Neuromuscular junction formation between human stem cell-derived motoneurons and human skeletal muscle in a defined system.

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Guo, Xiufang; Gonzalez, Mercedes; Stancescu, Maria; Vandenburgh, Herman H; Hickman, James J

2011-12-01

Functional in vitro models composed of human cells will constitute an important platform in the next generation of system biology and drug discovery. This study reports a novel human-based in vitro Neuromuscular Junction (NMJ) system developed in a defined serum-free medium and on a patternable non-biological surface. The motoneurons and skeletal muscles were derived from fetal spinal stem cells and skeletal muscle stem cells. The motoneurons and skeletal myotubes were completely differentiated in the co-culture based on morphological analysis and electrophysiology. NMJ formation was demonstrated by phase contrast microscopy, immunocytochemistry and the observation of motoneuron-induced muscle contractions utilizing time-lapse recordings and their subsequent quenching by d-Tubocurarine. Generally, functional human based systems would eliminate the issue of species variability during the drug development process and its derivation from stem cells bypasses the restrictions inherent with utilization of primary human tissue. This defined human-based NMJ system is one of the first steps in creating functional in vitro systems and will play an important role in understanding NMJ development, in developing high information content drug screens and as test beds in preclinical studies for spinal or muscular diseases/injuries such as muscular dystrophy, Amyotrophic lateral sclerosis and spinal cord repair. Copyright © 2011 Elsevier Ltd. All rights reserved.

Human induced pluripotent stem cell-derived vascular smooth muscle cells

DEFF Research Database (Denmark)

Ayoubi, Sohrab; Sheikh, Søren P; Eskildsen, Tilde V

2017-01-01

. To this end, human induced pluripotent stem cells (hiPSCs) have generated great enthusiasm, and have been a driving force for development of novel strategies in drug discovery and regenerative cell-therapy for the last decade. Hence, investigating the mechanisms underlying the differentiation of hi......PSCs into specialized cell types such as cardiomyocytes, endothelial cells, and vascular smooth muscle cells (VSMCs) may lead to a better understanding of developmental cardiovascular processes and potentiate progress of safe autologous regenerative therapies in pathological conditions. In this review, we summarize...

BMP9-Induced Osteogenetic Differentiation and Bone Formation of Muscle-Derived Stem Cells

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Li Xiang

2012-01-01

Full Text Available Efficient osteogenetic differentiation and bone formation from muscle-derived stem cells (MDSCs should have potential clinical applications in treating nonunion fracture healing or bone defects. Here, we investigate osteogenetic differentiation ability of MDSCs induced by bone morphogenetic protein 9 (BMP9 in vitro and bone formation ability in rabbit radius defects repairing model. Rabbit's MDSCs were extracted by type I collagenase and trypsin methods, and BMP9 was introduced into MDSCs by infection with recombinant adenovirus. Effects of BMP9-induced osteogenetic differentiation of MDSCs were identified with alkaline phosphatase (ALP activity and expression of later marker. In stem-cell implantation assay, MDSCs have also shown valuable potential bone formation ability induced by BMP9 in rabbit radius defects repairing test. Taken together, our findings suggest that MDSCs are potentiated osteogenetic stem cells which can be induced by BMP9 to treat large segmental bone defects, nonunion fracture, and/or osteoporotic fracture.

Aging, metabolism and stem cells: Spotlight on muscle stem cells.

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García-Prat, Laura; Muñoz-Cánoves, Pura

2017-04-15

All tissues and organs undergo a progressive regenerative decline as they age. This decline has been mainly attributed to loss of stem cell number and/or function, and both stem cell-intrinsic changes and alterations in local niches and/or systemic environment over time are known to contribute to the stem cell aging phenotype. Advancing in the molecular understanding of the deterioration of stem cell cells with aging is key for targeting the specific causes of tissue regenerative dysfunction at advanced stages of life. Here, we revise exciting recent findings on why stem cells age and the consequences on tissue regeneration, with a special focus on regeneration of skeletal muscle. We also highlight newly identified common molecular pathways affecting diverse types of aging stem cells, such as altered proteostasis, metabolism, or senescence entry, and discuss the questions raised by these findings. Finally, we comment on emerging stem cell rejuvenation strategies, principally emanating from studies on muscle stem cells, which will surely burst tissue regeneration research for future benefit of the increasing human aging population. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

Reconstruction of radical prostatectomy-induced urethral damage using skeletal muscle-derived multipotent stem cells.

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Hoshi, Akio; Tamaki, Tetsuro; Tono, Kayoko; Okada, Yoshinori; Akatsuka, Akira; Usui, Yukio; Terachi, Toshiro

2008-06-15

Postoperative damage of the urethral rhabdosphincter (URS) and neurovascular bundle (NVB) is a major operative complication of radical prostatectomy. It is generally recognized to be caused by unavoidable surgical damage to the muscle-nerve-blood vessel units around the urethra. We attempted to treat this damage using skeletal muscle-derived stem cells, which are able to reconstitute muscle-nerve-blood vessel units. Cells were enzymatically extracted and sorted by flow cytometry as CD34/45 (Sk-34) and CD34/45 (Sk-DN) cells from green fluorescent protein transgenic mice and rats. URS-NVB damage was induced by manually removing one-third of the total URS and unilateral invasion of NVB in wild-type Sprague-Dawley and node rats. Freshly isolated Sk-34, Sk-34+Sk-DN cells, and cultured Sk-DN cells were directly transplanted into the damaged portion. At 4 and 12 weeks after transplantation, urethral pressure profile by electrical stimulation through the sacral surface (L6-S1) was evaluated as functional recovery. The recovery ratio in the control and transplanted groups was 37.6% and 72.9%, at 4 weeks, and 41.6% and 78.4% at 12 weeks, respectively (Pcells differentiated into numerous skeletal muscle fibers having neuromuscular junctions (innervation) and nerve bundle-related Schwann cells and perineurium, and blood vessel-related endothelial cells and pericyte around the urethra. Thus, we conclude that transplantation of skeletal muscle-derived multipotent Sk-34 and Sk-DN cells is potentially useful for the reconstitution of postoperative damage of URS and NVB after radical prostatectomy.

Fetal stem cells and skeletal muscle regeneration: a therapeutic approach

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Michela ePozzobon

2014-08-01

Full Text Available More than 40% of the body mass is represented by muscle tissue, which possesses the innate ability to regenerate after damage through the activation of muscle specific stem cell, namely satellite cells. Muscle diseases, in particular chronic degenerative state of skeletal muscle such as dystrophies, lead to a perturbation of the regenerative process, which causes the premature exhaustion of satellite cell reservoir due to continue cycles of degeneration/regeneration. Nowadays, the research is focused on different therapeutic approaches, ranging from gene and cell to pharmacological therapy, but still there is not a definitive cure in particular for genetic muscle disease. Taking this in mind, in this article we will give special consideration to muscle diseases and the use of fetal derived stem cells as new approach for therapy. Cells of fetal origin, from cord blood to placenta and amniotic fluid, can be easily obtained without ethical concern, expanded and differentiated in culture, and possess immunemodulatory properties. The in vivo approach in animal models can be helpful to study the mechanism underneath the operating principle of the stem cell reservoir, namely the niche, which holds great potential to understand the onset of muscle pathologies.

Structural phenotyping of stem cell-derived cardiomyocytes.

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Pasqualini, Francesco Silvio; Sheehy, Sean Paul; Agarwal, Ashutosh; Aratyn-Schaus, Yvonne; Parker, Kevin Kit

2015-03-10

Structural phenotyping based on classical image feature detection has been adopted to elucidate the molecular mechanisms behind genetically or pharmacologically induced changes in cell morphology. Here, we developed a set of 11 metrics to capture the increasing sarcomere organization that occurs intracellularly during striated muscle cell development. To test our metrics, we analyzed the localization of the contractile protein α-actinin in a variety of primary and stem-cell derived cardiomyocytes. Further, we combined these metrics with data mining algorithms to unbiasedly score the phenotypic maturity of human-induced pluripotent stem cell-derived cardiomyocytes. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.

Satellite Cells and the Muscle Stem Cell Niche

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Yin, Hang; Price, Feodor

2013-01-01

Adult skeletal muscle in mammals is a stable tissue under normal circumstances but has remarkable ability to repair after injury. Skeletal muscle regeneration is a highly orchestrated process involving the activation of various cellular and molecular responses. As skeletal muscle stem cells, satellite cells play an indispensible role in this process. The self-renewing proliferation of satellite cells not only maintains the stem cell population but also provides numerous myogenic cells, which proliferate, differentiate, fuse, and lead to new myofiber formation and reconstitution of a functional contractile apparatus. The complex behavior of satellite cells during skeletal muscle regeneration is tightly regulated through the dynamic interplay between intrinsic factors within satellite cells and extrinsic factors constituting the muscle stem cell niche/microenvironment. For the last half century, the advance of molecular biology, cell biology, and genetics has greatly improved our understanding of skeletal muscle biology. Here, we review some recent advances, with focuses on functions of satellite cells and their niche during the process of skeletal muscle regeneration. PMID:23303905

Vascular Smooth Muscle Cells From Hypertensive Patient-Derived Induced Pluripotent Stem Cells to Advance Hypertension Pharmacogenomics.

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Biel, Nikolett M; Santostefano, Katherine E; DiVita, Bayli B; El Rouby, Nihal; Carrasquilla, Santiago D; Simmons, Chelsey; Nakanishi, Mahito; Cooper-DeHoff, Rhonda M; Johnson, Julie A; Terada, Naohiro

2015-12-01

Studies in hypertension (HTN) pharmacogenomics seek to identify genetic sources of variable antihypertensive drug response. Genetic association studies have detected single-nucleotide polymorphisms (SNPs) that link to drug responses; however, to understand mechanisms underlying how genetic traits alter drug responses, a biological interface is needed. Patient-derived induced pluripotent stem cells (iPSCs) provide a potential source for studying otherwise inaccessible tissues that may be important to antihypertensive drug response. The present study established multiple iPSC lines from an HTN pharmacogenomics cohort. We demonstrated that established HTN iPSCs can robustly and reproducibly differentiate into functional vascular smooth muscle cells (VSMCs), a cell type most relevant to vasculature tone control. Moreover, a sensitive traction force microscopy assay demonstrated that iPSC-derived VSMCs show a quantitative contractile response on physiological stimulus of endothelin-1. Furthermore, the inflammatory chemokine tumor necrosis factor α induced a typical VSMC response in iPSC-derived VSMCs. These studies pave the way for a large research initiative to decode biological significance of identified SNPs in hypertension pharmacogenomics. Treatment of hypertension remains suboptimal, and a pharmacogenomics approach seeks to identify genetic biomarkers that could be used to guide treatment decisions; however, it is important to understand the biological underpinnings of genetic associations. Mouse models do not accurately recapitulate individual patient responses based on their genetics, and hypertension-relevant cells are difficult to obtain from patients. Induced pluripotent stem cell (iPSC) technology provides a great interface to bring patient cells with their genomic data into the laboratory and to study hypertensive responses. As an initial step, the present study established an iPSC bank from patients with primary hypertension and demonstrated an effective

CD133+ cells derived from skeletal muscles of Duchenne muscular dystrophy patients have a compromised myogenic and muscle regenerative capability.

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Meng, Jinhong; Muntoni, Francesco; Morgan, Jennifer

2018-05-12

Cell-mediated gene therapy is a possible means to treat muscular dystrophies like Duchenne muscular dystrophy. Autologous patient stem cells can be genetically-corrected and transplanted back into the patient, without causing immunorejection problems. Regenerated muscle fibres derived from these cells will express the missing dystrophin protein, thus improving muscle function. CD133+ cells derived from normal human skeletal muscle contribute to regenerated muscle fibres and form muscle stem cells after their intra-muscular transplantation into an immunodeficient mouse model. But it is not known whether CD133+ cells derived from DMD patient muscles have compromised muscle regenerative function. To test this, we compared CD133+ cells derived from DMD and normal human muscles. DMD CD133+ cells had a reduced capacity to undergo myogenic differentiation in vitro compared with CD133+ cells derived from normal muscle. In contrast to CD133+ cells derived from normal human muscle, those derived from DMD muscle formed no satellite cells and gave rise to significantly fewer muscle fibres of donor origin, after their intra-muscular transplantation into an immunodeficient, non-dystrophic, mouse muscle. DMD CD133+ cells gave rise to more clones of smaller size and more clones that were less myogenic than did CD133+ cells derived from normal muscle. The heterogeneity of the progeny of CD133+ cells, combined with the reduced proliferation and myogenicity of DMD compared to normal CD133+ cells, may explain the reduced regenerative capacity of DMD CD133+ cells. Copyright © 2018 The Authors. Published by Elsevier B.V. All rights reserved.

Functional neuromuscular junctions formed by embryonic stem cell-derived motor neurons.

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Joy A Umbach

Full Text Available A key objective of stem cell biology is to create physiologically relevant cells suitable for modeling disease pathologies in vitro. Much progress towards this goal has been made in the area of motor neuron (MN disease through the development of methods to direct spinal MN formation from both embryonic and induced pluripotent stem cells. Previous studies have characterized these neurons with respect to their molecular and intrinsic functional properties. However, the synaptic activity of stem cell-derived MNs remains less well defined. In this study, we report the development of low-density co-culture conditions that encourage the formation of active neuromuscular synapses between stem cell-derived MNs and muscle cells in vitro. Fluorescence microscopy reveals the expression of numerous synaptic proteins at these contacts, while dual patch clamp recording detects both spontaneous and multi-quantal evoked synaptic responses similar to those observed in vivo. Together, these findings demonstrate that stem cell-derived MNs innervate muscle cells in a functionally relevant manner. This dual recording approach further offers a sensitive and quantitative assay platform to probe disorders of synaptic dysfunction associated with MN disease.

Generation of skeletal muscle from transplanted embryonic stem cells in dystrophic mice

International Nuclear Information System (INIS)

Bhagavati, Satyakam; Xu Weimin

2005-01-01

Embryonic stem (ES) cells have great therapeutic potential because of their capacity to proliferate extensively and to form any fully differentiated cell of the body, including skeletal muscle cells. Successful generation of skeletal muscle in vivo, however, requires selective induction of the skeletal muscle lineage in cultures of ES cells and following transplantation, integration of appropriately differentiated skeletal muscle cells with recipient muscle. Duchenne muscular dystrophy (DMD), a severe progressive muscle wasting disease due to a mutation in the dystrophin gene and the mdx mouse, an animal model for DMD, are characterized by the absence of the muscle membrane associated protein, dystrophin. Here, we show that co-culturing mouse ES cells with a preparation from mouse muscle enriched for myogenic stem and precursor cells, followed by injection into mdx mice, results occasionally in the formation of normal, vascularized skeletal muscle derived from the transplanted ES cells. Study of this phenomenon should provide valuable insights into skeletal muscle development in vivo from transplanted ES cells

Restoration of heart functions using human embryonic stem cells derived heart muscle cells.

Science.gov (United States)

Gepstein, Lior; Kehat, Izhak

2005-02-01

Extract: Recent advances in molecular and cellular biology and specifically in the areas of stem cell biology and tissue engineering have paved the way for the development of a new field in biomedicine, regenerative medicine. This exciting approach seeks to develop new biological solutions, using the mobilization of endogenous stem cells or delivery of exogenous cells to replace or modify the function of diseased, absent, or malfunctioning tissue. The adult heart represents an attractive candidate for these emerging technologies, since adult cardiomyocytes have limited regenerative capacity. Thus, any significant heart cell loss or dysfunction, such as occurs during heart attack, is mostly irreversible and may lead to the development of progressive heart failure, one of the leading causes of world-wide morbidity and mortality. Similarly, dysfunction of the specialized electrical conduction system within the heart may result in inefficient rhythm initiation or impulse conduction, leading to significant slowing of the heart rate, usually requiring the implantation of a permanent electronic pacemaker. Replacement of the dysfunctional myocardium (heart muscle) by implantation of external heart muscle cells is emerging as a novel paradigm for restoration of the myocardial electromechanical properties, but has been significantly hampered by the paucity of cell sources for human heart cells and by the relatively limited evidence for functional integration between grafted and host cells. The recently described human embryonic stem cell (hESC) lines may provide a possible solution for the aforementioned cell sourcing problem.

Aging induced loss of stemness with concomitant gain of myogenic properties of a pure population of CD34(+)/CD45(-) muscle derived stem cells.

Science.gov (United States)

Bose, Bipasha; Shenoy, P Sudheer

2016-01-01

Aging is accompanied by the functional decline of cells, tissues, and organs, as well as, a striking increase in susceptibility to a wide range of diseases. Within a tissue, both differentiated cells and adult stem cells are susceptible to intrinsic and extrinsic changes while aging. Muscle derived stem cells (MDSCs) are tissue specific stem cells which have been studied well for their multipotential nature. Although there are reports relating to diminished function and regenerative capacity of aged MDSCs as compared to their young counterparts, not much has been reported relating to the concomitant gain in unipotent nature of aged MDSCs. In this study, we report an inverse correlation between aging and expression of adult/mesenchymal stem cell markers and a direct correlation between aging and myogenecity in MDSCs. Aged MDSCs were able to generate a greater number of dystrophin positive myofibres, as compared to, the young MDSCs when transplanted in muscle of dystrophic mice. Our data, therefore, suggests that aging stress adds to the decline in stem cell characteristics with a concomitant increase in unipotency, in terms of, myogenecity of MDSCs. This study, hence, also opens the possibilities of using unipotent aged MDSCs as potential candidates for transplantation in patients with muscular dystrophies. Copyright © 2015. Published by Elsevier Ltd.

Extracellular matrix components direct porcine muscle stem cell behavior

International Nuclear Information System (INIS)

Wilschut, Karlijn J.; Haagsman, Henk P.; Roelen, Bernard A.J.

2010-01-01

In muscle tissue, extracellular matrix proteins, together with the vasculature system, muscle-residence cells and muscle fibers, create the niche for muscle stem cells. The niche is important in controlling proliferation and directing differentiation of muscle stem cells to sustain muscle tissue. Mimicking the extracellular muscle environment improves tools exploring the behavior of primary muscle cells. Optimizing cell culture conditions to maintain muscle commitment is important in stem cell-based studies concerning toxicology screening, ex vivo skeletal muscle tissue engineering and in the enhancement of clinical efficiency. We used the muscle extracellular matrix proteins collagen type I, fibronectin, laminin, and also gelatin and Matrigel as surface coatings of tissue culture plastic to resemble the muscle extracellular matrix. Several important factors that determine myogenic commitment of the primary muscle cells were characterized by quantitative real-time RT-PCR and immunofluorescence. Adhesion of high PAX7 expressing satellite cells was improved if the cells were cultured on fibronectin or laminin coatings. Cells cultured on Matrigel and laminin coatings showed dominant integrin expression levels and exhibited an activated Wnt pathway. Under these conditions both stem cell proliferation and myogenic differentiation capacity were superior if compared to cells cultured on collagen type I, fibronectin and gelatin. In conclusion, Matrigel and laminin are the preferred coatings to sustain the proliferation and myogenic differentiation capacity of the primary porcine muscle stem cells, when cells are removed from their natural environment for in vitro culture.

Extracellular matrix components direct porcine muscle stem cell behavior

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Wilschut, Karlijn J. [Department of Farm Animal Health, Faculty of Veterinary Medicine, Utrecht University, Yalelaan 104, 3584 CM, Utrecht (Netherlands); Haagsman, Henk P. [Department of Infectious Diseases and Immunology, Faculty of Veterinary Medicine, Utrecht University, Yalelaan 1, 3584 CL, Utrecht (Netherlands); Roelen, Bernard A.J., E-mail: b.a.j.roelen@uu.nl [Department of Farm Animal Health, Faculty of Veterinary Medicine, Utrecht University, Yalelaan 104, 3584 CM, Utrecht (Netherlands)

2010-02-01

In muscle tissue, extracellular matrix proteins, together with the vasculature system, muscle-residence cells and muscle fibers, create the niche for muscle stem cells. The niche is important in controlling proliferation and directing differentiation of muscle stem cells to sustain muscle tissue. Mimicking the extracellular muscle environment improves tools exploring the behavior of primary muscle cells. Optimizing cell culture conditions to maintain muscle commitment is important in stem cell-based studies concerning toxicology screening, ex vivo skeletal muscle tissue engineering and in the enhancement of clinical efficiency. We used the muscle extracellular matrix proteins collagen type I, fibronectin, laminin, and also gelatin and Matrigel as surface coatings of tissue culture plastic to resemble the muscle extracellular matrix. Several important factors that determine myogenic commitment of the primary muscle cells were characterized by quantitative real-time RT-PCR and immunofluorescence. Adhesion of high PAX7 expressing satellite cells was improved if the cells were cultured on fibronectin or laminin coatings. Cells cultured on Matrigel and laminin coatings showed dominant integrin expression levels and exhibited an activated Wnt pathway. Under these conditions both stem cell proliferation and myogenic differentiation capacity were superior if compared to cells cultured on collagen type I, fibronectin and gelatin. In conclusion, Matrigel and laminin are the preferred coatings to sustain the proliferation and myogenic differentiation capacity of the primary porcine muscle stem cells, when cells are removed from their natural environment for in vitro culture.

Enhanced elastin synthesis and maturation in human vascular smooth muscle tissue derived from induced-pluripotent stem cells.

Science.gov (United States)

Eoh, Joon H; Shen, Nian; Burke, Jacqueline A; Hinderer, Svenja; Xia, Zhiyong; Schenke-Layland, Katja; Gerecht, Sharon

2017-04-01

Obtaining vascular smooth muscle tissue with mature, functional elastic fibers is a key obstacle in tissue-engineered blood vessels. Poor elastin secretion and organization leads to a loss of specialization in contractile smooth muscle cells, resulting in over proliferation and graft failure. In this study, human induced-pluripotent stem cells (hiPSCs) were differentiated into early smooth muscle cells, seeded onto a hybrid poly(ethylene glycol) dimethacrylate/poly (l-lactide) (PEGdma-PLA) scaffold and cultured in a bioreactor while exposed to pulsatile flow, towards maturation into contractile smooth muscle tissue. We evaluated the effects of pulsatile flow on cellular organization as well as elastin expression and assembly in the engineered tissue compared to a static control through immunohistochemistry, gene expression and functionality assays. We show that culturing under pulsatile flow resulted in organized and functional hiPSC derived smooth muscle tissue. Immunohistochemistry analysis revealed hiPSC-smooth muscle tissue with robust, well-organized cells and elastic fibers and the supporting microfibril proteins necessary for elastic fiber assembly. Through qRT-PCR analysis, we found significantly increased expression of elastin, fibronectin, and collagen I, indicating the synthesis of necessary extracellular matrix components. Functionality assays revealed that hiPSC-smooth muscle tissue cultured in the bioreactor had an increased calcium signaling and contraction in response to a cholinergic agonist, significantly higher mature elastin content and improved mechanical properties in comparison to the static control. The findings presented here detail an effective approach to engineering elastic human vascular smooth muscle tissue with the functionality necessary for tissue engineering and regenerative medicine applications. Obtaining robust, mature elastic fibers is a key obstacle in tissue-engineered blood vessels. Human induced-pluripotent stem cells have

Slow-Adhering Stem Cells Derived from Injured Skeletal Muscle Have Improved Regenerative Capacity

Science.gov (United States)

2011-08-01

stress urinary incontinence . Urology 2006, 68:449–454 15. Chermansky CJ, Tarin T, Kwon DD, Jankowski RJ, Cannon TW, de Groat WC, Huard J, Chancellor...from control noninjured muscle. These data suggest that traumatic injury may modify stem cell characteristics through trophic factors and improve the...alter the microenvironment of resident muscle cells (ie, stimu- lating cell dedifferentiation on various trophic factors )20,21 and result in profound

Stem cell antigen-1 in skeletal muscle function.

Science.gov (United States)

Bernstein, Harold S; Samad, Tahmina; Cholsiripunlert, Sompob; Khalifian, Saami; Gong, Wenhui; Ritner, Carissa; Aurigui, Julian; Ling, Vivian; Wilschut, Karlijn J; Bennett, Stephen; Hoffman, Julien; Oishi, Peter

2013-08-15

Stem cell antigen-1 (Sca-1) is a member of the Ly-6 multigene family encoding highly homologous, glycosyl-phosphatidylinositol-anchored membrane proteins. Sca-1 is expressed on muscle-derived stem cells and myogenic precursors recruited to sites of muscle injury. We previously reported that inhibition of Sca-1 expression stimulated myoblast proliferation in vitro and regulated the tempo of muscle repair in vivo. Despite its function in myoblast expansion during muscle repair, a role for Sca-1 in normal, post-natal muscle has not been thoroughly investigated. We systematically compared Sca-1-/- (KO) and Sca-1+/+ (WT) mice and hindlimb muscles to elucidate the tissue, contractile, and functional effects of Sca-1 in young and aging animals. Comparison of muscle volume, fibrosis, myofiber cross-sectional area, and Pax7+ myoblast number showed little differences between ages or genotypes. Exercise protocols, however, demonstrated decreased stamina in KO versus WT mice, with young KO mice achieving results similar to aging WT animals. In addition, KO mice did not improve with practice, while WT animals demonstrated conditioning over time. Surprisingly, myomechanical analysis of isolated muscles showed that KO young muscle generated more force and experienced less fatigue. However, KO muscle also demonstrated incomplete relaxation with fatigue. These findings suggest that Sca-1 is necessary for muscle conditioning with exercise, and that deficient conditioning in Sca-1 KO animals becomes more pronounced with age.

Characterization of a PLGA sandwiched cell/fibrin tubular construct and induction of the adipose derived stem cells into smooth muscle cells

International Nuclear Information System (INIS)

Wang Xiaohong; Maekitie, Antti A.; Paloheimo, Kaija-Stiina; Tuomi, Jukka; Paloheimo, Markku; Sui Shaochun; Zhang Qiqing

2011-01-01

A poly(DL-lactic-co-glycolic acid) (PLGA) sandwiched adipose derived stem cell (ADSC)/fibrin tubular construct, fabricated using a step-by-step mold/extraction method, was characterized in this work. The ADSCs were also induced into smooth-muscle-like cells using growth factors such as hepatocyte growth factor (HGF), platelet-derived growth factor BB (PDGF-BB), transforming growth factor β1 (TGFβ1), and basic fibroblast growth factor (b-FGF). Compared with the non-induced cells, the proliferation ability of induced cells was much smaller. The PLGA sandwiched cell/hydrogel construct was shown to be useful for controlling the cellular microenvironment and cellular behaviors such as growth, migration, proliferation and differentiation. This strategy seems promising in tissue engineering and organ manufacturing.

Characterization of a PLGA sandwiched cell/fibrin tubular construct and induction of the adipose derived stem cells into smooth muscle cells

Energy Technology Data Exchange (ETDEWEB)

Wang Xiaohong, E-mail: wangxiaohong@tsinghua.edu.cn [BIT Research Centre, School of Science and Technology, Aalto University, P.O. Box 15500, 00076 Aalto (Finland); Key Laboratory for Advanced Materials Processing Technology, Ministry of Education and Center of Organ Manufacturing, Department of Mechanical Engineering, Tsinghua University, Beijing 100084 (China); Maekitie, Antti A.; Paloheimo, Kaija-Stiina; Tuomi, Jukka; Paloheimo, Markku [BIT Research Centre, School of Science and Technology, Aalto University, P.O. Box 15500, 00076 Aalto (Finland); Sui Shaochun [Key Laboratory for Advanced Materials Processing Technology, Ministry of Education and Center of Organ Manufacturing, Department of Mechanical Engineering, Tsinghua University, Beijing 100084 (China); Zhang Qiqing [Institute of Biomedical Engineering, Chinese Academy of Medical Science and Peking Union Medical College, Key Laboratory of Biomedical Material of Tianjin, Tianjin 300192 (China)

2011-05-10

A poly(DL-lactic-co-glycolic acid) (PLGA) sandwiched adipose derived stem cell (ADSC)/fibrin tubular construct, fabricated using a step-by-step mold/extraction method, was characterized in this work. The ADSCs were also induced into smooth-muscle-like cells using growth factors such as hepatocyte growth factor (HGF), platelet-derived growth factor BB (PDGF-BB), transforming growth factor {beta}1 (TGF{beta}1), and basic fibroblast growth factor (b-FGF). Compared with the non-induced cells, the proliferation ability of induced cells was much smaller. The PLGA sandwiched cell/hydrogel construct was shown to be useful for controlling the cellular microenvironment and cellular behaviors such as growth, migration, proliferation and differentiation. This strategy seems promising in tissue engineering and organ manufacturing.

Improved neurological outcome by intramuscular injection of human amniotic fluid derived stem cells in a muscle denervation model.

Directory of Open Access Journals (Sweden)

Chun-Jung Chen

Full Text Available The skeletal muscle develops various degrees of atrophy and metabolic dysfunction following nerve injury. Neurotrophic factors are essential for muscle regeneration. Human amniotic fluid derived stem cells (AFS have the potential to secrete various neurotrophic factors necessary for nerve regeneration. In the present study, we assess the outcome of neurological function by intramuscular injection of AFS in a muscle denervation and nerve anastomosis model.Seventy two Sprague-Dawley rats weighing 200-250 gm were enrolled in this study. Muscle denervation model was conducted by transverse resection of a sciatic nerve with the proximal end sutured into the gluteal muscle. The nerve anastomosis model was performed by transverse resection of the sciatic nerve followed by four stitches reconnection. These animals were allocated to three groups: control, electrical muscle stimulation, and AFS groups.NT-3 (Neurotrophin 3, BDNF (Brain derived neurotrophic factor, CNTF (Ciliary neurotrophic factor, and GDNF (Glia cell line derived neurotrophic factor were highly expressed in AFS cells and supernatant of culture medium. Intra-muscular injection of AFS exerted significant expression of several neurotrophic factors over the distal end of nerve and denervated muscle. AFS caused high expression of Bcl-2 in denervated muscle with a reciprocal decrease of Bad and Bax. AFS preserved the muscle morphology with high expression of desmin and acetylcholine receptors. Up to two months, AFS produced significant improvement in electrophysiological study and neurological functions such as SFI (sciatic nerve function index and Catwalk gait analysis. There was also significant preservation of the number of anterior horn cells and increased nerve myelination as well as muscle morphology.Intramuscular injection of AFS can protect muscle apoptosis and likely does so through the secretion of various neurotrophic factors. This protection furthermore improves the nerve

Adipose-derived stem cells enhance myogenic differentiation in the mdx mouse model of muscular dystrophy via paracrine signaling

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Ji-qing Cao

2016-01-01

Full Text Available Adipose-derived stem cells have been shown to promote peripheral nerve regeneration through the paracrine secretion of neurotrophic factors. However, it is unclear whether these cells can promote myogenic differentiation in muscular dystrophy. Adipose-derived stem cells (6 × 10 6 were injected into the gastrocnemius muscle of mdx mice at various sites. Dystrophin expression was found in the muscle fibers. Phosphorylation levels of Akt, mammalian target of rapamycin (mTOR, eIF-4E binding protein 1 and S6 kinase 1 were increased, and the Akt/mTOR pathway was activated. Simultaneously, myogenin levels were increased, whereas cleaved caspase 3 and vimentin levels were decreased. Necrosis and fibrosis were reduced in the muscle fibers. These findings suggest that adipose-derived stem cells promote the regeneration and survival of muscle cells by inhibiting apoptosis and fibrosis, thereby alleviating muscle damage in muscular dystrophy.

Advancements in stem cells treatment of skeletal muscle wasting

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mirella emeregalli

2014-02-01

Full Text Available Muscular dystrophies (MDs are a heterogeneous group of inherited disorders, in which progressive muscle wasting and weakness is often associated with exhaustion of muscle regeneration potential. Although physiological properties of skeletal muscle tissue are now well known, no treatments are effective for these diseases. Muscle regeneration was attempted by means transplantation of myogenic cells (from myoblast to embryonic stem cells and also by interfering with the malignant processes that originate in pathological tissues, such as uncontrolled fibrosis and inflammation. Taking into account the advances in the isolation of new subpopulation of stem cells and in the creation of artificial stem cell niches, we discuss how these emerging technologies offer great promises for therapeutic approaches to muscle diseases and muscle wasting associated with aging.

Molecular aging and rejuvenation of human muscle stem cells

DEFF Research Database (Denmark)

Carlson, Morgan E; Suetta, Charlotte; Conboy, Michael J

2009-01-01

. Our findings establish key evolutionarily conserved mechanisms of human stem cell aging. We find that satellite cells are maintained in aged human skeletal muscle, but fail to activate in response to muscle attrition, due to diminished activation of Notch compounded by elevated transforming growth...... factor beta (TGF-beta)/phospho Smad3 (pSmad3). Furthermore, this work reveals that mitogen-activated protein kinase (MAPK)/phosphate extracellular signal-regulated kinase (pERK) signalling declines in human muscle with age, and is important for activating Notch in human muscle stem cells. This molecular......Very little remains known about the regulation of human organ stem cells (in general, and during the aging process), and most previous data were collected in short-lived rodents. We examined whether stem cell aging in rodents could be extrapolated to genetically and environmentally variable humans...

Modulation of Stem Cell Differentiation and Myostatin as an Approach to Counteract Fibrosis in Muscle Dystrophy and Regeneration After Injury. Addendum

Science.gov (United States)

2012-03-01

ged mdx mouse a n ovel therapeutic approach for Duchenne’s muscular dystrophy (DMD) based on the implantation of muscle -derived stem cells (MDSC), and...with limb ischemia. 15. SUBJECT TERMS Myostatin, muscle dystrophy , stem cells, myogenesis, Oct-4; Duchenne ; fibrosis 16. SECURITY CLASSIFICATION...derived stem cells (MDSC) into myogenic, as opposed to lipofibrogenic lineages, is a promising therapeutic strategy for Duchenne muscular dystrophy (DMD

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

DEFF Research Database (Denmark)

Rasmussen, Jeppe; Frøbert, Ole; Holst-Hansen, Claus

2014-01-01

Background: Treatment of myocardial infarction 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...... myocardial infarction 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 myocardial infarction, using a fully...... grown non-immunecompromised rat model. Methods: 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...

Purified Human Skeletal Muscle-Derived Stem Cells Enhance the Repair and Regeneration in the Damaged Urethra.

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Nakajima, Nobuyuki; Tamaki, Tetsuro; Hirata, Maki; Soeda, Shuichi; Nitta, Masahiro; Hoshi, Akio; Terachi, Toshiro

2017-10-01

Postoperative damage of the urethral rhabdosphincter and nerve-vascular networks is a major complication of radical prostatectomy and generally causes incontinence and/or erectile dysfunction. The human skeletal muscle-derived stem cells, which have a synchronized reconstitution capacity of muscle-nerve-blood vessel units, were applied to this damage. Cells were enzymatically extracted from the human skeletal muscle, sorted using flow cytometry as CD34/45 (Sk-34) and CD29/34/45 (Sk-DN/29) fractions, and separately cultured/expanded in appropriate conditions within 2 weeks. Urethral damage was induced by manually removing one third of the wall of the muscle layer in nude rats. A mixture of expanded Sk-34 and Sk-DN/29 cells was applied on the damaged portion for the cell transplantation (CT) group. The same amount of media was used for the non-CT (NT) group. Urethral pressure profile was evaluated via electrical stimulation to assess functional recovery. Cell engraftments and differentiations were detected using immunohistochemistry and immunoelectron microscopy. Expression of angiogenic cytokines was also analyzed using reverse transcriptase-polymerase chain reaction and protein array. At 6 weeks after transplantation, the CT group showed a significantly higher functional recovery than the NT group (70.2% and 39.1%, respectively; P cells differentiated into skeletal muscle fibers, nerve-related Schwann cells, perineuriums, and vascular pericytes. Active paracrine angiogenic cytokines in the mixed cells were also detected with enhanced vascular formation in vivo. The transplantation of Sk-34 and Sk-DN/29 cells is potentially useful for the reconstitution of postoperative damage of the urethral rhabdosphincter and nerve-vascular networks.

Myostatin genetic inactivation inhibits myogenesis by muscle-derived stem cells in vitro but not when implanted in the mdx mouse muscle

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2013-01-01

Introduction Stimulating the commitment of implanted dystrophin+ muscle-derived stem cells (MDSCs) into myogenic, as opposed to lipofibrogenic lineages, is a promising therapeutic strategy for Duchenne muscular dystrophy (DMD). Methods To examine whether counteracting myostatin, a negative regulator of muscle mass and a pro-lipofibrotic factor, would help this process, we compared the in vitro myogenic and fibrogenic capacity of MDSCs from wild-type (WT) and myostatin knockout (Mst KO) mice under various modulators, the expression of key stem cell and myogenic genes, and the capacity of these MDSCs to repair the injured gastrocnemius in aged dystrophic mdx mice with exacerbated lipofibrosis. Results Surprisingly, the potent in vitro myotube formation by WT MDSCs was refractory to modulators of myostatin expression or activity, and the Mst KO MDSCs failed to form myotubes under various conditions, despite both MDSC expressing Oct 4 and various stem cell genes and differentiating into nonmyogenic lineages. The genetic inactivation of myostatin in MDSCs was associated with silencing of critical genes for early myogenesis (Actc1, Acta1, and MyoD). WT MDSCs implanted into the injured gastrocnemius of aged mdx mice significantly improved myofiber repair and reduced fat deposition and, to a lesser extent, fibrosis. In contrast to their in vitro behavior, Mst KO MDSCs in vivo also significantly improved myofiber repair, but had few effects on lipofibrotic degeneration. Conclusions Although WT MDSCs are very myogenic in culture and stimulate muscle repair after injury in the aged mdx mouse, myostatin genetic inactivation blocks myotube formation in vitro, but the myogenic capacity is recovered in vivo under the influence of the myostatin+ host-tissue environment, presumably by reactivation of key genes originally silenced in the Mst KO MDSCs. PMID:23295128

Requirement of myomaker-mediated stem cell fusion for skeletal muscle hypertrophy.

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Goh, Qingnian; Millay, Douglas P

2017-02-10

Fusion of skeletal muscle stem/progenitor cells is required for proper development and regeneration, however the significance of this process during adult muscle hypertrophy has not been explored. In response to muscle overload after synergist ablation in mice, we show that myomaker, a muscle specific membrane protein essential for myoblast fusion, is activated mainly in muscle progenitors and not myofibers. We rendered muscle progenitors fusion-incompetent through genetic deletion of myomaker in muscle stem cells and observed a complete reduction of overload-induced hypertrophy. This blunted hypertrophic response was associated with a reduction in Akt and p70s6k signaling and protein synthesis, suggesting a link between myonuclear accretion and activation of pro-hypertrophic pathways. Furthermore, fusion-incompetent muscle exhibited increased fibrosis after muscle overload, indicating a protective role for normal stem cell activity in reducing myofiber strain associated with hypertrophy. These findings reveal an essential contribution of myomaker-mediated stem cell fusion during physiological adult muscle hypertrophy.

Pluripotent Conversion of Muscle Stem Cells Without Reprogramming Factors or Small Molecules.

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Bose, Bipasha; Shenoy P, Sudheer

2016-02-01

Muscle derived stem cells (MDSCs) are multipotent stem cells that can differentiate into several lineages including skeletal muscle precursor cells. Here, we show that MDSCs from myostatin null mice (Mstn (-/-) ) can be readily induced into pluripotent stem cells without using reprogramming factors. Microarray studies revealed a strong upregulation of markers like Leukemia Inhibitory factor (LIF) and Leukemia Inhibitory factor receptor (LIFR) in Mstn (-/-) MDSCs as compared to wild type MDSCs (WT-MDSCs). Furthermore when cultured in mouse embryonic stem cell media with LIF for 95 days, Mstn (-/-) MDSCs formed embryonic stem cell (ES) like colonies. We termed such ES like cells as the culture-induced pluripotent stem cells (CiPSC). CiPSCs from Mstn (-/-) MDSCs were phenotypically similar to ESCs, expressed high levels of Oct4, Nanog, Sox2 and SSEA-1, maintained a normal karyotype. Furthermore, CiPSCs formed embryoid bodies and teratomas when injected into immunocompromised mice. In addition, CiPSCs differentiated into somatic cells of all three lineages. We further show that culturing in ES cell media, resulted in hypermethylation and downregulation of BMP2 in Mstn(-/-) MDSCs. Western blot further confirmed a down regulation of BMP2 signaling in Mstn (-/-) MDSCs in supportive of pluripotent reprogramming. Given that down regulation of BMP2 has been shown to induce pluripotency in cells, we propose that lack of myostatin epigenetically reprograms the MDSCs to become pluripotent stem cells. Thus, here we report the successful establishment of ES-like cells from adult stem cells of the non-germline origin under culture-induced conditions without introducing reprogramming genes.

Research progress in muscle-derived stem cells: Literature retrieval results based on international database.

