is known to signal via Activin receptor-Like Kinase (ALK) receptors ALK5 and ALK1. Since the specific role of these two TGFβ receptors in chondrogenesis is unknown, we investigated whether ALK5 and ALK1 are expressed in BMSCs and whether both receptors are required for chondrogenic differentiation of BMSCs. Materials & Methods ALK5 and ALK1 gene expression in human BMSCs was determined with RT-qPCR. To induce chondrogenesis, human BMSCs were pellet-cultured in serum-free chondrogenic medium containing TGFβ1. Chondrogenesis was evaluated by aggrecan and collagen type IIα1 RT-qPCR analysis, and histological stainings of proteoglycans and collagen type II. To overexpress constitutively active (ca) receptors, BMSCs were transduced either with caALK5 or caALK1. Expression of ALK5 and ALK1 was downregulated by transducing BMSCs with shRNA against ALK5 or ALK1. Results ALK5 and ALK1 were expressed in in vitro-expanded as well as in pellet-cultured BMSCs from five donors, but mRNA levels of both TGFβ receptors did not clearly associate with chondrogenic induction. TGFβ increased ALK5 and decreased ALK1 gene expression in chondrogenically differentiating BMSC pellets. Neither caALK5 nor caALK1 overexpression induced cartilage matrix formation as efficient as that induced by TGFβ. Moreover, short hairpin-mediated downregulation of either ALK5 or ALK1 resulted in a strong inhibition of TGFβ-induced chondrogenesis. Conclusion ALK5 as well as ALK1 are required for TGFβ-induced chondrogenic differentiation of BMSCs, and TGFβ not only directly induces chondrogenesis, but also modulates ALK5 and ALK1 receptor signaling in BMSCs. These results imply that optimizing cartilage formation by

  13. Lithium chloride attenuates the abnormal osteogenic/adipogenic differentiation of bone marrow-derived mesenchymal stem cells obtained from rats with steroid-related osteonecrosis by activating the β-catenin pathway

    PubMed Central

    YU, ZEFENG; FAN, LIHONG; LI, JIA; GE, ZHAOGANG; DANG, XIAOQIAN; WANG, KUNZHENG

    2015-01-01

    Steroid-related osteonecrosis of the femoral head (ONFH) may be a disease that results from the abnormal osteogenic/adipogenic differentiation of bone marrow-derived mesenchymal stem cells (BMMSCs). In the present study, we examined the possible use of lithium in an aim to reverse the abnormal osteogenic/adipogenic differentiation of BMMSCs isolated from rats with steroid-related ONFH (termed ONFH-BMMSCs). BMMSCs obtained from steroid-related ONFH rat femurs were cultured with or without lithium chloride (LiCl). BMMSCs obtained from normal rat femurs were cultured as controls. LiCl significantly increased the expression of osteocalcin and Runx2 in the ONFH-BMMSCs during osteogenic induction. The mineralization of ONFH-BMMSCs following osteogenic induction was also enhanced. Furthermore, LiCl exerted anti-adipogenic effects on the ONFH-BMMSCs by inhibiting the expression of peroxisome proliferator-activated receptor γ (PPARγ) and fatty acid binding protein 4 (Fabp4) during adipogenic induction, and decreasing lipid droplet formation at the end of adipogenic induction. These effects of LiCl on the ONFH-BMMSCs were associated with an increased expression of β-catenin and a decreased expression of phosphorylated GSK-3β at Tyr-216, and these effects were abolished by treatment with quercetin, an antagonist of the β-catenin pathway. The normal osteogenic/adipogenic activity of BMMSCs may be impaired in steroid-related ONFH. However, as demonstrated by our findings, LiCl reduces abnormal adipogenic activity and simultaneously increases the osteogenic differentiation of ONFH-BMMSCs by activating the β-catenin pathway. PMID:26352537

  14. Lithium chloride attenuates the abnormal osteogenic/adipogenic differentiation of bone marrow-derived mesenchymal stem cells obtained from rats with steroid-related osteonecrosis by activating the β-catenin pathway.

    PubMed

    Yu, Zefeng; Fan, Lihong; Li, Jia; Ge, Zhaogang; Dang, Xiaoqian; Wang, Kunzheng

    2015-11-01

    Steroid-related osteonecrosis of the femoral head (ONFH) may be a disease that results from the abnormal osteogenic/adipogenic differentiation of bone marrow-derived mesenchymal stem cells (BMMSCs). In the present study, we examined the possible use of lithium in an aim to reverse the abnormal osteogenic/adipogenic differentiation of BMMSCs isolated from rats with steroid-related ONFH (termed ONFH-BMMSCs). BMMSCs obtained from steroid‑related ONFH rat femurs were cultured with or without lithium chloride (LiCl). BMMSCs obtained from normal rat femurs were cultured as controls. LiCl significantly increased the expression of osteocalcin and Runx2 in the ONFH-BMMSCs during osteogenic induction. The mineralization of ONFH-BMMSCs following osteogenic induction was also enhanced. Furthermore, LiCl exerted anti-adipogenic effects on the ONFH-BMMSCs by inhibiting the expression of peroxisome proliferator-activated receptor γ (PPARγ) and fatty acid binding protein 4 (Fabp4) during adipogenic induction, and decreasing lipid droplet formation at the end of adipogenic induction. These effects of LiCl on the ONFH-BMMSCs were associated with an increased expression of β-catenin and a decreased expression of phosphorylated GSK-3β at Tyr-216, and these effects were abolished by treatment with quercetin, an antagonist of the β-catenin pathway. The normal osteogenic/adipogenic activity of BMMSCs may be impaired in steroid-related ONFH. However, as demonstrated by our findings, LiCl reduces abnormal adipogenic activity and simultaneously increases the osteogenic differentiation of ONFH-BMMSCs by activating the β-catenin pathway. PMID:26352537

  15. Optimal Seeding Densities for In Vitro Chondrogenesis of Two- and Three-Dimensional-Isolated and -Expanded Bone Marrow-Derived Mesenchymal Stromal Stem Cells Within a Porous Collagen Scaffold.

    PubMed

    Bornes, Troy D; Jomha, Nadr M; Mulet-Sierra, Aillette; Adesida, Adetola B

    2016-03-01

    Bone marrow-derived mesenchymal stromal stem cells (BMSCs) are a promising cell source for treating articular cartilage defects. The objective of this study was to assess the impact of cell seeding density within a collagen I scaffold on in vitro BMSC chondrogenesis following isolation and expansion in two-dimensional (2D) and three-dimensional (3D) environments. It was hypothesized that both expansion protocols would produce BMSCs capable of hyaline-like chondrogenesis with an optimal seeding density of 10 × 10(6) cells/cm(3). Ovine BMSCs were isolated in a 2D environment by plastic adherence, expanded to passage two in flasks containing an expansion medium, and seeded within collagen I scaffolds at densities of 50, 10, 5, 1, and 0.5 × 10(6) BMSCs/cm(3). For 3D isolation and expansion, aspirates containing known quantities of mononucleated cells (bone marrow-derived mononucleated cells [BMNCs]) were seeded on scaffolds at 50, 10, 5, 1, and 0.5 × 10(6) BMNCs/cm(3) and cultured in the expansion medium for an equivalent duration to 2D expansion. Constructs were differentiated in vitro in the chondrogenic medium for 21 days and assessed with reverse-transcription quantitative polymerase chain reaction, safranin O staining, histological scoring using the Bern Score, collagen immunofluorescence, and glycosaminoglycan (GAG) quantification. Two-dimensional-expanded BMSCs seeded at all densities were capable of proteoglycan production and displayed increased expressions of aggrecan and collagen II messenger RNA (mRNA) relative to predifferentiation controls. Collagen II deposition was apparent in scaffolds seeded at 0.5-10 × 10(6) BMSCs/cm(3). Chondrogenesis of 2D-expanded BMSCs was most pronounced in scaffolds seeded at 5-10 × 10(6) BMSCs/cm(3) based on aggrecan and collagen II mRNA, safranin O staining, Bern Score, total GAG, and GAG/deoxyribonucleic acid (DNA). For 3D-expanded BMSC-seeded scaffolds, increased aggrecan and collagen II m

  16. Optimal Seeding Densities for In Vitro Chondrogenesis of Two- and Three-Dimensional-Isolated and -Expanded Bone Marrow-Derived Mesenchymal Stromal Stem Cells Within a Porous Collagen Scaffold

    PubMed Central

    Bornes, Troy D.; Jomha, Nadr M.; Mulet-Sierra, Aillette; Adesida, Adetola B.

    2016-01-01

    Bone marrow-derived mesenchymal stromal stem cells (BMSCs) are a promising cell source for treating articular cartilage defects. The objective of this study was to assess the impact of cell seeding density within a collagen I scaffold on in vitro BMSC chondrogenesis following isolation and expansion in two-dimensional (2D) and three-dimensional (3D) environments. It was hypothesized that both expansion protocols would produce BMSCs capable of hyaline-like chondrogenesis with an optimal seeding density of 10 × 106 cells/cm3. Ovine BMSCs were isolated in a 2D environment by plastic adherence, expanded to passage two in flasks containing an expansion medium, and seeded within collagen I scaffolds at densities of 50, 10, 5, 1, and 0.5 × 106 BMSCs/cm3. For 3D isolation and expansion, aspirates containing known quantities of mononucleated cells (bone marrow-derived mononucleated cells [BMNCs]) were seeded on scaffolds at 50, 10, 5, 1, and 0.5 × 106 BMNCs/cm3 and cultured in the expansion medium for an equivalent duration to 2D expansion. Constructs were differentiated in vitro in the chondrogenic medium for 21 days and assessed with reverse-transcription quantitative polymerase chain reaction, safranin O staining, histological scoring using the Bern Score, collagen immunofluorescence, and glycosaminoglycan (GAG) quantification. Two-dimensional-expanded BMSCs seeded at all densities were capable of proteoglycan production and displayed increased expressions of aggrecan and collagen II messenger RNA (mRNA) relative to pre-differentiation controls. Collagen II deposition was apparent in scaffolds seeded at 0.5–10 × 106 BMSCs/cm3. Chondrogenesis of 2D-expanded BMSCs was most pronounced in scaffolds seeded at 5–10 × 106 BMSCs/cm3 based on aggrecan and collagen II mRNA, safranin O staining, Bern Score, total GAG, and GAG/deoxyribonucleic acid (DNA). For 3D-expanded BMSC-seeded scaffolds, increased aggrecan and collagen II mRNA expressions relative to controls were

  17. The fate of transplanted xenogeneic bone marrow-derived stem cells in rat intervertebral discs.

    PubMed

    Wei, Aiqun; Tao, Helen; Chung, Sylvia A; Brisby, Helena; Ma, David D; Diwan, Ashish D

    2009-03-01

    Intervertebral disc degeneration is a major cause and a risk factor for chronic low back pain. The potential of using stem cells to treat disc degeneration has been raised. The aims of our study were to assess whether xenogeneic bone-marrow derived stem cells could survive in a rat disc degeneration model and to determine which cell types, if any, survived and differentiated into disc-like cells. Human bone-marrow derived CD34(+) (hematopoietic progenitor cells) and CD34(-) (nonhematopoietic progenitor cells, including mesenchymal stem cells) cells were isolated, fluorescent-labeled, and injected into rat coccygeal discs. The rats were sacrificed at day 1, 10, 21, and 42. Treated discs were examined by histological and immunostaining techniques and compared to control discs. The survival of transplanted cells was further confirmed with a human nuclear specific marker. Fluorescent labeled CD34(-) cells were detected until day 42 in the nucleus pulposus of the injected discs. After 3 weeks these cells had differentiated into cells expressing chondrocytic phenotype (Collagen II and Sox-9). In contrast, the fluorescent labeled CD34(+) cells could not be detected after day 21. No fluorescence-positive cells were detected in the noninjected control discs. Further, no inflammatory cells infiltrated the nucleus pulposus, even though these animals had not received immunosuppressive treatment. Our data provide evidence that transplanted human BM CD34(-) cells survived and differentiated within the relative immune privileged nucleus pulposus of intervertebral disc degeneration. PMID:18853431

  18. Bone marrow-derived stem cells and radiation response.

    PubMed

    Greenberger, Joel S; Epperly, Michael

    2009-04-01

    The recovery of tissues and organs from ionizing irradiation is critically dependent on the repopulation of resident stem cells, defined as the subset of cells with capacity for both self-renewal and differentiation. Stem cells of both hematopoietic and epithelial origin reside in defined areas of the cellular microenvironment (recently defined as the stem cell "niche"). Experiments using serial repopulation assays in serial generations of total body irradiated mice receiving transplanted marrow and in continuous bone marrow cultures both identified specific microanatomic sites that comprise the bone marrow stem cell niche. Supportive cells of the hematopoietic microenvironment not only contribute to stem cell repopulation capacity but also to the maintenance of their quiescent or nonproliferative state, which allows the most primitive hematopoietic stem cells to stay in a noncycling state protected from both direct ionizing radiation-induced cell-cycle phase-specific killing and indirect cytokine and free radical mediated killing. Recent evidence has defined both cell contact and humoral mechanisms of protection of hematopoietic stem cells by stromal cells. There is also recent evidence for multilineage differentiation capacity of cells of the hematopoietic microenvironment termed bone marrow stromal cells (mesenchymal stem cells). Both hematopoietic stem cells and mesenchymal stem cell populations have been shown to be involved in the repair of ionizing irradiation damage of distant epithelial as well as other hematopoietic sites through their capacity to migrate through the circulation. The radiobiology of these 2 bone marrow stem cell populations is the subject of intense investigation. This review defines the status of research in the areas of stem cell quiescence, niche contact, and migratory responses to ionizing irradiation. PMID:19249651

  19. Human urokinase-type plasminogen activator gene-modified bone marrow-derived mesenchymal stem cells attenuate liver fibrosis in rats by down-regulating the Wnt signaling pathway

    PubMed Central

    Ma, Zhi-Gang; Lv, Xiao-Dan; Zhan, Ling-Ling; Chen, Lan; Zou, Qi-Yuan; Xiang, Ji-Qiao; Qin, Jiao-Li; Zhang, Wei-Wei; Zeng, Zhao-Jing; Jin, Hui; Jiang, Hai-Xing; Lv, Xiao-Ping

    2016-01-01

    AIM: To evaluate the therapeutic effects of bone marrow-derived mesenchymal stem cells (BMSCs) with human urokinase-type plasminogen activator (uPA) on liver fibrosis, and to investigate the mechanism of gene therapy. METHODS: BMSCs transfected with adenovirus-mediated human urokinase plasminogen activator (Ad-uPA) were transplanted into rats with CCl4-induced liver fibrosis. All rats were sacrificed after 8 wk, and their serum and liver tissue were collected for biochemical, histopathologic, and molecular analyzes. The degree of liver fibrosis was assessed by hematoxylin and eosin or Masson’s staining. Western blot and quantitative reverse transcription-polymerase chain reaction were used to determine protein and mRNA expression levels. RESULTS: Serum levels of alanine aminotransferase, aminotransferase, total bilirubin, hyaluronic acid, laminin, and procollagen type III were markedly decreased, whereas the levels of serum albumin were increased by uPA gene modified BMSCs treatment. Histopathology revealed that chronic CCl4-treatment resulted in significant fibrosis while uPA gene modified BMSCs treatment significantly reversed fibrosis. By quantitatively analysing the fibrosis area of liver tissue using Masson staining in different groups of animals, we found that model animals with CCl4-induced liver fibrosis had the largest fibrotic area (16.69% ± 1.30%), while fibrotic area was significantly decreased by BMSCs treatment (12.38% ± 2.27%) and was further reduced by uPA-BMSCs treatment (8.31% ± 1.21%). Both protein and mRNA expression of β-catenin, Wnt4 and Wnt5a was down-regulated in liver tissues following uPA gene modified BMSCs treatment when compared with the model animals. CONCLUSION: Transplantation of uPA gene modified BMSCs suppressed liver fibrosis and ameliorated liver function and may be a new approach to treating liver fibrosis. Furthermore, treatment with uPA gene modified BMSCs also resulted in a decrease in expression of molecules of the Wnt

  20. Bone marrow-derived stem cells and respiratory disease.

    PubMed

    Jones, Carla P; Rankin, Sara M

    2011-07-01

    Adult bone marrow contains a number of discrete populations of progenitor cells, including endothelial, mesenchymal, and epithelial progenitor cells and fibrocytes. In the context of a range of diseases, endothelial progenitor cells have been reported to promote angiogenesis, mesenchymal stem cells are potent immunosuppressors but can also contribute directly to tissue regeneration, and fibrocytes have been shown to induce tissue fibrosis. This article provides an overview of the basic biology of these different subsets of progenitor cells, reporting their distinct phenotypes and functional activities. The differences in their secretomes are highlighted, and the relative role of cellular differentiation vs paracrine effects of progenitor cells is considered. The article reviews the literature examining the contribution of progenitor cells to the pathogenesis of respiratory disease, and discusses recent studies using bone marrow progenitor cells as stem cell therapies in the context of pulmonary hypertension, COPD, and asthma. PMID:21729891

  1. Intravitreal Implantation of Genetically Modified Autologous Bone Marrow-Derived Stem Cells for Treating Retinal Disorders.

    PubMed

    Tracy, Christopher J; Sanders, Douglas N; Bryan, Jeffrey N; Jensen, Cheryl A; Castaner, Leilani J; Kirk, Mark D; Katz, Martin L

    2016-01-01

    A number of retinal degenerative diseases may be amenable to treatment with continuous intraocular delivery of therapeutic agents that cannot be delivered effectively to the retina via systemic or topical administration. Among these disorders are lysosomal storage diseases resulting from deficiencies in soluble lysosomal enzymes. Most cells, including those of the retina, are able to take up these enzymes and incorporate them in active form into their lysosomes. In theory, therefore, continuous intraocular administration of a normal form of a soluble lysosomal enzyme should be able to cure the molecular defect in the retinas of subjects lacking this enzyme. Experiments were conducted to determine whether genetically modified bone marrow-derived stem cells implanted into the vitreous could be used as -vehicles for continuous delivery of such enzymes to the retina. Bone marrow-derived mesenchymal stem cells (MSCs) from normal mice were implanted into the vitreous of mice undergoing retinal degeneration as a result of a mutation in the PPT1 gene. The implanted cells appeared to survive indefinitely in the vitreous without proliferating or invading the retina. This indicates that intravitreal implantation of MSCs is likely a safe means of long-term delivery of proteins synthesized by the implanted cells. Experiments have been initiated to test the efficacy of using genetically modified autologous MSCs to inhibit retinal degeneration in a canine model of neuronal ceroid lipofuscinosis. PMID:26427461

  2. Intravitreal Implantation of Genetically Modified Autologous Bone Marrow-Derived Stem Cells for Treating Retinal Disorders

    PubMed Central

    Tracy, Christopher J.; Sanders, Douglas N.; Bryan, Jeffrey N.; Jensen, Cheryl A.; Castaner, Leilani J.; Kirk, Mark D.; Katz, Martin L.

    2016-01-01

    A number of retinal degenerative diseases may be amenable to treatment with continuous intraocular delivery of therapeutic agents that cannot be delivered effectively to the retina via systemic or topical administration. Among these disorders are lysosomal storage diseases resulting from deficiencies in soluble lysosomal enzymes. Most cells, including those of the retina, are able to take up these enzymes and incorporate them in active form into their lysosomes. In theory, therefore, continuous intraocular administration of a normal form of a soluble lysosomal enzyme should be able to cure the molecular defect in the retinas of subjects lacking this enzyme. Experiments were conducted to determine whether genetically modified bone marrow-derived stem cells implanted into the vitreous could be used as vehicles for continuous delivery of such enzymes to the retina. Bone marrow-derived mesenchymal stem cells (MSCs) from normal mice were implanted into the vitreous of mice undergoing retinal degeneration as a result of a mutation in the PPT1 gene. The implanted cells appeared to survive indefinitely in the vitreous without proliferating or invading the retina. This indicates that intravitreal implantation of MSCs is likely a safe means of long-term delivery of proteins synthesized by the implanted cells. Experiments have been initiated to test the efficacy of using genetically modified autologous MSCs to inhibit retinal degeneration in a canine model of neuronal ceroid lipofuscinosis. PMID:26427461

  3. Intracoronary infusion of a combination of bone marrow-derived stem cells in dogs

    PubMed Central

    Minguell, José J; Florenzano, Fernando M; Ramírez, Manuel R; Martínez, Ramón F; Lasala, Gabriel P

    2010-01-01

    BACKGROUND: Infusion of diverse types of bone marrow cells, as a source of endothelial progenitor cells (EPCs), into the ischemic myocardium is emerging as a promising therapy for coronary ischemia, probably mediated by the formation of new blood vessels. Studies have shown that while the procedure is safe and feasible, efficacy results are contentious. The investigators in the present preclinical translation study hypothesized that the infusion of a combination cell product consisting of EPCs and other cell types, such as mesenchymal stem cells, promotes the formation of more stable and mature blood vessels resulting in improved clinical outcomes. The safety and feasibility of the intracoronary infusion of such a cell combination was assessed in a canine model. METHODS: A mixture of canine autologous mononuclear cells (as the source of EPCs) and ex vivo-expanded bone marrow-derived mesenchymal stem cells or a placebo solution were intracoronarily infused into healthy dogs. Follow-up after cell/placebo infusion included an electrocardiogram, serum cardiac enzyme testing, a transthoracic echocardiography and a histopathological heart examination. RESULTS: On follow-up at all time points after infusion, no significant changes or abnormalities in vital signs, electrocardiogram, transthoracic echocardiography and heart histology were detected. CONCLUSIONS: From a clinical perspective, the safety and feasibility of the protocol used in the present animal study demonstrated clinical relevance and provided direct evidence supporting the intracoronary infusion of combination stem/progenitor cell products. PMID:20631864

  4. Exendin-4 protects bone marrow-derived mesenchymal stem cells against oxygen/glucose and serum deprivation-induced apoptosis through the activation of the cAMP/PKA signaling pathway and the attenuation of ER stress

    PubMed Central

    HE, JIEQIONG; WANG, CHAO; SUN, YUNPENG; LU, BO; CUI, JINJIN; DONG, NANA; ZHANG, MAOMAO; LIU, YOUBING; YU, BO

    2016-01-01

    Exendin-4 (ex-4) is a long-acting glucagon-like peptide-1 receptor (GLP-1R) agonist which exerts beneficial effects on glycemic control and promotes cell viability. In the present study, we investigated the anti-apoptotic effects of ex-4, as well as the potential mechanisms responsible for these effects in rat bone marrow-derived mesenchymal stem cells (BM-MSCs) under conditions of oxygen, glucose and serum deprivation (OGD). The apoptosis of the MSCs was induced by subjecting the cells to OGD conditions for 4 h and was detected by Annexin V/PI and Hoechst 33258 staining. The MSCs were pre-conditioned with ex-4 for 12 h prior to being subjected to OGD conditions, and the expression levels of an apoptotic marker (cleaved caspase-3), endoplasmic reticulum (ER) stress markers [phosphorylated (p-)protein kinase RNA-like endoplasmic reticulum kinase (PERK), PERK, binding immunoglobulin protein (BIP), activating transcription factor 4 (ATF-4) and C/EBP homologous protein (CHOP)], as well as those of a survival marker (Bcl-2) were measured by western blot analysis. Furthermore, the mRNA levels of ATF-4 and CHOP were determined by RT-qPCR. ELISA was used to examine the activity of intracellular cAMP. Moreover, the GLP-1R antagonist, exendin9-39 (ex9-39), the protein kinase A (PKA) inhibitor, H89, and small interfering RNA (siRNA) targeting rat ATF-4 and CHOP were co-incubated with the MSCs. The apoptotic rate was markedly diminished following pre-conditioning with ex-4 in a dose-dependent manner (P<0.05). The ER stress markers, p-PERK, BIP, ATF-4 and CHOP, were upregulated in the cells subjected to OGD conditions. Ex-4 pre-conditioning significantly decreased the mRNA and protein levels of ATF-4 and CHOP (P<0.05), and increased the activity of intracellular cAMP (P<0.05). Furthermore, the anti-apoptotic effects of ex-4 were almost reversed by treatment with either H89 or ex9-39 (P<0.05); transfection with siRNA-CHOP significantly reduced the apoptotic rate of the MSCs and

  5. Bone marrow-derived stem cell therapy for metastatic brain cancers.

    PubMed

    Kaneko, Yuji; Tajiri, Naoki; Staples, Meaghan; Reyes, Stephanny; Lozano, Diego; Sanberg, Paul R; Freeman, Thomas B; van Loveren, Harry; Kim, Seung U; Borlongan, Cesar V

    2015-01-01

    We propose that stem cell therapy may be a potent treatment for metastatic melanoma in the brain. Here we discuss the key role of a leaky blood-brain barrier (BBB) that accompanies the development of brain metastases. We review the need to characterize the immunological and inflammatory responses associated with tumor-derived BBB damage in order to reveal the contribution of this brain pathological alteration to the formation and growth of brain metastatic cancers. Next, we discuss the potential repair of the BBB and attenuation of brain metastasis through transplantation of bone marrow-derived mesenchymal stem cells with the endothelial progenitor cell phenotype. In particular, we review the need for evaluation of the efficacy of stem cell therapy in repairing a disrupted BBB in an effort to reduce neuroinflammation, eventually attenuating brain metastatic cancers. The demonstration of BBB repair through augmented angiogenesis and vasculogenesis will be critical to establishing the potential of stem cell therapy for the treatment/prevention of metastatic brain tumors. The overarching hypothesis we advanced here is that BBB breakdown is closely associated with brain metastatic cancers of melanoma, exacerbating the inflammatory response of the brain during metastasis, and ultimately worsening the outcome of metastatic brain cancers. Abrogating this leaky BBB-mediated inflammation via stem cell therapy represents a paradigm-shifting approach to treating brain cancer. This review article discusses the pros and cons of cell therapy for melanoma brain metastases. PMID:25310691

  6. Genetic stability of bone marrow-derived human mesenchymal stromal cells in the Quantum System

    PubMed Central

    Jones, Mark; Varella-Garcia, Marileila; Skokan, Margaret; Bryce, Steven; Schowinsky, Jeffrey; Peters, Rebecca; Vang, Boah; Brecheisen, Michelle; Startz, Thomas; Frank, Nathan; Nankervis, Brian

    2014-01-01

    Background aims The Quantum® Cell Expansion System (Quantum; Terumo BCT, Inc, Lakewood, CO, USA) is a novel hollow fiber-based device that automates and closes the cell culture process, reducing labor intensive tasks such as manual cell culture feeding and harvesting. The manual cell selection and expansion processes for the production of clinical-scale quantities of bone marrow-derived human mesenchymal stromal cells (BM-hMSCs) have been successfully translated onto the Quantum platform previously. The formerly static, manual, in vitro process performed primarily on tissue culture polystyrene substrates may raise the question of whether BM-hMSCs cultured on a hollow fiber platform yields comparable cell quality. Methods A rigorous battery of assays was used to determine the genetic stability of BM-hMSCs selected and produced with the Quantum. In this study, genetic stability was determined by assessing spectral karyotype, micronucleus formation and tumorigenicity to resolve chromosomal aberrations in the stem cell population. Cell phenotype, adherent growth kinetics and tri-lineage differentiation were also evaluated. HMSC bone marrow aspirates, obtained from three approved donors, were expanded in parallel using T225 culture flasks and the Quantum. Results BM-hMSCs harvested from the Quantum demonstrated immunophenotype, morphology and tri-lineage differentiation capacity characteristics consistent with the International Society of Cell Therapy standard for hMSCs. Cell populations showed no malignant neoplastic formation in athymic mice 60 days post-transplant, no clonal chromosomal aberrations were observed and no DNA damage was found as measured by micronucleus formation. Conclusions Quantum-produced BM-hMSCs are of comparable quality and demonstrate analogous genetic stability to BM-hMSCs cultured on tissue culture polystyrene substrates. PMID:23992670

  7. Paracrine effects and heterogeneity of marrow-derived stem/progenitor cells: relevance for the treatment of respiratory diseases.

    PubMed

    Conese, Massimo; Carbone, Annalucia; Castellani, Stefano; Di Gioia, Sante

    2013-01-01

    Stem cell-based treatment may represent a hope for the treatment of acute lung injury and pulmonary fibrosis, and other chronic lung diseases, such as cystic fibrosis, asthma and chronic obstructive pulmonary disease (COPD). It is well established in preclinical models that bone marrow-derived stem and progenitor cells exert beneficial effects on inflammation, immune responses and repairing of damage in virtually all lung-borne diseases. While it was initially thought that the positive outcome was due to a direct engraftment of these cells into the lung as endothelial and epithelial cells, paracrine factors are now considered the main mechanism through which stem and progenitor cells exert their therapeutic effect. This knowledge has led to the clinical use of marrow cells in pulmonary hypertension with endothelial progenitor cells (EPCs) and in COPD with mesenchymal stromal (stem) cells (MSCs). Bone marrow-derived stem cells, including hematopoietic stem/progenitor cells, MSCs, EPCs and fibrocytes, encompass a wide array of cell subsets with different capacities of engraftment and injured tissue-regenerating potential. The characterization/isolation of the stem cell subpopulations represents a major challenge to improve the efficacy of transplantation protocols used in regenerative medicine and applied to lung disorders. PMID:23652321

  8. The Healing Effect of Bone Marrow-Derived Stem Cells in Knee Osteoarthritis: A Case Report.

    PubMed

    Mehrabani, Davood; Mojtahed Jaberi, Fereidoon; Zakerinia, Maryam; Hadianfard, Mohammad Javad; Jalli, Reza; Tanideh, Nader; Zare, Shahrokh

    2016-05-01

    Osteoarthritis (OA) is a prevalent chronic disease impacting on quality of life and has societal and economical burden increasing with age. Yet, no confirmed pharmacological, biological or surgical therapy could prevent the progressive destruction of OA joint. Mesenchymal stem cells (MSCs) with immunosuppressive activities emerged a potential therapy. We describe a magnetic resonance images (MRI) approved 47 years old nomad female suffering from a severe right knee OA. After intra-articular injection of 36×10(6) passage 2 of bone marrow-derived stem cells (BMSCs), the patient's functional status of the knee, the number of stairs she could climb, the pain on visual analog scale (VAS) and walking distance improved after two months post-transplantation. MRI revealed an extension of the repaired tissue over subchondral bone. So as MSC transplantation is a simple technique, resulted into pain relief, minimized donor-site morbidity, provided a better quality of life, significantly improved cartilage quality with no need to hospitalization or surgery, cell transplantation can be considered as a reliable alternative treatment for chronic knee OA. Therefore these findings can be added to the literature on using BMSCs for treatment of OA. PMID:27579273

  9. The Healing Effect of Bone Marrow-Derived Stem Cells in Knee Osteoarthritis: A Case Report

    PubMed Central

    Mehrabani, Davood; Mojtahed Jaberi, Fereidoon; Zakerinia, Maryam; Hadianfard, Mohammad Javad; Jalli, Reza; Tanideh, Nader; Zare, Shahrokh

    2016-01-01

    Osteoarthritis (OA) is a prevalent chronic disease impacting on quality of life and has societal and economical burden increasing with age. Yet, no confirmed pharmacological, biological or surgical therapy could prevent the progressive destruction of OA joint. Mesenchymal stem cells (MSCs) with immunosuppressive activities emerged a potential therapy. We describe a magnetic resonance images (MRI) approved 47 years old nomad female suffering from a severe right knee OA. After intra-articular injection of 36×106 passage 2 of bone marrow-derived stem cells (BMSCs), the patient’s functional status of the knee, the number of stairs she could climb, the pain on visual analog scale (VAS) and walking distance improved after two months post-transplantation. MRI revealed an extension of the repaired tissue over subchondral bone. So as MSC transplantation is a simple technique, resulted into pain relief, minimized donor-site morbidity, provided a better quality of life, significantly improved cartilage quality with no need to hospitalization or surgery, cell transplantation can be considered as a reliable alternative treatment for chronic knee OA. Therefore these findings can be added to the literature on using BMSCs for treatment of OA. PMID:27579273

  10. Donor age and cell passage affects differentiation potential of murine bone marrow-derived stem cells

    PubMed Central

    Kretlow, James D; Jin, Yu-Qing; Liu, Wei; Zhang, Wen Jie; Hong, Tan-Hui; Zhou, Guangdong; Baggett, L Scott; Mikos, Antonios G; Cao, Yilin

    2008-01-01

    Background Bone marrow-derived mesenchymal stem cells (BMSCs) are a widely researched adult stem cell population capable of differentiation into various lineages. Because many promising applications of tissue engineering require cell expansion following harvest and involve the treatment of diseases and conditions found in an aging population, the effect of donor age and ex vivo handling must be understood in order to develop clinical techniques and therapeutics based on these cells. Furthermore, there currently exists little understanding as to how these two factors may be influenced by one another. Results Differences in the adipogenic, chondrogenic, and osteogenic differentiation capacity of murine MSCs harvested from donor animals of different age and number of passages of these cells were observed. Cells from younger donors adhered to tissue culture polystyrene better and proliferated in greater number than those from older animals. Chondrogenic and osteogenic potential decreased with age for each group, and adipogenic differentiation decreased only in cells from the oldest donors. Significant decreases in differentiation potentials due to passage were observed as well for osteogenesis of BMSCs from the youngest donors and chondrogenesis of the cells from the oldest donors. Conclusion Both increasing age and the number of passages have lineage dependent effects on BMSC differentiation potential. Furthermore, there is an obvious interplay between donor age and cell passage that in the future must be accounted for when developing cell-based therapies for clinical use. PMID:18957087

  11. The healing effect of bone marrow-derived stem cells in acute radiation syndrome

    PubMed Central

    Mortazavi, Seyed Mohammad Javad; Shekoohi-Shooli, Fatemeh; Aghamir, Seyed Mahmood Reza; Mehrabani, Davood; Dehghanian, Amirreza; Zare, Shahrokh; Mosleh-Shirazi, Mohammad Amin

    2016-01-01

    Objectives: To determine the effect of bone marrow-derived mesenchymal stem cells (BMSCs) on regeneration of bone marrow and intestinal tissue and survival rate in experimental mice with acute radiation syndrome (ARS). Methods: Forty mice were randomly divided into two equal groups of A receiving no BMSC transplantation and B receiving BMSCs. BMSCs were isolated from the bone marrow and cultured in DMEM media. Both groups were irradiated with 10 Gy (dose rate 0.28 Gy/ min) 60CO during 35 minutes with a field size of 35×35 for all the body area. Twenty-four hours after γ irradiation, 150×103 cells of passage 5 in 150 µl medium were injected intravenously into the tail. Animals were euthanized one and two weeks after cell transplantation. They were evaluated histologically for any changes in bone marrow and intestinal tissues. The survival rate in mice were also determined. Results: A significant increase for bone marrow cell count and survival rate were observed in group B in comparison to group A. Histological findings denoted to a healing in sample tissues. Conclusion: BMSCs could significantly reduce the side effects of ARS and increase the survival rate and healing in injured tissue. As such their transplantation may open a window in treatment of patients with ARS. PMID:27375707

  12. Bone Marrow-Derived Stem Cells: a Mixed Blessing in the Multifaceted World of Diabetic Complications.

    PubMed

    Mangialardi, Giuseppe; Madeddu, Paolo

    2016-05-01

    Diabetes is one of the main economic burdens in health care, which threatens to worsen dramatically if prevalence forecasts are correct. What makes diabetes harmful is the multi-organ distribution of its microvascular and macrovascular complications. Regenerative medicine with cellular therapy could be the dam against life-threatening or life-altering complications. Bone marrow-derived stem cells are putative candidates to achieve this goal. Unfortunately, the bone marrow itself is affected by diabetes, as it can develop a microangiopathy and neuropathy similar to other body tissues. Neuropathy leads to impaired stem cell mobilization from marrow, the so-called mobilopathy. Here, we review the role of bone marrow-derived stem cells in diabetes: how they are affected by compromised bone marrow integrity, how they contribute to other diabetic complications, and how they can be used as a treatment for these. Eventually, we suggest new tactics to optimize stem cell therapy. PMID:27025211

  13. The role of the fibrocyte, a bone marrow-derived mesenchymal progenitor, in reactive and reparative fibroses.

    PubMed

    Bellini, Alberto; Mattoli, Sabrina

    2007-09-01

    Human fibrocytes are mesenchymal progenitors that exhibit mixed morphological and molecular characteristics of hematopoietic stem cells, monocytes and fibroblasts. They likely represent the obligate intermediate stage of differentiation into mature mesenchymal cells of a bone marrow-derived precursor of the monocyte lineage under permissive conditions. On in vitro stimulation with pro-fibrotic cytokines and growth factors, human fibrocytes produce large quantities of extracellular matrix components and further differentiate into cells identical to the contractile myofibroblasts that emerge at the tissue sites during repair processes and in some fibrotic lesions. Studies in various animal models of wound healing or fibrotic diseases have confirmed the ability of fibrocytes to differentiate into mature mesenchymal cells in vivo and have suggested a causal link between fibrocyte accumulation and ongoing tissue fibrogenesis or vascular remodeling in response to tissue damage or hypoxia. Fibrocytes synthesizing new collagen or acquiring myofibroblast markers have been detected in human hypertrophic scars, in the skin of patients affected by nephrogenic systemic fibrosis, in human atherosclerotic lesions, and in pulmonary diseases characterized by repeated cycles of inflammation and repair, like asthma. The presence of fibrocyte-like cells has been reported in human chronic pancreatitis and chronic cystitis. Similar cells also populate the stroma surrounding human benign tumors. The available data indicate that human fibrocytes serve as a source of mature mesenchymal cells during reparative processes and in fibrotic disorders or stromal reactions predominantly associated with a persistent inflammatory infiltrate or with the selective recruitment of monocytes induced by ischemic changes and tumor development. A deeper understanding of the mechanisms involved in fibrocyte differentiation in these pathological conditions may lead to the development of novel therapies for

  14. The Effect of Bone-Marrow-Derived Stem Cells and Adipose-Derived Stem Cells on Wound Contraction and Epithelization

    PubMed Central

    Uysal, Cagri A.; Tobita, Morikuni; Hyakusoku, Hiko; Mizuno, Hiroshi

    2014-01-01

    Objective: The relationship between the wound contraction and levels of α-smooth muscle actin (α-SMA) has been revealed in different studies. We aimed to investigate the effects of mesenchymal stem cells (MSCs), mainly bone-marrow-derived stem cells (BSCs) and adipose-derived stem cells (ASCs), and find out the α-SMA, fibroblast growth factor (FGF), transforming growth factor beta, and vascular endothelial growth factor (VEGF) levels on an in vivo acute wound healing model after the application of MSCs. Approach: Four circular skin defects were formed on the dorsum of Fisher rats (n=20). The defects were applied phosphate-buffered saline (PBS), ASCs, BSCs, and patchy skin graft, respectively. The healing time and scar area were noted. Results: There was a statistical decrease in the healing time in ASC, BSC, and skin graft groups (p<0.05). However, the scar was smaller in the PBS group (p<0.05). The α-SMA levels were statistically lower in ASC, BSC, and graft groups (p<0.05). The FGF levels were statistically higher in ASC and BSC groups (p<0.05). The differentiation of the injected MSCs to endothelial cells and keratinocytes was observed. Innovation and Conclusion: MSCs decrease the healing time and contraction of the wound while increasing the epithelization rate by increasing angiogenesis. PMID:24940554

  15. Different Balance of Wnt Signaling in Adult and Fetal Bone Marrow-Derived Mesenchymal Stromal Cells.

    PubMed

    Paciejewska, Maja M; Maijenburg, Marijke W; Gilissen, Christian; Kleijer, Marion; Vermeul, Kim; Weijer, Kees; Veltman, Joris A; von Lindern, Marieke; van der Schoot, C Ellen; Voermans, Carlijn

    2016-06-15

    Mesenchymal stromal cells (MSCs) are applied as novel therapeutics for their regenerative and immune-suppressive capacities. Clinical applications, however, require extensive expansion of MSCs. Fetal bone marrow-derived MSCs (FBMSCs) proliferate faster than adult bone marrow-derived MSC (ABMSCs). To optimize expansion and function of MSC in general, we explored the differences between ABMSC and FBMSC. Gene expression profiling implicated differential expression of genes encoding proteins in the Wnt signaling pathway, including excreted inhibitors of Wnt signaling, particularly by ABMSC. Both MSC types had a similar basal level of canonical Wnt signaling. Abrogation of autocrine Wnt production by inhibitor of Wnt production-2 (IWP2) reduced canonical Wnt signaling and cell proliferation of FBMSCs, but hardly affected ABMSC. Addition of exogenous Wnt3a, however, induced expression of the target genes lymphocyte enhancer-binding factor (LEF) and T-cell factor (TCF) faster and at lower Wnt3a levels in ABMSC compared to FBMSC. Medium replacement experiments indicated that ABMSC produce an inhibitor of Wnt signaling that is effective on ABMSC itself but not on FBMSC, whereas FBMSC excrete (Wnt) factors that stimulate proliferation of ABMSC. In contrast, FBMSC were not able to support hematopoiesis, whereas ABMSC displayed hematopoietic support sensitive to IWP2, the inhibitor of Wnt factor excretion. In conclusion, ABMSC and FBMSC differ in their Wnt signature. While FBMSC produced factors, including Wnt signals, that enhanced MSC proliferation, ABMSC produced Wnt factors in a setting that enhanced hematopoietic support. Thus, further unraveling the molecular basis of this phenomenon may lead to improvement of clinical expansion protocols of ABMSCs. PMID:27154244

  16. Data on nitric oxide production by human bone marrow-derived mesenchymal stromal cells.

    PubMed

    Najar, Mehdi; Fayyad-Kazan, Mohammad; Fayyad-Kazan, Hussein; Meuleman, Nathalie; Bron, Dominique; Lagneaux, Laurence

    2016-09-01

    Due to its anti-inflammatory and immunosuppressive potential, Nitric oxide (NO), a gaseous radical, is of special importance during graft-versus-host diseases (GVHD) and feoto-maternal tolerance. NO is a major mediator of murine mesenchymal stromal cells (MSCs)-immunosuppressive capacity. In this data article, we characterized NO production by human bone marrow-derived MSCs (hBMSCs). MSCs, isolated from healthy donors (n=5), were defined according to the International Society for cellular Therapy (ISCT) guidelines. Based on a fluorometric detection system, and upon using Nitrite ([Formula: see text])/Nitrate ( [Formula: see text]) Assay Kit, the amounts of NO metabolites ( [Formula: see text] and [Formula: see text]) produced by hBMSCs, being grown in a culture medium either lacking (constitutive condition) or containing IL-4, IL-10 or a pro-inflammatory cytokine cocktail made of IL-1β, TNF-α, IFN-α and IFN-γ, were assessed. All assays were carried out in triplicates and the mean values are reported. The data from this study supports and corroborates the discussion associated with our previously published work entitled "The Immunomodulatory Potential of Mesenchymal Stromal Cells: A Story of a Regulatory Network" (Najar et al., 2016) [1]. PMID:27536712

  17. Mesenchymal stem cells induce dermal fibroblast responses to injury

    SciTech Connect

    Smith, Andria N.; Willis, Elise; Chan, Vincent T.; Muffley, Lara A.; Isik, F. Frank; Gibran, Nicole S.; Hocking, Anne M.

    2010-01-01

    Although bone marrow-derived mesenchymal stem cells have been shown to promote repair when applied to cutaneous wounds, the mechanism for this response remains to be determined. The aim of this study was to determine the effects of paracrine signaling from mesenchymal stem cells on dermal fibroblast responses to injury including proliferation, migration and expression of genes important in wound repair. Dermal fibroblasts were co-cultured with bone marrow-derived mesenchymal stem cells grown in inserts, which allowed for paracrine interactions without direct cell contact. In this co-culture model, bone marrow-derived mesenchymal stem cells regulate dermal fibroblast proliferation, migration and gene expression. When co-cultured with mesenchymal stem cells, dermal fibroblasts show increased proliferation and accelerated migration in a scratch assay. A chemotaxis assay also demonstrated that dermal fibroblasts migrate towards bone marrow-derived mesenchymal stem cells. A PCR array was used to analyze the effect of mesenchymal stem cells on dermal fibroblast gene expression. In response to mesenchymal stem cells, dermal fibroblasts up-regulate integrin alpha 7 expression and down-regulate expression of ICAM1, VCAM1 and MMP11. These observations suggest that mesenchymal stem cells may provide an important early signal for dermal fibroblast responses to cutaneous injury.

  18. In vitro expanded bone marrow-derived murine (C57Bl/KaLwRij) mesenchymal stem cells can acquire CD34 expression and induce sarcoma formation in vivo

    SciTech Connect

    Xu, Song; De Becker, Ann; De Raeve, Hendrik; Van Camp, Ben; Vanderkerken, Karin; Van Riet, Ivan

    2012-08-03

    Highlights: Black-Right-Pointing-Pointer Murine MSCs can undergo spontaneously malignant transformation and form sarcoma. Black-Right-Pointing-Pointer Acquisition of CD34 is a transformation type for MSCs into sarcoma. Black-Right-Pointing-Pointer Notch/Hh/Wnt pathways are related to the malignant phenotype of transformed MSCs. -- Abstract: Mesenchymal stem cells (MSCs) have currently generated numerous interests in pre-clinical and clinical applications due to their multiple lineages differentiation potential and immunomodulary effects. However, accumulating evidence indicates that MSCs, especially murine MSCs (mMSCs), can undergo spontaneous transformation after long-term in vitro culturing, which might reduce the therapeutic application possibilities of these stem cells. In the present study, we observed that in vitro expanded bone marrow (BM) derived mMSCs from the C57Bl/KaLwRij mouse strain can lose their specific stem cells markers (CD90 and CD105) and acquire CD34 expression, accompanied with an altered morphology and an impaired tri-lineages differentiation capacity. Compared to normal mMSCs, these transformed mMSCs exhibited an increased proliferation rate, an enhanced colony formation and migration ability as well as a higher sensitivity to anti-tumor drugs. Transformed mMSCs were highly tumorigenic in vivo, resulting in aggressive sarcoma formation when transplanted in non-immunocompromised mice. Furthermore, we found that Notch signaling downstream genes (hey1, hey2 and heyL) were significantly upregulated in transformed mMSCs, while Hedgehog signaling downstream genes Gli1 and Ptch1 and the Wnt signaling downstream gene beta-catenin were all decreased. Taken together, we observed that murine in vitro expanded BM-MSCs can transform into CD34 expressing cells that induce sarcoma formation in vivo. We assume that dysregulation of the Notch(+)/Hh(-)/Wnt(-) signaling pathway is associated with the malignant phenotype of the transformed mMSCs.

  19. Dental mesenchymal stem cells.

    PubMed

    Sharpe, Paul T

    2016-07-01

    Mammalian teeth harbour mesenchymal stem cells (MSCs), which contribute to tooth growth and repair. These dental MSCs possess many in vitro features of bone marrow-derived MSCs, including clonogenicity, expression of certain markers, and following stimulation, differentiation into cells that have the characteristics of osteoblasts, chondrocytes and adipocytes. Teeth and their support tissues provide not only an easily accessible source of MSCs but also a tractable model system to study their function and properties in vivo In addition, the accessibility of teeth together with their clinical relevance provides a valuable opportunity to test stem cell-based treatments for dental disorders. This Review outlines some recent discoveries in dental MSC function and behaviour and discusses how these and other advances are paving the way for the development of new biologically based dental therapies. PMID:27381225

  20. The effect of telomerase template antagonist GRN163L on bone-marrow-derived rat mesenchymal stem cells is reversible and associated with altered expression of cyclin d1, cdk4 and cdk6.

