Fetal Myogenic Cells Research Articles

Research of stimulating efficiency of blood allogeneic plasma fractions on cell proliferation in vitro conditions

We conducted a comparative experiment on the effect of platelet rich plasma and platelet poor plasma of allogeneic blood of rats on the proliferative activity of different rat cell lines in experimental conditions in vitro. Rat cells such as umbilical cord mesenchymal stem cells, dermal fibroblasts, and fetal myogenic cells were used in the experiment. To compare the proliferative activity of cell lines, the density of the cell population - confluent - was estimated as the percentage of filling the bottom area of the culture vessel with attached and spread cells. As a result of the research it was found that plasma enriched with platelets in the form of 5% supplement to the growth medium has a pronounced stimulating effect on the proliferative activity of fibroblasts, but less effective in the case of cell culture of mesenchymal stem cells. Platelet poor plasma has a physiological effect on the cell cultures of mesenchymal stem cells and rat fibroblasts similar to fetal calf serum. Myogenic rat cells have a high proliferative potential and do not require additional stimulation of platelet rich plasma and platelet poor plasma at the concentrations studied by us.

Reduced satellite cell density and myogenesis in Wagyu compared with Angus cattle as a possible explanation of its high marbling

Mechanisms responsible for excellent marbling in Japanese black cattle, Wagyu, remain to be established. Because both muscle cells and intramuscular adipocytes are developed from mesenchymal progenitor cells during early muscle development, we hypothesized that intramuscular progenitor cells in Wagyu cattle have attenuated myogenic capacity in favor of adipogenesis, leading to high marbling but reduced muscle growth. Biceps femoris muscle biopsy samples were obtained from both Angus (n=3) and Wagyu (n=3) cattle at 12 months of age. Compared with Angus, the density of satellite cells was much lower in Wagyu muscle (by 45.8±10%, P<0.05). Consistently, the formation of myotubes from muscle-derived progenitor cells was also lower (by 64.2±12.9%, P<0.05), but adipogenic capacity was greater in Wagyu. The average muscle fiber diameter was larger in Wagyu (by 23.9±6.8%, P=0.089) despite less muscle mass, suggesting less muscle fiber formation in Wagyu compared with Angus cattle. Because satellite cells are derived from fetal myogenic cells, the reduction in satellite cell density together with lower muscle fiber formation suggests that myogenesis was attenuated during early muscle development in Wagyu cattle. Given the shared pool of mesenchymal progenitor cells, the attenuated myogenesis likely shifts progenitor cells to adipogenesis during early development, which may contribute to high intramuscular adipocyte formation in Wagyu cattle.

Embryonic and fetal limb myogenic cells are derived from developmentally distinct progenitors and have different requirements for β-catenin

Vertebrate muscle arises sequentially from embryonic, fetal, and adult myoblasts. Although functionally distinct, it is unclear whether these myoblast classes develop from common or different progenitors. Pax3 and Pax7 are expressed by somitic myogenic progenitors and are critical myogenic determinants. To test the developmental origin of embryonic and fetal myogenic cells in the limb, we genetically labeled and ablated Pax3(+) and Pax7(+) cells. Pax3(+)Pax7(-) cells contribute to muscle and endothelium, establish and are required for embryonic myogenesis, and give rise to Pax7(+) cells. Subsequently, Pax7(+) cells give rise to and are required for fetal myogenesis. Thus, Pax3(+) and Pax7(+) cells contribute differentially to embryonic and fetal limb myogenesis. To investigate whether embryonic and fetal limb myogenic cells have different genetic requirements we conditionally inactivated or activated beta-catenin, an important regulator of myogenesis, in Pax3- or Pax7-derived cells. beta-Catenin is necessary within the somite for dermomyotome and myotome formation and delamination of limb myogenic progenitors. In the limb, beta-catenin is not required for embryonic myoblast specification or myofiber differentiation but is critical for determining fetal progenitor number and myofiber number and type. Together, these studies demonstrate that limb embryonic and fetal myogenic cells develop from distinct, but related progenitors and have different cell-autonomous requirements for beta-catenin.