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Zhang, Li; Wang, Wei

2012-04-05

To identify global research trends of muscle-derived stem cells (MDSCs) using a bibliometric analysis of the Web of Science, Research Portfolio Online Reporting Tools of the National Institutes of Health (NIH), and the Clinical Trials registry database (ClinicalTrials.gov). We performed a bibliometric analysis of data retrievals for MDSCs from 2002 to 2011 using the Web of Science, NIH, and ClinicalTrials.gov. (1) Web of Science: (a) peer-reviewed articles on MDSCs that were published and indexed in the Web of Science. (b) Type of articles: original research articles, reviews, meeting abstracts, proceedings papers, book chapters, editorial material and news items. (c) Year of publication: 2002-2011. (d) Citation databases: Science Citation Index-Expanded (SCI-E), 1899-present; Conference Proceedings Citation Index-Science (CPCI-S), 1991-present; Book Citation Index-Science (BKCI-S), 2005-present. (2) NIH: (a) Projects on MDSCs supported by the NIH. (b) Fiscal year: 1988-present. (3) ClinicalTrials.gov: All clinical trials relating to MDSCs were searched in this database. (1) Web of Science: (a) Articles that required manual searching or telephone access. (b) We excluded documents that were not published in the public domain. (c) We excluded a number of corrected papers from the total number of articles. (d) We excluded articles from the following databases: Social Sciences Citation Index (SSCI), 1898-present; Arts & Humanities Citation Index (A&HCI), 1975-present; Conference Proceedings Citation Index - Social Science & Humanities (CPCI-SSH), 1991-present; Book Citation Index - Social Sciences & Humanities (BKCI-SSH), 2005-present; Current Chemical Reactions (CCR-EXPANDED), 1985-present; Index Chemicus (IC), 1993-present. (2) NIH: (a) We excluded publications related to MDSCs that were supported by the NIH. (b) We limited the keyword search to studies that included MDSCs within the title or abstract. (3) ClinicalTrials.gov: (a) We excluded clinical trials that were

Muscle Stem Cell Therapy for the Treatment of DMD Associated Cardiomyopathy

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2013-10-01

SUBTITLE Muscle Stem Cell Therapy for the Treatment of DMD Associated Cardiomyopathy 5a. CONTRACT NUMBER Subproject 1: Muscle Stem Cell Therapy...various muscle diseases, including Duchenne muscular dystrophy (DMD), develop progressive cardiomyopathy. Cellular cardiomyoplasty, which involves the

Functional dysregulation of stem cells during aging: a focus on skeletal muscle stem cells.

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García-Prat, Laura; Sousa-Victor, Pedro; Muñoz-Cánoves, Pura

2013-09-01

Aging of an organism is associated with the functional decline of tissues and organs, as well as a sharp decline in the regenerative capacity of stem cells. A prevailing view holds that the aging rate of an individual depends on the ratio of tissue attrition to tissue regeneration. Therefore, manipulations that favor the balance towards regeneration may prevent or delay aging. Skeletal muscle is a specialized tissue composed of postmitotic myofibers that contract to generate force. Satellite cells are the adult stem cells responsible for skeletal muscle regeneration. Recent studies on the biology of skeletal muscle and satellite cells in aging have uncovered the critical impact of systemic and niche factors on stem cell functionality and demonstrated the capacity of aged satellite cells to rejuvenate and increase their regenerative potential when exposed to a youthful environment. Here we review the current literature on the coordinated relationship between cell extrinsic and intrinsic factors that regulate the function of satellite cells, and ultimately determine tissue homeostasis and repair during aging, and which encourage the search for new anti-aging strategies. © 2013 The Authors Journal compilation © 2013 FEBS.

In vivo evaluation of adipose- and muscle-derived stem cells as a treatment for nonhealing diabetic wounds using multimodal microscopy

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Li, Joanne; Pincu, Yair; Marjanovic, Marina; Bower, Andrew J.; Chaney, Eric J.; Jensen, Tor; Boppart, Marni D.; Boppart, Stephen A.

2016-08-01

Impaired skin wound healing is a significant comorbid condition of diabetes, which often results in nonhealing diabetic ulcers due to poor peripheral microcirculation, among other factors. The effectiveness of the regeneration of adipose-derived stem cells (ADSCs) and muscle-derived stem cells (MDSCs) was assessed using an integrated multimodal microscopy system equipped with two-photon fluorescence and second-harmonic generation imaging. These imaging modalities, integrated in a single platform for spatial and temporal coregistration, allowed us to monitor in vivo changes in the collagen network and cell dynamics in a skin wound. Fluorescently labeled ADSCs and MDSCs were applied topically to the wound bed of wild-type and diabetic (db/db) mice following punch biopsy. Longitudinal imaging demonstrated that ADSCs and MDSCs provided remarkable capacity for improved diabetic wound healing, and integrated microscopy revealed a more organized collagen remodeling in the wound bed of treated mice. The results from this study verify the regenerative capacity of stem cells toward healing and, with multimodal microscopy, provide insight regarding their impact on the skin microenvironment. The optical method outlined in this study, which has the potential for in vivo human use, may optimize the care and treatment of diabetic nonhealing wounds.

Adipose-Derived Stem Cells Promote Peripheral Nerve Regeneration In Vivo without Differentiation into Schwann-Like Lineage.

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Sowa, Yoshihiro; Kishida, Tsunao; Imura, Tetsuya; Numajiri, Toshiaki; Nishino, Kenichi; Tabata, Yasuhiko; Mazda, Osam

2016-02-01

During recent decades, multipotent stem cells were found to reside in the adipose tissue, and these adipose-derived stem cells were shown to play beneficial roles, like those of Schwann cells, in peripheral nerve regeneration. However, it has not been well established whether adipose-derived stem cells offer beneficial effects to peripheral nerve injuries in vivo as Schwann cells do. Furthermore, the in situ survival and differentiation of adipose-derived stem cells after transplantation at the injured peripheral nerve tissue remain to be fully elucidated. Adipose-derived stem cells and Schwann cells were transplanted with gelatin hydrogel tubes at the artificially blunted sciatic nerve lesion in mice. Neuroregenerative abilities of them were comparably estimated. Cre-loxP-mediated fate tracking was performed to visualize survival in vivo of transplanted adipose-derived stem cells and to investigate whether they differentiated into Schwann linage cells at the peripheral nerve injury site. The transplantation of adipose-derived stem cells promoted regeneration of axons, formation of myelin, and restoration of denervation muscle atrophy to levels comparable to those achieved by Schwann cell transplantation. The adipose-derived stem cells survived for at least 4 weeks after transplantation without differentiating into Schwann cells. Transplanted adipose-derived stem cells did not differentiate into Schwann cells but promoted peripheral nerve regeneration at the injured site. The neuroregenerative ability was comparable to that of Schwann cells. Adipose-derived stem cells at an undifferentiated stage may be used as an alternative cell source for autologous cell therapy for patients with peripheral nerve injury.

Robust generation and expansion of skeletal muscle progenitors and myocytes from human pluripotent stem cells.

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Shelton, Michael; Kocharyan, Avetik; Liu, Jun; Skerjanc, Ilona S; Stanford, William L

2016-05-15

Human pluripotent stem cells provide a developmental model to study early embryonic and tissue development, tease apart human disease processes, perform drug screens to identify potential molecular effectors of in situ regeneration, and provide a source for cell and tissue based transplantation. Highly efficient differentiation protocols have been established for many cell types and tissues; however, until very recently robust differentiation into skeletal muscle cells had not been possible unless driven by transgenic expression of master regulators of myogenesis. Nevertheless, several breakthrough protocols have been published in the past two years that efficiently generate cells of the skeletal muscle lineage from pluripotent stem cells. Here, we present an updated version of our recently described 50-day protocol in detail, whereby chemically defined media are used to drive and support muscle lineage development from initial CHIR99021-induced mesoderm through to PAX7-expressing skeletal muscle progenitors and mature skeletal myocytes. Furthermore, we report an optional method to passage and expand differentiating skeletal muscle progenitors approximately 3-fold every 2weeks using Collagenase IV and continued FGF2 supplementation. Both protocols have been optimized using a variety of human pluripotent stem cell lines including patient-derived induced pluripotent stem cells. Taken together, our differentiation and expansion protocols provide sufficient quantities of skeletal muscle progenitors and myocytes that could be used for a variety of studies. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.

Effects of sildenafil and/or muscle derived stem cells on myocardial infarction

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Wang Judy SC

2012-08-01

Full Text Available Abstract Background Previous studies have shown that long-term oral daily PDE 5 inhibitors (PDE5i counteract fibrosis, cell loss, and the resulting dysfunction in tissues of various rat organs and that implantation of skeletal muscle-derived stem cells (MDSC exerts some of these effects. PDE5i and stem cells in combination were found to be more effective in non-MI cardiac repair than each treatment separately. We have now investigated whether sildenafil at lower doses and MDSC, alone or in combination are effective to attenuate LV remodeling after MI in rats. Methods MI was induced in rats by ligature of the left anterior descending coronary artery. Treatment groups were: “Series A”: 1 untreated; 2 oral sildenafil 3 mg/kg/day from day 1; and “Series B”: intracardiac injection at day 7 of: 3 saline; 4 rat MDSC (106 cells; 5 as #4, with sildenafil as in #2. Before surgery, and at 1 and 4 weeks, the left ventricle ejection fraction (LVEF was measured. LV sections were stained for collagen, myofibroblasts, apoptosis, cardiomyocytes, and iNOS, followed by quantitative image analysis. Western blots estimated angiogenesis and myofibroblast accumulation, as well as potential sildenafil tachyphylaxis by PDE 5 expression. Zymography estimated MMPs 2 and 9 in serum. Results As compared to untreated MI rats, sildenafil improved LVEF, reduced collagen, myofibroblasts, and circulating MMPs, and increased cardiac troponin T. MDSC replicated most of these effects and stimulated cardiac angiogenesis. Concurrent MDSC/sildenafil counteracted cardiomyocyte and endothelial cells loss, but did not improve LVEF or angiogenesis, and upregulated PDE 5. Conclusions Long-term oral sildenafil, or MDSC given separately, reduce the MI fibrotic scar and improve left ventricular function in this rat model. The failure of the treatment combination may be due to inducing overexpression of PDE5.

Generation of Equine-Induced Pluripotent Stem Cells and Analysis of Their Therapeutic Potential for Muscle Injuries.

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Lee, Eun-Mi; Kim, Ah-Young; Lee, Eun-Joo; Park, Jin-Kyu; Park, Se-Il; Cho, Ssang-Goo; Kim, Hong Kyun; Kim, Shin-Yoon; Jeong, Kyu-Shik

2016-11-01

Horse health has become a major concern with the expansion of horse-related industries and sports; the importance of healthy muscles for horse performance and daily activities is undisputed. Here we generated equine-induced pluripotent stem cells (E-iPSCs) by reprogramming equine adipose-derived stem cells (E-ADSCs) into iPSCs using a polycistronic lentiviral vector encoding four transcription factors (i.e., Oct4, Sox2, Klf4, and c-Myc) and then examined their pluripotent characteristics. Subsequently, established E-iPSCs were transplanted into muscle-injured Rag/ mdx mice. The histopathology results showed that E-iPSC-transplanted mice exhibited enhanced muscle regeneration compared to controls. In addition, E-iPSC-derived myofibers were observed in the injured muscles. In conclusion, we show that E-iPSCs could be successfully generated from equine ADSCs and transplanted into injured muscles and that E-iPSCs have the capacity to induce regeneration of injured muscles.

A Human Pluripotent Stem Cell Model of Facioscapulohumeral Muscular Dystrophy-Affected Skeletal Muscles.

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Caron, Leslie; Kher, Devaki; Lee, Kian Leong; McKernan, Robert; Dumevska, Biljana; Hidalgo, Alejandro; Li, Jia; Yang, Henry; Main, Heather; Ferri, Giulia; Petek, Lisa M; Poellinger, Lorenz; Miller, Daniel G; Gabellini, Davide; Schmidt, Uli

2016-09-01

: Facioscapulohumeral muscular dystrophy (FSHD) represents a major unmet clinical need arising from the progressive weakness and atrophy of skeletal muscles. The dearth of adequate experimental models has severely hampered our understanding of the disease. To date, no treatment is available for FSHD. Human embryonic stem cells (hESCs) potentially represent a renewable source of skeletal muscle cells (SkMCs) and provide an alternative to invasive patient biopsies. We developed a scalable monolayer system to differentiate hESCs into mature SkMCs within 26 days, without cell sorting or genetic manipulation. Here we show that SkMCs derived from FSHD1-affected hESC lines exclusively express the FSHD pathogenic marker double homeobox 4 and exhibit some of the defects reported in FSHD. FSHD1 myotubes are thinner when compared with unaffected and Becker muscular dystrophy myotubes, and differentially regulate genes involved in cell cycle control, oxidative stress response, and cell adhesion. This cellular model will be a powerful tool for studying FSHD and will ultimately assist in the development of effective treatments for muscular dystrophies. This work describes an efficient and highly scalable monolayer system to differentiate human pluripotent stem cells (hPSCs) into skeletal muscle cells (SkMCs) and demonstrates disease-specific phenotypes in SkMCs derived from both embryonic and induced hPSCs affected with facioscapulohumeral muscular dystrophy. This study represents the first human stem cell-based cellular model for a muscular dystrophy that is suitable for high-throughput screening and drug development. ©AlphaMed Press.

Unique proliferation response in odontoblastic cells derived from human skeletal muscle stem cells by cytokine-induced matrix metalloproteinase-3

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Ozeki, Nobuaki; Hase, Naoko; Kawai, Rie; Yamaguchi, Hideyuki; Hiyama, Taiki; Kondo, Ayami; Nakata, Kazuhiko; Mogi, Makio

2015-01-01

A pro-inflammatory cytokine mixture (CM: interleukin (IL)-1β, tumor necrosis factor-α and interferon-γ) and IL-1β-induced matrix metalloproteinase (MMP)-3 activity have been shown to increase the proliferation of rat dental pulp cells and murine stem cell-derived odontoblast-like cells. This suggests that MMP-3 may regulate wound healing and regeneration in the odontoblast-rich dental pulp. Here, we determined whether these results can be extrapolated to human dental pulp by investigating the effects of CM-induced MMP-3 up-regulation on the proliferation and apoptosis of purified odontoblast-like cells derived from human skeletal muscle stem cells. We used siRNA to specifically reduce MMP-3 expression. We found that CM treatment increased MMP-3 mRNA and protein levels as well as MMP-3 activity. Cell proliferation was also markedly increased, with no changes in apoptosis, upon treatment with CM and following the application of exogenous MMP-3. Endogenous tissue inhibitors of metalloproteinases were constitutively expressed during all experiments and unaffected by MMP-3. Although treatment with MMP-3 siRNA suppressed cell proliferation, it also unexpectedly increased apoptosis. This siRNA-mediated increase in apoptosis could be reversed by exogenous MMP-3. These results demonstrate that cytokine-induced MMP-3 activity regulates cell proliferation and suppresses apoptosis in human odontoblast-like cells. - Highlights: • Pro-inflammatory cytokines induce MMP-3 activity in human odontoblast-like cells. • Increased MMP-3 activity can promote cell proliferation in odontoblasts. • Specific loss of MMP-3 increases apoptosis in odontoblasts. • MMP-3 has potential as a promising new target for pupal repair and regeneration

Unique proliferation response in odontoblastic cells derived from human skeletal muscle stem cells by cytokine-induced matrix metalloproteinase-3

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Ozeki, Nobuaki; Hase, Naoko; Kawai, Rie; Yamaguchi, Hideyuki; Hiyama, Taiki [Department of Endodontics, School of Dentistry, Aichi Gakuin University, 2-11 Suemori-dori, Chikusa-ku, Nagoya 464-8651, Aichi (Japan); Kondo, Ayami [Department of Medicinal Biochemistry, School of Pharmacy, Aichi Gakuin University, 1-100 Kusumoto, Chikusa-ku, Nagoya 464-8650 (Japan); Nakata, Kazuhiko [Department of Endodontics, School of Dentistry, Aichi Gakuin University, 2-11 Suemori-dori, Chikusa-ku, Nagoya 464-8651, Aichi (Japan); Mogi, Makio, E-mail: makio@dpc.agu.ac.jp [Department of Medicinal Biochemistry, School of Pharmacy, Aichi Gakuin University, 1-100 Kusumoto, Chikusa-ku, Nagoya 464-8650 (Japan)

2015-02-01

A pro-inflammatory cytokine mixture (CM: interleukin (IL)-1β, tumor necrosis factor-α and interferon-γ) and IL-1β-induced matrix metalloproteinase (MMP)-3 activity have been shown to increase the proliferation of rat dental pulp cells and murine stem cell-derived odontoblast-like cells. This suggests that MMP-3 may regulate wound healing and regeneration in the odontoblast-rich dental pulp. Here, we determined whether these results can be extrapolated to human dental pulp by investigating the effects of CM-induced MMP-3 up-regulation on the proliferation and apoptosis of purified odontoblast-like cells derived from human skeletal muscle stem cells. We used siRNA to specifically reduce MMP-3 expression. We found that CM treatment increased MMP-3 mRNA and protein levels as well as MMP-3 activity. Cell proliferation was also markedly increased, with no changes in apoptosis, upon treatment with CM and following the application of exogenous MMP-3. Endogenous tissue inhibitors of metalloproteinases were constitutively expressed during all experiments and unaffected by MMP-3. Although treatment with MMP-3 siRNA suppressed cell proliferation, it also unexpectedly increased apoptosis. This siRNA-mediated increase in apoptosis could be reversed by exogenous MMP-3. These results demonstrate that cytokine-induced MMP-3 activity regulates cell proliferation and suppresses apoptosis in human odontoblast-like cells. - Highlights: • Pro-inflammatory cytokines induce MMP-3 activity in human odontoblast-like cells. • Increased MMP-3 activity can promote cell proliferation in odontoblasts. • Specific loss of MMP-3 increases apoptosis in odontoblasts. • MMP-3 has potential as a promising new target for pupal repair and regeneration.

Evaluation of Stem Cell-Derived Red Blood Cells as a Transfusion Product Using a Novel Animal Model.

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Shah, Sandeep N; Gelderman, Monique P; Lewis, Emily M A; Farrel, John; Wood, Francine; Strader, Michael Brad; Alayash, Abdu I; Vostal, Jaroslav G

2016-01-01

Reliance on volunteer blood donors can lead to transfusion product shortages, and current liquid storage of red blood cells (RBCs) is associated with biochemical changes over time, known as 'the storage lesion'. Thus, there is a need for alternative sources of transfusable RBCs to supplement conventional blood donations. Extracorporeal production of stem cell-derived RBCs (stemRBCs) is a potential and yet untapped source of fresh, transfusable RBCs. A number of groups have attempted RBC differentiation from CD34+ cells. However, it is still unclear whether these stemRBCs could eventually be effective substitutes for traditional RBCs due to potential differences in oxygen carrying capacity, viability, deformability, and other critical parameters. We have generated ex vivo stemRBCs from primary human cord blood CD34+ cells and compared them to donor-derived RBCs based on a number of in vitro parameters. In vivo, we assessed stemRBC circulation kinetics in an animal model of transfusion and oxygen delivery in a mouse model of exercise performance. Our novel, chronically anemic, SCID mouse model can evaluate the potential of stemRBCs to deliver oxygen to tissues (muscle) under resting and exercise-induced hypoxic conditions. Based on our data, stem cell-derived RBCs have a similar biochemical profile compared to donor-derived RBCs. While certain key differences remain between donor-derived RBCs and stemRBCs, the ability of stemRBCs to deliver oxygen in a living organism provides support for further development as a transfusion product.

Evaluation of Stem Cell-Derived Red Blood Cells as a Transfusion Product Using a Novel Animal Model.

Directory of Open Access Journals (Sweden)

Sandeep N Shah

Full Text Available Reliance on volunteer blood donors can lead to transfusion product shortages, and current liquid storage of red blood cells (RBCs is associated with biochemical changes over time, known as 'the storage lesion'. Thus, there is a need for alternative sources of transfusable RBCs to supplement conventional blood donations. Extracorporeal production of stem cell-derived RBCs (stemRBCs is a potential and yet untapped source of fresh, transfusable RBCs. A number of groups have attempted RBC differentiation from CD34+ cells. However, it is still unclear whether these stemRBCs could eventually be effective substitutes for traditional RBCs due to potential differences in oxygen carrying capacity, viability, deformability, and other critical parameters. We have generated ex vivo stemRBCs from primary human cord blood CD34+ cells and compared them to donor-derived RBCs based on a number of in vitro parameters. In vivo, we assessed stemRBC circulation kinetics in an animal model of transfusion and oxygen delivery in a mouse model of exercise performance. Our novel, chronically anemic, SCID mouse model can evaluate the potential of stemRBCs to deliver oxygen to tissues (muscle under resting and exercise-induced hypoxic conditions. Based on our data, stem cell-derived RBCs have a similar biochemical profile compared to donor-derived RBCs. While certain key differences remain between donor-derived RBCs and stemRBCs, the ability of stemRBCs to deliver oxygen in a living organism provides support for further development as a transfusion product.

Reconstruction of Multiple Facial Nerve Branches Using Skeletal Muscle-Derived Multipotent Stem Cell Sheet-Pellet Transplantation.

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Kosuke Saito

Full Text Available Head and neck cancer is often diagnosed at advanced stages, and surgical resection with wide margins is generally indicated, despite this treatment being associated with poor postoperative quality of life (QOL. We have previously reported on the therapeutic effects of skeletal muscle-derived multipotent stem cells (Sk-MSCs, which exert reconstitution capacity for muscle-nerve-blood vessel units. Recently, we further developed a 3D patch-transplantation system using Sk-MSC sheet-pellets. The aim of this study is the application of the 3D Sk-MSC transplantation system to the reconstitution of facial complex nerve-vascular networks after severe damage. Mouse experiments were performed for histological analysis and rats were used for functional examinations. The Sk-MSC sheet-pellets were prepared from GFP-Tg mice and SD rats, and were transplanted into the facial resection model (ST. Culture medium was transplanted as a control (NT. In the mouse experiment, facial-nerve-palsy (FNP scoring was performed weekly during the recovery period, and immunohistochemistry was used for the evaluation of histological recovery after 8 weeks. In rats, contractility of facial muscles was measured via electrical stimulation of facial nerves root, as the marker of total functional recovery at 8 weeks after transplantation. The ST-group showed significantly higher FNP (about three fold scores when compared to the NT-group after 2-8 weeks. Similarly, significant functional recovery of whisker movement muscles was confirmed in the ST-group at 8 weeks after transplantation. In addition, engrafted GFP+ cells formed complex branches of nerve-vascular networks, with differentiation into Schwann cells and perineurial/endoneurial cells, as well as vascular endothelial and smooth muscle cells. Thus, Sk-MSC sheet-pellet transplantation is potentially useful for functional reconstitution therapy of large defects in facial nerve-vascular networks.

Reconstruction of Multiple Facial Nerve Branches Using Skeletal Muscle-Derived Multipotent Stem Cell Sheet-Pellet Transplantation.

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Saito, Kosuke; Tamaki, Tetsuro; Hirata, Maki; Hashimoto, Hiroyuki; Nakazato, Kenei; Nakajima, Nobuyuki; Kazuno, Akihito; Sakai, Akihiro; Iida, Masahiro; Okami, Kenji

2015-01-01

Head and neck cancer is often diagnosed at advanced stages, and surgical resection with wide margins is generally indicated, despite this treatment being associated with poor postoperative quality of life (QOL). We have previously reported on the therapeutic effects of skeletal muscle-derived multipotent stem cells (Sk-MSCs), which exert reconstitution capacity for muscle-nerve-blood vessel units. Recently, we further developed a 3D patch-transplantation system using Sk-MSC sheet-pellets. The aim of this study is the application of the 3D Sk-MSC transplantation system to the reconstitution of facial complex nerve-vascular networks after severe damage. Mouse experiments were performed for histological analysis and rats were used for functional examinations. The Sk-MSC sheet-pellets were prepared from GFP-Tg mice and SD rats, and were transplanted into the facial resection model (ST). Culture medium was transplanted as a control (NT). In the mouse experiment, facial-nerve-palsy (FNP) scoring was performed weekly during the recovery period, and immunohistochemistry was used for the evaluation of histological recovery after 8 weeks. In rats, contractility of facial muscles was measured via electrical stimulation of facial nerves root, as the marker of total functional recovery at 8 weeks after transplantation. The ST-group showed significantly higher FNP (about three fold) scores when compared to the NT-group after 2-8 weeks. Similarly, significant functional recovery of whisker movement muscles was confirmed in the ST-group at 8 weeks after transplantation. In addition, engrafted GFP+ cells formed complex branches of nerve-vascular networks, with differentiation into Schwann cells and perineurial/endoneurial cells, as well as vascular endothelial and smooth muscle cells. Thus, Sk-MSC sheet-pellet transplantation is potentially useful for functional reconstitution therapy of large defects in facial nerve-vascular networks.

Aging of the skeletal muscle extracellular matrix drives a stem cell fibrogenic conversion.

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Stearns-Reider, Kristen M; D'Amore, Antonio; Beezhold, Kevin; Rothrauff, Benjamin; Cavalli, Loredana; Wagner, William R; Vorp, David A; Tsamis, Alkiviadis; Shinde, Sunita; Zhang, Changqing; Barchowsky, Aaron; Rando, Thomas A; Tuan, Rocky S; Ambrosio, Fabrisia

2017-06-01

Age-related declines in skeletal muscle regeneration have been attributed to muscle stem cell (MuSC) dysfunction. Aged MuSCs display a fibrogenic conversion, leading to fibrosis and impaired recovery after injury. Although studies have demonstrated the influence of in vitro substrate characteristics on stem cell fate, whether and how aging of the extracellular matrix (ECM) affects stem cell behavior has not been investigated. Here, we investigated the direct effect of the aged muscle ECM on MuSC lineage specification. Quantification of ECM topology and muscle mechanical properties reveals decreased collagen tortuosity and muscle stiffening with increasing age. Age-related ECM alterations directly disrupt MuSC responses, and MuSCs seeded ex vivo onto decellularized ECM constructs derived from aged muscle display increased expression of fibrogenic markers and decreased myogenicity, compared to MuSCs seeded onto young ECM. This fibrogenic conversion is recapitulated in vitro when MuSCs are seeded directly onto matrices elaborated by aged fibroblasts. When compared to young fibroblasts, fibroblasts isolated from aged muscle display increased nuclear levels of the mechanosensors, Yes-associated protein (YAP)/transcriptional coactivator with PDZ-binding motif (TAZ), consistent with exposure to a stiff microenvironment in vivo. Accordingly, preconditioning of young fibroblasts by seeding them onto a substrate engineered to mimic the stiffness of aged muscle increases YAP/TAZ nuclear translocation and promotes secretion of a matrix that favors MuSC fibrogenesis. The findings here suggest that an age-related increase in muscle stiffness drives YAP/TAZ-mediated pathogenic expression of matricellular proteins by fibroblasts, ultimately disrupting MuSC fate. © 2017 The Authors. Aging Cell published by the Anatomical Society and John Wiley & Sons Ltd.

The muscle stem cell niche : regulation of satellite cells during regeneration

NARCIS (Netherlands)

Boonen, K.J.M.; Post, M.J.

2008-01-01

Satellite cells are considered to be adult skeletal muscle stem cells. Their ability to regenerate large muscle defects is highly dependent on their specific niche. When these cells are cultured in vitro, the loss of this niche leads to a loss of proliferative capacity and defective regeneration

STEM CELLS IN THE TREATMENT OF URINARY STRESS INCONTINENCE – A VISION OF FUTURE?

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Adolf Lukanović

2018-02-01

Full Text Available Background. For one third of all women urinary incontinence is a health problem. Different strategies of treatment have been used, namely surgical, medical and physiotherapy. To this armamentarium a new method – adult stem cells therapy – has been added. Methods. The source of adult autologues multipotent stem cells consists of muscle-derived stem cells, adipose-derived stem cells and mesenchymed bone marrow cells. In facilities for stem cells with processing and in the presence of specific inducing factors, muscle derived stem cells can be differentiated to striated or smooth muscles. Clinical work is based on animal models that with immunohistochemical analysis demonstrated new skeletal-muscle fiber formation at the site of injection of stem cells. In stress urinary incontinence (SUI autologues bioptic material is obtained by using a musle needle biopsy device. From the biopsy speciment with dissociation muscle-derived stem cells are isolated and injected under local anaesthesia in the middle urethra and/or external urethral sphincter. Initial results of SUI treatment with adult muscle-derived stem cells suggest that perspectives of this method are encouraging. Conclusions. Stem cell therapy is promising to become minimally invasive method for reconstruction of the muscles and connective tissue of urethra and external urethral sphincter.

Injectable biomimetic liquid crystalline scaffolds enhance muscle stem cell transplantation

Science.gov (United States)

Sleep, Eduard; McClendon, Mark T.; Preslar, Adam T.; Chen, Charlotte H.; Sangji, M. Hussain; Pérez, Charles M. Rubert; Haynes, Russell D.; Meade, Thomas J.; Blau, Helen M.; Stupp, Samuel I.

2017-01-01

Muscle stem cells are a potent cell population dedicated to efficacious skeletal muscle regeneration, but their therapeutic utility is currently limited by mode of delivery. We developed a cell delivery strategy based on a supramolecular liquid crystal formed by peptide amphiphiles (PAs) that encapsulates cells and growth factors within a muscle-like unidirectionally ordered environment of nanofibers. The stiffness of the PA scaffolds, dependent on amino acid sequence, was found to determine the macroscopic degree of cell alignment templated by the nanofibers in vitro. Furthermore, these PA scaffolds support myogenic progenitor cell survival and proliferation and they can be optimized to induce cell differentiation and maturation. We engineered an in vivo delivery system to assemble scaffolds by injection of a PA solution that enabled coalignment of scaffold nanofibers with endogenous myofibers. These scaffolds locally retained growth factors, displayed degradation rates matching the time course of muscle tissue regeneration, and markedly enhanced the engraftment of muscle stem cells in injured and noninjured muscles in mice. PMID:28874575

Modeling Cerebrovascular Pathophysiology in Amyloid-β Metabolism using Neural-Crest-Derived Smooth Muscle Cells

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Christine Cheung

2014-10-01

Full Text Available Summary: There is growing recognition of cerebrovascular contributions to neurodegenerative diseases. In the walls of cerebral arteries, amyloid-beta (Aβ accumulation is evident in a majority of aged people and patients with cerebral amyloid angiopathy. Here, we leverage human pluripotent stem cells to generate vascular smooth muscle cells (SMCs from neural crest progenitors, recapitulating brain-vasculature-specific attributes of Aβ metabolism. We confirm that the lipoprotein receptor, LRP1, functions in our neural-crest-derived SMCs to mediate Aβ uptake and intracellular lysosomal degradation. Hypoxia significantly compromises the contribution of SMCs to Aβ clearance by suppressing LRP1 expression. This enabled us to develop an assay of Aβ uptake by using the neural crest-derived SMCs with hypoxia as a stress paradigm. We then tested several vascular protective compounds in a high-throughput format, demonstrating the value of stem-cell-based phenotypic screening for novel therapeutics and drug repurposing, aimed at alleviating amyloid burden. : The contribution of blood vessel pathologies to neurodegenerative disorders is relatively neglected, partly due to inadequate human tissues for research. By using human stem cells, Cheung et al. establish a method of generating vascular smooth muscle cells (SMCs from neural crest progenitors, the primary precursors that give rise to brain blood vessels. These stem-cell-derived SMCs display defective amyloid processing under chronic hypoxia, a phenomenon well documented in the cerebral vasculatures of aged people and patients with Alzheimer’s disease.

Mac-1low early myeloid cells in the bone marrow-derived SP fraction migrate into injured skeletal muscle and participate in muscle regeneration

International Nuclear Information System (INIS)

Ojima, Koichi; Uezumi, Akiyoshi; Miyoshi, Hiroyuki; Masuda, Satoru; Morita, Yohei; Fukase, Akiko; Hattori, Akihito; Nakauchi, Hiromitsu; Miyagoe-Suzuki, Yuko; Takeda, Shin'ichi

2004-01-01

Recent studies have shown that bone marrow (BM) cells, including the BM side population (BM-SP) cells that enrich hematopoietic stem cells (HSCs), are incorporated into skeletal muscle during regeneration, but it is not clear how and what kinds of BM cells contribute to muscle fiber regeneration. We found that a large number of SP cells migrated from BM to muscles following injury in BM-transplanted mice. These BM-derived SP cells in regenerating muscles expressed different surface markers from those of HSCs and could not reconstitute the mouse blood system. BM-derived SP/Mac-1 low cells increased in number in regenerating muscles following injury. Importantly, our co-culture studies with activated satellite cells revealed that this fraction carried significant potential for myogenic differentiation. By contrast, mature inflammatory (Mac-1 high ) cells showed negligible myogenic activities. Further, these BM-derived SP/Mac-1 low cells gave rise to mononucleate myocytes, indicating that their myogenesis was not caused by stochastic fusion with host myogenic cells, although they required cell-to-cell contact with myogenic cells for muscle differentiation. Taken together, our data suggest that neither HSCs nor mature inflammatory cells, but Mac-1 low early myeloid cells in the BM-derived SP fraction, play an important role in regenerating skeletal muscles

Reconstitution of the complete rupture in musculotendinous junction using skeletal muscle-derived multipotent stem cell sheet-pellets as a "bio-bond".

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Hashimoto, Hiroyuki; Tamaki, Tetsuro; Hirata, Maki; Uchiyama, Yoshiyasu; Sato, Masato; Mochida, Joji

2016-01-01

Background. Significant and/or complete rupture in the musculotendinous junction (MTJ) is a challenging lesion to treat because of the lack of reliable suture methods. Skeletal muscle-derived multipotent stem cell (Sk-MSC) sheet-pellets, which are able to reconstitute peripheral nerve and muscular/vascular tissues with robust connective tissue networks, have been applied as a "bio-bond". Methods. Sk-MSC sheet-pellets, derived from GFP transgenic-mice after 7 days of expansion culture, were detached with EDTA to maintain cell-cell connections. A completely ruptured MTJ model was prepared in the right tibialis anterior (TA) of the recipient mice, and was covered with sheet-pellets. The left side was preserved as a contralateral control. The control group received the same amount of the cell-free medium. The sheet-pellet transplantation (SP) group was further divided into two groups; as the short term (4-8 weeks) and long term (14-18 weeks) recovery group. At each time point after transplantation, tetanic tension output was measured through the electrical stimulation of the sciatic nerve. The behavior of engrafted GFP(+) tissues and cells was analyzed by fluorescence immunohistochemistry. Results. The SP short term recovery group showed average 64% recovery of muscle mass, and 36% recovery of tetanic tension output relative to the contralateral side. Then, the SP long term recovery group showed increased recovery of average muscle mass (77%) and tetanic tension output (49%). However, the control group showed no recovery of continuity between muscle and tendon, and demonstrated increased muscle atrophy, with coalescence to the tibia during 4-8 weeks after operation. Histological evidence also supported the above functional recovery of SP group. Engrafted Sk-MSCs primarily formed the connective tissues and muscle fibers, including nerve-vascular networks, and bridged the ruptured tendon-muscle fiber units, with differentiation into skeletal muscle cells, Schwann cells

Characterization Of Bovine Adipose-Derived Stem Cells

OpenAIRE

Daniel Cebo

2017-01-01

Bovine adipose-derived stem cells were obtained from the subcutaneous abdominal adipose tissue. The cells were cultured by the modified tissue-explants method developed in our laboratory and then analyzed using optical microscopy and flow cytometry. These cells were able to replicate in our cell culture conditions. cell Flow cytometry showed that bovine adipose-derived stem cells expressed mesenchymal stem cell markers CD73 and CD90. Meanwhile haematopoietic markers CD45 and CD34 are absent f...

DNA Methylation in Skeletal Muscle Stem Cell Specification, Proliferation, and Differentiation

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Rhianna C. Laker

2016-01-01

Full Text Available An unresolved and critically important question in skeletal muscle biology is how muscle stem cells initiate and regulate the genetic program during muscle development. Epigenetic dynamics are essential for cellular development and organogenesis in early life and it is becoming increasingly clear that epigenetic remodeling may also be responsible for the cellular adaptations that occur in later life. DNA methylation of cytosine bases within CpG dinucleotide pairs is an important epigenetic modification that reduces gene expression when located within a promoter or enhancer region. Recent advances in the field suggest that epigenetic regulation is essential for skeletal muscle stem cell identity and subsequent cell development. This review summarizes what is currently known about how skeletal muscle stem cells regulate the myogenic program through DNA methylation, discusses a novel role for metabolism in this process, and addresses DNA methylation dynamics in adult skeletal muscle in response to physical activity.

Skeletal muscle stem cells from animals I. Basic cell biology

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Skeletal muscle stem cells from food-producing animals have been of interest to agricultural life scientists seeking to develop a better understanding of the molecular regulation of lean tissue (skeletal muscle protein hypertrophy) and intramuscular fat (marbling) development. Enhanced understanding...

Differential Gene Expression Profiling of Dystrophic Dog Muscle after MuStem Cell Transplantation.

Science.gov (United States)

Robriquet, Florence; Lardenois, Aurélie; Babarit, Candice; Larcher, Thibaut; Dubreil, Laurence; Leroux, Isabelle; Zuber, Céline; Ledevin, Mireille; Deschamps, Jack-Yves; Fromes, Yves; Cherel, Yan; Guevel, Laetitia; Rouger, Karl

2015-01-01

Several adult stem cell populations exhibit myogenic regenerative potential, thus representing attractive candidates for therapeutic approaches of neuromuscular diseases such as Duchenne Muscular Dystrophy (DMD). We have recently shown that systemic delivery of MuStem cells, skeletal muscle-resident stem cells isolated in healthy dog, generates the remodelling of muscle tissue and gives rise to striking clinical benefits in Golden Retriever Muscular Dystrophy (GRMD) dog. This global effect, which is observed in the clinically relevant DMD animal model, leads us to question here the molecular pathways that are impacted by MuStem cell transplantation. To address this issue, we compare the global gene expression profile between healthy, GRMD and MuStem cell treated GRMD dog muscle, four months after allogenic MuStem cell transplantation. In the dystrophic context of the GRMD dog, disease-related deregulation is observed in the case of 282 genes related to various processes such as inflammatory response, regeneration, calcium ion binding, extracellular matrix organization, metabolism and apoptosis regulation. Importantly, we reveal the impact of MuStem cell transplantation on several molecular and cellular pathways based on a selection of 31 genes displaying signals specifically modulated by the treatment. Concomitant with a diffuse dystrophin expression, a histological remodelling and a stabilization of GRMD dog clinical status, we show that cell delivery is associated with an up-regulation of genes reflecting a sustained enhancement of muscle regeneration. We also identify a decreased mRNA expression of a set of genes having metabolic functions associated with lipid homeostasis and energy. Interestingly, ubiquitin-mediated protein degradation is highly enhanced in GRMD dog muscle after systemic delivery of MuStem cells. Overall, our results provide the first high-throughput characterization of GRMD dog muscle and throw new light on the complex molecular

Differential Gene Expression Profiling of Dystrophic Dog Muscle after MuStem Cell Transplantation.

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Florence Robriquet

Full Text Available Several adult stem cell populations exhibit myogenic regenerative potential, thus representing attractive candidates for therapeutic approaches of neuromuscular diseases such as Duchenne Muscular Dystrophy (DMD. We have recently shown that systemic delivery of MuStem cells, skeletal muscle-resident stem cells isolated in healthy dog, generates the remodelling of muscle tissue and gives rise to striking clinical benefits in Golden Retriever Muscular Dystrophy (GRMD dog. This global effect, which is observed in the clinically relevant DMD animal model, leads us to question here the molecular pathways that are impacted by MuStem cell transplantation. To address this issue, we compare the global gene expression profile between healthy, GRMD and MuStem cell treated GRMD dog muscle, four months after allogenic MuStem cell transplantation.In the dystrophic context of the GRMD dog, disease-related deregulation is observed in the case of 282 genes related to various processes such as inflammatory response, regeneration, calcium ion binding, extracellular matrix organization, metabolism and apoptosis regulation. Importantly, we reveal the impact of MuStem cell transplantation on several molecular and cellular pathways based on a selection of 31 genes displaying signals specifically modulated by the treatment. Concomitant with a diffuse dystrophin expression, a histological remodelling and a stabilization of GRMD dog clinical status, we show that cell delivery is associated with an up-regulation of genes reflecting a sustained enhancement of muscle regeneration. We also identify a decreased mRNA expression of a set of genes having metabolic functions associated with lipid homeostasis and energy. Interestingly, ubiquitin-mediated protein degradation is highly enhanced in GRMD dog muscle after systemic delivery of MuStem cells.Overall, our results provide the first high-throughput characterization of GRMD dog muscle and throw new light on the complex

Development of heart muscle-cell diversity: a help or a hindrance for phenotyping embryonic stem cell-derived cardiomyocytes

NARCIS (Netherlands)

Fijnvandraat, Arnoud C.; Lekanne Deprez, Ronald H.; Moorman, Antoon F. M.

2003-01-01

Despite the advances in cardiovascular treatment, cardiac disease remains a major cause of morbidity in all industrialized countries. The extraordinary potential of (embryonic) stem cells for therapeutic purposes has revolutionized ideas about cardiac repair of diseased cardiac muscle to exciting

Influence of exercise and aging on extracellular matrix composition in the skeletal muscle stem cell niche.

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Garg, Koyal; Boppart, Marni D

2016-11-01

Skeletal muscle is endowed with a remarkable capacity for regeneration, primarily due to the reserve pool of muscle resident satellite cells. The satellite cell is the physiologically quiescent muscle stem cell that resides beneath the basal lamina and adjacent to the sarcolemma. The anatomic location of satellite cells is in close proximity to vasculature where they interact with other muscle resident stem/stromal cells (e.g., mesenchymal stem cells and pericytes) through paracrine mechanisms. This mini-review describes the components of the muscle stem cell niche, as well as the influence of exercise and aging on the muscle stem cell niche. Although exercise promotes ECM reorganization and stem cell accumulation, aging is associated with dense ECM deposition and loss of stem cell function resulting in reduced regenerative capacity and strength. An improved understanding of the niche elements will be valuable to inform the development of therapeutic interventions aimed at improving skeletal muscle regeneration and adaptation over the life span. Copyright © 2016 the American Physiological Society.