    PubMed

    Tokcaer-Keskin, Zeynep; Dikmen, Zeliha G; Ayaloglu-Butun, Fatma; Gultekin, Sinan; Gryaznov, Sergei M; Akcali, Kamil Can

    2010-06-01

    Telomerase activity is essential for the continued growth and survival of malignant cells, therefore inhibition of this activity presents an attractive target for anti-cancer therapy. The telomerase inhibitor GRN163L, was shown to inhibit the growth of cancer cells both in vitro and in vivo. Mesenchymal stem cells (MSCs) also show telomerase activity in maintaining their self-renewal; therefore the effects of telomerase inhibitors on MSCs may be an issue of concern. MSCs are multipotent cells and are important for the homeostasis of the organism. In this study, we sought to demonstrate in vitro effects of GRN163L on rat MSCs. When MSCs were treated with 1 microM GRN163L, their phenotype changed from spindle-shaped cells to rounded ones and detached from the plate surface, similar to cancer cells. Quantitative-RT-PCR and immunoblotting results revealed that GRN163L holds MSCs at the G1 state of the cell cycle, with a drastic decrease in mRNA and protein levels of cyclin D1 and its cdk counterparts, cdk4 and cdk6. This effect was not observed when MSCs were treated with a mismatch control oligonucleotide. One week after GRN163L was removed, mRNA and protein expressions of the genes, as well as the phenotype of MSCs returned to those of untreated cells. Therefore, we concluded that GRN163L does not interfere with the self-renewal and differentiation of MSCs under short term in vitro culture conditions. Our study provides additional support for treating cancers by administrating GRN163L without depleting the body's stem cell pools. PMID:20180048

  1. Immune Dysfunction Associated with Abnormal Bone Marrow-Derived Mesenchymal Stroma Cells in Senescence Accelerated Mice

    PubMed Central

    Li, Ming; Guo, Kequan; Adachi, Yasushi; Ikehara, Susumu

    2016-01-01

    Senescence accelerated mice (SAM) are a group of mice that show aging-related diseases, and SAM prone 10 (SAMP10) show spontaneous brain atrophy and defects in learning and memory. Our previous report showed that the thymus and the percentage of T lymphocytes are abnormal in the SAMP10, but it was unclear whether the bone marrow-derived mesenchymal stroma cells (BMMSCs) were abnormal, and whether they played an important role in regenerative medicine. We thus compared BMMSCs from SAMP10 and their control, SAM-resistant (SAMR1), in terms of cell cycle, oxidative stress, and the expression of PI3K and mitogen-activated protein kinase (MAPK). Our cell cycle analysis showed that cell cycle arrest occurred in the G0/G1 phase in the SAMP10. We also found increased reactive oxygen stress and decreased PI3K and MAPK on the BMMSCs. These results suggested the BMMSCs were abnormal in SAMP10, and that this might be related to the immune system dysfunction in these mice. PMID:26840301

  2. Caffeine inhibits the viability and osteogenic differentiation of rat bone marrow-derived mesenchymal stromal cells

    PubMed Central

    Zhou, Y; Guan, XX; Zhu, ZL; Guo, J; Huang, YC; Hou, WW; Yu, HY

    2010-01-01

    BACKGROUND AND PURPOSE Caffeine is consumed extensively in Europe and North America. As a risk factor for osteoporosis, epidemiological studies have observed that caffeine can decrease bone mineral density, adversely affect calcium absorption and increase the risk of bone fracture. However, the exact mechanisms have not been fully investigated. Here, we examined the effects of caffeine on the viability and osteogenesis of rat bone marrow-derived mesenchymal stromal cells (rBMSCs). EXPERIMENTAL APPROACH Cell viability, apoptosis and necrosis were quantified using thymidine incorporation and flow cytometry. Sequential gene expressions in osteogenic process were measured by real-time PCR. cAMP, alkaline phosphatase and osteocalcin were assessed by immunoassay, spectrophotometry and radioimmunoassay, respectively. Mineralization was determined by calcium deposition. KEY RESULTS After treating BMSCs with high caffeine concentrations (0.1–1 mM), their viability decreased in a concentration-dependent manner. This cell death was primarily due to necrosis and, to a small extent, apoptosis. Genes and protein sequentially expressed in osteogenesis, including Cbfa1/Runx2, collagen I, alkaline phosphatase and its protein, were significantly downregulated except for osteocalcin and its protein. Moreover, caffeine inhibited calcium deposition in a concentration- and time-dependent manner, but increased intracellular cAMP in a concentration-dependent manner. CONCLUSIONS AND IMPLICATIONS By suppressing the commitment of BMSCs to the osteogenic lineage and selectively inhibiting gene expression, caffeine downregulated some important events in osteogenesis and ultimately affected bone mass. PMID:20726981

  3. Bone marrow-derived mesenchymal stromal cells inhibit Th2-mediated allergic airways inflammation in mice.

    PubMed

    Goodwin, Meagan; Sueblinvong, Viranuj; Eisenhauer, Philip; Ziats, Nicholas P; LeClair, Laurie; Poynter, Matthew E; Steele, Chad; Rincon, Mercedes; Weiss, Daniel J

    2011-07-01

    Bone marrow-derived mesenchymal stromal cells (BMSCs) mitigate inflammation in mouse models of acute lung injury. However, specific mechanisms of BMSC actions on CD4 T lymphocyte-mediated inflammation in vivo remain poorly understood. Limited data suggests promotion of Th2 phenotype in models of Th1-mediated diseases. However, whether this might alleviate or worsen Th2-mediated diseases such as allergic asthma is unknown. To ascertain the effects of systemic administration of BMSCs in a mouse model of Th2-mediated allergic airways inflammation, ovalbumin (OVA)-induced allergic airways inflammation was induced in wild-type C57BL/6 and BALB/c mice as well as in interferon-γ (IFNγ) receptor null mice. Effects of systemic administration during antigen sensitization of either syngeneic or allogeneic BMSC on airways hyperreactivity, lung inflammation, antigen-specific CD4 T lymphocytes, and serum immunoglobulins were assessed. Both syngeneic and allogeneic BMSCs inhibited airways hyperreactivity and lung inflammation through a mechanism partly dependent on IFNγ. However, contrary to existing data, BMSCs did not affect antigen-specific CD4 T lymphocyte proliferation but rather promoted Th1 phenotype in vivo as assessed by both OVA-specific CD4 T lymphocyte cytokine production and OVA-specific circulating immunoglobulins. BMSCs treated to prevent release of soluble mediators and a control cell population of primary dermal skin fibroblasts only partly mimicked the BMSC effects and in some cases worsened inflammation. In conclusion, BMSCs inhibit Th2-mediated allergic airways inflammation by influencing antigen-specific CD4 T lymphocyte differentiation. Promotion of a Th1 phenotype in antigen-specific CD4 T lymphocytes by BMSCs is sufficient to inhibit Th2-mediated allergic airways inflammation through an IFNγ-dependent process. PMID:21544902

  4. Real-time-guided bone regeneration around standardized critical size calvarial defects using bone marrow-derived mesenchymal stem cells and collagen membrane with and without using tricalcium phosphate: an in vivo micro-computed tomographic and histologic experiment in rats.

    PubMed

    Al-Hezaimi, Khalid; Ramalingam, Sundar; Al-Askar, Mansour; ArRejaie, Aws S; Nooh, Nasser; Jawad, Fawad; Aldahmash, Abdullah; Atteya, Muhammad; Wang, Cun-Yu

    2016-01-01

    The aim of the present real time in vivo micro-computed tomography (µCT) and histologic experiment was to assess the efficacy of guided bone regeneration (GBR) around standardized calvarial critical size defects (CSD) using bone marrow-derived mesenchymal stem cells (BMSCs), and collagen membrane (CM) with and without tricalcium phosphate (TCP) graft material. In the calvaria of nine female Sprague-Dawley rats, full-thickness CSD (diameter 4.6 mm) were created under general anesthesia. Treatment-wise, rats were divided into three groups. In group 1, CSD was covered with a resorbable CM; in group 2, BMSCs were filled in CSD and covered with CM; and in group 3, TCP soaked in BMSCs was placed in CSD and covered with CM. All defects were closed using resorbable sutures. Bone volume and bone mineral density of newly formed bone (NFB) and remaining TCP particles and rate of new bone formation was determined at baseline, 2, 4, 6, and 10 weeks using in vivo µCT. At the 10th week, the rats were killed and calvarial segments were assessed histologically. The results showed that the hardness of NFB was similar to that of the native bone in groups 1 and 2 as compared to the NFB in group 3. Likewise, values for the modulus of elasticity were also significantly higher in group 3 compared to groups 1 and 2. This suggests that TCP when used in combination with BMSCs and without CM was unable to form bone of significant strength that could possibly provide mechanical "lock" between the natural bone and NFB. The use of BMSCs as adjuncts to conventional GBR initiated new bone formation as early as 2 weeks of treatment compared to when GBR is attempted without adjunct BMSC therapy. PMID:27025260

  5. Real-time-guided bone regeneration around standardized critical size calvarial defects using bone marrow-derived mesenchymal stem cells and collagen membrane with and without using tricalcium phosphate: an in vivo micro-computed tomographic and histologic experiment in rats

    PubMed Central

    Al-Hezaimi, Khalid; Ramalingam, Sundar; Al-Askar, Mansour; ArRejaie, Aws S; Nooh, Nasser; Jawad, Fawad; Aldahmash, Abdullah; Atteya, Muhammad; Wang, Cun-Yu

    2016-01-01

    The aim of the present real time in vivo micro-computed tomography (µCT) and histologic experiment was to assess the efficacy of guided bone regeneration (GBR) around standardized calvarial critical size defects (CSD) using bone marrow-derived mesenchymal stem cells (BMSCs), and collagen membrane (CM) with and without tricalcium phosphate (TCP) graft material. In the calvaria of nine female Sprague-Dawley rats, full-thickness CSD (diameter 4.6 mm) were created under general anesthesia. Treatment-wise, rats were divided into three groups. In group 1, CSD was covered with a resorbable CM; in group 2, BMSCs were filled in CSD and covered with CM; and in group 3, TCP soaked in BMSCs was placed in CSD and covered with CM. All defects were closed using resorbable sutures. Bone volume and bone mineral density of newly formed bone (NFB) and remaining TCP particles and rate of new bone formation was determined at baseline, 2, 4, 6, and 10 weeks using in vivo µCT. At the 10th week, the rats were killed and calvarial segments were assessed histologically. The results showed that the hardness of NFB was similar to that of the native bone in groups 1 and 2 as compared to the NFB in group 3. Likewise, values for the modulus of elasticity were also significantly higher in group 3 compared to groups 1 and 2. This suggests that TCP when used in combination with BMSCs and without CM was unable to form bone of significant strength that could possibly provide mechanical “lock” between the natural bone and NFB. The use of BMSCs as adjuncts to conventional GBR initiated new bone formation as early as 2 weeks of treatment compared to when GBR is attempted without adjunct BMSC therapy. PMID:27025260

  6. The therapeutic potential of bone marrow-derived mesenchymal stromal cells on hepatocellular carcinoma.

    PubMed

    Bayo, Juan; Marrodán, Mariano; Aquino, Jorge B; Silva, Marcelo; García, Mariana G; Mazzolini, Guillermo

    2014-03-01

    Mesenchymal stromal cells (MSCs) are more often obtained from adult and extraembryonic tissues, with the latter sources being likely better from a therapeutic perspective. MSCs show tropism towards inflamed or tumourigenic sites. Mechanisms involved in MSC recruitment into tumours are comprehensively analysed, including chemoattractant signalling axes, endothelial adhesion and transmigration. In addition, signals derived from hepatocellular carcinoma (HCC) tumour microenvironment and their influence in MSC tropism and tumour recruitment are dissected, as well as the present controversy regarding their influence on tumour growth and/or metastasis. Finally, evidences available on the use of MSCs and other selected progenitor/stem cells as vehicles of antitumourigenic genes are discussed. A better knowledge of the mechanisms involved in progenitor/stem cell recruitment to HCC tumours is proposed in order to enhance their tumour targeting which may result in improvements in cell-based gene therapy strategies. PMID:24112437

  7. Bone marrow and bone marrow derived mononuclear stem cells therapy for the chronically ischemic myocardium

    SciTech Connect

    Waksman, Ron; Baffour, Richard

    2003-09-01

    Bone marrow stem cells have been shown to differentiate into various phenotypes including cardiomyocytes, vascular endothelial cells and smooth muscle. Bone marrow stem cells are mobilized and home in to areas of injured myocardium where they are involved in tissue repair. In addition, bone marrow secretes multiple growth factors, which are essential for angiogenesis and arteriogenesis. In some patients, these processes are not enough to avert clinical symptoms of ischemic disease. Therefore, in vivo administration of an adequate number of stem cells would be a significant therapeutic advance. Unfractionated bone marrow derived mononuclear stem cells, which contain both hematopoietic and nonhematopoietic cells may be more appropriate for cell therapy. Studies in animal models suggest that implantation of different types of stem cells improve angiogenesis and arteriogenesis, tissue perfusion as well as left ventricular function. Several unanswered questions remain. For example, the optimal delivery approach, dosage and timing of the administration of cell therapy as well as durability of improvements need to be studied. Early clinical studies have demonstrated safety and feasibility of various cell therapies in ischemic disease. Randomized, double blind and placebo-controlled clinical trials need to be completed to determine the effectiveness of stem cell.

  8. Impact of parathyroid hormone on bone marrow-derived stem cell mobilization and migration.

    PubMed

    Huber, Bruno C; Grabmaier, Ulrich; Brunner, Stefan

    2014-11-26

    Parathyroid hormone (PTH) is well-known as the principal regulator of calcium homeostasis in the human body and controls bone metabolism via actions on the survival and activation of osteoblasts. The intermittent administration of PTH has been shown to stimulate bone production in mice and men and therefore PTH administration has been recently approved for the treatment of osteoporosis. Besides to its physiological role in bone remodelling PTH has been demonstrated to influence and expand the bone marrow stem cell niche where hematopoietic stem cells, capable of both self-renewal and differentiation, reside. Moreover, intermittent PTH treatment is capable to induce mobilization of progenitor cells from the bone marrow into the bloodstream. This novel function of PTH on modulating the activity of the stem cell niche in the bone marrow as well as on mobilization and regeneration of bone marrow-derived stem cells offers new therapeutic options in bone marrow and stem cell transplantation as well as in the field of ischemic disorders. PMID:25426261

  9. Bone marrow derived stem cells in regenerative medicine as Advanced Therapy Medicinal Products

    PubMed Central

    Astori, Giuseppe; Soncin, Sabrina; Lo Cicero, Viviana; Siclari, Francesco; Sürder, Daniel; Turchetto, Lucia; Soldati, Gianni; Moccetti, Tiziano

    2010-01-01

    Bone marrow derived stem cells administered after minimal manipulation represent an important cell source for cellbased therapies. Clinical trial results, have revealed both safety and efficacy of the cell reinfusion procedure in many cardiovascular diseases. Many of these early clinical trials were performed in a period before the entry into force of the US and European regulation on cellbased therapies. As a result, conflicting data have been generated on the effectiveness of those therapies in certain conditions as acute myocardial infarction. As more academic medical centers and private companies move toward exploiting the full potential of cellbased medicinal products, needs arise for the development of the infrastructure necessary to support these investigations. This review describes the regulatory environment surrounding the production of cell based medicinal products and give practical aspects for cell isolation, characterization, production following Good Manufacturing Practice, focusing on the activities associated with the investigational new drug development. PMID:20589167

  10. Use of bone marrow derived stem cells in trauma and orthopaedics: A review of current concepts

    PubMed Central

    Pastides, Philip S; Welck, Matthew J; Khan, Wasim S

    2015-01-01

    There is a considerable amount of interest in the future role of bone marrow-derived stem cells (BMDSCs) and tissue engineering techniques to manage conditions within the musculoskeletal system. Repair of soft tissue and bone defects, in the early stages of injury, may lead to a reduction in progression of symptoms. Furthermore, troublesome soft tissue injuries that are notoriously fraught with problems either in healing or function, could be augmented with such techniques. The aim of this review paper is to look at the advances in such strategies to tackle these problems and assess how BMDSCs, with the aid of growth factors and scaffolds, are being used in vitro, animal and even human models to treat problems within the field of trauma and orthopaedics. There is plenty of evidence that the results are encouraging and thus gaining momentum toward their use in human studies. PMID:26191493

  11. Endometriosis Impairs Bone Marrow-Derived Stem Cell Recruitment to the Uterus Whereas Bazedoxifene Treatment Leads to Endometriosis Regression and Improved Uterine Stem Cell Engraftment

    PubMed Central

    Sakr, Sharif; Naqvi, Hanyia; Komm, Barry

    2014-01-01

    Endometriosis is a disease defined by the ectopic growth of uterine endometrium. Stem cells contribute to the generation of endometriosis as well as to repair and regeneration of normal endometrium. Here we demonstrate that the selective estrogen receptor modulator bazedoxifene (BZA), administered with conjugated estrogens (CEs), leads to regression of endometriosis lesions as well as reduction in stem cell recruitment to the lesions. Female mice underwent transplantation of male bone marrow. Endometrium was transplanted in the peritoneal cavity of half to create experimental endometriosis. Mice with or without experimental endometriosis were randomized to BZA/CE or vehicle treatment. Endometriosis lesions, bone marrow-derived mesenchymal stem cell engraftment of the lesions, and eutopic endometrium as well as ovarian stimulation were assessed. BZA treatment significantly reduced lesion size, gland number, and expression of proliferation marker proliferating cell nuclear antigen. Ovarian weight was not affected. Stem cells were recruited to the endometriosis lesions, and this recruitment was dramatically reduced by BZA/CE treatment. Stem cell engraftment was reduced in the uterus of animals with endometriosis; however the number of stem cells engrafting the uterus was completely restored by treatment with BZA/CE. Competition between endometriosis and the eutopic endometrium for a limited supply of stem cells and depletion of normal stem cells flux to the uterus is a novel mechanism by which endometriosis interferes with endometrial function and fertility. BZA/CE not only treats lesions of endometriosis, it also dramatically reduces stem cell recruitment to the lesions and restores stem cell engraftment of the uterine endometrium. PMID:24484171

  12. Human term placenta-derived mesenchymal stromal cells are less prone to osteogenic differentiation than bone marrow-derived mesenchymal stromal cells.

    PubMed

    Pilz, Gregor A; Ulrich, Christine; Ruh, Manuel; Abele, Harald; Schäfer, Richard; Kluba, Torsten; Bühring, Hans-Jörg; Rolauffs, Bernd; Aicher, Wilhelm K

    2011-04-01

    Mesenchymal stromal cells (MSC) can be isolated from different tissues. They are capable of differentiating in vitro, for example, to osteoblasts, chondrocytes, or adipocytes. In contrast to CD34 for hematopoietic stem cells, a distinct MSC-defining antibody is not available. Further, for hematopoietic cells lineage-defining antigens such as CD3 or CD20 are known. In contrast, for MSC-derived cells lineage-associated cell surface markers are far from being established. We therefore investigated expression of cell surface antigens on human term placenta-derived MSC (pMSC) in more detail and correlated expression pattern to the osteogenic differentiation capacity of the MSC. We report that pMSC expressed the typical cell surface antigens at levels comparable to bone marrow-derived MSC (bmMSC), including CD73, CD90, and CD105, but did not express CD11b, CD34, and CD45. Further, CD164, TNAP, and the W5C5 antigens were detected on pMSC, whereas CD349 was not observed. Some pMSC expressed CD146 at low or moderate levels, and their osteogenic differentiation potential was weak. In contrast, bmMSC expressed CD146 at high levels, expression of alkaline phosphatase was significantly higher, and they presented a pronounced osteogenic differentiation potential. We conclude that MSC from different sources differ in their expression of distinct markers, and that this may correlate in part with their lineage determination. Thus, a higher percentage of bmMSC expressed CD146 at prominent levels and such cells may be better suited for bone repair. In contrast, many pMSC expressed CD146 at low or moderate levels. They, therefore, may be suitable for applications in which osteogenic differentiation is undesirable. PMID:21047215

  13. Isolation and Manufacture of Clinical-Grade Bone Marrow-Derived Human Mesenchymal Stromal Cells.

    PubMed

    Miller, Renuka P; Hanley, Patrick J

    2016-01-01

    Mesenchymal stromal cells (MSCs) are multipotent cells with both regenerative and immunomodulatory capacities. These unique properties make them appealing as a biologic, with multiple phase 1-3 clinical trials currently testing their safety and efficacy. Although expanding MSCs does not require extensive manipulation, expanding MSCs for use in clinical trials does require the knowledge and safety that are delineated in current good manufacturing practices (GMPs). Here we briefly detail the characteristics of MSCs and considerations for expanding them for clinical use. We then include a step-by-step protocol for expanding MSCs for early phase clinical trials, with important notes to consider during the expansion of these MSCs. PMID:27236680

  14. Route of delivery influences biodistribution of human bone marrow-derived mesenchymal stromal cells following experimental bone marrow transplantation

    PubMed Central

    Wang, Fangjing; Eid, Saada; Dennis, James E; Cooke, Kenneth R; Auletta, Jeffery J; Lee, Zhenghong

    2015-01-01

    Mesenchymal stromal cells (MSCs) have shown promise as treatment for graft-versus-host disease (GvHD) following allogeneic bone marrow transplantation (alloBMT). Mechanisms mediating in vivo effects of MSCs remain largely unknown, including their biodistribution following infusion. To this end, human bone-marrow derived MSCs (hMSCs) were injected via carotid artery (IA) or tail vein (TV) into allogeneic and syngeneic BMT recipient mice. Following xenogeneic transplantation, MSC biodistribution was measured by bioluminescence imaging (BLI) using hMSCs transduced with a reporter gene system containing luciferase and by scintigraphic imaging using hMSCs labeled with [99mTc]-HMPAO. Although hMSCs initially accumulated in the lungs in both transplant groups, more cells migrated to organs in alloBMT recipient as measured by in vivo BLI and scintigraphy and confirmed by ex vivo BLI imaging, immunohistochemistry and quantitative RT-PCR. IA injection resulted in persistent whole–body hMSC distribution in alloBMT recipients, while hMSCs were rapidly cleared in the syngeneic animals within one week. In contrast, TV-injected hMSCs were mainly seen in the lungs with fewer cells traveling to other organs. Summarily, these results demonstrate the potential use of IA injection to alter hMSC biodistribution in order to more effectively deliver hMSCs to targeted tissues and microenvironments. PMID:27330253

  15. Bone regeneration in calvarial defects in a rat model by implantation of human bone marrow-derived mesenchymal stromal cell spheroids.

    PubMed

    Suenaga, Hideyuki; Furukawa, Katsuko S; Suzuki, Yukako; Takato, Tsuyoshi; Ushida, Takashi

    2015-11-01

    Mesenchymal stem cell (MSC) condensation contributes to membrane ossification by enhancing their osteodifferentiation. We investigated bone regeneration in rats using the human bone marrow-derived MSC-spheroids prepared by rotation culture, without synthetic or exogenous biomaterials. Bilateral calvarial defects (8 mm) were created in nude male rats; the left-sided defects were implanted with MSC-spheroids, β-tricalcium phosphate (β-TCP) granules, or β-TCP granules + MSC-spheroids, while the right-sided defects served as internal controls. Micro-computed tomography and immunohistochemical staining for osteocalcin/osteopontin indicated formation of new, full-thickness bones at the implantation sites, but not at the control sites in the MSC-spheroid group. Raman spectroscopy revealed similarity in the spectral properties of the repaired bone and native calvarial bone. Mechanical performance of the bones in the MSC-implanted group was good (50 and 60% those of native bones, respectively). All tests showed poor bone regeneration in the β-TCP and β-TCP + MSC-spheroid groups. Thus, significant bone regeneration was achieved with MSC-spheroid implantation into bone defects, justifying further investigation. PMID:26449444

  16. Clinical-scale expansion of a mixed population of bone-marrow-derived stem and progenitor cells for potential use in bone-tissue regeneration.

    PubMed

    Dennis, James E; Esterly, Kelly; Awadallah, Amad; Parrish, Christopher R; Poynter, Gregory M; Goltry, Kristin L

    2007-10-01

    Preclinical and clinical studies have demonstrated the ability of bone marrow derived stem and progenitor cells to regenerate many tissues, including bone. Methods to expand or enrich progenitors from bone marrow are common; however, these methods include many steps not amenable to clinical use. A closed automated cell production culture system was developed for clinical-scale ex vivo production of bone marrow-derived stem and progenitor cells for hematopoietic reconstitution. The current study tested the ability of this bioreactor system to produce progenitor cells, termed tissue repair cells (TRC), possessing osteogenic potential. Three TRC formulations were evaluated: (a) cells cultured without exogenous cytokines (TRC); (b) cells cultured with exogenous cytokines (TRC-C); and (c) an adherent subset of TRC-C (TRC-C(Ad)). Starting human bone marrow mononuclear cells (BM MNC) and TRC products were characterized for the expression of cell surface markers, in vitro colony forming ability, and in vivo osteogenic potential. Results showed significant expansion of mesenchymal progenitors (CD90+, CD105+, and CD166+) in each TRC formulation. In vivo bone formation, measured by histology, was highest in the TRC group, followed by TRC-C(Ad) and TRC-C. The TRC product outperformed starting BM MNC and had equivalent bone forming potential to purified MSCs at the same cell dose. Post hoc analysis revealed that the presence of CD90+, CD105+, and CD166+ correlated strongly with in vivo bone formation scores (r(2) > .95). These results demonstrate that this bioreactor system can be used to generate, in a single step, a population of progenitor cells with potent osteogenic potential. Disclosure of potential conflicts of interest is found at the end of this article. PMID:17585167

  17. Phenotypic differences in white-tailed deer antlerogenic progenitor cells and marrow-derived mesenchymal stromal cells.

    PubMed

    Daley, Ethan L H; Alford, Andrea I; Miller, Joshua D; Goldstein, Steven A

    2014-05-01

    Deer antlers are bony appendages that are annually cast and rapidly regrown in a seasonal process coupled to the reproductive cycle. Due to the uniqueness of this process among mammals, we reasoned that a fundamental characterization of antler progenitor cell behavior may provide insights that could lead to improved strategies for promoting bone repair. In this study, we investigated whether white-tailed deer antlerogenic progenitor cells (APC) conform to basic criteria defining mesenchymal stromal cells (MSC). In addition, we tested the effects of the artificial glucocorticoid dexamethasone (DEX) on osteogenic and chondrogenic differentiation as well as the degree of apoptosis during the latter. Comparisons were made to animal-matched marrow-derived MSC. APC and MSC generated similar numbers of colonies. APC cultures expanded less rapidly overall but experienced population recovery at later time points. In contrast to MSC, APC did not display adipogenic in vitro differentiation capacity. Under osteogenic culture conditions, APC and MSC exhibited different patterns of alkaline phosphatase activity over time. DEX increased APC alkaline phosphatase activity only initially but consistently led to decreased activity in MSC. APC and MSC in osteogenic culture underwent different time and DEX-dependent patterns of mineralization, yet APC and MSC achieved similar levels of mineral accrual in an ectopic ossicle model. During chondrogenic differentiation, APC exhibited high levels of apoptosis without a reduction in cell density. DEX decreased proteoglycan production and increased apoptosis in chondrogenic APC cultures but had the opposite effects in MSC. Our results suggest that APC and MSC proliferation and differentiation differ in their dependence on time, factors, and milieu. Antler tip APC may be more lineage-restricted osteo/chondroprogenitors with distinctly different responses to apoptotic and glucocorticoid stimuli. PMID:24313802

  18. Bone marrow-derived mesenchymal cells feature selective migration behavior on submicro- and nano-dimensional multi-patterned substrates.

    PubMed

    Klymov, Alexey; Bronkhorst, Ewald M; Te Riet, Joost; Jansen, John A; Walboomers, X Frank

    2015-04-01

    This study investigated whether cells have an intrinsic ability to recognize nanopatterns, which could lead to their accumulation or diminution on a biomaterial. A multi-patterned "biochip" was made, containing 36 differently designed surfaces, including squares and grooves varying in feature sizes between 10 and 1000 nm. The grooved patterns could additionally be subdivided into three groups having ridge to groove ratios of 1:1, 1:3 and 3:1. These substrates were used for culture of rat bone marrow derived mesenchymal cells. In time cells should accumulate on patterns of preference, while migrating away from patterns of disfavor. A regression analysis model was designed for the analysis of the obtained data. Results showed that strong differences existed between the tested patterns regarding the cellular affinity. All sizes of squares showed strong cell-repelling capacity, with the biggest sized squares displaying up to 40% less cells compared to the smooth surface. Among the nano-grooved patterns cell repelling was seen for the grooves with the ridge to groove ratio of 1:3, while grooves with the ridge to groove ratio of 3:1 partially showed cell attraction. Such effects were shown to be based on selective migration rather than proliferation. In conclusion, the use of a multi-patterned biochip setup allows for enhanced evaluation of cell behavior, as compared to uniformly patterned setups. Cells exhibit the ability to actively avoid or migrate to surfaces featuring certain topographies on nanometric scale. Such phenomena may be utilized for the development of biomaterials in regenerative medicine. PMID:25644453

  19. Gamma irradiation preserves immunosuppressive potential and inhibits clonogenic capacity of human bone marrow-derived mesenchymal stromal cells

    PubMed Central

    de Andrade, Ana Valéria Gouveia; Riewaldt, Julia; Wehner, Rebekka; Schmitz, Marc; Odendahl, Marcus; Bornhäuser, Martin; Tonn, Torsten

    2014-01-01

    Mesenchymal stromal cells (MSCs) are promising candidates for the treatment of graft-versus-host and autoimmune diseases. Here, by virtue of their immunosuppressive effects, they are discussed to exhibit inhibitory actions on various immune effector cells, including T lymphocytes that promote the underlying pathology. While it becomes apparent that MSCs exhibit their therapeutic effect in a transient manner, they are usually transplanted from third party donors into heavily immunocompromised patients. However, little is known about potential late complications of persisting third party MSCs in these patients. We therefore analysed the effect of gamma irradiation on the potency and proliferation of MSCs to elucidate an irradiation dose, which would allow inhibition of MSC proliferation while at the same time preserving their immunosuppressive function. Bone marrow-derived MSCs (BM-MSCs) were gamma-irradiated at increasing doses of 5, 10 and 30 Gy and subsequently assessed by colony formation unit (CFU)-assay, Annexin V-staining and in a mixed lymphocyte reaction, to assess colony growth, apoptosis and the immunosuppressive capacity, respectively. Complete loss of proliferative capacity measured by colony formation was observed after irradiation with a dose equal to or greater than 10 Gy. No significant decrease of viable cells was detected, as compared to non-irradiated BM-MSCs. Notably, irradiated BM-MSCs remained highly immunosuppressive in vitro for at least 5 days after irradiation. Gamma irradiation does not impair the immunosuppressive capacity of BM-MSCs in vitro and thus might increase the safety of MSC-based cell products in clinical applications. PMID:24655362

  20. Hypoxia/Reoxygenation-Preconditioned Human Bone Marrow-Derived Mesenchymal Stromal Cells Rescue Ischemic Rat Cortical Neurons by Enhancing Trophic Factor Release.

    PubMed

    Kim, Young Seo; Noh, Min Young; Cho, Kyung Ah; Kim, Hyemi; Kwon, Min-Soo; Kim, Kyung Suk; Kim, Juhan; Koh, Seong-Ho; Kim, Seung Hyun

    2015-08-01

    Bone marrow-derived mesenchymal stromal cells (BM-MSCs) represent a promising tool for stem cell-based therapies. However, the majority of MSCs fail to reach the injury site and have only minimal therapeutic effect. In this study, we assessed whether hypoxia/reoxygenation (H/R) preconditioning of human BM-MSCs could increase their functional capacity and beneficial effect on ischemic rat cortical neurons. Human BM-MSCs were cultured under hypoxia (1% O2) and with long-term reoxygenation for various times to identify the optimal conditions for increasing their viability and proliferation. The effects of H/R preconditioning on the BM-MSCs were assessed by analyzing the expression of prosurvival genes, trophic factors, and cell migration assays. The functionally improved BM-MSCs were cocultured with ischemic rat cortical neurons to compare with normoxic cultured BM-MSCs. Although the cell viability and proliferation of BM-MSCs were reduced after 1 day of hypoxic culture (1% O2), when this was followed by 5-day reoxygenation, the BM-MSCs recovered and multiplied extensively. The immunophenotype and trilineage differentiation of BM-MSCs were also maintained under this H/R preconditioning. In addition, the preconditioning enhanced the expression of prosurvival genes, the messenger RNA (mRNA) levels of various trophic factors and migration capacity. Finally, coculture with the H/R-preconditioned BM-MSCs promoted the survival of ischemic rat cortical neurons. H/R preconditioning of BM-MSCs increases prosurvival signals, trophic factor release, and cell migration and appears to increase their ability to rescue ischemic cortical neurons. This optimized H/R preconditioning procedure could provide the basis for a new strategy for stem cell therapy in ischemic stroke patients. PMID:25288154

  1. Anti-inflammatory/tissue repair macrophages enhance the cartilage-forming capacity of human bone marrow-derived mesenchymal stromal cells.

    PubMed

    Sesia, Sergio B; Duhr, Ralph; Medeiros da Cunha, Carolina; Todorov, Atanas; Schaeren, Stefan; Padovan, Elisabetta; Spagnoli, Giulio; Martin, Ivan; Barbero, Andrea

    2015-06-01

    Macrophages are key players in healing processes. However, little is known on their capacity to modulate the differentiation potential of mesenchymal stem/stromal cells (MSC). Here we investigated whether macrophages (Mf) with, respectively, pro-inflammatory and tissue-remodeling traits differentially modulate chondrogenesis of bone marrow derived-MSC (BM-MSC). We demonstrated that coculture in collagen scaffolds of BM-MSC with Mf derived from monocytes polarized with M-CSF (M-Mf), but not with GM-CSF (GM-Mf) resulted in significantly higher glycosaminoglycan (GAG) content than what would be expected from an equal number of BM-MSC alone (defined as chondro-induction). Moreover, type II collagen was expressed at significantly higher levels in BM-MSC/M-Mf as compared to BM-MSC/GM-Mf constructs, while type X collagen expression was unaffected. In order to understand the possible cellular mechanism accounting for chondro-induction, developing monoculture and coculture tissues were digested and the properties of the isolated BM-MSC analysed. We observed that as compared to monocultures, in coculture with M-Mf, BM-MSC decreased less markedly in number and exhibited higher clonogenic and chondrogenic capacity. Despite their chondro-inductive effect in vitro, M-Mf did not modulate the cartilage tissue maturation in subcutaneous pockets of nude mice, as evidenced by similar accumulation of type X collagen and calcified tissue. Our results demonstrate that coculture of BM-MSC with M-Mf results in synergistic cartilage tissue formation in vitro. Such effect seems to result from the survival of BM-MSC with high chondrogenic capacity. Studies in an orthotopic in vivo model are necessary to assess the clinical relevance of our findings in the context of cartilage repair. PMID:25413299

  2. Using the quantum cell expansion system for the automated expansion of clinical-grade bone marrow-derived human mesenchymal stromal cells.

    PubMed

    Martin-Manso, Gema; Hanley, Patrick J

    2015-01-01

    Bone marrow-derived human mesenchymal stromal cells (hMSCs) constitute a promising therapeutic approach. However, the extremely low frequency of hMSCs in bone marrow makes the translation of these regulatory cells to clinical therapies difficult for large patient populations. Here, we describe a good manufacturing practices-compliant procedure for the expansion of hMSCs using the Quantum Cell Expansion System. This closed and automated system allows the large-scale expansion of hMSCs while maintaining their multipotency, immunophenotype, morphology, and karyotype. PMID:25523809

  3. Bone marrow-derived CD11b+Jagged2+ cells promote epithelial-to-mesenchymal transition and metastasization in colorectal cancer.

    PubMed

    Caiado, Francisco; Carvalho, Tânia; Rosa, Isadora; Remédio, Leonor; Costa, Ana; Matos, João; Heissig, Beate; Yagita, Hideo; Hattori, Koichi; da Silva, João Pereira; Fidalgo, Paulo; Pereira, António Dias; Dias, Sérgio

    2013-07-15

    Timely detection of colorectal cancer metastases may permit improvements in their clinical management. Here, we investigated a putative role for bone marrow-derived cells in the induction of epithelial-to-mesenchymal transition (EMT) as a marker for onset of metastasis. In ectopic and orthotopic mouse models of colorectal cancer, bone marrow-derived CD11b(Itgam)(+)Jagged2 (Jag2)(+) cells infiltrated primary tumors and surrounded tumor cells that exhibited diminished expression of E-cadherin and increased expression of vimentin, 2 hallmarks of EMT. In vitro coculture experiments showed that the bone marrow-derived CD11b(+)Jag2(+) cells induced EMT through a Notch-dependent pathway. Using neutralizing antibodies, we imposed a blockade on CD11b(+) cells' recruitment to tumors, which decreased the tumor-infiltrating CD11b(+)Jag2(+) cell population of interest, decreasing tumor growth, restoring E-cadherin expression, and delaying EMT. In support of these results, we found that peripheral blood levels of CD11b(+)Jag2(+) cells in mouse models of colorectal cancer and in a cohort of untreated patients with colorectal cancer were indicative of metastatic disease. In patients with colorectal cancer, the presence of circulating CD11b(+)Jag2(+) cells was accompanied by loss of E-cadherin in the corresponding patient tumors. Taken together, our results show that bone marrow-derived CD11b(+)Jag2(+) cells, which infiltrate primary colorectal tumors, are sufficient to induce EMT in tumor cells, thereby triggering onset of metastasis. Furthermore, they argue that quantifying circulating CD11b(+)Jag2(+) cells in patients may offer an indicator of colorectal cancer progression to metastatic levels of the disease. PMID:23722542

  4. Probing the interaction forces of prostate cancer cells with collagen I and bone marrow derived stem cells on the single cell level.

    PubMed

    Sariisik, Ediz; Docheva, Denitsa; Padula, Daniela; Popov, Cvetan; Opfer, Jan; Schieker, Matthias; Clausen-Schaumann, Hauke; Benoit, Martin

    2013-01-01

    Adhesion of metastasizing prostate carcinoma cells was quantified for two carcinoma model cell lines LNCaP (lymph node-specific) and PC3 (bone marrow-specific). By time-lapse microscopy and force spectroscopy we found PC3 cells to preferentially adhere to bone marrow-derived mesenchymal stem cells (SCP1 cell line). Using atomic force microscopy (AFM) based force spectroscopy, the mechanical pattern of the adhesion to SCP1 cells was characterized for both prostate cancer cell lines and compared to a substrate consisting of pure collagen type I. PC3 cells dissipated more energy (27.6 aJ) during the forced de-adhesion AFM experiments and showed significantly more adhesive and stronger bonds compared to LNCaP cells (20.1 aJ). The characteristic signatures of the detachment force traces revealed that, in contrast to the LNCaP cells, PC3 cells seem to utilize their filopodia in addition to establish adhesive bonds. Taken together, our study clearly demonstrates that PC3 cells have a superior adhesive affinity to bone marrow mesenchymal stem cells, compared to LNCaP. Semi-quantitative PCR on both prostate carcinoma cell lines revealed the expression of two Col-I binding integrin receptors, α1β1 and α2β1 in PC3 cells, suggesting their possible involvement in the specific interaction to the substrates. Further understanding of the exact mechanisms behind this phenomenon might lead to optimized therapeutic applications targeting the metastatic behavior of certain prostate cancer cells towards bone tissue. PMID:23472100

  5. Probing the Interaction Forces of Prostate Cancer Cells with Collagen I and Bone Marrow Derived Stem Cells on the Single Cell Level

    PubMed Central

    Sariisik, Ediz; Docheva, Denitsa; Padula, Daniela; Popov, Cvetan; Opfer, Jan; Schieker, Matthias; Clausen-Schaumann, Hauke; Benoit, Martin

    2013-01-01

    Adhesion of metastasizing prostate carcinoma cells was quantified for two carcinoma model cell lines LNCaP (lymph node-specific) and PC3 (bone marrow-specific). By time-lapse microscopy and force spectroscopy we found PC3 cells to preferentially adhere to bone marrow-derived mesenchymal stem cells (SCP1 cell line). Using atomic force microscopy (AFM) based force spectroscopy, the mechanical pattern of the adhesion to SCP1 cells was characterized for both prostate cancer cell lines and compared to a substrate consisting of pure collagen type I. PC3 cells dissipated more energy (27.6 aJ) during the forced de-adhesion AFM experiments and showed significantly more adhesive and stronger bonds compared to LNCaP cells (20.1 aJ). The characteristic signatures of the detachment force traces revealed that, in contrast to the LNCaP cells, PC3 cells seem to utilize their filopodia in addition to establish adhesive bonds. Taken together, our study clearly demonstrates that PC3 cells have a superior adhesive affinity to bone marrow mesenchymal stem cells, compared to LNCaP. Semi-quantitative PCR on both prostate carcinoma cell lines revealed the expression of two Col-I binding integrin receptors, α1β1 and α2β1 in PC3 cells, suggesting their possible involvement in the specific interaction to the substrates. Further understanding of the exact mechanisms behind this phenomenon might lead to optimized therapeutic applications targeting the metastatic behavior of certain prostate cancer cells towards bone tissue. PMID:23472100

  6. Adult Bone Marrow-Derived Stem Cells in Muscle Connective Tissue and Satellite Cell Niches

    PubMed Central

    Dreyfus, Patrick A.; Chretien, Fabrice; Chazaud, Bénédicte; Kirova, Youlia; Caramelle, Philippe; Garcia, Luis; Butler-Browne, Gillian; Gherardi, Romain K.

    2004-01-01

    Skeletal muscle includes satellite cells, which reside beneath the muscle fiber basal lamina and mainly represent committed myogenic precursor cells, and multipotent stem cells of unknown origin that are present in muscle connective tissue, express the stem cell markers Sca-1 and CD34, and can differentiate into different cell types. We tracked bone marrow (BM)-derived stem cells in both muscle connective tissue and satellite cell niches of irradiated mice transplanted with green fluorescent protein (GFP)-expressing BM cells. An increasing number of GFP+ mononucleated cells, located both inside and outside of the muscle fiber basal lamina, were observed 1, 3, and 6 months after transplantation. Sublaminal cells expressed unambiguous satellite cell markers (M-cadherin, Pax7, NCAM) and fused into scattered GFP+ muscle fibers. In muscle connective tissue there were GFP+ cells located close to blood vessels that expressed the ScaI or CD34 stem-cell antigens. The rate of settlement of extra- and intralaminal compartments by BM-derived cells was compatible with the view that extralaminal cells constitute a reservoir of satellite cells. We conclude that both muscle satellite cells and stem cell marker-expressing cells located in muscle connective tissue can derive from BM in adulthood. PMID:14982831

  7. The role of Hibiscus sabdariffa L. (Roselle) in maintenance of ex vivo murine bone marrow-derived hematopoietic stem cells.

    PubMed

    Abdul Hamid, Zariyantey; Lin Lin, Winnie Hii; Abdalla, Basma Jibril; Bee Yuen, Ong; Latif, Elda Surhaida; Mohamed, Jamaludin; Rajab, Nor Fadilah; Paik Wah, Chow; Wak Harto, Muhd Khairul Akmal; Budin, Siti Balkis

    2014-01-01

    Hematopoietic stem cells- (HSCs-) based therapy requires ex vivo expansion of HSCs prior to therapeutic use. However, ex vivo culture was reported to promote excessive production of reactive oxygen species (ROS), exposing HSCs to oxidative damage. Efforts to overcome this limitation include the use of antioxidants. In this study, the role of Hibiscus sabdariffa L. (Roselle) in maintenance of cultured murine bone marrow-derived HSCs was investigated. Aqueous extract of Roselle was added at varying concentrations (0-1000 ng/mL) for 24 hours to the freshly isolated murine bone marrow cells (BMCs) cultures. Effects of Roselle on cell viability, reactive oxygen species (ROS) production, glutathione (GSH) level, superoxide dismutase (SOD) activity, and DNA damage were investigated. Roselle enhanced the survival (P < 0.05) of BMCs at 500 and 1000 ng/mL, increased survival of Sca-1(+) cells (HSCs) at 500 ng/mL, and maintained HSCs phenotype as shown from nonremarkable changes of surface marker antigen (Sca-1) expression in all experimental groups. Roselle increased (P < 0.05) the GSH level and SOD activity but the level of reactive oxygen species (ROS) was unaffected. Moreover, Roselle showed significant cellular genoprotective potency against H2O2-induced DNA damage. Conclusively, Roselle shows novel property as potential supplement and genoprotectant against oxidative damage to cultured HSCs. PMID:25405216

  8. Imaging and Tracking of Bone Marrow-Derived Immune and Stem Cells

    PubMed Central

    Zhao, Youbo; Bower, Andrew J.; Graf, Benedikt W.; Boppart, Marni D.; Boppart, Stephen A.

