67 PRODUCTION OF CLONED TRANSGENIC RABBITS FROM MESENCHYMAL STEM CELLS

2010 ◽  
Vol 22 (1) ◽  
pp. 192
Author(s):  
S. Li ◽  
T. Flisikowska ◽  
B. Kessler ◽  
T. Güngör ◽  
R. Kind ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Nuclear Transfer ◽  
Adult Stem Cells ◽  
Fluorescent Protein ◽  
Cell Clone ◽  
Serum Starvation ◽  
Cell Stage ◽  
Differentiation Potential ◽  
Transgenic Rabbits

Mesenchymal stem cells (MSC) are adult stem cells with fibroblast-like morphology, which can be easily isolated from bone marrow and expanded in culture. Mesenchymal stem cells are able to grow from a single cell into a cell clone, which makes them potentially useful for gene targeting. In our recent study we investigated the dynamics of epigenetic reprogramming following nuclear transfer (NT) with MSC and found that these cells can support development of cloned embryos as good as genetically identical fibroblasts (Brero et al. 2009 Cloning Stem Cells 11, 319-329). In the present study we tested whether live cloned rabbits can be produced from MSC. Nuclear donor cells were isolated from a 6-week-old transgenic Ali/Bas rabbit, expanded in culture, and assessed for their differentiation potential. Mesenchymal stem cells were transfected with a green fluorescent protein (GFP) reporter gene construct and stable cell clones were selected (GFP-MSC). The MSC and GFP-MSC were used for NT at passage 3 to 7 after serum starvation for 2 to 4 days. Nuclear transfer was performed essentially as described previously (Yang et al. 2007 Reproduction 133, 219-320). To assess the development to blastocyst, reconstructed embryos were cultured in B2 medium for 5 to 6 days, whereas for in vivo development embryos were cultured only overnight and then transferred into recipients at the 4- to 8-cell stage. In the MSC group, 844 oocytes were used, 793 (94%) of them fused, 698/786 (89%) cleaved, and 48/128 (38%) developed to blastocyst. After transfer of 483 cloned embryos into 13 recipients, 2 from 8 pregnant recipients gave birth to 10 (2.4%) rabbits, from which 2 and 1 survived for more than 7 days and 3 months, respectively. In the GFP-MSC group, 444 oocytes were used, 412 (93%) of them fused, 377/409 (92%) cleaved, and 97/178 (55%) developed to blastocyst. Transfer of 216 cloned embryos into 8 recipients resulted in 4 pregnancies. One recipient gave birth to 6 (3.7%) live and 2 stillborn rabbits, from which 2 and 1 rabbits survived for more than 3 days and 2 weeks, respectively. All cloned rabbits carried a GFP gene, and green fluorescence could be detected in the follicles of the skin under a fluorescence microscope (Zeiss Axiovert200, Carl Zeiss, Germany). Our study demonstrates that live cloned rabbits can be produced from genetically modified MSC, thus paving the way to generate gene targeted animals. This work is supported by Roche Diagnostic GmbH.

scholarly journals Epigenetic Regulation of Mesenchymal Stem Cells: A Focus on Osteogenic and Adipogenic Differentiation

2011 ◽  
Vol 2011 ◽  
pp. 1-18 ◽  
Author(s):  
Chad M. Teven ◽  
Xing Liu ◽  
Ning Hu ◽  
Ni Tang ◽  
Stephanie H. Kim ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Epigenetic Regulation ◽  
Adult Stem Cells ◽  
Es Cells ◽  
Adipogenic Differentiation ◽  
Embryonic Stem ◽  
Cell Types ◽  
Differentiation Potential ◽  
Msc Differentiation