Comparison of Gene Expression in Human Embryonic Stem Cells, hESC-Derived Mesenchymal Stem Cells and Human Mesenchymal Stem Cells.

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Barbet, Romain; Peiffer, Isabelle; Hatzfeld, Antoinette; Charbord, Pierre; Hatzfeld, Jacques A

2011-01-01

We present a strategy to identify developmental/differentiation and plasma membrane marker genes of the most primitive human Mesenchymal Stem Cells (hMSCs). Using sensitive and quantitative TaqMan Low Density Arrays (TLDA) methodology, we compared the expression of 381 genes in human Embryonic Stem Cells (hESCs), hESC-derived MSCs (hES-MSCs), and hMSCs. Analysis of differentiation genes indicated that hES-MSCs express the sarcomeric muscle lineage in addition to the classical mesenchymal lineages, suggesting they are more primitive than hMSCs. Transcript analysis of membrane antigens suggests that IL1R1(low), BMPR1B(low), FLT4(low), LRRC32(low), and CD34 may be good candidates for the detection and isolation of the most primitive hMSCs. The expression in hMSCs of cytokine genes, such as IL6, IL8, or FLT3LG, without expression of the corresponding receptor, suggests a role for these cytokines in the paracrine control of stem cell niches. Our database may be shared with other laboratories in order to explore the considerable clinical potential of hES-MSCs, which appear to represent an intermediate developmental stage between hESCs and hMSCs.

Muscle Stem Cell Fate Is Controlled by the Cell-Polarity Protein Scrib

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Yusuke Ono

2015-02-01

Full Text Available Satellite cells are resident skeletal muscle stem cells that supply myonuclei for homeostasis, hypertrophy, and repair in adult muscle. Scrib is one of the major cell-polarity proteins, acting as a potent tumor suppressor in epithelial cells. Here, we show that Scrib also controls satellite-cell-fate decisions in adult mice. Scrib is undetectable in quiescent cells but becomes expressed during activation. Scrib is asymmetrically distributed in dividing daughter cells, with robust accumulation in cells committed to myogenic differentiation. Low Scrib expression is associated with the proliferative state and preventing self-renewal, whereas high Scrib levels reduce satellite cell proliferation. Satellite-cell-specific knockout of Scrib in mice causes a drastic and insurmountable defect in muscle regeneration. Thus, Scrib is a regulator of tissue stem cells, controlling population expansion and self-renewal with Scrib expression dynamics directing satellite cell fate.

Skin appendage-derived stem cells: cell biology and potential for wound repair

OpenAIRE

Xie, Jiangfan; Yao, Bin; Han, Yutong; Huang, Sha; Fu, Xiaobing

2016-01-01

Stem cells residing in the epidermis and skin appendages are imperative for skin homeostasis and regeneration. These stem cells also participate in the repair of the epidermis after injuries, inducing restoration of tissue integrity and function of damaged tissue. Unlike epidermis-derived stem cells, comprehensive knowledge about skin appendage-derived stem cells remains limited. In this review, we summarize the current knowledge of skin appendage-derived stem cells, including their fundament...

Tissue-specific stem cells: Lessons from the skeletal muscle satellite cell

Science.gov (United States)

Brack, Andrew S.; Rando, Thomas A.

2012-01-01

In 1961, the satellite cell was first identified when electron microscopic examination of skeletal muscle demonstrated a cell wedged between the plasma membrane of the muscle fiber and the basement membrane. In recent years it has been conclusively demonstrated that the satellite cell is the primary cellular source for muscle regeneration and is equipped with the potential to self renew, thus functioning as a bone fide skeletal muscle stem cell (MuSC). As we move past the 50th anniversary of the satellite cell, we take this opportunity to discuss the current state of the art and dissect the unknowns in the MuSC field. PMID:22560074

Differential Gene Expression Profiling of Dystrophic Dog Muscle after MuStem Cell Transplantation

Science.gov (United States)

Babarit, Candice; Larcher, Thibaut; Dubreil, Laurence; Leroux, Isabelle; Zuber, Céline; Ledevin, Mireille; Deschamps, Jack-Yves; Fromes, Yves; Cherel, Yan; Guevel, Laetitia; Rouger, Karl

2015-01-01

Background Several adult stem cell populations exhibit myogenic regenerative potential, thus representing attractive candidates for therapeutic approaches of neuromuscular diseases such as Duchenne Muscular Dystrophy (DMD). We have recently shown that systemic delivery of MuStem cells, skeletal muscle-resident stem cells isolated in healthy dog, generates the remodelling of muscle tissue and gives rise to striking clinical benefits in Golden Retriever Muscular Dystrophy (GRMD) dog. This global effect, which is observed in the clinically relevant DMD animal model, leads us to question here the molecular pathways that are impacted by MuStem cell transplantation. To address this issue, we compare the global gene expression profile between healthy, GRMD and MuStem cell treated GRMD dog muscle, four months after allogenic MuStem cell transplantation. Results In the dystrophic context of the GRMD dog, disease-related deregulation is observed in the case of 282 genes related to various processes such as inflammatory response, regeneration, calcium ion binding, extracellular matrix organization, metabolism and apoptosis regulation. Importantly, we reveal the impact of MuStem cell transplantation on several molecular and cellular pathways based on a selection of 31 genes displaying signals specifically modulated by the treatment. Concomitant with a diffuse dystrophin expression, a histological remodelling and a stabilization of GRMD dog clinical status, we show that cell delivery is associated with an up-regulation of genes reflecting a sustained enhancement of muscle regeneration. We also identify a decreased mRNA expression of a set of genes having metabolic functions associated with lipid homeostasis and energy. Interestingly, ubiquitin-mediated protein degradation is highly enhanced in GRMD dog muscle after systemic delivery of MuStem cells. Conclusions Overall, our results provide the first high-throughput characterization of GRMD dog muscle and throw new light on the

Adipose-Derived Stem Cells and Application Areas

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Mujde Kivanc

2015-09-01

Full Text Available The use of stem cells derived from adipose tissue as an autologous and self-replenishing source for a variety of differentiated cell phenotypes, provides a great deal of promise for reconstructive surgery. The secret of the human body, stem cells are reserved. Stem cells are undifferentiated cells found in the human body placed in any body tissue characteristics that differentiate and win ever known to cross the tissue instead of more than 200 diseases and thus improve and, rejuvenates the tissues. So far, the cord blood of newborn babies are used as a source of stem cells, bone marrow, and twenty years after tooth stem cells in human adipose tissue, scientists studied more than other sources of stem cells in adipose tissue and discovered that. Increase in number of in vitro studies on adult stem cells, depending on many variables is that the stem cells directly to the desired soybean optimization can be performed.. We will conclude by assessing potential avenues for developing this incredibly promising field. The aim of this paper is to review the existing literature on applications of harvest, purification, characterization and cryopreservation of adipose-derived stem cells (ASCs. [Cukurova Med J 2015; 40(3.000: 399-408

Quantitative gene expression profiling of CD45+ and CD45- skeletal muscle-derived side population cells

DEFF Research Database (Denmark)

Ditte Caroline Andersen, Ditte Caroline; Kristiansen, Gitte Qvist; Jensen, Line

2012-01-01

The skeletal muscle-derived side population (mSP) which highly excludes Hoechst 33342 is composed of CD45(+) and CD45(-) subpopulations; yet, rareness of mSP cells in general has complicated extensive quantitative analysis of gene expression profiles in primarily isolated mSP cells. Here, we desc...... a satellite cell subpopulation) remain in the mSPCD45(-) fraction, and we show that these cells express high levels of many of the known myogenic precursor/stem cell related markers, including Pax7 and Myf5.......The skeletal muscle-derived side population (mSP) which highly excludes Hoechst 33342 is composed of CD45(+) and CD45(-) subpopulations; yet, rareness of mSP cells in general has complicated extensive quantitative analysis of gene expression profiles in primarily isolated mSP cells. Here, we...... describe the isolation of adult mouse normal skeletal muscle residing SPCD45(+) and SPCD45(-) cells from a parent mononuclear muscle-derived cell (MDC) population. Relative quantitative real time PCR (RT-PCR) of 64 genes revealed that mSPCD45(-) compared with mSPCD45(+) was enriched for cells expressing...

Adipose-derived mesenchymal stem cells and regenerative medicine.

Science.gov (United States)

Konno, Masamitsu; Hamabe, Atsushi; Hasegawa, Shinichiro; Ogawa, Hisataka; Fukusumi, Takahito; Nishikawa, Shimpei; Ohta, Katsuya; Kano, Yoshihiro; Ozaki, Miyuki; Noguchi, Yuko; Sakai, Daisuke; Kudoh, Toshihiro; Kawamoto, Koichi; Eguchi, Hidetoshi; Satoh, Taroh; Tanemura, Masahiro; Nagano, Hiroaki; Doki, Yuichiro; Mori, Masaki; Ishii, Hideshi

2013-04-01

Adipose tissue-derived mesenchymal stem cells (ADSCs) are multipotent and can differentiate into various cell types, including osteocytes, adipocytes, neural cells, vascular endothelial cells, cardiomyocytes, pancreatic β-cells, and hepatocytes. Compared with the extraction of other stem cells such as bone marrow-derived mesenchymal stem cells (BMSCs), that of ADSCs requires minimally invasive techniques. In the field of regenerative medicine, the use of autologous cells is preferable to embryonic stem cells or induced pluripotent stem cells. Therefore, ADSCs are a useful resource for drug screening and regenerative medicine. Here we present the methods and mechanisms underlying the induction of multilineage cells from ADSCs. © 2013 The Authors Development, Growth & Differentiation © 2013 Japanese Society of Developmental Biologists.

Vessel-associated stem cells from skeletal muscle: From biology to future uses in cell therapy.

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Sancricca, Cristina; Mirabella, Massimiliano; Gliubizzi, Carla; Broccolini, Aldobrando; Gidaro, Teresa; Morosetti, Roberta

2010-06-26

Over the last years, the existence of different stem cells with myogenic potential has been widely investigated. Besides the classical skeletal muscle progenitors represented by satellite cells, numerous multipotent and embryologically unrelated progenitors with a potential role in muscle differentiation and repair have been identified. In order to conceive a therapeutic approach for degenerative muscle disorders, it is of primary importance to identify an ideal stem cell endowed with all the features for a possible use in vivo. Among all emerging populations, vessel-associated stem cells are a novel and promising class of multipotent progenitors of mesodermal origin and with high myogenic potential which seem to best fit all the requirements for a possible cell therapy. In vitro and in vivostudies have already tested the effectiveness and safety of vessel-associated stem cells in animal models. This leads to the concrete possibility in the future to start pilot human clinical trials, hopefully opening the way to a turning point in the treatment of genetic and acquired muscle disorders.

The extraocular muscle stem cell niche is resistant to ageing and disease

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Luigi eFormicola

2014-12-01

Full Text Available Specific muscles are spared in many degenerative myopathies. Most notably, the extraocular muscles (EOMs do not show clinical signs of late stage myopathies including the accumulation of fibrosis and fat. It has been proposed that an altered stem cell niche underlies the resistance of EOMs in these pathologies, however, to date, no reports have provided a detailed characterization of the EOM stem cell niche. PW1/Peg3 is expressed in progenitor cells in all adult tissues including satellite cells and a subset of interstitial non-satellite cell progenitors in muscle. These PW1-positive interstitial cells (PICs include a fibroadipogenic progenitor population (FAPs that give rise to fat and fibrosis in late stage myopathies. PICs/FAPs are mobilized following injury and FAPs exert a promyogenic role upon myoblasts in vitro but require the presence of a minimal population of satellite cells in vivo. We and others recently described that FAPs express promyogenic factors while satellite cells express antimyogenic factors suggesting that PICs/FAPs act as support niche cells in skeletal muscle through paracrine interactions. We analyzed the EOM stem cell niche in young adult and aged wild-type mice and found that the balance between PICs and satellite cells within the EOM stem cell niche is maintained throughout life. Moreover, in the adult mdx mouse model for Duchenne muscular dystrophy, the EOM stem cell niche is unperturbed compared to normal mice, in contrast to Tibialis Anterior (TA muscle, which displays signs of ongoing degeneration/regeneration. Regenerating mdx TA shows increased levels of both PICs and satellite cells, comparable to normal unaffected EOMs. We propose that the increase in PICs that we observe in normal EOMs contributes to preserving the integrity of the myofibers and satellite cells. Our data suggest that molecular cues regulating muscle regeneration are intrinsic properties of EOMs.

PPARγ and MyoD are differentially regulated by myostatin in adipose-derived stem cells and muscle satellite cells

International Nuclear Information System (INIS)

Zhang, Feng; Deng, Bing; Wen, Jianghui; Chen, Kun; Liu, Wu; Ye, Shengqiang; Huang, Haijun; Jiang, Siwen; Xiong, Yuanzhu

2015-01-01

Myostatin (MSTN) is a secreted protein belonging to the transforming growth factor-β (TGF-β) family that is primarily expressed in skeletal muscle and also functions in adipocyte maturation. Studies have shown that MSTN can inhibit adipogenesis in muscle satellite cells (MSCs) but not in adipose-derived stem cells (ADSCs). However, the mechanism by which MSTN differently regulates adipogenesis in these two cell types remains unknown. Peroxisome proliferator-activated receptor-γ (PPARγ) and myogenic differentiation factor (MyoD) are two key transcription factors in fat and muscle cell development that influence adipogenesis. To investigate whether MSTN differentially regulates PPARγ and MyoD, we analyzed PPARγ and MyoD expression by assessing mRNA, protein and methylation levels in ADSCs and MSCs after treatment with 100 ng/mL MSTN for 0, 24, and 48 h. PPARγ mRNA levels were downregulated after 24 h and upregulated after 48 h of treatment in ADSCs, whereas in MSCs, PPARγ levels were downregulated at both time points. MyoD expression was significantly increased in ADSCs and decreased in MSCs. PPARγ and MyoD protein levels were upregulated in ADSCs and downregulated in MSCs. The CpG methylation levels of the PPARγ and MyoD promoters were decreased in ADSCs and increased in MSCs. Therefore, this study demonstrated that the different regulatory adipogenic roles of MSTN in ADSCs and MSCs act by differentially regulating PPARγ and MyoD expression. - Highlights: • PPARγ and MyoD mRNA and protein levels are upregulated by myostatin in ADSCs. • PPARγ and MyoD mRNA and protein levels are downregulated by myostatin in MSCs. • PPARγ exhibited different methylation levels in myostatin-treated ADSCs and MSCs. • MyoD exhibited different methylation levels in myostatin-treated ADSCs and MSCs. • PPARγ and MyoD are differentially regulated by myostatin in ADSCs and MSCs

PPARγ and MyoD are differentially regulated by myostatin in adipose-derived stem cells and muscle satellite cells

Energy Technology Data Exchange (ETDEWEB)

Zhang, Feng [Key Laboratory of Swine Genetics and Breeding of the Agricultural Ministry and Key Laboratory of Agricultural Animal Genetics, Breeding and Reproduction of the Ministry of Education, College of Animal Science and Technology, Huazhong Agricultural University, Wuhan, 430070 (China); Deng, Bing [Wuhan Institute of Animal Science and Veterinary Medicine, Wuhan Academy of Agricultural Science and Technology, Wuhan, Hubei, 430208 (China); Wen, Jianghui [Wu Han University of Technology, Wuhan 430074 (China); Chen, Kun [Key Laboratory of Swine Genetics and Breeding of the Agricultural Ministry and Key Laboratory of Agricultural Animal Genetics, Breeding and Reproduction of the Ministry of Education, College of Animal Science and Technology, Huazhong Agricultural University, Wuhan, 430070 (China); Liu, Wu; Ye, Shengqiang; Huang, Haijun [Wuhan Institute of Animal Science and Veterinary Medicine, Wuhan Academy of Agricultural Science and Technology, Wuhan, Hubei, 430208 (China); Jiang, Siwen, E-mail: jiangsiwen@mail.hzau.edu.cn [Key Laboratory of Swine Genetics and Breeding of the Agricultural Ministry and Key Laboratory of Agricultural Animal Genetics, Breeding and Reproduction of the Ministry of Education, College of Animal Science and Technology, Huazhong Agricultural University, Wuhan, 430070 (China); Xiong, Yuanzhu, E-mail: xiongyzhu@163.com [Key Laboratory of Swine Genetics and Breeding of the Agricultural Ministry and Key Laboratory of Agricultural Animal Genetics, Breeding and Reproduction of the Ministry of Education, College of Animal Science and Technology, Huazhong Agricultural University, Wuhan, 430070 (China)

2015-03-06

Myostatin (MSTN) is a secreted protein belonging to the transforming growth factor-β (TGF-β) family that is primarily expressed in skeletal muscle and also functions in adipocyte maturation. Studies have shown that MSTN can inhibit adipogenesis in muscle satellite cells (MSCs) but not in adipose-derived stem cells (ADSCs). However, the mechanism by which MSTN differently regulates adipogenesis in these two cell types remains unknown. Peroxisome proliferator-activated receptor-γ (PPARγ) and myogenic differentiation factor (MyoD) are two key transcription factors in fat and muscle cell development that influence adipogenesis. To investigate whether MSTN differentially regulates PPARγ and MyoD, we analyzed PPARγ and MyoD expression by assessing mRNA, protein and methylation levels in ADSCs and MSCs after treatment with 100 ng/mL MSTN for 0, 24, and 48 h. PPARγ mRNA levels were downregulated after 24 h and upregulated after 48 h of treatment in ADSCs, whereas in MSCs, PPARγ levels were downregulated at both time points. MyoD expression was significantly increased in ADSCs and decreased in MSCs. PPARγ and MyoD protein levels were upregulated in ADSCs and downregulated in MSCs. The CpG methylation levels of the PPARγ and MyoD promoters were decreased in ADSCs and increased in MSCs. Therefore, this study demonstrated that the different regulatory adipogenic roles of MSTN in ADSCs and MSCs act by differentially regulating PPARγ and MyoD expression. - Highlights: • PPARγ and MyoD mRNA and protein levels are upregulated by myostatin in ADSCs. • PPARγ and MyoD mRNA and protein levels are downregulated by myostatin in MSCs. • PPARγ exhibited different methylation levels in myostatin-treated ADSCs and MSCs. • MyoD exhibited different methylation levels in myostatin-treated ADSCs and MSCs. • PPARγ and MyoD are differentially regulated by myostatin in ADSCs and MSCs.

Skeletal muscle aging: stem cell function and tissue homeostasis

OpenAIRE

Victor, Pedro Sousa

2012-01-01

Muscle aging, in particular, is characterized by the reduction of tissue mass and function, which are particularly prominent in geriatric individuals undergoing sarcopenia. The age-associated muscle wasting is also associated with a decline in regenerative ability and a reduction in resident muscle stem cell (satellite cell) number and function. Although sarcopenia is one of the major contributors to the general loss of physiological function, the mechanisms involved in age-related loss of mu...

Skin appendage-derived stem cells: cell biology and potential for wound repair.

Science.gov (United States)

Xie, Jiangfan; Yao, Bin; Han, Yutong; Huang, Sha; Fu, Xiaobing

2016-01-01

Stem cells residing in the epidermis and skin appendages are imperative for skin homeostasis and regeneration. These stem cells also participate in the repair of the epidermis after injuries, inducing restoration of tissue integrity and function of damaged tissue. Unlike epidermis-derived stem cells, comprehensive knowledge about skin appendage-derived stem cells remains limited. In this review, we summarize the current knowledge of skin appendage-derived stem cells, including their fundamental characteristics, their preferentially expressed biomarkers, and their potential contribution involved in wound repair. Finally, we will also discuss current strategies, future applications, and limitations of these stem cells, attempting to provide some perspectives on optimizing the available therapy in cutaneous repair and regeneration.

Adipose-derived stem cells for treatment of chronic ulcers

DEFF Research Database (Denmark)

Holm, Jens Selch; Toyserkani, Navid Mohamadpour; Sorensen, Jens Ahm

2018-01-01

Chronic ulcers remain a difficult challenge in healthcare systems. While treatment options are limited, stem cells may be a novel alternative. Adipose-derived stem cells (ADSC) have become increasingly popular compared with bone marrow-derived stem cells as they are far easier to harvest...

Reconstitution of the complete rupture in musculotendinous junction using skeletal muscle-derived multipotent stem cell sheet-pellets as a “bio-bond”

Directory of Open Access Journals (Sweden)

Hiroyuki Hashimoto

2016-07-01

Full Text Available Background. Significant and/or complete rupture in the musculotendinous junction (MTJ is a challenging lesion to treat because of the lack of reliable suture methods. Skeletal muscle-derived multipotent stem cell (Sk-MSC sheet-pellets, which are able to reconstitute peripheral nerve and muscular/vascular tissues with robust connective tissue networks, have been applied as a “bio-bond”. Methods. Sk-MSC sheet-pellets, derived from GFP transgenic-mice after 7 days of expansion culture, were detached with EDTA to maintain cell–cell connections. A completely ruptured MTJ model was prepared in the right tibialis anterior (TA of the recipient mice, and was covered with sheet-pellets. The left side was preserved as a contralateral control. The control group received the same amount of the cell-free medium. The sheet-pellet transplantation (SP group was further divided into two groups; as the short term (4–8 weeks and long term (14–18 weeks recovery group. At each time point after transplantation, tetanic tension output was measured through the electrical stimulation of the sciatic nerve. The behavior of engrafted GFP+ tissues and cells was analyzed by fluorescence immunohistochemistry. Results. The SP short term recovery group showed average 64% recovery of muscle mass, and 36% recovery of tetanic tension output relative to the contralateral side. Then, the SP long term recovery group showed increased recovery of average muscle mass (77% and tetanic tension output (49%. However, the control group showed no recovery of continuity between muscle and tendon, and demonstrated increased muscle atrophy, with coalescence to the tibia during 4–8 weeks after operation. Histological evidence also supported the above functional recovery of SP group. Engrafted Sk-MSCs primarily formed the connective tissues and muscle fibers, including nerve-vascular networks, and bridged the ruptured tendon–muscle fiber units, with differentiation into skeletal

Catechins activate muscle stem cells by Myf5 induction and stimulate muscle regeneration.

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Kim, A Rum; Kim, Kyung Min; Byun, Mi Ran; Hwang, Jun-Ha; Park, Jung Il; Oh, Ho Taek; Kim, Hyo Kyeong; Jeong, Mi Gyeong; Hwang, Eun Sook; Hong, Jeong-Ho

2017-07-22

Muscle weakness is one of the most common symptoms in aged individuals and increases risk of mortality. Thus, maintenance of muscle mass is important for inhibiting aging. In this study, we investigated the effect of catechins, polyphenol compounds in green tea, on muscle regeneration. We found that (-)-epicatechin gallate (ECG) and (-)-epigallocatechin-3-gallate (EGCG) activate satellite cells by induction of Myf5 transcription factors. For satellite cell activation, Akt kinase was significantly induced after ECG treatment and ECG-induced satellite cell activation was blocked in the presence of Akt inhibitor. ECG also promotes myogenic differentiation through the induction of myogenic markers, including Myogenin and Muscle creatine kinase (MCK), in satellite and C2C12 myoblast cells. Finally, EGCG administration to mice significantly increased muscle fiber size for regeneration. Taken together, the results suggest that catechins stimulate muscle stem cell activation and differentiation for muscle regeneration. Copyright © 2017 Elsevier Inc. All rights reserved.

Neuromuscular junction formation between human stem-cell-derived motoneurons and rat skeletal muscle in a defined system.

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Guo, Xiufang; Das, Mainak; Rumsey, John; Gonzalez, Mercedes; Stancescu, Maria; Hickman, James

2010-12-01

To date, the coculture of motoneurons (MNs) and skeletal muscle in a defined in vitro system has only been described in one study and that was between rat MNs and rat skeletal muscle. No in vitro studies have demonstrated human MN to rat muscle synapse formation, although numerous studies have attempted to implant human stem cells into rat models to determine if they could be of therapeutic use in disease or spinal injury models, although with little evidence of neuromuscular junction (NMJ) formation. In this report, MNs differentiated from human spinal cord stem cells, together with rat skeletal myotubes, were used to build a coculture system to demonstrate that NMJ formation between human MNs and rat skeletal muscles is possible. The culture was characterized by morphology, immunocytochemistry, and electrophysiology, while NMJ formation was demonstrated by immunocytochemistry and videography. This defined system provides a highly controlled reproducible model for studying the formation, regulation, maintenance, and repair of NMJs. The in vitro coculture system developed here will be an important model system to study NMJ development, the physiological and functional mechanism of synaptic transmission, and NMJ- or synapse-related disorders such as amyotrophic lateral sclerosis, as well as for drug screening and therapy design.

Clone-derived human AF-amniotic fluid stem cells are capable of skeletal myogenic differentiation in vitro and in vivo.

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Ma, Xiaorong; Zhang, Shengli; Zhou, Junmei; Chen, Baisong; Shang, Yafeng; Gao, Tongbing; Wang, Xue; Xie, Hua; Chen, Fang

2012-08-01

Stem cell-based therapy may be the most promising method to cure skeletal muscle degenerative diseases such as Duchenne muscular dystrophy (DMD) and trauma in the future. Human amniotic fluid is enriched with early-stage stem cells from developing fetuses and these cells have cardiomyogenic potential both in vitro and in vivo. In the present study, we investigated the characteristics of human amniotic fluid-derived AF-type stem (HAF-AFS) cells by flow cytometry, immunofluorescence staining, reverse-transcription polymerase chain reaction, and osteogenic and adipogenic differentiation analysis. After confirming the stemness of HAF-AFS cells, we tested whether HAF-AFS cells could differentiate into skeletal myogenic cells in vitro and incorporate into regenerating skeletal muscle in vivo. By temporary exposure to the DNA demethylation agent 5-aza-2'-deoxycytidine (5-Aza dC) or co-cultured with C2C12 myoblasts, HAF-AFS cells differentiated into skeletal myogenic cells, expressing skeletal myogenic cell-specific markers such as Desmin, Troponin I (Tn I) and α-Actinin. Four weeks after transplantation into cardiotoxin-injured and X-ray-irradiated tibialis anterior (TA) muscles of NOD/SCID mice, HAF-AFS cells survived, differentiated into myogenic precursor cells and fused with host myofibres. The findings that HAF-AFS cells differentiate into myogenic cells in vitro and incorporate in skeletal muscle regeneration in vivo hold the promise of HAF-AFS cell-based therapy for skeletal muscle degenerative diseases. Copyright © 2012 John Wiley & Sons, Ltd.

TAK1 modulates satellite stem cell homeostasis and skeletal muscle repair

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Ogura, Yuji; Hindi, Sajedah M.; Sato, Shuichi; Xiong, Guangyan; Akira, Shizuo; Kumar, Ashok

2015-01-01

Satellite cells are resident adult stem cells that are required for regeneration of skeletal muscle. However, signalling mechanisms that regulate satellite cell function are less understood. Here we demonstrate that transforming growth factor-β-activated kinase 1 (TAK1) is important in satellite stem cell homeostasis and function. Inactivation of TAK1 in satellite cells inhibits muscle regeneration in adult mice. TAK1 is essential for satellite cell proliferation and its inactivation causes precocious differentiation. Moreover, TAK1-deficient satellite cells exhibit increased oxidative stress and undergo spontaneous cell death, primarily through necroptosis. TAK1 is required for the activation of NF-κB and JNK in satellite cells. Forced activation of NF-κB improves survival and proliferation of TAK1-deficient satellite cells. Furthermore, TAK1-mediated activation of JNK is essential to prevent oxidative stress and precocious differentiation of satellite cells. Collectively, our study suggests that TAK1 is required for maintaining the pool of satellite stem cells and for regenerative myogenesis. PMID:26648529

Derivation of Skeletal Myogenic Precursors from Human Pluripotent Stem Cells Using Conditional Expression of PAX7.

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Darabi, Radbod; Perlingeiro, Rita C R

2016-01-01

Cell-based therapies are considered as one of the most promising approaches for the treatment of degenerating pathologies including muscle disorders and dystrophies. Advances in the approach of reprogramming somatic cells into induced pluripotent stem (iPS) cells allow for the possibility of using the patient's own pluripotent cells to generate specific tissues for autologous transplantation. In addition, patient-specific tissue derivatives have been shown to represent valuable material for disease modeling and drug discovery. Nevertheless, directed differentiation of pluripotent stem cells into a specific lineage is not a trivial task especially in the case of skeletal myogenesis, which is generally poorly recapitulated during the in vitro differentiation of pluripotent stem cells.Here, we describe a practical and efficient method for the derivation of skeletal myogenic precursors from differentiating human pluripotent stem cells using controlled expression of PAX7. Flow cytometry (FACS) purified myogenic precursors can be expanded exponentially and differentiated in vitro into myotubes, enabling researchers to use these cells for disease modeling as well as therapeutic purposes.

The Potential for Synovium-derived Stem Cells in Cartilage Repair

DEFF Research Database (Denmark)

Kubosch, Eva Johanna; Lang, Gernot Michael; Fürst, David

2018-01-01

for the treatment of large, isolated, full thickness cartilage defects. Several disadvantages such as the need for two surgical procedures or hypertrophic regenerative cartilage, underline the need for alternative cell sources. OBJECTIVE: Mesenchymal stem cells, particularly synovium-derived mesenchymal stem cells......, represent a promising cell source. Synovium-derived mesenchymal stem cells have attracted considerable attention since they display great chondrogenic potential and less hypertrophic differentiation than mesenchymal stem cells derived from bone marrow. The aim of this review was to summarize the current...... knowledge on the chondrogenic potential for synovial stem cells in regard to cartilage repair purposes. RESULTS: A literature search was carried out identifying 260 articles in the databases up to January 2017. Several in vitro and initial animal in vivo studies of cartilage repair using synovia stem cell...

The Use Of Laser Irradiation To Stimulate Adipose Derived Stem Cell Proliferation And Differentiation For Use In Autologous Grafts

Science.gov (United States)

Abrahamse, Heidi

2009-09-01

Stem cells are characterized by the qualities of self-renewal, long term viability, and the ability to differentiate into various cell types. Historically, stem cells have been isolated from the inner cell mass of blastocysts and harvesting these cells resulted in the death of the embryo leading to religious, political and ethical issues. The identification and subsequent isolation of adult stem cells from bone marrow stroma have been welcomed as an alternate source for stem cells. The clinical use of Mesenchymal Stem Cells (MSCs) presented problems such as limited cell number, pain and morbidity upon isolation. Adipose tissue is derived from the mesenchyme, is easily isolated, a reliable source of stem cells and able to differentiate into different cell types including smooth muscle. Over the past few years, the identification and characterization of stem cells has led the potential use of these cells as a promising alternative to cell replacement therapy. Smooth muscle is a major component of human tissues and is essential for the normal functioning of many different organs. Low intensity laser irradiation has been shown to increase viability, protein expression and migration of stem cells in vitro, and to stimulate proliferation of various types of stem cells. In addition, the use of laser irradiation to stimulate differentiation in the absence of growth factors has also been demonstrated in normal human neural progenitor cells (NHNPCs) in vitro where NHNPCs are not only capable of being sustained by light in the absence of growth factors, but that they are also able to differentiate normally as assessed by neurite formation. Our work has focused on the ability of laser irradiation to proliferate adipose derived stem cells (ADSCs), maintain ADSC character and increase the rate and maintenance of differentiation of ADSCs into smooth muscle and skin fibroblast cells. Current studies are also investigating the effect of different irradiation wavelengths and

Umbilical Cord-Derived Mesenchymal Stem Cells for Hematopoietic Stem Cell Transplantation

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Yu-Hua Chao

2012-01-01

Full Text Available Hematopoietic stem cell transplantation (HSCT is becoming an effective therapeutic modality for a variety of diseases. Mesenchymal stem cells (MSCs can be used to enhance hematopoietic engraftment, accelerate lymphocyte recovery, reduce the risk of graft failure, prevent and treat graft-versus-host disease, and repair tissue damage in patients receiving HSCT. Till now, most MSCs for human clinical application have been derived from bone marrow. However, acquiring bone-marrow-derived MSCs involves an invasive procedure. Umbilical cord is rich with MSCs. Compared to bone-marrow-derived MSCs, umbilical cord-derived MSCs (UCMSCs are easier to obtain without harm to the donor and can proliferate faster. No severe adverse effects were noted in our previous clinical application of UCMSCs in HSCT. Accordingly, application of UCMSCs in humans appears to be feasible and safe. Further studies are warranted.

Mesenchymal Stem Cell-Like Cells Derived from Mouse Induced Pluripotent Stem Cells Ameliorate Diabetic Polyneuropathy in Mice

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Tatsuhito Himeno

2013-01-01

Full Text Available Background. Although pathological involvements of diabetic polyneuropathy (DPN have been reported, no dependable treatment of DPN has been achieved. Recent studies have shown that mesenchymal stem cells (MSCs ameliorate DPN. Here we demonstrate a differentiation of induced pluripotent stem cells (iPSCs into MSC-like cells and investigate the therapeutic potential of the MSC-like cell transplantation on DPN. Research Design and Methods. For induction into MSC-like cells, GFP-expressing iPSCs were cultured with retinoic acid, followed by adherent culture for 4 months. The MSC-like cells, characterized with flow cytometry and RT-PCR analyses, were transplanted into muscles of streptozotocin-diabetic mice. Three weeks after the transplantation, neurophysiological functions were evaluated. Results. The MSC-like cells expressed MSC markers and angiogenic/neurotrophic factors. The transplanted cells resided in hindlimb muscles and peripheral nerves, and some transplanted cells expressed S100β in the nerves. Impairments of current perception thresholds, nerve conduction velocities, and plantar skin blood flow in the diabetic mice were ameliorated in limbs with the transplanted cells. The capillary number-to-muscle fiber ratios were increased in transplanted hindlimbs of diabetic mice. Conclusions. These results suggest that MSC-like cell transplantation might have therapeutic effects on DPN through secreting angiogenic/neurotrophic factors and differentiation to Schwann cell-like cells.

Pathologic bladder microenvironment attenuates smooth muscle differentiation of skin derived precursor cells: implications for tissue regeneration.

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Cornelia Tolg

Full Text Available Smooth muscle cell containing organs (bladder, heart, blood vessels are damaged by a variety of pathological conditions necessitating surgery or organ replacement. Currently, regeneration of contractile tissues is hampered by lack of functional smooth muscle cells. Multipotent skin derived progenitor cells (SKPs can easily be isolated from adult skin and can be differentiated in vitro into contractile smooth muscle cells by exposure to FBS. Here we demonstrate an inhibitory effect of a pathologic contractile organ microenvironment on smooth muscle cell differentiation of SKPs. In vivo, urinary bladder strain induces microenvironmental changes leading to de-differentiation of fully differentiated bladder smooth muscle cells. Co-culture of SKPs with organoids isolated from ex vivo stretched bladders or exposure of SKPs to diffusible factors released by stretched bladders (e.g. bFGF suppresses expression of smooth muscle markers (alpha SMactin, calponin, myocardin, myosin heavy chain as demonstrated by qPCR and immunofluorescent staining. Rapamycin, an inhibitor of mTOR signalling, previously observed to prevent bladder strain induced de-differentiation of fully differentiated smooth muscle cells in vitro, inhibits FBS-induced smooth muscle cell differentiation of undifferentiated SKPs. These results suggest that intended precursor cell differentiation may be paradoxically suppressed by the disease context for which regeneration may be required. Organ-specific microenvironment contexts, particularly prevailing disease, may play a significant role in modulating or attenuating an intended stem cell phenotypic fate, possibly explaining the variable and inefficient differentiation of stem cell constructs in in vivo settings. These observations must be considered in drafting any regeneration strategies.

Muscle Tissue Engineering Using Gingival Mesenchymal Stem Cells Encapsulated in Alginate Hydrogels Containing Multiple Growth Factors.

Science.gov (United States)

Ansari, Sahar; Chen, Chider; Xu, Xingtian; Annabi, Nasim; Zadeh, Homayoun H; Wu, Benjamin M; Khademhosseini, Ali; Shi, Songtao; Moshaverinia, Alireza

2016-06-01

Repair and regeneration of muscle tissue following traumatic injuries or muscle diseases often presents a challenging clinical situation. If a significant amount of tissue is lost the native regenerative potential of skeletal muscle will not be able to grow to fill the defect site completely. Dental-derived mesenchymal stem cells (MSCs) in combination with appropriate scaffold material, present an advantageous alternative therapeutic option for muscle tissue engineering in comparison to current treatment modalities available. To date, there has been no report on application of gingival mesenchymal stem cells (GMSCs) in three-dimensional scaffolds for muscle tissue engineering. The objectives of the current study were to develop an injectable 3D RGD-coupled alginate scaffold with multiple growth factor delivery capacity for encapsulating GMSCs, and to evaluate the capacity of encapsulated GMSCs to differentiate into myogenic tissue in vitro and in vivo where encapsulated GMSCs were transplanted subcutaneously into immunocompromised mice. The results demonstrate that after 4 weeks of differentiation in vitro, GMSCs as well as the positive control human bone marrow mesenchymal stem cells (hBMMSCs) exhibited muscle cell-like morphology with high levels of mRNA expression for gene markers related to muscle regeneration (MyoD, Myf5, and MyoG) via qPCR measurement. Our quantitative PCR analyzes revealed that the stiffness of the RGD-coupled alginate regulates the myogenic differentiation of encapsulated GMSCs. Histological and immunohistochemical/fluorescence staining for protein markers specific for myogenic tissue confirmed muscle regeneration in subcutaneous transplantation in our in vivo animal model. GMSCs showed significantly greater capacity for myogenic regeneration in comparison to hBMMSCs (p alginate hydrogel with multiple growth factor delivery capacity is a promising candidate for muscle tissue engineering.

A novel paradigm links mitochondrial dysfunction with muscle stem cell impairment in sepsis.

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Chatre, Laurent; Verdonk, Franck; Rocheteau, Pierre; Crochemore, Clément; Chrétien, Fabrice; Ricchetti, Miria

2017-10-01

Sepsis is an acute systemic inflammatory response of the body to microbial infection and a life threatening condition associated with multiple organ failure. Survivors may display long-term disability with muscle weakness that remains poorly understood. Recent data suggest that long-term myopathy in sepsis survivors is due to failure of skeletal muscle stem cells (satellite cells) to regenerate the muscle. Satellite cells impairment in the acute phase of sepsis is linked to unusual mitochondrial dysfunctions, characterized by a dramatic reduction of the mitochondrial mass and hyperactivity of residual organelles. Survivors maintain the impairment of satellite cells, including alterations of the mitochondrial DNA (mtDNA), in the long-term. This condition can be rescued by treatment with mesenchymal stem cells (MSCs) that restore mtDNA alterations and mitochondrial function in satellite cells, and in fine their regenerative potential. Injection of MSCs in turn increases the force of isolated muscle fibers and of the whole animal, and improves the survival rate. These effects occur in the context of reduced inflammation markers that also raised during sepsis. Targeting muscle stem cells mitochondria, in a context of reduced inflammation, may represent a valuable strategy to reduce morbidity and long-term impairment of the muscle upon sepsis. Copyright © 2017 Elsevier B.V. All rights reserved.

Balancing Ethical Pros and Cons of Stem Cell Derived Gametes.

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Segers, Seppe; Mertes, Heidi; de Wert, Guido; Dondorp, Wybo; Pennings, Guido

2017-07-01

In this review we aim to provide an overview of the most important ethical pros and cons of stem cell derived gametes (SCD-gametes), as a contribution to the debate about reproductive tissue engineering. Derivation of gametes from stem cells holds promising applications both for research and for clinical use in assisted reproduction. We explore the ethical issues connected to gametes derived from embryonic stem cells (both patient specific and non-patient specific) as well as those related to gametes derived from induced pluripotent stem cells. The technology of SCD-gametes raises moral concerns of how reproductive autonomy relates to issues of embryo destruction, safety, access, and applications beyond clinical infertility.