    2014-01-01

    Bone marrow (BM)-derived stem and immune cells play critical roles in maintaining the health, regeneration, and repair of many tissues. Given their important functions in tissue regeneration and therapy, tracking the dynamic behaviors of BM-derived cells has been a long-standing research goal of both biologists and engineers. Because of the complex cellular-level processes involved, real-time imaging technologies that have sufficient spatial and temporal resolution to visualize them are needed. In addition, in order to track cellular dynamics, special attention is needed to account for changes in the microenvironment where the cells reside, for example, tissue contraction, stretching, development, etc. In this chapter, we introduce methods for real-time imaging and longitudinal tracking of BM-derived immune and stem cells in in vivo three-dimensional (3-D) tissue environments with an integrated optical microscope. The integrated microscope combines multiple imaging functions derived from optical coherence tomography (OCT) and multiphoton microscopy (MPM), including optical coherence microscopy (OCM), micro-vasculature imaging, two-photon excited fluorescence (TPEF), and second harmonic generation (SHG) microscopy. Short- and long-term tracking of the dynamic behavior of BM-derived cells involved in cutaneous wound healing and skin grafting in green fluorescent protein (GFP) BM-transplanted mice is demonstrated. Methods and algorithms for nonrigid registration of time-lapse images are introduced, which allows for long-term tracking of cell dynamics over several months. PMID:23737096

  9. Mesenchymal stem cells.

    PubMed

    Ding, Dah-Ching; Shyu, Woei-Cherng; Lin, Shinn-Zong

    2011-01-01

    Stem cells have two features: the ability to differentiate along different lineages and the ability of self-renewal. Two major types of stem cells have been described, namely, embryonic stem cells and adult stem cells. Embryonic stem cells (ESC) are obtained from the inner cell mass of the blastocyst and are associated with tumorigenesis, and the use of human ESCs involves ethical and legal considerations. The use of adult mesenchymal stem cells is less problematic with regard to these issues. Mesenchymal stem cells (MSCs) are stromal cells that have the ability to self-renew and also exhibit multilineage differentiation. MSCs can be isolated from a variety of tissues, such as umbilical cord, endometrial polyps, menses blood, bone marrow, adipose tissue, etc. This is because the ease of harvest and quantity obtained make these sources most practical for experimental and possible clinical applications. Recently, MSCs have been found in new sources, such as menstrual blood and endometrium. There are likely more sources of MSCs waiting to be discovered, and MSCs may be a good candidate for future experimental or clinical applications. One of the major challenges is to elucidate the mechanisms of differentiation, mobilization, and homing of MSCs, which are highly complex. The multipotent properties of MSCs make them an attractive choice for possible development of clinical applications. Future studies should explore the role of MSCs in differentiation, transplantation, and immune response in various diseases. PMID:21396235

  10. Bone marrow derived stem cells for the treatment of end-stage liver disease.

    PubMed

    Margini, Cristina; Vukotic, Ranka; Brodosi, Lucia; Bernardi, Mauro; Andreone, Pietro

    2014-07-21

    End-stage disease due to liver cirrhosis is an important cause of death worldwide. Cirrhosis results from progressive, extensive fibrosis and impaired hepatocyte regeneration. The only curative treatment is liver transplantation, but due to the several limitations of this procedure, the interest in alternative therapeutic strategies is increasing. In particular, the potential of bone marrow stem cell (BMSC) therapy in cirrhosis has been explored in different trials. In this article, we evaluate the results of 18 prospective clinical trials, and we provide a descriptive overview of recent advances in the research on hepatic regenerative medicine. The main message from the currently available data in the literature is that BMSC therapy is extremely promising in the context of liver cirrhosis. However, its application should be further explored in randomized, controlled trials with large cohorts and long follow-ups. PMID:25083082

  11. Bone reconstruction of large defects using bone marrow derived autologous stem cells.

    PubMed

    Lucarelli, Enrico; Donati, Davide; Cenacchi, Annarita; Fornasari, Pier Maria

    2004-04-01

    Bone is a tissue that has the ability to heal itself when fractured. Occasionally, a critical defect can be formed when part of the bone is lost or excised, in this case the bone fails to heal and requires bone reconstruction to prevent a non-union defect. Autogenous cancellous bone is the current gold standard treatment in bone loss. Because the amount of autogenous cancellous bone that can be harvested is limited, the expanding need for bone reconstruction is paired by the growth of interest in the discipline of tissue engineering. Labs worldwide are working to provide the right carrier and the right set of cells that, once retransplanted, will ensure bone repair. Several investigators have focused their attention on a subset of autologous non-hematopoietic stem/progenitor cells contained in the adult bone marrow stroma, referred to as stromal stem cells (SSC), as the appropriate cells to be transplanted. The use of autologous cells is facilitated by less stringent ethical and regulatory issues and does not require the patient to be immunologically suppressed. In pre-clinical and clinical protocols of critical defects in which SSC are employed, two approaches are mainly used: in the first, SSC are derived from bone marrow and directly introduced at the lesion site, in the second, SSC are derived from several sites and are expanded ex vivo before being implanted. Both approaches, equally correct in principle, will have to demonstrate, with definitive evidence of their efficacy, their capability of solving a critical clinical problem such as non-union. In this report we outline the difficulties of working with SSC. PMID:15062758

  12. Clumping and Viability of Bone Marrow Derived Mesenchymal Stromal Cells under Different Preparation Procedures: A Flow Cytometry-Based In Vitro Study

    PubMed Central

    Cui, Li-li; Kinnunen, Tuure; Boltze, Johannes; Nystedt, Johanna

    2016-01-01

    Complications of microocclusions have been reported after intra-arterial delivery of mesenchymal stromal cells. Hence, quantification and efficient limitation of cell clumps in suspension before transplantation is important to reduce the risk. We used a flow cytometry-based pulse-width assay to assess the effects of different cell suspension concentrations (0.2–2.0 × 106/mL), storage solutions (complete growth medium, Dulbecco's phosphate-buffered saline, and normal saline), storage time in suspension (0–9 h), and freeze-thawing procedure on the clumping of rat bone marrow derived mesenchymal stromal cells (BMMSCs) and also evaluated cell viability at the same time. Surprisingly, increasing the cell concentration did not result in more cell clumps in vitro. Freshly harvested (fresh) cells in normal saline had significantly fewer cell clumps and also displayed high viability (>90%). A time-dependent reduction in viability was observed for cells in all three storage solutions, without any significant change in the clumping tendency except for cells in medium. Fresh cells were more viable than their frozen-thawed counterparts, and fresh cells in normal saline had fewer cell clumps. In conclusion, cell clumping and viability could be affected by different cell preparation procedures, and quantification of cell clumping can be conducted using the flow cytometry-based pulse-width assay before intra-arterial cell delivery. PMID:27022399

  13. [Mesenchymal stem cells. A review.].

    PubMed

    Sigurjónsson, O E; Guðmundsson, K O; Guðmundsson, S

    2001-01-01

    The bone marrow contains various types of stem cells. Among them are hematopoietic stem cells, which are the precursors of all blood cells, and mesenchymal stem cells. Mesenchymal stem cells have recently received a lot of attention in biological research because of their capability to self renewal, to expand and transdifferentiate into many different cell types; bone cells, adipocytes, chondrocytes, tendocytes, neural cells and stromal cells of the bone marrow. Mesenchymal stem cells can be cultured in vitro although their differentiation potential is not yet fully understood. Several experiments have been conducted in animal models where mesenchymal stem cells have been transplanted in order to enhance hematopoiesis or to facilitate the repair of mesenchymal tissue. Similar experiments are being conducted in humans. Mesenchymal stem cells are believed to be able to enhance hematopoietic stem cells transplantation by rebuilding the bone marrow microenvironment which is damaged after radiation- and/or chemotherapy. Mesenchymal stem cells are promising as vehicles for gene transfer and therapy. It may prove possible to tranduce them with a gene coding for a defective protein i.e. collagen I in osteogenesis imperfecta. The cells could then be expanded ex vivo and transplanted to the patients where they home to the bone marrow, differentiate and produce the intact protein. Future medicine will probably involve mesenchymal stem cells in various treatment settings. PMID:17018999

  14. An axial distribution of seeding, proliferation, and osteogenic differentiation of MC3T3-E1 cells and rat bone marrow-derived mesenchymal stem cells across a 3D Thai silk fibroin/gelatin/hydroxyapatite scaffold in a perfusion bioreactor.

    PubMed

    Sinlapabodin, Salita; Amornsudthiwat, Phakdee; Damrongsakkul, Siriporn; Kanokpanont, Sorada

    2016-01-01

    In cell culture, a perfusion bioreactor provides effective transportation of nutrients, oxygen, and waste removal to and from the core of the scaffold. In addition, it provides mechanical stimuli for enhancing osteogenic differentiation. In this study, we used an axial distribution of cell numbers, alkaline phosphatase (ALP) enzyme activity, and calcium content across 4 cross-sections of 10mm thick scaffold, made of Thai silk fibroin (SF)/gelatin (G)/hydroxyapatite (HA), as a tool to evaluate the suitable perfusion flow rate. These evaluations cover all cellular developmental phases starting from seeding, to proliferation, and later osteogenic differentiation. Mouse pre-osteoblastic MC3T3-E1 cell lines were used as a cell model during seeding and proliferation. The bioreactor seeded scaffold provided more uniform cell distribution across the scaffold compared to centrifugal and agitation seeding, while the overall number of adhered cells from bioreactor seeding was slightly lower than agitation seeding. The dynamic culture using 1 ml/min perfusion flow rate (initial shear stress of 0.1 dyn/cm(2)) enabled statistically higher MC3T3-E1 proliferation, ALP activity, and calcium deposition than those observed in the static-culturing condition. However, the perfusion flow rate of 1 ml/min seemed not to be enough for enhancing ALP expression across all sections of the scaffold. Rat bone marrow derived stromal cells (rMSC) were used in the detachment test and osteogenic differentiation. It was found that perfusion flow rate of 5 ml/min caused statistically higher cell detachment than that of 1 and 3 ml/min. The perfusion flow rate of 3 ml/min gave the highest rMSC osteogenic differentiation on a SF/G/HA scaffold than other flow rates, as observed from the significantly highest number of ALP enzyme activity and the calcium content without any significant cell growth. In addition, all of these parameters were evenly distributed across all scaffold sections. PMID:26478392

  15. Effect of Metformin on Viability, Morphology, and Ultrastructure of Mouse Bone Marrow-Derived Multipotent Mesenchymal Stromal Cells and Balb/3T3 Embryonic Fibroblast Cell Line.

    PubMed

    Śmieszek, Agnieszka; Czyrek, Aleksandra; Basinska, Katarzyna; Trynda, Justyna; Skaradzińska, Aneta; Siudzińska, Anna; Marędziak, Monika; Marycz, Krzysztof

    2015-01-01

    Metformin, a popular drug used to treat diabetes, has recently gained attention as a potentially useful therapeutic agent for treating cancer. In our research metformin was added to in vitro cultures of bone marrow-derived multipotent mesenchymal stromal cells (BMSCs) and Balb/3T3 fibroblast at concentration of 1 mM, 5 mM, and 10 mM. Obtained results indicated that metformin negatively affected proliferation activity of investigated cells. The drug triggered the formation of autophagosomes and apoptotic bodies in all tested cultures. Additionally, we focused on determination of expression of genes involved in insulin-like growth factor 2 (IGF2) signaling pathway. The most striking finding was that the mRNA level of IGF2 was constant in both BMSCs and Balb/3T3. Further, the analysis of IGF2 concentration in cell supernatants showed that it decreased in BMSC cultures after 5 and 10 mM metformin treatments. In case of Balb/3T3 the concentration of IGF2 in culture supernatants decreased after 1 and 5 mM and increased after 10 mM of metformin. Our results suggest that metformin influences the cytophysiology of somatic cells in a dose- and time-dependent manner causing inhibition of proliferation and abnormalities of their morphology and ultrastructure. PMID:26064951

  16. Molecular Imaging for Comparison of Different Growth Factors on Bone Marrow-Derived Mesenchymal Stromal Cells' Survival and Proliferation In Vivo.

    PubMed

    Qiao, Hongyu; Zhang, Ran; Gao, Lina; Guo, Yanjie; Wang, Jinda; Zhang, Rongqing; Li, Xiujuan; Li, Congye; Chen, Yundai; Cao, Feng

    2016-01-01

    Introduction. Bone marrow-derived mesenchymal stromal cells (BMSCs) have emerged as promising cell candidates but with poor survival after transplantation. This study was designed to investigate the efficacy of VEGF, bFGF, and IGF-1 on BMSCs' viability and proliferation both in vivo and in vitro using bioluminescence imaging (BLI). Methods. BMSCs were isolated from β-actin-Fluc(+) transgenic FVB mice, which constitutively express firefly luciferase. Apoptosis was induced by hypoxia preconditioning for up to 24 h followed by flow cytometry and TUNEL assay. 10(6) BMSCs with/without growth factors were injected subcutaneously into wild type FVB mice's backs. Survival of BMSCs was longitudinally monitored using bioluminescence imaging (BLI) for 5 weeks. Protein expression of Akt, p-Akt, PARP, and caspase-3 was detected by Western blot. Results. Hypoxia-induced apoptosis was significantly attenuated by bFGF and IGF-1 compared with VEGF and control group in vitro (P < 0.05). When combined with matrigel, IGF-1 showed the most beneficial effects in protecting BMSCs from apoptosis in vivo. The phosphorylation of Akt had a higher ratio in the cells from IGF-1 group. Conclusion. IGF-1 could protect BMSCs from hypoxia-induced apoptosis through activation of p-Akt/Akt pathway. PMID:27419126

  17. Molecular Imaging for Comparison of Different Growth Factors on Bone Marrow-Derived Mesenchymal Stromal Cells' Survival and Proliferation In Vivo

    PubMed Central

    Qiao, Hongyu; Zhang, Ran; Gao, Lina; Guo, Yanjie; Wang, Jinda; Zhang, Rongqing; Li, Xiujuan; Li, Congye; Chen, Yundai; Cao, Feng

    2016-01-01

    Introduction. Bone marrow-derived mesenchymal stromal cells (BMSCs) have emerged as promising cell candidates but with poor survival after transplantation. This study was designed to investigate the efficacy of VEGF, bFGF, and IGF-1 on BMSCs' viability and proliferation both in vivo and in vitro using bioluminescence imaging (BLI). Methods. BMSCs were isolated from β-actin-Fluc+ transgenic FVB mice, which constitutively express firefly luciferase. Apoptosis was induced by hypoxia preconditioning for up to 24 h followed by flow cytometry and TUNEL assay. 106 BMSCs with/without growth factors were injected subcutaneously into wild type FVB mice's backs. Survival of BMSCs was longitudinally monitored using bioluminescence imaging (BLI) for 5 weeks. Protein expression of Akt, p-Akt, PARP, and caspase-3 was detected by Western blot. Results. Hypoxia-induced apoptosis was significantly attenuated by bFGF and IGF-1 compared with VEGF and control group in vitro (P < 0.05). When combined with matrigel, IGF-1 showed the most beneficial effects in protecting BMSCs from apoptosis in vivo. The phosphorylation of Akt had a higher ratio in the cells from IGF-1 group. Conclusion. IGF-1 could protect BMSCs from hypoxia-induced apoptosis through activation of p-Akt/Akt pathway. PMID:27419126

  18. Effect of Metformin on Viability, Morphology, and Ultrastructure of Mouse Bone Marrow-Derived Multipotent Mesenchymal Stromal Cells and Balb/3T3 Embryonic Fibroblast Cell Line

    PubMed Central

    Czyrek, Aleksandra; Basinska, Katarzyna; Trynda, Justyna; Skaradzińska, Aneta; Siudzińska, Anna; Marycz, Krzysztof

    2015-01-01

    Metformin, a popular drug used to treat diabetes, has recently gained attention as a potentially useful therapeutic agent for treating cancer. In our research metformin was added to in vitro cultures of bone marrow-derived multipotent mesenchymal stromal cells (BMSCs) and Balb/3T3 fibroblast at concentration of 1 mM, 5 mM, and 10 mM. Obtained results indicated that metformin negatively affected proliferation activity of investigated cells. The drug triggered the formation of autophagosomes and apoptotic bodies in all tested cultures. Additionally, we focused on determination of expression of genes involved in insulin-like growth factor 2 (IGF2) signaling pathway. The most striking finding was that the mRNA level of IGF2 was constant in both BMSCs and Balb/3T3. Further, the analysis of IGF2 concentration in cell supernatants showed that it decreased in BMSC cultures after 5 and 10 mM metformin treatments. In case of Balb/3T3 the concentration of IGF2 in culture supernatants decreased after 1 and 5 mM and increased after 10 mM of metformin. Our results suggest that metformin influences the cytophysiology of somatic cells in a dose- and time-dependent manner causing inhibition of proliferation and abnormalities of their morphology and ultrastructure. PMID:26064951

  19. Comparative miRNA-Based Fingerprinting Reveals Biological Differences in Human Olfactory Mucosa- and Bone-Marrow-Derived Mesenchymal Stromal Cells.

    PubMed

    Lindsay, Susan Louise; Johnstone, Steven Andrew; McGrath, Michael Anthony; Mallinson, David; Barnett, Susan Carol

    2016-05-10

    Previously we reported that nestin-positive human mesenchymal stromal cells (MSCs) derived from the olfactory mucosa (OM) enhanced CNS myelination in vitro to a greater extent than bone-marrow-derived MSCs (BM-MSCs). miRNA-based fingerprinting revealed the two MSCs were 64% homologous, with 26 miRNAs differentially expressed. We focused on miR-146a-5p and miR-140-5p due to their reported role in the regulation of chemokine production and myelination. The lower expression of miR-140-5p in OM-MSCs correlated with higher secretion of CXCL12 compared with BM-MSCs. Addition of CXCL12 and its pharmacological inhibitors to neural co-cultures supported these data. Studies on related miR-146a-5p targets demonstrated that OM-MSCs had lower levels of Toll-like receptors and secreted less pro-inflammatory cytokines, IL-6, IL-8, and CCL2. OM-MSCs polarized microglia to an anti-inflammatory phenotype, illustrating potential differences in their inflammatory response. Nestin-positive OM-MSCs could therefore offer a cell transplantation alternative for CNS repair, should these biological behaviors be translated in vivo. PMID:27117785

  20. Cell Therapy Using Bone Marrow-Derived Stem Cell Overexpressing BMP-7 for Degenerative Discs in a Rat Tail Disc Model.

    PubMed

    Liao, Jen-Chung

    2016-01-01

    Degenerative discs can cause low back pain. Cell-based transplantation or growth factors therapy have been suggested as a strategy to stimulate disc regeneration. Bone marrow-derived mesenchymal stem cells (BMDMSC) containing bone morphogenetic protein-7 (BMP-7) gene were constructed. We evaluated the effectiveness of these BMP-7 overexpressing cells on degenerative discs in rat tails. In vitro and in vivo studies were designed. In the first stage, the rats were divided into two group according to discs punctured by different needle gauges (18 gauge and 22 gauge). In the second stage, the ideal size of needle was used to induce rat tail disc degeneration. These animals are divided into three groups according to timing of treatment (zero-week, two-week, four-week). Each group was divided into three treating subgroups: control group, BMDMSC group, and Baculo-BMP-7-BMDMSC group. Each rat undergoes radiography examination every two weeks. After eight weeks, the discs were histologically examined with hematoxylin and eosin stain and Alcian blue stain. The 18-gauge group exhibited significant decrease in disc height index (%) than 22-gauge group at eight weeks at both Co6-7 (58.1% ± 2.8% vs. 63.7% ± 1.0%, p = 0.020) and Co8-9 discs (62.7% ± 2.8% vs. 62.8% ± 1.5%, p = 0.010). Baculo-BMP-7-BMDMSCs group showed significant difference in disc height index compared to the BMDMSCs group at both Co6-7 (93.7% ± 1.5% vs. 84.8% ± 1.0%, p = 0.011) and Co8-9 (86.0% ± 2.1% vs. 81.8% ± 1.7%, p = 0.012). In Baculo-BMP-7-BMDMSCs group, the zero-week treatment subgroup showed significant better in disc height index compared to two-week treatment group (p = 0.044), and four-week treatment group (p = 0.011). The zero-week treatment subgroup in Baculo-BMP-7-BMDMSCs group also had significant lower histology score than two-week treatment (4.3 vs. 5.7, p = 0.045) and four-week treatment (4.3 vs. 6.0, p = 0.031). In conclusion, Baculo-BMP-7-BMDMSC can slow down the progression of disc

  1. Cell Therapy Using Bone Marrow-Derived Stem Cell Overexpressing BMP-7 for Degenerative Discs in a Rat Tail Disc Model

    PubMed Central

    Liao, Jen-Chung

    2016-01-01

    Degenerative discs can cause low back pain. Cell-based transplantation or growth factors therapy have been suggested as a strategy to stimulate disc regeneration. Bone marrow-derived mesenchymal stem cells (BMDMSC) containing bone morphogenetic protein-7 (BMP-7) gene were constructed. We evaluated the effectiveness of these BMP-7 overexpressing cells on degenerative discs in rat tails. In vitro and in vivo studies were designed. In the first stage, the rats were divided into two group according to discs punctured by different needle gauges (18 gauge and 22 gauge). In the second stage, the ideal size of needle was used to induce rat tail disc degeneration. These animals are divided into three groups according to timing of treatment (zero-week, two-week, four-week). Each group was divided into three treating subgroups: control group, BMDMSC group, and Baculo-BMP-7-BMDMSC group. Each rat undergoes radiography examination every two weeks. After eight weeks, the discs were histologically examined with hematoxylin and eosin stain and Alcian blue stain. The 18-gauge group exhibited significant decrease in disc height index (%) than 22-gauge group at eight weeks at both Co6-7 (58.1% ± 2.8% vs. 63.7% ± 1.0%, p = 0.020) and Co8-9 discs (62.7% ± 2.8% vs. 62.8% ± 1.5%, p = 0.010). Baculo-BMP-7-BMDMSCs group showed significant difference in disc height index compared to the BMDMSCs group at both Co6-7 (93.7% ± 1.5% vs. 84.8% ± 1.0%, p = 0.011) and Co8-9 (86.0% ± 2.1% vs. 81.8% ± 1.7%, p = 0.012). In Baculo-BMP-7-BMDMSCs group, the zero-week treatment subgroup showed significant better in disc height index compared to two-week treatment group (p = 0.044), and four-week treatment group (p = 0.011). The zero-week treatment subgroup in Baculo-BMP-7-BMDMSCs group also had significant lower histology score than two-week treatment (4.3 vs. 5.7, p = 0.045) and four-week treatment (4.3 vs. 6.0, p = 0.031). In conclusion, Baculo-BMP-7-BMDMSC can slow down the progression of disc

  2. Pentosan polysulfate promotes proliferation and chondrogenic differentiation of adult human bone marrow-derived mesenchymal precursor cells

    PubMed Central

    2010-01-01

    Introduction This study was undertaken to determine whether the anti-osteoarthritis drug pentosan polysulfate (PPS) influenced mesenchymal precursor cell (MPC) proliferation and differentiation. Methods Human MPCs were maintained in monolayer, pellet or micromass cultures (MMC) for up to 10 days with PPS at concentrations of 0 to 20 μg/ml. MPC viability and proliferation was assessed using the WST-1 assay and 3H-thymidine incorporation into DNA, while apoptosis was monitored by flow cytometry. Proteoglycan (PG) biosynthesis was determined by 35SO42- incorporation and staining with Alcian blue. Proteoglycan and collagen type I and collagen type II deposition in pellet cultures was also examined by Toluidine blue and immunohistochemical staining, respectively. The production of hyaluronan (HA) by MPCs in MMC was assessed by ELISA. The relative outcome of PPS, HA, heparin or dextran sulfate (DS) on PG synthesis was compared in 5-day MMC. Gene expression of MPCs in 7-day and 10-day MMC was examined using real-time PCR. MPC differentiation was investigated by co-culturing with PPS in osteogenic or adipogenic inductive culture media for 28 days. Results Significant MPC proliferation was evident by day 3 at PPS concentrations of 1 to 5 μg/ml (P < 0.01). In the presence of 1 to 10 μg/ml PPS, a 38% reduction in IL-4/IFNγ-induced MPC apoptosis was observed. In 5-day MMC, 130% stimulation of PG synthesis occurred at 2.5 μg/ml PPS (P < 0.0001), while 5.0 μg/ml PPS achieved maximal stimulation in the 7-day and 10-day cultures (P < 0.05). HA and DS at ≥ 5 μg/ml inhibited PG synthesis (P < 0.05) in 5-day MMC. Collagen type II deposition by MMC was significant at ≥ 0.5 μg/ml PPS (P < 0.001 to 0.05). In MPC-PPS pellet cultures, more PG, collagen type II but less collagen type I was deposited than in controls. Real-time PCR results were consistent with the protein data. At 5 and 10 μg/ml PPS, MPC osteogenic differentiation was suppressed (P < 0.01). Conclusions This is

  3. Mesenchymal Stem Cells in Tissue Growth and Repair

    PubMed Central

    Kalinina, N.I.; Sysoeva, V.Yu.; Rubina, K.A.; Parfenova, Ye.V.; Tkachuk, V.A.

    2011-01-01

    It has been established in the recent several decades that stem cells play a crucial role in tissue renewal and regeneration. Mesenchymal stem cells (MSCs) are part of the most important population of adult stem cells. These cells have hereby been identified for the very first time and subsequently isolated from bone marrow stroma. Bone marrow-derived MSCs have been believed to play the role of a source of cells for the renewal and repair of connective tissues, including bone, cartilage and adipose tissues. Cells similar to bone marrow-derived MSCs have now been identified in all postnatal tissues. Data on the distribution and function of MSCsin vivocollected using novel approaches pertaining to the identification of MSCsin situ, to their isolation from tissues, and finally to the determination of their biological properties have enabled successful revision of the role of MSCs in various organs and tissues. This review summarizes our own, as well as others’, data concerning the role of MSCs in the regulation processes of tissue repair and regeneration. In our opinion, MSCs provide the connection between the blood-vascular, immune, endocrine, and nervous systems and tissue-specific stem cells in the body. PMID:22649702

  4. Evaluation of transport conditions for autologous bone marrow-derived mesenchymal stromal cells for therapeutic application in horses.

    PubMed

    Espina, Miguel; Jülke, Henriette; Brehm, Walter; Ribitsch, Iris; Winter, Karsten; Delling, Uta

    2016-01-01

    Background. Mesenchymal stromal cells (MSCs) are increasingly used for clinical applications in equine patients. For MSC isolation and expansion, a laboratory step is mandatory, after which the cells are sent back to the attending veterinarian. Preserving the biological properties of MSCs during this transport is paramount. The goal of the study was to compare transport-related parameters (transport container, media, temperature, time, cell concentration) that potentially influence characteristics of culture expanded equine MSCs. Methods. The study was arranged in three parts comparing (I) five different transport containers (cryotube, two types of plastic syringes, glass syringe, CellSeal), (II) seven different transport media, four temperatures (4 °C vs. room temperature; -20 °C vs. -80 °C), four time frames (24 h vs. 48 h; 48 h vs. 72 h), and (III) three MSC concentrations (5 × 10(6), 10 × 10(6), 20 × 10(6) MSC/ml). Cell viability (Trypan Blue exclusion; percent and total number viable cell), proliferation and trilineage differentiation capacity were assessed for each test condition. Further, the recovered volume of the suspension was determined in part I. Each condition was evaluated using samples of six horses (n = 6) and differentiation protocols were performed in duplicates. Results. In part I of the study, no significant differences in any of the parameters were found when comparing transport containers at room temperature. The glass syringe was selected for all subsequent evaluations (highest recoverable volume of cell suspension and cell viability). In part II, media, temperatures, or time frames had also no significant influence on cell viability, likely due to the large number of comparisons and small sample size. Highest cell viability was observed using autologous bone marrow supernatant as transport medium, and "transport" at 4 °C for 24 h (70.6% vs. control group 75.3%); this was not significant. Contrary, viability was unacceptably low (<40

  5. Evaluation of transport conditions for autologous bone marrow-derived mesenchymal stromal cells for therapeutic application in horses

    PubMed Central

    Espina, Miguel; Jülke, Henriette; Brehm, Walter; Ribitsch, Iris; Winter, Karsten

    2016-01-01

    Background. Mesenchymal stromal cells (MSCs) are increasingly used for clinical applications in equine patients. For MSC isolation and expansion, a laboratory step is mandatory, after which the cells are sent back to the attending veterinarian. Preserving the biological properties of MSCs during this transport is paramount. The goal of the study was to compare transport-related parameters (transport container, media, temperature, time, cell concentration) that potentially influence characteristics of culture expanded equine MSCs. Methods. The study was arranged in three parts comparing (I) five different transport containers (cryotube, two types of plastic syringes, glass syringe, CellSeal), (II) seven different transport media, four temperatures (4 °C vs. room temperature; −20 °C vs. −80 °C), four time frames (24 h vs. 48 h; 48 h vs. 72 h), and (III) three MSC concentrations (5 × 106, 10 × 106, 20 × 106 MSC/ml). Cell viability (Trypan Blue exclusion; percent and total number viable cell), proliferation and trilineage differentiation capacity were assessed for each test condition. Further, the recovered volume of the suspension was determined in part I. Each condition was evaluated using samples of six horses (n = 6) and differentiation protocols were performed in duplicates. Results. In part I of the study, no significant differences in any of the parameters were found when comparing transport containers at room temperature. The glass syringe was selected for all subsequent evaluations (highest recoverable volume of cell suspension and cell viability). In part II, media, temperatures, or time frames had also no significant influence on cell viability, likely due to the large number of comparisons and small sample size. Highest cell viability was observed using autologous bone marrow supernatant as transport medium, and “transport” at 4 °C for 24 h (70.6% vs. control group 75.3%); this was not significant. Contrary, viability was unacceptably low

  6. Osteogenic potential of sorted equine mesenchymal stem cell subpopulations

    PubMed Central

    Radtke, Catherine L.; Nino-Fong, Rodolfo; Rodriguez-Lecompte, Juan Carlos; Esparza Gonzalez, Blanca P.; Stryhn, Henrik; McDuffee, Laurie A.

    2015-01-01

    The objectives of this study were to use non-equilibrium gravitational field-flow fractionation (GrFFF), an immunotag-less method of sorting mesenchymal stem cells (MSCs), to sort equine muscle tissue-derived mesenchymal stem cells (MMSCs) and bone marrow-derived mesenchymal stem cells (BMSC) into subpopulations and to carry out assays in order to compare their osteogenic capabilities. Cells from 1 young adult horse were isolated from left semitendinosus muscle tissue and from bone marrow aspirates of the fourth and fifth sternebrae. Aliquots of 800 × 103 MSCs from each tissue source were sorted into 5 fractions using non-equilibrium GrFFF (GrFFF proprietary system). Pooled fractions were cultured and expanded for use in osteogenic assays, including flow cytometry, histochemistry, bone nodule assays, and real-time quantitative polymerase chain reaction (qPCR) for gene expression of osteocalcin (OCN), RUNX2, and osterix. Equine MMSCs and BMSCs were consistently sorted into 5 fractions that remained viable for use in further osteogenic assays. Statistical analysis confirmed strongly significant upregulation of OCN, RUNX2, and osterix for the BMSC fraction 4 with P < 0.00001. Flow cytometry revealed different cell size and granularity for BMSC fraction 4 and MMSC fraction 2 compared to unsorted controls and other fractions. Histochemisty and bone nodule assays revealed positive staining nodules without differences in average nodule area, perimeter, or stain intensity between tissues or fractions. As there are different subpopulations of MSCs with different osteogenic capacities within equine muscle- and bone marrow-derived sources, these differences must be taken into account when using equine stem cell therapy to induce bone healing in veterinary medicine. PMID:25852225

  7. Effects of in vitro chondrogenic priming time of bone-marrow-derived mesenchymal stromal cells on in vivo endochondral bone formation.

    PubMed

    Yang, Wanxun; Both, Sanne K; van Osch, Gerjo J V M; Wang, Yining; Jansen, John A; Yang, Fang

    2015-02-01

    Recapitulation of endochondral ossification leads to a new concept of bone tissue engineering via a cartilage intermediate as an osteoinductive template. In this study, we aimed to investigate the influence of in vitro chondrogenic priming time for the creation of cartilage template on the in vivo endochondral bone formation both qualitatively and quantitatively. To this end, rat bone-marrow-derived mesenchymal stromal cells (MSCs) were seeded onto two scaffolds with distinguished features: a fibrous poly(lactic-co-glycolic acid)/poly(ε-caprolactone) electrospun scaffold (PLGA/PCL) and a porous hydroxyapatite/tricalcium phosphate composite (HA/TCP). The constructs were then chondrogenically differentiated for 2, 3 and 4 weeks in vitro, followed by subcutaneous implantation in vivo for up to 8 weeks. A longer chondrogenic priming time resulted in a significantly increased amount and homogeneous deposition of the cartilage matrix on both the PLGA/PCL and HA/TCP scaffolds in vitro. In vivo, all implanted constructs gave rise to endochondral bone formation, whereas the bone volume was not affected by the length of priming time. An unpolarized woven bone-like structure, with significant amounts of cartilage remaining, was generated in fibrous PLGA/PCL scaffolds, while porous HA/TCP scaffolds supported progressive lamellar-like bone formation with mature bone marrow development. These data suggest that, by utilizing a chondrogenically differentiated MSC-scaffold construct as cartilage template, 2 weeks of in vitro priming time is sufficient to generate a substantial amount of vascularized endochondral bone in vivo. The structure of the bone depends on the chemical and structural cues provided by the scaffold design. PMID:25463490

  8. Establishment of a Cytocompatible Cell-Free Intervertebral Disc Matrix for Chondrogenesis with Human Bone Marrow-Derived Mesenchymal Stromal Cells.

    PubMed

    Huang, Zhao; Kohl, Benjamin; Kokozidou, Maria; Arens, Stephan; Schulze-Tanzil, Gundula

    2016-01-01

    Tissue-engineered intervertebral discs (IVDs) utilizing decellularized extracellular matrix (ECM) could be an option for the reconstruction of impaired IVDs due to degeneration or injury. The objective of this study was to prepare a cell-free decellularized human IVD scaffold and to compare neotissue formation in response to recellularization with human IVD cells (hIVDCs) or human bone marrow-derived (hBM) mesenchymal stromal cells (MSCs). IVDs were decellularized via freeze-thaw cycles, detergents and trypsin. Histological staining was performed to monitor cell removal and glycosaminoglycan (GAG) removal. The decellularized IVD was preconditioned using bovine serum albumin and fetal bovine serum before its cytocompatibility for dynamically cultured hBM-MSCs (chondrogenically induced or not) and hIVDCs was compared after 14 days. In addition, DNA, total collagen and GAG contents were assessed. The decellularization protocol achieved maximal cell removal, with only few remaining cell nuclei compared with native tissue, and low toxicity. The DNA content was significantly higher in scaffolds seeded with hIVDCs compared with native IVDs, cell-free and hBM-MSC-seeded scaffolds (p < 0.01). The GAG content in the native tissue was significantly higher compared to the others groups except for the scaffolds reseeded with chondrogenically induced hBM-MSCs (p < 0.05). In addition, there was a significantly increased total collagen content in the chondrogenically induced hBM-MSCs group (p < 0.01) compared with the native IVDs, cell-free and hIVDC-seeded scaffolds (p < 0.01); both recolonizing cell types were more evenly distributed on the scaffold surface, but only few cells penetrated the scaffold. The resulting decellularized ECM was cytocompatible and allowed hBM-MSCs/hIVDCs survival and ECM production. PMID:27160711

  9. Systemic Administration of Human Bone Marrow-Derived Mesenchymal Stromal Cell Extracellular Vesicles Ameliorates Aspergillus Hyphal Extract-Induced Allergic Airway Inflammation in Immunocompetent Mice

    PubMed Central

    Cruz, Fernanda F.; Borg, Zachary D.; Goodwin, Meagan; Sokocevic, Dino; Wagner, Darcy E.; Coffey, Amy; Antunes, Mariana; Robinson, Kristen L.; Mitsialis, S. Alex; Kourembanas, Stella; Thane, Kristen; Hoffman, Andrew M.; McKenna, David H.; Rocco, Patricia R.M.

    2015-01-01

    An increasing number of studies demonstrate that administration of either conditioned media (CM) or extracellular vesicles (EVs) released by mesenchymal stromal cells (MSCs) derived from bone marrow and other sources are as effective as the MSCs themselves in mitigating inflammation and injury. The goal of the current study was to determine whether xenogeneic administration of CM or EVs from human bone marrow-derived MSCs would be effective in a model of mixed Th2/Th17, neutrophilic-mediated allergic airway inflammation, reflective of severe refractory asthma, induced by repeated mucosal exposure to Aspergillus hyphal extract (AHE) in immunocompetent C57Bl/6 mice. Systemic administration of both CM and EVs isolated from human and murine MSCs, but not human lung fibroblasts, at the onset of antigen challenge in previously sensitized mice significantly ameliorated the AHE-provoked increases in airway hyperreactivity (AHR), lung inflammation, and the antigen-specific CD4 T-cell Th2 and Th17 phenotype. Notably, both CM and EVs from human MSCs (hMSCs) were generally more potent than those from mouse MSCs (mMSCs) in most of the outcome measures. The weak cross-linking agent 1-ethyl-3-[3-dimethylaminopropyl]carbodiimide hydrochloride was found to inhibit release of both soluble mediators and EVs, fully negating effects of systemically administered hMSCs but only partly inhibited the ameliorating effects of mMSCs. These results demonstrate potent xenogeneic effects of CM and EVs from hMSCs in an immunocompetent mouse model of allergic airway inflammation and they also show differences in mechanisms of action of hMSCs versus mMSCs to mitigate AHR and lung inflammation in this model. Significance There is a growing experience demonstrating benefit of mesenchymal stromal cell (MSC)-based cell therapies in preclinical models of asthma. In the current study, conditioned media (CM) and, in particular, the extracellular vesicle fraction obtained from the CM were as potent as the

  10. Intermittent Hypoxia Mobilizes Bone Marrow-Derived Very Small Embryonic-Like Stem Cells and Activates Developmental Transcriptional Programs in Mice

    PubMed Central

    Gharib, Sina A.; Dayyat, Ehab A.; Khalyfa, Abdelnaby; Kim, Jinkwan; Clair, Heather B.; Kucia, Magdalena; Gozal, David

    2010-01-01

    Background: Obstructive sleep apnea is a prevalent disorder associated with cognitive dysfunction and cardiovascular and metabolic morbidity and is characterized by recurrent episodes of hypoxia during sleep. Bone marrow-derived very small embryonic-like (VSEL) pluripotent stem cells represent a recruitable pool that may play an important role in organ repair after injury. We hypothesized that exposure to intermittent hypoxia (IH) can mobilize VSELs from the bone marrow (BM) to peripheral blood (PB) in mice and can activate distinct transcriptional programs. Methods: Adult mice were exposed to IH or normoxia for 48 hours. VSELs were sorted from BM and PB using flow cytometry. Plasma levels of stem cell chemokines, stromal cell derived factor-1 (SDF-1), hepatocyte growth factor (HGF), and leukemia inhibitory factor (LIF) were measured. Transcriptional profiling of VSELs was performed, and differentially expressed genes were mapped to enriched functional categories and genetic networks. Results: Exposure to IH elicited migration of VSELs from BM to PB and elevations in plasma levels of chemokines. More than 1100 unique genes were differentially expressed in VSELs in response to IH. Gene Ontology and network analysis revealed the activation of organ-specific developmental programs among these genes. Conclusions: Exposure to IH mobilizes VSELs from the BM to PB and activates distinct transcriptional programs in VSELs that are enriched in developmental pathways, including central nervous system development and angiogenesis. Thus, VSELs may serve as a reserve mobile pool of pluripotent stem cells that can be recruited into PB and may play an important role in promoting end-organ repair during IH. Citation: Gharib SA; Dayyat EA; Khalyfa A; Kim J; Clair HB; Kucia M; Gozal D. Intermittent hypoxia mobilizes bone marrow-derived very small embryonic-like stem cells and activates developmental transcriptional programs in mice. SLEEP 2010;33(11):1439-1446. PMID:21102985

  11. Mesenchymal Stem Cell Transplantation in Multiple Sclerosis

    PubMed Central

    Cohen, Jeffrey A.

    2013-01-01

    Mesenchymal stem cells (MSCs) are a pluripotent non-hematopoietic precursor cells that can be isolated from bone marrow and numerous other tissues, culture-expanded to purity, and induced to differentiate in vitro and in vivo into mesodermal derivatives. MSCs exhibit many phenotypic and functional similarities to pericytes. The immunomodulatory, tissue protective, and repair-promoting properties of MSCs demonstrated both in vitro and in animal models make them an attractive potential therapy for MS and other conditions characterized by inflammation and/or tissue injury. Other potential advantages of MSCs as a therapeutic include the relative ease of culture expansion, relative immunoprivilege allowing allogeneic transplantation, and their ability to traffic from blood to areas of tissue allowing intravascular administration. The overall published experience with MSC transplantation in MS is modest, but several small case series and preliminary studies yielded promising results. Several groups, including us, recently initiated formal studies of autologous, culture-expanded, bone-marrow-derived MSC transplantation in MS. Although there are several potential safety concerns, to date, the procedure has been well tolerated. Future studies that more definitively assess efficacy also will need to address several technical issues. PMID:23294498

  12. What Makes Umbilical Cord Tissue-Derived Mesenchymal Stromal Cells Superior Immunomodulators When Compared to Bone Marrow Derived Mesenchymal Stromal Cells?

    PubMed Central

    Bárcia, R. N.; Santos, J. M.; Filipe, M.; Teixeira, M.; Martins, J. P.; Almeida, J.; Água-Doce, A.; Almeida, S. C. P.; Varela, A.; Pohl, S.; Dittmar, K. E. J.; Calado, S.; Simões, S. I.; Gaspar, M. M.; Cruz, M. E. M.; Lindenmaier, W.; Graça, L.; Cruz, H.; Cruz, P. E.

    2015-01-01

    MSCs derived from the umbilical cord tissue, termed UCX, were investigated for their immunomodulatory properties and compared to bone marrow-derived MSCs (BM-MSCs), the gold-standard in immunotherapy. Immunogenicity and immunosuppression were assessed by mixed lymphocyte reactions, suppression of lymphocyte proliferation and induction of regulatory T cells. Results showed that UCX were less immunogenic and showed higher immunosuppression activity than BM-MSCs. Further, UCX did not need prior activation or priming to exert their immunomodulatory effects. This was further corroborated in vivo in a model of acute inflammation. To elucidate the potency differences observed between UCX and BM-MSCs, gene expression related to immune modulation was analysed in both cell types. Several gene expression profile differences were found between UCX and BM-MSCs, namely decreased expression of HLA-DRA, HO-1, IGFBP1, 4 and 6, ILR1, IL6R and PTGES and increased expression of CD200, CD273, CD274, IL1B, IL-8, LIF and TGFB2. The latter were confirmed at the protein expression level. Overall, these results show that UCX seem to be naturally more potent immunosuppressors and less immunogenic than BM-MSCs. We propose that these differences may be due to increased levels of immunomodulatory surface proteins such as CD200, CD273, CD274 and cytokines such as IL1β, IL-8, LIF and TGFβ2. PMID:26064137

  13. Sonic hedgehog and retinoic Acid induce bone marrow-derived stem cells to differentiate into glutamatergic neural cells.

    PubMed

    Yu, Zhenhai; Wu, Shixing; Liu, Zhen; Lin, Haiyan; Chen, Lei; Yuan, Xinli; Zhang, Zhiying; Liu, Fang; Zhang, Chuansen

    2015-01-01

    Studies have showed that transplanted stem cells in the inner ear won't regenerate to replace the damaged sensory hair cells. They can spontaneously differentiate into mesenchymal cells and fibrocytes in the damaged inner ear. Only mature sensory cells of MSCs-derived possess the great potency for cell transplantation in the treatment of sensorineural hearing loss. So, we try to establish an efficient generation of the glutamatergic sensory neural phenotype for the cell transplantation of the hearing loss. We isolated MSCs from femoral and tibial bones according to their adherence to culture dishes. After purification, proliferation, and passaged, cells became homogeneous in appearance, showing more uniformity and grew in a monolayer with a typical spindle-shape morphology. The cell surface markers were assessed using FACS to characterize the isolated cells. For neural induction to harvest the glutamatergic sensory neurons, passage 3 MSCs were incubated with preinduced medium for 24 hr, and neural-induced medium for an additional 14 days. The cells exhibit a typical neural shape. RT-PCR analysis indicated that the mRNA levels of the neural cell marker nestin, Tau, MAP-2, β-tubulin III, GluR-3, and GluR-4 were higher compared with primary MSCs. Immunohistochemistry and western-blotting proofed that nestin, MAP-2, β-tubulin III, and GluR-4 proteins indeed exhibit their expression difference in the induced cells compared to the MSCs. We show an efficient protocol by the combined applications of Sonic Hedgehog (Shh) and Retinoic Acid (RA) to induce MSCs to differentiate into the glutamatergic sensory neuron which were identified from the morphological, biochemical, and molecular characteristics. PMID:24547891

  14. Mesenchymal stem cells for cardiac cell therapy.

    PubMed

    Choi, Yeong-Hoon; Kurtz, Andreas; Stamm, Christof

    2011-01-01

    Despite refinements of medical and surgical therapies, heart failure remains a fatal disease. Myocardial infarction is the most common cause of heart failure, and only palliative measures are available to relieve symptoms and prolong the patient's life span. Because mammalian cardiomyocytes irreversibly exit the cell cycle at about the time of birth, the heart has traditionally been considered to lack any regenerative capacity. This paradigm, however, is currently shifting, and the cellular composition of the myocardium is being targeted by various regeneration strategies. Adult progenitor and stem cell treatment of diseased human myocardium has been carried out for more than 10 years (Menasche et al., 2001; Stamm et al., 2003), and it has become clear that, in humans, the regenerative capacity of hematopoietic stem cells and endothelial progenitor cells, despite potent proangiogenic effects, is limited (Stamm et al., 2009). More recently, mesenchymal stem cells (MSCs) and related cell types are being evaluated in preclinical models of heart disease as well as in clinical trials (see Published Clinical Trials, below). MSCs have the capacity to self-renew and to differentiate into lineages that normally originate from the embryonic mesenchyme (connective tissues, blood vessels, blood-related organs) (Caplan, 1991; Prockop, 1997; Pittenger et al., 1999). The current definition of MSCs includes plastic adherence in cell culture, specific surface antigen expression (CD105(+)/CD90(+)/CD73(+), CD34(-)/CD45(-)/CD11b(-) or CD14(-)/CD19(-) or CD79α(-)/HLA-DR1(-)), and multilineage in vitro differentiation potential (osteogenic, chondrogenic, and adipogenic) (Dominici et al., 2006 ). If those criteria are not met completely, the term "mesenchymal stromal cells" should be used for marrow-derived adherent cells, or other terms for MSC-like cells of different origin. For the purpose of this review, MSCs and related cells are discussed in general, and cell type

  15. Transplantation of mesenchymal stem cells improves type 1 diabetes mellitus.

    PubMed

    Li, Lisha; Li, Furong; Gao, Feng; Yang, Yali; Liu, Yuanyuan; Guo, Pingping; Li, Yulin

    2016-05-01

    Bone-marrow-derived stem cells can regenerate pancreatic tissue in a model of type 1 diabetes mellitus. Mesenchymal stem cells (MSCs) form the main part of bone marrow. We show that the intrapancreatic transplantation of MSCs elevates serum insulin and C-peptide, while decreasing blood glucose. MSCs engrafted into the damaged rat pancreas become distributed into the blood vessels, acini, ducts, and islets. Renascent islets, islet-like clusters, and a small number of MSCs expressing insulin protein have been observed in the pancreas of diabetic rats. Intrapancreatic transplantation of MSCs triggers a series of molecular and cellular events, including differentiation towards the pancreas directly and the provision of a niche to start endogenous pancreatic regeneration, which ameliorates hypoinsulinemia and hyperglycemia caused by streptozotocin. These data establish the many roles of MSCs in the restoration of the function of an injured organ. PMID:26650464

  16. Nicorandil enhances the efficacy of mesenchymal stem cell therapy in isoproterenol-induced heart failure in rats.

    PubMed

    Mohamed, Sarah S; Ahmed, Lamiaa A; Attia, Wael A; Khattab, Mahmoud M

    2015-12-01

    Stem cell transplantation has emerged as a promising technique for regenerative medicine in cardiovascular therapeutics. However, the results have been less than optimal. The aim of the present study was to investigate whether nicorandil could offer an additional benefit over bone marrow-derived mesenchymal stem cell therapy in isoproterenol-induced myocardial damage and its progression to heart failure in rats. Isoproterenol was injected subcutaneously for 2 consecutive days at doses of 85 and 170 mg/kg/day, respectively. Nicorandil (3 mg/kg/day) was then given orally with or without a single intravenous bone marrow-derived mesenchymal stem cell administration. Electrocardiography and echocardiography were recorded 2 weeks after the beginning of treatment. Rats were then sacrificed and the ventricle was isolated for estimation of tumor necrosis factor-alpha, vascular endothelial growth factor and transforming growth factor-beta. Moreover, protein expressions of caspase-3, connexin-43 as well as endothelial and inducible nitric oxide synthases were evaluated. Finally, histological studies of myocardial fibrosis and blood vessel density were performed and cryosections were done for estimation cell homing. Combined nicorandil/bone marrow-derived mesenchymal stem cell therapy provided an additional improvement compared to cell therapy alone toward reducing isoproterenol-induced cardiac hypertrophy, fibrosis and inflammation. Notably, combined therapy induced significant increase in angiogenesis and cell homing and prevented isoproterenol-induced changes in contractility and apoptotic markers. In conclusion, combined nicorandil/bone marrow-derived mesenchymal stem cell therapy was superior to cell therapy alone toward preventing isoproterenol-induced heart failure in rats through creation of a supportive environment for mesenchymal stem cells. PMID:26453143

  17. The Effects of Graphene Nanostructures on Mesenchymal Stem Cells

    PubMed Central

    Lalwani, Gaurav; Kanakia, Shruti; Sitharaman, Balaji

    2014-01-01

    We report the effects of two-dimensional graphene nanostructures; graphene nano-onions (GNOs), graphene oxide nanoribbons (GONRs), and graphene oxide nanoplatelets (GONPs) on viability, and differentiation of human mesenchymal stem cells (MSCs). Cytotoxicity of GNOs, GONRs, and GONPs dispersed in distearoyl-sn-glycero-3-phosphoethanolamine-N-[amino(polyethylene glycol)] (DSPE-PEG), on adipose derived mesenchymal stem cells (adMSCs), and bone marrow-derived mesenchymal stem cells (bmMSCs) was assessed by AlamarBlue and Calcein AM viability assays at concentrations ranging from 5–300 μg/ml for 24 or 72 hours. Cytotoxicity of the 2D graphene nanostructures was found to be dose dependent, not time dependent, with concentrations less than 50 μg/ml showing no significant differences compared to untreated controls. Differentiation potential of adMSCs to adipocytes and osteoblasts, --characterized by Oil Red O staining and elution, alkaline phosphatase activity, calcium matrix deposition and Alizarin Red S staining-- did not change significantly when treated with the three graphene nanoparticles at a low (10 μg/ml) and high (50 μg/ml) concentration for 24 hours. Transmission electron microscopy (TEM) and confocal Raman spectroscopy indicated cellular uptake of only GNOs and GONPs. The results lay the foundation for the use of these nanoparticles at potentially safe doses as ex vivo labels for MSC-based imaging and therapy. PMID:24674462

  18. Mobilization of endogenous bone marrow-derived stem cells in a thioacetamide-induced mouse model of liver fibrosis.

    PubMed

    El-Akabawy, Gehan; El-Mehi, Abeer

    2015-06-01

    The clinical significance of enhancing endogenous circulating haematopoietic stem cells is becoming increasingly recognized, and the augmentation of circulating stem cells using granulocyte-colony stimulating factor (G-CSF) has led to promising preclinical and clinical results for several liver fibrotic conditions. However, this approach is largely limited by cost and the infeasibility of maintaining long-term administration. Preclinical studies have reported that StemEnhance, a mild haematopoietic stem cell mobilizer, promotes cardiac muscle regeneration and remedies the manifestation of diabetes. However, the effectiveness of StemEnhance in ameliorating liver cirrhosis has not been studied. This study is the first to evaluate the beneficial effect of StemEnhance administration in a thioacetamide-induced mouse model of liver fibrosis. StemEnhance augmented the number of peripheral CD34-positive cells, reduced hepatic fibrosis, improved histopathological changes, and induced endogenous liver proliferation. In addition, VEGF expression was up-regulated, while TNF-α expression was down-regulated in thioacetamide-induced fibrotic livers after StemEnhance intake. These data suggest that StemEnhance may be useful as a potential therapeutic candidate for liver fibrosis by inducing reparative effects via mobilization of haematopoietic stem cells. PMID:25857836

  19. Secretome From Mesenchymal Stem Cells Induces Angiogenesis Via Cyr61

    PubMed Central

    ESTRADA, ROSENDO; LI, NA; SAROJINI, HARSHINI; AN, JIN; LEE, MENQ-JER; WANG, EUGENIA

    2010-01-01

    It is well known that bone marrow-derived mesenchymal stem cells (MSCs) are involved in wound healing and regeneration responses. In this study, we globally profiled the proteome of MSCs to investigate critical factor(s) that may promote wound healing. Cysteine-rich protein 61 (Cyr61) was found to be abundantly present in MSCs. The presence of Cyr61 was confirmed by immunofluorescence staining and immunoblot analysis. Moreover, we showed that Cyr61 is present in the culture medium (secretome) of MSCs. The secretome of MSCs stimulates angiogenic response in vitro, and neovascularization in vivo. Depletion of Cyr61 completely abrogates the angiogenic-inducing capability of the MSC secretome. Importantly, addition of recombinant Cyr61 polypeptides restores the angiogenic activity of Cyr61-depleted secretome. Collectively, these data demonstrate that Cyr61 polypeptide in MSC secretome contributes to the angiogenesis-promoting activity, a key event needed for regeneration and repair of injured tissues. PMID:19170074

  20. VEGF improves survival of mesenchymal stem cells in infarcted hearts

    SciTech Connect

    Pons, Jennifer; Huang Yu; Arakawa-Hoyt, Janice; Washko, Daniel; Takagawa, Junya; Ye, Jianqin; Grossman, William; Su Hua

    2008-11-14

    Bone marrow-derived mesenchymal stem cells (MSC) are a promising source for cell-based treatment of myocardial infarction (MI), but existing strategies are restricted by low cell survival and engraftment. We examined whether vascular endothelial growth factor (VEGF) improve MSC viability in infracted hearts. We found long-term culture increased MSC-cellular stress: expressing more cell cycle inhibitors, p16{sup INK}, p21 and p19{sup ARF}. VEGF treatment reduced cellular stress, increased pro-survival factors, phosphorylated-Akt and Bcl-xL expression and cell proliferation. Co-injection of MSCs with VEGF to MI hearts increased cell engraftment and resulted in better improvement of cardiac function than that injected with MSCs or VEGF alone. In conclusion, VEGF protects MSCs from culture-induce cellular stress and improves their viability in ischemic myocardium, which results in improvements of their therapeutic effect for the treatment of MI.