Human muscular fetal cells: a potential cell source for muscular therapies

Myoblast transfer therapy has been extensively studied for a wide range of clinical applications, such as tissue engineering for muscular loss, cardiac surgery or Duchenne Muscular Dystrophy treatment. However, this approach has been hindered by numerous limitations, including early myoblast death after injection and specific immune response after transplantation with allogenic cells. Different cell sources have been analyzed to overcome some of these limitations. The object of our study was to investigate the growth potential, characterization and integration in vivo of human primary fetal skeletal muscle cells. These data together show the potential for the creation of a cell bank to be used as a cell source for muscle cell therapy and tissue engineering. For this purpose, we developed primary muscular cell cultures from biopsies of human male thigh muscle from a 16-week-old fetus and from donors of 13 and 30 years old. We show that fetal myogenic cells can be successfully isolated and expanded in vitro from human fetal muscle biopsies, and that fetal cells have higher growth capacities when compared to young and adult cells. We confirm lineage specificity by comparing fetal muscle cells to fetal skin and bone cells in vitro by immunohistochemistry with desmin and 5.1 H11 antibodies. For the feasibility of the cell bank, we ensured that fetal muscle cells retained intrinsic characteristics after 5 years cryopreservation. Finally, human fetal muscle cells marked with PKH26 were injected in normal C57BL/6 mice and were found to be present up to 4 days. In conclusion we estimate that a human fetal skeletal muscle cell bank can be created for potential muscle cell therapy and tissue engineering.

Melanoma cell adhesion molecule is a novel marker for human fetal myogenic cells and affects myoblast fusion

Myoblast fusion is a highly regulated process that is important during muscle development and myofiber repair and is also likely to play a key role in the incorporation of donor cells in myofibers for cell-based therapy. Although several proteins involved in muscle cell fusion in Drosophila are known, less information is available on the regulation of this process in vertebrates, including humans. To identify proteins that are regulated during fusion of human myoblasts, microarray studies were performed on samples obtained from human fetal skeletal muscle of seven individuals. Primary muscle cells were isolated, expanded, induced to fuse in vitro, and gene expression comparisons were performed between myoblasts and early or late myotubes. Among the regulated genes, melanoma cell adhesion molecule (M-CAM) was found to be significantly downregulated during human fetal muscle cell fusion. M-CAM expression was confirmed on activated myoblasts, both in vitro and in vivo, and on myoendothelial cells (M-CAM(+) CD31(+)), which were positive for the myogenic markers desmin and MyoD. Lastly, in vitro functional studies using M-CAM RNA knockdown demonstrated that inhibition of M-CAM expression enhances myoblast fusion. These studies identify M-CAM as a novel marker for myogenic progenitors in human fetal muscle and confirm that downregulation of this protein promotes myoblast fusion.

Secretion and differential localization of the proteolytic cleavage products Aβ40 and Aβ42 of the Alzheimer amyloid precursor protein in human fetal myogenic cells

Abeta peptides are major components of the amyloid plaques that characterize Alzheimer's disease. These peptides are proteolytic cleavage products of the amyloid precursor protein (APP) and are generated by beta- and gamma-secretases. Here we show by multiparameter immunofluorescence imaging in muscle cells that localization of the Abeta40 and Abeta42 cleavage products reveals different myocyte types in a three-dimensional culture system. These myocyte types are heterogeneous by selective intracellular concentration of either Abeta40 or Abeta42 in vesicular structures, whilst only the Abeta40 peptide is secreted as indicated by Western blot analysis. This cellular pattern of APP proteolysis and Abeta peptide secretion correlates with lack of L-APP mRNA splice isoforms. Differential secretion and intracellular accumulation of Abeta peptides is characteristic for the early myocyte development and might be related to cell fusion.

