The development of skeletal muscle is a highly regulated, multi-step process in which pluripotent mesodermal cells give rise to myoblasts that subsequently withdraw from the cell cycle and differentiate into myotubes as well as myofibers. The plasticity of myoblasts plays a critical role in maintaining skeletal muscle structure and function by myoblast activation, migration, adhesion, membrane reorganization, nuclear fusion, finally forming myotubes/myofibers. Our studies demonstrate that the local hypoxic microenvironment, a great diversity of regulatory factors such as IL-6 superfamily factors (IL-6, LIF, CNTF) and TGF-beta1 could regulate the myoblast plasticity. The aim of this paper is to review the previous studies focused on the regulation of myoblast plasticity and its mechanism in our laboratory. Knowledge about the microenvironment or factors involved in regulating the myoblast plasticity will help develop the prevention and cure measures of skeletal muscle diseases.
Cell Differentiation ; Cellular Microenvironment ; Humans ; Hypoxia ; Muscle Fibers, Skeletal ; cytology ; Muscle, Skeletal ; cytology ; Myoblasts ; cytology

Skeletal muscle possesses a remarkable ability for its regeneration and injured tissue repair. This ability depends on the activity and contributions of muscle satellite cells. Proliferating satellite cells, termed myogenic precursor cells or myoblasts, are activated and driven out of their quiescent state upon muscle injury. In this summary, we present a review to summarize the molecular regulation in skeletal satellite cells to light on the satellite cells' self-renewal mechanism.
Cell Proliferation ; Humans ; Muscle, Skeletal ; Regeneration ; Satellite Cells, Skeletal Muscle ; cytology ; Soft Tissue Injuries

Duchenne muscular dystrophy (DMD) is a fatal, genetic neuromuscular disorders that manifests as progressive muscle wasting. Although there has been enormous progress in the studies of the molecular mechanism of muscular dystrophy, there is still no cure. Cell-based therapy is a promiseful option. This review will focus on the present status of cell-based therapy. Myoblast transfer therapy is hindered by minimal distribution of cells after injection, immune rejection, and poor cell survival. The drawback of bone marrow-derived stem cell transplantation is the low efficiency of transdifferentiation. Compared with them, the injection of postnatal muscle-derived stem cells (MDSC) results in a superior regeneration of dystrophin-expressing myofibers.
Animals ; Bone Marrow Cells ; cytology ; Humans ; Muscle, Skeletal ; cytology ; Muscular Dystrophy, Duchenne ; therapy ; Myoblasts, Skeletal ; transplantation ; Stem Cell Transplantation ; methods

OBJECTIVETo establish the methods for purification, culture, and identification of adult human skeletal muscle stem cells in vitro and to explore the biological properties of the cells.

METHODSMuscle stem cells were obtained by reformed enzymatic digestion of muscle tissue from the consenting donors and cultured in serum-free medium. The morphology was inspected by an inverted phase contrast microscope. Phenotypic characteristics of the cells and expression of cell-specific markers were determined using reverse transcription-polymerase chain reaction (RT-PCR) and immunocytochemistry. The growth of single cells in suspension culture was observed and recorded continuously. The cells were analyzed for their multi-lineage differentiation potential into osteoblastic, adipocyte, and smooth muscle cell lineages.

RESULTSPrimary cultured human skeletal muscle stem cells proliferated and formed the big spheres when cultured with serum free medium. Immunofluorescence staining displayed Pax7 and ALDH1 positive expression in the cell spheres. Furthermore, Myod and Desmin showed positive expression in the monolayer cells derived from the spheres. The gene expressions of Oct3/4, Nanog, Sox2 and Pax7 in the cells were determined by RT-PCR. The cell clones formed from single cells grew well. In addition, they were capable of spontaneous differentiation into myotubes in differentiation medium and into other mesodermal cell lineages in induction medium.

CONCLUSIONSHuman muscle stem cells with properties of self-renewal capacity and multi-differentiation could be successfully isolated and expanded in vitro.