  1. Isolation of functionally distinct mesenchymal stem cell subsets using antibodies against CD56, CD271, and mesenchymal stem cell antigen-1

    PubMed Central

    Battula, Venkata Lokesh; Treml, Sabrina; Bareiss, Petra M.; Gieseke, Friederike; Roelofs, Helene; de Zwart, Peter; Müller, Ingo; Schewe, Bernhard; Skutella, Thomas; Fibbe, Willem E.; Kanz, Lothar; Bühring, Hans-Jörg

    2009-01-01

    Background Conventionally, mesenchymal stem cells are functionally isolated from primary tissue based on their capacity to adhere to a plastic surface. This isolation procedure is hampered by the unpredictable influence of co-cultured hematopoietic and/or other unrelated cells and/or by the elimination of a late adhering mesenchymal stem cells subset during removal of undesired cells. To circumvent these limitations, several antibodies have been developed to facilitate the prospective isolation of mesenchymal stem cells. Recently, we described a panel of monoclonal antibodies with superior selectivity for mesenchymal stem cells, including the monoclonal antibodies W8B2 against human mesenchymal stem cell antigen-1 (MSCA-1) and 39D5 against a CD56 epitope, which is not expressed on natural killer cells. Design and Methods Bone marrow derived mesenchymal stem cells from healthy donors were analyzed and isolated by flow cytometry using a large panel of antibodies against surface antigens including CD271, MSCA-1, and CD56. The growth of mesenchymal stem cells was monitored by colony formation unit fibroblast (CFU-F) assays. The differentiation of mesenchymal stem cells into defined lineages was induced by culture in appropriate media and verified by immunostaining. Results Multicolor cell sorting and CFU-F assays showed that mesenchymal stem cells were ~90-fold enriched in the MSCA-1+CD56− fraction and ~180-fold in the MSCA-1+CD56+ fraction. Phenotype analysis revealed that the expression of CD10, CD26, CD106, and CD146 was restricted to the MSCA-1+CD56− mesenchymal stem cells subset and CD166 to MSCA-1+CD56± mesenchymal stem cells. Further differentiation of these subsets showed that chondrocytes and pancreatic-like islets were predominantly derived from MSCA-1+CD56± cells whereas adipocytes emerged exclusively from MSCA-1+CD56− cells. The culture of single sorted MSCA-1+CD56+ cells resulted in the appearance of phenotypically heterogeneous clones with

  2. The Effects of Secretion Factors from Umbilical Cord Derived Mesenchymal Stem Cells on Osteogenic Differentiation of Mesenchymal Stem Cells

    PubMed Central

    Wang, Kui-Xing; Xu, Liang-Liang; Rui, Yun-Feng; Huang, Shuo; Lin, Si-En; Xiong, Jiang-Hui; Li, Ying-Hui; Lee, Wayne Yuk-Wai; Li, Gang

    2015-01-01

    Factors synthesized by mesenchymal stem cells (MSCs) contain various growth factors, cytokines, exosomes and microRNAs, which may affect the differentiation abilities of MSCs. In the present study, we investigated the effects of secretion factors of human umbilical cord derived mesenchymal stem cells (hUCMSCs) on osteogenesis of human bone marrow derived MSCs (hBMSCs). The results showed that 20 μg/ml hUCMSCs secretion factors could initiate osteogenic differentiation of hBMSCs without osteogenic induction medium (OIM), and the amount of calcium deposit (stained by Alizarin Red) was significantly increased after the hUCMSCs secretion factors treatment. Real time quantitative reverse transcription-polymerase chain reaction (real time qRT-PCR) demonstrated that the expression of osteogenesis-related genes including ALP, BMP2, OCN, Osterix, Col1α and Runx2 were significantly up-regulated following hUCMSCs secretion factors treatment. In addition, we found that 10 μg hUCMSCs secretion factors together with 2×105 hBMSCs in the HA/TCP scaffolds promoted ectopic bone formation in nude mice. Local application of 10 μg hUCMSCs secretion factors with 50 μl 2% hyaluronic acid hydrogel and 1×105 rat bone marrow derived MSCs (rBMSCs) also significantly enhanced the bone repair of rat calvarial bone critical defect model at both 4 weeks and 8 weeks. Moreover, the group that received the hUCMSCs secretion factors treatment had more cartilage and bone regeneration in the defect areas than those in the control group. Taken together, these findings suggested that hUCMSCs secretion factors can initiate osteogenesis of bone marrow MSCs and promote bone repair. Our study indicates that hUCMSCs secretion factors may be potential sources for promoting bone regeneration. PMID:25799169

  3. Migration of Bone Marrow-Derived Very Small Embryonic-Like Stem Cells toward An Injured Spinal Cord

    PubMed Central

    Golipoor, Zoleikha; Mehraein, Fereshteh; Zafari, Fariba; Alizadeh, Akram; Ababzadeh, Shima; Baazm, Maryam

    2016-01-01

    Objective Bone marrow (BM) is one of the major hematopoietic organs in postnatal life that consists of a heterogeneous population of stem cells which have been previously described. Recently, a rare population of stem cells that are called very small embryonic-like (VSEL) stem cells has been found in the BM. These cells express several developmental markers of pluri- potent stem cells and can be mobilized into peripheral blood (PB) in response to tissue injury. In this study we have attempted to investigate the ability of these cells to migrate toward an injured spinal cord after transplantation through the tail vein in a rat model. Materials and Methods In this experimental study, VSELs were isolated from total BM cells using a fluorescent activated cell sorting (FACS) system and sca1 and stage specific embryonic antigen (SSEA-1) antibodies. After isolation, VSELs were cultured for 7 days on C2C12 as the feeder layer. Then, VSELs were labeled with 1,1´-dioctadecyl-3,3,3´,3´- tetramethylindocarbocyanine perchlorate (DiI) and transplanted into the rat spinal cord injury (SCI) model via the tail vein. Finally, we sought to determine the presence of VSELs in the lesion site. Results We isolated a high number of VSELs from the BM. After cultivation, the VSELs colonies were positive for SSEA-1, Oct4 and Sca1. At one month after transplantation, real-time polymerase chain reaction analysis confirmed a significantly increased expres- sion level of Oct4 and SSEA-1 positive cells at the injury site. Conclusion VSELs have the capability to migrate and localize in an injured spinal cord after transplantation. PMID:26862523

  4. Mesenchymal stem cells are sensitive to bleomycin treatment

    PubMed Central

    Nicolay, Nils H.; Rühle, Alexander; Perez, Ramon Lopez; Trinh, Thuy; Sisombath, Sonevisay; Weber, Klaus-Josef; Ho, Anthony D.; Debus, Jürgen; Saffrich, Rainer; Huber, Peter E.

    2016-01-01

    Mesenchymal stem cells (MSCs) have been shown to attenuate pulmonary damage induced by bleomycin-based anticancer treatments, but the influence of bleomycin on the stem cells themselves remains largely unknown. Here, we demonstrate that human bone marrow-derived MSCs are relatively sensitive to bleomycin exposure compared to adult fibroblasts. MSCs revealed increased levels of apoptosis after bleomycin treatment, while cellular morphology, stem cell surface marker expression and the ability for adhesion and migration remained unchanged. Bleomycin treatment also resulted in a reduced adipogenic differentiation potential of these stem cells. MSCs were found to efficiently repair DNA double strand breaks induced by bleomycin, mostly through non-homologous end joining repair. Low mRNA and protein expression levels of the inactivating enzyme bleomycin hydrolase were detected in MSCs that may contribute to the observed bleomycin-sensitive phenotype of these cells. The sensitivity of MSCs against bleomycin needs to be taken into consideration for ongoing and future treatment protocols investigating these stem cells as a potential treatment option for bleomycin-induced pulmonary damage in the clinic. PMID:27215195

  5. Ameliorative effect of bone marrow-derived stem cells on injured liver of mice infected with Schistosoma mansoni.

    PubMed

    El-Mahdi, Magda M; Mansour, Wafaa A; Hammam, Olfat; Mehana, Noha A; Hussein, Taghreed M

    2014-04-01

    The technique of stem cells or hepatocytes transplantation has recently improved in order to bridge the time before whole-organ liver transplantation. In the present study, unfractionated bone marrow stem cells (BMSCs) were harvested from the tibial and femoral marrow compartments of male mice, which were cultured in Dulbecco's modified Eagle's medium (DMEM) with and without hepatocyte growth factor (HGF), and then transplanted into Schistosoma mansoni-infected female mice on their 8th week post-infection. Mice were sacrificed monthly until the third month of bone marrow transplantation, serum was collected, and albumin concentration, ALT, AST, and alkaline phosphatase (ALP) activities were assayed. On the other hand, immunohistopathological and immunohistochemical changes of granuloma size and number, collagen content, and cells expressing OV-6 were detected for identification of liver fibrosis. BMSCs were shown to differentiate into hepatocyte-like cells. Serum ALT, AST, and ALP were markedly reduced in the group of mice treated with BMSCs than in the untreated control group. Also, granuloma showed a marked decrease in size and number as compared to the BMSCs untreated group. Collagen content showed marked decrease after the third month of treatment with BMSCs. On the other hand, the expression of OV-6 increased detecting the presence of newly formed hepatocytes after BMSCs treatment. BMSCs with or without HGF infusion significantly enhanced hepatic regeneration in S. mansoni-induced fibrotic liver model and have pathologic and immunohistopathologic therapeutic effects. Also, this new therapeutic trend could generate new hepatocytes to improve the overall liver functions. PMID:24850958

  6. Ameliorative Effect of Bone Marrow-Derived Stem Cells on Injured Liver of Mice Infected with Schistosoma mansoni

    PubMed Central

    Mansour, Wafaa A.; Hammam, Olfat; Mehana, Noha A.; Hussein, Taghreed M.

    2014-01-01

    The technique of stem cells or hepatocytes transplantation has recently improved in order to bridge the time before whole-organ liver transplantation. In the present study, unfractionated bone marrow stem cells (BMSCs) were harvested from the tibial and femoral marrow compartments of male mice, which were cultured in Dulbecco's modified Eagle's medium (DMEM) with and without hepatocyte growth factor (HGF), and then transplanted into Schistosoma mansoni-infected female mice on their 8th week post-infection. Mice were sacrificed monthly until the third month of bone marrow transplantation, serum was collected, and albumin concentration, ALT, AST, and alkaline phosphatase (ALP) activities were assayed. On the other hand, immunohistopathological and immunohistochemical changes of granuloma size and number, collagen content, and cells expressing OV-6 were detected for identification of liver fibrosis. BMSCs were shown to differentiate into hepatocyte-like cells. Serum ALT, AST, and ALP were markedly reduced in the group of mice treated with BMSCs than in the untreated control group. Also, granuloma showed a marked decrease in size and number as compared to the BMSCs untreated group. Collagen content showed marked decrease after the third month of treatment with BMSCs. On the other hand, the expression of OV-6 increased detecting the presence of newly formed hepatocytes after BMSCs treatment. BMSCs with or without HGF infusion significantly enhanced hepatic regeneration in S. mansoni-induced fibrotic liver model and have pathologic and immunohistopathologic therapeutic effects. Also, this new therapeutic trend could generate new hepatocytes to improve the overall liver functions. PMID:24850958

  7. The potential for resident lung mesenchymal stem cells to promote functional tissue regeneration: understanding microenvironmental cues.

    PubMed

    Foronjy, Robert F; Majka, Susan M

    2012-12-01

    Tissue resident mesenchymal stem cells (MSCs) are important regulators of tissue repair or regeneration, fibrosis, inflammation, angiogenesis and tumor formation. Bone marrow derived mesenchymal stem cells (BM-MSCs) and endothelial progenitor cells (EPC) are currently being considered and tested in clinical trials as a potential therapy in patients with such inflammatory lung diseases including, but not limited to, chronic lung disease, pulmonary arterial hypertension (PAH), pulmonary fibrosis (PF), chronic obstructive pulmonary disease (COPD)/emphysema and asthma. However, our current understanding of tissue resident lung MSCs remains limited. This review addresses how environmental cues impact on the phenotype and function of this endogenous stem cell pool. In addition, it examines how these local factors influence the efficacy of cell-based treatments for lung diseases. PMID:23626909

  8. Bone Marrow Derived Eosinophil Cultures

    PubMed Central

    Lu, Thomas X.; Rothenberg, Marc E.

    2016-01-01

    Eosinophils are multifunctional effector cells implicated in the pathogenesis of a variety of diseases including asthma, eosinophil gastrointestinal disorders and helminth infection. Mouse bone marrow derived progenitor cells can be differentiated into eosinophils following IL-5 exposure. These bone marrow derived eosinophils are fully differentiated at the end of a 14 day culture based on morphology and expression of molecular markers.

  9. Osteogenic Potency of Nacre on Human Mesenchymal Stem Cells

    PubMed Central

    Green, David W.; Kwon, Hyuk-Jae; Jung, Han-Sung

    2015-01-01

    Nacre seashell is a natural osteoinductive biomaterial with strong effects on osteoprogenitors, osteoblasts, and osteoclasts during bone tissue formation and morphogenesis. Although nacre has shown, in one study, to induce bridging of new bone across large non-union bone defects in 8 individual human patients, there have been no succeeding human surgical studies to confirm this outstanding potency. But the molecular mechanisms associated with nacre osteoinduction and the influence on bone marrow-derived mesenchymal stem cells (BMSC’s), skeletal stem cells or bone marrow stromal cells remain elusive. In this study we highlight the phenotypic and biochemical effects of Pinctada maxima nacre chips and the global nacre soluble protein matrix (SPM) on primary human bone marrow-derived stromal cells (hBMSCs) in vitro. In static co-culture with nacre chips, the hBMSCs secreted Alkaline phosphatase (ALP) at levels that exceeded bone morphogenetic protein (rhBMP-2) treatment. Concentrated preparation of SPM applied to Stro-1 selected hBMSC’s led to rapid ALP secretions, at concentrations exceeding the untreated controls even in osteogenic conditions. Within 21 days the same population of Stro-1 selected hBMSCs proliferated and secreted collagens I–IV, indicating the premature onset of an osteoblast phenotype. The same SPM was found to promote unselected hBMSC differentiation with osteocalcin detected at 7 days, and proliferation increased at 7 days in a dose-dependent manner. In conclusion, nacre particles and nacre SPM induced the early stages of human bone cell differentiation, indicating that they may be promising soluble factors with osteoinductive capacity in primary human bone cell progenitors such as, hBMSC’s. PMID:25666352

  10. Mesenchymal markers on human adipose stem/progenitor cells

    PubMed Central

    Zimmerlin, Ludovic; Donnenberg, Vera S.; Rubin, J. Peter; Donnenberg, Albert D.

    2014-01-01

    The stromal-vascular fraction (SVF) of adipose tissue is a rich source of multipotent stem cells. We and others have described 3 major populations of stem/progenitor cells in this fraction, all closely associated with small blood vessels: endothelial progenitor cells (EPC, CD45−/CD31+/CD34+), pericytes (CD45−/CD31−/CD146+) and supra-adventitial adipose stromal cells (SA-ASC, CD45−/CD31−/CD146−/CD34+). EPC are luminal, pericytes are adventitial and SA-ASC surround the vessel like a sheath. The multipotency of the pericytes and SA-ASC compartments is strikingly similar to that of CD45−/CD34−/CD73+/CD105+/CD90+ bone marrow-derived mesenchymal stem cells (BM-MSC). Here we determine the extent to which this mesenchymal expression pattern is expressed on the 3 adipose stem/progenitor populations. Eight independent adipose tissue samples were analyzed in a single tube (CD105-FITC/CD73-PE/CD146-PETXR/CD14-PECY5/CD33-PECY5/CD235A-PECY5/CD31-PECY7/CD90-APC/CD34-A700/CD45-APCCY7/DAPI). Adipose EPC were highly proliferative with 14.3±2.8% (mean ± SEM) having >2N DNA. About half (53.1±7.6%) coexpressed CD73 and CD105, and 71.9±7.4% expressed CD90. Pericytes were less proliferative (8.2±3.4% >2N DNA) with a smaller proportion (29.6±6.9% CD73+/CD105+, 60.5±10.2% CD90+) expressing mesenchymal associated markers. However, the CD34+ subset of CD146+ pericytes, were both highly proliferative (15.1±3.6% with >2N DNA) and of uniform mesenchymal phenotype (93.3±3.7% CD73+/CD105+, 97.8±0.7% CD90+), suggesting transit amplifying progenitor cells. SA-ASC were the least proliferative (3.7 ± 0.8%>2N DNA) but were also highly mesenchymal in phenotype (94.4±3.2% CD73+/CD105+, 95.5±1.2% CD90+). These data imply a progenitor/progeny relationship between pericytes and SA-ASC, the most mesenchymal of SVF cells. Despite phenotypic and functional similarities to BM-MSC, SA-ASC are distinguished by CD34 expression. PMID:23184564

  11. Human mesenchymal stem-cell behaviour on direct laser micropatterned electrospun scaffolds with hierarchical structures.

    PubMed

    Li, Huaqiong; Wong, Yee Shan; Wen, Feng; Ng, Kee Woei; Ng, Gary Ka Lai; Venkatraman, Subbu S; Boey, Freddy Yin Chiang; Tan, Lay Poh

    2013-03-01

    Direct laser machining and electrospinning are utilized to obtain a bi-layered hybrid scaffold with hierarchical topographical features to mimic extracellular matrix-like microenvironment of cells. Adult bone marrow derived human mesenchymal stem cells (hMSCs) are cultured in vitro in these hybrid scaffolds, and cell orientation, proliferation, viability, and differentiation are evaluated. The results show that this novel hybrid scaffold not only supports cell growth like traditional scaffolds, but also elicits positive responses from the cells, like lineage commitment and alignment, which are essential features of future scaffolds. PMID:23233197

  12. Chitosan and Chitin Hexamers affect expansion and differentiation of mesenchymal stem cells differently.

    PubMed

    Lieder, Ramona; Thormodsson, Finnbogi; Ng, C-H; Einarsson, Jon M; Gislason, Johannes; Petersen, Petur H; Sigurjonsson, Olafur E

    2012-11-01

    Chitooligosaccharides are of interest as potential drugs due to their bioactivity and water solubility. We compared the effect of acetylated and deacetylated chitooligomers (Hexamers) on short-term expansion (7 days) and osteogenic differentiation of bone-marrow derived, human mesenchymal stem cells in terms of gene expression, cytokine secretion and quality of osteogenic differentiation. We show that chitooligomers affect hMSC gene expression and cytokine secretion, but not mineralization. The effect of chitooligomers was shown to be dependent on the acetylation degree, with significantly stronger effects when cells are stimulated with chitin-derived Hexamers (N-Acetyl Chitohexaose) than with Chitosan Hexamers (Chitohexaose). PMID:22790025

  13. Generation of mesenchymal stromal cells in the presence of platelet lysate: a phenotypic and functional comparison of umbilical cord blood- and bone marrow-derived progenitors

    PubMed Central

    Avanzini, Maria Antonietta; Bernardo, Maria Ester; Cometa, Angela Maria; Perotti, Cesare; Zaffaroni, Nadia; Novara, Francesca; Visai, Livia; Moretta, Antonia; Del Fante, Claudia; Villa, Raffaella; Ball, Lynne M.; Fibbe, Willem E.; Maccario, Rita; Locatelli, Franco

    2009-01-01

    Background Mesenchymal stromal cells are employed in various different clinical settings in order to modulate immune response. However, relatively little is known about the mechanisms responsible for their immunomodulatory effects, which could be influenced by both the cell source and culture conditions. Design and Methods We tested the ability of a 5% platelet lysate-supplemented medium to support isolation and ex vivo expansion of mesenchymal stromal cells from full-term umbilical-cord blood. We also investigated the biological/functional properties of umbilical cord blood mesenchymal stromal cells, in comparison with platelet lysate-expanded bone marrow mesenchymal stromal cells. Results The success rate of isolation of mesenchymal stromal cells from umbilical cord blood was in the order of 20%. These cells exhibited typical morphology, immunophenotype and differentiation capacity. Although they have a low clonogenic efficiency, umbilical cord blood mesenchymal stromal cells may possess high proliferative potential. The genetic stability of these cells from umbilical cord blood was demonstrated by a normal molecular karyotype; in addition, these cells do not express hTERT and telomerase activity, do express p16ink4a protein and do not show anchorage-independent cell growth. Concerning alloantigen-specific immune responses, umbilical cord blood mesenchymal stromal cells were able to: (i) suppress T- and NK-lymphocyte proliferation, (ii) decrease cytotoxic activity and (iii) only slightly increase interleukin-10, while decreasing interferon-γ secretion, in mixed lymphocyte culture supernatants. While an indoleamine 2,3-dioxygenase-specific inhibitor did not reverse mesenchymal stromal cell-induced suppressive effects, a prostaglandin E2-specific inhibitor hampered the suppressive effect of both umbilical cord blood- and bone marrow-mesenchymal stromal cells on alloantigen-induced cytotoxic activity. Mesenchymal stromal cells from both sources expressed HLA

  14. Snail1 controls TGF-β responsiveness and differentiation of Mesenchymal Stem Cells

    PubMed Central

    Batlle, Raquel; Alba-Castellón, Lorena; Loubat-Casanovas, Jordina; Armenteros, Elena; Francí, Clara; Stanisavljevic, Jelena; Banderas, Rodrigo; Martin-Caballero, Juan; Bonilla, Félix; Baulida, Josep; Casal, J. Ignacio; Gridley, Thomas; de Herreros, Antonio García

    2012-01-01

    The Snail1 transcriptional repressor plays a key role in triggering epithelial to mesenchymal transition. Although Snail1 is widely expressed in early development, in adult animals it is limited to a subset of mesenchymal cells where it has a largely unknown function. Using a mouse model with inducible depletion of Snail1, here we demonstrate that Snail1 is required to maintain mesenchymal stem cells (MSCs). This effect is associated to the responsiveness to TGF-β1 which shows a strong Snail1 dependence. Snail1-depletion in conditional knock-out adult animals causes a significant decrease in the number of bone marrow-derived MSCs. In culture, Snail1-deficient MSCs prematurely differentiate to osteoblasts or adipocytes and, in contrast to controls, are resistant to the TGF-β1-induced differentiation block. These results demonstrate a new role for Snail1 in TGF-β response and MSC maintenance. PMID:22869142

  15. Antitumor immune response of dendritic cells (DCs) expressing tumor-associated antigens derived from induced pluripotent stem cells: in comparison to bone marrow-derived DCs.

    PubMed

    Iwamoto, Hiromitsu; Ojima, Toshiyasu; Hayata, Keiji; Katsuda, Masahiro; Miyazawa, Motoki; Iida, Takeshi; Nakamura, Masaki; Nakamori, Mikihito; Iwahashi, Makoto; Yamaue, Hiroki

    2014-01-15

    It is generally accepted that the difficulty in obtaining a sufficient number of functional dendritic cells (DCs) is a serious problem in DC-based immunotherapy. Therefore, we used the induced pluripotent stem (iPS) cell-derived DCs (iPSDCs). If the therapeutic efficacy of iPSDCs is equivalent to that of bone marrow-derived DCs (BMDCs), then the aforementioned problems may be solved. In our study, we induced iPSDCs from iPS cells and examined the capacity for maturation of iPSDCs compared to that of BMDCs in addition to the capacity for migration of iPSDCs to regional lymph nodes. We adenovirally transduced the hgp100 gene, natural tumor antigens, into DCs and immunized mice once with the genetically modified DCs. The cytotoxic activity of CD8 (+) cytotoxic T lymphocytes (CTLs) was assayed using a (51) Cr-release assay. The therapeutic efficacy of the vaccination was examined in a subcutaneous tumor model. Our results showed that iPSDCs have an equal capacity to BMDCs in terms of maturation and migration. Furthermore, hgp100-specific CTLs were generated in mice immunized with genetically modified iPSDCs. These CTLs exhibited as high a level of cytotoxicity against B16 cells as BMDCs. Moreover, vaccination with the genetically modified iPSDCs achieved as high a level of therapeutic efficacy as vaccination with BMDCs. Our study clarified experimentally that genetically modified iPSDCs have an equal capacity to BMDCs in terms of tumor-associated antigen-specific therapeutic antitumor immunity. This vaccination strategy may therefore be useful for future clinical application as a cancer vaccine. PMID:23824921

  16. The Great Migration of Bone Marrow-Derived Stem Cells Toward the Ischemic Brain: Therapeutic Implications for Stroke and Other Neurological Disorders

    PubMed Central

    Borlongan, Cesar V.; Glover, Loren E.; Tajiri, Naoki; Kaneko, Yuji; Freeman, Thomas B.

    2011-01-01

    Accumulating laboratory studies have implicated the mobilization of bone marrow (BM)-derived stem cells in brain plasticity and stroke therapy. This mobilization of bone cells to the brain is an essential concept in regenerative medicine. Over the past ten years, mounting data have shown the ability of bone marrow–derived stem cells to mobilize from BM to the peripheral blood (PB) and eventually enter the injured brain. This homing action is exemplified in BM stem cell mobilization following ischemic brain injury. Various BM-derived cells, such as hematopoietic stem cells (HSCs), mesenchymal stem cells (MSCs), endothelial progenitor cells (EPCs) and very small embryonic-like cells (VSELs) have been demonstrated to exert therapeutic benefits in stroke. Here, we discuss the current status of these BM-derived stem cells in stroke therapy, with emphasis on possible cellular and molecular mechanisms of action that mediate the cells’ beneficial effects in the ischemic brain. When possible, we also discuss the relevance of this therapeutic regimen in other central nervous system (CNS) disorders. PMID:21903148

  17. Human mesenchymal stem cells: New sojourn of bacterial pathogens.

    PubMed

    Kohli, Sakshi; Singh, Yadvir; Sowpati, Divya Tej; Ehtesham, Nasreen Z; Dobrindt, Ulrich; Hacker, Jörg; Hasnain, Seyed E

    2015-05-01

    Tuberculosis (TB), caused by Mycobacterium tuberculosis (M. tuberculosis), is the leading infectious disease which claims one human life every 15-20s globally. The persistence of this deadly disease in human population can be attributed to the ability of the bacterium to stay in latent form. M. tuberculosis possesses a plethora of mechanisms not only to survive latently under harsh conditions inside the host but also modulate the host immune cells in its favour. Various M. tuberculosis gene families have also been described to play a role in this process. Recently, human bone marrow derived mesenchymal stem cells (MSCs) have been reported as a niche for dormant M. tuberculosis. MSCs possess abilities to alter the host immune response. The bacterium finds this self-renewal and immune privileged nature of MSCs very favourable not only to modulate the host immune system, with some help from its own genes, but also to avoid the external drug pressure. We suggest that the MSCs not only provide a resting place for M. tuberculosis but could also, by virtue of their intrinsic ability to disseminate in the body, explain the genesis of extra-pulmonary TB. A similar exploitation of stem cells by other bacterial pathogens is a distinct possibility. It may be likely that other intracellular bacterial pathogens adopt this strategy to 'piggy-back' on to ovarian stem cells to ensure vertical transmission and successful propagation to the next generation. PMID:25648374

  18. Mesenchymal Stem Cells as Therapeutics

    PubMed Central

    Parekkadan, Biju; Milwid, Jack M.

    2013-01-01

    Mesenchymal stem cells (MSCs) are multipotent cells that are being clinically explored as a new therapeutic for treating a variety of immune-mediated diseases. First heralded as a regenerative therapy for skeletal tissue repair, MSCs have recently been shown to modulate endogenous tissue and immune cells. Preclinical studies of the mechanism of action suggest that the therapeutic effects afforded by MSC transplantation are short-lived and related to dynamic, paracrine interactions between MSCs and host cells. Therefore, representations of MSCs as drug-loaded particles may allow for pharmacokinetic models to predict the therapeutic activity of MSC transplants as a function of drug delivery mode. By integrating principles of MSC biology, therapy, and engineering, the field is armed to usher in the next generation of stem cell therapeutics. PMID:20415588

  19. Non-invasive characterization of the adipogenic differentiation of human bone marrow-derived mesenchymal stromal cells by HS-SPME/GC-MS

    PubMed Central

    Lee, Dong-Kyu; Yi, TacGhee; Park, Kyung-Eun; Lee, Hyun-Joo; Cho, Yun-Kyoung; Lee, Seul Ji; Lee, Jeongmi; Park, Jeong Hill; Lee, Mi-Young; Song, Sun U.; Kwon, Sung Won

    2014-01-01

    A non-invasive method to characterize human mesenchymal stromal cells during adipogenic differentiation was developed for the first time. Seven fatty acid methyl esters (FAMEs), including methyl laurate, methyl myristate, methyl palmitate, methyl linoleate, methyl oleate, methyl elaidate and methyl stearate, were used for characterizing adipogenic differentiation using headspace solid-phase microextraction (HS-SPME) which is a very simple and non-invasive method for the extraction of volatile compounds. Glassware was used for culturing mesenchymal stromal cells rather than the common plasticware to minimize contamination by volatile impurities. The optimal SPME fiber was selected by comparing diverse fibers containing two pure liquid polymers (PDMS and PA) and two porous solids (PDMS/DVB and CAR/PDMS). Using optimized procedures, we discovered that seven FAMEs were only detected in adipogenic differentiated mesenchymal stromal cells and not in the mesenchymal stromal cells before differentiation. These data could support the quality control of clinical mesenchymal stromal cell culture in the pharmaceutical industry in addition to the development of many clinical applications using mesenchymal stromal cells. PMID:25298091

  20. Non-invasive characterization of the adipogenic differentiation of human bone marrow-derived mesenchymal stromal cells by HS-SPME/GC-MS.

    PubMed

    Lee, Dong-Kyu; Yi, TacGhee; Park, Kyung-Eun; Lee, Hyun-Joo; Cho, Yun-Kyoung; Lee, Seul Ji; Lee, Jeongmi; Park, Jeong Hill; Lee, Mi-Young; Song, Sun U; Kwon, Sung Won

    2014-01-01

    A non-invasive method to characterize human mesenchymal stromal cells during adipogenic differentiation was developed for the first time. Seven fatty acid methyl esters (FAMEs), including methyl laurate, methyl myristate, methyl palmitate, methyl linoleate, methyl oleate, methyl elaidate and methyl stearate, were used for characterizing adipogenic differentiation using headspace solid-phase microextraction (HS-SPME) which is a very simple and non-invasive method for the extraction of volatile compounds. Glassware was used for culturing mesenchymal stromal cells rather than the common plasticware to minimize contamination by volatile impurities. The optimal SPME fiber was selected by comparing diverse fibers containing two pure liquid polymers (PDMS and PA) and two porous solids (PDMS/DVB and CAR/PDMS). Using optimized procedures, we discovered that seven FAMEs were only detected in adipogenic differentiated mesenchymal stromal cells and not in the mesenchymal stromal cells before differentiation. These data could support the quality control of clinical mesenchymal stromal cell culture in the pharmaceutical industry in addition to the development of many clinical applications using mesenchymal stromal cells. PMID:25298091

  1. Expression of CD24 in Human Bone Marrow-Derived Mesenchymal Stromal Cells Is Regulated by TGFβ3 and Induces a Myofibroblast-Like Genotype

    PubMed Central

    Schäck, Luisa Marilena; Buettner, Manuela; Wirth, Alexander; Krettek, Christian; Hoffmann, Andrea; Noack, Sandra

    2016-01-01

    Human bone marrow-derived stromal cells (hBMSCs) derived from the adult organism hold great promise for diverse settings in regenerative medicine. Therefore a more complete understanding of hBMSC biology to fully exploit the cells' potential for clinical settings is important. The protein CD24 has been reported to be involved in a diverse range of processes such as cancer, adaptive immunity, inflammation, and autoimmune diseases in other cell types. Its expression in hBMSCs, which has not yet been analyzed, may add an important aspect in the understanding of hBMSC biology. The present study therefore analyzes the expression, regulation, and functional implication of the surface protein CD24 in hBMSCs. Methods used are stimulation studies with TGF beta as well as shRNA-mediated knockdown and overexpression of CD24 followed by microarray, immunocytochemistry, and flow cytometric analyses. To our knowledge, we demonstrate for the first time that the expression of CD24 is an inherent property of hBMSCs. Importantly, the data links the upregulation of CD24 to the adoption of a myofibroblast-like gene expression pattern in hBMSCs. We demonstrate that CD24 is an important modulator in transforming growth factor beta 3 (TGFβ3) signaling with a reciprocal regulatory relationship between these two proteins. PMID:26788063

  2. Transplantation of expanded bone marrow-derived very small embryonic-like stem cells (VSEL-SCs) improves left ventricular function and remodelling after myocardial infarction

    PubMed Central

    Zuba-Surma, Ewa K; Guo, Yiru; Taher, Hisham; Sanganalmath, Santosh K; Hunt, Greg; Vincent, Robert J; Kucia, Magda; Abdel-Latif, Ahmed; Tang, Xian-Liang; Ratajczak, Mariusz Z; Dawn, Buddhadeb; Bolli, Roberto

    2011-01-01

    Abstract Adult bone marrow-derived very small embryonic-like stem cells (VSEL-SCs) exhibit a Sca-1+/Lin–/CD45– phenotype and can differentiate into various cell types, including cardiomyocytes and endothelial cells. We have previously reported that transplantation of a small number (1 × 106) of freshly isolated, non-expanded VSEL-SCs into infarcted mouse hearts resulted in improved left ventricular (LV) function and anatomy. Clinical translation, however, will require large numbers of cells. Because the frequency of VSEL-SCs in the marrow is very low, we examined whether VSEL-SCs can be expanded in culture without loss of therapeutic efficacy. Mice underwent a 30 min. coronary occlusion followed by reperfusion and, 48 hrs later, received an intramyocardial injection of vehicle (group I, n= 11), 1 × 105 enhanced green fluorescent protein (EGFP)-labelled expanded untreated VSEL-SCs (group II, n= 7), or 1 × 105 EGFP-labelled expanded VSEL-SCs pre-incubated in a cardiogenic medium (group III, n= 8). At 35 days after myocardial infarction (MI), mice treated with pre-incubated VSEL-SCs exhibited better global and regional LV systolic function and less LV hypertrophy compared with vehicle-treated controls. In contrast, transplantation of expanded but untreated VSEL-SCs did not produce appreciable reparative benefits. Scattered EGFP+ cells expressing α-sarcomeric actin, platelet endothelial cell adhesion molecule (PECAM)-1, or von Willebrand factor were present in VSEL-SC-treated mice, but their numbers were very small. No tumour formation was observed. We conclude that VSEL-SCs expanded in culture retain the ability to alleviate LV dysfunction and remodelling after a reperfused MI provided that they are exposed to a combination of cardiomyogenic growth factors and cytokines prior to transplantation. Counter intuitively, the mechanism whereby such pre-incubation confers therapeutic efficacy does not involve differentiation into new cardiac cells. These results

  3. Bile acids induce hepatic differentiation of mesenchymal stem cells

    PubMed Central

    Sawitza, Iris; Kordes, Claus; Götze, Silke; Herebian, Diran; Häussinger, Dieter

    2015-01-01

    Mesenchymal stem cells (MSC) have the potential to differentiate into multiple cell lineages and their therapeutic potential has become obvious. In the liver, MSC are represented by stellate cells which have the potential to differentiate into hepatocytes after stimulation with growth factors. Since bile acids can promote liver regeneration, their influence on liver-resident and bone marrow-derived MSC was investigated. Physiological concentrations of bile acids such as tauroursodeoxycholic acid were able to initiate hepatic differentiation of MSC via the farnesoid X receptor and transmembrane G-protein-coupled bile acid receptor 5 as investigated with knockout mice. Notch, hedgehog, transforming growth factor-β/bone morphogenic protein family and non-canonical Wnt signalling were also essential for bile acid-mediated differentiation, whereas β-catenin-dependent Wnt signalling was able to attenuate this process. Our findings reveal bile acid-mediated signalling as an alternative way to induce hepatic differentiaion of stem cells and highlight bile acids as important signalling molecules during liver regeneration. PMID:26304833

  4. Mesenchymal Stem Cells as Cellular Vectors for Pediatric Neurological Disorders

    PubMed Central

    Phinney, Donald G.; Isakova, Iryna A.

