Isolation of a slowly adhering cell fraction containing stem cells from murine skeletal muscle by the preplate technique

2008 ◽  
Vol 3 (9) ◽  
pp. 1501-1509 ◽  
Author(s):  
Burhan Gharaibeh ◽  
Aiping Lu ◽  
Jessica Tebbets ◽  
Bo Zheng ◽  
Joe Feduska ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Stem Cells ◽  
Cell Fraction ◽  
Murine Skeletal Muscle

scholarly journals Dynamic Support Culture of Murine Skeletal Muscle-Derived Stem Cells Improves Their Cardiogenic PotentialIn Vitro

2015 ◽  
Vol 2015 ◽  
pp. 1-12 ◽  
Author(s):  
Klaus Neef ◽  
Philipp Treskes ◽  
Guoxing Xu ◽  
Florian Drey ◽  
Sureshkumar Perumal Srinivasan ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Stem Cells ◽  
Cluster Formation ◽  
Mrna Level ◽  
Fatal Outcome ◽  
Heart Tissue ◽  
Culture Conditions ◽  
Whole Cell Patch Clamp ◽  
Specific Culture ◽  
Murine Skeletal Muscle

Ischemic heart disease is the main cause of death in western countries and its burden is increasing worldwide. It typically involves irreversible degeneration and loss of myocardial tissue leading to poor prognosis and fatal outcome. Autologous cells with the potential to regenerate damaged heart tissue would be an ideal source for cell therapeutic approaches. Here, we compared different methods of conditional culture for increasing the yield and cardiogenic potential of murine skeletal muscle-derived stem cells. A subpopulation of nonadherent cells was isolated from skeletal muscle by preplating and applying cell culture conditions differing in support of cluster formation. In contrast to static culture conditions, dynamic culture with or without previous hanging drop preculture led to significantly increased cluster diameters and the expression of cardiac specific markers on the protein and mRNA level. Whole-cell patch-clamp studies revealed similarities to pacemaker action potentials and responsiveness to cardiac specific pharmacological stimuli. This data indicates that skeletal muscle-derived stem cells are capable of adopting enhanced cardiac muscle cell-like properties by applying specific culture conditions. Choosing this route for the establishment of a sustainable, autologous source of cells for cardiac therapies holds the potential of being clinically more acceptable than transgenic manipulation of cells.

Comparative Single Cell Analysis of Side Population (SP) / CD45+ Cells from Marrow and Skeletal Muscle Reveals Evidence of Genuine Stem Cell Function and Multilineage Differentiation Ability in Muscle-Resident Stem Cells.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 2688-2688
Author(s):  
Celine Haond ◽  
Francoise Farace ◽  
Martine Guillier ◽  
Yann Lecluse ◽  
Ludmilla Mecaj ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Stem Cells ◽  
Stem Cell ◽  
Cell Function ◽  
Side Population ◽  
Single Muscle ◽  
Multilineage Differentiation ◽  
Resident Stem Cells ◽  
Murine Skeletal Muscle

Abstract Murine skeletal muscle harbors hematopoietic stem cells. It has been suggested that these cells of hematopoietic origin have an altered stem cell function possibly due to their inadeqaute environment as compared to marrow-resident stem cells. The comparative quantitative and qualitative analysis of marrow and muscle-resident stem cells at the single cell level has not been performed so far. To this end, we have performed in vitro and in vivo stem cell detection assays using highly purified CD45+ cells, side population (SP) cells and SP/CD45 +cells. Muscle and marrow were found to contain 1–3 % and 0.2– 0.5 % of SP cells, respectively. The frequency of SP/CD45+ phenotype was 0.1–0.4% for the marrow and 0.2–0.5% in the muscle. Hematopoietic clonogenic cell efficiency from total nucleated cells was 1/500 for marrow and 1/10000 for muscle. Clonogenic efficiency of muscle CD45+ cells was about 1/3rd of that of marrow but with preserved erythroid and granulocytic differentiation ability. The use of SP/CD45+ cells from both muscle and marrow allowed an enrichment of clonogenic capacity by 60-fold in marrow and 360-fold in the muscle. In limiting dilution assays performed in MS-5 cells over 5 weeks, LTC-IC frequency was found to be 1/100 for marrow SP/CD45+ cells and 1/550 for muscle SP/CD45+ cells. To determine cloning and differentiation abilities of single SP/CD45+ cells purified from muscle and marrow, we have cultured single FACS-sorted cells in the presence of SCF, l-Flt3, IL-7, IL-11 and / or the OP-9 stroma (which promotes hematopoietic cell differentiation from embryonic stem cells) for 14– 21 days. Single SP/CD45+ cell cloning efficiency was 14% for marrow (109 wells + / 768) and 3% for muscle (42 wells+ / 1248). Despite this difference, single muscle-derived SP/CD45+ cells exhibited very robust proliferative activity, with 8– 13 cell doubling being obtained in 8 days in the presence of either cytokines alone or OP9 cells + cytokines, leading to absolute numbers of up to 60000/well. More importantly, like marrow SP/CD45+ cells, individual muscle-derived cells exhibited multilineage differentiation ability, with evidence of myeloid, B, NK and dendritic cell differentiation at day 14–21. In in vivo reconstitution experiments, the mean % of Ly5.1 chimerism generated after transplantation of of highly purified marrow SP/CD45+ cells ( 30 – 5700 cells/mouse, n=25 mice) and muscle SP/CD45+ cells ( 300–6500 cells / mouse) was 60% and 9 %, respectively (+ 8 months). To determine if this difference could be due to homing characteristics, SP+/CD45+ cells of marrow (300 /mouse) and muscle origin (300–500/mouse) were transplanted in lethally irradiated Ly5.2 mice by intrafemoral injection. In these assays, muscle-derived SP+/CD45+ cells gave rise also to persistent but lower Ly5.1 chimerisms as compared to marrow (+ 3 months). Thus, our results demonstrate that murine skeletal muscle harbors true stem cells with extensive proliferative and multilineage differentiation ability but as compared to marrow this population occurs with lower frequency. This heterogeneity, explaining an apparently reduced stem cell function in vivo, is not due to homing inability. Experiments underway will determine the in vivo potential of single muscle-resident stem cells.

scholarly journals Preferential and Comprehensive Reconstitution of Severely Damaged Sciatic Nerve Using Murine Skeletal Muscle-Derived Multipotent Stem Cells

PLoS ONE ◽  
2014 ◽  
Vol 9 (3) ◽  
pp. e91257 ◽  
Author(s):  
Tetsuro Tamaki ◽  
Maki Hirata ◽  
Shuichi Soeda ◽  
Nobuyuki Nakajima ◽  
Kosuke Saito ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Stem Cells ◽  
Sciatic Nerve ◽  
Multipotent Stem Cells ◽  
Murine Skeletal Muscle
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