Stem cells are characterized by their capability to self-renew and terminally differentiate into multiple cell types. Somatic or adult stem cells have a finite self-renewal capacity and are lineage-restricted. The use of adult stem cells for therapeutic purposes has been a topic of recent interest given the ethical considerations associated with embryonic stem (ES) cells. Mesenchymal stem cells (MSCs) are adult stem cells that can differentiate into osteogenic, adipogenic, chondrogenic, or myogenic lineages. Owing to their ease of isolation and unique characteristics, MSCs have been widely regarded as potential candidates for tissue engineering and repair. While various signaling molecules important to MSC differentiation have been identified, our complete understanding of this process is lacking. Recent investigations focused on the role of epigenetic regulation in lineage-specific differentiation of MSCs have shown that unique patterns of DNA methylation and histone modifications play an important role in the induction of MSC differentiation toward specific lineages. Nevertheless, MSC epigenetic profiles reflect a more restricted differentiation potential as compared to ES cells. Here we review the effect of epigenetic modifications on MSC multipotency and differentiation, with a focus on osteogenic and adipogenic differentiation. We also highlight clinical applications of MSC epigenetics and nuclear reprogramming.

scholarly journals Sources and Clinical Applications of Mesenchymal Stem Cells: State-of-the-art review

2018 ◽  
Vol 18 (3) ◽  
pp. 264 ◽  
Author(s):  
Roberto Berebichez-Fridman ◽  
Pablo R. Montero-Olvera
Keyword(s):  
Tissue Engineering ◽  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Regenerative Medicine ◽  
Adult Stem Cells ◽  
Current Knowledge ◽  
Clinical Applications ◽  
The Body ◽  
Differentiation Potential ◽  
Advantages And Disadvantages

First discovered by Friedenstein in 1976, mesenchymal stem cells (MSCs) are adult stem cells found throughout the body that share a fixed set of characteristics. Discovered initially in the bone marrow, this cell source is considered the gold standard for clinical research, although various other sources—including adipose tissue, dental pulp, mobilised peripheral blood and birth-derived tissues—have since been identified. Although similar, MSCs derived from different sources possess distinct characteristics, advantages and disadvantages, including their differentiation potential and proliferation capacity, which influence their applicability. Hence, they may be used for specific clinical applications in the fields of regenerative medicine and tissue engineering. This review article summarises current knowledge regarding the various sources, characteristics and therapeutic applications of MSCs.Keywords: Mesenchymal Stem Cells; Adult Stem Cells; Regenerative Medicine; Cell Differentiation; Tissue Engineering.

scholarly journals Evaluation of isolation method in remaining of differentiation potential of perivascular human umbilical cord mesenchymal stem cells toward male germ cell-like

2021 ◽  
Vol 23 (2) ◽  
pp. 81-86
Author(s):  
Ali Shojaeian ◽  
Ameneh Mehri-Ghahfarrokhi ◽  
Shima Rahmati-Dehkordi ◽  
Mehdi Banitalebi-Dehkordi
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Germ Cell ◽  
Umbilical Cord ◽  
Adult Stem Cells ◽  
Male Germ Cell ◽  
Human Umbilical Cord ◽  
Differentiation Potential ◽  
Protein Levels ◽  
Old Mice

Background and aims: Infertility is one of the most common problems among couples. Generation of male germ cells from adult stem cells is a current promising priority of researchers. This study aimed to investigate the potential of human umbilical cord mesenchymal stem cells (hUMSCs) on the expression of male germ cell markers after isolating by this method. Methods: The hUMSCs was incubated with retinoic acid, testosterone, and conditioned medium (prepared from testicular cell cultures of 7-day-old mice) during 3 days. The bands were visualized and densitometry was accomplished using LI-COR Biosciences software. Results: The high expression levels of C-KIT, DAZL, PIWIL2, and DDX4 in mRNA and protein levels were observed in treated hUMSCs. Conclusion: Results of reverse transcription polymerase chain reaction (RT-PCR) and western blotting showed that method of isolation had no adverse effects on differentiation potential of hUMSCs.

scholarly journals Adipose Derived Mesenchymal Stem Cells Efficiently Rescue Carbon Tetrachloride-Induced Acute Liver Failure in Mouse

2014 ◽  
Vol 2014 ◽  
pp. 1-8 ◽  
Author(s):  
Lisi Deng ◽  
Guangze Liu ◽  
Xin Wu ◽  
Yaping Wang ◽  
Minghua Tong ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Carbon Tetrachloride ◽  
Liver Failure ◽  
Acute Liver Failure ◽  
Fluorescent Protein ◽  
Survival Rates ◽  
Control Group ◽  
Differentiation Potential ◽  
Green Fluorescent