    2014-01-01

    Lysosomal storage diseases are a heterogeneous group of hereditary disorders characterized by a deficiency in lysosomal function. Although these disorders differ in their etiology and phenotype those that affect the nervous system generally manifest as a profound deterioration in neurologic function with age. Over the past several decades implementation of various treatment regimens including bone marrow and cord blood cell transplantation, enzyme replacement, and substrate reduction therapy have proved effective for managing some clinical manifestations of these diseases but their ability to ameliorate neurologic complications remains unclear. Consequently, there exists a need to develop alternative therapies that more effectively target the central nervous system. Recently, direct intracranial transplantation of tissue-specific stem and progenitor cells has been explored as a means to reconstitute metabolic deficiencies in the CNS. In this chapter we discuss the merits of bone marrow-derived mesenchymal stem cells (MSCs) for this purpose. Originally identified as progenitors of connective tissue cell lineages, recent findings have revealed several novel aspects of MSC biology that make them attractive as therapeutic agents in the CNS. We relate these advances in MSC biology to their utility as cellular vectors for treating neurologic sequelae associated with pediatric neurologic disorders. PMID:24858930

  5. Sources of adult mesenchymal stem cells for ligament and tendon tissue engineering.

    PubMed

    Dhinsa, Baljinder S; Mahapatra, Anant N; Khan, Wasim S

    2014-01-01

    Tendon and ligament injuries are common, and repair slowly with reduced biomechanical properties. With increasing financial demands on the health service and patients to recover from tendon and ligament injuries faster, and with less morbidity, health professionals are exploring new treatment options. Tissue engineering may provide the answer, with its unlimited source of natural cells that in the correct environment may improve repair and regeneration of tendon and ligament tissue. Mesenchymal stem cells have demonstrated the ability to self renew and have multilineage differentiation potential. The use of bone marrow-derived mesenchymal stem cells has been reported, however significant in vitro culture expansion is required due to the low yield of cells, which has financial implications. Harvesting of bone marrow cells also has associated morbidity. Several studies have looked at alternative sources for mesenchymal stem cells. Reports in literature from animal studies have been encouraging, however further work is required. This review assesses the potential sources of mesenchymal stem cells for tissue engineering in tendons and ligaments. PMID:25012740

  6. Mesenchymal stem cells protect from hypoxia-induced alveolar epithelial-mesenchymal transition.

    PubMed

    Uzunhan, Yurdagül; Bernard, Olivier; Marchant, Dominique; Dard, Nicolas; Vanneaux, Valérie; Larghero, Jérôme; Gille, Thomas; Clerici, Christine; Valeyre, Dominique; Nunes, Hilario; Boncoeur, Emilie; Planès, Carole

    2016-03-01

    Administration of bone marrow-derived human mesenchymal stem cells (hMSC) reduces lung inflammation, fibrosis, and mortality in animal models of lung injury, by a mechanism not completely understood. We investigated whether hMSC would prevent epithelial-mesenchymal transition (EMT) induced by hypoxia in primary rat alveolar epithelial cell (AEC). In AEC cultured on semipermeable filters, prolonged hypoxic exposure (1.5% O2 for up to 12 days) induced phenotypic changes consistent with EMT, i.e., a change in cell morphology, a decrease in transepithelial resistance (Rte) and in the expression of epithelial markers [zonula occludens-1 (ZO-1), E-cadherin, AQP-5, TTF-1], together with an increase in mesenchymal markers [vimentin, α-smooth muscle actin (α-SMA)]. Expression of transcription factors driving EMT such as SNAIL1, ZEB1, and TWIST1 increased after 2, 24, and 48 h of hypoxia, respectively. Hypoxia also induced TGF-β1 mRNA expression and the secretion of active TGF-β1 in apical medium, and the expression of connective tissue growth factor (CTGF), two inducers of EMT. Coculture of AEC with hMSC partially prevented the decrease in Rte and in ZO-1, E-cadherin, and TTF-1 expression, and the increase in vimentin expression induced by hypoxia. It also abolished the increase in TGF-β1 expression and in TGF-β1-induced genes ZEB1, TWIST1, and CTGF. Finally, incubation with human recombinant KGF at a concentration similar to what was measured in hMSC-conditioned media restored the expression of TTF-1 and prevented the increase in TWIST1, TGF-β1, and CTGF in hypoxic AEC. Our results indicate that hMSC prevent hypoxia-induced alveolar EMT through the paracrine modulation of EMT signaling pathways and suggest that this effect is partly mediated by KGF. PMID:26702148

  7. Mesenchymal Stem Cells in Cardiology.

    PubMed

    White, Ian A; Sanina, Cristina; Balkan, Wayne; Hare, Joshua M

    2016-01-01

    Cardiovascular disease (CVD) accounts for more deaths globally than any other single disease. There are on average 1.5 million episodes of myocardial infarction (heart attack) each year in the United States alone with roughly one-third resulting in death. There is therefore a major need for developing new and effective strategies to promote cardiac repair. Intramyocardial transplantation of mesenchymal stem cells (MSCs) has emerged as a leading contender in the pursuit of clinical intervention and therapy. MSCs are potent mediators of cardiac repair and are therefore an attractive tool in the development of preclinical and clinical trials. MSCs are capable of secreting a large array of soluble factors, which have had demonstrated effects on pathogenic cardiac remolding, fibrosis, immune activation, and cardiac stem cell proliferation within the damaged heart. MSCs are also capable of differentiation into cardiomyocytes, endothelial cells, and vascular smooth muscle cells, although the relative contribution of trilineage differentiation and paracrine effectors on cardiac repair remains the subject of active investigation. PMID:27236666

  8. Mesenchymal stem cells in regenerative rehabilitation

    PubMed Central

    Nurkovic, Jasmin; Dolicanin, Zana; Mustafic, Fahrudin; Mujanovic, Rifat; Memic, Mensur; Grbovic, Vesna; Skevin, Aleksandra Jurisic; Nurkovic, Selmina

    2016-01-01

    [Purpose] Regenerative medicine and rehabilitation contribute in many ways to a specific plan of care based on a patient’s medical status. The intrinsic self-renewing, multipotent, regenerative, and immunosuppressive properties of mesenchymal stem cells offer great promise in the treatment of numerous autoimmune, degenerative, and graft-versus-host diseases, as well as tissue injuries. As such, mesenchymal stem cells represent a therapeutic fortune in regenerative medicine. The aim of this review is to discuss possibilities, limitations, and future clinical applications of mesenchymal stem cells. [Subjects and Methods] The authors have identified and discussed clinically and scientifically relevant articles from PubMed that have met the inclusion criteria. [Results] Direct treatment of muscle injuries, stroke, damaged peripheral nerves, and cartilage with mesenchymal stem cells has been demonstrated to be effective, with synergies seen between cellular and physical therapies. Over the past few years, several researchers, including us, have shown that there are certain limitations in the use of mesenchymal stem cells. Aging and spontaneous malignant transformation of mesenchymal stem cells significantly affect the functionality of these cells. [Conclusion] Definitive conclusions cannot be made by these studies because limited numbers of patients were included. Studies clarifying these results are expected in the near future. PMID:27390452

  9. Mesenchymal stem cells in regenerative rehabilitation.

    PubMed

    Nurkovic, Jasmin; Dolicanin, Zana; Mustafic, Fahrudin; Mujanovic, Rifat; Memic, Mensur; Grbovic, Vesna; Skevin, Aleksandra Jurisic; Nurkovic, Selmina

    2016-06-01

    [Purpose] Regenerative medicine and rehabilitation contribute in many ways to a specific plan of care based on a patient's medical status. The intrinsic self-renewing, multipotent, regenerative, and immunosuppressive properties of mesenchymal stem cells offer great promise in the treatment of numerous autoimmune, degenerative, and graft-versus-host diseases, as well as tissue injuries. As such, mesenchymal stem cells represent a therapeutic fortune in regenerative medicine. The aim of this review is to discuss possibilities, limitations, and future clinical applications of mesenchymal stem cells. [Subjects and Methods] The authors have identified and discussed clinically and scientifically relevant articles from PubMed that have met the inclusion criteria. [Results] Direct treatment of muscle injuries, stroke, damaged peripheral nerves, and cartilage with mesenchymal stem cells has been demonstrated to be effective, with synergies seen between cellular and physical therapies. Over the past few years, several researchers, including us, have shown that there are certain limitations in the use of mesenchymal stem cells. Aging and spontaneous malignant transformation of mesenchymal stem cells significantly affect the functionality of these cells. [Conclusion] Definitive conclusions cannot be made by these studies because limited numbers of patients were included. Studies clarifying these results are expected in the near future. PMID:27390452

  10. Generation of functional islets from human umbilical cord and placenta derived mesenchymal stem cells.

    PubMed

    Kadam, Sachin; Govindasamy, Vijayendran; Bhonde, Ramesh

    2012-01-01

    Bone marrow-derived mesenchymal stem cells (BM-MSCs) have been used for allogeneic application in tissue engineering but have certain drawbacks. Therefore, mesenchymal stem cells (MSCs) derived from other adult tissue sources have been considered as an alternative. The human umbilical cord and placenta are easily available noncontroversial sources of human tissue, which are often discarded as biological waste, and their collection is noninvasive. These sources of MSCs are not subjected to ethical constraints, as in the case of embryonic stem cells. MSCs derived from umbilical cord and placenta are multipotent and have the ability to differentiate into various cell types crossing the lineage boundary towards endodermal lineage. The aim of this chapter is to provide a detailed reproducible cookbook protocol for the isolation, propagation, characterization, and differentiation of MSCs derived from human umbilical cord and placenta with special reference to harnessing their potential towards pancreatic/islet lineage for utilization as a cell therapy product. We show here that mesenchymal stromal cells can be extensively expanded from umbilical cord and placenta of human origin retaining their multilineage differentiation potential in vitro. Our report indicates that postnatal tissues obtained as delivery waste represent a rich source of mesenchymal stromal cells, which can be differentiated into functional islets employing three-stage protocol developed by our group. These islets could be used as novel in vitro model for screening hypoglycemics/insulin secretagogues, thus reducing animal experimentation for this purpose and for the future human islet transplantation programs to treat diabetes. PMID:22610566

  11. Mesenchymal Stem Cells from Human Extra Ocular Muscle Harbor Neuroectodermal Differentiation Potential.

    PubMed

    Mawrie, Darilang; Kumar, Atul; Magdalene, Damaris; Bhattacharyya, Jina; Jaganathan, Bithiah Grace

    2016-01-01

    Mesenchymal stem cells (MSC) have been proposed as suitable candidates for cell therapy for neurological disorderssince they exhibit good neuronal differentiation capacity. However, for better therapeutic outcomes, it is necessary to isolate MSC from a suitable tissue sourcethat posses high neuronal differentiation. In this context, we isolated MSC from extra ocular muscle (EOM) tissue and tested the in vitro neuronal differentiation potential. In the current study, EOM tissue derived MSC were characterized and compared with bone marrow derived MSC. We found that EOM derived MSC proliferated as a monolayer and showed similarities in morphology, growth properties and cell surface marker expression with bone marrow derived MSC and expressed high levels of NES, OCT4, NANOG and SOX2 in its undifferentiated state. They also expressed embryonic cell surface marker SSEA4 and their intracellular mitochondrial distribution pattern was similar to that of multipotent stem cells. Although EOM derived MSC differentiated readily into adipocytes, osteocytes and chondrocytes, they differentiated more efficiently into neuroectodermal cells. The differentiation into neuroectodermal cellswas confirmed by the expression of neuronal markers NGFR and MAP2B. Thus, EOM derived MSC might be good candidates for stem cell based therapies for treating neurodegenerative diseases. PMID:27248788

  12. Mesenchymal Stem Cells from Human Extra Ocular Muscle Harbor Neuroectodermal Differentiation Potential

    PubMed Central

    Magdalene, Damaris; Bhattacharyya, Jina; Jaganathan, Bithiah Grace

    2016-01-01

    Mesenchymal stem cells (MSC) have been proposed as suitable candidates for cell therapy for neurological disorderssince they exhibit good neuronal differentiation capacity. However, for better therapeutic outcomes, it is necessary to isolate MSC from a suitable tissue sourcethat posses high neuronal differentiation. In this context, we isolated MSC from extra ocular muscle (EOM) tissue and tested the in vitro neuronal differentiation potential. In the current study, EOM tissue derived MSC were characterized and compared with bone marrow derived MSC. We found that EOM derived MSC proliferated as a monolayer and showed similarities in morphology, growth properties and cell surface marker expression with bone marrow derived MSC and expressed high levels of NES, OCT4, NANOG and SOX2 in its undifferentiated state. They also expressed embryonic cell surface marker SSEA4 and their intracellular mitochondrial distribution pattern was similar to that of multipotent stem cells. Although EOM derived MSC differentiated readily into adipocytes, osteocytes and chondrocytes, they differentiated more efficiently into neuroectodermal cells. The differentiation into neuroectodermal cellswas confirmed by the expression of neuronal markers NGFR and MAP2B. Thus, EOM derived MSC might be good candidates for stem cell based therapies for treating neurodegenerative diseases. PMID:27248788

  13. Mesenchymal Stem Cells Respond to Hypoxia by Increasing Diacylglycerols.

    PubMed

    Lakatos, Kinga; Kalomoiris, Stefanos; Merkely, Béla; Nolta, Jan A; Fierro, Fernando A

    2016-02-01

    Mesenchymal stem cells (MSC) are currently being tested clinically for a plethora of conditions, with most approaches relying on the secretion of paracrine signals by MSC to modulate the immune system, promote wound healing, and induce angiogenesis. Hypoxia has been shown to affect MSC proliferation, differentiation, survival and secretory profile. Here, we investigate changes in the lipid composition of human bone marrow-derived MSC after exposure to hypoxia. Using mass spectrometry, we compared the lipid profiles of MSC derived from five different donors, cultured for two days in either normoxia (control) or hypoxia (1% oxygen). Hypoxia induced a significant increase of total triglycerides, fatty acids and diacylglycerols (DG). Remarkably, reduction of DG levels using the phosphatidylcholine-specific phospholipase C inhibitor D609 inhibited the secretion of VEGF and Angiopoietin-2, but increased the secretion of interleukin-8, without affecting significantly their respective mRNA levels. Functionally, incubation of MSC in hypoxia with D609 inhibited the potential of the cells to promote migration of human endothelial cells in a wound/scratch assay. Hence, we show that hypoxia induces in MSC an increase of DG that may affect the angiogenic potential of these cells. PMID:26212931

  14. Mesenchymal Stem Cell (MSC) Aggregate Formation in vivo

    PubMed Central

    Bartosh, Thomas J.; Ylostalo, Joni H.

    2016-01-01

    Human mesenchymal stem/progenitor cells (MSCs) isolated from various adult tissues show remarkable therapeutic potential and are being employed in clinical trials for the treatment of numerous diseases (Prockop et al., 2010). While routes of cell administration vary, profound beneficial effects of MSCs in animal models have been observed following intraperitoneal injections of the cells (Roddy et al., 2011). Similar to MSC spheres formed in culture under conditions where attachment to plastic is not permitted (Bartosh et al., 2010), MSCs injected into the peritoneum of mice spontaneously aggregate into 3D sphere-like structures (Bartosh et al., 2013). During the process of sphere assembly and compaction, MSCs upregulate expression of numerous therapeutic anti-inflammatory and immune modulatory factors. Here we describe the method we previously used for the generation of human bone marrow-derived MSC aggregates/spheres in vivo (Bartosh et al., 2013). By tagging the MSCs with green fluorescent protein (GFP), the aggregates formed can be easily visualized, collected and analyzed for changes in cellular properties and interactions with host immune cells.

  15. Enhancement of mesenchymal stem cell angiogenic capacity and stemness by a biomimetic hydrogel scaffold

    PubMed Central

    Rustad, Kristine C.; Wong, Victor W.; Sorkin, Michael; Glotzbach, Jason P.; Major, Melanie R.; Rajadas, Jayakumar; Longaker, Michael T.; Gurtner, Geoffrey C.

    2014-01-01

    In this study, we examined the capacity of a biomimetic pullulan–collagen hydrogel to create a functional biomaterial-based stem cell niche for the delivery of mesenchymal stem cells (MSCs) into wounds. Murine bone marrow-derived MSCs were seeded into hydrogels and compared to MSCs grown in standard culture conditions. Hydrogels induced MSC secretion of angiogenic cytokines and expression of transcription factors associated with maintenance of pluripotency and self-renewal (Oct4, Sox2, Klf4) when compared to MSCs grown in standard conditions. An excisonal wound healing model was used to compare the ability of MSC-hydrogel constructs versus MSC injection alone to accelerate wound healing. Injection of MSCs did not significantly improve time to wound closure. In contrast, wounds treated with MSC-seeded hydrogels showed significantly accelerated healing and a return of skin appendages. Bioluminescence imaging and FACS analysis of luciferase+/GFP+ MSCs indicated that stem cells delivered within the hydrogel remained viable longer and demonstrated enhanced engraftment efficiency than those delivered via injection. Engrafted MSCs were found to differentiate into fibroblasts, pericytes and endothelial cells but did not contribute to the epidermis. Wounds treated with MSC-seeded hydrogels demonstrated significantly enhanced angiogenesis, which was associated with increased levels of VEGF and other angiogenic cytokines within the wounds. Our data suggest that biomimetic hydrogels provide a functional niche capable of augmenting MSC regenerative potential and enhancing wound healing. PMID:21963148

  16. Behaviour of human mesenchymal stem cells on chemically synthesized HA-PCL scaffolds for hard tissue regeneration.

    PubMed

    D'Antò, Vincenzo; Raucci, Maria Grazia; Guarino, Vincenzo; Martina, Stefano; Valletta, Rosa; Ambrosio, Luigi

    2016-02-01

    Our goal was to characterize the response of human mesenchymal stem cells (hMSCs) to a novel composite scaffold for bone tissue engineering. The hydroxyapatite-polycaprolactone (HA-PCL) composite scaffolds were prepared by a sol-gel method at room temperature and the scaffold morphology was investigated by scanning electron microscopy (SEM)/energy-dispersive spectroscopy (EDS) to validate the synthesis process. The response of two different lines of hMSCs, bone-marrow-derived human mesenchymal stem cells (BMSCs) and dental pulp stem cells (DPSCs) in terms of cell proliferation and differentiation into the osteoblastic phenotype, was evaluated using Alamar blue assay, SEM, histology and alkaline phosphatase activity. Our results indicate that tissue engineering by means of composite HA-PCL scaffolds may represent a new therapeutic strategy to repair craniofacial bone defects. PMID:23723157

  17. Potential mechanisms underlying the Runx2 induced osteogenesis of bone marrow mesenchymal stem cells

    PubMed Central

    Xu, Jiahai; Li, Zhanghua; Hou, Yudong; Fang, Weijun

    2015-01-01

    Bone marrow derived mesenchymal stem cells (BM-MSCs) belong a type of pluripotent stem cells and can be induced to differentiate into osteoblasts (OB). Runt-related transcription factor 2 (Runx2) is an osteogenesis specific transcription factor and plays an important role in osteogenesis of BM-MSCs. It can promote the expression of osteogenesis related genes, regulate cell cycle progression, improve bone microenvironment and affect functions of chondrocytes and osteoclasts, which have involvement of a large amount of signal molecules including TGF-β, BMP, Notch, Wnt, Hedgehog, FGF and microRNA. In this paper, we summarize the mechanisms underlying the Runx2 induced osteogenesis of BM-MSCs. PMID:26885254

  18. Mesenchymal Stem Cells Improve Motor Functions and Decrease Neurodegeneration in Ataxic Mice

    PubMed Central

    Jones, Jonathan; Estirado, Alicia; Redondo, Carolina; Pacheco-Torres, Jesus; Sirerol-Piquer, Maria-Salomé; Garcia-Verdugo, José M; Martinez, Salvador

    2015-01-01

    The main objective of this work is to demonstrate the feasibility of using bone marrow-derived stem cells in treating a neurodegenerative disorder such as Friedreich's ataxia. In this disease, the dorsal root ganglia of the spinal cord are the first to degenerate. Two groups of mice were injected intrathecally with mesenchymal stem cells isolated from either wild-type or Fxntm1Mkn/Tg(FXN)YG8Pook (YG8) mice. As a result, both groups presented improved motor skills compared to nontreated mice. Also, frataxin expression was increased in the dorsal root ganglia of the treated groups, along with lower expression of the apoptotic markers analyzed. Furthermore, the injected stem cells expressed the trophic factors NT3, NT4, and BDNF, which bind to sensory neurons of the dorsal root ganglia and increase their survival. The expression of antioxidant enzymes indicated that the stem cell-treated mice presented higher levels of catalase and GPX-1, which are downregulated in the YG8 mice. There were no significant differences in the use of stem cells isolated from wild-type and YG8 mice. In conclusion, bone marrow mesenchymal stem cell transplantation, both autologous and allogeneic, is a feasible therapeutic option to consider in delaying the neurodegeneration observed in the dorsal root ganglia of Friedreich's ataxia patients. PMID:25070719

  19. Tumor necrosis factor alpha promotes the expression of immunosuppressive proteins and enhances the cell growth in a human bone marrow-derived stem cell culture.

    PubMed

    Miettinen, Johanna A; Pietilä, Mika; Salonen, Riikka J; Ohlmeier, Steffen; Ylitalo, Kari; Huikuri, Heikki V; Lehenkari, Petri

    2011-04-01

    Mesenchymal stem cells (MSCs) are widely used in experimental treatments for various conditions that involve normal tissue regeneration via inflammatory repair. It is known that MSCs can secrete multiple soluble factors and suppress inflammation. Even though the effect of MSCs on inflammation has been extensively studied, the effect of inflammation on MSCs is poorly understood. One of the major cytokines released at the site of inflammation is tumor necrosis factor alpha (TNF-α) which is known to induce MSC invasion and proliferation. Therefore, we wanted to test the effects of TNF-α exposure on MSCs derived from human bone marrow. We found, as expected, that cell proliferation was significantly enhanced during TNF-α exposure. However, according to the cell surface marker analysis, the intensity of several antigens in the minimum criteria panel for MSCs proposed by International Society of Cellular Therapy (ISCT) was decreased dramatically, and in certain cases, the criteria for MSCs were not fulfilled. In addition, TNF-α exposure resulted in a significant but transient increase in human leukocyte antigen and CD54 expression. Additional proteomic analysis by two-dimensional difference gel electrophoresis and mass spectrometry revealed three proteins whose expression levels decreased and 8 proteins whose expression levels increased significantly during TNF-α exposure. The majority of these proteins could be linked to immunosuppressive and signalling pathways. These results strongly support reactive and immunosuppressive activation of MSCs during TNF-α exposure, which might influence MSC differentiation stage and capacity. PMID:21182837

  20. Tumor necrosis factor alpha promotes the expression of immunosuppressive proteins and enhances the cell growth in a human bone marrow-derived stem cell culture

    SciTech Connect

    Miettinen, Johanna A.; Pietilae, Mika; Salonen, Riikka J.; Ohlmeier, Steffen; Ylitalo, Kari; Huikuri, Heikki V.; Lehenkari, Petri

    2011-04-01

    Mesenchymal stem cells (MSCs) are widely used in experimental treatments for various conditions that involve normal tissue regeneration via inflammatory repair. It is known that MSCs can secrete multiple soluble factors and suppress inflammation. Even though the effect of MSCs on inflammation has been extensively studied, the effect of inflammation on MSCs is poorly understood. One of the major cytokines released at the site of inflammation is tumor necrosis factor alpha (TNF-{alpha}) which is known to induce MSC invasion and proliferation. Therefore, we wanted to test the effects of TNF-{alpha} exposure on MSCs derived from human bone marrow. We found, as expected, that cell proliferation was significantly enhanced during TNF-{alpha} exposure. However, according to the cell surface marker analysis, the intensity of several antigens in the minimum criteria panel for MSCs proposed by International Society of Cellular Therapy (ISCT) was decreased dramatically, and in certain cases, the criteria for MSCs were not fulfilled. In addition, TNF-{alpha} exposure resulted in a significant but transient increase in human leukocyte antigen and CD54 expression. Additional proteomic analysis by two-dimensional difference gel electrophoresis and mass spectrometry revealed three proteins whose expression levels decreased and 8 proteins whose expression levels increased significantly during TNF-{alpha} exposure. The majority of these proteins could be linked to immunosuppressive and signalling pathways. These results strongly support reactive and immunosuppressive activation of MSCs during TNF-{alpha} exposure, which might influence MSC differentiation stage and capacity.

  1. A robust and reproducible animal serum-free culture method for clinical-grade bone marrow-derived mesenchymal stromal cells.

    PubMed

    Laitinen, Anita; Oja, Sofia; Kilpinen, Lotta; Kaartinen, Tanja; Möller, Johanna; Laitinen, Saara; Korhonen, Matti; Nystedt, Johanna

    2016-08-01

    Efficient xenofree expansion methods to replace fetal bovine serum (FBS)-based culture methods are strongly encouraged by the regulators and are needed to facilitate the adoption of mesenchymal stromal cell (MSC)-based therapies. In the current study we established a clinically-compliant and reproducible animal serum-free culture protocol for bone marrow-(BM-) MSCs based on an optimized platelet-derived supplement. Our study compared two different platelet-derived supplements, platelet lysate PL1 versus PL2, produced by two different methods and lysed with different amounts of freeze-thaw cycles. Our study also explored the effect of a low oxygen concentration on BM-MSCs. FBS-supplemented BM-MSC culture served as control. Growth kinetics, differentiation and immunomodulatory potential, morphology, karyotype and immunophenotype was analysed. Growth kinetics in long-term culture was also studied. Based on the initial results, we chose to further process develop the PL1-supplemented culture protocol at 20 % oxygen. The results from 11 individual BM-MSC batches expanded in the chosen condition were consistent, yielding 6.60 × 10(9) ± 4.74 × 10(9) cells from only 20 ml of bone marrow. The cells suppressed T-cell proliferation, displayed normal karyotype and typical MSC differentiation potential and phenotype. The BM-MSCs were, however, consistently HLA-DR positive when cultured in platelet lysate (7.5-66.1 %). We additionally show that culture media antibiotics and sterile filtration of the platelet lysate can be successfully omitted. We present a robust and reproducible clinically-compliant culture method for BM-MSCs based on platelet lysate, which enables high quantities of HLA-DR positive MSCs at a low passage number (p2) and suitable for clinical use. PMID:25777046

  2. Canine Platelet Lysate Is Inferior to Fetal Bovine Serum for the Isolation and Propagation of Canine Adipose Tissue- and Bone Marrow-Derived Mesenchymal Stromal Cells

    PubMed Central

    Russell, Keith A.; Gibson, Thomas W. G.; Chong, Andrew; Co, Carmon; Koch, Thomas G.

    2015-01-01

    Background Mesenchymal stromal cells (MSC) are increasingly investigated for their clinical utility in dogs. Fetal bovine serum (FBS) is a common culture supplement used for canine MSC expansion. However, FBS content is variable, its clinical use carries risk of an immune response, and its cost is increasing due to global demand. Platelet lysate (PL) has proven to be a suitable alternative to FBS for expansion of human MSC. Hypothesis and Objectives We hypothesized that canine adipose tissue (AT) and bone marrow (BM) MSC could be isolated and expanded equally in PL and FBS at conventionally-used concentrations with differentiation of these MSC unaffected by choice of supplement. Our objectives were to evaluate the use of canine PL in comparison with FBS at four stages: 1) isolation, 2) proliferation, 3) spontaneous differentiation, and 4) directed differentiation. Results 1) Medium with 10% PL was unable to isolate MSC. 2) MSC, initially isolated in FBS-supplemented media, followed a dose-dependent response with no significant difference between PL and FBS cultures at up to 20% (AT) or 30% (BM) enrichment. Beyond these respective peaks, proliferation fell in PL cultures only, while a continued dose-dependent proliferation response was noted in FBS cultures. 3) Further investigation indicated PL expansion culture was inducing spontaneous adipogenesis in concentrations as low as 10% and as early as 4 days in culture. 4) MSC isolated in FBS, but expanded in either FBS or PL, maintained ability to undergo directed adipogenesis and osteogenesis, but not chondrogenesis. Conclusions/Significance Canine PL did not support establishment of MSC colonies from AT and BM, nor expansion of MSC, which appear to undergo spontaneous adipogenesis in response to PL exposure. In vivo studies are warranted to determine if concurrent use of MSC with any platelet-derived products such as platelet-rich plasma are associated with synergistic, neutral or antagonistic effects. PMID:26353112

  3. Biological, functional and genetic characterization of bone marrow-derived mesenchymal stromal cells from pediatric patients affected by acute lymphoblastic leukemia.

    PubMed

    Conforti, Antonella; Biagini, Simone; Del Bufalo, Francesca; Sirleto, Pietro; Angioni, Adriano; Starc, Nadia; Li Pira, Giuseppina; Moretta, Francesca; Proia, Alessandra; Contoli, Benedetta; Genovese, Silvia; Ciardi, Claudia; Avanzini, Maria Antonietta; Rosti, Vittorio; Lo-Coco, Francesco; Locatelli, Franco; Bernardo, Maria Ester

    2013-01-01

    Alterations in hematopoietic microenvironment of acute lymphoblastic leukemia patients have been claimed to occur, but little is known about the components of marrow stroma in these patients. In this study, we characterized mesenchymal stromal cells (MSCs) isolated from bone marrow (BM) of 45 pediatric patients with acute lymphoblastic leukemia (ALL-MSCs) at diagnosis (day+0) and during chemotherapy treatment (days: +15; +33; +78), the time points being chosen according to the schedule of BM aspirates required by the AIEOP-BFM ALL 2009 treatment protocol. Morphology, proliferative capacity, immunophenotype, differentiation potential, immunomodulatory properties and ability to support long-term hematopoiesis of ALL-MSCs were analysed and compared with those from 41 healthy donors (HD-MSCs). ALL-MSCs were also genetically characterized through array-CGH, conventional karyotyping and FISH analysis. Moreover, we compared ALL-MSCs generated at day+0 with those isolated during chemotherapy. Morphology, immunophenotype, differentiation potential and in vitro life-span did not differ between ALL-MSCs and HD-MSCs. ALL-MSCs showed significantly lower proliferative capacity (p<0.001) and ability to support in vitro hematopoiesis (p = 0.04) as compared with HD-MSCs, while they had similar capacity to inhibit in vitro mitogen-induced T-cell proliferation (p = N.S.). ALL-MSCs showed neither the typical translocations carried by the leukemic clone (when present), nor other genetic abnormalities acquired during ex vivo culture. Our findings indicate that ALL-MSCs display reduced ability to proliferate and to support long-term hematopoiesis in vitro. ALL-MSCs isolated at diagnosis do not differ from those obtained during treatment. PMID:24244271

  4. Biological, Functional and Genetic Characterization of Bone Marrow-Derived Mesenchymal Stromal Cells from Pediatric Patients Affected by Acute Lymphoblastic Leukemia

    PubMed Central

    Conforti, Antonella; Biagini, Simone; Del Bufalo, Francesca; Sirleto, Pietro; Angioni, Adriano; Starc, Nadia; Li Pira, Giuseppina; Moretta, Francesca; Proia, Alessandra; Contoli, Benedetta; Genovese, Silvia; Ciardi, Claudia; Avanzini, Maria Antonietta; Rosti, Vittorio; Lo-Coco, Francesco; Locatelli, Franco; Bernardo, Maria Ester

    2013-01-01

    Alterations in hematopoietic microenvironment of acute lymphoblastic leukemia patients have been claimed to occur, but little is known about the components of marrow stroma in these patients. In this study, we characterized mesenchymal stromal cells (MSCs) isolated from bone marrow (BM) of 45 pediatric patients with acute lymphoblastic leukemia (ALL-MSCs) at diagnosis (day+0) and during chemotherapy treatment (days: +15; +33; +78), the time points being chosen according to the schedule of BM aspirates required by the AIEOP-BFM ALL 2009 treatment protocol. Morphology, proliferative capacity, immunophenotype, differentiation potential, immunomodulatory properties and ability to support long-term hematopoiesis of ALL-MSCs were analysed and compared with those from 41 healthy donors (HD-MSCs). ALL-MSCs were also genetically characterized through array-CGH, conventional karyotyping and FISH analysis. Moreover, we compared ALL-MSCs generated at day+0 with those isolated during chemotherapy. Morphology, immunophenotype, differentiation potential and in vitro life-span did not differ between ALL-MSCs and HD-MSCs. ALL-MSCs showed significantly lower proliferative capacity (p<0.001) and ability to support in vitro hematopoiesis (p = 0.04) as compared with HD-MSCs, while they had similar capacity to inhibit in vitro mitogen-induced T-cell proliferation (p = N.S.). ALL-MSCs showed neither the typical translocations carried by the leukemic clone (when present), nor other genetic abnormalities acquired during ex vivo culture. Our findings indicate that ALL-MSCs display reduced ability to proliferate and to support long-term hematopoiesis in vitro. ALL-MSCs isolated at diagnosis do not differ from those obtained during treatment. PMID:24244271

  5. Canine bone marrow-derived mesenchymal stromal cells suppress alloreactive lymphocyte proliferation in vitro but fail to enhance engraftment in canine bone marrow transplantation.

    PubMed

    Lee, Won Sik; Suzuki, Yasuhiro; Graves, Scott S; Iwata, Mineo; Venkataraman, G M; Mielcarek, Marco; Peterson, Laura J; Ikehara, Susumu; Torok-Storb, Beverly; Storb, Rainer

    2011-04-01

    Stable mixed hematopoietic chimerism has been consistently established in dogs who were mildly immunosuppressed by 200 cGy of total body irradiation (TBI) before undergoing dog leukocyte antigen (DLA)-identical bone marrow (BM) transplantation and who received a brief course of immunosuppression with mycophenolate mofetil (28 days) and cyclosporine (35 days) after transplantation. However, when TBI was reduced from 200 to 100 cGy, grafts were nearly uniformly rejected within 3-12 weeks. Here, we asked whether stable engraftment could be accomplished after a suboptimal dose of 100 cGy TBI with host immunosuppression enhanced by donor-derived mesenchymal stromal cells (MSCs) given after transplantation. MSCs were cultured from BM cells and evaluated in vitro for antigen expression. They showed profound immunosuppressive properties in mixed lymphocyte reactions (MLRs) in a cell dose-dependent manner not restricted by DLA. MSC and lymphocyte contact was not required, indicating that immunosuppression was mediated by soluble factors. Prostaglandin E2 was increased in culture supernatant when MSCs were cocultured in MLRs. The addition of indomethacin restored lymphocyte proliferation in cultures containing MSCs. MSCs expressed CD10, CD13, CD29, CD44, CD73/SH-3, CD90/Thy-1, and CD106/VCAM-1. For in vivo studies, MSCs were injected on the day of BM grafting and on day 35, the day of discontinuation of posttransplantation cyclosporine. MSCs derived from the respective BM donors failed to avert BM graft rejection in 4 dogs who received DLA-identical grafts after nonmyeloablative conditioning with 100 cGy TBI in a time course not significantly different from that of control dogs not given MSCs. Although the MSCs displayed in vitro characteristics similar to those reported for MSCs from other species, their immunosuppressive qualities failed to sustain stable BM engraftment in vivo in this canine model. PMID:20457265

  6. Equine bone marrow-derived mesenchymal stromal cells are heterogeneous in MHC class II expression and capable of inciting an immune response in vitro

    PubMed Central

    2014-01-01

    Introduction The horse is a valuable species to assess the effect of allogeneic mesenchymal stromal cells (MSCs) in regenerative treatments. No studies to date have examined recipient response to major histocompatibility complex (MHC)-mismatched equine MSCs. The purposes of this study were to immunophenotype MSCs from horses of known MHC haplotype and to compare the immunogenicity of MSCs with differing MHC class II expression. Methods MSCs and peripheral blood leukocytes (PBLs) were obtained from Thoroughbred horses (n = 10) of known MHC haplotype (ELA-A2, -A3, and -A9 homozygotes). MSCs were cultured through P8; cells from each passage (P2 to P8) were cryopreserved until used. Immunophenotyping of MHC class I and II, CD44, CD29, CD90, LFA-1, and CD45RB was performed by using flow cytometry. Tri-lineage differentiation assays were performed to confirm MSC multipotency. Recombinant equine IFN-γ was used to stimulate MHC class II negative MSCs in culture, after which expression of MHC class II was re-examined. To assess the ability of MHC class II negative or positive MSCs to stimulate an immune response, modified one-way mixed leukocyte reactions (MLRs) were performed by using MHC-matched and mismatched responder PBLs and stimulator PBLs or MSCs. Proliferation of gated CFSE-labeled CD3+ responder T cells was evaluated via CFSE attenuation by using flow cytometry and reported as the number of cells in the proliferating T-cell gate. Results MSCs varied widely in MHC class II expression despite being homogenous in terms of “stemness” marker expression and ability to undergo trilineage differentiation. Stimulation of MHC class II negative MSCs with IFN-γ resulted in markedly increased expression of MHC class II. MLR results revealed that MHC-mismatched MHC class II-positive MSCs caused significantly increased responder T-cell proliferation compared with MHC-mismatched MHC class II-negative and MHC-matched MSCs, and equivalent to that of the positive control of

  7. Functional Role of FcγRIIB in the Regulation of Mesenchymal Stem Cell Function

    PubMed Central

    Zhu, Tianyi; Chen, Ruohua; Li, Zeng; Tian, Jun; Deng, Changwen; Zhang, Xingxing; Zhang, Koudong; Tong, Linrong; Yu, Yizhi; Bai, Chong

    2016-01-01

    Mesenchymal stem cells (MSCs) derived from bone marrow are plural-potent stem cells with immune regulatory functions. We aimed to evaluate role of FcγRIIB in the regulation of bone marrow-derived MSC function. MSCs were prepared from mouse bone marrow derived from wild-type (WT) or FcγRIIB-deficient (FcγRIIB-/-) mice. MSCs were co-cultured with bone marrow-derived dendritic cells (BMDCs), and BMDC maturation and function were evaluated by flow cytometric analysis and carboxyfluorescein succinimidyl ester-labeled OT-II T-cell addition. An acute asthma model was established by aeresol ovalbumin challenge in mice. Mice received WT or FcγRIIB-/- MSC therapy. Lung function was evaluated by histological examination and cytokine production measurement. mRNA and protein expression levels of target genes were examined by real-time quantitative polymerase chain reactionor western blotting. We found that MSCs derived from bone marrow exhibit a high level of FcγRIIB expression. FcγRIIB deficiency impaired the suppressive function of MSCs, as FcγRIIB deficiency efficiently reversed the inhibitory effect of MSCs on BMDC maturation and function. Additionally, FcγRIIB-/-MSCs were less potent at suppressing asthma in model mice, possibly through reduced expression of Smad2, Smad3, Cox-2, and prostaglandin E2 in FcγRIIB-/-MSCs. FcγRIIB might play an essential role in regulating the inhibitory effects of MSCs derived from bone marrow. PMID:26941575

  8. Functional Role of FcγRIIB in the Regulation of Mesenchymal Stem Cell Function.

    PubMed

    Zhu, Tianyi; Chen, Ruohua; Li, Zeng; Tian, Jun; Deng, Changwen; Zhang, Xingxing; Zhang, Koudong; Tong, Linrong; Yu, Yizhi; Bai, Chong

    2016-01-01

    Mesenchymal stem cells (MSCs) derived from bone marrow are plural-potent stem cells with immune regulatory functions. We aimed to evaluate role of FcγRIIB in the regulation of bone marrow-derived MSC function. MSCs were prepared from mouse bone marrow derived from wild-type (WT) or FcγRIIB-deficient (FcγRIIB-/-) mice. MSCs were co-cultured with bone marrow-derived dendritic cells (BMDCs), and BMDC maturation and function were evaluated by flow cytometric analysis and carboxyfluorescein succinimidyl ester-labeled OT-II T-cell addition. An acute asthma model was established by aeresol ovalbumin challenge in mice. Mice received WT or FcγRIIB-/- MSC therapy. Lung function was evaluated by histological examination and cytokine production measurement. mRNA and protein expression levels of target genes were examined by real-time quantitative polymerase chain reactionor western blotting. We found that MSCs derived from bone marrow exhibit a high level of FcγRIIB expression. FcγRIIB deficiency impaired the suppressive function of MSCs, as FcγRIIB deficiency efficiently reversed the inhibitory effect of MSCs on BMDC maturation and function. Additionally, FcγRIIB-/-MSCs were less potent at suppressing asthma in model mice, possibly through reduced expression of Smad2, Smad3, Cox-2, and prostaglandin E2 in FcγRIIB-/-MSCs. FcγRIIB might play an essential role in regulating the inhibitory effects of MSCs derived from bone marrow. PMID:26941575

  9. SWI/SNF-Mediated Lineage Determination in Mesenchymal Stem Cells Confers Resistance to Osteoporosis.

    PubMed

    Nguyen, Kevin Hong; Xu, Fuhua; Flowers, Stephen; Williams, Edek A J; Fritton, J Christopher; Moran, Elizabeth

    2015-10-01

    Redirecting the adipogenic potential of bone marrow-derived mesenchymal stem cells to other lineages, particularly osteoblasts, is a key goal in regenerative medicine. Controlling lineage selection through chromatin remodeling complexes such as SWI/SNF, which act coordinately to establish new patterns of gene expression, would be a desirable intervention point, but the requirement for the complex in essentially every lineage pathway has generally precluded selectivity. However, a novel approach now appears possible by targeting the subset of SWI/SNF powered by the alternative ATPase, mammalian brahma (BRM). BRM is not required for development, which has hindered understanding of its contributions, but knockdown genetics here, designed to explore the hypothesis that BRM-SWI/SNF has different regulatory roles in different mesenchymal stem cell lineages, shows that depleting BRM from mesenchymal stem cells has a dramatic effect on the balance of lineage selection between osteoblasts and adipocytes. BRM depletion enhances the proportion of cells expressing markers of osteoblast precursors at the expense of cells able to differentiate along the adipocyte lineage. This effect is evident in primary bone marrow stromal cells as well as in established cell culture models. The altered precursor balance has major physiological significance, which becomes apparent as protection against age-related osteoporosis and as reduced bone marrow adiposity in adult BRM-null mice. PMID:26059320

  10. Endotoxins affect bioactivity of chitosan derivatives in cultures of bone marrow-derived human mesenchymal stem cells.

    PubMed

    Lieder, Ramona; Gaware, Vivek S; Thormodsson, Finnbogi; Einarsson, Jon M; Ng, Chuen-How; Gislason, Johannes; Masson, Mar; Petersen, Petur H; Sigurjonsson, Olafur E

    2013-01-01

    Biomaterials research has been expanding over the last decade, in part to provide improved medical devices for the treatment of orthopedic tissue injuries. In the quest to provide the best performance combined with low cost for medical implants, an increasing number of non-chemists have entered the field of biomaterials research without the profound knowledge of chemistry needed to understand the complex interaction mechanisms and characteristics of natural substances. Likewise, non-biologists often lack understanding when it comes to the presence of the contaminating biota frequently found in natural substances. This lack of knowledge by researchers in the field, combined with sensitive in vitro cell-based assays, can lead to inaccurate evaluation of biomaterials. Hence, there should be both an active effort to assemble multi-disciplinary teams and a genuine concern for the possible effects of contamination on in vitro assays. Here, we show that the presence of bacterial endotoxins in chitosan derivatives can result in false-positive results, profoundly altering product performance in in vitro assays. False-positive results through uncritical use of natural substances in vitro can be avoided by proper endotoxin testing and careful evaluation of cytokine secretion patterns. PMID:22947323

  11. Regeneration of articular cartilage by adipose tissue derived mesenchymal stem cells: perspectives from stem cell biology and molecular medicine.

    PubMed

    Wu, Ling; Cai, Xiaoxiao; Zhang, Shu; Karperien, Marcel; Lin, Yunfeng

    2013-05-01

    Adipose-derived stem cells (ASCs) have been discovered for more than a decade. Due to the large numbers of cells that can be harvested with relatively little donor morbidity, they are considered to be an attractive alternative to bone marrow derived mesenchymal stem cells. Consequently, isolation and differentiation of ASCs draw great attention in the research of tissue engineering and regenerative medicine. Cartilage defects cause big therapeutic problems because of their low self-repair capacity. Application of ASCs in cartilage regeneration gives hope to treat cartilage defects with autologous stem cells. In recent years, a lot of studies have been performed to test the possibility of using ASCs to re-construct damaged cartilage tissue. In this article, we have reviewed the most up-to-date articles utilizing ASCs for cartilage regeneration in basic and translational research. Our topic covers differentiation of adipose tissue derived mesenchymal stem cells into chondrocytes, increased cartilage formation by co-culture of ASCs with chondrocytes and enhancing chondrogenic differentiation of ASCs by gene manipulation. PMID:23042088

  12. Hematopoietic and mesenchymal stem cells for the treatment of chronic respiratory diseases: role of plasticity and heterogeneity.

    PubMed

    Conese, Massimo; Piro, Donatella; Carbone, Annalucia; Castellani, Stefano; Di Gioia, Sante

    2014-01-01

    Chronic lung diseases, such as cystic fibrosis (CF), asthma, and chronic obstructive pulmonary disease (COPD) are incurable and represent a very high social burden. Stem cell-based treatment may represent a hope for the cure of these diseases. In this paper, we revise the overall knowledge about the plasticity and engraftment of exogenous marrow-derived stem cells into the lung, as well as their usefulness in lung repair and therapy of chronic lung diseases. The lung is easily accessible and the pathophysiology of these diseases is characterized by injury, inflammation, and eventually by remodeling of the airways. Bone marrow-derived stem cells, including hematopoietic stem/progenitor cells (HSPCs) and mesenchymal stromal (stem) cells (MSCs), encompass a wide array of cell subsets with different capacities of engraftment and injured tissue regenerating potential. Proof-of-principle that marrow cells administered locally may engraft and give rise to specialized epithelial cells has been given, but the efficiency of this conversion is too limited to give a therapeutic effect. Besides the identification of plasticity mechanisms, the characterization/isolation of the stem cell subpopulations represents a major challenge to improving the efficacy of transplantation protocols used in regenerative medicine for lung diseases. PMID:24563632

  13. Clonal Analysis and Hierarchy of Human Bone Marrow Mesenchymal Stem and Progenitor Cells

    PubMed Central

    Lee, C. Chang I.; Christensen, Jared E.; Yoder, Mervin C.; Tarantal, Alice F.

    2009-01-01

    Objective This study was performed to assess adult human bone marrow mesenchymal stem/progenitor cells at a single cell level and to determine a hierarchy based on proliferative potential. Methods Adult bone marrow mesenchymal cells expressing the enhanced green fluorescent protein (EGFP) were sorted as single cells into 24-well plates, each well confirmed with single EGFP-positive cells by fluorescence microscopy, and counted every three days. Colonies derived from single cells were expanded then sorted and evaluated using established differentiation protocols for adipogenic, chondrogenic, and osteogenic lineages. Cells were further analyzed by real-time RT-PCR (PPARγ2, LEP, LPL, LUM, COMP, BIG, CBFA1, IBSP, BGLAP) and immunocytochemistry (PPARγ1/2, Collagen II, Bone Sialoprotein II) specific for tri-lineage differentiation. Results Bone marrow mesenchymal cells were found to contain high proliferative potential-mesenchymal colony-forming cells (HPP-MCFC, 7%), low proliferative potential-mesenchymal colony-forming cells (LPP-MCFC, 29%), mesenchymal cell clusters (MCC, 26%), and mature mesenchymal cells (MMC, 38%). All LPP-MCFC, MCC, and MMC colonies reached senescence at the end of the evaluation period. However, HPP-MCFC continued to grow, showed differentiation toward all three lineages, and demonstrated the capacity to give rise to secondary HPP-MCFC upon replating at a clonal level. Conclusion These findings suggest that there is a low frequency of bone marrow derived HPP-MCFC that can both self-renew at a single cell level and differentiate toward multiple lineages of mesenchymal origin. PMID:19900502

  14. Epigallocatechin-3-gallate protects against tumor necrosis factor alpha induced inhibition of osteogenesis of mesenchymal stem cells.

    PubMed

    Liu, Wei; Fan, Jian-Bo; Xu, Da-Wei; Zhang, Jie; Cui, Zhi-Ming

    2016-03-01

    Anabolic bone accruement through osteogenic differentiation is important for the maintenance of physiological bone mass and often disrupted in various inflammatory diseases. Epigallocatechin-3-gallate, as an antioxidant and anti-inflammatory agent, has been suggested for potential therapeutic use in this context, possibly by the inhibition of bone resorption as well as the enhancement of bone formation through directly activating osteoblast differentiation. However, the reported effects of epigallocatechin-3-gallate modulating osteoblast differentiation are mixed, and the underlying molecular mechanism is still elusive. Moreover, there is limited information regarding the effects of epigallocatechin-3-gallate on osteogenic potential of mesenchymal stem cell in inflammation. Here, we examined the in vitro osteogenic differentiation of human mesenchymal stem cells. We found that the cell viability and osteoblast differentiation of human bone marrow-derived mesenchymal stem cells are significantly inhibited by inflammatory cytokine TNFα treatment. Epigallocatechin-3-gallate is able to enhance the cell viability and osteoblast differentiation of mesenchymal stem cells and is capable of reversing the TNFα-induced inhibition. Notably, only low doses of epigallocatechin-3-gallate have such benefits, which potentially act through the inhibition of NF-κB signaling that is stimulated by TNFα. These data altogether clarify the controversy on epigallocatechin-3-gallate promoting osteoblast differentiation and further provide molecular basis for the putative clinical use of epigallocatechin-3-gallate in stem cell-based bone regeneration for inflammatory bone loss diseases, such as rheumatoid arthritis and prosthetic osteolysis. PMID:26748399

  15. Viscoelastic properties of human mesenchymally-derived stem cells and primary osteoblasts, chondrocytes, and adipocytes

    PubMed Central

    Darling, Eric M.; Topel, Matthew; Zauscher, Stefan; Vail, Thomas P.; Guilak, Farshid

    2010-01-01

    The mechanical properties of single cells play important roles in regulating cell-matrix interactions, potentially influencing the process of mechanotransduction. Recent studies also suggest that cellular mechanical properties may provide novel biological markers, or “biomarkers,” of cell phenotype, reflecting specific changes that occur with disease, differentiation, or cellular transformation. Of particular interest in recent years has been the identification of such biomarkers that can be used to determine specific phenotypic characteristics of stem cells that separate them from primary, differentiated cells. The goal of this study was to determine the elastic and viscoelastic properties of three primary cell types of mesenchymal lineage (chondrocytes, osteoblasts, and adipocytes) and to test the hypothesis that primary differentiated cells exhibit distinct mechanical properties compared to adult stem cells (adipose-derived or bone marrow-derived mesenchymal stem cells). In an adherent, spread configuration, chondrocytes, osteoblasts, and adipocytes all exhibited significantly different mechanical properties, with osteoblasts being stiffer than chondrocytes and both being stiffer than adipocytes. Adipose-derived and mesenchymal stem cells exhibited similar properties to each other, but were mechanically distinct from primary cells, particularly when comparing a ratio of elastic to relaxed moduli. These findings will help more accurately model the cellular mechanical environment in mesenchymal tissues, which could assist in describing injury thresholds and disease progression or even determining the influence of mechanical loading for tissue engineering efforts. Furthermore, the identification of mechanical properties distinct to stem cells could result in more successful sorting procedures to enrich multipotent progenitor cell populations. PMID:17825308

  16. Monolayered mesenchymal stem cells repair scarred myocardium after myocardial infarction.

    PubMed

    Miyahara, Yoshinori; Nagaya, Noritoshi; Kataoka, Masaharu; Yanagawa, Bobby; Tanaka, Koichi; Hao, Hiroyuki; Ishino, Kozo; Ishida, Hideyuki; Shimizu, Tatsuya; Kangawa, Kenji; Sano, Shunji; Okano, Teruo; Kitamura, Soichiro; Mori, Hidezo

    2006-04-01

    Mesenchymal stem cells are multipotent cells that can differentiate into cardiomyocytes and vascular endothelial cells. Here we show, using cell sheet technology, that monolayered mesenchymal stem cells have multipotent and self-propagating properties after transplantation into infarcted rat hearts. We cultured adipose tissue-derived mesenchymal stem cells characterized by flow cytometry using temperature-responsive culture dishes. Four weeks after coronary ligation, we transplanted the monolayered mesenchymal stem cells onto the scarred myocardium. After transplantation, the engrafted sheet gradually grew to form a thick stratum that included newly formed vessels, undifferentiated cells and few cardiomyocytes. The mesenchymal stem cell sheet also acted through paracrine pathways to trigger angiogenesis. Unlike a fibroblast cell sheet, the monolayered mesenchymal stem cells reversed wall thinning in the scar area and improved cardiac function in rats with myocardial infarction. Thus, transplantation of monolayered mesenchymal stem cells may be a new therapeutic strategy for cardiac tissue regeneration. PMID:16582917

  17. Mesenchymal Stem Cells Migration Homing and Tracking

    PubMed Central

    Verfaillie, Catherine M.