Stem cells in bone tissue engineering

Energy Technology Data Exchange (ETDEWEB)

Seong, Jeong Min [Department of Preventive and Social Dentistry and Institute of Oral Biology, College of Dentistry, Kyung Hee University, Seoul 130-701 (Korea, Republic of); Kim, Byung-Chul; Park, Jae-Hong; Kwon, Il Keun; Hwang, Yu-Shik [Department of Maxillofacial Biomedical Engineering and Institute of Oral Biology, College of Dentistry, Kyung Hee University, Seoul 130-701 (Korea, Republic of); Mantalaris, Anathathios, E-mail: yshwang@khu.ac.k [Department of Chemical Engineering, Imperial College London, South Kensington Campus, London SW7 2AZ (United Kingdom)

2010-12-15

Bone tissue engineering has been one of the most promising areas of research, providing a potential clinical application to cure bone defects. Recently, various stem cells including embryonic stem cells (ESCs), bone marrow-derived mesenchymal stem cells (BM-MSCs), umbilical cord blood-derived mesenchymal stem cells (UCB-MSCs), adipose tissue-derived stem cells (ADSCs), muscle-derived stem cells (MDSCs) and dental pulp stem cells (DPSCs) have received extensive attention in the field of bone tissue engineering due to their distinct biological capability to differentiate into osteogenic lineages. The application of these stem cells to bone tissue engineering requires inducing in vitro differentiation of these cells into bone forming cells, osteoblasts. For this purpose, efficient in vitro differentiation towards osteogenic lineage requires the development of well-defined and proficient protocols. This would reduce the likelihood of spontaneous differentiation into divergent lineages and increase the available cell source for application to bone tissue engineering therapies. This review provides a critical examination of the various experimental strategies that could be used to direct the differentiation of ESC, BM-MSC, UCB-MSC, ADSC, MDSC and DPSC towards osteogenic lineages and their potential applications in tissue engineering, particularly in the regeneration of bone. (topical review)

Stem cells in bone tissue engineering

International Nuclear Information System (INIS)

Seong, Jeong Min; Kim, Byung-Chul; Park, Jae-Hong; Kwon, Il Keun; Hwang, Yu-Shik; Mantalaris, Anathathios

2010-01-01

Bone tissue engineering has been one of the most promising areas of research, providing a potential clinical application to cure bone defects. Recently, various stem cells including embryonic stem cells (ESCs), bone marrow-derived mesenchymal stem cells (BM-MSCs), umbilical cord blood-derived mesenchymal stem cells (UCB-MSCs), adipose tissue-derived stem cells (ADSCs), muscle-derived stem cells (MDSCs) and dental pulp stem cells (DPSCs) have received extensive attention in the field of bone tissue engineering due to their distinct biological capability to differentiate into osteogenic lineages. The application of these stem cells to bone tissue engineering requires inducing in vitro differentiation of these cells into bone forming cells, osteoblasts. For this purpose, efficient in vitro differentiation towards osteogenic lineage requires the development of well-defined and proficient protocols. This would reduce the likelihood of spontaneous differentiation into divergent lineages and increase the available cell source for application to bone tissue engineering therapies. This review provides a critical examination of the various experimental strategies that could be used to direct the differentiation of ESC, BM-MSC, UCB-MSC, ADSC, MDSC and DPSC towards osteogenic lineages and their potential applications in tissue engineering, particularly in the regeneration of bone. (topical review)

Stem Cell-Derived Exosome in Cardiovascular Diseases: Macro Roles of Micro Particles.

Science.gov (United States)

Yuan, Ye; Du, Weijie; Liu, Jiaqi; Ma, Wenya; Zhang, Lai; Du, Zhimin; Cai, Benzhi

2018-01-01

The stem cell-based therapy has emerged as the promising therapeutic strategies for cardiovascular diseases (CVDs). Recently, increasing evidence suggest stem cell-derived active exosomes are important communicators among cells in the heart via delivering specific substances to the adjacent/distant target cells. These exosomes and their contents such as certain proteins, miRNAs and lncRNAs exhibit huge beneficial effects on preventing heart damage and promoting cardiac repair. More importantly, stem cell-derived exosomes are more effective and safer than stem cell transplantation. Therefore, administration of stem cell-derived exosomes will expectantly be an alternative stem cell-based therapy for the treatment of CVDs. Furthermore, modification of stem cell-derived exosomes or artificial synthesis of exosomes will be the new therapeutic tools for CVDs in the future. In addition, stem cell-derived exosomes also have been implicated in the diagnosis and prognosis of CVDs. In this review, we summarize the current advances of stem cell-derived exosome-based treatment and prognosis for CVDs, including their potential benefits, underlying mechanisms and limitations, which will provide novel insights of exosomes as a new tool in clinical therapeutic translation in the future.

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

International Nuclear Information System (INIS)

Waksman, Ron; Baffour, Richard

2003-01-01

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

Plasticity of the Muscle Stem Cell Microenvironment.

Science.gov (United States)

Dinulovic, Ivana; Furrer, Regula; Handschin, Christoph

2017-01-01

Satellite cells (SCs) are adult muscle stem cells capable of repairing damaged and creating new muscle tissue throughout life. Their functionality is tightly controlled by a microenvironment composed of a wide variety of factors, such as numerous secreted molecules and different cell types, including blood vessels, oxygen, hormones, motor neurons, immune cells, cytokines, fibroblasts, growth factors, myofibers, myofiber metabolism, the extracellular matrix and tissue stiffness. This complex niche controls SC biology-quiescence, activation, proliferation, differentiation or renewal and return to quiescence. In this review, we attempt to give a brief overview of the most important players in the niche and their mutual interaction with SCs. We address the importance of the niche to SC behavior under physiological and pathological conditions, and finally survey the significance of an artificial niche both for basic and translational research purposes.

Cardiotoxicity evaluation using human embryonic stem cells and induced pluripotent stem cell-derived cardiomyocytes.

Science.gov (United States)

Zhao, Qi; Wang, Xijie; Wang, Shuyan; Song, Zheng; Wang, Jiaxian; Ma, Jing

2017-03-09

Cardiotoxicity remains an important concern in drug discovery. Human pluripotent stem cell-derived cardiomyocytes (hPSC-CMs) have become an attractive platform to evaluate cardiotoxicity. However, the consistency between human embryonic stem cell-derived cardiomyocytes (hESC-CMs) and human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) in prediction of cardiotoxicity has yet to be elucidated. Here we screened the toxicities of four representative drugs (E-4031, isoprenaline, quinidine, and haloperidol) using both hESC-CMs and hiPSC-CMs, combined with an impedance-based bioanalytical method. It showed that both hESC-CMs and hiPSC-CMs can recapitulate cardiotoxicity and identify the effects of well-characterized compounds. The combined platform of hPSC-CMs and an impedance-based bioanalytical method could improve preclinical cardiotoxicity screening, holding great potential for increasing drug development accuracy.

Isolation and expansion of adipose-derived stem cells for tissue engineering

DEFF Research Database (Denmark)

Fink, Trine; Rasmussen, Jeppe Grøndahl; Lund, Pia

2011-01-01

For treatment of cardiac failure with bone marrow-derived mesenchymal stem cells, several clinical trials are ongoing. However, more attention is gathering on the use of adipose tissue-derived stem cells (ASCs). This paper describes the optimization of isolation and propagation of ASCs for subseq......For treatment of cardiac failure with bone marrow-derived mesenchymal stem cells, several clinical trials are ongoing. However, more attention is gathering on the use of adipose tissue-derived stem cells (ASCs). This paper describes the optimization of isolation and propagation of ASCs...

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

International Nuclear Information System (INIS)

Alexanian, Arshak R.

2005-01-01

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 β-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

Determination of osteogenic or adipogenic lineages in muscle-derived stem cells (MDSCs) by a collagen-binding peptide (CBP) derived from bone sialoprotein (BSP)

Energy Technology Data Exchange (ETDEWEB)

Choi, Yoon Jung [Dental Regenerative Biotechnology Major, School of Dentistry and Dental Research Institute, Seoul National University, Seoul 110-749 (Korea, Republic of); Lee, Jue Yeon [Research Institute, Nano Intelligent Biomedical Engineering Corporation (NIBEC), Seoul (Korea, Republic of); Lee, Seung Jin [Department of Industrial Pharmacy, College of Pharmacy, Ewha Womans University, Seoul (Korea, Republic of); Chung, Chong-Pyoung, E-mail: ccpperio@snu.ac.kr [Research Institute, Nano Intelligent Biomedical Engineering Corporation (NIBEC), Seoul (Korea, Republic of); Department of Periodontology, School of Dentistry and Dental Research Institute, Seoul National University, Seoul (Korea, Republic of); Park, Yoon Jeong, E-mail: parkyj@snu.ac.kr [Dental Regenerative Biotechnology Major, School of Dentistry and Dental Research Institute, Seoul National University, Seoul 110-749 (Korea, Republic of); Research Institute, Nano Intelligent Biomedical Engineering Corporation (NIBEC), Seoul (Korea, Republic of)

2012-03-09

Highlights: Black-Right-Pointing-Pointer CBP sequence is identified from BSP and has collagen binding activity. Black-Right-Pointing-Pointer CBP directly activates the MAPK signaling, especially ERK1/2. Black-Right-Pointing-Pointer CBP increase osteoblastic differentiation by the activation of Runx2. Black-Right-Pointing-Pointer CBP decrease adipogenic differentiation by the inhibition of PPAR{gamma}. -- Abstract: Bone sialoprotein (BSP) is a mineralized, tissue-specific, non-collagenous protein that is normally expressed only in mineralized tissues such as bone, dentin, cementum, and calcified cartilage, and at sites of new mineral formation. The binding of BSP to collagen is thought to be important for initiating bone mineralization and bone cell adhesion to the mineralized matrix. Several recent studies have isolated stem cells from muscle tissue, but their functional properties are still unclear. In this study, we examined the effects of a synthetic collagen-binding peptide (CBP) on the differentiation efficiency of muscle-derived stem cells (MDSCs). The CBP sequence (NGVFKYRPRYYLYKHAYFYPHLKRFPVQ) corresponds to residues 35-62 of bone sialoprotein (BSP), which are located within the collagen-binding domain in BSP. Interestingly, this synthetic CBP inhibited adipogenic differentiation but increased osteogenic differentiation in MDSCs. The CBP also induced expression of osteoblastic marker proteins, including alkaline phosphatase (ALP), type I collagen, Runt-related transcription factor 2 (Runx2), and osteocalcin; prevented adipogenic differentiation in MDSCs; and down-regulated adipose-specific mRNAs, such as adipocyte protein 2 (aP2) and peroxisome proliferator-activated receptor {gamma}. The CBP increased Extracellular signal-regulated kinases (ERK) 1/2 protein phosphorylation, which is important in lineage determination. These observations suggest that this CBP determines the osteogenic or adipogenic lineage in MDSCs by activating ERK1/2. Taken together, a

Efficient derivation of multipotent neural stem/progenitor cells from non-human primate embryonic stem cells.

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Hiroko Shimada

Full Text Available The common marmoset (Callithrix jacchus is a small New World primate that has been used as a non-human primate model for various biomedical studies. We previously demonstrated that transplantation of neural stem/progenitor cells (NS/PCs derived from mouse and human embryonic stem cells (ESCs and induced pluripotent stem cells (iPSCs promote functional locomotor recovery of mouse spinal cord injury models. However, for the clinical application of such a therapeutic approach, we need to evaluate the efficacy and safety of pluripotent stem cell-derived NS/PCs not only by xenotransplantation, but also allotransplantation using non-human primate models to assess immunological rejection and tumorigenicity. In the present study, we established a culture method to efficiently derive NS/PCs as neurospheres from common marmoset ESCs. Marmoset ESC-derived neurospheres could be passaged repeatedly and showed sequential generation of neurons and astrocytes, similar to that of mouse ESC-derived NS/PCs, and gave rise to functional neurons as indicated by calcium imaging. Although marmoset ESC-derived NS/PCs could not differentiate into oligodendrocytes under default culture conditions, these cells could abundantly generate oligodendrocytes by incorporating additional signals that recapitulate in vivo neural development. Moreover, principal component analysis of microarray data demonstrated that marmoset ESC-derived NS/PCs acquired similar gene expression profiles to those of fetal brain-derived NS/PCs by repeated passaging. Therefore, marmoset ESC-derived NS/PCs may be useful not only for accurate evaluation by allotransplantation of NS/PCs into non-human primate models, but are also applicable to analysis of iPSCs established from transgenic disease model marmosets.

Induced Pluripotent Stem Cell Derived Mesenchymal Stem Cells for Attenuating Age-Related Bone Loss

Science.gov (United States)

2012-07-01

Mesenchymal stem cell (MSC) differentiation towards the bone forming osteoblastic lineage decreases as a function of age and may contribute to age-related...problem of age-related reduced availability of MSC we propose to examine the bone anabolic potential of induced pluripotent stem cell (iPS) derived MSC

In Vitro Derivation and Propagation of Spermatogonial Stem Cell Activity from Mouse Pluripotent Stem Cells.

Science.gov (United States)

Ishikura, Yukiko; Yabuta, Yukihiro; Ohta, Hiroshi; Hayashi, Katsuhiko; Nakamura, Tomonori; Okamoto, Ikuhiro; Yamamoto, Takuya; Kurimoto, Kazuki; Shirane, Kenjiro; Sasaki, Hiroyuki; Saitou, Mitinori

2016-12-06

The in vitro derivation and propagation of spermatogonial stem cells (SSCs) from pluripotent stem cells (PSCs) is a key goal in reproductive science. We show here that when aggregated with embryonic testicular somatic cells (reconstituted testes), primordial germ cell-like cells (PGCLCs) induced from mouse embryonic stem cells differentiate into spermatogonia-like cells in vitro and are expandable as cells that resemble germline stem cells (GSCs), a primary cell line with SSC activity. Remarkably, GSC-like cells (GSCLCs), but not PGCLCs, colonize adult testes and, albeit less effectively than GSCs, contribute to spermatogenesis and fertile offspring. Whole-genome analyses reveal that GSCLCs exhibit aberrant methylation at vulnerable regulatory elements, including those critical for spermatogenesis, which may restrain their spermatogenic potential. Our study establishes a strategy for the in vitro derivation of SSC activity from PSCs, which, we propose, relies on faithful epigenomic regulation. Copyright © 2016 The Author(s). Published by Elsevier Inc. All rights reserved.

In Situ Immunofluorescent Staining of Autophagy in Muscle Stem Cells

KAUST Repository

Castagnetti, Francesco

2017-06-13

Increasing evidence points to autophagy as a crucial regulatory process to preserve tissue homeostasis. It is known that autophagy is involved in skeletal muscle development and regeneration, and the autophagic process has been described in several muscular pathologies and agerelated muscle disorders. A recently described block of the autophagic process that correlates with the functional exhaustion of satellite cells during muscle repair supports the notion that active autophagy is coupled with productive muscle regeneration. These data uncover the crucial role of autophagy in satellite cell activation during muscle regeneration in both normal and pathological conditions, such as muscular dystrophies. Here, we provide a protocol to monitor the autophagic process in the adult Muscle Stem Cell (MuSC) compartment during muscle regenerative conditions. This protocol describes the setup methodology to perform in situ immunofluorescence imaging of LC3, an autophagy marker, and MyoD, a myogenic lineage marker, in muscle tissue sections from control and injured mice. The methodology reported allows for monitoring the autophagic process in one specific cell compartment, the MuSC compartment, which plays a central role in orchestrating muscle regeneration.

Adipose stem cells for bone tissue repair

OpenAIRE

Ciuffi, Simone; Zonefrati, Roberto; Brandi, Maria Luisa

2017-01-01

Adipose-derived stem/stromal cells (ASCs), together with adipocytes, vascular endothelial cells, and vascular smooth muscle cells, are contained in fat tissue. ASCs, like the human bone marrow stromal/stem cells (BMSCs), can differentiate into several lineages (adipose cells, fibroblast, chondrocytes, osteoblasts, neuronal cells, endothelial cells, myocytes, and cardiomyocytes). They have also been shown to be immunoprivileged, and genetically stable in long-term cultures. Nevertheless, unlik...

Derivation and characterization of a pig embryonic stem cell-derived exocrine pancreatic cell line

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The establishment and initial characterization of a pig embryonic stem cell-derived pancreatic cell line, PICM-31, and a colony-cloned derivative cell line, PICM-31A, is described. The cell lines were propagated for several months at split ratios of 1:3 or 1:5 at each passage on STO feeder cells af...

Transplantation of Human Dental Pulp-Derived Stem Cells or Differentiated Neuronal Cells from Human Dental Pulp-Derived Stem Cells Identically Enhances Regeneration of the Injured Peripheral Nerve.

Science.gov (United States)

Ullah, Imran; Park, Ju-Mi; Kang, Young-Hoon; Byun, June-Ho; Kim, Dae-Geon; Kim, Joo-Heon; Kang, Dong-Ho; Rho, Gyu-Jin; Park, Bong-Wook

2017-09-01

Human dental mesenchymal stem cells isolated from the dental follicle, pulp, and root apical papilla of extracted wisdom teeth have been known to exhibit successful and potent neurogenic differentiation capacity. In particular, human dental pulp-derived stem cells (hDPSCs) stand out as the most prominent source for in vitro neuronal differentiation. In this study, to evaluate the in vivo peripheral nerve regeneration potential of hDPSCs and differentiated neuronal cells from DPSCs (DF-DPSCs), a total of 1 × 10 6 hDPSCs or DF-hDPSCs labeled with PKH26 tracking dye and supplemented with fibrin glue scaffold and collagen tubulization were transplanted into the sciatic nerve resection (5-mm gap) of rat models. At 12 weeks after cell transplantation, both hDPSC and DF-hDPSC groups showed notably increased behavioral activities and higher muscle contraction forces compared with those in the non-cell transplanted control group. In immunohistochemical analysis of regenerated nerve specimens, specific markers for angiogenesis, axonal fiber, and myelin sheath increased in both the cell transplantation groups. Pretransplanted labeled PKH26 were also distinctly detected in the regenerated nerve tissues, indicating that transplanted cells were well-preserved and differentiated into nerve cells. Furthermore, no difference was observed in the nerve regeneration potential between the hDPSC and DF-hDPSC transplanted groups. These results demonstrate that dental pulp tissue is an excellent stem cell source for nerve regeneration, and in vivo transplantation of the undifferentiated hDPSCs could exhibit sufficient and excellent peripheral nerve regeneration potential.

Transcriptional and functional differences in stem cell populations isolated from Extraocular and Limb muscles

DEFF Research Database (Denmark)

Pacheco-Pinedo, Eugenia Cristina; Budak, Murat T; Zeiger, Ulrike

2008-01-01

The extraocular muscles (EOMs) are a distinct muscle group that displays an array of unique contractile, structural and regenerative properties. They also have differential sensitivity to certain diseases and are enigmatically spared in Duchenne muscular dystrophy (DMD). The EOMs are so distinct...... from other skeletal muscles that the term: allotype has been coined to highlight EOM-group-specific properties. We hypothesized that increased and distinct stem cells may underlie the continual myogenesis noted in EOM. The side population (SP) stem cells were isolated and studied. EOMs had 15x higher...... SP cell content compared to limb muscles. Expression profiling revealed 348 transcripts that define the EOM-SP transcriptome. Over 92% of transcripts were SP-specific, as they were absent in previous whole-muscle microarray studies. Cultured EOM-SP cells revealed superior in vitro proliferative...

Tumorigenicity studies for human pluripotent stem cell-derived products.

Science.gov (United States)

Kuroda, Takuya; Yasuda, Satoshi; Sato, Yoji

2013-01-01

Human pluripotent stem cells (hPSCs), i.e. human embryonic stem cells and human induced pluripotent stem cells, are able to self-renew and differentiate into multiple cell types. Because of these abilities, numerous attempts have been made to utilize hPSCs in regenerative medicine/cell therapy. hPSCs are, however, also tumorigenic, that is, they can give rise to the progressive growth of tumor nodules in immunologically unresponsive animals. Therefore, assessing and managing the tumorigenicity of all final products is essential in order to prevent ectopic tissue formation, tumor development, and/or malignant transformation elicited by residual pluripotent stem cells after implantation. No detailed guideline for the tumorigenicity testing of hPSC-derived products has yet been issued for regenerative medicine/cell therapy, despite the urgent necessity. Here, we describe the current situations and issues related to the tumorigenicity testing of hPSC-derived products and we review the advantages and disadvantages of several types of tumorigenicity-associated tests. We also refer to important considerations in the execution and design of specific studies to monitor the tumorigenicity of hPSC-derived products.

Translational applications of adult stem cell-derived organoids

NARCIS (Netherlands)

Drost, Jarno; Clevers, Hans

2017-01-01

Adult stem cells from a variety of organs can be expanded long-term in vitro as three-dimensional organotypic structures termed organoids. These adult stem cell-derived organoids retain their organ identity and remain genetically stable over long periods of time. The ability to grow organoids from

Role of bone marrow-derived stem cells, renal progenitor cells and ...

African Journals Online (AJOL)

It remains the leading cause of late allograft loss. Bone marrow derived stem cells are undifferentiated cells typically characterized by their capacity for self renewal, ability to give rise to multiple differentiated cellular population, including hematopoietic (HSCs) and mesenchymal stem cells (MSCs). Characterization of HSCs ...

Comparison of Gene Expression in Human Embryonic Stem Cells, hESC-Derived Mesenchymal Stem Cells and Human Mesenchymal Stem Cells

OpenAIRE

Romain Barbet; Isabelle Peiffer; Antoinette Hatzfeld; Pierre Charbord; Jacques A. Hatzfeld

2011-01-01

We present a strategy to identify developmental/differentiation and plasma membrane marker genes of the most primitive human Mesenchymal Stem Cells (hMSCs). Using sensitive and quantitative TaqMan Low Density Arrays (TLDA) methodology, we compared the expression of 381 genes in human Embryonic Stem Cells (hESCs), hESC-derived MSCs ...

Notch signaling is required for maintaining stem-cell features of neuroprogenitor cells derived from human embryonic stem cells

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Chung Hyung-Min

2009-08-01

Full Text Available Abstract Background Studies have provided important findings about the roles of Notch signaling in neural development. Unfortunately, however, most of these studies have investigated the neural stem cells (NSCs of mice or other laboratory animals rather than humans, mainly owing to the difficulties associated with obtaining human brain samples. It prompted us to focus on neuroectodermal spheres (NESs which are derived from human embryonic stem cell (hESC and densely inhabited by NSCs. We here investigated the role of Notch signaling with the hESC-derived NESs. Results From hESCs, we derived NESs, the in-vitro version of brain-derived neurospheres. NES formation was confirmed by increased levels of various NSC marker genes and the emergence of rosette structures in which neuroprogenitors are known to reside. We found that Notch signaling, which maintains stem cell characteristics of in-vivo-derived neuroprogenitors, is active in these hESC-derived NESs, similar to their in-vivo counterpart. Expression levels of Notch signaling molecules such as NICD, DLLs, JAG1, HES1 and HES5 were increased in the NESs. Inhibition of the Notch signaling by a γ-secretase inhibitor reduced rosette structures, expression levels of NSC marker genes and proliferation potential in the NESs, and, if combined with withdrawal of growth factors, triggered differentiation toward neurons. Conclusion Our results indicate that the hESC-derived NESs, which share biochemical features with brain-derived neurospheres, maintain stem cell characteristics mainly through Notch signaling, which suggests that the hESC-derived NESs could be an in-vitro model for in-vivo neurogenesis.

Comparative study of adipose-derived stem cells and bone marrow-derived stem cells in similar microenvironmental conditions

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Guneta, Vipra [Division of Materials Technology, School of Materials Science and Engineering, Nanyang Technological University, 50 Nanyang Avenue, Singapore 639798 (Singapore); Tan, Nguan Soon [School of Biological Science, Nanyang Technological University, 60 Nanyang Drive, Singapore 637551 (Singapore); KK Research Centre, KK Women' s and Children Hospital, 100 Bukit Timah Road, Singapore 229899 (Singapore); Institute of Molecular and Cell Biology, Agency for Science Technology & Research - A*STAR, 61 Biopolis Drive, Proteos, Singapore 138673 (Singapore); Chan, Soon Kiat Jeremy [School of Biological Science, Nanyang Technological University, 60 Nanyang Drive, Singapore 637551 (Singapore); Tanavde, Vivek [Bioinformatics Institute, Agency for Science Technology & Research - A*STAR, 30 Biopolis Street, Matrix, Singapore 138671 (Singapore); Lim, Thiam Chye [Division of Plastic, Reconstructive and Aesthetic Surgery, Department of Surgery, National University Hospital (NUH) and National University of Singapore (NUS), Kent Ridge Wing, Singapore 119074 (Singapore); Wong, Thien Chong Marcus [Plastic, Reconstructive and Aesthetic Surgery Section, Tan Tock Seng Hospital (TTSH), 11, Jalan Tan Tock Seng, Singapore 308433 (Singapore); Choong, Cleo, E-mail: cleochoong@ntu.edu.sg [Division of Materials Technology, School of Materials Science and Engineering, Nanyang Technological University, 50 Nanyang Avenue, Singapore 639798 (Singapore); KK Research Centre, KK Women' s and Children Hospital, 100 Bukit Timah Road, Singapore 229899 (Singapore)

2016-11-01

Mesenchymal stem cells (MSCs), which were first isolated from the bone marrow, are now being extracted from various other tissues in the body, including the adipose tissue. The current study presents systematic evidence of how the adipose tissue-derived stem cells (ASCs) and bone marrow-derived mesenchymal stem cells (Bm-MSCs) behave when cultured in specific pro-adipogenic microenvironments. The cells were first characterized and identified as MSCs in terms of their morphology, phenotypic expression, self-renewal capabilities and multi-lineage potential. Subsequently, the proliferation and gene expression profiles of the cell populations cultured on two-dimensional (2D) adipose tissue extracellular matrix (ECM)-coated tissue culture plastic (TCP) and in three-dimensional (3D) AlgiMatrix® microenvironments were analyzed. Overall, it was found that adipogenesis was triggered in both cell populations due to the presence of adipose tissue ECM. However, in 3D microenvironments, ASCs and Bm-MSCs were predisposed to the adipogenic and osteogenic lineages respectively. Overall, findings from this study will contribute to ongoing efforts in adipose tissue engineering as well as provide new insights into the role of the ECM and cues provided by the immediate microenvironment for stem cell differentiation. - Highlights: • Native adipose tissue ECM coated on 2D TCP triggers adipogenesis in both ASCs and Bm-MSCs. • A 3D microenvironment with similar stiffness to adipose tissue induces adipogenic differentiation of ASCs. • ASCs cultured in 3D alginate scaffolds exhibit predisposition to adipogenesis. • Bm-MSCs cultured in 3D alginate scaffolds exhibit predisposition to osteogenesis. • The native microenvironment of the cells affects their differentiation behaviour in vitro.

Comparative study of adipose-derived stem cells and bone marrow-derived stem cells in similar microenvironmental conditions

International Nuclear Information System (INIS)

Guneta, Vipra; Tan, Nguan Soon; Chan, Soon Kiat Jeremy; Tanavde, Vivek; Lim, Thiam Chye; Wong, Thien Chong Marcus; Choong, Cleo

2016-01-01

Mesenchymal stem cells (MSCs), which were first isolated from the bone marrow, are now being extracted from various other tissues in the body, including the adipose tissue. The current study presents systematic evidence of how the adipose tissue-derived stem cells (ASCs) and bone marrow-derived mesenchymal stem cells (Bm-MSCs) behave when cultured in specific pro-adipogenic microenvironments. The cells were first characterized and identified as MSCs in terms of their morphology, phenotypic expression, self-renewal capabilities and multi-lineage potential. Subsequently, the proliferation and gene expression profiles of the cell populations cultured on two-dimensional (2D) adipose tissue extracellular matrix (ECM)-coated tissue culture plastic (TCP) and in three-dimensional (3D) AlgiMatrix® microenvironments were analyzed. Overall, it was found that adipogenesis was triggered in both cell populations due to the presence of adipose tissue ECM. However, in 3D microenvironments, ASCs and Bm-MSCs were predisposed to the adipogenic and osteogenic lineages respectively. Overall, findings from this study will contribute to ongoing efforts in adipose tissue engineering as well as provide new insights into the role of the ECM and cues provided by the immediate microenvironment for stem cell differentiation. - Highlights: • Native adipose tissue ECM coated on 2D TCP triggers adipogenesis in both ASCs and Bm-MSCs. • A 3D microenvironment with similar stiffness to adipose tissue induces adipogenic differentiation of ASCs. • ASCs cultured in 3D alginate scaffolds exhibit predisposition to adipogenesis. • Bm-MSCs cultured in 3D alginate scaffolds exhibit predisposition to osteogenesis. • The native microenvironment of the cells affects their differentiation behaviour in vitro.

Potential antitumor therapeutic strategies of human amniotic membrane and amniotic fluid-derived stem cells.

Science.gov (United States)

Kang, N-H; Hwang, K-A; Kim, S U; Kim, Y-B; Hyun, S-H; Jeung, E-B; Choi, K-C

2012-08-01

As stem cells are capable of self-renewal and can generate differentiated progenies for organ development, they are considered as potential source for regenerative medicine and tissue replacement after injury or disease. Along with this capacity, stem cells have the therapeutic potential for treating human diseases including cancers. According to the origins, stem cells are broadly classified into two types: embryonic stem cells (ESCs) and adult stem cells. In terms of differentiation potential, ESCs are pluripotent and adult stem cells are multipotent. Amnion, which is a membranous sac that contains the fetus and amniotic fluid and functions in protecting the developing embryo during gestation, is another stem cell source. Amnion-derived stem cells are classified as human amniotic membrane-derived epithelial stem cells, human amniotic membrane-derived mesenchymal stem cells and human amniotic fluid-derived stem cells. They are in an intermediate stage between pluripotent ESCs and lineage-restricted adult stem cells, non-tumorigenic, and contribute to low immunogenicity and anti-inflammation. Furthermore, they are easily available and do not cause any controversial issues in their recovery and applications. Not only are amnion-derived stem cells applicable in regenerative medicine, they have anticancer capacity. In non-engineered stem cells transplantation strategies, amnion-derived stem cells effectively target the tumor and suppressed the tumor growth by expressing cytotoxic cytokines. Additionally, they also have a potential as novel delivery vehicles transferring therapeutic genes to the cancer formation sites in gene-directed enzyme/prodrug combination therapy. Owing to their own advantageous properties, amnion-derived stem cells are emerging as a new candidate in anticancer therapy.

The Phenotypic Fate of Bone Marrow-Derived Stem Cells in Acute Kidney Injury

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Guowei Feng

2013-11-01

Full Text Available Background: Despite increasing attention on the role of bone marrow derived stem cells in repair or rejuvenation of tissues and organs, cellular mechanisms of such cell-based therapy remain poorly understood. Methods: We reconstituted hematopoiesis in recipient C57BL/6J mice by transplanting syngeneic GFP+ bone marrow (BM cells. Subsequently, the recipients received subcutaneous injection of granulocyte-colony stimulating factor (G-CSF and were subjected to acute renal ischemic injury. Flow cytometry and immunostaining were performed at various time points to assess engraftment and phenotype of BM derived stem cells. Results: Administration of G-CSF increased the release of BM derived stem cells into circulation and enhanced the ensuing recruitment of BM derived stem cells into injured kidney. During the second month post injury, migrated BM derived stem cells lost hematopoietic phenotype (CD45 but maintained the expression of other markers (Sca-1, CD133 and CD44, suggesting their potential of transdifferentiation into renal stem cells. Moreover, G-CSF treatment enhanced the phenotypic conversion. Conclusion: Our work depicted a time-course dependent transition of phenotypic characteristics of BM derived stem cells, demonstrated the existence of BM derived stem cells in damaged kidney and revealed the effects of G-CSF on cell transdifferentiation.

Adipose Derived Mesenchymal Stem Cells In Wound Healing: A Clinical Review

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Gunalp Uzun

2014-08-01

Full Text Available The aim of this article is to review clinical studies on the use of adipose derived mesenchymal stem cells in the treatment of chronic wounds. A search on PubMed was performed on April 30th, 2014 to identify the relevant clinical studies. We reviewed 13 articles that reported the use adipose derived stem cells in the treatment of different types of wounds. Adipose derived stem cells have the potential to be used in the treatment of chronic wounds. However, standard methods for isolation, storage and application of these cells are needed. New materials to transfer these stem cells to injured tissues should be investigated. [Dis Mol Med 2014; 2(4.000: 57-64

In vitro cementoblast-like differentiation of postmigratory neural crest-derived p75+ stem cells with dental follicle cell conditioned medium

International Nuclear Information System (INIS)

Wen, Xiujie; Liu, Luchuan; Deng, Manjing; Liu, Rui; Zhang, Li; Nie, Xin

2015-01-01

Cranial neural crest-derived cells (CNCCs) play important role in epithelial–mesenchymal interactions during tooth morphogenesis. However, the heterogeneity of CNCCs and their tendency to spontaneously differentiate along smooth muscle or osteoblast lineages in vitro limit further understanding of their biological properties. We studied the differentiation properties of isolated rat embryonic postmigratory CNCCs, expressing p75 neurotrophin receptor (p75NTR). These p75NTR positive (p75 + ) CNCCs, isolated using fluorescence activated cell sorter, exhibited fibroblast-like morphology and characteristics of mesenchymal stem cells. Incubation of p75 + CNCCs in dental follicle cell conditioned medium (DFCCM) combined with dentin non-collagenous proteins (dNCPs), altered their morphological features to cementoblast-like appearance. These cells also showed low proliferative activity, high ALP activity and significantly increased calcified nodule formation. Markers related to mineralization or specific to cementoblast lineage were highly expressed in dNCPs/DFCCM-treated p75 + cells, suggesting their differentiation along cementoblast-like lineage. p75 + stem cells selected from postmigratory CNCCs represent a pure stem cell population and could be used as a stem cell model for in vitro studies due to their intrinsic ability to differentiate to neuronal cells and transform from neuroectoderm to ectomesenchyme. They can provide a potential stem cell resource for tooth engineering studies and help to further investigate mechanisms of epithelial–mesenchymal interactions in tooth morphogenesis. - Highlights: • Cranial neural crest-derived cells (CNCCs) take part in tooth morphogenesis. • positive (p75 + ) CNCCs are fibroblast-like and resemble mesenchymal stem cells. • p75 + CNCCs in dental follicle cell medium (DFCCM/dNCP) appear like cementoblasts. • DFCCM/dNCP-treated p75 + cells express cementoblast specific mineralization markers. • p75 + cells are pure stem

Time- and dose-dependent effects of total-body ionizing radiation on muscle stem cells

Science.gov (United States)

Masuda, Shinya; Hisamatsu, Tsubasa; Seko, Daiki; Urata, Yoshishige; Goto, Shinji; Li, Tao-Sheng; Ono, Yusuke

2015-01-01

Exposure to high levels of genotoxic stress, such as high-dose ionizing radiation, increases both cancer and noncancer risks. However, it remains debatable whether low-dose ionizing radiation reduces cellular function, or rather induces hormetic health benefits. Here, we investigated the effects of total-body γ-ray radiation on muscle stem cells, called satellite cells. Adult C57BL/6 mice were exposed to γ-radiation at low- to high-dose rates (low, 2 or 10 mGy/day; moderate, 50 mGy/day; high, 250 mGy/day) for 30 days. No hormetic responses in proliferation, differentiation, or self-renewal of satellite cells were observed in low-dose radiation-exposed mice at the acute phase. However, at the chronic phase, population expansion of satellite cell-derived progeny was slightly decreased in mice exposed to low-dose radiation. Taken together, low-dose ionizing irradiation may suppress satellite cell function, rather than induce hormetic health benefits, in skeletal muscle in adult mice. PMID:25869487

Role of adipose-derived stem cells in wound healing.

Science.gov (United States)

Hassan, Waqar Ul; Greiser, Udo; Wang, Wenxin

2014-01-01

Impaired wound healing remains a challenge to date and causes debilitating effects with tremendous suffering. Recent advances in tissue engineering approaches in the area of cell therapy have provided promising treatment options to meet the challenges of impaired skin wound healing such as diabetic foot ulcers. Over the last few years, stem cell therapy has emerged as a novel therapeutic approach for various diseases including wound repair and tissue regeneration. Several different types of stem cells have been studied in both preclinical and clinical settings such as bone marrow-derived stem cells, adipose-derived stem cells (ASCs), circulating angiogenic cells (e.g., endothelial progenitor cells), human dermal fibroblasts, and keratinocytes for wound healing. Adipose tissue is an abundant source of mesenchymal stem cells, which have shown an improved outcome in wound healing studies. ASCs are pluripotent stem cells with the ability to differentiate into different lineages and to secrete paracrine factors initiating tissue regeneration process. The abundant supply of fat tissue, ease of isolation, extensive proliferative capacities ex vivo, and their ability to secrete pro-angiogenic growth factors make them an ideal cell type to use in therapies for the treatment of nonhealing wounds. In this review, we look at the pathogenesis of chronic wounds, role of stem cells in wound healing, and more specifically look at the role of ASCs, their mechanism of action and their safety profile in wound repair and tissue regeneration. © 2014 by the Wound Healing Society.

Foetal stem cell derivation & characterization for osteogenic lineage

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A Mangala Gowri

2013-01-01

Full Text Available Background & objectives: Mesencymal stem cells (MSCs derived from foetal tissues present a multipotent progenitor cell source for application in tissue engineering and regenerative medicine. The present study was carried out to derive foetal mesenchymal stem cells from ovine source and analyze their differentiation to osteogenic linage to serve as an animal model to predict human applications. Methods: Isolation and culture of sheep foetal bone marrow cells were done and uniform clonally derived MSC population was collected. The cells were characterized using cytochemical, immunophenotyping, biochemical and molecular analyses. The cells with defined characteristics were differentiated into osteogenic lineages and analysis for differentiated cell types was done. The cells were analyzed for cell surface marker expression and the gene expression in undifferentiated and differentiated osteoblast was checked by reverse transcriptase PCR (RT PCR analysis and confirmed by sequencing using genetic analyzer. Results: Ovine foetal samples were processed to obtain mononuclear (MNC cells which on culture showed spindle morphology, a characteristic oval body with the flattened ends. MSC population CD45 - /CD14 - was cultured by limiting dilution to arrive at uniform spindle morphology cells and colony forming units. The cells were shown to be positive for surface markers such as CD44, CD54, integrinβ1, and intracellular collagen type I/III and fibronectin. The osteogenically induced MSCs were analyzed for alkaline phosphatase (ALP activity and mineral deposition. The undifferentiated MSCs expressed RAB3B, candidate marker for stemness in MSCs. The osteogenically induced and uninduced MSCs expressed collagen type I and MMP13 gene in osteogenic induced cells. Interpretation & conclusions: The protocol for isolation of ovine foetal bone marrow derived MSCs was simple to perform, and the cultural method of obtaining pure spindle morphology cells was established

Study of muscle cell dedifferentiation after skeletal muscle injury of mice with a Cre-Lox system.

Science.gov (United States)

Mu, Xiaodong; Peng, Hairong; Pan, Haiying; Huard, Johnny; Li, Yong

2011-02-03

Dedifferentiation of muscle cells in the tissue of mammals has yet to be observed. One of the challenges facing the study of skeletal muscle cell dedifferentiation is the availability of a reliable model that can confidentially distinguish differentiated cell populations of myotubes and non-fused mononuclear cells, including stem cells that can coexist within the population of cells being studied. In the current study, we created a Cre/Lox-β-galactosidase system, which can specifically tag differentiated multinuclear myotubes and myotube-generated mononuclear cells based on the activation of the marker gene, β-galactosidase. By using this system in an adult mouse model, we found that β-galactosidase positive mononuclear cells were generated from β-galactosidase positive multinuclear myofibers upon muscle injury. We also demonstrated that these mononuclear cells can develop into a variety of different muscle cell lineages, i.e., myoblasts, satellite cells, and muscle derived stem cells. These novel findings demonstrated, for the first time, that cellular dedifferentiation of skeletal muscle cells actually occurs in mammalian skeletal muscle following traumatic injury in vivo.

Safety and immune regulatory properties of canine induced pluripotent stem cell-derived mesenchymal stem cells.

Science.gov (United States)

Chow, Lyndah; Johnson, Valerie; Regan, Dan; Wheat, William; Webb, Saiphone; Koch, Peter; Dow, Steven

2017-12-01

Mesenchymal stem cells (MSCs) exhibit broad immune modulatory activity in vivo and can suppress T cell proliferation and dendritic cell activation in vitro. Currently, most MSC for clinical usage are derived from younger donors, due to ease of procurement and to the superior immune modulatory activity. However, the use of MSC from multiple unrelated donors makes it difficult to standardize study results and compare outcomes between different clinical trials. One solution is the use of MSC derived from induced pluripotent stem cells (iPSC); as iPSC-derived MSC have nearly unlimited proliferative potential and exhibit in vitro phenotypic stability. Given the value of dogs as a spontaneous disease model for pre-clinical evaluation of stem cell therapeutics, we investigated the functional properties of canine iPSC-derived MSC (iMSC), including immune modulatory properties and potential for teratoma formation. We found that canine iMSC downregulated expression of pluripotency genes and appeared morphologically similar to conventional MSC. Importantly, iMSC retained a stable phenotype after multiple passages, did not form teratomas in immune deficient mice, and did not induce tumor formation in dogs following systemic injection. We concluded therefore that iMSC were phenotypically stable, immunologically potent, safe with respect to tumor formation, and represented an important new source of cells for therapeutic modulation of inflammatory disorders. Copyright © 2017. Published by Elsevier B.V.

Template DNA-strand co-segregation and asymmetric cell division in skeletal muscle stem cells.

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Shinin, Vasily; Gayraud-Morel, Barbara; Tajbakhsh, Shahragim

2009-01-01

Stem cells are present in all tissues and organs, and are crucial for normal regulated growth. How the pool size of stem cells and their progeny is regulated to establish the tissue prenatally, then maintain it throughout life, is a key question in biology and medicine. The ability to precisely locate stem and progenitors requires defining lineage progression from stem to differentiated cells, assessing the mode of cell expansion and self-renewal and identifying markers to assess the different cell states within the lineage. We have shown that during lineage progression from a quiescent adult muscle satellite cell to a differentiated myofibre, both symmetric and asymmetric divisions take place. Furthermore, we provide evidence that a sub-population of label retaining satellite cells co-segregate template DNA strands to one daughter cell. These findings provide a means of identifying presumed stem and progenitor cells within the lineage. In addition, asymmetric segregation of template DNA and the cytoplasmic protein Numb provides a landmark to define cell behaviour as self-renewal and differentiation decisions are being executed.