Cell Culture Techniques ; methods ; Cell Differentiation ; Cell Proliferation ; Cells, Cultured ; Humans ; Muscle, Skeletal ; cytology ; Satellite Cells, Skeletal Muscle ; cytology ; Stem Cells ; cytology

Muscle-derived stem cells (MDSC) are defined as myogenic stem cells endowed with their ability to self-renew and differentiate into multiple cell types of their derivative tissue, and are proved to be over 10 times more efficient in hematopoiesis than hematopoietic stem cells (HSC). Although the mechanism which MDSC differentiate into blood cells is still unclear, MDSC were considered to replace HSC to treat the patients suffering from bone marrow diseases such as aplastic anemia and tumor. MDSC are different from HSC in a variety aspects like biological characteristics, protein expression and cell proliferation. On the other hand, MDSC contain multiple distinct stem cell populations. Among these, there is only a small part with the ability to repopulate hematopoietic cells, and it is still uncertain whether their origin is same as HSC. This review summarizes the difference between MDSC and HSC, the ability of MDSC to repopulate hematopoietic cells, and the prospect of MDSCs' transplantation.
Anemia, Aplastic ; Cell Differentiation ; Cell Proliferation ; Hematopoiesis ; Hematopoietic Stem Cells ; cytology ; Humans ; Muscle, Skeletal ; cytology

OBJECTIVETo reveal the effects of three days' repeated exhausted eccentric exercise on the skeletal muscle apoptosis and proliferation in rats.

METHODSFifty male SD rats aged at 8 week old were randomly divided into control group (C) and training groups (B1, B2, B3, B4) (n = 10), the training groups ran on a treadmill every day till exhausted. After they had been trained repeatedly for three days, their medial head of triceps brachii muscle cell apoptosis was detected in paraffin section by the TUNEL, expression of proliferating cell nuclear antigen (PCNA) protein was examined by immunohistochemistry.

RESULTS(1) The apoptosis appeared sequential change, and it was consistent with the exercise-induced skeletal muscle micro-injury (EIMmI). The apoptosis index in the training group after exercise was much greater than that in the control group (P < 0.05), and it reached the peak at 24 h after exercise, then it reduced at 48 h after exercise. (2) The express of PCNA exhibited a sequential change after exercise, the proliferation index in the training group after exercise was greater than that in the control group (P < 0.05), it increased after exercise immediately, but it reduced at 3 h after exercise, then was reached the peak at 24 h after exercise, the proliferation index was moderately correlated with the apoptosis index (P < 0.05).

CONCLUSION(1) Cell apoptosis can induce the delayed skeletal muscle damage. (2) Apoptosis may be a start factor of skeletal muscle regeneration.

Animals ; Apoptosis ; Cell Proliferation ; Male ; Muscle, Skeletal ; cytology ; Physical Conditioning, Animal ; Rats ; Rats, Sprague-Dawley

OBJECTIVETo study the effect of different access routes on autologous satellite cell implantation to stimulate myocardial regeneration.

METHODSSatellite cells were procured from skeletal muscle (gluteus max) of adult mongrel canine, cultured, proliferated and labeled with 4', 6-diamidino-2-phenylindone (DAPI) in vitro. The cells were autologously implanted into the site of acute myocardial infarction by local injection or perfusion through the ligated distal left anterior descending coronary artery. Specimens were harvested 2, 4 and 8 weeks later for histological study.

RESULTSThe labeling efficiency of satellite cells with DAPI was close to 100%. Fluorescent cells were found at the infarcted zone, papillary muscle and local injection site. Some of these cells had progressively differentiated into striated muscle fibers connected to intercalated discs. The infant cells appeared different from the mature myocardium under an electron microscope. Satellite cells implanted by perfusion through the coronary artery were arranged in order of consistency with host myocardial fibers. The satellite cells, implanted by local injection, were found growing in a disordered way.

CONCLUSIONSatellite cells, implanted by coronary artery perfusion, can progressively differentiate into striated muscle fibers, arranging in order and disseminating over the infarcted zone. This approach seems more favorable for the recovery of myocardial contractile function than that of local injection.

Animals ; Cell Differentiation ; physiology ; Dogs ; Myocardial Infarction ; pathology ; therapy ; Myocardium ; cytology ; Regeneration ; Satellite Cells, Skeletal Muscle ; cytology ; transplantation ; Transplantation, Autologous

OBJECTIVETo study the effects of skeletal muscle satellite cells implanted into infarcted myocardium on the volume of remnant myocytes.

METHODSThirty-six adult mongrel canines were divided randomly into implantation group and control group. In the implantation group, skeletal muscle satellite cells taken from the gluteus maximus muscles of the dogs were cultured, proliferated and labeled with 4',6-diamidino-2-phenylindone (DAPI) in vitro. In both groups, a model of acute myocardial infarction was established in every dog. In the implantation group, each dog was injected with M199 solution containing autologous skeletal muscle satellite cells. The dogs in the control group received M199 solution without skeletal muscle satellite cells. The dogs of both groups were killed 2, 4 and 8 weeks after implantation (six dogs in a separate group each time). Both infarcted myocardium and normal myocytes distal from the infracted regions isolated were observed under optical and fluorescent microscope. Their volumes were determined using a confocal microscopy image analysis system and analyzed using SAS. A P < 0.05 was considered significant.