    2013-01-01

    In this review, we discuss the migration and homing ability of mesenchymal stem cells (MSCs) and MSC-like cells and factors influencing this. We also discuss studies related to the mechanism of migration and homing and the approaches undertaken to enhance it. Finally, we describe the different methods available and frequently used to track and identify the injected cells in vivo. PMID:24194766

  18. Liver-derived human mesenchymal stem cells: a novel therapeutic source for liver diseases.

    PubMed

    Wang, Yini; Yu, Xiaopeng; Chen, Ermei; Li, Lanuan

    2016-01-01

    Mesenchymal stem cells (MSCs) represent an attractive cell type for research and therapy due to their ability to proliferate, differentiate, modulate immune reactions, and secrete trophic factors. MSCs exist in a multitude of tissues, including bone marrow, umbilical cord, and adipose tissues. Moreover, MSCs have recently been isolated from the liver. Compared with other MSC types, liver-derived human MSCs (LHMSCs) possess general morphologies, immune functions, and differentiation capacities. Interestingly, LHMCSs produce higher levels of pro-angiogenic, anti-inflammatory, and anti-apoptotic cytokines than those of bone marrow-derived MSCs. Thus, these cells may be a promising therapeutic source for liver diseases. This paper summarizes the biological characteristics of LHMSCs and their potential benefits and risks for the treatment of liver diseases. PMID:27176654

  19. Mesenchymal Stem Cells for Regenerative Therapy: Optimization of Cell Preparation Protocols

    PubMed Central

    2014-01-01

    Administration of bone marrow-derived mesenchymal stem cells (MSCs) is an innovative approach for the treatment of a range of diseases that are not curable by current therapies including heart failure. A number of clinical trials have been completed and many others are ongoing; more than 2,000 patients worldwide have been administered with culture-expanded allogeneic or autologous MSCs for the treatment of various diseases, showing feasibility and safety (and some efficacy) of this approach. However, protocols for isolation and expansion of donor MSCs vary widely between these trials, which could affect the efficacy of the therapy. It is therefore important to develop international standards of MSC production, which should be evidence-based, regulatory authority-compliant, of good medical practice grade, cost-effective, and clinically practical, so that this innovative approach becomes an established widely adopted treatment. This review article summarizes protocols to isolate and expand bone marrow-derived MSCs in 47 recent clinical trials of MSC-based therapy, which were published after 2007 onwards and provided sufficient methodological information. Identified issues and possible solutions associated with the MSC production methods, including materials and protocols for isolation and expansion, are discussed with reference to relevant experimental evidence with aim of future clinical success of MSC-based therapy. PMID:24511552

  20. RUNX1 is essential for mesenchymal stem cell proliferation and myofibroblast differentiation.

    PubMed

    Kim, Woosook; Barron, David A; San Martin, Rebeca; Chan, Keith S; Tran, Linda L; Yang, Feng; Ressler, Steven J; Rowley, David R

    2014-11-18

    Myofibroblasts are a key cell type in wound repair, cardiovascular disease, and fibrosis and in the tumor-promoting microenvironment. The high accumulation of myofibroblasts in reactive stroma is predictive of the rate of cancer progression in many different tumors, yet the cell types of origin and the mechanisms that regulate proliferation and differentiation are unknown. We report here, for the first time to our knowledge, the characterization of normal human prostate-derived mesenchymal stem cells (MSCs) and the TGF-β1-regulated pathways that modulate MSC proliferation and myofibroblast differentiation. Human prostate MSCs combined with prostate cancer cells expressing TGF-β1 resulted in commitment to myofibroblasts. TGF-β1-regulated runt-related transcription factor 1 (RUNX1) was required for cell cycle progression and proliferation of progenitors. RUNX1 also inhibited, yet did not block, differentiation. Knockdown of RUNX1 in prostate or bone marrow-derived MSCs resulted in cell cycle arrest, attenuated proliferation, and constitutive differentiation to myofibroblasts. These data show that RUNX1 is a key transcription factor for MSC proliferation and cell fate commitment in myofibroblast differentiation. This work also shows that the normal human prostate gland contains tissue-derived MSCs that exhibit multilineage differentiation similar to bone marrow-derived MSCs. Targeting RUNX1 pathways may represent a therapeutic approach to affect myofibroblast proliferation and biology in multiple disease states. PMID:25313057

  1. Use of mesenchymal stem cells for cutaneous repair and skin substitute elaboration.

    PubMed

    Laverdet, B; Micallef, L; Lebreton, C; Mollard, J; Lataillade, J-J; Coulomb, B; Desmoulière, A

    2014-04-01

    Human mesenchymal stem cells (MSCs) are a heterogeneous population of fibroblast-like cells, which are present in different locations, including bone marrow, adipose tissue, extra-foetal tissues, gingiva and dermis. MSCs, which present multipotency capacities, important expansive potential and immunotolerance properties, remain an attractive tool for tissue repair and regenerative medicine. Currently, several studies and clinical trials highlight the use of MSCs in cutaneous repair underlining that their effects are essentially due to the numerous factors that they release. MSCs are also used in skin substitute development. In this study, we will first discuss the different sources of MSCs actually available. We will then present results showing that bone marrow-derived MSCs prepared according to Good Manufacturing Practices and included in a dermal equivalent are able to promote appropriate epidermis growth and differentiation. These data demonstrate that bone marrow-derived MSCs represent a satisfactory alternative to dermal fibroblasts in order to develop skin substitute. In addition, MSCs could provide a useful alternative to sustain epidermis development and to promote wound healing. PMID:24661975

  2. [Glioma treatment strategies using mesenchymal stem cells].

    PubMed

    Namba, Hiroki

    2010-10-01

    Because of the growth characteristics of malignant gliomas that are highly invasive and deeply infiltrate the surrounding brain area; the surgical resection of these gliomas with preservation of neural functions is almost always noncurative. The residual tumor cells are usually resistant to standard adjuvant radiochemotherapy, and therefore, the tumors inevitably recur after a certain period and finally cause the death of the patients. Neural and mesenchymal stem cells have been extensively studied for the development of new strategies for treating malignant gliomas because of these cells possess the intrinsic property of homing toward tumor cells. By using neural and mesenchymal stem cells as vehicles for drug carriers, it is possible to deliver anticancer drugs to the tumor cells that infiltrate functioning normal brain tissue and are difficult to remove. Several cytokines and suicide genes have been tested, and promising results have been reported in animal brain tumor models. However, further studies involving safety issues such as secondary cancer formation are required before human trials of stem cell therapies. In the present paper, the author has reviewed the recent concepts involved in the treatment of malignant gliomas with stem cells, especially mesenchymal stem cells that are much easier to obtain from the patients themselves. PMID:20940507

  3. Colonization of collagen scaffolds by adipocytes derived from mesenchymal stem cells of the common marmoset monkey

    SciTech Connect

    Bernemann, Inga; Mueller, Thomas; Blasczyk, Rainer; Glasmacher, Birgit; Hofmann, Nicola

    2011-07-29

    Highlights: {yields} Marmoset bone marrow-derived MSCs differentiate in suspension into adipogenic, osteogenic and chondrogenic lineages. {yields} Marmoset MSCs integrate in collagen type I scaffolds and differentiate excellently into adipogenic cells. {yields} Common marmoset monkey is a suitable model for soft tissue engineering in human regenerative medicine. -- Abstract: In regenerative medicine, human cell replacement therapy offers great potential, especially by cell types differentiated from immunologically and ethically unproblematic mesenchymal stem cells (MSCs). In terms of an appropriate carrier material, collagen scaffolds with homogeneous pore size of 65 {mu}m were optimal for cell seeding and cultivating. However, before clinical application and transplantation of MSC-derived cells in scaffolds, the safety and efficiency, but also possible interference in differentiation due to the material must be preclinically tested. The common marmoset monkey (Callithrix jacchus) is a preferable non-human primate animal model for this aim due to its genetic and physiological similarities to the human. Marmoset bone marrow-derived MSCs were successfully isolated, cultured and differentiated in suspension into adipogenic, osteogenic and chondrogenic lineages by defined factors. The differentiation capability could be determined by FACS. Specific marker genes for all three cell types could be detected by RT-PCR. Furthermore, MSCs seeded on collagen I scaffolds differentiated in adipogenic lineage showed after 28 days of differentiation high cell viability and homogenous distribution on the material which was validated by calcein AM and EthD staining. As proof of adipogenic cells, the intracellular lipid vesicles in the cells were stained with Oil Red O. The generation of fat vacuoles was visibly extensive distinguishable and furthermore determined on the molecular level by expression of specific marker genes. The results of the study proved both the differential

  4. Mesenchymal stem cells induce epithelial proliferation within the inflamed stomach.

    PubMed

    Donnelly, Jessica M; Engevik, Amy; Feng, Rui; Xiao, Chang; Boivin, Gregory P; Li, Jing; Houghton, JeanMarie; Zavros, Yana

    2014-06-15

    Bone marrow-derived mesenchymal stem cells (MSCs) sustain cancer cells by creating a microenvironment favorable for tumor growth. In particular, MSCs have been implicated in gastric cancer development. There is extensive evidence suggesting that Hedgehog signaling regulates tumor growth. However, very little is known regarding the precise roles of Hedgehog signaling and MSCs in tumor development within the stomach. The current study tests that hypothesis that Sonic Hedgehog (Shh), secreted from MSCs, provides a proliferative stimulus for the gastric epithelium in the presence of inflammation. Red fluorescent protein-expressing MSCs transformed in vitro (stMSCs) were transduced with lentiviral constructs containing a vector control (stMSC(vect)) or short hairpin RNA (shRNA) targeting the Shh gene (stMSC(ShhKO)). Gastric submucosal transplantation of wild-type MSCs (wtMSCs), wild-type MSCs overexpressing Shh (wtMSC(Shh)), stMSC(vect), or stMSC(ShhKO) cells in C57BL/6 control (BL/6) or gastrin-deficient (GKO) mice was performed and mice analyzed 30 and 60 days posttransplantation. Compared with BL/6 mice transplanted with wtMSC(Shh) and stMSC(vect) cells, inflamed GKO mice developed aggressive gastric tumors. Tumor development was not observed in mouse stomachs transplanted with wtMSC or stMSC(ShhKO) cells. Compared with stMSC(ShhKO)-transplanted mice, within the inflamed GKO mouse stomach, Shh-expressing stMSC(vect)- and wtMSC(Shh)-induced proliferation of CD44-positive cells. CD44-positive cells clustered in gland-like structures within the tumor stroma and were positive for Patched (Ptch) expression. We conclude that Shh, secreted from MSCs, provides a proliferative stimulus for the gastric epithelium that is associated with tumor development, a response that is sustained by chronic inflammation. PMID:24789207

  5. Mesenchymal stem cells enhance wound healing through differentiation and angiogenesis.

    PubMed

    Wu, Yaojiong; Chen, Liwen; Scott, Paul G; Tredget, Edward E

    2007-10-01

    Although chronic wounds are common, treatment for these disabling conditions remains limited and largely ineffective. In this study, we examined the benefit of bone marrow-derived mesenchymal stem cells (BM-MSCs) in wound healing. Using an excisional wound splinting model, we showed that injection around the wound and application to the wound bed of green fluorescence protein (GFP)(+) allogeneic BM-MSCs significantly enhanced wound healing in normal and diabetic mice compared with that of allogeneic neonatal dermal fibroblasts or vehicle control medium. Fluorescence-activated cell sorting analysis of cells derived from the wound for GFP-expressing BM-MSCs indicated engraftments of 27% at 7 days, 7.6% at 14 days, and 2.5% at 28 days of total BM-MSCs administered. BM-MSC-treated wounds exhibited significantly accelerated wound closure, with increased re-epithelialization, cellularity, and angiogenesis. Notably, BM-MSCs, but not CD34(+) bone marrow cells in the wound, expressed the keratinocyte-specific protein keratin and formed glandular structures, suggesting a direct contribution of BM-MSCs to cutaneous regeneration. Moreover, BM-MSC-conditioned medium promoted endothelial cell tube formation. Real-time polymerase chain reaction and Western blot analysis revealed high levels of vascular endothelial growth factor and angiopoietin-1 in BM-MSCs and significantly greater amounts of the proteins in BM-MSC-treated wounds. Thus, our data suggest that BM-MSCs promote wound healing through differentiation and release of proangiogenic factors. Disclosure of potential conflicts of interest is found at the end of this article. PMID:17615264

  6. Comparison of mesenchymal stem cells and leukocytes from Large White and Göttingen Minipigs: Clues for stem cell-based immunomodulatory therapies.

    PubMed

    Álvarez, Verónica; Sánchez-Margallo, Francisco-Miguel; Blázquez, Rebeca; Tarazona, Raquel; Casado, Javier G

    2016-10-15

    The mesenchymal stem cells (MSCs) are one of the most promising cell types for human and veterinary use and their therapeutic effect is associated with their immunomodulatory properties. Farm animal models, such as pigs, have become a valuable tool to evaluate the safety and efficacy of adoptively transferred MSCs in the setting of veterinary medicine. In order to evaluate the immunomodulatory effect of stem cell-based therapies in porcine breeds, a deep analysis and comparison of MSCs and leukocyte subsets are absolutely necessary. Here we provide a detailed analysis of bone-marrow derived MSCs and leukocyte subsets from Large White pigs and Göttingen Minipigs. Significant differences were observed between the two pig breeds in terms of T cell subsets that need to be considered for immune monitoring of stem cell-based therapies. PMID:27590427

  7. Human Wharton's Jelly Mesenchymal Stem Cells plasticity augments scar-free skin wound healing with hair growth.

    PubMed

    Sabapathy, Vikram; Sundaram, Balasubramanian; V M, Sreelakshmi; Mankuzhy, Pratheesh; Kumar, Sanjay

    2014-01-01

    Human mesenchymal stem cells (MSCs) are a promising candidate for cell-based transplantation and regenerative medicine therapies. Thus in the present study Wharton's Jelly Mesenchymal Stem Cells (WJ-MSCs) have been derived from extra embryonic umbilical cord matrix following removal of both arteries and vein. Also, to overcome the clinical limitations posed by fetal bovine serum (FBS) supplementation because of xenogeneic origin of FBS, usual FBS cell culture supplement has been replaced with human platelet lysate (HPL). Apart from general characteristic features of bone marrow-derived MSCs, wharton jelly-derived MSCs have the ability to maintain phenotypic attributes, cell growth kinetics, cell cycle pattern, in vitro multilineage differentiation plasticity, apoptotic pattern, normal karyotype-like intrinsic mesenchymal stem cell properties in long-term in vitro cultures. Moreover, the WJ-MSCs exhibited the in vitro multilineage differentiation capacity by giving rise to differentiated cells of not only mesodermal lineage but also to the cells of ectodermal and endodermal lineage. Also, WJ-MSC did not present any aberrant cell state upon in vivo transplantation in SCID mice and in vitro soft agar assays. The immunomodulatory potential assessed by gene expression levels of immunomodulatory factors upon exposure to inflammatory cytokines in the fetal WJ-MSCs was relatively higher compared to adult bone marrow-derived MSCs. WJ-MSCs seeded on decellularized amniotic membrane scaffold transplantation on the skin injury of SCID mice model demonstrates that combination of WJ-MSCs and decellularized amniotic membrane scaffold exhibited significantly better wound-healing capabilities, having reduced scar formation with hair growth and improved biomechanical properties of regenerated skin compared to WJ-MSCs alone. Further, our experimental data indicate that indocyanin green (ICG) at optimal concentration can be resourcefully used for labeling of stem cells and in vivo

  8. Morphological and functional characterization of femoral head drilling-derived mesenchymal stem cells.

    PubMed

    Tatu, Romulus Fabian; Anuşca, Dan Nelu; Groza, Sabine Ştefania; Marusciac, Laura; Bojin, Florina Maria; Tatu, Carmen; Hurmuz, Mihai; Păunescu, Virgil

    2014-01-01

    Adult mesenchymal stem cells (MSCs) were primary identified as bone marrow-derived cells, fibroblast-like morphology, and adherent to plastic surfaces of in vitro culture plate. Their identification criteria evolved in time to a well-established panel of markers (expression of CD73, CD90, and CD105) and functional characteristics (adipogenic, osteogenic, and chondrogenic trilineage differentiation ability), which can be applied to adult mesenchymal stem cells obtained from other tissue sources. We tried to assess the potential stemness of femoral head drilling-derived cells as a new source of mesenchymal stem cells (FH-MSCs). For this purpose, we used the morphological and ultrastructural characteristics defined by scanning and transmission electron microscopy and spindle-shape cellular body, fibroblast-like, with few thick elongations (lamellipodia) and numerous fine, thin cytoplasmic projections (filopodia) that extend beyond the edge of lamellipodia. Immunophenotypical analysis was performed by flow cytometry and immunocytochemical methods and we showed that FH-MSCs share the characteristic markers of MSCs, expressing CD73, CD90, CD105, and being positive for vimentin, and c-kit (CD117). Proliferation rate of these cells was moderate, as revealed by Ki67 immunostaining. Regarding the functional characteristics of FH-MSCs, after appropriate time of induction in specific culture media, the cells were able to prove their trilineage potential and differentiated towards adipocytic, osteogenic, and chondrogenic lineage, as revealed by immunofluorescent staining. We may conclude that femoral head drilling-derived cells can be used as a novel source of stem cells, and employed in diverse clinical settings. PMID:25611275

  9. Nuclear Export of Smads by RanBP3L Regulates Bone Morphogenetic Protein Signaling and Mesenchymal Stem Cell Differentiation

    PubMed Central

    Chen, Fenfang; Lin, Xia; Xu, Pinglong; Zhang, Zhengmao; Chen, Yanzhen; Wang, Chao; Han, Jiahuai; Zhao, Bin; Xiao, Mu

    2015-01-01

    Bone morphogenetic proteins (BMPs) play vital roles in regulating stem cell maintenance and differentiation. BMPs can induce osteogenesis and inhibit myogenesis of mesenchymal stem cells. Canonical BMP signaling is stringently controlled through reversible phosphorylation and nucleocytoplasmic shuttling of Smad1, Smad5, and Smad8 (Smad1/5/8). However, how the nuclear export of Smad1/5/8 is regulated remains unclear. Here we report that the Ran-binding protein RanBP3L acts as a nuclear export factor for Smad1/5/8. RanBP3L directly recognizes dephosphorylated Smad1/5/8 and mediates their nuclear export in a Ran-dependent manner. Increased expression of RanBP3L blocks BMP-induced osteogenesis of mouse bone marrow-derived mesenchymal stem cells and promotes myogenic induction of C2C12 mouse myoblasts, whereas depletion of RanBP3L expression enhances BMP-dependent stem cell differentiation activity and transcriptional responses. In conclusion, our results demonstrate that RanBP3L, as a nuclear exporter for BMP-specific Smads, plays a critical role in terminating BMP signaling and regulating mesenchymal stem cell differentiation. PMID:25755279

  10. Mesenchymal stem cell-based therapy.

    PubMed

    Mundra, Vaibhav; Gerling, Ivan C; Mahato, Ram I

    2013-01-01

    Mesenchymal stem cells (MSCs) are multipotent adult stem cells which have self-renewal capacity and differentiation potential into several mesenchymal lineages including bones, cartilages, adipose tissues and tendons. MSCs may repair tissue injuries and prevent immune cell activation and proliferation. Immunomodulation and secretion of growth factors by MSCs have led to realizing the true potential of MSC-based cell therapy. The use of MSCs as immunomodulators has been explored in cell/organ transplant, tissue repair, autoimmune diseases, and prevention of graft vs host disease (GVHD). This review focuses on the clinical applications of MSC-based cell therapy, with particular emphasis on islet transplantation for treating type I diabetes. PMID:23215004

  11. Mesenchymal Stem Cell-Based Therapy

    PubMed Central

    Mundra, Vaibhav; Gerling, Ivan C.; Mahato, Ram I.

    2012-01-01

    Mesenchymal stem cells (MSCs) are multipotent adult stem cells which have self-renewal capacity and differentiation potential into several mesenchymal lineages including bones, cartilages, adipose tissues and tendons. MSCs may repair tissue injuries and prevent immune cell activation and proliferation. Immunomodulation and secretion of growth factors by MSCs have led to realizing the true potential of MSC-based cell therapy. The use of MSCs as immunomdulators has been explored in cell/organ transplant, tissue repair, autoimmune diseases and prevention of graft vs. host disease (GVHD). This review focuses on the clinical applications of MSC-based cell therapy, with particular emphasis on islet transplantation for treating type I diabetes. PMID:23215004

  12. Bone Marrow-Derived Cells as a Therapeutic Approach to Optic Nerve Diseases.

    PubMed

    Mesentier-Louro, Louise A; Zaverucha-do-Valle, Camila; Rosado-de-Castro, Paulo H; Silva-Junior, Almir J; Pimentel-Coelho, Pedro M; Mendez-Otero, Rosalia; Santiago, Marcelo F

    2016-01-01

    Following optic nerve injury associated with acute or progressive diseases, retinal ganglion cells (RGCs) of adult mammals degenerate and undergo apoptosis. These diseases have limited therapeutic options, due to the low inherent capacity of RGCs to regenerate and due to the inhibitory milieu of the central nervous system. Among the numerous treatment approaches investigated to stimulate neuronal survival and axonal extension, cell transplantation emerges as a promising option. This review focuses on cell therapies with bone marrow mononuclear cells and bone marrow-derived mesenchymal stem cells, which have shown positive therapeutic effects in animal models of optic neuropathies. Different aspects of available preclinical studies are analyzed, including cell distribution, potential doses, routes of administration, and mechanisms of action. Finally, published and ongoing clinical trials are summarized. PMID:26649049

  13. Bone Marrow-Derived Cells as a Therapeutic Approach to Optic Nerve Diseases

    PubMed Central

    Mesentier-Louro, Louise A.; Zaverucha-do-Valle, Camila; Rosado-de-Castro, Paulo H.; Silva-Junior, Almir J.; Pimentel-Coelho, Pedro M.; Mendez-Otero, Rosalia; Santiago, Marcelo F.

    2016-01-01

    Following optic nerve injury associated with acute or progressive diseases, retinal ganglion cells (RGCs) of adult mammals degenerate and undergo apoptosis. These diseases have limited therapeutic options, due to the low inherent capacity of RGCs to regenerate and due to the inhibitory milieu of the central nervous system. Among the numerous treatment approaches investigated to stimulate neuronal survival and axonal extension, cell transplantation emerges as a promising option. This review focuses on cell therapies with bone marrow mononuclear cells and bone marrow-derived mesenchymal stem cells, which have shown positive therapeutic effects in animal models of optic neuropathies. Different aspects of available preclinical studies are analyzed, including cell distribution, potential doses, routes of administration, and mechanisms of action. Finally, published and ongoing clinical trials are summarized. PMID:26649049

  14. Human pluripotent stem cell-derived mesenchymal stem cells prevent allergic airway inflammation in mice.

    PubMed

    Sun, Yue-Qi; Deng, Meng-Xia; He, Jia; Zeng, Qing-Xiang; Wen, Weiping; Wong, David S H; Tse, Hung-Fat; Xu, Geng; Lian, Qizhou; Shi, Jianbo; Fu, Qing-Ling

    2012-12-01

    We previously found that mesenchymal stem cells (MSCs) derived from human-induced pluripotent stem cells (iPSCs) exerted immunomodulatory effects on Th2-mediated allergic rhinitis in vitro. However, their contribution to the asthma and allergic rhinitis in animal models remains unclear. In this study, we developed a mouse model of ovalbumin (OVA)-induced allergic inflammation in both the upper and lower airways and evaluated the effects of the systemic administration of human iPSC-MSCs and bone marrow-derived MSCs (BM-MSCs) on allergic inflammation. Our results showed that treatments with both the iPSC-MSCs and BM-MSCs before the challenge phase protected the animals from the majority of allergy-specific pathological changes. This protection included an inhibition of inflammatory cell infiltration and mucus production in the lung, a reduction in eosinophil infiltration in the nose, and a decrease in inflammatory cell infiltration in both the bronchoalveolar and nasal lavage fluids. In addition, treatment with iPSC-MSCs or BM-MSCs before the challenge phase resulted in reduced serum levels of Th2 immunoglobulins (e.g., IgE) and decreased levels of Th2 cytokines including interleukin (IL)-4, IL-5, or IL-13 in the bronchoalveolar and/or nasal lavage fluids. Similar therapeutic effects were observed when the animals were pretreated with human iPSC-MSCs before the sensitization phase. These data suggest that iPSC-MSCs may be used as an alternative strategy to adult MSCs in the treatment of asthma and allergic rhinitis. PMID:22987325

  15. Mesenchymal stem cells expressing neural antigens instruct a neurogenic cell fate on neural stem cells.

    PubMed

    Croft, Adam P; Przyborski, Stefan A

    2009-04-01

    The neurogenic response to injury in the postnatal brain is limited and insufficient for restoration of function. Recent evidence suggests that transplantation of mesenchymal stem cells (MSCs) into the injured brain is associated with improved functional recovery, mediated in part through amplification in the endogenous neurogenic response to injury. In the current study we investigate the interactions between bone marrow-derived MSCs and embryonic neural stem cells (NSCs) plus their differentiated progeny using an in vitro co-culture system. Two populations of MSCs were used, MSCs induced to express neural antigens (nestin+, Tuj-1+, GFAP+) and neural antigen negative MSCs. Following co-culture of induced MSCs with differentiating NSC/progenitor cells a significant increase in Tuj-1+ neurons was detected compared to co-cultures of non-induced MSCs in which an increase in astrocyte (GFAP+) differentiation was observed. The effect was mediated by soluble interactions between the two cell populations and was independent of any effect on cell death and proliferation. Induced and non-induced MSCs also promoted the survival of Tuj-1+ cell progeny in long-term cultures and both promoted axonal growth, an effect also seen in differentiating neuroblastoma cells. Therefore, MSCs provide instructive signals that are able to direct the differentiation of NSCs and promote axonal development in neuronal progeny. The data indicates that the nature of MSC derived signals is dependent not only on their microenvironment but on the developmental status of the MSCs. Pre-manipulation of MSCs prior to transplantation in vivo may be an effective means of enhancing the endogenous neurogenic response to injury. PMID:19159625

  16. Effect of heparin on the biological properties and molecular signature of human mesenchymal stem cells.

    PubMed

    Ling, Ling; Camilleri, Emily T; Helledie, Torben; Samsonraj, Rebekah M; Titmarsh, Drew M; Chua, Ren Jie; Dreesen, Oliver; Dombrowski, Christian; Rider, David A; Galindo, Mario; Lee, Ian; Hong, Wanjin; Hui, James H; Nurcombe, Victor; van Wijnen, Andre J; Cool, Simon M

    2016-01-15

    Chronic use of heparin as an anti-coagulant for the treatment of thrombosis or embolism invokes many adverse systemic events including thrombocytopenia, vascular reactions and osteoporosis. Here, we addressed whether adverse effects might also be directed to mesenchymal stem cells that reside in the bone marrow compartment. Harvested human bone marrow-derived mesenchymal stem cells (hMSCs) were exposed to varying doses of heparin and their responses profiled. At low doses (<200 ng/ml), serial passaging with heparin exerted a variable effect on hMSC proliferation and multipotentiality across multiple donors, while at higher doses (≥ 100 μg/ml), heparin supplementation inhibited cell growth and increased both senescence and cell size. Gene expression profiling using cDNA arrays and RNA-seq analysis revealed pleiotropic effects of low-dose heparin on signaling pathways essential to hMSC growth and differentiation (including the TGFβ/BMP superfamily, FGFs, and Wnts). Cells serially passaged in low-dose heparin possess a donor-dependent gene signature that reflects their altered phenotype. Our data indicate that heparin supplementation during the culturing of hMSCs can alter their biological properties, even at low doses. This warrants caution in the application of heparin as a culture supplement for the ex vivo expansion of hMSCs. It also highlights the need for careful evaluation of the bone marrow compartment in patients receiving chronic heparin treatment. PMID:26484394

  17. A Refractory Celiac Patient Successfully Treated With Mesenchymal Stem Cell Infusions.

    PubMed

    Ciccocioppo, Rachele; Gallia, Alessandra; Avanzini, Maria A; Betti, Elena; Picone, Cristina; Vanoli, Alessandro; Paganini, Chiara; Biagi, Federico; Maccario, Rita; Corazza, Gino R

    2016-06-01

    Type II refractory celiac disease (RCD), as defined according to the amount of aberrant intraepithelial lymphocytes, is a condition characterized by severe malabsorption syndrome and poor prognosis, with no effective treatment. Based on the regenerative and immunomodulatory properties of mesenchymal stem cells (MSCs), we investigated the feasibility, safety, and efficacy of serial infusions of autologous bone marrow-derived MSCs in a 51-year-old woman with type II RCD. Mesenchymal stem cells were isolated, expanded, and characterized following standard protocols. Monitoring of the patient's malabsorption indexes, mucosal architecture, and percentage of aberrant intraepithelial lymphocytes was scheduled for the time of enrollment, at each infusion, and after 6 months. Determination of mucosal expression of interleukin (IL)-15 and its receptor was also performed. Expansion of MSCs was feasible, and the patient underwent 4 systemic infusions of 2 × 10(6) MSCs/kg body weight 4 months apart, without adverse effects. During the treatment period, she experienced gradual and durable amelioration of her general condition, with normalization of stool frequency, body mass index, laboratory test results, and mucosal architecture. Remarkably, the expression of IL-15 and its receptor almost completely disappeared. Thus, treatment of RCD with serial MSC infusions seems promising, leading to recovery from the life-threatening condition while blocking the IL-15 pathogenic pathway. PMID:27087453

  18. Inhibition of adipocytogenesis by canonical WNT signaling in human mesenchymal stem cells

    SciTech Connect

    Shen, Longxiang; Glowacki, Julie; Zhou, Shuanhu

    2011-08-01

    The WNT signaling pathway plays important roles in the self-renewal and differentiation of mesenchymal stem cells (MSCs). Little is known about WNT signaling in adipocyte differentiation of human MSCs. In this study, we tested the hypothesis that canonical and non-canonical WNTs differentially regulate in vitro adipocytogenesis in human MSCs. The expression of adipocyte gene PPAR{gamma}2, lipoprotein lipase, and adipsin increased during adipocytogenesis of hMSCs. Simultaneously, the expression of canonical WNT2, 10B, 13, and 14 decreased, whereas non-canonical WNT4 and 11 increased, and WNT5A was unchanged. A small molecule WNT mimetic, SB-216763, increased accumulation of {beta}-catenin protein, inhibited induction of WNT4 and 11 and inhibited adipocytogenesis. In contrast, knockdown of {beta}-catenin with siRNA resulted in spontaneous adipocytogenesis. These findings support the view that canonical WNT signaling inhibits and non-canonical WNT signaling promotes adipocytogenesis in adult human marrow-derived mesenchymal stem cells.

  19. Mesenchymal stem cells preconditioned with trimetazidine promote neovascularization of hearts under hypoxia/reoxygenation injury

    PubMed Central

    Hu, Xiaowu; Yang, Junjie; Wang, Ying; Zhang, You; Ii, Masaaki; Shen, Zhenya; Hui, Jie

    2015-01-01

    Background: Cell-based angiogenesis is a promising treatment for ischemic diseases; however, survival of implanted cells is impaired by the ischemic microenvironment. In this study, mesenchymal stem cells (MSCs) for cell transplantation were preconditioned with trimetazidine (TMZ). We hypothesized that TMZ enhances the survival rate of MSCs under hypoxic stimuli through up-regulation of HIF1-α. Methods and results: Bone marrow-derived rat mesenchymal stem cells were preconditioned with 10 μM TMZ for 6 h. TMZ preconditioning of MSCs remarkably increased cell viability and the expression of HIF1-α and Bcl-2, when cells were under hypoxia/reoxygenation (H/R) stimuli. But the protective effects of TMZ were abolished after knocking down of HIF-1α. Three days after implantation of the cells into the peri-ischemic zone of rat myocardial ischemia-reperfusion (I/R) injury model, survival of the TMZ-preconditioned MSCs was high. Furthermore, capillary density and cardiac function were significantly better in the rats implanted with TMZ-preconditioned MSCs 28 days after cell injection. Conclusions: TMZ preconditioning increased the survival rate of MSCs, through up-regulation of HIF1-α, thus contributing to neovascularization and improved cardiac function of rats subjected to myocardial I/R injury. PMID:26629255

  20. Cartilage Engineering from Mesenchymal Stem Cells

    NASA Astrophysics Data System (ADS)

    Goepfert, C.; Slobodianski, A.; Schilling, A. F.; Adamietz, P.; Pörtner, R.

    Mesenchymal progenitor cells known as multipotent mesenchymal stromal cells or mesenchymal stem cells (MSC) have been isolated from various tissues. Since they are able to differentiate along the mesenchymal lineages of cartilage and bone, they are regarded as promising sources for the treatment of skeletal defects. Tissue regeneration in the adult organism and in vitro engineering of tissues is hypothesized to follow the principles of embryogenesis. The embryonic development of the skeleton has been studied extensively with respect to the regulatory mechanisms governing morphogenesis, differentiation, and tissue formation. Various concepts have been designed for engineering tissues in vitro based on these developmental principles, most of them involving regulatory molecules such as growth factors or cytokines known to be the key regulators in developmental processes. Growth factors most commonly used for in vitro cultivation of cartilage tissue belong to the fibroblast growth factor (FGF) family, the transforming growth factor-beta (TGF-β) super-family, and the insulin-like growth factor (IGF) family. In this chapter, in vivo actions of members of these growth factors described in the literature are compared with in vitro concepts of cartilage engineering making use of these growth factors.

  1. Repair mechanisms of bone marrow mesenchymal stem cells in myocardial infarction.

    PubMed

    Wen, Zhuzhi; Zheng, Shaoxin; Zhou, Changqing; Wang, Jingfeng; Wang, Tong

    2011-05-01

    The prognosis of patients with myocardial infarction (MI) and resultant chronic heart failure remains extremely poor despite advances in optimal medical therapy and interventional procedures. Animal experiments and clinical trials using adult stem cell therapy following MI have shown a global improvement of myocardial function. Bone marrow-derived mesenchymal stem cells (MSCs) hold promise for cardiac repair following MI, due to their multilineage, self-renewal and proliferation potential. In addition, MSCs can be easily isolated, expanded in culture, and have immunoprivileged properties to the host tissue. Experimental studies and clinical trials have revealed that MSCs not only differentiate into cardiomyocytes and vascular cells, but also secrete amounts of growth factors and cytokines which may mediate endogenous regeneration via activation of resident cardiac stem cells and other stem cells, as well as induce neovascularization, anti-inflammation, anti-apoptosis, anti-remodelling and cardiac contractility in a paracrine manner. It has also been postulated that the anti-arrhythmic and cardiac nerve sprouting potential of MSCs may contribute to their beneficial effects in cardiac repair. Most molecular and cellular mechanisms involved in the MSC-based therapy after MI are still unclear at present. This article reviews the potential repair mechanisms of MSCs in the setting of MI. PMID:21199333

  2. Spheroid Culture of Mesenchymal Stem Cells

    PubMed Central

    Cesarz, Zoe; Tamama, Kenichi

    2016-01-01

    Compared with traditional 2D adherent cell culture, 3D spheroidal cell aggregates, or spheroids, are regarded as more physiological, and this technique has been exploited in the field of oncology, stem cell biology, and tissue engineering. Mesenchymal stem cells (MSCs) cultured in spheroids have enhanced anti-inflammatory, angiogenic, and tissue reparative/regenerative effects with improved cell survival after transplantation. Cytoskeletal reorganization and drastic changes in cell morphology in MSC spheroids indicate a major difference in mechanophysical properties compared with 2D culture. Enhanced multidifferentiation potential, upregulated expression of pluripotency marker genes, and delayed replicative senescence indicate enhanced stemness in MSC spheroids. Furthermore, spheroid formation causes drastic changes in the gene expression profile of MSC in microarray analyses. In spite of these significant changes, underlying molecular mechanisms and signaling pathways triggering and sustaining these changes are largely unknown. PMID:26649054

  3. A Comparative Study of the Therapeutic Potential of Mesenchymal Stem Cells and Limbal Epithelial Stem Cells for Ocular Surface Reconstruction

    PubMed Central

    Trosan, Peter; Cejka, Cestmir; Javorkova, Eliska; Zajicova, Alena; Hermankova, Barbora; Chudickova, Milada; Cejkova, Jitka

    2015-01-01

    Stem cell-based therapy has become an attractive and promising approach for the treatment of severe injuries or thus-far incurable diseases. However, the use of stem cells is often limited by a shortage of available tissue-specific stem cells; therefore, other sources of stem cells are being investigated and tested. In this respect, mesenchymal stromal/stem cells (MSCs) have proven to be a promising stem cell type. In the present study, we prepared MSCs from bone marrow (BM-MSCs) or adipose tissue (Ad-MSCs) as well as limbal epithelial stem cells (LSCs), and their growth, differentiation, and secretory properties were compared. The cells were grown on nanofiber scaffolds and transferred onto the alkali-injured eye in a rabbit model, and their therapeutic potential was characterized. We found that BM-MSCs and tissue-specific LSCs had similar therapeutic effects. Clinical characterization of the healing process, as well as the evaluation of corneal thickness, re-epithelialization, neovascularization, and the suppression of a local inflammatory reaction, were comparable in the BM-MSC- and LSC-treated eyes, but results were significantly better than in injured, untreated eyes or in eyes treated with a nanofiber scaffold alone or with a nanofiber scaffold seeded with Ad-MSCs. Taken together, the results show that BM-MSCs’ therapeutic effect on healing of injured corneal surface is comparable to that of tissue-specific LSCs. We suggest that BM-MSCs can be used for ocular surface regeneration in cases when autologous LSCs are absent or difficult to obtain. Significance Damage of ocular surface represents one of the most common causes of impaired vision or even blindness. Cell therapy, based on transplantation of stem cells, is an optimal treatment. However, if limbal stem cells (LSCs) are not available, other sources of stem cells are tested. Mesenchymal stem cells (MSCs) are a convenient type of cell for stem cell therapy. The therapeutic potential of LSCs and MSCs was

  4. Toll-Like Receptors on Human Mesenchymal Stem Cells Drive their Migration and Immunomodulating Responses

    PubMed Central

    Tomchuck, Suzanne L.; Zwezdaryk, Kevin J.; Coffelt, Seth B.; Waterman, Ruth S.; Danka, Elizabeth S.; Scandurro, Aline B.

    2009-01-01

    Adult human bone marrow-derived mesenchymal stem cells (hMSCs) are under study as therapeutic delivery agents that assist in the repair of damaged tissues. To achieve the desired clinical outcomes for this strategy requires a better understanding of the mechanisms that drive the recruitment, migration and engraftment of hMSCs to the targeted tissues. It is known that hMSCs are recruited to sites of stress or inflammation to fulfill their repair function. It is recognized that toll-like receptors (TLRs) mediate stress responses of other bone marrow-derived cells. This study explored the role of TLRs in mediating stress responses of hMSCs. Accordingly, the presence of TLRs in hMSCs was established initially by RT-PCR assays. Flow cytometry and fluorescence immunocytochemical analyses confirmed these findings. The stimulation of hMSCs with TLR agonists led to the activation of downstream signaling pathways, including NF-κB, AKT and MAPK. Consequently, activation of these pathways triggered the induction and secretion of cytokines, chemokines and related TLR gene products as established from cDNA array, immunoassay and cytokine antibody array analyses. Interestingly, the unique patterns of affected genes, cytokines and chemokines measured, identify these receptors as critical players in the clinically established immunomodulation, observed for hMSCs. Lastly, hMSCs migration was promoted by TLR ligand exposure as demonstrated by transwell migration assays. Conversely, disruption of TLRs by neutralizing TLR antibodies compromised hMSCs migration. This study defines a novel TLR-driven stress and immune modulating response for hMSCs that is critical to consider in the design of stem cell-based therapies. PMID:17916800

  5. Toll-like receptors on human mesenchymal stem cells drive their migration and immunomodulating responses.

    PubMed

    Tomchuck, Suzanne L; Zwezdaryk, Kevin J; Coffelt, Seth B; Waterman, Ruth S; Danka, Elizabeth S; Scandurro, Aline B

    2008-01-01

    Adult human bone marrow-derived mesenchymal stem cells (hMSCs) are under study as therapeutic delivery agents that assist in the repair of damaged tissues. To achieve the desired clinical outcomes for this strategy requires a better understanding of the mechanisms that drive the recruitment, migration, and engraftment of hMSCs to the targeted tissues. It is known that hMSCs are recruited to sites of stress or inflammation to fulfill their repair function. It is recognized that toll-like receptors (TLRs) mediate stress responses of other bone marrow-derived cells. This study explored the role of TLRs in mediating stress responses of hMSCs. Accordingly, the presence of TLRs in hMSCs was initially established by reverse transcription-polymerase chain reaction assays. Flow cytometry and fluorescence immunocytochemical analyses confirmed these findings. The stimulation of hMSCs with TLR agonists led to the activation of downstream signaling pathways, including nuclear factor kappaB, AKT, and MAPK. Consequently, activation of these pathways triggered the induction and secretion of cytokines, chemokines, and related TLR gene products as established from cDNA array, immunoassay, and cytokine antibody array analyses. Interestingly, the unique patterns of affected genes, cytokines, and chemokines measured identify these receptors as critical players in the clinically established immunomodulation observed for hMSCs. Lastly, hMSC migration was promoted by TLR ligand exposure as demonstrated by transwell migration assays. Conversely, disruption of TLRs by neutralizing TLR antibodies compromised hMSC migration. This study defines a novel TLR-driven stress and immune modulating response for hMSCs that is critical to consider in the design of stem cell-based therapies. PMID:17916800

  6. Mesenchymal stem cells in osteoarticular diseases.

    PubMed

    Jorgensen, Christian; Noël, Danièle

    2011-11-01

    Multipotent mesenchymal stromal cells or mesenchymal stem cells (MSCs) are mainly isolated from bone marrow or fat tissue. Owing to their potential for multilineage differentiation towards bone, cartilage and fat tissue, they were initially evaluated in innovative strategies for tissue engineering. More recently, they have gained interest for their immunomodulatory properties and have been tested in various clinical trials that aim to modulate the host immune response in graft-versus-host disease or autoimmune diseases. MSC-mediated immunomodulation occurs through the secretion of soluble mediators. The clinical applications of MSCs for rheumatic diseases focus on their potential to promote tissue repair/regeneration and prevent inflammation. This article will focus on the mechanisms by which MSCs might exhibit a therapeutic potential in rheumatology. Special attention is given to their potential for innovative future strategies. PMID:21999261

  7. In Vitro Differentiation of Insulin Secreting Cells from Mouse Bone Marrow Derived Stage-Specific Embryonic Antigen 1 Positive Stem Cells

    PubMed Central

    Abouzaripour, Morteza; Pasbakhsh, Parichehr; Atlasi, Nader; Shahverdi, Abdol Hossein; Mahmoudi, Reza; Kashani, Iraj Ragerdi

    2016-01-01

    Objective Bone marrow has recently been recognized as a novel source of stem cells for the treatment of wide range of diseases. A number of studies on murine bone mar- row have shown a homogenous population of rare stage-specific embryonic antigen 1 (SSEA-1) positive cells that express markers of pluripotent stem cells. This study focuses on SSEA-1 positive cells isolated from murine bone marrow in an attempt to differentiate them into insulin-secreting cells (ISCs) in order to investigate their differentiation potential for future use in cell therapy. Materials and Methods This study is an experimental research. Mouse SSEA-1 positive cells were isolated by Magnetic-activated cell sorting (MACS) followed by characteriza- tion with flow cytometry. Induced SSEA-1 positive cells were differentiated into ISCs with specific differentiation media. In order to evaluate differentiation quality and analysis, dithizone (DTZ) staining was use, followed by reverse transcription polymerase chain reaction (RT-PCR), immunocytochemistry and insulin secretion assay. Statistical results were analyzed by one-way ANOVA. Results The results achieved in this study reveal that mouse bone marrow contains a population of SSEA-1 positive cells that expresses pluripotent stem cells markers such as SSEA-1, octamer-binding transcription factor 4 (OCT-4) detected by immunocytochem- istry and C-X-C chemokine receptor type 4 (CXCR4) and stem cell antigen-1 (SCA-1) detected by flow cytometric analysis. SSEA-1 positive cells can differentiate into ISCs cell clusters as evidenced by their DTZ positive staining and expression of genes such as Pdx1 (pancreatic transcription factors), Ngn3 (endocrine progenitor marker), Insulin1 and Insulin2 (pancreaticβ-cell markers). Additionally, our results demonstrate expression of Pdx1 and Glut2 protein and insulin secretion in response to a glucose challenge in the differentiated cells. Conclusion Our study clearly demonstrates the potential of SSEA-1 positive

  8. HORSE SPECIES SYMPOSIUM: Use of mesenchymal stem cells in fracture repair in horses.

    PubMed

    Govoni, K E

    2015-03-01

    Equine bone fractures are often catastrophic, potentially fatal, and costly to repair. Traditional methods of healing fractures have limited success, long recovery periods, and a high rate of reinjury. Current research in the equine industry has demonstrated that stem cell therapy is a promising novel therapy to improve fracture healing and reduce the incidence of reinjury; however, reports of success in horses have been variable and limited. Stem cells can be derived from embryonic, fetal, and adult tissue. Based on the ease of collection, opportunity for autologous cells, and proven success in other models, adipose- or bone marrow-derived mesenchymal stem cells (MSC) are often used in equine therapies. Methods for isolation, proliferation, and differentiation of MSC are well established in rodent and human models but are not well characterized in horses. There is recent evidence that equine bone marrow MSC are able to proliferate in culture for several passages in the presence of autologous and fetal bovine serum, which is important for expansion of cells. Mesenchymal stem cells have the capacity to differentiate into osteoblasts, the bone forming cells, and this complex process is regulated by a number of transcription factors including runt-related transcription factor 2 (Runx2) and osterix (Osx). However, it has not been well established if equine MSC are regulated in a similar manner. The data presented in this review support the view that equine bone marrow MSC are regulated by the same transcription factors that control the differentiation of rodent and human MSC into osteoblasts. Although stem cell therapy is promising in equine bone repair, additional research is needed to identify optimal methods for reintroduction and potential manipulations to improve their ability to form new bone. PMID:26020865

  9. Allogeneic mesenchymal stem cell infusion for treatment of metachromatic leukodystrophy (MLD) and Hurler syndrome (MPS-IH).