Background and Aim.Adipose derived mesenchymal stem cells (ADMSCs) may be an attractive source for acute and chronic liver injury because they are abundant and easy to obtain. We aim to investigate the efficacy of ADMSCs transplantation in the acute liver failure (ALF) caused by carbon tetrachloride (CCl4) in mice.Methods.ADMSCs were isolated from inguinal fat pads of enhanced green fluorescent protein (EGFP) transgenic mice and their surface markers and differentiation potential were analyzed. ALF models were established by infusion of CCl4and divided into two groups: control group; EGFP-ADMSCs transplantation group. The restoration of biological functions of the livers receiving transplantation was assessed via a variety of approaches such as survival rates, live function parameters, histological localization of EGFP-ADMSCs, and Immunofluorescence analysis.Results.ADMSCs were positive for CD105, CD44 but negative for CD45, CD34 and had adipogenic, osteogenic differentiation potential. The survival rate of transplantation group significantly increased compared to PBS group. Furthermore, the transplanted cells were well integrated into injured livers and produced albumin, cytokeratin-18.Conclusion.Direct transplantation of ADMSCs is an effective treatment for ALF. The transplanted ADMSCs exhibit the potential to differentiate into hepatocyte-like cells in the injured livers.

scholarly journals GFP Labeling and Hepatic Differentiation Potential of Human Placenta-Derived Mesenchymal Stem Cells

2015 ◽  
Vol 35 (6) ◽  
pp. 2299-2308 ◽  
Author(s):  
Jiong Yu ◽  
Xiaoru Su ◽  
Chengxing Zhu ◽  
Qiaoling Pan ◽  
Jinfeng Yang ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Human Placenta ◽  
Fluorescent Protein ◽  
Gene Transduction ◽  
Hepatic Differentiation ◽  
Differentiation Potential ◽  
Gfp Gene ◽  
Cell Based Therapy

Background: Stem cell-based therapy in liver diseases has received increasing interest over the past decade, but direct evidence of the homing and implantation of transplanted cells is conflicting. Reliable labeling and tracking techniques are essential but lacking. The purpose of this study was to establish human placenta-derived mesenchymal stem cells (hPMSCs) expressing green fluorescent protein (GFP) and to assay their hepatic functional differentiation in vitro. Methods: The GFP gene was transduced into hPMSCs using a lentivirus to establish GFP+ hPMSCs. GFP+ hPMSCs were analyzed for their phenotypic profile, viability and adipogenic, osteogenic and hepatic differentiation. The derived GFP+ hepatocyte-like cells were evaluated for their metabolic, synthetic and secretory functions, respectively. Results: GFP+ hPMSCs expressed high levels of HLA I, CD13, CD105, CD73, CD90, CD44 and CD29, but were negative for HLA II, CD45, CD31, CD34, CD133, CD271 and CD79. They possessed adipogenic, osteogenic and hepatic differentiation potential. Hepatocyte-like cells derived from GFP+ hPMSCs showed typical hepatic phenotypes. Conclusions: GFP gene transduction has no adverse influences on the cellular or biochemical properties of hPMSCs or markers. GFP gene transduction using lentiviral vectors is a reliable labeling and tracking method. GFP+ hPMSCs can therefore serve as a tool to investigate the mechanisms of MSC-based therapy, including hepatic disease therapy.

scholarly journals Quiescent Human Mesenchymal Stem Cells Are More Resistant to Heat Stress than Cycling Cells

2018 ◽  
Vol 2018 ◽  
pp. 1-15 ◽  
Author(s):  
L. L. Alekseenko ◽  
M. A. Shilina ◽  
O. G. Lyublinskaya ◽  
J. S. Kornienko ◽  
O. V. Anatskaya ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Heat Stress ◽  
Stress Responses ◽  
Adult Stem Cells ◽  
Serum Starvation ◽  
Ros Production ◽  
Proliferating Cells ◽  
Spontaneous Transformation ◽  
Quiescent Cells