Autologous Pluripotent Stem Cell-Derived β-Like Cells for Diabetes Cellular Therapy.

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Millman, Jeffrey R; Pagliuca, Felicia W

2017-05-01

Development of stem cell technologies for cell replacement therapy has progressed rapidly in recent years. Diabetes has long been seen as one of the first applications for stem cell-derived cells because of the loss of only a single cell type-the insulin-producing β-cell. Recent reports have detailed strategies that overcome prior hurdles to generate functional β-like cells from human pluripotent stem cells in vitro, including from human induced pluripotent stem cells (hiPSCs). Even with this accomplishment, addressing immunological barriers to transplantation remains a major challenge for the field. The development of clinically relevant hiPSC derivation methods from patients and demonstration that these cells can be differentiated into β-like cells presents a new opportunity to treat diabetes without immunosuppression or immunoprotective encapsulation or with only targeted protection from autoimmunity. This review focuses on the current status in generating and transplanting autologous β-cells for diabetes cell therapy, highlighting the unique advantages and challenges of this approach. © 2017 by the American Diabetes Association.

Three-Dimensional Human iPSC-Derived Artificial Skeletal Muscles Model Muscular Dystrophies and Enable Multilineage Tissue Engineering

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Sara Martina Maffioletti

2018-04-01

Full Text Available Summary: Generating human skeletal muscle models is instrumental for investigating muscle pathology and therapy. Here, we report the generation of three-dimensional (3D artificial skeletal muscle tissue from human pluripotent stem cells, including induced pluripotent stem cells (iPSCs from patients with Duchenne, limb-girdle, and congenital muscular dystrophies. 3D skeletal myogenic differentiation of pluripotent cells was induced within hydrogels under tension to provide myofiber alignment. Artificial muscles recapitulated characteristics of human skeletal muscle tissue and could be implanted into immunodeficient mice. Pathological cellular hallmarks of incurable forms of severe muscular dystrophy could be modeled with high fidelity using this 3D platform. Finally, we show generation of fully human iPSC-derived, complex, multilineage muscle models containing key isogenic cellular constituents of skeletal muscle, including vascular endothelial cells, pericytes, and motor neurons. These results lay the foundation for a human skeletal muscle organoid-like platform for disease modeling, regenerative medicine, and therapy development. : Maffioletti et al. generate human 3D artificial skeletal muscles from healthy donors and patient-specific pluripotent stem cells. These human artificial muscles accurately model severe genetic muscle diseases. They can be engineered to include other cell types present in skeletal muscle, such as vascular cells and motor neurons. Keywords: skeletal muscle, pluripotent stem cells, iPS cells, myogenic differentiation, tissue engineering, disease modeling, muscular dystrophy, organoids

Enhanced neuro-therapeutic potential of Wharton's Jelly-derived mesenchymal stem cells in comparison with bone marrow mesenchymal stem cells culture.

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Drela, Katarzyna; Lech, Wioletta; Figiel-Dabrowska, Anna; Zychowicz, Marzena; Mikula, Michał; Sarnowska, Anna; Domanska-Janik, Krystyna

2016-04-01

Substantial inconsistencies in mesenchymal stem (stromal) cell (MSC) therapy reported in early translational and clinical studies may indicate need for selection of the proper cell population for any particular therapeutic purpose. In the present study we have examined stromal stem cells derived either from umbilical cord Wharton's Jelly (WJ-MSC) or bone marrow (BM-MSC) of adult, healthy donors. The cells characterized in accordance with the International Society for Cellular Therapy (ISCT) indications as well as other phenotypic and functional parameters have been compared under strictly controlled culture conditions. WJ-MSC, in comparison with BM-MSC, exhibited a higher proliferation rate, a greater expansion capability being additionally stimulated under low-oxygen atmosphere, enhanced neurotrophic factors gene expression and spontaneous tendency toward a neural lineage differentiation commitment confirmed by protein and gene marker induction. Our data suggest that WJ-MSC may represent an example of immature-type "pre-MSC," where a substantial cellular component is embryonic-like, pluripotent derivatives with the default neural-like differentiation. These cells may contribute in different extents to nearly all classical MSC populations adversely correlated with the age of cell donors. Our data suggest that neuro-epithelial markers, like nestin, stage specific embryonic antigens-4 or α-smooth muscle actin expressions, may serve as useful indicators of MSC culture neuro-regeneration-associated potency. Copyright © 2016 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

Adipose-derived mesenchymal stem cells accelerate nerve regeneration and functional recovery in a rat model of recurrent laryngeal nerve injury

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Yun Li

2017-01-01

Full Text Available Medialization thyroplasty or injection laryngoplasty for unilateral vocal fold paralysis cannot restore mobility of the vocal fold. Recent studies have shown that transplantation of mesenchymal stem cells is effective in the repair of nerve injuries. This study investigated whether adipose-derived stem cell transplantation could repair recurrent laryngeal nerve injury. Rat models of recurrent laryngeal nerve injury were established by crushing with micro forceps. Adipose-derived mesenchymal stem cells (ADSCs; 8 × 105 or differentiated Schwann-like adipose-derived mesenchymal stem cells (dADSCs; 8 × 105 or extracellular matrix were injected at the site of injury. At 2, 4 and 6 weeks post-surgery, a higher density of myelinated nerve fiber, thicker myelin sheath, improved vocal fold movement, better recovery of nerve conduction capacity and reduced thyroarytenoid muscle atrophy were found in ADSCs and dADSCs groups compared with the extracellular matrix group. The effects were more pronounced in the ADSCs group than in the dADSCs group. These experimental results indicated that ADSCs transplantation could be an early interventional strategy to promote regeneration after recurrent laryngeal nerve injury.

Electrophysiological properties of neurosensory progenitors derived from human embryonic stem cells

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Karina Needham

2014-01-01

Full Text Available In severe cases of sensorineural hearing loss where the numbers of auditory neurons are significantly depleted, stem cell-derived neurons may provide a potential source of replacement cells. The success of such a therapy relies upon producing a population of functional neurons from stem cells, to enable precise encoding of sound information to the brainstem. Using our established differentiation assay to produce sensory neurons from human stem cells, patch-clamp recordings indicated that all neurons examined generated action potentials and displayed both transient sodium and sustained potassium currents. Stem cell-derived neurons reliably entrained to stimuli up to 20 pulses per second (pps, with 50% entrainment at 50 pps. A comparison with cultured primary auditory neurons indicated similar firing precision during low-frequency stimuli, but significant differences after 50 pps due to differences in action potential latency and width. The firing properties of stem cell-derived neurons were also considered relative to time in culture (31–56 days and revealed no change in resting membrane potential, threshold or firing latency over time. Thus, while stem cell-derived neurons did not entrain to high frequency stimulation as effectively as mammalian auditory neurons, their electrical phenotype was stable in culture and consistent with that reported for embryonic auditory neurons.

Efficient and rapid derivation of primitive neural stem cells and generation of brain subtype neurons from human pluripotent stem cells.

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Yan, Yiping; Shin, Soojung; Jha, Balendu Shekhar; Liu, Qiuyue; Sheng, Jianting; Li, Fuhai; Zhan, Ming; Davis, Janine; Bharti, Kapil; Zeng, Xianmin; Rao, Mahendra; Malik, Nasir; Vemuri, Mohan C

2013-11-01

Human pluripotent stem cells (hPSCs), including human embryonic stem cells and human induced pluripotent stem cells, are unique cell sources for disease modeling, drug discovery screens, and cell therapy applications. The first step in producing neural lineages from hPSCs is the generation of neural stem cells (NSCs). Current methods of NSC derivation involve the time-consuming, labor-intensive steps of an embryoid body generation or coculture with stromal cell lines that result in low-efficiency derivation of NSCs. In this study, we report a highly efficient serum-free pluripotent stem cell neural induction medium that can induce hPSCs into primitive NSCs (pNSCs) in 7 days, obviating the need for time-consuming, laborious embryoid body generation or rosette picking. The pNSCs expressed the neural stem cell markers Pax6, Sox1, Sox2, and Nestin; were negative for Oct4; could be expanded for multiple passages; and could be differentiated into neurons, astrocytes, and oligodendrocytes, in addition to the brain region-specific neuronal subtypes GABAergic, dopaminergic, and motor neurons. Global gene expression of the transcripts of pNSCs was comparable to that of rosette-derived and human fetal-derived NSCs. This work demonstrates an efficient method to generate expandable pNSCs, which can be further differentiated into central nervous system neurons and glia with temporal, spatial, and positional cues of brain regional heterogeneity. This method of pNSC derivation sets the stage for the scalable production of clinically relevant neural cells for cell therapy applications in good manufacturing practice conditions.

Delivery of adipose-derived stem cells in poloxamer hydrogel improves peripheral nerve regeneration.

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Allbright, Kassandra O; Bliley, Jacqueline M; Havis, Emmanuelle; Kim, Deok-Yeol; Dibernardo, Gabriella A; Grybowski, Damian; Waldner, Matthias; James, Isaac B; Sivak, Wesley N; Rubin, J Peter; Marra, Kacey G

2018-02-06

Peripheral nerve damage is associated with high long-term morbidity. Because of beneficial secretome, immunomodulatory effects, and ease of clinical translation, transplantation with adipose-derived stem cells (ASC) represents a promising therapeutic modality. Effect of ASC delivery in poloxamer hydrogel was assessed in a rat sciatic nerve model of critical-sized (1.5 cm) peripheral nerve injury. Nerve/muscle unit regeneration was assessed via immunostaining explanted nerve, quantitative polymerase chain reaction (qPCR), and histological analysis of reinnervating gastrocnemius muscle. On the basis of viability data, 10% poloxamer hydrogel was selected for in vivo study. Six weeks after transection and repair, the group treated with poloxamer delivered ASCs demonstrated longest axonal regrowth. The qPCR results indicated that the inclusion of ASCs appeared to result in expression of factors that aid in reinnervating muscle tissue. Delivery of ASCs in poloxamer addresses multiple facets of the complexity of nerve/muscle unit regeneration, representing a promising avenue for further study. Muscle Nerve, 2018. © 2018 Wiley Periodicals, Inc.

Single Stem Cell Imaging and Analysis Reveals Telomere Length Differences in Diseased Human and Mouse Skeletal Muscles

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Elisia D. Tichy

2017-10-01

Full Text Available Muscle stem cells (MuSCs contribute to muscle regeneration following injury. In many muscle disorders, the repeated cycles of damage and repair lead to stem cell dysfunction. While telomere attrition may contribute to aberrant stem cell functions, methods to accurately measure telomere length in stem cells from skeletal muscles have not been demonstrated. Here, we have optimized and validated such a method, named MuQ-FISH, for analyzing telomere length in MuSCs from either mice or humans. Our analysis showed no differences in telomere length between young and aged MuSCs from uninjured wild-type mice, but MuSCs isolated from young dystrophic mice exhibited significantly shortened telomeres. In corroboration, we demonstrated that telomere attrition is present in human dystrophic MuSCs, which underscores its importance in diseased regenerative failure. The robust technique described herein provides analysis at a single-cell resolution and may be utilized for other cell types, especially rare populations of cells.

Donor-derived stem-cells and epithelial mesenchymal transition in squamous cell carcinoma in transplant recipients.

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Verneuil, Laurence; Leboeuf, Christophe; Bousquet, Guilhem; Brugiere, Charlotte; Elbouchtaoui, Morad; Plassa, Louis-François; Peraldi, Marie-Noelle; Lebbé, Celeste; Ratajczak, Philippe; Janin, Anne

2015-12-08

Skin squamous-cell-carcinoma (SCC), is the main complication in long-term kidney-transplant recipients, and it can include donor-derived cells. Preclinical models demonstrated the involvement of epithelial mesenchymal transition (EMT) in the progression of skin SCC, and the role of Snail, an EMT transcription factor, in cancer stem-cell survival and expansion.Here, we studied stem-cells and EMT expression in SCCs and concomitant actinic keratoses (AK) in kidney-transplant recipients. In SCC and AK in 3 female recipients of male kidney-transplants, donor-derived Y chromosome in epidermal stem cells was assessed using combined XY-FISH/CD133 immunostaining, and digital-droplet-PCR on laser-microdissected CD133 expressing epidermal cells.For EMT study, double immunostainings of CD133 with vimentin or snail and slug, electron microscopy and immunostainings of keratinocytes junctions were performed. Digital droplet PCR was used to check CDH1 (E-cadherin) expression level in laser-microdissected cells co-expressing CD133 and vimentin or snail and slug.The numbers of Y-chromosome were assessed using digital droplet PCR in laser-microdissected cells co-expressing CD133 and vimentin, or snail and slug, and in CD133 positive cells not expressing any EMT maker. We identified donor-derived stem-cells in basal layers and invasive areas in all skin SCCs and in concomitant AKs, but not in surrounding normal skin.The donor-derived stem-cells expressed the EMT markers, vimentin, snail and slug in SCCs but not in AKs. The expression of the EMT transcription factor, SNAI1, was higher in stem-cells when they expressed vimentin. They were located in invasive areas of SCCs. In these areas, the expressions of claudin-1 and desmoglein 1 were reduced or absent, and within the basal layer there were features of basal membrane disappearance.Donor-derived stem cells were in larger numbers in stem cells co-expressing vimentin or snail and slug than in stem cells not expressing any EMT marker

Preclinical Studies of Induced Pluripotent Stem Cell-Derived Astrocyte Transplantation in ALS

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2012-10-01

Pluripotent Stem Cell -Derived Astrocyte Transplantation in ALS PRINCIPAL INVESTIGATOR: Nicholas J. Maragakis, M.D...Pluripotent Stem Cell -Derived Astrocyte Transplantation in ALS 5b. GRANT NUMBER W81XWH-10-1-0520 5c. PROGRAM ELEMENT NUMBER 6. AUTHOR(S) 5d...into astrocytes following transplantation. 15. SUBJECT TERMS Stem Cells , iPS cells, astrocytes, familial ALS 16. SECURITY CLASSIFICATION OF

In vitro cementoblast-like differentiation of postmigratory neural crest-derived p75{sup +} stem cells with dental follicle cell conditioned medium

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Wen, Xiujie; Liu, Luchuan; Deng, Manjing; Liu, Rui; Zhang, Li; Nie, Xin, E-mail: dr.xinnie@gmail.com

2015-09-10

Cranial neural crest-derived cells (CNCCs) play important role in epithelial–mesenchymal interactions during tooth morphogenesis. However, the heterogeneity of CNCCs and their tendency to spontaneously differentiate along smooth muscle or osteoblast lineages in vitro limit further understanding of their biological properties. We studied the differentiation properties of isolated rat embryonic postmigratory CNCCs, expressing p75 neurotrophin receptor (p75NTR). These p75NTR positive (p75{sup +}) CNCCs, isolated using fluorescence activated cell sorter, exhibited fibroblast-like morphology and characteristics of mesenchymal stem cells. Incubation of p75{sup +} CNCCs in dental follicle cell conditioned medium (DFCCM) combined with dentin non-collagenous proteins (dNCPs), altered their morphological features to cementoblast-like appearance. These cells also showed low proliferative activity, high ALP activity and significantly increased calcified nodule formation. Markers related to mineralization or specific to cementoblast lineage were highly expressed in dNCPs/DFCCM-treated p75{sup +} cells, suggesting their differentiation along cementoblast-like lineage. p75{sup +} stem cells selected from postmigratory CNCCs represent a pure stem cell population and could be used as a stem cell model for in vitro studies due to their intrinsic ability to differentiate to neuronal cells and transform from neuroectoderm to ectomesenchyme. They can provide a potential stem cell resource for tooth engineering studies and help to further investigate mechanisms of epithelial–mesenchymal interactions in tooth morphogenesis. - Highlights: • Cranial neural crest-derived cells (CNCCs) take part in tooth morphogenesis. • positive (p75{sup +}) CNCCs are fibroblast-like and resemble mesenchymal stem cells. • p75{sup +} CNCCs in dental follicle cell medium (DFCCM/dNCP) appear like cementoblasts. • DFCCM/dNCP-treated p75{sup +} cells express cementoblast specific mineralization

Differentiation and molecular profiling of human embryonic stem cell-derived corneal epithelial cells.

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Brzeszczynska, J; Samuel, K; Greenhough, S; Ramaesh, K; Dhillon, B; Hay, D C; Ross, J A

2014-06-01

It has been suggested that the isolation of scalable populations of limbal stem cells may lead to radical changes in ocular therapy. In particular, the derivation and transplantation of corneal stem cells from these populations may result in therapies providing clinical normality of the diseased or damaged cornea. Although feasible in theory, the lack of donor material in sufficient quantity and quality currently limits such a strategy. A potential scalable source of corneal cells could be derived from pluripotent stem cells (PSCs). We developed an in vitro and serum-free corneal differentiation model which displays significant promise. Our stepwise differentiation model was designed with reference to development and gave rise to cells which displayed similarities to epithelial progenitor cells which can be specified to cells displaying a corneal epithelial phenotype. We believe our approach is novel, provides a robust model of human development and in the future, may facilitate the generation of corneal epithelial cells that are suitable for clinical use. Additionally, we demonstrate that following continued cell culture, stem cell-derived corneal epithelial cells undergo transdifferentiation and exhibit squamous metaplasia and therefore, also offer an in vitro model of disease.

Large Scale Production of Stem Cells and Their Derivatives

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Zweigerdt, Robert

Stem cells have been envisioned to become an unlimited cell source for regenerative medicine. Notably, the interest in stem cells lies beyond direct therapeutic applications. They might also provide a previously unavailable source of valuable human cell types for screening platforms, which might facilitate the development of more efficient and safer drugs. The heterogeneity of stem cell types as well as the numerous areas of application suggests that differential processes are mandatory for their in vitro culture. Many of the envisioned applications would require the production of a high number of stem cells and their derivatives in scalable, well-defined and potentially clinical compliant manner under current good manufacturing practice (cGMP). In this review we provide an overview on recent strategies to develop bioprocesses for the expansion, differentiation and enrichment of stem cells and their progenies, presenting examples for adult and embryonic stem cells alike.

Differentiation defect in neural crest-derived smooth muscle cells in patients with aortopathy associated with bicuspid aortic valves

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Jiao Jiao

2016-08-01

Full Text Available Individuals with bicuspid aortic valves (BAV are at a higher risk of developing thoracic aortic aneurysms (TAA than patients with trileaflet aortic valves (TAV. The aneurysms associated with BAV most commonly involve the ascending aorta and spare the descending aorta. Smooth muscle cells (SMCs in the ascending and descending aorta arise from neural crest (NC and paraxial mesoderm (PM, respectively. We hypothesized defective differentiation of the neural crest stem cells (NCSCs-derived SMCs but not paraxial mesoderm cells (PMCs-derived SMCs contributes to the aortopathy associated with BAV. When induced pluripotent stem cells (iPSCs from BAV/TAA patients were differentiated into NCSC-derived SMCs, these cells demonstrated significantly decreased expression of marker of SMC differentiation (MYH11 and impaired contraction compared to normal control. In contrast, the PMC-derived SMCs were similar to control cells in these aspects. The NCSC-SMCs from the BAV/TAA also showed decreased TGF-β signaling based on phosphorylation of SMAD2, and increased mTOR signaling. Inhibition of mTOR pathway using rapamycin rescued the aberrant differentiation. Our data demonstrates that decreased differentiation and contraction of patient's NCSC-derived SMCs may contribute to that aortopathy associated with BAV.

Impaired cell surface expression of HLA-B antigens on mesenchymal stem cells and muscle cell progenitors

DEFF Research Database (Denmark)

Isa, Adiba; Nehlin, Jan; Sabir, Hardee Jawad

2010-01-01

HLA class-I expression is weak in embryonic stem cells but increases rapidly during lineage progression. It is unknown whether all three classical HLA class-I antigens follow the same developmental program. In the present study, we investigated allele-specific expression of HLA-A, -B, and -C...... at the mRNA and protein levels on human mesenchymal stem cells from bone marrow and adipose tissue as well as striated muscle satellite cells and lymphocytes. Using multicolour flow cytometry, we found high cell surface expression of HLA-A on all stem cells and PBMC examined. Surprisingly, HLA-B was either...... undetectable or very weakly expressed on all stem cells protecting them from complement-dependent cytotoxicity (CDC) using relevant human anti-B and anti-Cw sera. IFNgamma stimulation for 48-72 h was required to induce full HLA-B protein expression. Quantitative real-time RT-PCR showed that IFNgamma induced...

Transiently Active Wnt/β-Catenin Signaling Is Not Required but Must Be Silenced for Stem Cell Function during Muscle Regeneration

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Malea M. Murphy

2014-09-01

Full Text Available Adult muscle’s exceptional capacity for regeneration is mediated by muscle stem cells, termed satellite cells. As with many stem cells, Wnt/β-catenin signaling has been proposed to be critical in satellite cells during regeneration. Using new genetic reagents, we explicitly test in vivo whether Wnt/β-catenin signaling is necessary and sufficient within satellite cells and their derivatives for regeneration. We find that signaling is transiently active in transit-amplifying myoblasts, but is not required for regeneration or satellite cell self-renewal. Instead, downregulation of transiently activated β-catenin is important to limit the regenerative response, as continuous regeneration is deleterious. Wnt/β-catenin activation in adult satellite cells may simply be a vestige of their developmental lineage, in which β-catenin signaling is critical for fetal myogenesis. In the adult, surprisingly, we show that it is not activation but rather silencing of Wnt/β-catenin signaling that is important for muscle regeneration.

Induced pluripotent stem cells-derived myeloid-derived suppressor cells regulate the CD8+ T cell response

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Daniel Joyce

2018-05-01

Full Text Available Myeloid-derived suppressor cells (MDSCs are markedly increased in cancer patients and tumor-bearing mice and promote tumor growth and survival by inhibiting host innate and adaptive immunity. In this study, we generated and characterized MDSCs from murine-induced pluripotent stem cells (iPSCs. The iPSCs were co-cultured with OP9 cells, stimulated with GM-CSF, and became morphologically heterologous under co-culturing with hepatic stellate cells. Allogeneic and OVA-specific antigen stimulation demonstrated that iPS-MDSCs have a T-cell regulatory function. Furthermore, a popliteal lymph node assay and autoimmune hepatitis model showed that iPS-MDSCs also regulate immune responsiveness in vivo and have a therapeutic effect against hepatitis. Taken together, our results demonstrated a method of generating functional MDSCs from iPSCs and highlighted the potential of iPS-MDSCs as a key cell therapy resource for transplantation and autoimmune diseases. Keywords: Myeloid-derived suppressor cells, Induced pluripotent stem cells, T cell response

Skeletal myogenic differentiation of human urine-derived cells as a potential source for skeletal muscle regeneration.

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Chen, Wei; Xie, Minkai; Yang, Bin; Bharadwaj, Shantaram; Song, Lujie; Liu, Guihua; Yi, Shanhong; Ye, Gang; Atala, Anthony; Zhang, Yuanyuan

2017-02-01

Stem cells are regarded as possible cell therapy candidates for skeletal muscle regeneration. However, invasive harvesting of those cells can cause potential harvest-site morbidity. The goal of this study was to assess whether human urine-derived stem cells (USCs), obtained through non-invasive procedures, can differentiate into skeletal muscle linage cells (Sk-MCs) and potentially be used for skeletal muscle regeneration. In this study, USCs were harvested from six healthy individuals aged 25-55. Expression profiles of cell-surface markers were assessed by flow cytometry. To optimize the myogenic differentiation medium, we selected two from four different types of myogenic differentiation media to induce the USCs. Differentiated USCs were identified with myogenic markers by gene and protein expression. USCs were implanted into the tibialis anterior muscles of nude mice for 1 month. The results showed that USCs displayed surface markers with positive staining for CD24, CD29, CD44, CD73, CD90, CD105, CD117, CD133, CD146, SSEA-4 and STRO-1, and negative staining for CD14, CD31, CD34 and CD45. After myogenic differentiation, a change in morphology was observed from 'rice-grain'-like cells to spindle-shaped cells. The USCs expressed specific Sk-MC transcripts and protein markers (myf5, myoD, myosin, and desmin) after being induced with different myogenic culture media. Implanted cells expressed Sk-MC markers stably in vivo. Our findings suggest that USCs are able to differentiate into the Sk-MC lineage in vitro and after being implanted in vivo. Thus, they might be a potential source for cell injection therapy in the use of skeletal muscle regeneration. Copyright © 2014 John Wiley & Sons, Ltd. Copyright © 2014 John Wiley & Sons, Ltd.

The niche-derived glial cell line-derived neurotrophic factor (GDNF induces migration of mouse spermatogonial stem/progenitor cells.

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Lisa Dovere

Full Text Available In mammals, the biological activity of the stem/progenitor compartment sustains production of mature gametes through spermatogenesis. Spermatogonial stem cells and their progeny belong to the class of undifferentiated spermatogonia, a germ cell population found on the basal membrane of the seminiferous tubules. A large body of evidence has demonstrated that glial cell line-derived neurotrophic factor (GDNF, a Sertoli-derived factor, is essential for in vivo and in vitro stem cell self-renewal. However, the mechanisms underlying this activity are not completely understood. In this study, we show that GDNF induces dose-dependent directional migration of freshly selected undifferentiated spermatogonia, as well as germline stem cells in culture, using a Boyden chamber assay. GDNF-induced migration is dependent on the expression of the GDNF co-receptor GFRA1, as shown by migration assays performed on parental and GFRA1-transduced GC-1 spermatogonial cell lines. We found that the actin regulatory protein vasodilator-stimulated phosphoprotein (VASP is specifically expressed in undifferentiated spermatogonia. VASP belongs to the ENA/VASP family of proteins implicated in actin-dependent processes, such as fibroblast migration, axon guidance, and cell adhesion. In intact seminiferous tubules and germline stem cell cultures, GDNF treatment up-regulates VASP in a dose-dependent fashion. These data identify a novel role for the niche-derived factor GDNF, and they suggest that GDNF may impinge on the stem/progenitor compartment, affecting the actin cytoskeleton and cell migration.

Neuromuscular electrical stimulation as a method to maximize the beneficial effects of muscle stem cells transplanted into dystrophic skeletal muscle.

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Giovanna Distefano

Full Text Available Cellular therapy is a potential approach to improve the regenerative capacity of damaged or diseased skeletal muscle. However, its clinical use has often been limited by impaired donor cell survival, proliferation and differentiation following transplantation. Additionally, functional improvements after transplantation are all-too-often negligible. Because the host microenvironment plays an important role in the fate of transplanted cells, methods to modulate the microenvironment and guide donor cell behavior are warranted. The purpose of this study was to investigate whether the use of neuromuscular electrical stimulation (NMES for 1 or 4 weeks following muscle-derived stem cell (MDSC transplantation into dystrophic skeletal muscle can modulate the fate of donor cells and enhance their contribution to muscle regeneration and functional improvements. Animals submitted to 4 weeks of NMES after transplantation demonstrated a 2-fold increase in the number of dystrophin+ myofibers as compared to control transplanted muscles. These findings were concomitant with an increased vascularity in the MDSC+NMES group when compared to non-stimulated counterparts. Additionally, animals subjected to NMES (with or without MDSC transplantation presented an increased maximal specific tetanic force when compared to controls. Although cell transplantation and/or the use of NMES resulted in no changes in fatigue resistance, the combination of both MDSC transplantation and NMES resulted in a faster recovery from fatigue, when compared to non-injected and non-stimulated counterparts. We conclude that NMES is a viable method to improve MDSC engraftment, enhance dystrophic muscle strength, and, in combination with MDSC transplantation, improve recovery from fatigue. These findings suggest that NMES may be a clinically-relevant adjunct approach for cell transplantation into skeletal muscle.

A Long-Gap Peripheral Nerve Injury Therapy Using Human Skeletal Muscle-Derived Stem Cells (Sk-SCs: An Achievement of Significant Morphological, Numerical and Functional Recovery.

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Tetsuro Tamaki

Full Text Available Losses in vital functions of the somatic motor and sensory nervous system are induced by severe long-gap peripheral nerve transection injury. In such cases, autologous nerve grafts are the gold standard treatment, despite the unavoidable sacrifice of other healthy functions, whereas the prognosis is not always favorable. Here, we use human skeletal muscle-derived stem cells (Sk-SCs to reconstitute the function after long nerve-gap injury. Muscles samples were obtained from the amputated legs from 9 patients following unforeseen accidents. The Sk-SCs were isolated using conditioned collagenase solution, and sorted as CD34+/45- (Sk-34 and CD34-/45-/29+ (Sk-DN/29+ cells. Cells were separately cultured/expanded under optimal conditions for 2 weeks, then injected into the athymic nude mice sciatic nerve long-gap model (7-mm bridging an acellular conduit. After 8-12 weeks, active cell engraftment was observed only in the Sk-34 cell transplanted group, showing preferential differentiation into Schwann cells and perineurial/endoneurial cells, as well as formation of the myelin sheath and perineurium/endoneurium surrounding regenerated axons, resulted in 87% of numerical recovery. Differentiation into vascular cell lineage (pericyte and endothelial cells were also observed. A significant tetanic tension recovery (over 90% of downstream muscles following electrical stimulation of the sciatic nerve (at upper portion of the gap was also achieved. In contrast, Sk-DN/29+ cells were completely eliminated during the first 4 weeks, but relatively higher numerical (83% vs. 41% in axon and functional (80% vs. 60% in tetanus recovery than control were observed. Noteworthy, significant increase in the formation of vascular networks in the conduit during the early stage (first 2 weeks of recovery was observed in both groups with the expression of key factors (mRNA and protein levels, suggesting the paracrine effects to angiogenesis. These results suggested that the

A Long-Gap Peripheral Nerve Injury Therapy Using Human Skeletal Muscle-Derived Stem Cells (Sk-SCs): An Achievement of Significant Morphological, Numerical and Functional Recovery.

Science.gov (United States)

Tamaki, Tetsuro; Hirata, Maki; Nakajima, Nobuyuki; Saito, Kosuke; Hashimoto, Hiroyuki; Soeda, Shuichi; Uchiyama, Yoshiyasu; Watanabe, Masahiko

2016-01-01

Losses in vital functions of the somatic motor and sensory nervous system are induced by severe long-gap peripheral nerve transection injury. In such cases, autologous nerve grafts are the gold standard treatment, despite the unavoidable sacrifice of other healthy functions, whereas the prognosis is not always favorable. Here, we use human skeletal muscle-derived stem cells (Sk-SCs) to reconstitute the function after long nerve-gap injury. Muscles samples were obtained from the amputated legs from 9 patients following unforeseen accidents. The Sk-SCs were isolated using conditioned collagenase solution, and sorted as CD34+/45- (Sk-34) and CD34-/45-/29+ (Sk-DN/29+) cells. Cells were separately cultured/expanded under optimal conditions for 2 weeks, then injected into the athymic nude mice sciatic nerve long-gap model (7-mm) bridging an acellular conduit. After 8-12 weeks, active cell engraftment was observed only in the Sk-34 cell transplanted group, showing preferential differentiation into Schwann cells and perineurial/endoneurial cells, as well as formation of the myelin sheath and perineurium/endoneurium surrounding regenerated axons, resulted in 87% of numerical recovery. Differentiation into vascular cell lineage (pericyte and endothelial cells) were also observed. A significant tetanic tension recovery (over 90%) of downstream muscles following electrical stimulation of the sciatic nerve (at upper portion of the gap) was also achieved. In contrast, Sk-DN/29+ cells were completely eliminated during the first 4 weeks, but relatively higher numerical (83% vs. 41% in axon) and functional (80% vs. 60% in tetanus) recovery than control were observed. Noteworthy, significant increase in the formation of vascular networks in the conduit during the early stage (first 2 weeks) of recovery was observed in both groups with the expression of key factors (mRNA and protein levels), suggesting the paracrine effects to angiogenesis. These results suggested that the human Sk

Stem-cell-derived products: an FDA update.

Science.gov (United States)

Moos, Malcolm

2008-12-01

The therapeutic potential of products derived from stem cells of various types has prompted increasing research and development and public attention. Initiation of human clinical trials in the not-too-distant future is now a realistic possibility. It is, therefore, important to weigh the potential benefits against known, theoretical and totally unsuspected risks in light of current knowledge to ensure that subjects participating in these trials are afforded the most reasonable balance possible between potential risks and potential benefits. There are no apparent differences in fundamental, qualitative biological characteristics between stem-cell-derived products and other cellular therapies regulated by the United States Food and Drug Administration (FDA). Existing authorities can, therefore, be applied. Nevertheless, these products do have properties that require careful evaluation.

Myocardial regeneration potential of adipose tissue-derived stem cells

Energy Technology Data Exchange (ETDEWEB)

Bai, Xiaowen, E-mail: baixw01@yahoo.com [Department of Molecular Pathology, The University of Texas M.D. Anderson Cancer Center, 1515 Holcombe, Houston, TX 77030 (United States); Alt, Eckhard, E-mail: ealt@mdanderson.org [Department of Molecular Pathology, The University of Texas M.D. Anderson Cancer Center, 1515 Holcombe, Houston, TX 77030 (United States)

2010-10-22

Research highlights: {yields} Various tissue resident stem cells are receiving tremendous attention from basic scientists and clinicians and hold great promise for myocardial regeneration. {yields} For practical reasons, human adipose tissue-derived stem cells are attractive stem cells for future clinical application in repairing damaged myocardium. {yields} This review summarizes the characteristics of cultured and freshly isolated stem cells obtained from adipose tissue, their myocardial regeneration potential and the, underlying mechanisms, and safety issues. -- Abstract: Various tissue resident stem cells are receiving attention from basic scientists and clinicians as they hold promise for myocardial regeneration. For practical reasons, adipose tissue-derived stem cells (ASCs) are attractive cells for clinical application in repairing damaged myocardium based on the following advantages: abundant adipose tissue in most patients and easy accessibility with minimally invasive lipoaspiration procedure. Several recent studies have demonstrated that both cultured and freshly isolated ASCs could improve cardiac function in animal model of myocardial infarction. The mechanisms underlying the beneficial effect of ASCs on myocardial regeneration are not fully understood. Growing evidence indicates that transplantation of ASCs improve cardiac function via the differentiation into cardiomyocytes and vascular cells, and through paracrine pathways. Paracrine factors secreted by injected ASCs enhance angiogenesis, reduce cell apoptosis rates, and promote neuron sprouts in damaged myocardium. In addition, Injection of ASCs increases electrical stability of the injured heart. Furthermore, there are no reported cases of arrhythmia or tumorigenesis in any studies regarding myocardial regeneration with ASCs. This review summarizes the characteristics of both cultured and freshly isolated stem cells obtained from adipose tissue, their myocardial regeneration potential, and the

Myocardial regeneration potential of adipose tissue-derived stem cells

International Nuclear Information System (INIS)

Bai, Xiaowen; Alt, Eckhard

2010-01-01

Research highlights: → Various tissue resident stem cells are receiving tremendous attention from basic scientists and clinicians and hold great promise for myocardial regeneration. → For practical reasons, human adipose tissue-derived stem cells are attractive stem cells for future clinical application in repairing damaged myocardium. → This review summarizes the characteristics of cultured and freshly isolated stem cells obtained from adipose tissue, their myocardial regeneration potential and the, underlying mechanisms, and safety issues. -- Abstract: Various tissue resident stem cells are receiving attention from basic scientists and clinicians as they hold promise for myocardial regeneration. For practical reasons, adipose tissue-derived stem cells (ASCs) are attractive cells for clinical application in repairing damaged myocardium based on the following advantages: abundant adipose tissue in most patients and easy accessibility with minimally invasive lipoaspiration procedure. Several recent studies have demonstrated that both cultured and freshly isolated ASCs could improve cardiac function in animal model of myocardial infarction. The mechanisms underlying the beneficial effect of ASCs on myocardial regeneration are not fully understood. Growing evidence indicates that transplantation of ASCs improve cardiac function via the differentiation into cardiomyocytes and vascular cells, and through paracrine pathways. Paracrine factors secreted by injected ASCs enhance angiogenesis, reduce cell apoptosis rates, and promote neuron sprouts in damaged myocardium. In addition, Injection of ASCs increases electrical stability of the injured heart. Furthermore, there are no reported cases of arrhythmia or tumorigenesis in any studies regarding myocardial regeneration with ASCs. This review summarizes the characteristics of both cultured and freshly isolated stem cells obtained from adipose tissue, their myocardial regeneration potential, and the underlying

Regenerative capacity of old muscle stem cells declines without significant accumulation of DNA damage.

Directory of Open Access Journals (Sweden)

Wendy Cousin

Full Text Available The performance of adult stem cells is crucial for tissue homeostasis but their regenerative capacity declines with age, leading to failure of multiple organs. In skeletal muscle this failure is manifested by the loss of functional tissue, the accumulation of fibrosis, and reduced satellite cell-mediated myogenesis in response to injury. While recent studies have shown that changes in the composition of the satellite cell niche are at least in part responsible for the impaired function observed with aging, little is known about the effects of aging on the intrinsic properties of satellite cells. For instance, their ability to repair DNA damage and the effects of a potential accumulation of DNA double strand breaks (DSBs on their regenerative performance remain unclear. This work demonstrates that old muscle stem cells display no significant accumulation of DNA DSBs when compared to those of young, as assayed after cell isolation and in tissue sections, either in uninjured muscle or at multiple time points after injury. Additionally, there is no significant difference in the expression of DNA DSB repair proteins or globally assayed DNA damage response genes, suggesting that not only DNA DSBs, but also other types of DNA damage, do not significantly mark aged muscle stem cells. Satellite cells from DNA DSB-repair-deficient SCID mice do have an unsurprisingly higher level of innate DNA DSBs and a weakened recovery from gamma-radiation-induced DNA damage. Interestingly, they are as myogenic in vitro and in vivo as satellite cells from young wild type mice, suggesting that the inefficiency in DNA DSB repair does not directly correlate with the ability to regenerate muscle after injury. Overall, our findings suggest that a DNA DSB-repair deficiency is unlikely to be a key factor in the decline in muscle regeneration observed upon aging.

Mechanosensing of matrix by stem cells: From matrix heterogeneity, contractility, and the nucleus in pore-migration to cardiogenesis and muscle stem cells in vivo.

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Smith, Lucas; Cho, Sangkyun; Discher, Dennis E

2017-11-01

Stem cells are particularly 'plastic' cell types that are induced by various cues to become specialized, tissue-functional lineages by switching on the expression of specific gene programs. Matrix stiffness is among the cues that multiple stem cell types can sense and respond to. This seminar-style review focuses on mechanosensing of matrix elasticity in the differentiation or early maturation of a few illustrative stem cell types, with an intended audience of biologists and physical scientists. Contractile forces applied by a cell's acto-myosin cytoskeleton are often resisted by the extracellular matrix and transduced through adhesions and the cytoskeleton ultimately into the nucleus to modulate gene expression. Complexity is added by matrix heterogeneity, and careful scrutiny of the evident stiffness heterogeneity in some model systems resolves some controversies concerning matrix mechanosensing. Importantly, local stiffness tends to dominate, and 'durotaxis' of stem cells toward stiff matrix reveals a dependence of persistent migration on myosin-II force generation and also rigid microtubules that confer directionality. Stem and progenitor cell migration in 3D can be further affected by matrix porosity as well as stiffness, with nuclear size and rigidity influencing niche retention and fate choices. Cell squeezing through rigid pores can even cause DNA damage and genomic changes that contribute to de-differentiation toward stem cell-like states. Contraction of acto-myosin is the essential function of striated muscle, which also exhibit mechanosensitive differentiation and maturation as illustrated in vivo by beating heart cells and by the regenerative mobilization of skeletal muscle stem cells. Copyright © 2017 Elsevier Ltd. All rights reserved.

Adipose tissue-derived stem cells in oral mucosa tissue engineering ...

African Journals Online (AJOL)

Jane

2011-10-10

Oct 10, 2011 ... stem cells (ADSCs) may play an important role in this field. In this research ..... Adipose tissue is derived from embryonic mesodermal precursors and .... Clonogenic multipotent stem cells in human adipose tissue differentiate ...

Labeling and Imaging Mesenchymal Stem Cells with Quantum Dots

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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...

BMP signaling regulates satellite cell-dependent postnatal muscle growth.

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Stantzou, Amalia; Schirwis, Elija; Swist, Sandra; Alonso-Martin, Sonia; Polydorou, Ioanna; Zarrouki, Faouzi; Mouisel, Etienne; Beley, Cyriaque; Julien, Anaïs; Le Grand, Fabien; Garcia, Luis; Colnot, Céline; Birchmeier, Carmen; Braun, Thomas; Schuelke, Markus; Relaix, Frédéric; Amthor, Helge

2017-08-01

Postnatal growth of skeletal muscle largely depends on the expansion and differentiation of resident stem cells, the so-called satellite cells. Here, we demonstrate that postnatal satellite cells express components of the bone morphogenetic protein (BMP) signaling machinery. Overexpression of noggin in postnatal mice (to antagonize BMP ligands), satellite cell-specific knockout of Alk3 (the gene encoding the BMP transmembrane receptor) or overexpression of inhibitory SMAD6 decreased satellite cell proliferation and accretion during myofiber growth, and ultimately retarded muscle growth. Moreover, reduced BMP signaling diminished the adult satellite cell pool. Abrogation of BMP signaling in satellite cell-derived primary myoblasts strongly diminished cell proliferation and upregulated the expression of cell cycle inhibitors p21 and p57 In conclusion, these results show that BMP signaling defines postnatal muscle development by regulating satellite cell-dependent myofiber growth and the generation of the adult muscle stem cell pool. © 2017. Published by The Company of Biologists Ltd.