    PubMed

    Koç, O N; Day, J; Nieder, M; Gerson, S L; Lazarus, H M; Krivit, W

    2002-08-01

    Patients with Hurler syndrome (mucopolysaccharidosis type-IH) and metachromatic leukodystrophy (MLD) develop significant skeletal and neurologic defects that limit their survival. Transplantation of allogeneic hematopoietic stem cells results in partial correction of the clinical manifestations. We postulated that some of these defects may be corrected by infusion of allogeneic, multipotential, bone marrow-derived mesenchymal stem cells (MSC). Patients with Hurler syndrome (n = 5) or MLD (n = 6) who previously underwent successful bone marrow transplantation from an HLA-identical sibling were infused with 2-10 x 10(6)/kg MSCs, isolated and expanded from a bone marrow aspirate of the original donor. There was no infusion-related toxicity. In most recipients culture-purified MSCs at 2 days, 30-60 days and 6-24 months after MSC infusion remained of host type. In two patients the bone marrow-derived MSCs contained 0.4 and 2% donor MSCs by FISH 60 days after MSC infusion. In four patients with MLD there were significant improvements in nerve conduction velocities after MSC infusion. The bone mineral density was either maintained or slightly improved in all patients. There was no clinically apparent change in patients' overall health, mental and physical development after MSC infusion. We conclude that donor allogeneic MSC infusion is safe and may be associated with reversal of disease pathophysiology in some tissues. The role of MSCs in the management of Hurler syndrome and MLD should be further evaluated. PMID:12203137

  10. Adult Mesenchymal Stem Cells and Radiation Injury.

    PubMed

    Kiang, Juliann G

    2016-08-01

    Recent understanding of the cellular and molecular signaling activations in adult mesenchymal stem cells (MSCs) has provided new insights into their potential clinical applications, particularly for tissue repair and regeneration. This review focuses on these advances, specifically in the context of self-renewal for tissue repair and recovery after radiation injury. Thus far, MSCs have been characterized extensively and shown to be useful in mitigation and therapy for acute radiation syndrome and cognitive dysfunction. Use of MSCs for treating radiation injury alone or in combination with additional trauma is foreseeable. PMID:27356065

  11. Mesenchymal stem cells and cardiac repair

    PubMed Central

    Nesselmann, Catharina; Ma, Nan; Bieback, Karen; Wagner, Wolfgang; Ho, Anthony; Konttinen, Yrjö T; Zhang, Hao; Hinescu, Mihail E; Steinhoff, Gustav

    2008-01-01

    Accumulating clinical and experimental evidence indicates that mesenchymal stem cells (MSCs) are promising cell types in the treatment of cardiac dysfunction. They may trigger production of reparative growth factors, replace damaged cells and create an environment that favours endogenous cardiac repair. However, identifying mechanisms which regulate the role of MSCs in cardiac repair is still at work. To achieve the maximal clinical benefits, ex vivo manipulation can further enhance MSC therapeutic potential. This review focuses on the mechanism of MSCs in cardiac repair, with emphasis on ex vivo manipulation. PMID:18684237

  12. Identification of New Rat Bone Marrow-Derived Population of Very Small Stem Cell with Oct-4A and Nanog Expression by Flow Cytometric Platforms

    PubMed Central

    Labedz-Maslowska, Anna; Kamycka, Elzbieta; Bobis-Wozowicz, Sylwia; Madeja, Zbigniew; Zuba-Surma, Ewa K.

    2016-01-01

    Very small embryonic-like stem cells (VSELs) represent a unique rare population of adult stem cells (SCs) sharing several structural, genetic, biochemical, and functional properties with embryonic SCs and have been identified in several adult murine and human tissues. However, rat bone marrow- (BM-) derived SCs closely resembling murine or human VSELs have not been described. Thus, we employed multi-instrumental flow cytometric approach including classical and imaging cytometry and we established that newly identified population of nonhematopoietic cells expressing CD106 (VCAM-I) antigen contains SCs with very small size, expressing markers of pluripotency (Oct-4A and Nanog) on both mRNA and protein levels that indicate VSEL population. Based on our experience in both murine and human VSEL isolation procedures by fluorescence-activated cell sorting (FACS), we also optimized sorting protocol for separation of CD45−/Lin−/CD106+ rat BM-derived VSELs from wild type and eGFP-expressing rats, which are often used as donor animals for cell transplantations in regenerative studies in vivo. Thus, this is a first study identifying multiantigenic phenotype and providing sorting protocols for isolation VSELs from rat BM tissue for further examining of their functional properties in vitro as well as regenerative capacity in distinct in vivo rat models of tissue injury. PMID:26633976

  13. Transplantation of Bone Marrow-Derived Very Small Embryonic-Like Stem Cells Attenuates Left Ventricular Dysfunction and Remodeling After Myocardial Infarction

    PubMed Central

    Dawn, Buddhadeb; Tiwari, Sumit; Kucia, Magdalena J.; Zuba-Surma, Ewa K.; Guo, Yiru; SanganalMath, Santosh K.; Abdel-Latif, Ahmed; Hunt, Greg; Vincent, Robert J.; Taher, Hisham; Reed, Nathan J.; Ratajczak, Mariusz Z.; Bolli, Roberto

    2013-01-01

    Adult bone marrow (BM) contains Sca-1+/Lin−/CD45− very small embryonic-like stem cells (VSELs) that express markers of several lineages, including cardiac markers, and differentiate into cardiomyocytes in vitro. We examined whether BM-derived VSELs promote myocardial repair after a reperfused myocardial infarction (MI). Mice underwent a 30-minute coronary occlusion followed by reperfusion and received intramyocardial injection of vehicle (n = 11), 1 × 105 Sca-1+/Lin−/CD45+ enhanced green fluorescent protein (EGFP)-labeled hematopoietic stem cells (n = 13 [cell control group]), or 1 × 104 Sca-1+/Lin−/CD45− EGFP-labeled cells (n = 14 [VSEL-treated group]) at 48 hours after MI. At 35 days after MI, VSEL-treated mice exhibited improved global and regional left ventricular (LV) systolic function (echocardiography) and attenuated myocyte hypertrophy in surviving tissue (histology and echocardiography) compared with vehicle-treated controls. In contrast, transplantation of Sca-1+/Lin−/CD45+ cells failed to confer any functional or structural benefits. Scattered EGFP+ myocytes and capillaries were present in the infarct region in VSEL-treated mice, but their numbers were very small. These results indicate that transplantation of a relatively small number of CD45− VSELs is sufficient to improve LV function and alleviate myocyte hypertrophy after MI, supporting the potential therapeutic utility of these cells for cardiac repair. PMID:18420834

  14. Mesenchymal Stem/Stromal Cells Derived from Induced Pluripotent Stem Cells Support CD34pos Hematopoietic Stem Cell Propagation and Suppress Inflammatory Reaction

    PubMed Central

    Moslem, Mohsen; Eberle, Irina; Weber, Iuliia; Henschler, Reinhard; Cantz, Tobias

    2015-01-01

    Mesenchymal stem/stromal cells (MSCs) represent a promising cell source for research and therapeutic applications, but their restricted ex vivo propagation capabilities limit putative applications. Substantial self-renewing of stem cells can be achieved by reprogramming cells into induced pluripotent stem cells (iPSCs) that can be easily expanded as undifferentiated cells even in mass culture. Here, we investigated a differentiation protocol enabling the generation and selection of human iPSC-derived MSCs exhibiting relevant surface marker expression profiles (CD105 and CD73) and functional characteristics. We generated such iPSC-MSCs from fibroblasts and bone marrow MSCs utilizing two different reprogramming constructs. All such iPSC-MSCs exhibited the characteristics of normal bone marrow-derived (BM) MSCs. In direct comparison to BM-MSCs our iPSC-MSCs exhibited a similar surface marker expression profile but shorter doubling times without reaching senescence within 20 passages. Considering functional capabilities, iPSC-MSCs provided supportive feeder layer for CD34+ hematopoietic stem cells' self-renewal and colony forming capacities. Furthermore, iPSC-MSCs gained immunomodulatory function to suppress CD4+ cell proliferation, reduce proinflammatory cytokines in mixed lymphocyte reaction, and increase regulatory CD4+/CD69+/CD25+ T-lymphocyte population. In conclusion, we generated fully functional MSCs from various iPSC lines irrespective of their starting cell source or reprogramming factor composition and we suggest that such iPSC-MSCs allow repetitive cell applications for advanced therapeutic approaches. PMID:26185499

  15. Mesenchymal Stem/Stromal Cells Derived from Induced Pluripotent Stem Cells Support CD34(pos) Hematopoietic Stem Cell Propagation and Suppress Inflammatory Reaction.

    PubMed

    Moslem, Mohsen; Eberle, Irina; Weber, Iuliia; Henschler, Reinhard; Cantz, Tobias

    2015-01-01

    Mesenchymal stem/stromal cells (MSCs) represent a promising cell source for research and therapeutic applications, but their restricted ex vivo propagation capabilities limit putative applications. Substantial self-renewing of stem cells can be achieved by reprogramming cells into induced pluripotent stem cells (iPSCs) that can be easily expanded as undifferentiated cells even in mass culture. Here, we investigated a differentiation protocol enabling the generation and selection of human iPSC-derived MSCs exhibiting relevant surface marker expression profiles (CD105 and CD73) and functional characteristics. We generated such iPSC-MSCs from fibroblasts and bone marrow MSCs utilizing two different reprogramming constructs. All such iPSC-MSCs exhibited the characteristics of normal bone marrow-derived (BM) MSCs. In direct comparison to BM-MSCs our iPSC-MSCs exhibited a similar surface marker expression profile but shorter doubling times without reaching senescence within 20 passages. Considering functional capabilities, iPSC-MSCs provided supportive feeder layer for CD34(+) hematopoietic stem cells' self-renewal and colony forming capacities. Furthermore, iPSC-MSCs gained immunomodulatory function to suppress CD4(+) cell proliferation, reduce proinflammatory cytokines in mixed lymphocyte reaction, and increase regulatory CD4(+)/CD69(+)/CD25(+) T-lymphocyte population. In conclusion, we generated fully functional MSCs from various iPSC lines irrespective of their starting cell source or reprogramming factor composition and we suggest that such iPSC-MSCs allow repetitive cell applications for advanced therapeutic approaches. PMID:26185499

  16. Comprehensive transcriptomic and proteomic characterization of human mesenchymal stem cells reveals source specific cellular markers.

    PubMed

    Billing, Anja M; Ben Hamidane, Hisham; Dib, Shaima S; Cotton, Richard J; Bhagwat, Aditya M; Kumar, Pankaj; Hayat, Shahina; Yousri, Noha A; Goswami, Neha; Suhre, Karsten; Rafii, Arash; Graumann, Johannes

    2016-01-01

    Mesenchymal stem cells (MSC) are multipotent cells with great potential in therapy, reflected by more than 500 MSC-based clinical trials registered with the NIH. MSC are derived from multiple tissues but require invasive harvesting and imply donor-to-donor variability. Embryonic stem cell-derived MSC (ESC-MSC) may provide an alternative, but how similar they are to ex vivo MSC is unknown. Here we performed an in depth characterization of human ESC-MSC, comparing them to human bone marrow-derived MSC (BM-MSC) as well as human embryonic stem cells (hESC) by transcriptomics (RNA-seq) and quantitative proteomics (nanoLC-MS/MS using SILAC). Data integration highlighted and validated a central role of vesicle-mediated transport and exosomes in MSC biology and also demonstrated, through enrichment analysis, their versatility and broad application potential. Particular emphasis was placed on comparing profiles between ESC-MSC and BM-MSC and assessing their equivalency. Data presented here shows that differences between ESC-MSC and BM-MSC are similar in magnitude to those reported for MSC of different origin and the former may thus represent an alternative source for therapeutic applications. Finally, we report an unprecedented coverage of MSC CD markers, as well as membrane associated proteins which may benefit immunofluorescence-based applications and contribute to a refined molecular description of MSC. PMID:26857143

  17. Comprehensive transcriptomic and proteomic characterization of human mesenchymal stem cells reveals source specific cellular markers

    PubMed Central

    Billing, Anja M.; Ben Hamidane, Hisham; Dib, Shaima S.; Cotton, Richard J.; Bhagwat, Aditya M.; Kumar, Pankaj; Hayat, Shahina; Yousri, Noha A.; Goswami, Neha; Suhre, Karsten; Rafii, Arash; Graumann, Johannes

    2016-01-01

    Mesenchymal stem cells (MSC) are multipotent cells with great potential in therapy, reflected by more than 500 MSC-based clinical trials registered with the NIH. MSC are derived from multiple tissues but require invasive harvesting and imply donor-to-donor variability. Embryonic stem cell-derived MSC (ESC-MSC) may provide an alternative, but how similar they are to ex vivo MSC is unknown. Here we performed an in depth characterization of human ESC-MSC, comparing them to human bone marrow-derived MSC (BM-MSC) as well as human embryonic stem cells (hESC) by transcriptomics (RNA-seq) and quantitative proteomics (nanoLC-MS/MS using SILAC). Data integration highlighted and validated a central role of vesicle-mediated transport and exosomes in MSC biology and also demonstrated, through enrichment analysis, their versatility and broad application potential. Particular emphasis was placed on comparing profiles between ESC-MSC and BM-MSC and assessing their equivalency. Data presented here shows that differences between ESC-MSC and BM-MSC are similar in magnitude to those reported for MSC of different origin and the former may thus represent an alternative source for therapeutic applications. Finally, we report an unprecedented coverage of MSC CD markers, as well as membrane associated proteins which may benefit immunofluorescence-based applications and contribute to a refined molecular description of MSC. PMID:26857143

  18. Proteomic Definitions of Mesenchymal Stem Cells

    PubMed Central

    Maurer, Martin H.

    2011-01-01

    Mesenchymal stem cells (MSCs) are pluripotent cells isolated from the bone marrow and various other organs. They are able to proliferate and self-renew, as well as to give rise to progeny of at least the osteogenic, chondrogenic, and adipogenic lineages. Despite this functional definition, MSCs can also be defined by their expression of a distinct set of cell surface markers. In the current paper, studies investigating the proteome of human MSCs are reviewed with the aim to identify common protein markers of MSCs. The proteomic analysis of MSCs revealed a distinct set of proteins representing the basic molecular inventory, including proteins for (i) cell surface markers, (ii) the responsiveness to growth factors, (iii) the reuse of developmental signaling cascades in adult stem cells, (iv) the interaction with molecules of the extracellular matrix, (v) the expression of genes regulating transcription and translation, (vi) the control of the cell number, and (vii) the protection against cellular stress. PMID:21437194

  19. Bioprinting Organotypic Hydrogels with Improved Mesenchymal Stem Cell Remodeling and Mineralization Properties for Bone Tissue Engineering.

    PubMed

    Duarte Campos, Daniela Filipa; Blaeser, Andreas; Buellesbach, Kate; Sen, Kshama Shree; Xun, Weiwei; Tillmann, Walter; Fischer, Horst

    2016-06-01

    3D-manufactured hydrogels with precise contours and biological adhesion motifs are interesting candidates in the regenerative medicine field for the culture and differentiation of human bone-marrow-derived mesenchymal stem cells (MSCs). 3D-bioprinting is a powerful technique to approach one step closer the native organization of cells. This study investigates the effect of the incorporation of collagen type I in 3D-bioprinted polysaccharide-based hydrogels to the modulation of cell morphology, osteogenic remodeling potential, and mineralization. By combining thermo-responsive agarose hydrogels with collagen type I, the mechanical stiffness and printing contours of printed constructs can be improved compared to pure collagen hydrogels which are typically used as standard materials for MSC osteogenic differentiation. The results presented here show that MSC not only survive the 3D-bioprinting process but also maintain the mesenchymal phenotype, as proved by live/dead staining and immunocytochemistry (vimentin positive, CD34 negative). Increased solids concentrations of collagen in the hydrogel blend induce changes in cell morphology, namely, by enhancing cell spreading, that ultimately contribute to enhanced and directed MSC osteogenic differentiation. 3D-bioprinted agarose-collagen hydrogels with high-collagen ratio are therefore feasible for MSC osteogenic differentiation, contrarily to low-collagen blends, as proved by two-photon microscopy, Alizarin Red staining, and real-time polymerase chain reaction. PMID:27072652

  20. Micropatterned 3-Dimensional Hydrogel System to Study Human Endothelial-Mesenchymal Stem Cell Interactions

    PubMed Central

    Trkov, Sasa; Eng, George; di Liddo, Rosa; Parnigotto, Pier Paolo; Vunjak-Novakovic, Gordana

    2009-01-01

    The creation of vascularized engineered tissues of clinically relevant size is a major challenge of tissue engineering. While it is known that endothelial and mural vascular cells are integral to the formation of stable blood vessels, the specific cell type and optimal conditions for engineered vascular networks are poorly understood. To this end, we investigated the vasculogenic potential of human mesenchymal stem cell (MSC) populations derived from three different sources: (i) bone marrow aspirates, (ii) perivascular cells from umbilical cord vein, and (iii) perivascular cells from umbilical cord artery. Cell populations were isolated and identified as MSCs according to their phenotypes and differentiation potential. Human umbilical vein endothelial cells (HUVEC) were used as a standard for endothelial cells. A novel co-culture system was developed to study cell-cell interactions in a spatially controlled three-dimensional (3D) fibrin hydrogel model. Using microfluidic patterning, it was possible to localize hydrogel-encapsulated HUVECs and MSCs within separate channels spaced at 500, 1000 or 2000 μm. All three MSC populations had similar expression profiles of mesenchymal cell markers, and similar capacity for osteogenic and adipogenic differentiation. However, bone marrow-derived MSCs (but not umbilical vein or artery derived MSCs) showed strong distance-dependent migration toward HUVECs and supported the formation of stable vascular networks resembling capillary-like vasculature. The presented approach provides a simple and robust model to study cell-cell communication of relevance to engineering vascularized tissues. PMID:19998330

  1. Viability of mesenchymal stem cells during electrospinning

    PubMed Central

    Zanatta, G.; Steffens, D.; Braghirolli, D.I.; Fernandes, R.A.; Netto, C.A.; Pranke, P.

    2011-01-01

    Tissue engineering is a technique by which a live tissue can be re-constructed and one of its main goals is to associate cells with biomaterials. Electrospinning is a technique that facilitates the production of nanofibers and is commonly used to develop fibrous scaffolds to be used in tissue engineering. In the present study, a different approach for cell incorporation into fibrous scaffolds was tested. Mesenchymal stem cells were extracted from the wall of the umbilical cord and mononuclear cells from umbilical cord blood. Cells were re-suspended in a 10% polyvinyl alcohol solution and subjected to electrospinning for 30 min under a voltage of 21 kV. Cell viability was assessed before and after the procedure by exclusion of dead cells using trypan blue staining. Fiber diameter was observed by scanning electron microscopy and the presence of cells within the scaffolds was analyzed by confocal laser scanning microscopy. After electrospinning, the viability of mesenchymal stem cells was reduced from 88 to 19.6% and the viability of mononuclear cells from 99 to 8.38%. The loss of viability was possibly due to the high viscosity of the polymer solution, which reduced the access to nutrients associated with electric and mechanical stress during electrospinning. These results suggest that the incorporation of cells during fiber formation by electrospinning is a viable process that needs more investigation in order to find ways to protect cells from damage. PMID:22183245

  2. Addition of Adipose-Derived Stem Cells to Mesenchymal Stem Cell Sheets Improves Bone Formation at an Ectopic Site

    PubMed Central

    Wang, Zhifa; Li, Zhijin; Dai, Taiqiang; Zong, Chunlin; Liu, Yanpu; Liu, Bin

    2016-01-01

    To determine the effect of adipose-derived stem cells (ADSCs) added to bone marrow-derived mesenchymal stem cell (MSC) sheets on bone formation at an ectopic site. We isolated MSCs and ADSCs from the same rabbits. We then prepared MSC sheets for implantation with or without ADSCs subcutaneously in the backs of severe combined immunodeficiency (SCID) mice. We assessed bone formation at eight weeks after implantation by micro-computed tomography and histological analysis. In osteogenic medium, MSCs grew to form multilayer sheets containing many calcium nodules. MSC sheets without ADSCs formed bone-like tissue; although neo-bone and cartilage-like tissues were sparse and unevenly distributed by eight weeks after implantation. In comparison, MSC sheets with ADSCs promoted better bone regeneration as evidenced by the greater density of bone, increased mineral deposition, obvious formation of blood vessels, large number of interconnected ossified trabeculae and woven bone structures, and greater bone volume/total volume within the composite constructs. Our results indicate that although sheets of only MSCs have the potential to form tissue engineered bone at an ectopic site, the addition of ADSCs can significantly increase the osteogenic potential of MSC sheets. Thus, the combination of MSC sheets with ADSCs may be regarded as a promising therapeutic strategy to stimulate bone regeneration. PMID:26848656

  3. Hepatoma SK Hep-1 Cells Exhibit Characteristics of Oncogenic Mesenchymal Stem Cells with Highly Metastatic Capacity

    PubMed Central

    Zhang, Yanling; Zhang, Yanhong; Tschudy-Seney, Benjamin; Ramsamooj, Rajen; Wan, Yu-Jui Yvonne; Theise, Neil D.; Zern, Mark A.; Duan, Yuyou

    2014-01-01

    Background SK Hep-1 cells (SK cells) derived from a patient with liver adenocarcinoma have been considered a human hepatoma cell line with mesenchymal origin characteristics, however, SK cells do not express liver genes and exhibit liver function, thus, we hypothesized whether mesenchymal cells might contribute to human liver primary cancers. Here, we characterized SK cells and its tumourigenicity. Methods and Principal Findings We found that classical mesenchymal stem cell (MSC) markers were presented on SK cells, but endothelial marker CD31, hematopoietic markers CD34 and CD45 were negative. SK cells are capable of differentiate into adipocytes and osteoblasts as adipose-derived MSC (Ad-MSC) and bone marrow-derived MSC (BM-MSC) do. Importantly, a single SK cell exhibited a substantial tumourigenicity and metastatic capacity in immunodefficient mice. Metastasis not only occurred in circulating organs such as lung, liver, and kidneys, but also in muscle, outer abdomen, and skin. SK cells presented greater in vitro invasive capacity than those of Ad-MSC and BM-MSC. The xenograft cells from subcutaneous and metastatic tumors exhibited a similar tumourigenicity and metastatic capacity, and showed the same relatively homogenous population with MSC characteristics when compared to parental SK cells. SK cells could unlimitedly expand in vitro without losing MSC characteristics, its tumuorigenicity and metastatic capacity, indicating that SK cells are oncogenic MSC with enhanced self-renewal capacity. We believe that this is the first report that human MSC appear to be transformed into cancer stem cells (CSC), and that their derivatives also function as CSCs. Conclusion Our findings demonstrate that SK cells represent a transformation mechanism of normal MSC into an enhanced self-renewal CSC with metastasis capacity, SK cells and their xenografts represent a same relative homogeneity of CSC with substantial metastatic capacity. Thus, it represents a novel mechanism of

  4. Recruitment of mesenchymal stem cells into prostate tumours promotes metastasis.

    PubMed

    Jung, Younghun; Kim, Jin Koo; Shiozawa, Yusuke; Wang, Jingcheng; Mishra, Anjali; Joseph, Jeena; Berry, Janice E; McGee, Samantha; Lee, Eunsohl; Sun, Hongli; Wang, Jianhua; Jin, Taocong; Zhang, Honglai; Dai, Jinlu; Krebsbach, Paul H; Keller, Evan T; Pienta, Kenneth J; Taichman, Russell S

    2013-01-01

    Tumours recruit mesenchymal stem cells to facilitate healing, which induces their conversion into cancer-associated fibroblasts that facilitate metastasis. However, this process is poorly understood on the molecular level. Here we show that CXCL16, a ligand for CXCR6, facilitates mesenchymal stem cell or very small embryonic-like cells recruitment into prostate tumours. CXCR6 signalling stimulates the conversion of mesenchymal stem cells into cancer-associated fibroblasts, which secrete stromal-derived factor-1, also known as CXCL12. CXCL12 expressed by cancer-associated fibroblasts then binds to CXCR4 on tumour cells and induces an epithelial-to-mesenchymal transition, which ultimately promotes metastasis to secondary tumour sites. Our results provide the molecular basis for mesenchymal stem cell recruitment into tumours and how this process leads to tumour metastasis. PMID:23653207

  5. MiR-124 inhibits myogenic differentiation of mesenchymal stem cells via targeting Dlx5.

    PubMed

    Qadir, Abdul S; Woo, Kyung Mi; Ryoo, Hyun-Mo; Yi, TacGhee; Song, Sun U; Baek, Jeong-Hwa

    2014-09-01

    MicroRNAs (miRNAs), including miR-1, miR-133, and miR-206, play a crucial role in muscle development by regulating muscle cell proliferation and differentiation. The aim of the present study was to define the effect of miR-124 on myogenic differentiation of mesenchymal stem cells (MSCs). The expression level of miR-124 in skeletal muscles was much lower than those in primary cultured bone marrow-derived MSCs and the bone, fat and brain tissues obtained from C57BL/6 mice. Myogenic stimuli significantly decreased the expression levels of miR-124 in mouse bone marrow-derived MSCs and C2C12 cells. Forced expression of miR-124 suppressed the expression of myogenic marker genes such as Myf5, Myod1, myogenin and myosin heavy chain and multinucleated myotube formation. Blockade of endogenous miR-124 with a hairpin inhibitor enhanced myogenic marker gene expression and myotube formation. During myogenic differentiation of MSCs and C2C12 cells, the levels of Dlx5, a known target of miR-124, were inversely regulated with those of miR-124. Furthermore, overexpression of Dlx5 increased myogenic differentiation, whereas knockdown of Dlx5 using siRNA inhibited myogenesis in C2C12 cells. These results suggest that miR-124 is a negative regulator of myogenic differentiation of MSCs and that upregulation of Dlx5 accompanied with downregulation of miR-124 by myogenic stimuli is necessary for the proper progression of myogenic differentiation. PMID:24733577

  6. FOXP3 gene expression in multiple sclerosis patients pre- and post mesenchymal stem cell therapy.

    PubMed

    Mohajeri, Maryam; Farazmand, Ali; Mohyeddin Bonab, Mandana; Nikbin, Behrooz; Minagar, Alireza

    2011-09-01

    Multiple Sclerosis (MS) is an inflammatory demyelinating and neurodegenerative disorder of the central nervous system (CNS), which mainly affects young adults. Activated T lymphocytes promote the neuro-inflammatory cascade of MS by secreting pro-inflammatory cytokines and play a significant role in its pathogenesis. T lymphocytes may trigger the inflammation, which in turn leads to axonal loss and neurodegeneration observed in the course of MS. Currently, there is no cure for MS, however, one of the most promising neuroprotective research tools consists of the use of bone marrow derived mesenchymal stem cells (MSC). This method promotes immune system regulation and possibly induces neurological repair and re-myelination of the damaged axons. Recent studies have shown that MSC exert an immune regulatory function and induce T regulatory-cell proliferation, therefore, it may serve as a potentially useful treatment for immune-mediated diseases such as MS. In this pilot study a group of MS patients underwent MSC therapy and we assayed the expression of an X-linked transcription factor, FoxP3, as a specific marker of T Regulatory cells in peripheral blood, prior to and after the treatment. Using q RT-PCR for measurement of expression of FoxP3 by peripheral blood mononuclear cells, we found that in all subjects, except for one, the expression of FoxP3 at 6 months after intrathecal injection of MSC was significantly higher than the levels prior to treatment. Such significant enhanced expression of FoxP3 associated with clinical stability. Findings from this pilot study further support the potential of bone marrow derived MSC for treatment of MS patients. PMID:21891821

  7. Culture and properties of adipose-derived mesenchymal stem cells: characteristics in vitro and immunosuppression in vivo

    PubMed Central

    Cao, Fujiang; Liu, Tao; Xu, Yunqiang; Xu, Dongdong; Feng, Shiqing

    2015-01-01

    Objective: To compare the two sources of adipose and bone marrow derived mesenchymal stem cells (BMSCs and AMSCs) in immune regulation and to evaluate the therapeutic effects of AMSCs on Con A induced hepatitis and the possible mechanism involved in it. Methods: We isolated bone marrow and adipose derived mesenchymal stem cells respectively and compared their differences on T lymphocyte activation, proliferation and suppression. We also test the anti-apoptosis ability of AMSCs on LO2 cell line. The effects of intravenous infusion of AMSCs on liver damage were also tested and we detected donor AMSCs in liver of recipient and their effects on the activity of intrahepatic NKT cells. Results: BMSCs and AMSCs were similar in cell phenotype and the difference existed only in the expression of CD106. The results showed that the capacity of suppressing T cells proliferation and activation was weakened in AMSCs. AMSCs ameliorated liver damage and this effect was time and dose dependent. We detected donor AMSCs in liver of recipient which suggested tissue damage could be a clue for AMSCs migration. We also found AMSCs suppress the activity of intrahepatic NKT cells, but this suppress effects was not restricted in liver only, but the whole body. Conclusion: Cell origin and abundance are decisive factors in stem cells applications and with the same premise of AMSCs and BMSCs, adipose tissue is a more promising origin source of stem cells. The immunoregulatory features of MSCs might play an important role in various MSCs cellular therapies. PMID:26339336

  8. Transplantation of mesenchymal stem cells promotes the functional recovery of the central nervous system following cerebral ischemia by inhibiting myelin-associated inhibitor expression and neural apoptosis

    PubMed Central

    FENG, NIANPING; HAO, GUANG; YANG, FENGGANG; QU, FUJUN; ZHENG, HAIHONG; LIANG, SONGLAN; JIN, YONGHUA

    2016-01-01

    Cerebral ischemia, which may lead to cerebral hypoxia and damage of the brain tissue, is a leading cause of human mortality and adult disability. Mesenchymal stem cells (MSCs) are a class of adult progenitor cells with the ability to differentiate into multiple cell types. The transplantation of bone marrow-derived MSCs is a potential therapeutic strategy for cerebral ischemia. However, the underlying mechanism has yet to be elucidated. In the present study, primary MSCs were isolated from healthy rats, labeled and transplanted into the brains of middle cerebral artery occlusion rat models. The location of the labeled MSCs in the rat brains were determined by fluorescent microscopy, and the neurological functions of the rats were scored. Immunohistochemical analyses demonstrated that the protein expression levels of myelin-associated inhibitors of regeneration, including Nogo-A, oligodendrocyte myelin glycoprotein and myelin-associated glycoprotein, were decreased following transplantation of the bone marrow-derived MSCs. Furthermore, the mRNA expression levels of Capase-3 and B-cell lymphoma 2, as determined by reverse transcription-quantitative polymerase chain reactions, were downregulated and upregulated, respectively, in the MSC-transplanted rats; thus suggesting that neural apoptosis was inhibited. The results of the present study suggested that the transplantation of bone marrow-derived MSCs was able to promote the functional recovery of the central nervous system following cerebral ischemia. Accordingly, inhibitors targeting myelin-associated inhibitors and apoptosis may be of clinical significance for cerebral ischemia in the future. PMID:27168778

  9. Cotransplantation of ex vivo expanded mesenchymal stem cells accelerates lymphocyte recovery and may reduce the risk of graft failure in haploidentical hematopoietic stem-cell transplantation.

    PubMed

    Ball, Lynne M; Bernardo, Maria Ester; Roelofs, Helene; Lankester, Arjan; Cometa, Angela; Egeler, R Maarten; Locatelli, Franco; Fibbe, Willem E

    2007-10-01

    Haploidentical hematopoietic stem-cell transplantation (HSCT) is associated with an increased risk of graft failure. Adult bone marrow-derived mesenchymal stromal cells (MSCs) have been shown to support in vivo normal hematopoiesis and to display potent immune suppressive effects. We cotransplanted donor MSCs in 14 children undergoing transplantation of HLA-disparate CD34(+) cells from a relative. While we observed a graft failure rate of 15% in 47 historic controls, all patients given MSCs showed sustained hematopoietic engraftment without any adverse reaction. In particular, children given MSCs did not experience more infections compared with controls. These data suggest that MSCs, possibly thanks to their potent immunosuppressive effect on alloreactive host T lymphocytes escaping the preparative regimen, reduce the risk of graft failure in haploidentical HSC transplant recipients. PMID:17638847

  10. Combined effects of flow-induced shear stress and electromagnetic field on neural differentiation of mesenchymal stem cells.

    PubMed

    Mascotte-Cruz, Juan Uriel; Ríos, Amelia; Escalante, Bruno

    2016-01-01

    Differentiation of bone marrow-derived mesenchymal stem cells (MSCs) into neural phenotype has been induced by either flow-induced shear stress (FSS) or electromagnetic fields (EMF). However, procedures are still expensive and time consuming. In the present work, induction for 1 h with the combination of both forces showed the presence of the neural precursor nestin as early as 9 h in culture after treatment and this result lasted for the following 6 d. In conclusion, the use of a combination of FSS and EMF for a short-time renders in neurite-like cells, although further investigation is required to analyze cell functionality. PMID:26325339

  11. Potential of bone marrow mesenchymal stem cells in management of Alzheimer's disease in female rats.

    PubMed

    Salem, Ahmed M; Ahmed, Hanaa H; Atta, Hazem M; Ghazy, Mohamed A; Aglan, Hadeer A

    2014-12-01

    Alzheimer's disease (AD) has been called the disease of the century with significant clinical and socioeconomic impacts. Pharmacological treatment has limited efficacy and only provides symptomatic relief without long-term cure. Accordingly, there is an urgent need to develop novel and effective medications for AD. Stem cell-based therapy is a promising approach to handling neurodegenerative diseases. Therefore, the current study aimed to explore the possible therapeutic role of single intravenous injection of bone marrow derived mesenchymal stem cells (BM-MSCs) after 4 months in management of AD in the experimental model. The work also extended to compare the therapeutic potential of BM-MSCs with 2 conventional therapies of AD; rivastigmine and cerebrolysin administered daily. BM-MSCs were able to home at the injured brains and produced significant increases in the number of positive cells for choline acetyltransferase (ChAT) and survivin expression, as well as selective AD indicator-1 (seladin-1) and nestin gene expression. Histopathological examination indicated that BM-MSCs could remove beta-amyloid plaques from hippocampus. Significant improvement in these biomarkers was similar to or better sometimes than the reference drugs, clearly showing the potential therapeutic role of BM-MSCs against AD through their anti-apoptotic, neurogenic and immunomodulatory properties. PMID:25044885

  12. Olfactory ecto-mesenchymal stem cells possess immunoregulatory function and suppress autoimmune arthritis

    PubMed Central

    Rui, Ke; Zhang, Zhijiang; Tian, Jie; Lin, Xiang; Wang, Xiaohui; Ma, Jie; Tang, Xinyi; Xu, Huaxi; Lu, Liwei; Wang, Shengjun

    2016-01-01

    Recent studies have identified olfactory ecto-mesenchymal stem cells (OE-MSCs) as a new type of resident stem cell in the olfactory lamina propria. However, it remains unclear whether OE-MSCs possess any immunoregulatory functions. In this study, we found that mouse OE-MSCs expressed higher transforming growth factor-beta and interleukin-10 levels than bone marrow-derived MSCs. In culture, OE-MSCs exerted their immunosuppressive capacity via directly suppressing effector T-cell proliferation and increasing regulatory T (Treg) cell expansion. In mice with collagen-induced arthritis, adoptive transfer of OE-MSCs markedly suppressed arthritis onset and disease severity, which was accompanied by increased Treg cells and reduced Th1/Th17 cell responses in vivo. Taken together, our findings identified a novel function of OE-MSCs in regulating T-cell responses, indicating that OE-MSCs may represent a new cell therapy for the treatment of rheumatoid arthritis and other autoimmune diseases. PMID:26388237

  13. Myocyte-Depleted Engineered Cardiac Tissues Support Therapeutic Potential of Mesenchymal Stem Cells

    PubMed Central

    Serrao, Gregory W.; Turnbull, Irene C.; Ancukiewicz, Damian; Kim, Do Eun; Kao, Evan; Cashman, Timothy J.; Hadri, Lahouaria; Hajjar, Roger J.

    2012-01-01

    The therapeutic potential of mesenchymal stem cells (MSCs) for restoring cardiac function after cardiomyocyte loss remains controversial. Engineered cardiac tissues (ECTs) offer a simplified three-dimensional in vitro model system to evaluate stem cell therapies. We hypothesized that contractile properties of dysfunctional ECTs would be enhanced by MSC treatment. ECTs were created from neonatal rat cardiomyocytes with and without bone marrow-derived adult rat MSCs in a type-I collagen and Matrigel scaffold using custom elastomer molds with integrated cantilever force sensors. Three experimental groups included the following: (1) baseline condition ECT consisting only of myocytes, (2) 50% myocyte-depleted ECT, modeling a dysfunctional state, and (3) 50% myocyte-depleted ECT plus 10% MSC, modeling dysfunctional myocardium with intervention. Developed stress (DS) and pacing threshold voltage (VT) were measured using 2-Hz field stimulation at 37°C on culture days 5, 10, 15, and 20. By day 5, DS of myocyte-depleted ECTs was significantly lower than baseline, and VT was elevated. In MSC-supplemented ECTs, DS and VT were significantly better than myocyte-depleted values, approaching baseline ECTs. Findings were similar through culture day 15, but lost significance at day 20. Trends in DS were partly explained by changes in the cell number and alignment with time. Thus, supplementing myocyte-depleted ECTs with MSCs transiently improved contractile function and compensated for a 50% loss of cardiomyocytes, mimicking recent animal studies and clinical trials and supporting the potential of MSCs for myocardial therapy. PMID:22500611

  14. Efficient gene silencing in mesenchymal stem cells by substrate-mediated RNA interference.

    PubMed

    Hsu, Shan-Hui; Huang, Guo-Shiang; Ho, Tung-Tso; Feng, Fuh

    2014-11-01

    We described a novel substrate-mediated RNA interference (RNAi) technology to investigate the effect of neural crest marker expression on the multipotency of human gingival fibroblasts (HGFs). HGFs showed significantly higher neural and chondrogenic differentiation potentials compared with adult bone-marrow-derived mesenchymal stem cells and stem cells from human exfoliated deciduous teeth. By sending target-specific RNAi agents with the conventional vehicle (PolyFect), we observed that the multipotency of HGFs was closely associated with the expression of neural crest marker gene Forkhead box D3 (FoxD3). Using the novel chitosan substrate-mediated method, we successfully delivered short-hairpin RNA constructs to HGFs grown on chitosan without the use of conventional vehicles. The delivery efficiency measured by flow cytometry showed a 10-fold increase for HGFs on chitosan versus those on culture dish, and the cell viability was >95%. Moreover, HGFs with FoxD3 gene knockdown did not form spheroids on chitosan. Based on this working principle, we further selected the gene-silenced population from HGFs. The nonsilenced HGFs showed much higher neural differentiation ability with the nestin expression 40-fold greater than FoxD3-silenced population after induction, suggesting the feasibility of the method to silence genes. The new substrate-mediated gene silencing platform that combines the use of substrate and RNAi can be used to clarify the functions of important genes without suffering the toxicity. PMID:24624901

  15. Innovative Strategy for MicroRNA Delivery in Human Mesenchymal Stem Cells via Magnetic Nanoparticles

    PubMed Central

    Schade, Anna; Delyagina, Evgenya; Scharfenberg, Dorothee; Skorska, Anna; Lux, Cornelia; David, Robert; Steinhoff, Gustav

    2013-01-01

    Bone marrow derived human mesenchymal stem cells (hMSCs) show promising potential in regeneration of defective tissue. Recently, gene silencing strategies using microRNAs (miR) emerged with the aim to expand the therapeutic potential of hMSCs. However, researchers are still searching for effective miR delivery methods for clinical applications. Therefore, we aimed to develop a technique to efficiently deliver miR into hMSCs with the help of a magnetic non-viral vector based on cationic polymer polyethylenimine (PEI) bound to iron oxide magnetic nanoparticles (MNP). We tested different magnetic complex compositions and determined uptake efficiency and cytotoxicity by flow cytometry. Additionally, we monitored the release, processing and functionality of delivered miR-335 with confocal laser scanning microscopy, real-time PCR and live cell imaging, respectively. On this basis, we established parameters for construction of magnetic non-viral vectors with optimized uptake efficiency (~75%) and moderate cytotoxicity in hMSCs. Furthermore, we observed a better transfection performance of magnetic complexes compared to PEI complexes 72 h after transfection. We conclude that MNP-mediated transfection provides a long term effect beneficial for successful genetic modification of stem cells. Hence, our findings may become of great importance for future in vivo applications. PMID:23702843

  16. Transplantation of placenta-derived mesenchymal stem cells in the EAE mouse model of MS.

    PubMed

    Fisher-Shoval, Yonit; Barhum, Yael; Sadan, Ofer; Yust-Katz, Shlomit; Ben-Zur, Tali; Lev, Nirit; Benkler, Chen; Hod, Moshe; Melamed, Eldad; Offen, Daniel

    2012-09-01

    Stem cell-based regenerative medicine raises great hope for the treatment of multiple sclerosis (MS). Bone marrow-derived mesenchymal stem cells (BM-MSCs) are being tested in clinical trials. Bone marrow is the traditional source of human MSCs, but human term placenta appears to be an excellent alternative because of its availability, without ethical issues. In this study, the therapeutic effect of human placental MSCs (PL-MSCs) was evaluated in experimental autoimmune encephalomyelitis (EAE), the mice model of MS. EAE mice were transplanted intra-cerebrally with PL-MSCs or with the vehicle saline 5 or 10 days after first MOG injection. The mice were monitored for a month after therapy. A daily EAE score revealed a decrease in disease severity in the transplanted animals when compared to saline. Survival was significantly higher in the transplanted animals. In vitro experiments demonstrated that conditioned media from LPS-activated astrocytes stimulated PL-MSCs to express the gene TNF-α-stimulated gene/protein 6 (TSG-6). The same mRNA expression was obtained when PL-MSCs were exposed to TNF-α or IL1-β. These results demonstrate that PL-MSCs have a therapeutic effect in the EAE mice model. We assume that this effect is caused by reduction of the anti-inflammatory protein, TSG-6, of the inflammatory damage. PMID:22638856

  17. Intraventricular injections of mesenchymal stem cells activate endogenous functional remyelination in a chronic demyelinating murine model

    PubMed Central

    Cruz-Martinez, P; González-Granero, S; Molina-Navarro, M M; Pacheco-Torres, J; García-Verdugo, J M; Geijo-Barrientos, E; Jones, J; Martinez, S

    2016-01-01

    Current treatments for demyelinating diseases are generally only capable of ameliorating the symptoms, with little to no effect in decreasing myelin loss nor promoting functional recovery. Mesenchymal stem cells (MSCs) have been shown by many researchers to be a potential therapeutic tool in treating various neurodegenerative diseases, including demyelinating disorders. However, in the majority of the cases, the effect was only observed locally, in the area surrounding the graft. Thus, in order to achieve general remyelination in various brain structures simultaneously, bone marrow-derived MSCs were transplanted into the lateral ventricles (LVs) of the cuprizone murine model. In this manner, the cells may secrete soluble factors into the cerebrospinal fluid (CSF) and boost the endogenous oligodendrogenic potential of the subventricular zone (SVZ). As a result, oligodendrocyte progenitor cells (OPCs) were recruited within the corpus callosum (CC) over time, correlating with an increased myelin content. Electrophysiological studies, together with electron microscopy (EM) analysis, indicated that the newly formed myelin correctly enveloped the demyelinated axons and increased signal transduction through the CC. Moreover, increased neural stem progenitor cell (NSPC) proliferation was observed in the SVZ, possibly due to the tropic factors released by the MSCs. In conclusion, the findings of this study revealed that intraventricular injections of MSCs is a feasible method to elicit a paracrine effect in the oligodendrogenic niche of the SVZ, which is prone to respond to the factors secreted into the CSF and therefore promoting oligodendrogenesis and functional remyelination. PMID:27171265

  18. Myocyte-depleted engineered cardiac tissues support therapeutic potential of mesenchymal stem cells.

    PubMed

    Serrao, Gregory W; Turnbull, Irene C; Ancukiewicz, Damian; Kim, Do Eun; Kao, Evan; Cashman, Timothy J; Hadri, Lahouaria; Hajjar, Roger J; Costa, Kevin D

    2012-07-01

    The therapeutic potential of mesenchymal stem cells (MSCs) for restoring cardiac function after cardiomyocyte loss remains controversial. Engineered cardiac tissues (ECTs) offer a simplified three-dimensional in vitro model system to evaluate stem cell therapies. We hypothesized that contractile properties of dysfunctional ECTs would be enhanced by MSC treatment. ECTs were created from neonatal rat cardiomyocytes with and without bone marrow-derived adult rat MSCs in a type-I collagen and Matrigel scaffold using custom elastomer molds with integrated cantilever force sensors. Three experimental groups included the following: (1) baseline condition ECT consisting only of myocytes, (2) 50% myocyte-depleted ECT, modeling a dysfunctional state, and (3) 50% myocyte-depleted ECT plus 10% MSC, modeling dysfunctional myocardium with intervention. Developed stress (DS) and pacing threshold voltage (VT) were measured using 2-Hz field stimulation at 37°C on culture days 5, 10, 15, and 20. By day 5, DS of myocyte-depleted ECTs was significantly lower than baseline, and VT was elevated. In MSC-supplemented ECTs, DS and VT were significantly better than myocyte-depleted values, approaching baseline ECTs. Findings were similar through culture day 15, but lost significance at day 20. Trends in DS were partly explained by changes in the cell number and alignment with time. Thus, supplementing myocyte-depleted ECTs with MSCs transiently improved contractile function and compensated for a 50% loss of cardiomyocytes, mimicking recent animal studies and clinical trials and supporting the potential of MSCs for myocardial therapy. PMID:22500611

  19. Persistent circulating human insulin in sheep transplanted in utero with human mesenchymal stem cells

    PubMed Central

    Ersek, Adel; Pixley, John S.; Goodrich, A. Daisy; Porada, Christopher D.; Almeida-Porada, Graca; Thain, David S.; Zanjani, Esmail D.

    2010-01-01

    Objective To determine if mesenchymal stem cells (MSC) derived from human fetal pancreatic tissue (pMSC) would engraft and differentiate in sheep pancreas following transplantation in utero. Methods A three-step culture system was established for generating human fetal pMSC. Sheep fetuses were transplanted during the fetal transplant receptivity period with human pMSC and evaluated for in situ and functional engraftment in their pancreas, liver and bone marrow. Results Isolation and expansion of adherent cells from the human fetal pancreas yielded a cell population with morphologic and phenotypic characteristics similar to MSC derived from bone marrow. This putative stem cell population could undergo multilineage differentiation in vitro. Three to 27 months after fetal transplantation, the pancreatic engraftment frequency (chimeric index) was 79% while functional engraftment was noted in 50% of transplanted sheep. Hepatic and marrow engraftment and expression was noted as well. Conclusion We have established a procedure for isolation of human fetal pMSC that display characteristics similar to bone marrow derived MSC. In vivo results suggest the pMSC engraft, differentiate and secrete human insulin from the sheep pancreas. PMID:20170708

  20. Concise Review: Mesenchymal Stem Cells for Diabetes

    PubMed Central

    Domínguez-Bendala, Juan; Lanzoni, Giacomo

    2012-01-01

    Mesenchymal stem cells (MSCs) have already made their mark in the young field of regenerative medicine. Easily derived from many adult tissues, their therapeutic worth has already been validated for a number of conditions. Unlike embryonic stem cells, neither their procurement nor their use is deemed controversial. Here we review the potential use of MSCs for the treatment of type 1 diabetes mellitus, a devastating chronic disease in which the insulin-producing cells of the pancreas (the β-cells) are the target of an autoimmune process. It has been hypothesized that stem cell-derived β-cells may be used to replenish the islet mass in diabetic patients, making islet transplantation (a form of cell therapy that has already proven effective at clinically restoring normoglycemia) available to millions of prospective patients. Here we review the most current advances in the design and application of protocols for the differentiation of transplantable β-cells, with a special emphasis in analyzing MSC potency according to their tissue of origin. Although no single method appears to be ripe enough for clinical trials yet, recent progress in reprogramming (a biotechnological breakthrough that relativizes the thus far insurmountable barriers between embryonal germ layers) bodes well for the rise of MSCs as a potential weapon of choice to develop personalized therapies for type 1 diabetes. PMID:23197641

  1. Concise review: mesenchymal stem cells for diabetes.

    PubMed

    Domínguez-Bendala, Juan; Lanzoni, Giacomo; Inverardi, Luca; Ricordi, Camillo

    2012-01-01

    Mesenchymal stem cells (MSCs) have already made their mark in the young field of regenerative medicine. Easily derived from many adult tissues, their therapeutic worth has already been validated for a number of conditions. Unlike embryonic stem cells, neither their procurement nor their use is deemed controversial. Here we review the potential use of MSCs for the treatment of type 1 diabetes mellitus, a devastating chronic disease in which the insulin-producing cells of the pancreas (the β-cells) are the target of an autoimmune process. It has been hypothesized that stem cell-derived β-cells may be used to replenish the islet mass in diabetic patients, making islet transplantation (a form of cell therapy that has already proven effective at clinically restoring normoglycemia) available to millions of prospective patients. Here we review the most current advances in the design and application of protocols for the differentiation of transplantable β-cells, with a special emphasis in analyzing MSC potency according to their tissue of origin. Although no single method appears to be ripe enough for clinical trials yet, recent progress in reprogramming (a biotechnological breakthrough that relativizes the thus far insurmountable barriers between embryonal germ layers) bodes well for the rise of MSCs as a potential weapon of choice to develop personalized therapies for type 1 diabetes. PMID:23197641

  2. Epigenetic Mechanisms Regulating Mesenchymal Stem Cell Differentiation

    PubMed Central

    Pérez-Campo, Flor M.; Riancho, José A.