Quiescence is the prevailing state of many cell types under homeostatic conditions. Yet, surprisingly, little is known about how quiescent cells respond to environmental challenges. The aim of the present study is to compare stress responses of cycling and quiescent mesenchymal stem cells (MSC). Human endometrial mesenchymal cells (eMSС) were employed as adult stem cells. eMSC quiescence was modeled by serum starvation. Sublethal heat shock (HS) was used as a stress factor. Both quiescent and cycling cells were heated at 45°C for 30 min and then returned to standard culture conditions for their recovery. HS response was monitored by DNA damage response, stress-induced premature senescence (SIPS), cell proliferation activity, and oxidative metabolism. It has been found that quiescent cells repair DNA more rapidly, resume proliferation, and undergo SIPS less than proliferating cells. HS-enforced ROS production in heated cycling cells was accompanied with increased expression of genes regulating redox-active proteins. Quiescent cells exposed to HS did not intensify the ROS production, and genes involved in antioxidant defense were mostly silent. Altogether, the results have shown that quiescent cells are more resistant to heat stress than cycling cells. Next-generation sequencing (NGS) demonstrates that HS-survived cells retain differentiation capacity and do not exhibit signs of spontaneous transformation.

scholarly journals Evidence for Bone Marrow Adult Stem Cell Plasticity: Properties, Molecular Mechanisms, Negative Aspects, and Clinical Applications of Hematopoietic and Mesenchymal Stem Cells Transdifferentiation

2013 ◽  
Vol 2013 ◽  
pp. 1-11 ◽  
Author(s):  
Ivana Catacchio ◽  
Simona Berardi ◽  
Antonia Reale ◽  
Annunziata De Luisi ◽  
Vito Racanelli ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Adult Stem Cells ◽  
Molecular Mechanisms ◽  
Embryonic Stem ◽  
Clinical Importance ◽  
The Body ◽  
Differentiation Potential ◽  
New Findings

In contrast to the pluripotentembryonic stem cells(ESCs) which are able to give rise to all cell types of the body, mammalianadult stem cells(ASCs) appeared to be more limited in their differentiation potential and to be committed to their tissue of origin. Recently, surprising new findings have contradicted central dogmas of commitment of ASCs by showing their plasticity to differentiate across tissue lineage boundaries, irrespective of classical germ layer designations. The present paper supports the plasticity of thebone marrow stem cells(BMSCs), bringing the most striking and the latest evidences of the transdifferentiation properties of thebone marrow hematopoietic and mesenchymal stem cells(BMHSCs, and BMMSCs), the two BM populations of ASCs better characterized. In addition, we report the possible mechanisms that may explain these events, outlining the clinical importance of these phenomena and the relative problems.

Mesenchymal Stem Cells in Veterinary Regenerative Therapy: Basic Physiology to Clinical Applications

2021 ◽  
Vol 16 ◽  
Author(s):  
Vikash Chandra ◽  
Pratheesh Mankuzhy ◽  
Taru Sharma G.
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Adult Stem Cells ◽  
Es Cells ◽  
Embryonic Stem ◽  
The Body ◽  
Ligament Injury ◽  
Repair System ◽  
Regenerative Therapy ◽  
Differentiation Potential

Background: The consistent, self-renewal capability and wide-ranging differentiation potential during specific physiologic conditions mark stem cells as a novel candidate not only for biomedical research and regenerative therapy but also as an alternative source in research related to life sciences. This vital and distinct characteristic of stem cells, enable them to offer unprecedented hope in treating many diseases and disorders, which are otherwise difficult to treat. Several efforts are still being undertaken to enhance the efficiency of MSCs for better therapeutic applications. Objective: In recent past several studies have been conducted regarding isolation of stem cells from diverse sources and are being used clinically in veterinary regenerative therapy. But till date only a few systemic studies are available. This study provides a comprehensive analysis of the findings from basic and applied research conducted in stem cell therapeutics with particular emphasis on animals. Result: On the basis of their sources, stem cells can be classified as adult or embryonic stem (ES) cells. Physiologically, the ES cells have capability to differentiate into all body cells and develop into normal adult organism; whereas, adult stem cells serve as repair system by restoring damaged tissues of the body. The adult stem cells referred as Mesenchymal stem cells (MSCs) can be derived from various adult body organs whereas embryos give rise to embryonic stem cells. MSCs, passes unique property of proliferation, trans-differentiation and secretion of important biomolecules to create microenvironment; which is immunosuppressive and stimulate native MSCs of damaged tissue. MSCs being immunocompromised cells can be used in autologous as well as in allogenic mode. Conclusion: In Veterinary therapeutics, MSCs equipped with engineering and pharmaceutical modifications offer a potentially candidate in the treatment of wound healing, nerve injury, bone/ligament injury etc. and also bear a great hope in improvement of udder health and milk production in animals.