Derivation of Stromal (Skeletal and Mesenchymal) Stem-Like Cells from Human Embryonic Stem Cells

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Harkness, Linda; Abdallah, Basem M.; Elsafadi, Mona; Al-Nbaheen, May S.; Aldahmash, Abdullah; Kassem, Moustapha

2012-01-01

Derivation of bone forming cells (osteoblasts) from human embryonic stem cells (hESCs) is a prerequisite for their use in clinical applications. However, there is no standard protocol for differentiating hESCs into osteoblastic cells. The aim of this study was to identify the emergence of a human stromal (mesenchymal and skeletal) stem cell (hMSC)-like population, known to be osteoblastic cell precursors and to test their osteoblastic differentiation capacity in ex vivo cultures and in vivo. We cultured hESCs in a feeder-free environment using serum replacement and as suspension aggregates (embryoid bodies; hEBs). Over a 20 day developmental period, the hEBs demonstrated increasing enrichment for cells expressing hMSC markers: CD29, CD44, CD63, CD56, CD71, CD73, CD105, CD106, and CD166 as revealed by immunohistochemical staining and flow cytometry (fluorescence-activated cell sorting) analysis. Ex vivo differentiation of hEBs using bone morphogenic protein 2 (BMP2) combined with standard osteoblast induction medium led to weak osteoblastic induction. Conversely, subcutaneous in vivo implantation of day 20 hEBs in immune deficient mice, mixed with hydroxyapatite/tricalcium phosphate (HA/TCP) as an osteoconductive scaffold, revealed bone and cartilage, and fibrous tissue elements after 8 weeks. These tissues were of human origin and there was no evidence of differentiation to nonmesodermal tissues. hEBs implanted in the absence of HA/TCP formed vacuolated tissue containing glandular, fibrous and muscle-like tissue elements. Conversely, implantation of undifferentiated hESCs resulted in the formation of a teratoma containing a mixture of endodermal, mesodermal, and ectodermal tissues. Our study demonstrates that hMSC-like cells can be obtained from hESCs and they can be induced to form skeletal tissues in vivo when combined with HA/TCP. These findings are relevant for tissue engineering and suggest that differentiated hEBs can provide an unlimited source for

Delivery of Placenta-Derived Mesenchymal Stem Cells Ameliorates Ischemia Induced Limb Injury by Immunomodulation

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Bo Zhang

2014-11-01

Full Text Available Background: Peripheral artery disease (PAD is a major health burden in the world. Stem cell-based therapy has emerged as an attractive treatment option in regenerative medicine. In this study, we sought to test the hypothesis that stem cell-based therapy can ameliorate ischemia induced limb injury. Methods: We isolated mesenchymal stem cells derived from human placentas (PMSCs and intramuscularly transplanted them into injured hind limbs. Treatment with PMSCs reduced acute muscle fibers apoptosis induced by ischemia. Results: PMSC treatment significantly enhanced regeneration of the injured hind limb by reducing fibrosis and enhancing running capacity when the animals were subjected to treadmill training. Mechanistically, injected PMSCs can modulate acute inflammatory responses by reducing neutrophil and macrophage infiltration following limb ischemia. ELISA assays further confirmed that PMSC treatment can also reduce pro-inflammatory cytokines, TNF-α and IL-6, and enhance anti-inflammatory cytokine, IL-10 at the injury sites. Conclusion: Taken together, our results demonstrated that PMSCs can be a potential effective therapy for treatment of PAD via immunomodulation.

Embryonic stem cell-like cells derived from adult human testis

NARCIS (Netherlands)

Mizrak, S. C.; Chikhovskaya, J. V.; Sadri-Ardekani, H.; van Daalen, S.; Korver, C. M.; Hovingh, S. E.; Roepers-Gajadien, H. L.; Raya, A.; Fluiter, K.; de Reijke, Th M.; de la Rosette, J. J. M. C. H.; Knegt, A. C.; Belmonte, J. C.; van der Veen, F.; de rooij, D. G.; Repping, S.; van Pelt, A. M. M.

2010-01-01

Given the significant drawbacks of using human embryonic stem (hES) cells for regenerative medicine, the search for alternative sources of multipotent cells is ongoing. Studies in mice have shown that multipotent ES-like cells can be derived from neonatal and adult testis. Here we report the

Regeneration of articular cartilage by adipose tissue derived mesenchymal stem cells: perspectives from stem cell biology and molecular medicine.

Science.gov (United States)

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. Copyright © 2012 Wiley Periodicals, Inc.

Role of bone marrow-derived stem cells, renal progenitor cells and stem cell factor in chronic renal allograft nephropathy

OpenAIRE

Hayam Abdel Meguid El Aggan; Mona Abdel Kader Salem; Nahla Mohamed Gamal Farahat; Ahmad Fathy El-Koraie; Ghaly Abd Al-Rahim Mohammed Kotb

2013-01-01

Introduction: Chronic allograft nephropathy (CAN) is a poorly understood clinico-pathological entity associated with chronic allograft loss due to immunologic and non-immunologic causes. It remains the leading cause of late allograft loss. Bone marrow derived stem cells are undifferentiated cells typically characterized by their capacity for self renewal, ability to give rise to multiple differentiated cellular population, including hematopoietic (HSCs) and mesenchymal stem cells (MSCs). Char...

mTOR signaling plays a critical role in the defects observed in muscle-derived stem/progenitor cells isolated from a murine model of accelerated aging.

Science.gov (United States)

Takayama, Koji; Kawakami, Yohei; Lavasani, Mitra; Mu, Xiaodong; Cummins, James H; Yurube, Takashi; Kuroda, Ryosuke; Kurosaka, Masahiro; Fu, Freddie H; Robbins, Paul D; Niedernhofer, Laura J; Huard, Johnny

2017-07-01

Mice expressing reduced levels of ERCC1-XPF (Ercc1 -/Δ mice) demonstrate premature onset of age-related changes due to decreased repair of DNA damage. Muscle-derived stem/progenitor cells (MDSPCs) isolated from Ercc1 -/Δ mice have an impaired capacity for cell differentiation. The mammalian target of rapamycin (mTOR) is a critical regulator of cell growth in response to nutrient, hormone, and oxygen levels. Inhibition of the mTOR pathway extends the lifespan of several species. Here, we examined the role of mTOR in regulating the MDSPC dysfunction that occurs with accelerated aging. We show that mTOR signaling pathways are activated in Ercc1 -/Δ MDSPCs compared with wild-type (WT) MDSPCs. Additionally, inhibiting mTOR with rapamycin promoted autophagy and improved the myogenic differentiation capacity of the Ercc1 -/Δ MDSPCs. The percent of apoptotic and senescent cells in Ercc1 -/Δ MDSPC cultures was decreased upon mTOR inhibition. These results establish that mTOR signaling contributes to stem cell dysfunction and cell fate decisions in response to endogenous DNA damage. Therefore, mTOR represents a potential therapeutic target for improving defective, aged stem cells. © 2016 The Authors. Journal of Orthopaedic Research Published by Wiley Periodicals, Inc. on behalf of Orthopaedic Research Society. J Orthop Res 35:1375-1382, 2017. © 2016 The Authors. Journal of Orthopaedic Research Published by Wiley Periodicals, Inc. on behalf of Orthopaedic Research Society.

Mesenchymal Stem Cell-Derived Factors Restore Function to Human Frataxin-Deficient Cells.

Science.gov (United States)

Kemp, Kevin; Dey, Rimi; Cook, Amelia; Scolding, Neil; Wilkins, Alastair

2017-08-01

Friedreich's ataxia is an inherited neurological disorder characterised by mitochondrial dysfunction and increased susceptibility to oxidative stress. At present, no therapy has been shown to reduce disease progression. Strategies being trialled to treat Friedreich's ataxia include drugs that improve mitochondrial function and reduce oxidative injury. In addition, stem cells have been investigated as a potential therapeutic approach. We have used siRNA-induced knockdown of frataxin in SH-SY5Y cells as an in vitro cellular model for Friedreich's ataxia. Knockdown of frataxin protein expression to levels detected in patients with the disorder was achieved, leading to decreased cellular viability, increased susceptibility to hydrogen peroxide-induced oxidative stress, dysregulation of key anti-oxidant molecules and deficiencies in both cell proliferation and differentiation. Bone marrow stem cells are being investigated extensively as potential treatments for a wide range of neurological disorders, including Friedreich's ataxia. The potential neuroprotective effects of bone marrow-derived mesenchymal stem cells were therefore studied using our frataxin-deficient cell model. Soluble factors secreted by mesenchymal stem cells protected against cellular changes induced by frataxin deficiency, leading to restoration in frataxin levels and anti-oxidant defences, improved survival against oxidative stress and stimulated both cell proliferation and differentiation down the Schwann cell lineage. The demonstration that mesenchymal stem cell-derived factors can restore cellular homeostasis and function to frataxin-deficient cells further suggests that they may have potential therapeutic benefits for patients with Friedreich's ataxia.

Neural stem cell-derived exosomes mediate viral entry

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Sims B

2014-10-01

Full Text Available Brian Sims,1,2,* Linlin Gu,3,* Alexandre Krendelchtchikov,3 Qiana L Matthews3,4 1Division of Neonatology, Department of Pediatrics, 2Department of Cell, Developmental, and Integrative Biology, 3Division of Infectious Diseases, Department of Medicine, 4Center for AIDS Research, University of Alabama at Birmingham, Birmingham, AL, USA *These authors contributed equally to this work Background: Viruses enter host cells through interactions of viral ligands with cellular receptors. Viruses can also enter cells in a receptor-independent fashion. Mechanisms regarding the receptor-independent viral entry into cells have not been fully elucidated. Exosomal trafficking between cells may offer a mechanism by which viruses can enter cells.Methods: To investigate the role of exosomes on cellular viral entry, we employed neural stem cell-derived exosomes and adenovirus type 5 (Ad5 for the proof-of-principle study. Results: Exosomes significantly enhanced Ad5 entry in Coxsackie virus and adenovirus receptor (CAR-deficient cells, in which Ad5 only had very limited entry. The exosomes were shown to contain T-cell immunoglobulin mucin protein 4 (TIM-4, which binds phosphatidylserine. Treatment with anti-TIM-4 antibody significantly blocked the exosome-mediated Ad5 entry.Conclusion: Neural stem cell-derived exosomes mediated significant cellular entry of Ad5 in a receptor-independent fashion. This mediation may be hampered by an antibody specifically targeting TIM-4 on exosomes. This set of results will benefit further elucidation of virus/exosome pathways, which would contribute to reducing natural viral infection by developing therapeutic agents or vaccines. Keywords: neural stem cell-derived exosomes, adenovirus type 5, TIM-4, viral entry, phospholipids

Therapeutic effect of adipose-derived mesenchymal stem cells on radiation enteritis

International Nuclear Information System (INIS)

Chang Pengyu; Cui Shuang; Luo Jinghua; Qu Chao; Jiang Xin; Qu Yaqin; Dong Lihua

2014-01-01

Objective: To evaluate the therapeutic effect of adipose-derived mesenchymal stem cells on radiation enteritis. Methods: A total of 52 male Sprague-Dawley rats were used in the present study. Herein, 46 rats were randomly selected and irradiated with a dose of 15 Gy at their abdomens. Two hours post-irradiation, 23 rats were randomly selected and infused intraperitoneally with adipose-derived mesenchymal stem cells in passage 6 from young-female donor. The other 23 rats were intraperitoneally infused with PBS. The rest 6 rats were set as normal control. During the first 10 days post-irradiation, peripheral blood-samples from irradiated rats were harvested for testing the levels of IL-10 in serum using ELISA assay. Additionally, after isolating the thymic cells and peripheral blood mononuclear cells, the percentages of CD4/CD25/Foxp(3)-positive regulatory T cells in thymus and peripheral blood were tested by flow-cytometry. Finally, infiltration of inflammatory cells and deposition of collagens within irradiated small intestine were analyzed by H&E staining and Masson Trichrome staining, respectively. Based on the MPO-immunohistochemistry staining, the type of infiltrated cells was identified. The Kaplan-Meier method was used for analyzing the survival rate of irradiated rats. Results: During a period of 30 days post-irradiation, the irradiated rats receiving adipose-derived mesenchymal stem cells survived longer than those receiving PBS (t = 4.53, P < 0.05). Compared to the irradiated rats with PBS-treatment, adipose-derived mesenchymal stem cells could elevate the level of IL-10 in serum (7 d: t = 13.93, P < 0.05) and increase the percentages of CD4/CD25/Foxp(3)-positive regulatory T cells in both peripheral blood (3.5 d: t = 7.72, 7 d: t = 11.11, 10 d: t = 6.99, P < 0.05) and thymus (7 d: t = 16.17, 10 d: t = 12.12, P < 0.05). Moreover, infiltration of inflammatory cells and deposition of collagens within irradiated small intestine were mitigated by adipose-derived

Human dental pulp pluripotent-like stem cells promote wound healing and muscle regeneration.

Science.gov (United States)

Martínez-Sarrà, Ester; Montori, Sheyla; Gil-Recio, Carlos; Núñez-Toldrà, Raquel; Costamagna, Domiziana; Rotini, Alessio; Atari, Maher; Luttun, Aernout; Sampaolesi, Maurilio

2017-07-27

Dental pulp represents an easily accessible autologous source of adult stem cells. A subset of these cells, named dental pulp pluripotent-like stem cells (DPPSC), shows high plasticity and can undergo multiple population doublings, making DPPSC an appealing tool for tissue repair or maintenance. DPPSC were harvested from the dental pulp of third molars extracted from young patients. Growth factors released by DPPSC were analysed using antibody arrays. Cells were cultured in specific differentiation media and their endothelial, smooth and skeletal muscle differentiation potential was evaluated. The therapeutic potential of DPPSC was tested in a wound healing mouse model and in two genetic mouse models of muscular dystrophy (Scid/mdx and Sgcb-null Rag2-null γc-null). DPPSC secreted several growth factors involved in angiogenesis and extracellular matrix deposition and improved vascularisation in all three murine models. Moreover, DPPSC stimulated re-epithelialisation and ameliorated collagen deposition and organisation in healing wounds. In dystrophic mice, DPPSC engrafted in the skeletal muscle of both dystrophic murine models and showed integration in muscular fibres and vessels. In addition, DPPSC treatment resulted in reduced fibrosis and collagen content, larger cross-sectional area of type II fast-glycolytic fibres and infiltration of higher numbers of proangiogenic CD206 + macrophages. Overall, DPPSC represent a potential source of stem cells to enhance the wound healing process and slow down dystrophic muscle degeneration.

Patient-derived stem cells: pathways to drug discovery for brain diseases

Directory of Open Access Journals (Sweden)

Alan eMackay-Sim

2013-03-01

Full Text Available The concept of drug discovery through stem cell biology is based on technological developments whose genesis is now coincident. The first is automated cell microscopy with concurrent advances in image acquisition and analysis, known as high content screening (HCS. The second is patient-derived stem cells for modelling the cell biology of brain diseases. HCS has developed from the requirements of the pharmaceutical industry for high throughput assays to screen thousands of chemical compounds in the search for new drugs. HCS combines new fluorescent probes with automated microscopy and computational power to quantify the effects of compounds on cell functions. Stem cell biology has advanced greatly since the discovery of genetic reprogramming of somatic cells into induced pluripotent stem cells (iPSCs. There is now a rush of papers describing their generation from patients with various diseases of the nervous system. Although the majority of these have been genetic diseases, iPSCs have been generated from patients with complex diseases (schizophrenia and sporadic Parkinson’s disease. Some genetic diseases are also modelled in embryonic stem cells generated from blastocysts rejected during in vitro fertilisation. Neural stem cells have been isolated from post-mortem brain of Alzheimer’s patients and neural stem cells generated from biopsies of the olfactory organ of patients is another approach. These olfactory neurosphere-derived cells demonstrate robust disease-specific phenotypes in patients with schizophrenia and Parkinson’s disease. High content screening is already in use to find small molecules for the generation and differentiation of embryonic stem cells and induced pluripotent stem cells. The challenges for using stem cells for drug discovery are to develop robust stem cell culture methods that meet the rigorous requirements for repeatable, consistent quantities of defined cell types at the industrial scale necessary for high

A CREB-MPP7-AMOT Regulatory Axis Controls Muscle Stem Cell Expansion and Self-Renewal Competence

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Lydia Li

2017-10-01

Full Text Available Summary: Skeletal muscle regeneration requires resident muscle stem cells, termed satellite cells (SCs. SCs are largely quiescent during homeostasis yet become activated upon injury to supply myonuclei and self-renewed SCs. Molecular mechanisms underlying the competence of SCs to proliferate and self-renew in response to injury remain unclear. Here, we show that CREB activity establishes proliferative potential during SC quiescence. SCs with inhibited CREB activity remain quiescent and positioned in their niche, but upon injury, they cannot enter or maintain a proliferative state for expansion and self-renewal. We demonstrate mechanistically that Mpp7 is a CREB target and its functional mediator. MPP7 loss affects the level and sub-cellular localization of AMOT and YAP1 in quiescent SCs. Furthermore, MPP7 and AMOT are required for YAP1 nuclear accumulation, and the three are individually required for a proliferative state in myoblasts. We propose that the CREB-MPP7-AMOT-YAP1 axis establishes the competence of quiescent SCs to expand and self-renew, thereby preserving stem cell function. : Satellite cells are quiescent muscle stem cells that have the ability to regenerate muscles after injury. Li and Fan reveal an MPP7-AMOT-YAP1 regulatory axis that acts downstream of CREB to instill satellite cell competence. They also show how this regulatory axis prepares satellite cells for robust muscle regeneration after injury.

Evaluation of the osteogenic differentiation of gingiva-derived stem cells grown on culture plates or in stem cell spheroids: Comparison of two- and three-dimensional cultures.

Science.gov (United States)

Lee, Sung-Il; Ko, Youngkyung; Park, Jun-Beom

2017-09-01

Three-dimensional cell culture systems provide a convenient in vitro model for the study of complex cell-cell and cell-matrix interactions in the absence of exogenous substrates. The current study aimed to evaluate the osteogenic differentiation potential of gingiva-derived stem cells cultured in two-dimensional or three-dimensional systems. To the best of our knowledge, the present study is the first to compare the growth of gingiva-derived stem cells in monolayer culture to a three-dimensional culture system with microwells. For three-dimensional culture, gingiva-derived stem cells were isolated and seeded into polydimethylsiloxane-based concave micromolds. Alkaline phosphatase activity and alizarin red S staining assays were then performed to evaluate osteogenesis and the degree of mineralization, respectively. Stem cell spheroids had a significantly increased level of alkaline phosphatase activity and mineralization compared with cells from the two-dimensional culture. In addition, an increase in mineralized deposits was observed with an increase in the loading cell number. The results of present study indicate that gingiva-derived stem cell spheroids exhibit an increased osteogenic potential compared with stem cells from two-dimensional culture. This highlights the potential of three-dimensional culture systems using gingiva-derived stem cells for regenerative medicine applications requiring stem cells with osteogenic potential.

Generation of a transplantable erythropoietin-producer derived from human mesenchymal stem cells.

Science.gov (United States)

Yokoo, Takashi; Fukui, Akira; Matsumoto, Kei; Ohashi, Toya; Sado, Yoshikazu; Suzuki, Hideaki; Kawamura, Tetsuya; Okabe, Masataka; Hosoya, Tatsuo; Kobayashi, Eiji

2008-06-15

Differentiation of autologous stem cells into functional transplantable tissue for organ regeneration is a promising regenerative therapeutic approach for cancer, diabetes, and many human diseases. Yet to be established, however, is differentiation into tissue capable of producing erythropoietin (EPO), which has a critical function in anemia. We report a novel EPO-producing organ-like structure (organoid) derived from human mesenchymal stem cells. Using our previously established relay culture system, a human mesenchymal stem cell-derived, human EPO-competent organoid was established in rat omentum. The organoid-derived levels of human EPO increased in response to anemia induced by rapid blood withdrawal. In addition, the presence of an organoid in rats suppressed for native (rat) EPO production enhanced recovery from anemia when compared with control animals lacking the organoid. Together these results confirmed the generation of a stem cell-derived organoid that is capable of producing EPO and sensitive to physiological regulation.

Synovium-derived stem cells: a tissue-specific stem cell for cartilage engineering and regeneration.

Science.gov (United States)

Jones, Brendan A; Pei, Ming

2012-08-01

Articular cartilage is difficult to heal once injury or disease occurs. Autologous chondrocyte transplantation is a biological treatment with good prognosis, but donor site morbidity and limited cell source are disadvantages. Currently, mesenchymal stem cells (MSCs) are a promising approach for cartilage regeneration. Despite there being various sources, the best candidate for cartilage regeneration is the one with the greatest chondrogenic potential and the least hypertrophic differentiation. These properties are able to insure that the regenerated tissue is hyaline cartilage of high quality. This review article will summarize relevant literature to justify synovium-derived stem cells (SDSCs) as a tissue-specific stem cell for chondrogenesis by comparing synovium and cartilage with respect to anatomical location and functional structure, comparing the growth characterization and chondrogenic capacity of SDSCs and MSCs, evaluating the application of SDSCs in regenerative medicine and diseases, and discussing potential future directions.

Human Pluripotent Stem Cell-Derived Cardiomyocytes as Research and Therapeutic Tools

Directory of Open Access Journals (Sweden)

Ivana Acimovic

2014-01-01

Full Text Available Human pluripotent stem cells (hPSCs, namely, embryonic stem cells (ESCs and induced pluripotent stem cells (iPSCs, with their ability of indefinite self-renewal and capability to differentiate into cell types derivatives of all three germ layers, represent a powerful research tool in developmental biology, for drug screening, disease modelling, and potentially cell replacement therapy. Efficient differentiation protocols that would result in the cell type of our interest are needed for maximal exploitation of these cells. In the present work, we aim at focusing on the protocols for differentiation of hPSCs into functional cardiomyocytes in vitro as well as achievements in the heart disease modelling and drug testing on the patient-specific iPSC-derived cardiomyocytes (iPSC-CMs.

Differential gene expression profiling of human adipose stem cells differentiating into smooth muscle-like cells by TGFβ1/BMP4

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Elçin, Ayşe Eser; Parmaksiz, Mahmut; Dogan, Arin; Seker, Sukran; Durkut, Serap; Dalva, Klara; Elçin, Yaşar Murat, E-mail: elcinmurat@gmail.com

2017-03-15

Regenerative repair of the vascular system is challenging from the perspectives of translational medicine and tissue engineering. There are fundamental hurdles in front of creating bioartificial arteries, which involve recaputilation of the three-layered structure under laboratory settings. Obtaining and maintaining smooth muscle characteristics is an important limitation, as the transdifferentiated cells fail to display mature phenotype. This study aims to shed light on the smooth muscle differentiation of human adipose stem cells (hASCs). To this end, we first acquired hASCs from lipoaspirate samples. Upon characterization, the cells were induced to differentiate into smooth muscle (SM)-like cells using a variety of inducer combinations. Among all, TGFβ1/BMP4 combination had the highest differentiation efficiency, based on immunohistochemical analyses. hSM-like cell samples were compared to hASCs and to the positive control, human coronary artery-smooth muscle cells (hCA-SMCs) through gene transcription profiling. Microarray findings revealed the activation of gene groups that function in smooth muscle differentiation, signaling pathways, extracellular modeling and cell proliferation. Our results underline the effectiveness of the growth factors and suggest some potential variables for detecting the SM-like cell characteristics. Evidence in transcriptome level was used to evaluate the TGFβ1/BMP4 combination as a previously unexplored effector for the smooth muscle differentiation of adipose stem cells. - Highlights: • Human adipose stem cells (hASCs) were isolated, characterized and cultured. • Growth factor combinations were evaluated for their effectiveness in differentiation using IHC. • hASCs were differentiated into smooth muscle (SM)-like cells using TGF-β1 and BMP4 combination. • Microarray analysis was performed for hASCs, SM-like cells and coronary artery-SMCs. • Microarray data was used to perform hierarchical clustering and interpretation

Generation of insulin-producing cells from gnotobiotic porcine skin-derived stem cells

International Nuclear Information System (INIS)

Yang, Ji Hoon; Lee, Sung Ho; Heo, Young Tae; Uhm, Sang Jun; Lee, Hoon Taek

2010-01-01

A major problem in the treatment of type 1 diabetes mellitus is the limited availability of alternative sources of insulin-producing cells for islet transplantation. In this study, we investigated the effect of bone morphogenetic protein 4 (BMP-4) treatments of gnotobiotic porcine skin-derived stem cells (gSDSCs) on their reprogramming and subsequent differentiation into insulin-producing cells (IPCs). We isolated SDSCs from the ear skin of a gnotobiotic pig. During the proliferation period, the cells expressed stem-cell markers Oct-4, Sox-2, and CD90; nestin expression also increased significantly. The cells could differentiate into IPCs after treatments with activin-A, glucagon-like peptide-1 (GLP-1), and nicotinamide. After 15 days in the differentiation medium, controlled gSDSCs began expressing endocrine progenitor genes and proteins (Ngn3, Neuro-D, PDX-1, NKX2.2, NKX6.1, and insulin). The IPCs showed increased insulin synthesis after glucose stimulation. The results indicate that stem cells derived from the skin of gnotobiotic pigs can differentiate into IPCs under the appropriate conditions in vitro. Our three-stage induction protocol could be applied without genetic modification to source IPCs from stem cells in the skin of patients with diabetes for autologous transplantation.

Generation of insulin-producing cells from gnotobiotic porcine skin-derived stem cells

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Yang, Ji Hoon; Lee, Sung Ho; Heo, Young Tae [Department of Bioscience and Biotechnology, Bio-Organ Research Center, Konkuk University, Seoul 143-701 (Korea, Republic of); Uhm, Sang Jun [Department of Animal Biotechnology, Bio-Organ Research Center, Konkuk University, Seoul 143-701 (Korea, Republic of); Lee, Hoon Taek, E-mail: htl3675@konkuk.ac.kr [Department of Animal Biotechnology, Bio-Organ Research Center, Konkuk University, Seoul 143-701 (Korea, Republic of)

2010-07-09

A major problem in the treatment of type 1 diabetes mellitus is the limited availability of alternative sources of insulin-producing cells for islet transplantation. In this study, we investigated the effect of bone morphogenetic protein 4 (BMP-4) treatments of gnotobiotic porcine skin-derived stem cells (gSDSCs) on their reprogramming and subsequent differentiation into insulin-producing cells (IPCs). We isolated SDSCs from the ear skin of a gnotobiotic pig. During the proliferation period, the cells expressed stem-cell markers Oct-4, Sox-2, and CD90; nestin expression also increased significantly. The cells could differentiate into IPCs after treatments with activin-A, glucagon-like peptide-1 (GLP-1), and nicotinamide. After 15 days in the differentiation medium, controlled gSDSCs began expressing endocrine progenitor genes and proteins (Ngn3, Neuro-D, PDX-1, NKX2.2, NKX6.1, and insulin). The IPCs showed increased insulin synthesis after glucose stimulation. The results indicate that stem cells derived from the skin of gnotobiotic pigs can differentiate into IPCs under the appropriate conditions in vitro. Our three-stage induction protocol could be applied without genetic modification to source IPCs from stem cells in the skin of patients with diabetes for autologous transplantation.

Derivation of Stromal (Skeletal, Mesenchymal) Stem-like cells from Human Embryonic Stem Cells

DEFF Research Database (Denmark)

Mahmood, Amer; Harkness, Linda; Abdallah, Basem

2012-01-01

EBs using BMP2 (bone morphogenic protein 2) combined with standard osteoblast induction medium led to weak osteoblastic induction. Conversely, subcutaneous in vivo implantation of day 20 hEBs in immune deficient mice, mixed with hydroxyapatite/tricalcium phosphate (HA/TCP) as an osteoconductive scaffold......Derivation of bone forming cells (osteoblasts) from human embryonic stem cells (hESC) is a pre-requisite for their use in clinical applications. However, there is no standard protocol for differentiating hESC into osteoblastic cells. The aim of this study was to identify the emergence of a human...... stromal (mesenchymal, skeletal) stem cell (hMSC)-like population, known to be osteoblastic cell precursors and to test their osteoblastic differentiation capacity in ex vivo cultures and in vivo. We cultured hESC in a feeder-free environment using serum replacement and as suspension aggregates (embryoid...

Functional Differences in Engineered Myocardium from Embryonic Stem Cell-Derived versus Neonatal Cardiomyocytes

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Feinberg, Adam W.; Ripplinger, Crystal M.; van der Meer, Peter; Sheehy, Sean P.; Domian, Ibrahim; Chien, Kenneth R.; Parker, Kevin Kit

2013-01-01

Stem cell-derived cardiomyocytes represent unique tools for cell-and tissue-based regenerative therapies, drug discovery and safety, and studies of fundamental heart-failure mechanisms. However, the degree to which stem cell-derived cardiomyocytes compare to mature cardiomyocytes is often debated.

Age-Associated Methylation Suppresses SPRY1, Leading to a Failure of Re-quiescence and Loss of the Reserve Stem Cell Pool in Elderly Muscle

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Anne Bigot

2015-11-01

Full Text Available The molecular mechanisms by which aging affects stem cell number and function are poorly understood. Murine data have implicated cellular senescence in the loss of muscle stem cells with aging. Here, using human cells and by carrying out experiments within a strictly pre-senescent division count, we demonstrate an impaired capacity for stem cell self-renewal in elderly muscle. We link aging to an increased methylation of the SPRY1 gene, a known regulator of muscle stem cell quiescence. Replenishment of the reserve cell pool was modulated experimentally by demethylation or siRNA knockdown of SPRY1. We propose that suppression of SPRY1 by age-associated methylation in humans inhibits the replenishment of the muscle stem cell pool, contributing to a decreased regenerative response in old age. We further show that aging does not affect muscle stem cell senescence in humans.

Endothelial juxtaposition of distinct adult stem cells activates angiogenesis signaling molecules in endothelial cells.

Science.gov (United States)

Mohammadi, Elham; Nassiri, Seyed Mahdi; Rahbarghazi, Reza; Siavashi, Vahid; Araghi, Atefeh

2015-12-01

Efficacy of therapeutic angiogenesis needs a comprehensive understanding of endothelial cell (EC) function and biological factors and cells that interplay with ECs. Stem cells are considered the key components of pro- and anti-angiogenic milieu in a wide variety of physiopathological states, and interactions of EC-stem cells have been the subject of controversy in recent years. In this study, the potential effects of three tissue-specific adult stem cells, namely rat marrow-derived mesenchymal stem cells (rBMSCs), rat adipose-derived stem cells (rADSCs) and rat muscle-derived satellite cells (rSCs), on the endothelial activation of key angiogenic signaling molecules, including VEGF, Ang-2, VEGFR-2, Tie-2, and Tie2-pho, were investigated. Human umbilical vein endothelial cells (HUVECs) and rat lung microvascular endothelial cells (RLMECs) were cocultured with the stem cells or incubated with the stem cell-derived conditioned media on Matrigel. Following HUVEC-stem cell coculture, CD31-positive ECs were flow sorted and subjected to western blotting to analyze potential changes in the expression of the pro-angiogenic signaling molecules. Elongation and co-alignment of the stem cells were seen along the EC tubes in the EC-stem cell cocultures on Matrigel, with cell-to-cell dye communication in the EC-rBMSC cocultures. Moreover, rBMSCs and rADSCs significantly improved endothelial tubulogenesis in both juxtacrine and paracrine manners. These two latter stem cells dynamically up-regulated VEGF, Ang-2, VREGR-2, and Tie-2 but down-regulated Tie2-pho and the Tie2-pho/Tie-2 ratio in HUVECs. Induction of pro-angiogenic signaling in ECs by marrow- and adipose-derived MSCs further indicates the significance of stem cell milieu in angiogenesis dynamics.

Adult hippocampus derived soluble factors induce a neuronal-like phenotype in mesenchymal stem cells.

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Rivera, Francisco J; Sierralta, Walter D; Minguell, Jose J; Aigner, Ludwig

2006-10-02

Bone marrow-derived mesenchymal stem cells (MSCs) are not restricted in their differentiation fate to cells of the mesenchymal lineage. They acquire a neural phenotype in vitro and in vivo after transplantation in the central nervous system. Here we investigated whether soluble factors derived from different brain regions are sufficient to induce a neuronal phenotype in MSCs. We incubated bone marrow-derived MSCs in conditioned medium (CM) derived from adult hippocampus (HCM), cortex (CoCM) or cerebellum (CeCM) and analyzed the cellular morphology and the expression of neuronal and glial markers. In contrast to muscle derived conditioned medium, which served as control, conditioned medium derived from the different brain regions induced a neuronal morphology and the expression of the neuronal markers GAP-43 and neurofilaments in MSCs. Hippocampus derived conditioned medium had the strongest activity. It was independent of NGF or BDNF; and it was restricted to the neuronal differentiation fate, since no induction of the astroglial marker GFAP was observed. The work indicates that soluble factors present in the brain are sufficient to induce a neuronal phenotype in MSCs.

One-step derivation of mesenchymal stem cell (MSC-like cells from human pluripotent stem cells on a fibrillar collagen coating.

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Yongxing Liu

Full Text Available Controlled differentiation of human embryonic stem cells (hESCs and induced pluripotent stem cells (iPSCs into cells that resemble adult mesenchymal stem cells (MSCs is an attractive approach to obtain a readily available source of progenitor cells for tissue engineering. The present study reports a new method to rapidly derive MSC-like cells from hESCs and hiPSCs, in one step, based on culturing the cells on thin, fibrillar, type I collagen coatings that mimic the structure of physiological collagen. Human H9 ESCs and HDFa-YK26 iPSCs were singly dissociated in the presence of ROCK inhibitor Y-27632, plated onto fibrillar collagen coated plates and cultured in alpha minimum essential medium (alpha-MEM supplemented with 10% fetal bovine serum, 50 uM magnesium L-ascorbic acid phosphate and 100 nM dexamethasone. While fewer cells attached on the collagen surface initially than standard tissue culture plastic, after culturing for 10 days, resilient colonies of homogenous spindle-shaped cells were obtained. Flow cytometric analysis showed that a high percentage of the derived cells expressed typical MSC surface markers including CD73, CD90, CD105, CD146 and CD166 and were negative as expected for hematopoietic markers CD34 and CD45. The MSC-like cells derived from pluripotent cells were successfully differentiated in vitro into three different lineages: osteogenic, chondrogenic, and adipogenic. Both H9 hES and YK26 iPS cells displayed similar morphological changes during the derivation process and yielded MSC-like cells with similar properties. In conclusion, this study demonstrates that bioimimetic, fibrillar, type I collagen coatings applied to cell culture plates can be used to guide a rapid, efficient derivation of MSC-like cells from both human ES and iPS cells.

Stem cells: progressions and applications in clinical medicine

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Ali Hosseini Bereshneh

2016-05-01

Full Text Available Stem cells are undifferentiated and multi pluripotent cells which can differentiate into a variety of mature cells and tissues such as nervous tissue, muscle tissue, epithelial tissue, skeletal tissue and etc. Stem cells from all different source have three unique features: 1 Proliferative capability: Stem cells are capable of self dividing and self renewing for long periods or more than six months at least that called immortalization. 2 Undifferentiated nature: It’s considered as one of the essential characteristics of stem cell, so it doesn't have any tissue-specific construction. 3 Differentiation to the different cells from all organs: This ability can Induced by tissue specific transcription factors. Because of that, they are so important in prevention and treatment of human disease. Depending on the sources from which they derive, they have different types which can be used to produce special cells and tissues. The most significant types of stem cells are; embryonic stem cells (ESCs which are derived from embryos, adult stem cells (ASCs which are derived from differentiated cells in a specific tissue, induced pluripotent stem cells (iPSs which are produced from adult differentiated cells that have been genetically reprogrammed to act resemble to an embryonic stem cell and cord blood stem cells which contains haematopoietic stem cells and derived from the umbilical cord after gestation. By providing a medium containing of special growth factor, it is possible to orientated stem cell differentiation pathway and gained certain cells from them. The important uses of stem cells includes damaged heart tissue cells improvements and bone tissue repairing, cancer treatment, damaged neurological and spinal tissue repairing, improving burns and injuries and the treatment of diabetes, infertility and spermatogenesis dysfunction. Furthermore, the application of them in gene therapy is an important issue in the modern medicine science due to the role

Adipose-derived mesenchymal stem cells promote cell proliferation and invasion of epithelial ovarian cancer

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Chu, Yijing; Tang, Huijuan; Guo, Yan; Guo, Jing; Huang, Bangxing; Fang, Fang; Cai, Jing, E-mail: caijingmmm@hotmail.com; Wang, Zehua, E-mail: zehuawang@163.net

2015-09-10

Adipose-derived mesenchymal stem cell (ADSC) is an important component of tumor microenvironment. However, whether ADSCs have a hand in ovarian cancer progression remains unclear. In this study, we investigated the impact of human ADSCs derived from the omentum of normal donors on human epithelial ovarian cancer (EOC) cells in vitro and in vivo. Direct and indirect co-culture models including ADSCs and human EOC cell lines were established and the effects of ADSCs on EOC cell proliferation were evaluated by EdU incorporation and flow cytometry. Transwell migration assays and detection of MMPs were performed to assess the invasion activity of EOC cells in vitro. Mouse models were established by intraperitoneal injection of EOC cells with or without concomitant ADSCs to investigate the role of ADSCs in tumor progression in vivo. We found that ADSCs significantly promoted proliferation and invasion of EOC cells in both direct and indirect co-culture assays. In addition, after co-culture with ADSCs, EOC cells secreted higher levels of matrix metalloproteinases (MMPs), and inhibition of MMP2 and MMP9 partially relieved the tumor-promoting effects of ADSCs in vitro. In mouse xenograft models, we confirmed that ADSCs promoted EOC growth and metastasis and elevated the expression of MMP2 and MMP9. Our findings indicate that omental ADSCs play a promotive role during ovarian cancer progression. - Highlights: • Omental adipose derived stem cells enhanced growth and invasion properties of ovarian cancer cells. • Adipose derived stem cells promoted the growth and metastasis of ovarian cancer in mice models. • Adipose derived stem cells promoted MMPs expression and secretion of ovarian cancer cells. • Elevated MMPs mediated the tumor promoting effects of ADSCs.

Adipose-derived mesenchymal stem cells promote cell proliferation and invasion of epithelial ovarian cancer

International Nuclear Information System (INIS)

Chu, Yijing; Tang, Huijuan; Guo, Yan; Guo, Jing; Huang, Bangxing; Fang, Fang; Cai, Jing; Wang, Zehua

2015-01-01

Adipose-derived mesenchymal stem cell (ADSC) is an important component of tumor microenvironment. However, whether ADSCs have a hand in ovarian cancer progression remains unclear. In this study, we investigated the impact of human ADSCs derived from the omentum of normal donors on human epithelial ovarian cancer (EOC) cells in vitro and in vivo. Direct and indirect co-culture models including ADSCs and human EOC cell lines were established and the effects of ADSCs on EOC cell proliferation were evaluated by EdU incorporation and flow cytometry. Transwell migration assays and detection of MMPs were performed to assess the invasion activity of EOC cells in vitro. Mouse models were established by intraperitoneal injection of EOC cells with or without concomitant ADSCs to investigate the role of ADSCs in tumor progression in vivo. We found that ADSCs significantly promoted proliferation and invasion of EOC cells in both direct and indirect co-culture assays. In addition, after co-culture with ADSCs, EOC cells secreted higher levels of matrix metalloproteinases (MMPs), and inhibition of MMP2 and MMP9 partially relieved the tumor-promoting effects of ADSCs in vitro. In mouse xenograft models, we confirmed that ADSCs promoted EOC growth and metastasis and elevated the expression of MMP2 and MMP9. Our findings indicate that omental ADSCs play a promotive role during ovarian cancer progression. - Highlights: • Omental adipose derived stem cells enhanced growth and invasion properties of ovarian cancer cells. • Adipose derived stem cells promoted the growth and metastasis of ovarian cancer in mice models. • Adipose derived stem cells promoted MMPs expression and secretion of ovarian cancer cells. • Elevated MMPs mediated the tumor promoting effects of ADSCs

Platelet-Rich Plasma Derived Growth Factors Contribute to Stem Cell Differentiation in Musculoskeletal Regeneration

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Yun Qian

2017-10-01

Full Text Available Stem cell treatment and platelet-rich plasma (PRP therapy are two significant issues in regenerative medicine. Stem cells such as bone marrow mesenchymal stem cells, adipose-derived stem cells and periodontal ligament stem cells can be successfully applied in the field of tissue regeneration. PRP, a natural product isolated from whole blood, can secrete multiple growth factors (GFs for regulating physiological activities. These GFs can stimulate proliferation and differentiation of different stem cells in injury models. Therefore, combination of both agents receives wide expectations in regenerative medicine, especially in bone, cartilage and tendon repair. In this review, we thoroughly discussed the interaction and underlying mechanisms of PRP derived GFs with stem cells, and assessed their functions in cell differentiation for musculoskeletal regeneration.