Chapter 8 Skeletal Muscle Satellite Cell Cultures

Publisher Summary Skeletal muscle satellite cell culture techniques have provided a means for coupling muscle structural and physiological observations to cellular and molecular explanations. Intrinsic properties of these myogenic cells have been studied in vitro , and it has become apparent that the satellite cell has properties that are distinctly different from those of embryonic or fetal myogenic cells. This chapter describes two types of satellite cell culture procedures—(1) monolayer mass cultures of dissociated satellite cells and (2) single muscle fiber cultures with their associated satellite cells—and discusses their application. In both cases, cells and fibers are isolated directly from experimental subjects. The general cellular processes that have been studied in satellite cell culture systems include activation from the quiescent state, migration, proliferation, and differentiation or return to quiescence. There are advantages and disadvantages to each of the culture systems and one single culture system may not be adequate for addressing all experimental questions.

The distal limb environment regulates MyoD accumulation and muscle differentiation in mouse-chick chimæric limbs

Differentiation of muscle and cartilage within developing vertebrate limbs occurs in a proximodistal progression. To investigate the cues responsible for regulating muscle pattern, mouse myoblasts were implanted into early chick wings prior to endogenous chick muscle differentiation. Fetal myogenic cells originating from transgenic mice carrying a lacZ reporter were readily detected in vivo after implantation and their state of differentiation determined with species-specific antibodies to MyoD and myosin heavy chain. When mouse myogenic cells are implanted at the growing tip of early stage 21 limbs MyoD expression is suppressed and little differentiation of the mouse cells is detected initially. At later stages ectopically implanted mouse cells come to lie within muscle masses, re-express MyoD and differentiate in parallel with differentiating chick myoblasts. However, if mouse cells are implanted either proximally at stage 21 or into the limb tip at stage 24, situations in which mouse cells encounter endogenous differentiating chick myoblasts earlier, MyoD suppression is not detected and a higher proportion of mouse cells differentiate. Mouse cells that remain distal to endogenous differentiating myogenic cells are more likely to remain undifferentiated than myoblasts that lie within differentiated chick muscle. Undifferentiated distal mouse cells are still capable of differentiating if explanted in vitro, suggesting that myoblast differentiation is inhibited in vivo. In vitro, MyoD is suppressed in primary mouse myoblasts by the addition of FGF2 and FGF4 to the culture media. Taken together, our data suggest that the inhibition of myogenic differentiation in the distal limb involves MyoD suppression in myoblasts, possibly through an FGF-like activity.

Differential expression of muscle-specific enolase in embryonic and fetal myogenic cells during mouse development

Abstract Three isoforms of the glycolytic enzyme enolase are present in mammals and birds. During development, a switch from the alpha to the beta form takes place in skeletal muscle. In order to investigate the molecular basis of this developmental transition of enolase isoforms, we extracted total RNA from limbs of mouse embryos of different ages, and from cultures of embryonic and fetal myogenic cells. The beta message was detected in limbs from 16-day-old fetuses by Northern-blot analysis and its level was found to increase in newborn and adult muscle; no significant amount of beta mRNA was present in samples from earlier developmental stages, which did however express high levels of the muscle-specific actin mRNA. Analysis of RNA extracted from embryonic and fetal myoblasts differentiated in culture revealed that the level of beta mRNA is about 9-fold higher in fetal myotubes than in embryonic myo-tubes, although the level of muscle actin is comparable in both types of myotubes. These results were confirmed by S1 nuclease protection experiments. Our data show that the appearence of beta enolase transcripts temporally correlates with the formation of the second generation of muscle fibers and suggest that the developmental transition from alpha to beta enolase is linked to a developmental program which takes place in fetal but not in embryonic muscle.

Differential expression of MSE in embryonic and fetal myogenic cells

Differential expression of MSE in embryonic and fetal myogenic cells