RESULTSA portion of the implanted cells differentiated into muscle fiber with striations and were connected with intercalated discs. Cross-sectional area and cell volume were increased in normal myocardium. Hypertrophy of remnant myocytes in the infarcted site after skeletal muscle cell implantation was much more evident than in the control group. Cross-sectional area, cell area and cell volume differed significantly from those of the control group (P < 0.05). Hypertrophy of the cells occurred predominantly in terms of width and thickness, whereas cell length remained unchanged.

CONCLUSIONSkeletal muscle satellite cells implanted into infarct myocardium, could induce the hypertrophy of remnant myocyte cells in the infarcted site and could also aid in the recovery of the contractile force of the infarcted myocardium.

Animals ; Cell Size ; Dogs ; Myocardial Infarction ; pathology ; Myocardium ; pathology ; Myocytes, Cardiac ; cytology ; Random Allocation ; Satellite Cells, Skeletal Muscle ; cytology ; physiology

This study sought to find out a good way for the cryopreservation of tendon seeding cells so as to facilitate the preparation of tissue engineering tendons as products. The related questions are how different factors affect cell survival rate at the procedure of preservation and whether cryopreservation affects seeding cells' biological characters as well as collagen secretive function. The results of experiment indicate that DMSO is a more effective cryoprotectant in cryopreservation of tissue engineered tendon seeding cells. Blood serum nourishment is very important in cell culture, preservation and treatment. The same sustenance after cryopreservation increases cell survival rate. In the process of cryopreservation, the concentration of cells is important to cell survival rate; cell survival rate will decrease when it is less than 1.0 x 10(6)/ml. In the process of cryopreservation, the cooling speed is also important to cell survival rate, slow cooling method achieves higher cell survival rate than does the rapid cooling method. Cryopreservation by use of 10%DMSO+15%FCS+75%DMEM does not affect seeding cells' collagen secretive function greatly and does not affect seeding cells' growth curve, cell cycle and chromosome mode obviously. The prescription of 10%DMSO +15%FCS+75%DMEM is suited for the cryopreservation of tendon seeding cells.
Cell Count ; Cell Survival ; Cryopreservation ; methods ; Dimethyl Sulfoxide ; Humans ; Muscle, Skeletal ; cytology ; Tendons ; cytology ; Tissue Engineering ; Tissue Preservation ; methods

To prepare scaffolds for heart valve tissue engineering, porcine heart valves were treated with varied concentrations of trypsin for 32, 56, 80 and 104 h or followed with DNase. And then the structure of acellular valves was observed under light microscope, scanning and transmission electron microscope. Porcine endothelial cells, human endothelial cells, and canine myofibroblasts were reseeded onto the acellularized porcine heart valve scaffolds once a day for 3 days. The valves were analyzed by immunohistochemical staining and electron microscopy. Results show that all endothelial cells and the majority of interstitial cells were removed from the heart valves after digestion with trypsin for 104 h, and the collagen fiber structure remains intact, but the space between collagen fibers increased slightly. Incubation with trypsin for 80 h and then with DNase almost removed all cells, and the collagen fiber structure and the space between the fibers remain intact. After reseeding, human endothelial cells almost fully cover the valve scaffold surface as shown by H-E staining and platelet endothelial cell adhesion molecules (PECAM-1) staining. Xenogeneic porcine endothelial cells also adhered to and grew on the scaffolds. As shown by H-E staining and actin staining, canine myofibroblasts not only adhered to the surface of valve scaffold but also migrated to the inner part of matrix after one week culture. These results suggest that the digestion of porcine heart valves with trypsin combining with DNase is a suitable method to remove cells. The acellular porcine heart valve scaffolds have a quite favorable biocompatibility with human and porcine endothelial cells as well as canine myofibroblasts.
Animals ; Bioprosthesis ; Cells, Cultured ; Endothelium, Vascular ; cytology ; transplantation ; Fibroblasts ; cytology ; Heart Valve Prosthesis ; Heart Valves ; cytology ; Humans ; Muscle Fibers, Skeletal ; cytology ; Swine ; Tissue Engineering