Platelet-rich plasma derived growth factors contribute to stem cell differentiation in musculoskeletal regeneration

Science.gov (United States)

Qian, Yun; Han, Qixin; Chen, Wei; Song, Jialin; Zhao, Xiaotian; Ouyang, Yuanming; Yuan, Weien; Fan, Cunyi

2017-10-01

Stem cell treatment and platelet-rich plasma (PRP) therapy are two significant issues in regenerative medicine. Stem cells such as bone marrow mesenchymal stem cells, adipose-derived stem cells and periodontal ligament stem cells can be successfully applied in the field of tissue regeneration. PRP, a natural product isolated from whole blood, can secrete multiple growth factors (GFs) for regulating physiological activities. These GFs can stimulate proliferation and differentiation of different stem cells in injury models. Therefore, combination of both agents receives wide expectations in regenerative medicine, especially in bone, cartilage and tendon repair. In this review, we thoroughly discussed the interaction and underlying mechanisms of platelet-rich plasma derived growth factors with stem cells, and assessed their functions in cell differentiation for musculoskeletal regeneration.

PITX2 Enhances the Regenerative Potential of Dystrophic Skeletal Muscle Stem Cells.

Science.gov (United States)

Vallejo, Daniel; Hernández-Torres, Francisco; Lozano-Velasco, Estefanía; Rodriguez-Outeiriño, Lara; Carvajal, Alejandra; Creus, Carlota; Franco, Diego; Aránega, Amelia Eva

2018-04-10

Duchenne muscular dystrophy (DMD), one of the most lethal genetic disorders, involves progressive muscle degeneration resulting from the absence of DYSTROPHIN. Lack of DYSTROPHIN expression in DMD has critical consequences in muscle satellite stem cells including a reduced capacity to generate myogenic precursors. Here, we demonstrate that the c-isoform of PITX2 transcription factor modifies the myogenic potential of dystrophic-deficient satellite cells. We further show that PITX2c enhances the regenerative capability of mouse DYSTROPHIN-deficient satellite cells by increasing cell proliferation and the number of myogenic committed cells, but importantly also increasing dystrophin-positive (revertant) myofibers by regulating miR-31. These PITX2-mediated effects finally lead to improved muscle function in dystrophic (DMD/mdx) mice. Our studies reveal a critical role for PITX2 in skeletal muscle repair and may help to develop therapeutic strategies for muscular disorders. Copyright © 2018 The Author(s). Published by Elsevier Inc. All rights reserved.

Deriving Dorsal Spinal Sensory Interneurons from Human Pluripotent Stem Cells

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Sandeep Gupta

2018-02-01

Full Text Available Summary: Cellular replacement therapies for neurological conditions use human embryonic stem cell (hESC- or induced pluripotent stem cell (hiPSC-derived neurons to replace damaged or diseased populations of neurons. For the spinal cord, significant progress has been made generating the in-vitro-derived motor neurons required to restore coordinated movement. However, there is as yet no protocol to generate in-vitro-derived sensory interneurons (INs, which permit perception of the environment. Here, we report on the development of a directed differentiation protocol to derive sensory INs for both hESCs and hiPSCs. Two developmentally relevant factors, retinoic acid in combination with bone morphogenetic protein 4, can be used to generate three classes of sensory INs: the proprioceptive dI1s, the dI2s, and mechanosensory dI3s. Critical to this protocol is the competence state of the neural progenitors, which changes over time. This protocol will facilitate developing cellular replacement therapies to reestablish sensory connections in injured patients. : In this article, Gupta and colleagues describe a robust protocol to derive spinal dorsal sensory interneurons from human pluripotent stem cells using the sequential addition of RA and BMP4. They find that neural progenitors must be in the correct competence state to respond to RA/BMP4 as dorsalizing signals. This competence state changes over time and determines the efficiency of the protocol. Keywords: spinal cord, neurons, sensory interneurons, proprioception, mechanosensation, human embryonic stem cells, induced pluripotent stem cells, directed differentiation, primate spinal cord, mouse spinal cord

In vitro cardiomyogenic potential of human umbilical vein-derived mesenchymal stem cells

International Nuclear Information System (INIS)

Kadivar, Mehdi; Khatami, Shohreh; Mortazavi, Yousef; Shokrgozar, Mohammad Ali; Taghikhani, Mohammad; Soleimani, Masoud

2006-01-01

Cardiomyocyte loss in the ischemically injured human heart often leads to irreversible defects in cardiac function. Recently, cellular cardiomyoplasty with mesenchymal stem cells, which are multipotent cells with the ability to differentiate into specialized cells under appropriate stimuli, has emerged as a new approach for repairing damaged myocardium. In the present study, the potential of human umbilical cord-derived mesenchymal stem cells to differentiate into cells with characteristics of cardiomyocyte was investigated. Mesenchymal stem cells were isolated from endothelial/subendothelial layers of the human umbilical cords using a method similar to that of human umbilical vein endothelial cell isolation. Isolated cells were characterized by transdifferentiation ability to adipocytes and osteoblasts, and also with flow cytometry analysis. After treatment with 5-azacytidine, the human umbilical cord-derived mesenchymal stem cells were morphologically transformed into cardiomyocyte-like cells and expressed cardiac differentiation markers. During the differentiation, cells were monitored by a phase contrast microscope and their morphological changes were demonstrated. Immunostaining of the differentiated cells for sarcomeric myosin (MF20), desmin, cardiac troponin I, and sarcomeric α-actinin was positive. RT-PCR analysis showed that these differentiated cells express cardiac-specific genes. Transmission electron microscopy revealed a cardiomyocyte-like ultrastructure and typical sarcomers. These observations confirm that human umbilical cord-derived mesenchymal stem cells can be chemically transformed into cardiomyocytes and can be considered as a source of cells for cellular cardiomyoplasty

Gene expression profiling and secretome analysis differentiate adult-derived human liver stem/progenitor cells and human hepatic stellate cells.

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Silvia Berardis

Full Text Available Adult-derived human liver stem/progenitor cells (ADHLSC are obtained after primary culture of the liver parenchymal fraction. The cells are of fibroblastic morphology and exhibit a hepato-mesenchymal phenotype. Hepatic stellate cells (HSC derived from the liver non-parenchymal fraction, present a comparable morphology as ADHLSC. Because both ADHLSC and HSC are described as liver stem/progenitor cells, we strived to extensively compare both cell populations at different levels and to propose tools demonstrating their singularity. ADHLSC and HSC were isolated from the liver of four different donors, expanded in vitro and followed from passage 5 until passage 11. Cell characterization was performed using immunocytochemistry, western blotting, flow cytometry, and gene microarray analyses. The secretion profile of the cells was evaluated using Elisa and multiplex Luminex assays. Both cell types expressed α-smooth muscle actin, vimentin, fibronectin, CD73 and CD90 in accordance with their mesenchymal origin. Microarray analysis revealed significant differences in gene expression profiles. HSC present high expression levels of neuronal markers as well as cytokeratins. Such differences were confirmed using immunocytochemistry and western blotting assays. Furthermore, both cell types displayed distinct secretion profiles as ADHLSC highly secreted cytokines of therapeutic and immuno-modulatory importance, like HGF, interferon-γ and IL-10. Our study demonstrates that ADHLSC and HSC are distinct liver fibroblastic cell populations exhibiting significant different expression and secretion profiles.

Identification of rabbit annulus fibrosus-derived stem cells.

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Chen Liu

Full Text Available Annulus fibrosus (AF injuries can lead to substantial deterioration of intervertebral disc (IVD which characterizes degenerative disc disease (DDD. However, treatments for AF repair/regeneration remain challenging due to the intrinsic heterogeneity of AF tissue at cellular, biochemical, and biomechanical levels. In this study, we isolated and characterized a sub-population of cells from rabbit AF tissue which formed colonies in vitro and could self-renew. These cells showed gene expression of typical surface antigen molecules characterizing mesenchymal stem cells (MSCs, including CD29, CD44, and CD166. Meanwhile, they did not express negative markers of MSCs such as CD4, CD8, and CD14. They also expressed Oct-4, nucleostemin, and SSEA-4 proteins. Upon induced differentiation they showed typical osteogenesis, chondrogenesis, and adipogenesis potential. Together, these AF-derived colony-forming cells possessed clonogenicity, self-renewal, and multi-potential differentiation capability, the three criteria characterizing MSCs. Such AF-derived stem cells may potentially be an ideal candidate for DDD treatments using cell therapies or tissue engineering approaches.

Paracrine Mechanisms of Intravenous Bone Marrow-Derived Mononuclear Stem Cells in Chronic Ischemic Stroke

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Ashu Bhasin

2016-10-01

Full Text Available Background: The emerging role of stem cell technology and transplantation has helped scientists to study their potential role in neural repair and regeneration. The fate of stem cells is determined by their niche, consisting of surrounding cells and the secreted trophic growth factors. This interim report evaluates the safety, feasibility and efficacy (if any of bone marrow-derived mononuclear stem cells (BM-MNC in chronic ischemic stroke by studying the release of serum vascular endothelial growth factor (VEGF and brain-derived neurotrophic growth factor (BDNF. Methods: Twenty stroke patients and 20 age-matched healthy controls were recruited with the following inclusion criteria: 3 months to 1.5 years from the index event, Medical Research Council (MRC grade of hand muscles of at least 2, Brunnstrom stage 2-5, conscious, and comprehendible. They were randomized to one group receiving autologous BM-MNC (mean 60-70 million and to another group receiving saline infusion (placebo. All patients were administered a neuromotor rehabilitation regime for 8 weeks. Clinical assessments [Fugl Meyer scale (FM, modified Barthel index (mBI, MRC grade, Ashworth tone scale] were carried out and serum VEGF and BDNF levels were assessed at baseline and at 8 weeks. Results: No serious adverse events were observed during the study. There was no statistically significant clinical improvement between the groups (FM: 95% CI 15.2-5.35, p = 0.25; mBI: 95% CI 14.3-4.5, p = 0.31. VEGF and BDNF expression was found to be greater in group 1 compared to group 2 (VEGF: 442.1 vs. 400.3 pg/ml, p = 0.67; BDNF: 21.3 vs. 19.5 ng/ml without any statistically significant difference. Conclusion: Autologous mononuclear stem cell infusion is safe and tolerable by chronic ischemic stroke patients. The released growth factors (VEGF and BDNF in the microenvironment could be due to the paracrine hypothesis of stem cell niche and neurorehabilitation regime.

Role of bone marrow-derived stem cells, renal progenitor cells and stem cell factor in chronic renal allograft nephropathy

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Hayam Abdel Meguid El Aggan

2013-09-01

Full Text Available Introduction: Chronic allograft nephropathy (CAN is a poorly understood clinico-pathological entity associated with chronic allograft loss due to immunologic and non-immunologic causes. It remains the leading cause of late allograft loss. Bone marrow derived stem cells are undifferentiated cells typically characterized by their capacity for self renewal, ability to give rise to multiple differentiated cellular population, including hematopoietic (HSCs and mesenchymal stem cells (MSCs. Characterization of HSCs includes their multipotency, expression of typical surface markers such as CD34 and CD45, while characterization of MSC includes their multipotency, expression of typical surface markers such as CD90 and CD105, and the absence of hemopoietic lineage markers. Aim & methods: The aim of the present work was to study the role of bone marrow-derived HSCs and MSCs, renal progenitor cells and SCF in chronic renal allograft nephropathy in relation to renal hemodynamics and histopathological changes. We studied 30 patients with kidney transplantation for more than 6 months, divided into 15 patients with stable serum creatinine and 15 patients who developed CAN. Detection of HSCs and MSCs in the peripheral blood using flow cytometry via detection of CD34, CD45, CD117 and CD106, as well as immunohistochemical detection of CD34, CD133, VEGF and αSMA in transplanted kidney biopsies of patients with CAN were done. Results: There was a significant increase in the levels of SCF, number of peripheral blood HSCs and MSCs in both transplanted patient groups than the controls and they were higher in patients of group Ia than patients of group Ib, (F = 39.73, P < 0.001, (F = 13.28, P < 0.001, (F = 11.94, P < 0.001, respectively and this was accompanied by evident expression of markers of renal repair. Conclusion: Stem cells might have a role in renal regeneration in CAN and this may pave the way toward the use of stem cells in correction of CAN. KEYWORDS

β-catenin functions pleiotropically in differentiation and tumorigenesis in mouse embryo-derived stem cells.

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Noriko Okumura

Full Text Available The canonical Wnt/β-catenin signaling pathway plays a crucial role in the maintenance of the balance between proliferation and differentiation throughout embryogenesis and tissue homeostasis. β-Catenin, encoded by the Ctnnb1 gene, mediates an intracellular signaling cascade activated by Wnt. It also plays an important role in the maintenance of various types of stem cells including adult stem cells and cancer stem cells. However, it is unclear if β-catenin is required for the derivation of mouse embryo-derived stem cells. Here, we established β-catenin-deficient (β-cat(Δ/Δ mouse embryo-derived stem cells and showed that β-catenin is not essential for acquiring self-renewal potential in the derivation of mouse embryonic stem cells (ESCs. However, teratomas formed from embryo-derived β-cat(Δ/Δ ESCs were immature germ cell tumors without multilineage differentiated cell types. Re-expression of functional β-catenin eliminated their neoplastic, transformed phenotype and restored pluripotency, thereby rescuing the mutant ESCs. Our findings demonstrate that β-catenin has pleiotropic effects in ESCs; it is required for the differentiation of ESCs and prevents them from acquiring tumorigenic character. These results highlight β-catenin as the gatekeeper in differentiation and tumorigenesis in ESCs.

Molecular characterisation of stromal populations derived from human embryonic stem cells

DEFF Research Database (Denmark)

Harkness, L.; Twine, N. A.; Abu Dawud, R.

2015-01-01

Human bone marrow-derived stromal (skeletal) stem cells (BM-hMSC) are being employed in an increasing number of clinical trials for tissue regeneration. A limiting factor for their clinical use is the inability to obtain sufficient cell numbers. Human embryonic stem cells (hESC) can provide an un...

Establishment and cryopreservation of a giant panda skeletal muscle-derived cell line.

Science.gov (United States)

Yu, Fang-Jian; Zeng, Chang-Jun; Zhang, Yan; Wang, Cheng-Dong; Xiong, Tie-Yi; Fang, Sheng-Guo; Zhang, He-Min

2015-06-01

The giant panda Ailuropoda melanoleuca is an endangered species and is a symbol for wildlife conservation. Although efforts have been made to protect this rare and endangered species through breeding and conservative biology, the long-term preservation of giant panda genome resources (gametes, tissues, organs, genomic libraries, etc.) is still a practical option. In this study, the giant panda skeletal muscle-derived cell line was successfully established via primary explants culture and cryopreservation techniques. The population doubling time of giant panda skeletal cells was approximately 33.8 h, and this population maintained a high cell viability before and after cryopreservation (95.6% and 90.7%, respectively). The two skeletal muscle-specific genes SMYD1 and MYF6 were expressed and detected by RT-PCR in the giant panda skeletal muscle-derived cell line. Karyotyping analysis revealed that the frequencies of giant panda skeletal muscle cells showing a chromosome number of 2n=42 ranged from 90.6∼94.2%. Thus, the giant panda skeletal muscle-derived cell line provides a vital resource and material platform for further studies and is likely to be useful for the protection of this rare and endangered species.

PEDF-derived peptide promotes skeletal muscle regeneration through its mitogenic effect on muscle progenitor cells.

Science.gov (United States)

Ho, Tsung-Chuan; Chiang, Yi-Pin; Chuang, Chih-Kuang; Chen, Show-Li; Hsieh, Jui-Wen; Lan, Yu-Wen; Tsao, Yeou-Ping

2015-08-01

In response injury, intrinsic repair mechanisms are activated in skeletal muscle to replace the damaged muscle fibers with new muscle fibers. The regeneration process starts with the proliferation of satellite cells to give rise to myoblasts, which subsequently differentiate terminally into myofibers. Here, we investigated the promotion effect of pigment epithelial-derived factor (PEDF) on muscle regeneration. We report that PEDF and a synthetic PEDF-derived short peptide (PSP; residues Ser(93)-Leu(112)) induce satellite cell proliferation in vitro and promote muscle regeneration in vivo. Extensively, soleus muscle necrosis was induced in rats by bupivacaine, and an injectable alginate gel was used to release the PSP in the injured muscle. PSP delivery was found to stimulate satellite cell proliferation in damaged muscle and enhance the growth of regenerating myofibers, with complete regeneration of normal muscle mass by 2 wk. In cell culture, PEDF/PSP stimulated C2C12 myoblast proliferation, together with a rise in cyclin D1 expression. PEDF induced the phosphorylation of ERK1/2, Akt, and STAT3 in C2C12 myoblasts. Blocking the activity of ERK, Akt, or STAT3 with pharmacological inhibitors attenuated the effects of PEDF/PSP on the induction of C2C12 cell proliferation and cyclin D1 expression. Moreover, 5-bromo-2'-deoxyuridine pulse-labeling demonstrated that PEDF/PSP stimulated primary rat satellite cell proliferation in myofibers in vitro. In summary, we report for the first time that PSP is capable of promoting the regeneration of skeletal muscle. The signaling mechanism involves the ERK, AKT, and STAT3 pathways. These results show the potential utility of this PEDF peptide for muscle regeneration. Copyright © 2015 the American Physiological Society.

Procedures for Derivation and Characterisation of Human Embryonic Stem Cells from Odense, Denmark

DEFF Research Database (Denmark)

Harkness, Linda; Kassem, Moustapha

2012-01-01

In 1998, a development occurred in stem cell biology with the fi rst report of the derivation of a human embryonic stem cell (hESC) line. Since then a number of techniques have been used to derive and characterise hESCs. Here, we describe the derivation methods used by our laboratory for isolatio...

Conversion of adipose-derived stem cells into natural killer-like cells with anti-tumor activities in nude mice.

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Hongxiu Ning

Full Text Available Efforts to develop peripheral blood-derived nature killer (NK cells into therapeutic products have been hampered by these cells' low abundance and histoincompatibility. On the other hand, derivation of NK-like cells from more abundant cell sources such as embryonic stem cells (ESCs and umbilical cord blood (UCB requires the selection of rare CD34+ cells. Thus, we sought to convert adipose-derived stem cells (ADSCs, which are abundant and natively CD34+, into NK-like cells. When grown in hematopoietic induction medium, ADSCs formed sphere clusters and expressed hematopoietic markers CD34, CD45, and KDR. Further induction in NK cell-specific medium resulted in a population of cells that expressed NK cell marker CD56, and thus termed ADSC-NK. Alternatively, the hematopoietically induced ADSCs were transduced with NK cell-specific transcription factor E4BP4 prior to induction in NK cell-specific medium. This latter population of cells, termed ADSC-NKE, expressed CD56 and additional NK cell markers such as CD16, CD94, CD158, CD314, FasL, and NKp46. ADSC-NKE was as potent as NK leukemia cell NKL in killing breast cancer cell MCF7 and prostate cancer cells DU145, PC3, LnCap, DuPro, C4-2 and CWR22, but exhibited no killing activity toward normal endothelial and smooth muscle cells. In nude mice test ADSC-NKE was able to significantly delay the progression of tumors formed by MCF7 and PC3. When injected into immunocompetent rats, ADSC-NKE was detectable in bone marrow and spleen for at least 5 weeks. Together, these results suggest that ADSCs can be converted into NK-like cells with anti-tumor activities.

Comparison of multiple transcriptomes exposes unified and divergent features of quiescent and activated skeletal muscle stem cells

DEFF Research Database (Denmark)

Pietrosemoli, Natalia; Mella, Sébastien; Yennek, Siham

2017-01-01

of identifying quiescent markers. Here, we focused on the quiescent cell state and generated new transcriptome profiles that include subfractionations of adult satellite cell populations, and an artificially induced prenatal quiescent state, to identify core signatures for quiescent and proliferating. Methods......Background: Skeletal muscle satellite (stem) cells are quiescent in adult mice and can undergo multiple rounds of proliferation and self-renewal following muscle injury. Several labs have profiled transcripts of myogenic cells during the developmental and adult myogenesis with the aim...... with true in vivo quiescence from those that are first responding genes due to disruption of the stem cell niche....

Research on human placenta-derived mesenchymal stem cells ...

African Journals Online (AJOL)

Research on human placenta-derived mesenchymal stem cells transfected with pIRES2-EGFP-VEGF165 using liposome. ... African Journal of Biotechnology. Journal Home · ABOUT THIS JOURNAL · Advanced Search · Current Issue ...

Purification of human induced pluripotent stem cell-derived neural precursors using magnetic activated cell sorting.

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Rodrigues, Gonçalo M C; Fernandes, Tiago G; Rodrigues, Carlos A V; Cabral, Joaquim M S; Diogo, Maria Margarida

2015-01-01

Neural precursor (NP) cells derived from human induced pluripotent stem cells (hiPSCs), and their neuronal progeny, will play an important role in disease modeling, drug screening tests, central nervous system development studies, and may even become valuable for regenerative medicine treatments. Nonetheless, it is challenging to obtain homogeneous and synchronously differentiated NP populations from hiPSCs, and after neural commitment many pluripotent stem cells remain in the differentiated cultures. Here, we describe an efficient and simple protocol to differentiate hiPSC-derived NPs in 12 days, and we include a final purification stage where Tra-1-60+ pluripotent stem cells (PSCs) are removed using magnetic activated cell sorting (MACS), leaving the NP population nearly free of PSCs.

Ex Vivo Gene Therapy Using Human Mesenchymal Stem Cells to Deliver Growth Factors in the Skeletal Muscle of a Familial ALS Rat Model.

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Suzuki, Masatoshi; Svendsen, Clive N

2016-01-01

Therapeutic protein and molecule delivery to target sites by transplanted human stem cells holds great promise for ex vivo gene therapy. Our group has demonstrated the therapeutic benefits of ex vivo gene therapy targeting the skeletal muscles in a transgenic rat model of familial amyotrophic lateral sclerosis (ALS). We used human mesenchymal stem cells (hMSCs) and genetically modified them to release neuroprotective growth factors such as glial cell line-derived neurotrophic factor (GDNF) and vascular endothelial growth factor (VEGF). Intramuscular growth factor delivery via hMSCs can enhance neuromuscular innervation and motor neuron survival in a rat model of ALS (SOD1(G93A) transgenic rats). Here, we describe the protocol of ex vivo delivery of growth factors via lentiviral vector-mediated genetic modification of hMSCs and hMSC transplantation into the skeletal muscle of a familial ALS rat model.

Bone Marrow Stromal Cells Generate Muscle Cells and Repair Muscle Degeneration

Science.gov (United States)

Dezawa, Mari; Ishikawa, Hiroto; Itokazu, Yutaka; Yoshihara, Tomoyuki; Hoshino, Mikio; Takeda, Shin-ichi; Ide, Chizuka; Nabeshima, Yo-ichi

2005-07-01

Bone marrow stromal cells (MSCs) have great potential as therapeutic agents. We report a method for inducing skeletal muscle lineage cells from human and rat general adherent MSCs with an efficiency of 89%. Induced cells differentiated into muscle fibers upon transplantation into degenerated muscles of rats and mdx-nude mice. The induced population contained Pax7-positive cells that contributed to subsequent regeneration of muscle upon repetitive damage without additional transplantation of cells. These MSCs represent a more ready supply of myogenic cells than do the rare myogenic stem cells normally found in muscle and bone marrow.

Tumor cell-derived PDGF-B potentiates mouse mesenchymal stem cells-pericytes transition and recruitment through an interaction with NRP-1

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Haque Inamul

2010-08-01

Full Text Available Abstract Background New blood vessel formation, or angiogenic switch, is an essential event in the development of solid tumors and their metastatic growth. Tumor blood vessel formation and remodeling is a complex and multi-step processes. The differentiation and recruitment of mural cells including vascular smooth muscle cells and pericytes are essential steps in tumor angiogenesis. However, the role of tumor cells in differentiation and recruitment of mural cells has not yet been fully elucidated. This study focuses on the role of human tumor cells in governing the differentiation of mouse mesenchymal stem cells (MSCs to pericytes and their recruitment in the tumor angiogenesis process. Results We show that C3H/10T1/2 mouse embryonic mesenchymal stem cells, under the influence of different tumor cell-derived conditioned media, differentiate into mature pericytes. These differentiated pericytes, in turn, are recruited to bind with capillary-like networks formed by endothelial cells on the matrigel under in vitro conditions and recruited to bind with blood vessels on gel-foam under in vivo conditions. The degree of recruitment of pericytes into in vitro neo-angiogenesis is tumor cell phenotype specific. Interestingly, invasive cells recruit less pericytes as compared to non-invasive cells. We identified tumor cell-secreted platelet-derived growth factor-B (PDGF-B as a crucial factor controlling the differentiation and recruitment processes through an interaction with neuropilin-1 (NRP-1 in mesenchymal stem cells. Conclusion These new insights into the roles of tumor cell-secreted PDGF-B-NRP-1 signaling in MSCs-fate determination may help to develop new antiangiogenic strategies to prevent the tumor growth and metastasis and result in more effective cancer therapies.

A Bilayer Construct Controls Adipose-Derived Stem Cell Differentiation into Endothelial Cells and Pericytes without Growth Factor Stimulation

Science.gov (United States)

2011-01-01

A Bilayer Construct Controls Adipose-Derived Stem Cell Differentiation into Endothelial Cells and Pericytes Without Growth Factor Stimulation...Ph.D.3 This work describes the differentiation of adipose-derived mesenchymal stem cells (ASC) in a composite hy- drogel for use as a vascularized...tissue from a single population of ASC. This work underscores the importance of the extracellular matrix in controlling stem cell phenotype. It is our

Extracellular matrix-derived hydrogels for dental stem cell delivery.

Science.gov (United States)

Viswanath, Aiswarya; Vanacker, Julie; Germain, Loïc; Leprince, Julian G; Diogenes, Anibal; Shakesheff, Kevin M; White, Lisa J; des Rieux, Anne

2017-01-01

Decellularized mammalian extracellular matrices (ECM) have been widely accepted as an ideal substrate for repair and remodelling of numerous tissues in clinical and pre-clinical studies. Recent studies have demonstrated the ability of ECM scaffolds derived from site-specific homologous tissues to direct cell differentiation. The present study investigated the suitability of hydrogels derived from different source tissues: bone, spinal cord and dentine, as suitable carriers to deliver human apical papilla derived mesenchymal stem cells (SCAP) for spinal cord regeneration. Bone, spinal cord, and dentine ECM hydrogels exhibited distinct structural, mechanical, and biological characteristics. All three hydrogels supported SCAP viability and proliferation. However, only spinal cord and bone derived hydrogels promoted the expression of neural lineage markers. The specific environment of ECM scaffolds significantly affected the differentiation of SCAP to a neural lineage, with stronger responses observed with spinal cord ECM hydrogels, suggesting that site-specific tissues are more likely to facilitate optimal stem cell behavior for constructive spinal cord regeneration. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 319-328, 2017. © 2016 Wiley Periodicals, Inc.

Potential differentiation of islet-like cells from pregnant cow-derived placental stem cells.

Science.gov (United States)

Peng, Shao-Yu; Chou, Chien-Wen; Kuo, Yu-Hsuan; Shen, Perng-Chih; Shaw, S W Steven

2017-06-01

Type 1 diabetes is an autoimmune disease that destroys islet cells and results in insufficient insulin secretion by pancreatic β-cells. Islet transplantation from donors is an approach used for treating patients with diabetes; however, this therapy is difficult to implement because of the lack of donors. Nevertheless, several stem cells have the potential to differentiate from islet-like cells and enable insulin secretion for treating diabetes in animal models. For example, placenta is considered a waste material and can be harvested noninvasively during delivery without ethical or moral concerns. To date, the differentiation of islet-like cells from cow-derived placental stem cells (CPSCs) has yet to be demonstrated. The investigation of potential differentiation of islet-like cells from CPSCs was conducted by supplementation with nicotinamide, exendin-4, glucose, and poly-d-lysine and was detected through reverse transcription polymerase chain reaction, dithizone staining, and immunocytochemical methods. Our results indicated that CPSCs are established and express mesenchymal stem cell surface antigen markers, such as CD73, CD166, β-integrin, and Oct-4, but not hematopoietic stem cell surface antigen markers, such as CD45. After induction, the CPSCs successfully differentiated into islet-like cells. The CPSC-derived islet-like cells expressed islet cell development-related genes, such as insulin, glucagon, pax-4, Nkx6.1, pax-6, and Fox. Moreover, CPSC-derived islet-like cells can be stained with zinc ions, which are widely distributed in the islet cells and enable insulin secretion. Altogether, islet-like cells have the potential to be differentiated from CPSCs without gene manipulation, and can be used in diabetic animal models in the future for preclinical and drug testing trial investigations. Copyright © 2017. Published by Elsevier B.V.

Stem Cell Therapy: An emerging science

International Nuclear Information System (INIS)

Khan, Muhammad M.

2007-01-01

The research on stem cells is advancing knowledge about the development of an organism from a single cell and to how healthy cells replace damaged cells in adult organisms. Stem cell therapy is emerging rapidly nowadays as a technical tool for tissue repair and replacement. The purpose of this review to provide a framework of understanding for the challenges behind translating fundamental stem cell biology and its potential use into clinical therapies, also to give an overview on stem cell research to the scientists of Saudi Arabia in general. English language MEDLINE publications from 1980 through January 2007 for experimental, observational and clinical studies having relation with stem cells with different diseases were reviewed. Approximately 85 publications were reviewed based on the relevance, strength and quality of design and methods, 36 publications were selected for inclusion. Stem cells reside in a specific area of each tissue where they may remain undivided for several years until they are activated by disease or tissue injury. The embryonic stem cells are typically derived from four or five days old embryos and they are pluripotent. The adult tissues reported to contain stem cells brain, bone marrow, peripheral blood, blood vessels, skeletal muscle, skin and liver. The promise of stem cell therapies is an exciting one, but significant technical hurdles remain that will only be overcome through years of intensive research. (author)

Directed Differentiation of Human-Induced Pluripotent Stem Cells to Mesenchymal Stem Cells.

Science.gov (United States)

Lian, Qizhou; Zhang, Yuelin; Liang, Xiaoting; Gao, Fei; Tse, Hung-Fat

2016-01-01

Multipotent stromal cells, also known as mesenchymal stem cells (MSCs), possess great potential to generate a wide range of cell types including endothelial cells, smooth muscle cells, bone, cartilage, and lipid cells. This protocol describes in detail how to perform highly efficient, lineage-specific differentiation of human-induced pluripotent stem cells (iPSCs) with an MSCs fate. The approach uses a clinically compliant protocol with chemically defined media, feeder-free conditions, and a CD105 positive and CD24 negative selection to achieve a single cell-based MSCs derivation from differentiating human pluripotent cells in approximately 20 days. Cells generated with this protocol express typical MSCs surface markers and undergo adipogenesis, osteogenesis, and chondrogenesis similar to adult bone marrow-derived MSCs (BM-MSCs). Nonetheless, compared with adult BM-MSCs, iPSC-MSCs display a higher proliferative capacity, up to 120 passages, without obvious loss of self-renewal potential and constitutively express MSCs surface antigens. MSCs generated with this protocol have numerous applications, including expansion to large scale cell numbers for tissue engineering and the development of cellular therapeutics. This approach has been used to rescue limb ischemia, allergic disorders, and cigarette smoke-induced lung damage and to model mesenchymal and vascular disorders of Hutchinson-Gilford progeria syndrome (HGPS).

Human-induced pluripotent stem cell-derived cardiomyocytes from cardiac progenitor cells: effects of selective ion channel blockade.

Science.gov (United States)

Altomare, Claudia; Pianezzi, Enea; Cervio, Elisabetta; Bolis, Sara; Biemmi, Vanessa; Benzoni, Patrizia; Camici, Giovanni G; Moccetti, Tiziano; Barile, Lucio; Vassalli, Giuseppe

2016-12-01

Human-induced pluripotent stem cell (hiPSC)-derived cardiomyocytes are likely to revolutionize electrophysiological approaches to arrhythmias. Recent evidence suggests the somatic cell origin of hiPSCs may influence their differentiation potential. Owing to their cardiomyogenic potential, cardiac-stromal progenitor cells (CPCs) are an interesting cellular source for generation of hiPSC-derived cardiomyocytes. The effect of ionic current blockade in hiPSC-derived cardiomyocytes generated from CPCs has not been characterized yet. Human-induced pluripotent stem cell-derived cardiomyocytes were generated from adult CPCs and skin fibroblasts from the same individuals. The effect of selective ionic current blockade on spontaneously beating hiPSC-derived cardiomyocytes was assessed using multi-electrode arrays. Cardiac-stromal progenitor cells could be reprogrammed into hiPSCs, then differentiated into hiPSC-derived cardiomyocytes. Human-induced pluripotent stem cell-derived cardiomyocytes of cardiac origin showed higher upregulation of cardiac-specific genes compared with those of fibroblastic origin. Human-induced pluripotent stem cell-derived cardiomyocytes of both somatic cell origins exhibited sensitivity to tetrodotoxin, a blocker of Na +  current (I Na ), nifedipine, a blocker of L-type Ca 2+  current (I CaL ), and E4031, a blocker of the rapid component of delayed rectifier K +  current (I Kr ). Human-induced pluripotent stem cell-derived cardiomyocytes of cardiac origin exhibited sensitivity to JNJ303, a blocker of the slow component of delayed rectifier K +  current (I Ks ). In hiPSC-derived cardiomyocytes of cardiac origin, I Na , I CaL , I Kr , and I Ks were present as tetrodotoxin-, nifedipine-, E4031-, and JNJ303-sensitive currents, respectively. Although cardiac differentiation efficiency was improved in hiPSCs of cardiac vs. non-cardiac origin, no major functional differences were observed between hiPSC-derived cardiomyocytes of different somatic

Diploid, but not haploid, human embryonic stem cells can be derived from microsurgically repaired tripronuclear human zygotes

Science.gov (United States)

Fan, Yong; Li, Rong; Huang, Jin; Yu, Yang; Qiao, Jie

2013-01-01

Human embryonic stem cells have shown tremendous potential in regenerative medicine, and the recent progress in haploid embryonic stem cells provides new insights for future applications of embryonic stem cells. Disruption of normal fertilized embryos remains controversial; thus, the development of a new source for human embryonic stem cells is important for their usefulness. Here, we investigated the feasibility of haploid and diploid embryo reconstruction and embryonic stem cell derivation using microsurgically repaired tripronuclear human zygotes. Diploid and haploid zygotes were successfully reconstructed, but a large proportion of them still had a tripolar spindle assembly. The reconstructed embryos developed to the blastocyst stage, although the loss of chromosomes was observed in these zygotes. Finally, triploid and diploid human embryonic stem cells were derived from tripronuclear and reconstructed zygotes (from which only one pronucleus was removed), but haploid human embryonic stem cells were not successfully derived from the reconstructed zygotes when two pronuclei were removed. Both triploid and diploid human embryonic stem cells showed the general characteristics of human embryonic stem cells. These results indicate that the lower embryo quality resulting from abnormal spindle assembly contributed to the failure of the haploid embryonic stem cell derivation. However, the successful derivation of diploid embryonic stem cells demonstrated that microsurgical tripronuclear zygotes are an alternative source of human embryonic stem cells. In the future, improving spindle assembly will facilitate the application of triploid zygotes to the field of haploid embryonic stem cells. PMID:23255130

In vitro chondrogenic differentiation of human adipose-derived stem cells with silk scaffolds

Directory of Open Access Journals (Sweden)

Hyeon Joo Kim

2012-12-01

Full Text Available Human adipose-derived stem cells have shown chondrogenic differentiation potential in cartilage tissue engineering in combination with natural and synthetic biomaterials. In the present study, we hypothesized that porous aqueous-derived silk protein scaffolds would be suitable for chondrogenic differentiation of human adipose-derived stem cells. Human adipose-derived stem cells were cultured up to 6 weeks, and cell proliferation and chondrogenic differentiation were investigated and compared with those in conventional micromass culture. Cell proliferation, glycosaminoglycan, and collagen levels in aqueous-derived silk scaffolds were significantly higher than in micromass culture. Transcript levels of SOX9 and type II collagen were also upregulated in the cell–silk constructs at 6 weeks. Histological examination revealed that the pores of the silk scaffolds were filled with cells uniformly distributed. In addition, chondrocyte-specific lacunae formation was evident and distributed in the both groups. The results suggest the biodegradable and biocompatible three-dimensional aqueous-derived silk scaffolds provided an improved environment for chondrogenic differentiation compared to micromass culture.

Expanding the Tissue Toolbox : Deriving Colon Tissue from Human Pluripotent Stem Cells

NARCIS (Netherlands)

Bruens, Lotte; Snippert, Hugo J.G.

2017-01-01

Organoid technology holds great potential for disease modeling and regenerative medicine. In this issue of Cell Stem Cell, Múnera et al. (2017) establish the generation of pluripotent stem cell-derived colon organoids that upon transplantation in mice, resembling human colon to a large extent,

Characterization of human adipose-derived stem cells and expression of chondrogenic genes during induction of cartilage differentiation.

Science.gov (United States)

Hamid, Adila A; Idrus, Ruszymah Bt Hj; Saim, Aminuddin Bin; Sathappan, Somasumdaram; Chua, Kien-Hui

2012-01-01

Understanding the changes in chondrogenic gene expression that are involved in the differentiation of human adipose-derived stem cells to chondrogenic cells is important prior to using this approach for cartilage repair. The aims of the study were to characterize human adipose-derived stem cells and to examine chondrogenic gene expression after one, two, and three weeks of induction. Human adipose-derived stem cells at passage 4 were evaluated by flow cytometry to examine the expression of surface markers. These adipose-derived stem cells were tested for adipogenic and osteogenic differentiation capacity. Ribonucleic acid was extracted from the cells for quantitative polymerase chain reaction analysis to determine the expression levels of chondrogenic genes after chondrogenic induction. Human adipose-derived stem cells were strongly positive for the mesenchymal markers CD90, CD73, CD44, CD9, and histocompatibility antigen and successfully differentiated into adipogenic and osteogenic lineages. The human adipose-derived stem cells aggregated and formed a dense matrix after chondrogenic induction. The expression of chondrogenic genes (collagen type II, aggrecan core protein, collagen type XI, COMP, and ELASTIN) was significantly higher after the first week of induction. However, a significantly elevated expression of collagen type X was observed after three weeks of chondrogenic induction. Human adipose-derived stem cells retain stem cell characteristics after expansion in culture to passage 4 and serve as a feasible source of cells for cartilage regeneration. Chondrogenesis in human adipose-derived stem cells was most prominent after one week of chondrogenic induction.

Tissue-Engineered Vascular Rings from Human iPSC-Derived Smooth Muscle Cells

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Biraja C. Dash

2016-07-01

Full Text Available There is an urgent need for an efficient approach to obtain a large-scale and renewable source of functional human vascular smooth muscle cells (VSMCs to establish robust, patient-specific tissue model systems for studying the pathogenesis of vascular disease, and for developing novel therapeutic interventions. Here, we have derived a large quantity of highly enriched functional VSMCs from human induced pluripotent stem cells (hiPSC-VSMCs. Furthermore, we have engineered 3D tissue rings from hiPSC-VSMCs using a facile one-step cellular self-assembly approach. The tissue rings are mechanically robust and can be used for vascular tissue engineering and disease modeling of supravalvular aortic stenosis syndrome. Our method may serve as a model system, extendable to study other vascular proliferative diseases for drug screening. Thus, this report describes an exciting platform technology with broad utility for manufacturing cell-based tissues and materials for various biomedical applications.

Adipose Tissue-Derived Stem Cell Secreted IGF-1 Protects Myoblasts from the Negative Effect of Myostatin

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Sebastian Gehmert

2014-01-01

Full Text Available Myostatin, a TGF-β family member, is associated with inhibition of muscle growth and differentiation and might interact with the IGF-1 signaling pathway. Since IGF-1 is secreted at a bioactive level by adipose tissue-derived mesenchymal stem cells (ASCs, these cells (ASCs provide a therapeutic option for Duchenne Muscular Dystrophy (DMD. But the protective effect of stem cell secreted IGF-1 on myoblast under high level of myostatin remains unclear. In the present study murine myoblasts were exposed to myostatin under presence of ASCs conditioned medium and investigated for proliferation and apoptosis. The protective effect of IGF-1 was further examined by using IGF-1 neutralizing and receptor antibodies as well as gene silencing RNAi technology. MyoD expression was detected to identify impact of IGF-1 on myoblasts differentiation when exposed to myostatin. IGF-1 was accountable for 43.6% of the antiapoptotic impact and 48.8% for the proliferative effect of ASCs conditioned medium. Furthermore, IGF-1 restored mRNA and protein MyoD expression of myoblasts under risk. Beside fusion and transdifferentiation the beneficial effect of ASCs is mediated by paracrine secreted cytokines, particularly IGF-1. The present study underlines the potential of ASCs as a therapeutic option for Duchenne muscular dystrophy and other dystrophic muscle diseases.

Stromal vascular stem cell treatment decreases muscle fibrosis following chronic rotator cuff tear.