    2015-01-01

    Human Mesenchymal Stem Cells (hMSCs) have emerged in the last few years as one of the most promising therapeutic cell sources and, in particular, as an important tool for regenerative medicine of skeletal tissues. Although they present a more restricted potency than Embryonic Stem (ES) cells, the use of hMCS in regenerative medicine avoids many of the drawbacks characteristic of ES cells or induced pluripotent stem cells. The challenge in using these cells lies into developing precise protocols for directing cellular differentiation to generate a specific cell lineage. In order to achieve this goal, it is of the upmost importance to be able to control de process of fate decision and lineage commitment. This process requires the coordinate regulation of different molecular layers at transcriptional, posttranscriptional and translational levels. At the transcriptional level, switching on and off different sets of genes is achieved not only through transcriptional regulators, but also through their interplay with epigenetic modifiers. It is now well known that epigenetic changes take place in an orderly way through development and are critical in the determination of lineage-specific differentiation. More importantly, alteration of these epigenetic changes would, in many cases, lead to disease generation and even tumour formation. Therefore, it is crucial to elucidate how epigenetic factors, through their interplay with transcriptional regulators, control lineage commitment in hMSCs. PMID:27019612

  3. Mechanical regulation of mesenchymal stem cell differentiation.

    PubMed

    Steward, Andrew J; Kelly, Daniel J

    2015-12-01

    Biophysical cues play a key role in directing the lineage commitment of mesenchymal stem cells or multipotent stromal cells (MSCs), but the mechanotransductive mechanisms at play are still not fully understood. This review article first describes the roles of both substrate mechanics (e.g. stiffness and topography) and extrinsic mechanical cues (e.g. fluid flow, compression, hydrostatic pressure, tension) on the differentiation of MSCs. A specific focus is placed on the role of such factors in regulating the osteogenic, chondrogenic, myogenic and adipogenic differentiation of MSCs. Next, the article focuses on the cellular components, specifically integrins, ion channels, focal adhesions and the cytoskeleton, hypothesized to be involved in MSC mechanotransduction. This review aims to illustrate the strides that have been made in elucidating how MSCs sense and respond to their mechanical environment, and also to identify areas where further research is needed. PMID:25382217

  4. Proteomic Analysis of Mesenchymal Stem Cells.

    PubMed

    Faça, Vitor Marcel; Orellana, Maristela Delgado; Greene, Lewis Joel; Covas, Dimas Tadeu

    2016-01-01

    Mesenchymal stem or stromal cells (MSCs) are of great interest in biomedical sciences and disease treatment because of their multipotency and wide range of applications for tissue repair and suppression of the immune system. Proteomic analysis of these unique cells has contributed to the identification of important pathways utilized by MSCs to differentiate into distinct tissues as well as important proteins responsible for their special function in vivo and in vitro. However, comparison of proteomic studies in MSCs still suffers from the heterogeneity of MSC preparations. In addition, as proteomics technology advances, several studies can be revisited in order to increase the depth of analysis and, therefore, elucidate more refined mechanisms involved in MSC functionalities. Here, we present detailed protocols to obtain MSCs, as well as protocols to perform in-depth profiling and quantification of alterations in MSC proteomes. PMID:27236693

  5. Mesenchymal Stem Cells: Angels or Demons?

    PubMed Central

    Wong, Rebecca S. Y.

    2011-01-01

    Mesenchymal stem cells (MSCs) have been used in cell-based therapy in various disease conditions such as graft-versus-host and heart diseases, osteogenesis imperfecta, and spinal cord injuries, and the results have been encouraging. However, as MSC therapy gains popularity among practitioners and researchers, there have been reports on the adverse effects of MSCs especially in the context of tumour modulation and malignant transformation. These cells have been found to enhance tumour growth and metastasis in some studies and have been related to anticancer-drug resistance in other instances. In addition, various studies have also reported spontaneous malignant transformation of MSCs. The mechanism of the modulatory behaviour and the tumorigenic potential of MSCs, warrant urgent exploration, and the use of MSCs in patients with cancer awaits further evaluation. However, if MSCs truly play a role in tumour modulation, they can also be potential targets of cancer treatment. PMID:21822372

  6. Role of Human Corneal Stroma-Derived Mesenchymal-Like Stem Cells in Corneal Immunity and Wound Healing

    PubMed Central

    Veréb, Zoltán; Póliska, Szilárd; Albert, Réka; Olstad, Ole Kristoffer; Boratkó, Anita; Csortos, Csilla; Moe, Morten C.; Facskó, Andrea; Petrovski, Goran

    2016-01-01

    Corneal tissue regeneration is of crucial importance for maintaining normal vision. We aimed to isolate and cultivate human corneal stroma-derived mesenchymal stem-like cells (CSMSCs) from the central part of cadaver corneas and study their phenotype, multipotency, role in immunity and wound healing. The isolated cells grew as monolayers in vitro, expressed mesenchymal- and stemness-related surface markers (CD73, CD90, CD105, CD140b), and were negative for hematopoietic markers as determined by flow cytometry. CSMSCs were able to differentiate in vitro into fat, bone and cartilage. Their gene expression profile was closer to bone marrow-derived MSCs (BMMSCs) than to limbal epithelial stem cells (LESC) as determined by high-throughput screening. The immunosuppressive properties of CSMSCs were confirmed by a mixed lymphocyte reaction (MLR), while they could inhibit proliferation of activated immune cells. Treatment of CSMSCs by pro-inflammatory cytokines and toll-like receptor ligands significantly increased the secreted interleukin-6 (IL-6), interleukin-8 (IL-8) and C-X-C motif chemokine 10 (CXCL-10) levels, as well as the cell surface adhesion molecules. CSMSCs were capable of closing a wound in vitro under different stimuli. These cells thus contribute to corneal tissue homeostasis and play an immunomodulatory and regenerative role with possible implications in future cell therapies for treating sight-threatening corneal diseases. PMID:27195722

  7. Role of Human Corneal Stroma-Derived Mesenchymal-Like Stem Cells in Corneal Immunity and Wound Healing.

    PubMed

    Veréb, Zoltán; Póliska, Szilárd; Albert, Réka; Olstad, Ole Kristoffer; Boratkó, Anita; Csortos, Csilla; Moe, Morten C; Facskó, Andrea; Petrovski, Goran

    2016-01-01

    Corneal tissue regeneration is of crucial importance for maintaining normal vision. We aimed to isolate and cultivate human corneal stroma-derived mesenchymal stem-like cells (CSMSCs) from the central part of cadaver corneas and study their phenotype, multipotency, role in immunity and wound healing. The isolated cells grew as monolayers in vitro, expressed mesenchymal- and stemness-related surface markers (CD73, CD90, CD105, CD140b), and were negative for hematopoietic markers as determined by flow cytometry. CSMSCs were able to differentiate in vitro into fat, bone and cartilage. Their gene expression profile was closer to bone marrow-derived MSCs (BMMSCs) than to limbal epithelial stem cells (LESC) as determined by high-throughput screening. The immunosuppressive properties of CSMSCs were confirmed by a mixed lymphocyte reaction (MLR), while they could inhibit proliferation of activated immune cells. Treatment of CSMSCs by pro-inflammatory cytokines and toll-like receptor ligands significantly increased the secreted interleukin-6 (IL-6), interleukin-8 (IL-8) and C-X-C motif chemokine 10 (CXCL-10) levels, as well as the cell surface adhesion molecules. CSMSCs were capable of closing a wound in vitro under different stimuli. These cells thus contribute to corneal tissue homeostasis and play an immunomodulatory and regenerative role with possible implications in future cell therapies for treating sight-threatening corneal diseases. PMID:27195722

  8. Labeling and Imaging Mesenchymal Stem Cells with Quantum Dots

    EPA Science Inventory

    Mesenchymal stem cells (MSCs) are multipotent cells with the potential to differentiate into bone, cartilage, adipose and muscle cells. Adult derived MSCs are being actively investigated because of their potential to be utilized for therapeutic cell-based transplantation. Methods...

  9. Mesenchymal stem cell therapy for heart disease.

    PubMed

    Gnecchi, Massimiliano; Danieli, Patrizia; Cervio, Elisabetta

    2012-08-19

    Mesenchymal stem cells (MSC) are adult stem cells with capacity for self-renewal and multi-lineage differentiation. Initially described in the bone marrow, MSC are also present in other organs and tissues. From a therapeutic perspective, because of their easy preparation and immunologic privilege, MSC are emerging as an extremely promising therapeutic agent for tissue regeneration and repair. Studies in animal models of myocardial infarction have demonstrated the ability of transplanted MSC to engraft and differentiate into cardiomyocytes and vascular cells. Most importantly, engrafted MSC secrete a wide array of soluble factors that mediate beneficial paracrine effects and may greatly contribute to cardiac repair. Together, these properties can be harnessed to both prevent and reverse remodeling in the ischemically injured ventricle. In proof-of-concept and phase I clinical trials, MSC therapy improved left ventricular function, induced reverse remodeling, and decreased scar size. In this review we will focus on the current understanding of MSC biology and MSC mechanism of action in cardiac repair. PMID:22521741

  10. A fat option for the pig: Hepatocytic differentiated mesenchymal stem cells for translational research

    SciTech Connect

    Brückner, Sandra; Tautenhahn, Hans-Michael; Winkler, Sandra; Stock, Peggy; Dollinger, Matthias; Christ, Bruno

    2014-02-15

    Study background: Extended liver resection is the only curative treatment option of liver cancer. Yet, the residual liver may not accomplish the high metabolic and regenerative capacity needed, which frequently leads to acute liver failure. Because of their anti-inflammatory and -apoptotic as well as pro-proliferative features, mesenchymal stem cells differentiated into hepatocyte-like cells might provide functional and regenerative compensation. Clinical translation of basic research requires pre-clinical approval in large animals. Therefore, we characterized porcine mesenchymal stem cells (MSC) from adipose tissue and bone marrow and their hepatocyte differentiation potential for future assessment of functional liver support after surgical intervention in the pig model. Methods: Mesenchymal surface antigens and multi-lineage differentiation potential of porcine MSC isolated by collagenase digestion either from bone marrow or adipose tissue (subcutaneous/visceral) were assessed by flow cytometry. Morphology and functional properties (urea-, glycogen synthesis and cytochrome P450 activity) were determined during culture under differentiation conditions and compared with primary porcine hepatocytes. Results: MSC from porcine adipose tissue and from bone marrow express the typical mesenchymal markers CD44, CD29, CD90 and CD105 but not haematopoietic markers. MSC from both sources displayed differentiation into the osteogenic as well as adipogenic lineage. After hepatocyte differentiation, expression of CD105 decreased significantly and cells adopted the typical polygonal morphology of hepatocytes. Glycogen storage was comparable in adipose tissue- and bone marrow-derived cells. Urea synthesis was about 35% lower in visceral than in subcutaneous adipose tissue-derived MSC. Cytochrome P450 activity increased significantly during differentiation and was twice as high in hepatocyte-like cells generated from bone marrow as from adipose tissue. Conclusion: The hepatocyte

  11. Mesenchymal stem cell implantation in atrophic nonunion of the long bones

    PubMed Central

    Phedy, P.; Kholinne, E.; Djaja, Y. P.; Kusnadi, Y.; Merlina, M.; Yulisa, N. D.

    2016-01-01

    Objectives To explore the therapeutic potential of combining bone marrow-derived mesenchymal stem cells (BM-MSCs) and hydroxyapatite (HA) granules to treat nonunion of the long bone. Methods Ten patients with an atrophic nonunion of a long bone fracture were selectively divided into two groups. Five subjects in the treatment group were treated with the combination of 15 million autologous BM-MSCs, 5g/cm3 (HA) granules and internal fixation. Control subjects were treated with iliac crest autograft, 5g/cm3 HA granules and internal fixation. The outcomes measured were post-operative pain (visual analogue scale), level of functionality (LEFS and DASH), and radiograph assessment. Results Post-operative pain evaluation showed no significant differences between the two groups. The treatment group demonstrated faster initial radiographic and functional improvements. Statistically significant differences in functional scores were present during the first (p = 0.002), second (p = 0.005) and third (p = 0.01) month. Both groups achieved similar outcomes by the end of one-year follow-up. No immunologic or neoplastic side effects were reported. Conclusions All cases of nonunion of a long bone presented in this study were successfully treated using autologous BM-MSCs. The combination of autologous BM-MSCs and HA granules is a safe method for treating nonunion. Patients treated with BM-MSCs had faster initial radiographic and functional improvements. By the end of 12 months, both groups had similar outcomes. Cite this article: H.D. Ismail, P. Phedy, E. Kholinne, Y. P. Djaja, Y. Kusnadi, M. Merlina, N. D. Yulisa. Mesenchymal stem cell implantation in atrophic nonunion of the long bones: A translational study. Bone Joint Res 2016;5:287–293. DOI: 10.1302/2046-3758.57.2000587. PMID:27412657

  12. Single-cell-derived mesenchymal stem cells overexpressing Csx/Nkx2.5 and GATA4 undergo the stochastic cardiomyogenic fate and behave like transient amplifying cells

    SciTech Connect

    Yamada, Yoji; Sakurada, Kazuhiro; Takeda, Yukiji; Gojo, Satoshi; Umezawa, Akihiro . E-mail: umezawa@1985.jukuin.keio.ac.jp

    2007-02-15

    Bone marrow-derived stromal cells can give rise to cardiomyocytes as well as adipocytes, osteocytes, and chondrocytes in vitro. The existence of mesenchymal stem cells has been proposed, but it remains unclear if a single-cell-derived stem cell stochastically commits toward a cardiac lineage. By single-cell marking, we performed a follow-up study of individual cells during the differentiation of 9-15c mesenchymal stromal cells derived from bone marrow cells. Three types of cells, i.e., cardiac myoblasts, cardiac progenitors and multipotent stem cells were differentiated from a single cell, implying that cardiomyocytes are generated stochastically from a single-cell-derived stem cell. We also demonstrated that overexpression of Csx/Nkx2.5 and GATA4, precardiac mesodermal transcription factors, enhanced cardiomyogenic differentiation of 9-15c cells, and the frequency of cardiomyogenic differentiation was increased by co-culturing with fetal cardiomyocytes. Single-cell-derived mesenchymal stem cells overexpressing Csx/Nkx2.5 and GATA4 behaved like cardiac transient amplifying cells, and still retained their plasticity in vivo.

  13. Mesenchymal stem cells: From stem cells to sarcomas.

    PubMed

    Lye, Kwan Liang; Nordin, Norshariza; Vidyadaran, Sharmili; Thilakavathy, Karuppiah

    2016-06-01

    Mesenchymal stem cells (MSCs) have garnered vast interests in clinical settings, especially in regenerative medicine due to their unique properties-they are reliably isolated and expanded from various tissue sources; they are able to differentiate into mesodermal tissues such as bones, cartilages, adipose tissues, and muscles; and they have unique immunosuppressive properties. However, there are some concerns pertaining to the role of MSCs in the human body. On one hand, they are crucial component in the regeneration and repair of the human body. On the contrary, they are shown to transform into sarcomas. Although the exact mechanisms are still unknown, many new leads have pointed to the belief that MSCs do play a role in sarcomagenesis. This review focuses on the current updates and findings of the role of MSCs in their transformation process into sarcomas. PMID:26992453

  14. An Investigation of Equine Mesenchymal Stem Cell Characteristics from Different Harvest Sites: More Similar Than Not

    PubMed Central

    Lombana, Karla G.; Goodrich, Laurie R.; Phillips, Jennifer Nikki; Kisiday, John David; Ruple-Czerniak, Audrey; McIlwraith, C. Wayne

    2015-01-01

    Diseases of the musculoskeletal system are a major cause of loss of use and retirement in sport horses. The use of bone marrow-derived mesenchymal stem cells (BMDMSCs) for healing of traumatized tissue has gained substantial favor in clinical settings and can assist healing and tissue regeneration in orthopedic injuries. There are two common sites of harvest of BMDMSCs, the sternum and the ilium. Our objective was to determine if any differences exist in BMDMSCs acquired from the sternum and the ilium. We compared the two harvest sites in their propensity to undergo multilineage differentiation, differences in cell surface markers, or gene transduction efficiencies. BMDMSCs were isolated and culture-expanded from 5 ml aspirates of bone marrow from sternum and ilium. The cells were then plated and cultured with appropriate differentiation medium to result in multi-lineage differentiation and cell characteristics were compared between sternal and ilial samples. Cell surface antibody expression of CD11a/18, CD34, CD44, and CD90 were evaluated using flow cytometry, and gene transduction efficiencies were evaluated using GFP scAAV. There were no statistically significant differences in cell characteristics between MSCs cultured from the sternum and the ilium under any circumstances. PMID:26664993

  15. Allogeneic Mesenchymal Stem Cells in Combination with Hyaluronic Acid for the Treatment of Osteoarthritis in Rabbits

    PubMed Central

    Chiang, En-Rung; Ma, Hsiao-Li; Wang, Jung-Pan; Liu, Chien-Lin; Chen, Tain-Hsiung; Hung, Shih-Chieh

    2016-01-01

    Mesenchymal stem cell (MSC)-based therapies may aid in the repair of articular cartilage defects. The purpose of this study was to investigate the effects of intraarticular injection of allogeneic MSCs in an in vivo anterior cruciate ligament transection (ACLT) model of osteoarthritis in rabbits. Allogeneic bone marrow-derived MSCs were isolated and cultured under hypoxia (1% O2). After 8 weeks following ACLT, MSCs suspended in hyaluronic acid (HA) were injected into the knees, and the contralateral knees were injected with HA alone. Additional controls consisted of a sham operation group as well as an untreated osteoarthritis group. The tissues were analyzed by macroscopic examination as well as histologic and immunohistochemical methods at 6 and 12 weeks post-transplantation. At 6 and 12 weeks, the joint surface showed less cartilage loss and surface abrasion after MSC injection as compared to the tissues receiving HA injection alone. Significantly better histological scores and cartilage content were observed with the MSC transplantation. Furthermore, engraftment of allogenic MSCs were evident in surface cartilage. Thus, injection of the allogeneic MSCs reduced the progression of osteoarthritis in vivo. PMID:26915044

  16. Effect of microfabricated microgroove-surface devices on the morphology of mesenchymal stem cells.

    PubMed

    Zhang, Xiangkai; Aoyama, Tomoki; Yasuda, Takashi; Oike, Makoto; Ito, Akira; Tajino, Junichi; Nagai, Momoko; Fujioka, Rune; Iijima, Hirotaka; Yamaguchi, Shoki; Kakinuma, Norihiro; Kuroki, Hiroshi

    2015-12-01

    The surface of a material that is in contact with cells is known to affect cell morphology and function. To develop an appropriate surface for tendon engineering, we used zigzag microgroove surfaces, which are similar to the tenocyte microenvironment. The purpose of this study was to investigate the effect of microgroove surfaces with different ridge angles (RAs), ridge lengths (RLs), ridge widths (RWs), and groove widths (GWs) on human bone marrow-derived mesenchymal stem cell (MSC) shape. Dishes with microgroove surfaces were fabricated using cyclic olefin polymer by injection-compression molding. The other parameters were fixed, and effects of different RAs (180 - 30 °), RLs (5 - 500 μm), RWs (5 - 500 μm), and GWs (5 - 500 μm) were examined. Changes in the zigzag shape of the cell due to different RAs, RLs, RWs, and GWs were observed by optical microscopy and scanning electron microscopy. Cytoskeletal changes were investigated using Phalloidin immunofluorescence staining. As observed by optical microscopy, MSCs changed to a zigzag shape in response to microgroove surfaces with different ridge and groove properties. . As observed by scanning electron microscopy, the cell shape changed at turns in the microgroove surface. Phalloidin immunofluorescence staining indicated that F-actin, not only in cell filopodia but also inside the cell body, changed orientation to conform to the microgrooves. In conclusion, the use of zigzag microgroove surfaces microfabricated by injection-compression molding demonstrated the property of MSCs to alter their shapes to fit the surface. PMID:26573821

  17. Rapid fusion between mesenchymal stem cells and cardiomyocytes yields electrically active, non-contractile hybrid cells

    PubMed Central

    Shadrin, Ilya Y.; Yoon, Woohyun; Li, Liqing; Shepherd, Neal; Bursac, Nenad

    2015-01-01

    Cardiac cell therapies involving bone marrow-derived human mesenchymal stem cells (hMSCs) have shown promising results, although their mechanisms of action are still poorly understood. Here, we investigated direct interactions between hMSCs and cardiomyocytes in vitro. Using a genetic Ca2+ indicator gCaMP3 to efficiently label hMSCs in co-cultures with neonatal rat ventricular myocytes (NRVMs), we determined that 25–40% of hMSCs (from 4 independent donors) acquired periodic Ca2+ transients and cardiac markers through spontaneous fusion with NRVMs. Sharp electrode and voltage-clamp recordings in fused cells showed action potential properties and Ca2+ current amplitudes in between those of non-fused hMSCs and NRVMs. Time-lapse video-microscopy revealed the first direct evidence of active fusion between hMSCs and NRVMs within several hours of co-culture. Application of blebbistatin, nifedipine or verapamil caused complete and reversible inhibition of fusion, suggesting potential roles for actomyosin bridging and Ca2+ channels in the fusion process. Immunostaining for Cx43, Ki67, and sarcomeric α-actinin showed that fused cells remain strongly coupled to surrounding NRVMs, but downregulate sarcomeric structures over time, acquiring a non-proliferative and non-contractile phenotype. Overall, these results describe the phenotype and mechanisms of hybrid cell formation via fusion of hMSCs and cardiomyocytes with potential implications for cardiac cell therapy. PMID:26159124

  18. Activated Wnt signaling induces myofibroblast differentiation of mesenchymal stem cells, contributing to pulmonary fibrosis.

    PubMed

    Sun, Zhaorui; Wang, Cong; Shi, Chaowen; Sun, Fangfang; Xu, Xiaomeng; Qian, Weiping; Nie, Shinan; Han, Xiaodong

    2014-05-01

    Acute lung injury may lead to fibrogenesis. However, no treatment is currently available. This study was conducted to determine the effects of bone marrow-derived mesenchymal stem cells (MSCs) in a model of HCl-induced acute lung injury in Sprague-Dawley (SD) rats. Stromal cell-derived factor (SDF)-1 and its receptor CXC chemokine receptor (CXCR)4 have been shown to participate in mobilizing MSCs. Adenovirus carrying the CXCR4 gene was used to transfect MSCs in order to increase the engraftment numbers of MSCs at injured sites. Histological examination data demonstrated that the engraftment of MSCs did not attenuate lung injury and pulmonary fibrosis. The results showed that engraftment of MSCs almost differentiated into myofibroblasts, but rarely differentiated into lung epithelial cells. Additionally, it was demonstrated that activated canonical Wnt/β-catenin signaling in injured lung tissue regulated the myofibroblast differentiation of MSCs in vivo. The in vitro study results demonstrated that activation of the Wnt/β-catenin signaling stimulated MSCs to express myofibroblast markers; however, this process was attenuated by Wnt antagonist DKK1. Therefore, the results demonstrated that the aberrant activation of Wnt signaling induces the myofibroblast differentiation of engrafted MSCs, thus contributing to pulmonary fibrosis following lung injury. PMID:24573542

  19. Effect of cell density on adipogenic differentiation of mesenchymal stem cells

    SciTech Connect

    Lu, Hongxu; Biomaterials Center, National Institute for Materials Science, 1-1 Namiki, Tsukuba, Ibaraki 305-0044 ; Guo, Likun; National Engineering Research Center for Biomaterials, Sichuan University, 29 Wangjiang Road, Chengdu 610064 ; Wozniak, Michal J.; Kawazoe, Naoki; International Center for Materials Nanoarchitectonics , National Institute for Materials Science, 1-1 Namiki, Tsukuba, Ibaraki 305-0044 ; Tateishi, Tetsuya; Zhang, Xingdong; Chen, Guoping; Biomaterials Center, National Institute for Materials Science, 1-1 Namiki, Tsukuba, Ibaraki 305-0044; International Center for Materials Nanoarchitectonics , National Institute for Materials Science, 1-1 Namiki, Tsukuba, Ibaraki 305-0044

    2009-04-10

    The effect of cell density on the adipogenic differentiation of human bone marrow-derived mesenchymal stem cells (MSCs) was investigated by using a patterning technique to induce the formation of a cell density gradient on a micropatterned surface. The adipogenic differentiation of MSCs at a density gradient from 5 x 10{sup 3} to 3 x 10{sup 4} cells/cm{sup 2} was examined. Lipid vacuoles were observed at all cell densities after 1-3 weeks of culture in adipogenic differentiation medium although the lipid vacuoles were scarce at the low cell density and abundant at the high cell density. Real-time RT-PCR analysis showed that adipogenesis marker genes encoding peroxisome proliferator-activated receptor {gamma}2 (PPAR{gamma}2), lipoprotein lipase (LPL), and fatty acid binding protein-4 (FABP4) were detected in the MSCs cultured at all cell densities. The results suggest that there was no apparent effect of cell density on the adipogenic differentiation of human MSCs.

  20. Temporal Analyses of the Response of Intervertebral Disc Cells and Mesenchymal Stem Cells to Nutrient Deprivation.

    PubMed

    Turner, Sarah A; Wright, Karina T; Jones, Philip N; Balain, Birender; Roberts, Sally

    2016-01-01

    Much emphasis has been placed recently on the repair of degenerate discs using implanted cells, such as disc cells or bone marrow derived mesenchymal stem cells (MSCs). This study examines the temporal response of bovine and human nucleus pulposus (NP) cells and MSCs cultured in monolayer following exposure to altered levels of glucose (0, 3.15, and 4.5 g/L) and foetal bovine serum (0, 10, and 20%) using an automated time-lapse imaging system. NP cells were also exposed to the cell death inducers, hydrogen peroxide and staurosporine, in comparison to serum starvation. We have demonstrated that human NP cells show an initial "shock" response to reduced nutrition (glucose). However, as time progresses, NP cells supplemented with serum recover with minimal evidence of cell death. Human NP cells show no evidence of proliferation in response to nutrient supplementation, whereas MSCs showed greater response to increased nutrition. When specifically inducing NP cell death with hydrogen peroxide and staurosporine, as expected, the cell number declined. These results support the concept that implanted NP cells or MSCs may be capable of survival in the nutrient-poor environment of the degenerate human disc, which has important clinical implications for the development of IVD cell therapies. PMID:26977156

  1. Mesenchymal Stem Cells Enhance Nerve Regeneration in a Rat Sciatic Nerve Repair and Hindlimb Transplant Model

    PubMed Central

    Cooney, Damon S.; Wimmers, Eric G.; Ibrahim, Zuhaib; Grahammer, Johanna; Christensen, Joani M.; Brat, Gabriel A.; Wu, Lehao W.; Sarhane, Karim A.; Lopez, Joseph; Wallner, Christoph; Furtmüller, Georg J.; Yuan, Nance; Pang, John; Sarkar, Kakali; Lee, W. P. Andrew; Brandacher, Gerald

    2016-01-01

    This study investigates the efficacy of local and intravenous mesenchymal stem cell (MSC) administration to augment neuroregeneration in both a sciatic nerve cut-and-repair and rat hindlimb transplant model. Bone marrow-derived MSCs were harvested and purified from Brown-Norway (BN) rats. Sciatic nerve transections and repairs were performed in three groups of Lewis (LEW) rats: negative controls (n = 4), local MSCs (epineural) injection (n = 4), and systemic MSCs (intravenous) injection (n = 4). Syngeneic (LEW-LEW) (n = 4) and allogeneic (BN-LEW) (n = 4) hindlimb transplants were performed and assessed for neuroregeneration after local or systemic MSC treatment. Rats undergoing sciatic nerve cut-and-repair and treated with either local or systemic injection of MSCs had significant improvement in the speed of recovery of compound muscle action potential amplitudes and axon counts when compared with negative controls. Similarly, rats undergoing allogeneic hindlimb transplants treated with local injection of MSCs exhibited significantly increased axon counts. Similarly, systemic MSC treatment resulted in improved nerve regeneration following allogeneic hindlimb transplants. Systemic administration had a more pronounced effect on electromotor recovery while local injection was more effective at increasing fiber counts, suggesting different targets of action. Local and systemic MSC injections significantly improve the pace and degree of nerve regeneration after nerve injury and hindlimb transplantation. PMID:27510321

  2. Differential Mechanical Response of Mesenchymal Stem Cells and Fibroblasts to Tumor-Secreted Soluble Factors

    PubMed Central

    McGrail, Daniel J.; Ghosh, Deepraj; Quach, Nhat D.; Dawson, Michelle R.

    2012-01-01

    The progression of neoplastic malignancies is a complex process resulting not only from the accumulation of mutations within tumor cells, but also modulation of the tumor microenvironment. Recent advances have shown that the recruitment and subsequent heterotypic interactions of stromal cells—including fibroblasts and bone marrow-derived mesenchymal stem cells (MSCs)—are crucial for carcinogenesis. Though extensive work has been done analyzing the signals that recruit these cells, the governing mechanical properties have not been fully investigated. Here, we report that despite their initial similarities, MSCs respond not only faster but also more dramatically to pro-migratory tumor-secreted soluble factors. Utilizing multiple particle tracking microrheology to probe the cytoskeletal mechanical properties, we show that MSCs stiffen completely within one hour, three times faster than fibroblasts. In addition, unlike fibroblasts, MSCs exposed to tumor-secreted soluble factors display a functionally different phenotype characterized by morphological elongation, decreased actin stress fiber density, and decreased adhesion. Quantitative real-time PCR indicates these phenomena occur based on differential expression of small GTPases RhoA and Cdc42, but not Rac1. These findings demonstrate a fundamental difference in the recruitment of fibroblasts and MSCs. PMID:22438903

  3. Thiosemicarbazone-Pt(II) Complex Causes a Growth Inhibitory Effect on Human Mesenchymal Stem Cells.

    PubMed

    Garcia-Ruiz, Josefa Predestinacion; Matesanz Garcia, Ana Isabel; Souza, Ana Perez; Castelo, Pilar Souza

    2015-01-01

    We showed di[3,5-diacetyl-1,2,4-triazolbis(4-cyclohexylthiosemicarbazonato) platinum(II)] complex, (W8), endowed with important antitumor properties. Here, we analysed whether W8 can affect human bone marrow-derived Mesenchymal Stem Cells, (hMSCs), involved in tissue repair, immunomodulatory properties and also capacity for homing to injure-tumor sites in ovarian cancer. Specifically, we analysed the effect of W8 on cell proliferation, response to scratch, and whether copper-derived cellular mechanism is used by this platinum(II) complex being studied. Results showed that W8 causes a significant inhibition of cell proliferation at µM concentration. This effect is directly related to the alteration of cytoskeletal proteins and inhibition of the response to scratch induced by the presence of foetal bovine serum. This strongly supports the notion of W8 triggers the energetic metabolism of hMSCs and adds an extra support by the results showing W8 relationship with the cellular copper ions. W8, acting in hMSCs, regulates in addition the inhibition of cell proliferation, the inhibition of tumor angiogenesis and metastasis. PMID:25974080

  4. The Osteogenic Priming of Mesenchymal Stem Cells is Impaired in Experimental Diabetes.

    PubMed

    Silva, J C; Sampaio, P; Fernandes, M H; Gomes, P S

    2015-08-01

    Diabetes mellitus encompasses a group of metabolic conditions embracing the dysfunction and failure of various tissues and organs, including bone. Sustained bone alterations seem to result from anabolic, rather than catabolic processes, and suggest a decreased osteoblastic recruitment and activity. Current knowledge on the cellular and molecular mechanisms were provided by studies performed with osteogenic populations cultured in diabetic-simulated conditions, and osteogenic-induced precursor populations harvested from diabetic animals, sustaining an impaired cellular behavior in terms of osteogenic responsiveness and function. However, the reasons leaning to this impairment remain essentially unknown, as the priming capability and functionality of undifferentiated precursors, developed within the diabetic environment, have not been addressed. Accordingly, this work aims to evaluate the functionality and osteogenic priming capability of bone marrow-derived mesenchymal stem cells (MSCs), harvested from animals with experimental diabetes, and grown in the absence of any given differentiation factor. MSCs developed within a diabetic microenvironment displayed an impaired behavior, with diminished cell viability and proliferation, altered cytoskeleton organization, impaired osteogenic priming, and increased adipogenic activation. Further, the osteogenic induction of diabetic MSCs resulted in an impaired osteogenic commitment. The modified cell phenotype may be related, at least in part, with altered activity of ERK WNT and p38 signaling pathways in diabetic-derived cultures. Specific strategies, aiming the modulation of the verified hindrances, may be of therapeutic value to enhance the functionality of diabetic MSCs and sustain an improved outcome in the metabolism and regeneration of the bone tissue in diabetic conditions. PMID:25704854

  5. Differentiation of bone mesenchymal stem cells into hepatocyte-like cells induced by liver tissue homogenate.

    PubMed

    Xing, X K; Feng, H G; Yuan, Z Q

    2016-01-01

    This study investigated the efficacy and feasibility of inducing the differentiation of bone marrow-derived mesenchymal stem cells (BMSCs) into hepatocyte-like cells in vitro using Sprague Dawley rats, as a model of hepatocyte generation for cell transplantation. BMSCs were isolated and grown using the adherent method and exposed to 5 or 10% liver tissue homogenate, before being collected for analysis after 0, 7, 14, and 21 days. Immunofluorescence and western blotting were employed to detect the liver-specific markers a-fetoprotein (AFP) and albumin (ALB). Supernatant urea content was also measured to verify that differentiation had been induced. After 7 days in the presence of 10% liver tissue homogenate, BMSCs demonstrated hepatocyte-like morphological characteristics, and with prolonged culture time, liver-specific markers were gradually produced at levels indicating cell maturation. AFP expression peaked at 14 days then began to decrease, while both urea and ALB levels increased with induction time. Overall, marker expression in the 5% homogenate group was less than or equal to the 10% group at each time point. Thus, in a rat model, liver tissue homogenate obtained from partial hepatectomy can induce the differentiation of BMSCs into hepatocyte-like cells. This method is simple, feasible, and has remarkable real-world application potential. PMID:27525848

  6. Concise review: optimizing expansion of bone marrow mesenchymal stem/stromal cells for clinical applications.

    PubMed

    Hoch, Allison I; Leach, J Kent

    2014-05-01

    Bone marrow-derived mesenchymal stem/stromal cells (MSCs) have demonstrated success in the clinical treatment of hematopoietic pathologies and cardiovascular disease and are the focus of treating other diseases of the musculoskeletal, digestive, integumentary, and nervous systems. However, during the requisite two-dimensional (2D) expansion to achieve a clinically relevant number of cells, MSCs exhibit profound degeneration in progenitor potency. Proliferation, multilineage potential, and colony-forming efficiency are fundamental progenitor properties that are abrogated by extensive monolayer culture. To harness the robust therapeutic potential of MSCs, a consistent, rapid, and minimally detrimental expansion method is necessary. Alternative expansion efforts have exhibited promise in the ability to preserve MSC progenitor potency better than the 2D paradigm by mimicking features of the native bone marrow niche. MSCs have been successfully expanded when stimulated by growth factors, under reduced oxygen tension, and in three-dimensional bioreactors. MSC therapeutic value can be optimized for clinical applications by combining system inputs to tailor culture parameters for recapitulating the niche with probes that nondestructively monitor progenitor potency. The purpose of this review is to explore how modulations in the 2D paradigm affect MSC progenitor properties and to highlight recent efforts in alternative expansion techniques. PMID:24682286

  7. Mesenchymal Stem Cells Enhance Nerve Regeneration in a Rat Sciatic Nerve Repair and Hindlimb Transplant Model.

    PubMed

    Cooney, Damon S; Wimmers, Eric G; Ibrahim, Zuhaib; Grahammer, Johanna; Christensen, Joani M; Brat, Gabriel A; Wu, Lehao W; Sarhane, Karim A; Lopez, Joseph; Wallner, Christoph; Furtmüller, Georg J; Yuan, Nance; Pang, John; Sarkar, Kakali; Lee, W P Andrew; Brandacher, Gerald

    2016-01-01

    This study investigates the efficacy of local and intravenous mesenchymal stem cell (MSC) administration to augment neuroregeneration in both a sciatic nerve cut-and-repair and rat hindlimb transplant model. Bone marrow-derived MSCs were harvested and purified from Brown-Norway (BN) rats. Sciatic nerve transections and repairs were performed in three groups of Lewis (LEW) rats: negative controls (n = 4), local MSCs (epineural) injection (n = 4), and systemic MSCs (intravenous) injection (n = 4). Syngeneic (LEW-LEW) (n = 4) and allogeneic (BN-LEW) (n = 4) hindlimb transplants were performed and assessed for neuroregeneration after local or systemic MSC treatment. Rats undergoing sciatic nerve cut-and-repair and treated with either local or systemic injection of MSCs had significant improvement in the speed of recovery of compound muscle action potential amplitudes and axon counts when compared with negative controls. Similarly, rats undergoing allogeneic hindlimb transplants treated with local injection of MSCs exhibited significantly increased axon counts. Similarly, systemic MSC treatment resulted in improved nerve regeneration following allogeneic hindlimb transplants. Systemic administration had a more pronounced effect on electromotor recovery while local injection was more effective at increasing fiber counts, suggesting different targets of action. Local and systemic MSC injections significantly improve the pace and degree of nerve regeneration after nerve injury and hindlimb transplantation. PMID:27510321

  8. The Effects of Naproxen on Chondrogenesis of Human Mesenchymal Stem Cells.

    PubMed

    Antoniou, John; Wang, Hong Tian; Hadjab, Insaf; Aldebeyan, Sultan; Alaqeel, Motaz A; Meij, Björn P; Tryfonidou, Marianna A; Mwale, Fackson

    2015-07-01

    Currently, there are no established treatments to prevent, stop, or even retard the degeneration of articular cartilage in osteoarthritis (OA). Biological repair of the degenerating articular cartilage would be preferable to surgery. There is no benign site where autologous chondrocytes can be harvested and used as a cell source for cartilage repair, leaving mesenchymal stem cells (MSCs) as an attractive option. However, MSCs from OA patients have been shown to constitutively express collagen type X (COL-X), a marker of late-stage chondrocyte hypertrophy. We recently found that naproxen (Npx), but not other nonsteroidal anti-inflammatory drugs, can induce collagen type X alpha 1 (COL10A1) gene expression in bone marrow-derived MSCs from healthy and OA donors. In this study, we determined the effect of Npx on COL10A1 expression and investigated the intracellular signaling pathways that mediate such effect in normal human MSCs during chondrogenesis. MSCs were cultured in standard chondrogenic differentiation media supplemented with or without Npx. Our results show that Npx can regulate chondrogenic differentiation by affecting the gene expression of both Indian hedgehog and parathyroid hormone/parathyroid hormone-related protein signaling pathways in a time-dependent manner, suggesting a complex interaction of different signaling pathways during the process. PMID:25873236

  9. Temporal Analyses of the Response of Intervertebral Disc Cells and Mesenchymal Stem Cells to Nutrient Deprivation

    PubMed Central

    Turner, Sarah A.; Wright, Karina T.; Jones, Philip N.; Balain, Birender; Roberts, Sally

    2016-01-01

    Much emphasis has been placed recently on the repair of degenerate discs using implanted cells, such as disc cells or bone marrow derived mesenchymal stem cells (MSCs). This study examines the temporal response of bovine and human nucleus pulposus (NP) cells and MSCs cultured in monolayer following exposure to altered levels of glucose (0, 3.15, and 4.5 g/L) and foetal bovine serum (0, 10, and 20%) using an automated time-lapse imaging system. NP cells were also exposed to the cell death inducers, hydrogen peroxide and staurosporine, in comparison to serum starvation. We have demonstrated that human NP cells show an initial “shock” response to reduced nutrition (glucose). However, as time progresses, NP cells supplemented with serum recover with minimal evidence of cell death. Human NP cells show no evidence of proliferation in response to nutrient supplementation, whereas MSCs showed greater response to increased nutrition. When specifically inducing NP cell death with hydrogen peroxide and staurosporine, as expected, the cell number declined. These results support the concept that implanted NP cells or MSCs may be capable of survival in the nutrient-poor environment of the degenerate human disc, which has important clinical implications for the development of IVD cell therapies. PMID:26977156

  10. The Mutual Interactions between Mesenchymal Stem Cells and Myoblasts in an Autologous Co-Culture Model

    PubMed Central

    Szczepanska, Izabela; Zarychta-Wisniewska, Weronika; Pajak, Beata; Bojarczuk, Kamil; Dybowski, Bartosz; Paczek, Leszek

    2016-01-01

    Both myoblasts and mesenchymal stem cells (MSC) take part in the muscle tissue regeneration and have been used as experimental cellular therapy in muscular disorders treatment. It is possible that co-transplantation approach could improve the efficacy of this treatment. However, the relations between those two cell types are not clearly defined. The aim of this study was to determine the reciprocal interactions between myoblasts and MSC in vitro in terms of the features important for the muscle regeneration process. Primary caprine muscle-derived cells (MDC) and bone marrow-derived MSC were analysed in autologous settings. We found that MSC contribute to myotubes formation by fusion with MDC when co-cultured directly, but do not acquire myogenic phenotype if exposed to MDC-derived soluble factors only. Experiments with exposure to hydrogen peroxide showed that MSC are significantly more resistant to oxidative stress than MDC, but a direct co-culture with MSC does not diminish the cytotoxic effect of H2O2 on MDC. Cell migration assay demonstrated that MSC possess significantly greater migration ability than MDC which is further enhanced by MDC-derived soluble factors, whereas the opposite effect was not found. MSC-derived soluble factors significantly enhanced the proliferation of MDC, whereas MDC inhibited the division rate of MSC. To conclude, presented results suggest that myogenic precursors and MSC support each other during muscle regeneration and therefore myoblasts-MSC co-transplantation could be an attractive approach in the treatment of muscular disorders. PMID:27551730

  11. Adult human mesenchymal stem cells enhance breast tumorigenesis and promote hormone independence

    PubMed Central

    Rhodes, Lyndsay V.; Muir, Shannon E.; Elliott, Steven; Guillot, Lori M.; Antoon, James W.; Penfornis, Patrice; Tilghman, Syreeta L.; Salvo, Virgilio A.; Fonseca, Juan P.; Lacey, Michelle R.; Beckman, Barbara S.; McLachlan, John A.; Rowan, Brian G.; Pochampally, Radhika

    2016-01-01

    Adult human mesenchymal stem cells (hMSCs) have been shown to home to sites of breast cancer and integrate into the tumor stroma. We demonstrate here the effect of hMSCs on primary breast tumor growth and the progression of these tumors to hormone independence. Co-injection of bone marrow-derived hMSCs enhances primary tumor growth of the estrogen receptor-positive, hormone-dependent breast carcinoma cell line MCF-7 in the presence or absence of estrogen in SCID/beige mice. We also show hormone-independent growth of MCF-7 cells when co-injected with hMSCs. These effects were found in conjunction with increased immunohistochemical staining of the progesterone receptor in the MCF-7/hMSC tumors as compared to MCF-7 control tumors. This increase in PgR expression indicates a link between MCF-7 cells and MSCs through ER-mediated signaling. Taken together, our data reveal the relationship between tumor microenvironment and tumor growth and the progression to hormone independence. This tumor stroma-cell interaction may provide a novel target for the treatment of estrogen receptor-positive, hormone-independent, and endocrine-resistant breast carcinoma. PMID:19597705

  12. Differential mechanical response of mesenchymal stem cells and fibroblasts to tumor-secreted soluble factors.

    PubMed

    McGrail, Daniel J; Ghosh, Deepraj; Quach, Nhat D; Dawson, Michelle R

    2012-01-01

    The progression of neoplastic malignancies is a complex process resulting not only from the accumulation of mutations within tumor cells, but also modulation of the tumor microenvironment. Recent advances have shown that the recruitment and subsequent heterotypic interactions of stromal cells--including fibroblasts and bone marrow-derived mesenchymal stem cells (MSCs)--are crucial for carcinogenesis. Though extensive work has been done analyzing the signals that recruit these cells, the governing mechanical properties have not been fully investigated. Here, we report that despite their initial similarities, MSCs respond not only faster but also more dramatically to pro-migratory tumor-secreted soluble factors. Utilizing multiple particle tracking microrheology to probe the cytoskeletal mechanical properties, we show that MSCs stiffen completely within one hour, three times faster than fibroblasts. In addition, unlike fibroblasts, MSCs exposed to tumor-secreted soluble factors display a functionally different phenotype characterized by morphological elongation, decreased actin stress fiber density, and decreased adhesion. Quantitative real-time PCR indicates these phenomena occur based on differential expression of small GTPases RhoA and Cdc42, but not Rac1. These findings demonstrate a fundamental difference in the recruitment of fibroblasts and MSCs. PMID:22438903

  13. Mesenchymal stem cells attenuate inflammatory processes in the heart and lung via inhibition of TNF signaling.

    PubMed

    Martire, Alessandra; Bedada, Fikru B; Uchida, Shizuka; Pöling, Jochen; Krüger, Marcus; Warnecke, Henning; Richter, Manfred; Kubin, Thomas; Herold, Susanne; Braun, Thomas

    2016-09-01

    Mesenchymal stem cells (MSC) have been used to treat different clinical conditions although the mechanisms by which pathogenetic processes are affected are still poorly understood. We have previously analyzed the homing of bone marrow-derived MSC to diseased tissues characterized by a high degree of mononuclear cell infiltration and postulated that MSC might modulate inflammatory responses. Here, we demonstrate that MSC mitigate adverse tissue remodeling, improve organ function, and extend lifespan in a mouse model of inflammatory dilative cardiomyopathy (DCM). Furthermore, MSC attenuate Lipopolysaccharide-induced acute lung injury indicating a general role in the suppression of inflammatory processes. We found that MSC released sTNF-RI, which suppressed activation of the NFκBp65 pathway in cardiomyocytes during DCM in vivo. Substitution of MSC by recombinant soluble TNF-R partially recapitulated the beneficial effects of MSC while knockdown of TNF-R prevented MSC-mediated suppression of the NFκBp65 pathway and improvement of tissue pathology. We conclude that sTNF-RI is a major part of the paracrine machinery by which MSC effect local inflammatory reactions. PMID:27435289