scholarly journals In Vitro Osteogenic Potential of Green Fluorescent Protein Labelled Human Embryonic Stem Cell-Derived Osteoprogenitors

2016 ◽  
Vol 2016 ◽  
pp. 1-9 ◽  
Author(s):  
Intekhab Islam ◽  
Gopu Sriram ◽  
Mingming Li ◽  
Yu Zou ◽  
Lulu Li ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Green Fluorescent Protein ◽  
Bone Regeneration ◽  
Osteogenic Differentiation ◽  
Fluorescent Protein ◽  
Embryonic Stem ◽  
Differentiation Potential ◽  
Green Fluorescent

Cellular therapy using stem cells in bone regeneration has gained increasing interest. Various studies suggest the clinical utility of osteoprogenitors-like mesenchymal stem cells in bone regeneration. However, limited availability of mesenchymal stem cells and conflicting evidence on their therapeutic efficacy limit their clinical application. Human embryonic stem cells (hESCs) are potentially an unlimited source of healthy and functional osteoprogenitors (OPs) that could be utilized for bone regenerative applications. However, limited ability to track hESC-derived progenies in vivo greatly hinders translational studies. Hence, in this study, we aimed to establish hESC-derived OPs (hESC-OPs) expressing green fluorescent protein (GFP) and to investigate their osteogenic differentiation potential in vitro. We fluorescently labelled H9-hESCs using a plasmid vector encoding GFP. The GFP-expressing hESCs were differentiated into hESC-OPs. The hESC-OPsGFP+ stably expressed high levels of GFP, CD73, CD90, and CD105. They possessed osteogenic differentiation potential in vitro as demonstrated by increased expression of COL1A1, RUNX2, OSTERIX, and OPG transcripts and mineralized nodules positive for Alizarin Red and immunocytochemical expression of osteocalcin, alkaline phosphatase, and collagen-I. In conclusion, we have demonstrated that fluorescently labelled hESC-OPs can maintain their GFP expression for the long term and their potential for osteogenic differentiation in vitro. In future, these fluorescently labelled hESC-OPs could be used for noninvasive assessment of bone regeneration, safety, and therapeutic efficacy.

scholarly journals Mesenchymal Stem Cells Isolated from the Human Bone Marrow: Cultivation, Phenotypic Analysis and Changes in Proliferation Kinetics

2006 ◽  
Vol 49 (1) ◽  
pp. 27-33 ◽  
Author(s):  
Tomáš Soukup ◽  
Jaroslav Mokrý ◽  
Jana Karbanová ◽  
Robert Pytlík ◽  
Petr Suchomel ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mesenchymal Stem Cells ◽  
Progenitor Cells ◽  
Adult Stem Cells ◽  
Biological Properties ◽  
Limb Bud ◽  
Phenotypic Analysis ◽  
Differentiation Potential ◽  
Visceral Mesoderm

Mesenchymal Stem Cells (MSCs) are rare elements living in various organs (e.g., bone marrow), able to differentiate into specialized tissues, such as bone, cartilage, tendon, and myocardium. Since the first description of MSCs by Fridenshtein, several investigators have shown that these cells can also differentiate into chondrocytes, adipocytes, and, at least, in rodents into skeletal myoblasts. Later on, more primitive progenitor cells were characterized, able to give rise not only to limb-bud mesoderm, but also to cells of visceral mesoderm. Those cells were named mesodermal progenitor cells (MPCs). The aim of our study was to characterize and compare the biological properties and spontaneous differentiation potential of two different cell types (MSCs and MPCs) isolated from the human vertebral body bone marrow. The results of our experiments proved that the MPCs can be expanded beyond Hayflick’s limit and differed from MSCs in morphology, biological and phenotypic characteristics. Because of their high proliferative and differentiation potential, MPCs can become more attractive source of adult stem cells for therapeutic purposes.

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