Science.gov (United States)

Gumucio, Jonathan P; Flood, Michael D; Roche, Stuart M; Sugg, Kristoffer B; Momoh, Adeyiza O; Kosnik, Paul E; Bedi, Asheesh; Mendias, Christopher L

2016-04-01

Rotator cuff injuries are associated with atrophy and fat infiltration into the muscle, commonly referred to as "fatty degeneration." As the poor function of chronically torn muscles may limit recovery after surgical repair, there is considerable interest in finding therapies to enhance muscle regeneration. Stromal vascular fraction stem cells (SVFCs) can improve muscle regeneration in other chronic injury states, and our objective was to evaluate the ability of SVFCs to reduce fibrosis and fat accumulation, and enhance muscle fibre specific force production after chronic rotator cuff tear. Chronic supraspinatus tears were induced in adult immunodeficient rats, and repaired one month following tear. Rats received vehicle control, or injections of 3 × 10(5) or 3 × 10(6) human SVFCs into supraspinatus muscles. Two weeks following repair, we detected donor human DNA and protein in SVFC treated muscles. There was a 40 % reduction in fibrosis in the treated groups compared to controls (p = 0.03 for 3 × 10(5), p = 0.04 for 3 × 10(6)), and no differences between groups for lipid content or force production were observed. As there has been much interest in the use of stem cell-based therapies in musculoskeletal regenerative medicine, the reduction in fibrosis and trend towards an improvement in single fiber contractility suggest that SVFCs may be beneficial to enhance the treatment and recovery of patients with chronic rotator cuff tears.

Three-Dimensional Human iPSC-Derived Artificial Skeletal Muscles Model Muscular Dystrophies and Enable Multilineage Tissue Engineering.

Science.gov (United States)

Maffioletti, Sara Martina; Sarcar, Shilpita; Henderson, Alexander B H; Mannhardt, Ingra; Pinton, Luca; Moyle, Louise Anne; Steele-Stallard, Heather; Cappellari, Ornella; Wells, Kim E; Ferrari, Giulia; Mitchell, Jamie S; Tyzack, Giulia E; Kotiadis, Vassilios N; Khedr, Moustafa; Ragazzi, Martina; Wang, Weixin; Duchen, Michael R; Patani, Rickie; Zammit, Peter S; Wells, Dominic J; Eschenhagen, Thomas; Tedesco, Francesco Saverio

2018-04-17

Generating human skeletal muscle models is instrumental for investigating muscle pathology and therapy. Here, we report the generation of three-dimensional (3D) artificial skeletal muscle tissue from human pluripotent stem cells, including induced pluripotent stem cells (iPSCs) from patients with Duchenne, limb-girdle, and congenital muscular dystrophies. 3D skeletal myogenic differentiation of pluripotent cells was induced within hydrogels under tension to provide myofiber alignment. Artificial muscles recapitulated characteristics of human skeletal muscle tissue and could be implanted into immunodeficient mice. Pathological cellular hallmarks of incurable forms of severe muscular dystrophy could be modeled with high fidelity using this 3D platform. Finally, we show generation of fully human iPSC-derived, complex, multilineage muscle models containing key isogenic cellular constituents of skeletal muscle, including vascular endothelial cells, pericytes, and motor neurons. These results lay the foundation for a human skeletal muscle organoid-like platform for disease modeling, regenerative medicine, and therapy development. Copyright © 2018 The Author(s). Published by Elsevier Inc. All rights reserved.

The differentiation potential of adipose tissue-derived mesenchymal stem cells into cell lineage related to male germ cells

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P. Bräunig

Full Text Available ABSTRACT The adipose tissue is a reliable source of Mesenchymal stem cells (MSCs showing a higher plasticity and transdifferentiation potential into multilineage cells. In the present study, adipose tissue-derived mesenchymal stem cells (AT-MSCs were isolated from mice omentum and epididymis fat depots. The AT-MSCs were initially compared based on stem cell surface markers and on the mesodermal trilineage differentiation potential. Additionally, AT-MSCs, from both sources, were cultured with differentiation media containing retinoic acid (RA and/or testicular cell-conditioned medium (TCC. The AT-MSCs expressed mesenchymal surface markers and differentiated into adipogenic, chondrogenic and osteogenic lineages. Only omentum-derived AT-MSCs expressed one important gene marker related to male germ cell lineages, after the differentiation treatment with RA. These findings reaffirm the importance of adipose tissue as a source of multipotent stromal-stem cells, as well as, MSCs source regarding differentiation purpose.

Platelet-Rich Plasma Derived Growth Factors Contribute to Stem Cell Differentiation in Musculoskeletal Regeneration

OpenAIRE

Yun Qian; Yun Qian; Qixin Han; Wei Chen; Wei Chen; Jialin Song; Jialin Song; Xiaotian Zhao; Yuanming Ouyang; Yuanming Ouyang; Weien Yuan; Cunyi Fan

2017-01-01

Stem cell treatment and platelet-rich plasma (PRP) therapy are two significant issues in regenerative medicine. Stem cells such as bone marrow mesenchymal stem cells, adipose-derived stem cells and periodontal ligament stem cells can be successfully applied in the field of tissue regeneration. PRP, a natural product isolated from whole blood, can secrete multiple growth factors (GFs) for regulating physiological activities. These GFs can stimulate proliferation and differentiation of differen...

Chitosan derived co-spheroids of neural stem cells and mesenchymal stem cells for neural regeneration.

Science.gov (United States)

Han, Hao-Wei; Hsu, Shan-Hui

2017-10-01

Chitosan has been considered as candidate biomaterials for neural applications. The effective treatment of neurodegeneration or injury to the central nervous system (CNS) is still in lack nowadays. Adult neural stem cells (NSCs) represents a promising cell source to treat the CNS diseases but they are limited in number. Here, we developed the core-shell spheroids of NSCs (shell) and mesenchymal stem cells (MSCs, core) by co-culturing cells on the chitosan surface. The NSCs in chitosan derived co-spheroids displayed a higher survival rate than those in NSC homo-spheroids. The direct interaction of NSCs with MSCs in the co-spheroids increased the Notch activity and differentiation tendency of NSCs. Meanwhile, the differentiation potential of MSCs in chitosan derived co-spheroids was significantly enhanced toward neural lineages. Furthermore, NSC homo-spheroids and NSC/MSC co-spheroids derived on chitosan were evaluated for their in vivo efficacy by the embryonic and adult zebrafish brain injury models. The locomotion activity of zebrafish receiving chitosan derived NSC homo-spheroids or NSC/MSC co-spheroids was partially rescued in both models. Meanwhile, the higher survival rate was observed in the group of adult zebrafish implanted with chitosan derived NSC/MSC co-spheroids as compared to NSC homo-spheroids. These evidences indicate that chitosan may provide an extracellular matrix-like environment to drive the interaction and the morphological assembly between NSCs and MSCs and promote their neural differentiation capacities, which can be used for neural regeneration. Copyright © 2017 Elsevier B.V. All rights reserved.

Anti-aging effects of vitamin C on human pluripotent stem cell-derived cardiomyocytes.

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Kim, Yoon Young; Ku, Seung-Yup; Huh, Yul; Liu, Hung-Ching; Kim, Seok Hyun; Choi, Young Min; Moon, Shin Yong

2013-10-01

Human pluripotent stem cells (hPSCs) have arisen as a source of cells for biomedical research due to their developmental potential. Stem cells possess the promise of providing clinicians with novel treatments for disease as well as allowing researchers to generate human-specific cellular metabolism models. Aging is a natural process of living organisms, yet aging in human heart cells is difficult to study due to the ethical considerations regarding human experimentation as well as a current lack of alternative experimental models. hPSC-derived cardiomyocytes (CMs) bear a resemblance to human cardiac cells and thus hPSC-derived CMs are considered to be a viable alternative model to study human heart cell aging. In this study, we used hPSC-derived CMs as an in vitro aging model. We generated cardiomyocytes from hPSCs and demonstrated the process of aging in both human embryonic stem cell (hESC)- and induced pluripotent stem cell (hiPSC)-derived CMs. Aging in hESC-derived CMs correlated with reduced membrane potential in mitochondria, the accumulation of lipofuscin, a slower beating pattern, and the downregulation of human telomerase RNA (hTR) and cell cycle regulating genes. Interestingly, the expression of hTR in hiPSC-derived CMs was not significantly downregulated, unlike in hESC-derived CMs. In order to delay aging, vitamin C was added to the cultured CMs. When cells were treated with 100 μM of vitamin C for 48 h, anti-aging effects, specifically on the expression of telomere-related genes and their functionality in aging cells, were observed. Taken together, these results suggest that hPSC-derived CMs can be used as a unique human cardiomyocyte aging model in vitro and that vitamin C shows anti-aging effects in this model.

Linking incomplete reprogramming to the improved pluripotency of murine embryonal carcinoma cell-derived pluripotent stem cells.

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Gang Chang

Full Text Available Somatic cell nuclear transfer (SCNT has been proved capable of reprogramming various differentiated somatic cells into pluripotent stem cells. Recently, induced pluripotent stem cells (iPS have been successfully derived from mouse and human somatic cells by the over-expression of a combination of transcription factors. However, the molecular mechanisms underlying the reprogramming mediated by either the SCNT or iPS approach are poorly understood. Increasing evidence indicates that many tumor pathways play roles in the derivation of iPS cells. Embryonal carcinoma (EC cells have the characteristics of both stem cells and cancer cells and thus they might be the better candidates for elucidating the details of the reprogramming process. Although previous studies indicate that EC cells cannot be reprogrammed into real pluripotent stem cells, the reasons for this remain unclear. Here, nuclei from mouse EC cells (P19 were transplanted into enucleated oocytes and pluripotent stem cells (P19 NTES cells were subsequently established. Interestingly, P19 NTES cells prolonged the development of tetraploid aggregated embryos compared to EC cells alone. More importantly, we found that the expression recovery of the imprinted H19 gene was dependent on the methylation state in the differential methylation region (DMR. The induction of Nanog expression, however, was independent of the promoter region DNA methylation state in P19 NTES cells. A whole-genome transcriptome analysis further demonstrated that P19 NTES cells were indeed the intermediates between P19 cells and ES cells and many interesting genes were uncovered that may be responsible for the failed reprogramming of P19 cells. To our knowledge, for the first time, we linked incomplete reprogramming to the improved pluripotency of EC cell-derived pluripotent stem cells. The candidate genes we discovered may be useful not only for understanding the mechanisms of reprogramming, but also for deciphering the

Characterization of human adipose-derived stem cells and expression of chondrogenic genes during induction of cartilage differentiation

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Adila A Hamid

2012-01-01

Full Text Available OBJECTIVES: Understanding the changes in chondrogenic gene expression that are involved in the differentiation of human adipose-derived stem cells to chondrogenic cells is important prior to using this approach for cartilage repair. The aims of the study were to characterize human adipose-derived stem cells and to examine chondrogenic gene expression after one, two, and three weeks of induction. MATERIALS AND METHODS: Human adipose-derived stem cells at passage 4 were evaluated by flow cytometry to examine the expression of surface markers. These adipose-derived stem cells were tested for adipogenic and osteogenic differentiation capacity. Ribonucleic acid was extracted from the cells for quantitative polymerase chain reaction analysis to determine the expression levels of chondrogenic genes after chondrogenic induction. RESULTS: Human adipose-derived stem cells were strongly positive for the mesenchymal markers CD90, CD73, CD44, CD9, and histocompatibility antigen and successfully differentiated into adipogenic and osteogenic lineages. The human adipose-derived stem cells aggregated and formed a dense matrix after chondrogenic induction. The expression of chondrogenic genes (collagen type II, aggrecan core protein, collagen type XI, COMP, and ELASTIN was significantly higher after the first week of induction. However, a significantly elevated expression of collagen type X was observed after three weeks of chondrogenic induction. CONCLUSION: Human adipose-derived stem cells retain stem cell characteristics after expansion in culture to passage 4 and serve as a feasible source of cells for cartilage regeneration. Chondrogenesis in human adiposederived stem cells was most prominent after one week of chondrogenic induction.

Efficient and Rapid Derivation of Primitive Neural Stem Cells and Generation of Brain Subtype Neurons From Human Pluripotent Stem Cells

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Yan, Yiping; Shin, Soojung; Jha, Balendu Shekhar; Liu, Qiuyue; Sheng, Jianting; Li, Fuhai; Zhan, Ming; Davis, Janine; Bharti, Kapil; Zeng, Xianmin; Rao, Mahendra; Malik, Nasir; Vemuri, Mohan C.

2013-01-01

This study developed a highly efficient serum-free pluripotent stem cell (PSC) neural induction medium that can induce human PSCs into primitive neural stem cells (NSCs) in 7 days, obviating the need for time-consuming, laborious embryoid body generation or rosette picking. This method of primitive NSC derivation sets the stage for the scalable production of clinically relevant neural cells for cell therapy applications in good manufacturing practice conditions.

Derivation of Rabbit Embryonic Stem Cells from Vitrified–Thawed Embryos

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Chen, Chien-Hong; Li, Yi; Hu, Yeshu; An, Li-You; Yang, Lan; Zhang, Jifeng; Chen, Y. Eugene

2015-01-01

Abstract The rabbit is a useful animal model for regenerative medicine. We previously developed pluripotent rabbit embryonic stem cell (rbESC) lines using fresh embryos. We also successfully cryopreserved rabbit embryos by vitrification. In the present work, we combined these two technologies to derive rbESCs using vitrified–thawed (V/T) embryos. We demonstrate that V/T blastocysts (BLs) can be used to derive pluripotent rbESCs with efficiencies comparable to those using fresh BLs. These ESCs are undistinguishable from the ones derived from fresh embryos. We tested the developmental capacity of rbESCs derived from V/T embryos by BL injection experiments and produced chimeric kits. Our work adds cryopreservation to the toolbox of rabbit stem cell research and applications and will greatly expand the available research materials for regenerative medicine in a clinically relevant animal model. PMID:26579970

Comparison of exosomes secreted by induced pluripotent stem cell-derived mesenchymal stem cells and synovial membrane-derived mesenchymal stem cells for the treatment of osteoarthritis.

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Zhu, Yu; Wang, Yuchen; Zhao, Bizeng; Niu, Xin; Hu, Bin; Li, Qing; Zhang, Juntao; Ding, Jian; Chen, Yunfeng; Wang, Yang

2017-03-09

Osteoarthritis (OA) is the most common joint disease worldwide. In the past decade, mesenchymal stem cells (MSCs) have been used widely for the treatment of OA. A potential mechanism of MSC-based therapies has been attributed to the paracrine secretion of trophic factors, in which exosomes may play a major role. In this study, we aimed to compare the effectiveness of exosomes secreted by synovial membrane MSCs (SMMSC-Exos) and exosomes secreted by induced pluripotent stem cell-derived MSCs (iMSC-Exos) on the treatment of OA. Induced pluripotent stem cell-derived MSCs and synovial membrane MSCs were characterized by flow cytometry. iMSC-Exos and SMMSC-Exos were isolated using an ultrafiltration method. Tunable resistive pulse-sensing analysis, transmission electron microscopy, and western blots were used to identify exosomes. iMSC-Exos and SMMSC-Exos were injected intra-articularly in a mouse model of collagenase-induced OA and the efficacy of exosome injections was assessed by macroscopic, histological, and immunohistochemistry analysis. We also evaluated the effects of iMSC-Exos and SMMSC-Exos on proliferation and migration of human chondrocytes by cell-counting and scratch assays, respectively. The majority of iMSC-Exos and SMMSC-Exos were approximately 50-150 nm in diameter and expressed CD9, CD63, and TSG101. The injection of iMSC-Exos and SMMSC-Exos both attenuated OA in the mouse OA model, but iMSC-Exos had a superior therapeutic effect compared with SMMSC-Exos. Similarly, chondrocyte migration and proliferation were stimulated by both iMSC-Exos and SMMSC-Exos, with iMSC-Exos exerting a stronger effect. The present study demonstrated that iMSC-Exos have a greater therapeutic effect on OA than SMMSC-Exos. Because autologous iMSCs are theoretically inexhaustible, iMSC-Exos may represent a novel therapeutic approach for the treatment of OA.

Evaluation of hollow fiber culture for large-scale production of mouse embryonic stem cell-derived hematopoietic stem cells.

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Nakano, Yu; Iwanaga, Shinya; Mizumoto, Hiroshi; Kajiwara, Toshihisa

2018-03-03

Hematopoietic stem cells (HSCs) have the ability to differentiate into all types of blood cells and can be transplanted to treat blood disorders. However, it is difficult to obtain HSCs in large quantities because of the shortage of donors. Recent efforts have focused on acquiring HSCs by differentiation of pluripotent stem cells. As a conventional differentiation method of pluripotent stem cells, the formation of embryoid bodies (EBs) is often employed. However, the size of EBs is limited by depletion of oxygen and nutrients, which prevents them from being efficient for the production of HSCs. In this study, we developed a large-scale hematopoietic differentiation approach for mouse embryonic stem (ES) cells by applying a hollow fiber (HF)/organoid culture method. Cylindrical organoids, which had the potential for further spontaneous differentiation, were established inside of hollow fibers. Using this method, we improved the proliferation rate of mouse ES cells to produce an increased HSC population and achieved around a 40-fold higher production volume of HSCs in HF culture than in conventional EB culture. Therefore, the HF/organoid culture method may be a new mass culture method to acquire pluripotent stem cell-derived HSCs.

Patient-specific cardiovascular progenitor cells derived from integration-free induced pluripotent stem cells for vascular tissue regeneration.

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Hu, Jiang; Wang, Yongyu; Jiao, Jiao; Liu, Zhongning; Zhao, Chao; Zhou, Zhou; Zhang, Zhanpeng; Forde, Kaitlynn; Wang, Lunchang; Wang, Jiangang; Baylink, David J; Zhang, Xiao-Bing; Gao, Shaorong; Yang, Bo; Chen, Y Eugene; Ma, Peter X

2015-12-01

Tissue-engineered blood vessels (TEBVs) are promising in regenerating a live vascular replacement. However, the vascular cell source is limited, and it is crucial to develop a scaffold that accommodates new type of vascular progenitor cells and facilitates in vivo lineage specification of the cells into functional vascular smooth muscle cells (VSMCs) to regenerate vascular tissue. In the present study, integration-free human induced pluripotent stem cells (hiPSCs) were established from patient peripheral blood mononuclear cells through episomal vector nucleofection of reprogramming factors. The established hiPSCs were then induced into mesoderm-originated cardiovascular progenitor cells (CVPCs) with a highly efficient directed lineage specification method. The derived CVPCs were demonstrated to be able to differentiate into functional VSMCs. Subcutaneous implantation of CVPCs seeded on macroporous nanofibrous poly(l-lactide) scaffolds led to in vivo VSMC lineage specification and matrix deposition inside the scaffolds. In summary, we established integration-free patient-specific hiPSCs from peripheral blood mononuclear cells, derived CVPCs through directed lineage specification, and developed an advanced scaffold for these progenitor cells to further differentiate in vivo into VSMCs and regenerate vascular tissue in a subcutaneous implantation model. This study has established an efficient patient-specific approach towards in vivo regeneration of vascular tissue. Copyright © 2015 Elsevier Ltd. All rights reserved.

Adult Tissue-Derived Stem Cells and Tolerance Induction in Nonhuman Primates for Vascularized Composite Allograft Transplantation

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2017-10-01

AWARD NUMBER: W81XWH-16-2-0042 TITLE: Adult Tissue-Derived Stem Cells and Tolerance Induction in Nonhuman Primates for Vascularized Composite...2017 2. REPORT TYPE Annual 3. DATES COVERED 30 Sep 2016 - 29 Sep 2017 4. TITLE AND SUBTITLE Adult Tissue-Derived Stem Cells and Tolerance Induction...Distribution Unlimited 13. SUPPLEMENTARY NOTES The utilization of adult derived adipose stem cells administration in composite tissue transplantation

Induced Pluripotent Stem Cell-Derived Endothelial Cells in Insulin Resistance and Metabolic Syndrome.

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Carcamo-Orive, Ivan; Huang, Ngan F; Quertermous, Thomas; Knowles, Joshua W

2017-11-01

Insulin resistance leads to a number of metabolic and cellular abnormalities including endothelial dysfunction that increase the risk of vascular disease. Although it has been particularly challenging to study the genetic determinants that predispose to abnormal function of the endothelium in insulin-resistant states, the possibility of deriving endothelial cells from induced pluripotent stem cells generated from individuals with detailed clinical phenotyping, including accurate measurements of insulin resistance accompanied by multilevel omic data (eg, genetic and genomic characterization), has opened new avenues to study this relationship. Unfortunately, several technical barriers have hampered these efforts. In the present review, we summarize the current status of induced pluripotent stem cell-derived endothelial cells for modeling endothelial dysfunction associated with insulin resistance and discuss the challenges to overcoming these limitations. © 2017 American Heart Association, Inc.

Comprehensive characterization of genomic instability in pluripotent stem cells and their derived neuroprogenitor cell lines

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Nestor Luis Lopez Corrales

2012-12-01

Full Text Available The genomic integrity of two human pluripotent stem cells and their derived neuroprogenitor cell lines was studied, applying a combination of high-resolution genetic methodologies. The usefulness of combining array-comparative genomic hybridization (aCGH and multiplex fluorescence in situ hybridization (M-FISH techniques should be delineated to exclude/detect a maximum of possible genomic structural aberrations. Interestingly, in parts different genomic imbalances at chromosomal and subchromosomal levels were detected in pluripotent stem cells and their derivatives. Some of the copy number variations were inherited from the original cell line, whereas other modifications were presumably acquired during the differentiation and manipulation procedures. These results underline the necessity to study both pluripotent stem cells and their differentiated progeny by as many approaches as possible in order to assess their genomic stability before using them in clinical therapies.

Implanted hair follicle stem cells form Schwann cells that support repair of severed peripheral nerves.

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Amoh, Yasuyuki; Li, Lingna; Campillo, Raul; Kawahara, Katsumasa; Katsuoka, Kensei; Penman, Sheldon; Hoffman, Robert M

2005-12-06

The hair follicle bulge area is an abundant, easily accessible source of actively growing, pluripotent adult stem cells. Nestin, a protein marker for neural stem cells, also is expressed in follicle stem cells and their immediate, differentiated progeny. The fluorescent protein GFP, whose expression is driven by the nestin regulatory element in transgenic mice, served to mark the follicle cell fate. The pluripotent nestin-driven GFP stem cells are positive for the stem cell marker CD34 but negative for keratinocyte marker keratin 15, suggesting their relatively undifferentiated state. These cells can differentiate into neurons, glia, keratinocytes, smooth muscle cells, and melanocytes in vitro. In vivo studies show the nestin-driven GFP hair follicle stem cells can differentiate into blood vessels and neural tissue after transplantation to the subcutis of nude mice. Equivalent hair follicle stem cells derived from transgenic mice with beta-actin-driven GFP implanted into the gap region of a severed sciatic nerve greatly enhance the rate of nerve regeneration and the restoration of nerve function. The follicle cells transdifferentiate largely into Schwann cells, which are known to support neuron regrowth. Function of the rejoined sciatic nerve was measured by contraction of the gastrocnemius muscle upon electrical stimulation. After severing the tibial nerve and subsequent transplantation of hair follicle stem cells, walking print length and intermediate toe spread significantly recovered, indicating that the transplanted mice recovered the ability to walk normally. These results suggest that hair follicle stem cells provide an important, accessible, autologous source of adult stem cells for regenerative medicine.

Cell culture density affects the stemness gene expression of adipose tissue-derived mesenchymal stem cells.

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Kim, Dae Seong; Lee, Myoung Woo; Lee, Tae-Hee; Sung, Ki Woong; Koo, Hong Hoe; Yoo, Keon Hee

2017-03-01

The results of clinical trials using mesenchymal stem cells (MSCs) are controversial due to the heterogeneity of human MSCs and differences in culture conditions. In this regard, it is important to identify gene expression patterns according to culture conditions, and to determine how the cells are expanded and when they should be clinically used. In the current study, stemness gene expression was investigated in adipose tissue-derived MSCs (AT-MSCs) harvested following culture at different densities. AT-MSCs were plated at a density of 200 or 5,000 cells/cm 2 . After 7 days of culture, stemness gene expression was examined by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) analysis. The proliferation rate of AT-MSCs harvested at a low density (~50% confluent) was higher than that of AT-MSCs harvested at a high density (~90% confluent). Although there were differences in the expression levels of stemness gene, such as octamer-binding transcription factor 4, nanog homeobox ( Nanog ), SRY-box 2, Kruppel like factor 4, v-myc avian myelocytomatosis viral oncogene homolog ( c-Myc ), and lin-28 homolog A, in the AT-MSCs obtained from different donors, RT-qPCR analysis demonstrated differential gene expression patterns according to the cell culture density. Expression levels of stemness genes, particularly Nanog and c-Myc , were upregulated in AT-MSCs harvested at a low density (~50% confluent) in comparison to AT-MSCs from the same donor harvested at a high density (~90% confluent). These results imply that culture conditions, such as the cell density at harvesting, modulate the stemness gene expression and proliferation of MSCs.

Pluripotent stem cell-derived neural stem cells: From basic research to applications

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Otsu, Masahiro; Nakayama, Takashi; Inoue, Nobuo

2014-01-01

Basic research on pluripotent stem cells is designed to enhance understanding of embryogenesis, whereas applied research is designed to develop novel therapies and prevent diseases. Attainment of these goals has been enhanced by the establishment of embryonic stem cell lines, the technological development of genomic reprogramming to generate induced-pluripotent stem cells, and improvements in vitro techniques to manipulate stem cells. This review summarizes the techniques required to generate...

Drug-loaded nanoparticles induce gene expression in human pluripotent stem cell derivatives

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Gajbhiye, Virendra; Escalante, Leah; Chen, Guojun; Laperle, Alex; Zheng, Qifeng; Steyer, Benjamin; Gong, Shaoqin; Saha, Krishanu

2013-12-01

Tissue engineering and advanced manufacturing of human stem cells requires a suite of tools to control gene expression spatiotemporally in culture. Inducible gene expression systems offer cell-extrinsic control, typically through addition of small molecules, but small molecule inducers typically contain few functional groups for further chemical modification. Doxycycline (DXC), a potent small molecule inducer of tetracycline (Tet) transgene systems, was conjugated to a hyperbranched dendritic polymer (Boltorn H40) and subsequently reacted with polyethylene glycol (PEG). The resulting PEG-H40-DXC nanoparticle exhibited pH-sensitive drug release behavior and successfully controlled gene expression in stem-cell-derived fibroblasts with a Tet-On system. While free DXC inhibited fibroblast proliferation and matrix metalloproteinase (MMP) activity, PEG-H40-DXC nanoparticles maintained higher fibroblast proliferation levels and MMP activity. The results demonstrate that the PEG-H40-DXC nanoparticle system provides an effective tool to controlling gene expression in human stem cell derivatives.Tissue engineering and advanced manufacturing of human stem cells requires a suite of tools to control gene expression spatiotemporally in culture. Inducible gene expression systems offer cell-extrinsic control, typically through addition of small molecules, but small molecule inducers typically contain few functional groups for further chemical modification. Doxycycline (DXC), a potent small molecule inducer of tetracycline (Tet) transgene systems, was conjugated to a hyperbranched dendritic polymer (Boltorn H40) and subsequently reacted with polyethylene glycol (PEG). The resulting PEG-H40-DXC nanoparticle exhibited pH-sensitive drug release behavior and successfully controlled gene expression in stem-cell-derived fibroblasts with a Tet-On system. While free DXC inhibited fibroblast proliferation and matrix metalloproteinase (MMP) activity, PEG-H40-DXC nanoparticles maintained

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

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Tomar, Geetanjali B.; Srivastava, Rupesh K.; Gupta, Navita; Barhanpurkar, Amruta P.; Pote, Satish T. [National Center for Cell Science, University of Pune Campus, Pune 411 007 (India); Jhaveri, Hiral M. [Department of Periodontics and Oral Implantology, Dr. D.Y. Patil Dental College and Hospital, Pune (India); Mishra, Gyan C. [National Center for Cell Science, University of Pune Campus, Pune 411 007 (India); Wani, Mohan R., E-mail: mohanwani@nccs.res.in [National Center for Cell Science, University of Pune Campus, Pune 411 007 (India)

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.

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

International Nuclear Information System (INIS)

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

2010-01-01

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.

Human induced pluripotent stem cell-derived models to investigate human cytomegalovirus infection in neural cells.

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Leonardo D'Aiuto

Full Text Available Human cytomegalovirus (HCMV infection is one of the leading prenatal causes of congenital mental retardation and deformities world-wide. Access to cultured human neuronal lineages, necessary to understand the species specific pathogenic effects of HCMV, has been limited by difficulties in sustaining primary human neuronal cultures. Human induced pluripotent stem (iPS cells now provide an opportunity for such research. We derived iPS cells from human adult fibroblasts and induced neural lineages to investigate their susceptibility to infection with HCMV strain Ad169. Analysis of iPS cells, iPS-derived neural stem cells (NSCs, neural progenitor cells (NPCs and neurons suggests that (i iPS cells are not permissive to HCMV infection, i.e., they do not permit a full viral replication cycle; (ii Neural stem cells have impaired differentiation when infected by HCMV; (iii NPCs are fully permissive for HCMV infection; altered expression of genes related to neural metabolism or neuronal differentiation is also observed; (iv most iPS-derived neurons are not permissive to HCMV infection; and (v infected neurons have impaired calcium influx in response to glutamate.

Adipose Tissue-Derived Mesenchymal Stem Cells as a New Host Cell in Latent Leishmaniasis

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Allahverdiyev, Adil M.; Bagirova, Melahat; Elcicek, Serhat; Koc, Rabia Cakir; Baydar, Serap Yesilkir; Findikli, Necati; Oztel, Olga N.

2011-01-01

Some protozoan infections such as Toxoplasma, Cryptosporidium, and Plasmodium can be transmitted through stem cell transplantations. To our knowledge, so far, there is no study about transmission of Leishmania parasites in stem cell transplantation and interactions between parasites and stem cells in vitro. Therefore, the aim of this study was to investigate the interaction between different species of Leishmania parasites and adipose tissue-derived mesenchymal stem cells (ADMSCs). ADMSCs have been isolated, cultured, characterized, and infected with different species of Leishmania parasites (L. donovani, L. major, L. tropica, and L. infantum). Infectivity was examined by Giemsa staining, microculture, and polymerase chain reaction methods. As a result, infectivity of ADMSCs by Leishmania parasites has been determined for the first time in this study. According to our findings, it is very important that donors are screened for Leishmania parasites before stem cell transplantations in regions where leishmaniasis is endemic. PMID:21896818

Generation of glucose-responsive, insulin-producing cells from human umbilical cord blood-derived mesenchymal stem cells.

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Prabakar, Kamalaveni R; Domínguez-Bendala, Juan; Molano, R Damaris; Pileggi, Antonello; Villate, Susana; Ricordi, Camillo; Inverardi, Luca

2012-01-01

We sought to assess the potential of human cord blood-derived mesenchymal stem cells (CB-MSCs) to derive insulin-producing, glucose-responsive cells. We show here that differentiation protocols based on stepwise culture conditions initially described for human embryonic stem cells (hESCs) lead to differentiation of cord blood-derived precursors towards a pancreatic endocrine phenotype, as assessed by marker expression and in vitro glucose-regulated insulin secretion. Transplantation of these cells in immune-deficient animals shows human C-peptide production in response to a glucose challenge. These data suggest that human cord blood may be a promising source for regenerative medicine approaches for the treatment of diabetes mellitus.

Production of hemizygous and homozygous embryonic stem cell-derived neural progenitor cells from the transgenic alszheimer göttingen minipis

DEFF Research Database (Denmark)

Hall, Vanessa Jane; Jacobsen, J.; Gunnarsson, A.

2011-01-01

Production of hemizygous and homozygous embryonic stem cell-derived neural progenitor cells from the transgenic alszheimer göttingen minipis......Production of hemizygous and homozygous embryonic stem cell-derived neural progenitor cells from the transgenic alszheimer göttingen minipis...

Functional Studies of Missense TREM2 Mutations in Human Stem Cell-Derived Microglia

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Philip W. Brownjohn

2018-04-01

Full Text Available Summary: The derivation of microglia from human stem cells provides systems for understanding microglial biology and enables functional studies of disease-causing mutations. We describe a robust method for the derivation of human microglia from stem cells, which are phenotypically and functionally comparable with primary microglia. We used stem cell-derived microglia to study the consequences of missense mutations in the microglial-expressed protein triggering receptor expressed on myeloid cells 2 (TREM2, which are causal for frontotemporal dementia-like syndrome and Nasu-Hakola disease. We find that mutant TREM2 accumulates in its immature form, does not undergo typical proteolysis, and is not trafficked to the plasma membrane. However, in the absence of plasma membrane TREM2, microglia differentiate normally, respond to stimulation with lipopolysaccharide, and are phagocytically competent. These data indicate that dementia-associated TREM2 mutations have subtle effects on microglia biology, consistent with the adult onset of disease in individuals with these mutations. : Brownjohn and colleagues report methods to generate microglia from induced pluripotent human stem cells, which they demonstrate are highly similar to cultured primary human microglia. Microglia differentiated from patient-derived stem cells carrying neurological disease-causing mutations in the TREM2 receptor differentiate normally and respond appropriately to pathogenic stimuli, despite the absence of functional TREM2 receptor on the plasma membrane. Keywords: dementia, microglia, TREM2, Nasu-Hakola disease, frontotemporal dementia, iPSC-microglia, neuroinflammation

Comparative study of human-induced pluripotent stem cells derived from bone marrow cells, hair keratinocytes, and skin fibroblasts.

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Streckfuss-Bömeke, Katrin; Wolf, Frieder; Azizian, Azadeh; Stauske, Michael; Tiburcy, Malte; Wagner, Stefan; Hübscher, Daniela; Dressel, Ralf; Chen, Simin; Jende, Jörg; Wulf, Gerald; Lorenz, Verena; Schön, Michael P; Maier, Lars S; Zimmermann, Wolfram H; Hasenfuss, Gerd; Guan, Kaomei

2013-09-01

Induced pluripotent stem cells (iPSCs) provide a unique opportunity for the generation of patient-specific cells for use in disease modelling, drug screening, and regenerative medicine. The aim of this study was to compare human-induced pluripotent stem cells (hiPSCs) derived from different somatic cell sources regarding their generation efficiency and cardiac differentiation potential, and functionalities of cardiomyocytes. We generated hiPSCs from hair keratinocytes, bone marrow mesenchymal stem cells (MSCs), and skin fibroblasts by using two different virus systems. We show that MSCs and fibroblasts are more easily reprogrammed than keratinocytes. This corresponds to higher methylation levels of minimal promoter regions of the OCT4 and NANOG genes in keratinocytes than in MSCs and fibroblasts. The success rate and reprogramming efficiency was significantly higher by using the STEMCCA system than the OSNL system. All analysed hiPSCs are pluripotent and show phenotypical characteristics similar to human embryonic stem cells. We studied the cardiac differentiation efficiency of generated hiPSC lines (n = 24) and found that MSC-derived hiPSCs exhibited a significantly higher efficiency to spontaneously differentiate into beating cardiomyocytes when compared with keratinocyte-, and fibroblast-derived hiPSCs. There was no significant difference in the functionalities of the cardiomyocytes derived from hiPSCs with different origins, showing the presence of pacemaker-, atrial-, ventricular- and Purkinje-like cardiomyocytes, and exhibiting rhythmic Ca2+ transients and Ca2+ sparks in hiPSC-derived cardiomyocytes. Furthermore, spontaneously and synchronously beating and force-developing engineered heart tissues were generated. Human-induced pluripotent stem cells can be reprogrammed from all three somatic cell types, but with different efficiency. All analysed iPSCs can differentiate into cardiomyocytes, and the functionalities of cardiomyocytes derived from different cell

Induction of Skin-Derived Precursor Cells from Human Induced Pluripotent Stem Cells.

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Sugiyama-Nakagiri, Yoriko; Fujimura, Tsutomu; Moriwaki, Shigeru

2016-01-01

The generation of full thickness human skin from dissociated cells is an attractive approach not only for treating skin diseases, but also for treating many systemic disorders. However, it is currently not possible to obtain an unlimited number of skin dermal cells. The goal of this study was to develop a procedure to produce skin dermal stem cells from induced pluripotent stem cells (iPSCs). Skin-derived precursor cells (SKPs) were isolated as adult dermal precursors that could differentiate into both neural and mesodermal progenies and could reconstitute the dermis. Thus, we attempted to generate SKPs from iPSCs that could reconstitute the skin dermis. Human iPSCs were initially cultured with recombinant noggin and SB431542, an inhibitor of activin/nodal and TGFβ signaling, to induce neural crest progenitor cells. Those cells were then treated with SKP medium that included CHIR99021, a WNT signal activator. The induction efficacy from neural crest progenitor cells to SKPs was more than 97%. No other modifiers tested were able to induce those cells. Those human iPSC-derived SKPs (hiPSC-SKPs) showed a similar gene expression signature to SKPs isolated from human skin dermis. Human iPSC-SKPs differentiated into neural and mesodermal progenies, including adipocytes, skeletogenic cell types and Schwann cells. Moreover, they could be induced to follicular type keratinization when co-cultured with human epidermal keratinocytes. We here provide a new efficient protocol to create human skin dermal stem cells from hiPSCs that could contribute to the treatment of various skin disorders.

[Characterization of stem cells derived from the neonatal auditory sensory epithelium].

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Diensthuber, M; Heller, S

2010-11-01

In contrast to regenerating hair cell-bearing organs of nonmammalian vertebrates the adult mammalian organ of Corti appears to have lost its ability to maintain stem cells. The result is a lack of regenerative ability and irreversible hearing loss following auditory hair cell death. Unexpectedly, the neonatal auditory sensory epithelium has recently been shown to harbor cells with stem cell features. The origin of these cells within the cochlea's sensory epithelium is unknown. We applied a modified neurosphere assay to identify stem cells within distinct subregions of the neonatal mouse auditory sensory epithelium. Sphere cells were characterized by multiple markers and morphologic techniques. Our data reveal that both the greater and the lesser epithelial ridge contribute to the sphere-forming stem cell population derived from the auditory sensory epithelium. These self-renewing sphere cells express a variety of markers for neural and otic progenitor cells and mature inner ear cell types. Stem cells can be isolated from specific regions of the auditory sensory epithelium. The distinct features of these cells imply a potential application in the development of a cell replacement therapy to regenerate the damaged sensory epithelium.

Biomechanics of stem cells

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Spector, A. A.; Yuan, D.; Somers, S.; Grayson, W. L.

2018-04-01

Stem cells play a key role in the healthy development and maintenance of organisms. They are also critically important in medical treatments of various diseases. It has been recently demonstrated that the mechanical factors such as forces, adhesion, stiffness, relaxation, etc. have significant effects on stem cell functions. Under physiological conditions, cells (stem cells) in muscles, heart, and blood vessels are under the action of externally applied strains. We consider the stem cell microenvironment and performance associated with their conversion (differentiation) into skeletal muscle cells. Two problems are studied by using mathematical models whose parameters are then optimized by fitting experiments. First, we present our analysis of the process of stem cell differentiation under the application of cyclic unidirectional strain. This process is interpreted as a transition through several (six) stages where each of them is defined in terms of expression of a set of factors typical to skeletal muscle cells. The stem cell evolution toward muscle cells is described by a system of nonlinear ODEs. The parameters of the model are determined by fitting the experimental data on the time course of expression of the factors under consideration. Second, we analyse the mechanical (relaxation) properties of a scaffold that serves as the microenvironment for stem cells differentiation into skeletal muscle cells. This scaffold (surrounded by a liquid solution) is composed of unidirectional fibers with pores between them. The relaxation properties of the scaffold are studied in an experiment where a long cylindrical specimen is loaded by the application of ramp displacement until the strain reaches a prescribed value. The magnitude of the corresponding load is recorded. The specimen is considered as transversely isotropic poroelastic cylinder whose force relaxation is associated with liquid diffusion through the pores. An analytical solution for the total force applied to

Characterization and comparison of osteoblasts derived from mouse embryonic stem cells and induced pluripotent stem cells.

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Ma, Ming-San; Kannan, Vishnu; de Vries, Anneriek E; Czepiel, Marcin; Wesseling, Evelyn M; Balasubramaniyan, Veerakumar; Kuijer, Roel; Vissink, Arjan; Copray, Sjef C V M; Raghoebar, Gerry M

2017-01-01

New developments in stem cell biology offer alternatives for the reconstruction of critical-sized bone defects. One of these developments is the use of induced pluripotent stem (iPS) cells. These stem cells are similar to embryonic stem (ES) cells, but can be generated from adult somatic cells and therefore do not raise ethical concerns. Proper characterization of iPS-derived osteoblasts is important for future development of safe clinical applications of these cells. For this reason, we differentiated mouse ES and iPS cells toward osteoblasts using osteogenic medium and compared their functionality. Immunocytochemical analysis showed significant expression of bone markers (osteocalcin and collagen type I) in osteoblasts differentiated from ES and iPS cells on days 7 and 30. An in vitro mineralization assay confirmed the functionality of osteogenically differentiated ES and iPS cells. Gene expression arrays focusing on osteogenic differentiation were performed in order to compare the gene expression pattern in both differentiated and undifferentiated ES cells and iPS cells. We observed a significant upregulation of osteogenesis-related genes such as Runx2, osteopontin, collagen type I, Tnfsf11, Csf1, and alkaline phosphatase upon osteogenic differentiation of the ES and iPS cells. We further validated the expression of key osteogenic genes Runx2, osteopontin, osteocalcin, collagen type I, and osterix in both differentiated and undifferentiated ES and iPS cells by means of quantified real-time polymerase chain reaction. We conclude that ES and iPS cells are similar in their osteogenic differentiation capacities, as well as in their gene expression patterns.