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Differentiation-dependent PTPIP51 expression in human skeletal muscle cell culture.

ABSTRACT: Protein tyrosine phosphatase-interacting protein 51 (PTPIP51) expression was analyzed in proliferating and differentiating human myogenic cells cultured in vitro. Satellite cell cultures derived from four different individuals were used in this study. To analyze the expression of PTPIP51, myoblasts were cultured under conditions promoting either proliferation or differentiation. In addition, further differentiation of already-differentiated myobtubes was inhibited by resubmitting the cells to conditions promoting proliferation. PTPIP51 protein and mRNA were investigated in samples taken at defined time intervals by immunostaining, immunoblotting, in situ hybridization, and PCR. Image analyses of fluorescence immunostainings were used to quantify PTPIP51 in cultured myoblasts and myotubes. Myoblasts grown in the presence of epidermal and fibroblast growth factors (EGF and FGF), both promoting proliferation, expressed PTPIP51 on a basic level. Differentiation to multinuclear myotubes displayed a linear increase in PTPIP51 expression. The rise in PTPIP51 protein was paralleled by an augmented expression of muscle-specific proteins, namely, sarcoplasmic reticulum Ca(2+) ATPase and myosin heavy-chain protein, both linked to a progressive state of myotubal differentiation. This differentiation-induced increase in PTPIP51 was partly reversible by resubmission of differentiated myotubes to conditions boosting proliferation. The results clearly point toward a strong association between PTPIP51 expression and differentiation in human muscle cells.

SUBMITTER: Barop J

PROVIDER: S-EPMC2675071 | biostudies-literature | 2009 May

REPOSITORIES: biostudies-literature

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Differentiation-dependent PTPIP51 expression in human skeletal muscle cell culture.

Barop Justus J Sauer Heinrich H Steger Klaus K Wimmer Monika M

The journal of histochemistry and cytochemistry : official journal of the Histochemistry Society 20090105 5

Protein tyrosine phosphatase-interacting protein 51 (PTPIP51) expression was analyzed in proliferating and differentiating human myogenic cells cultured in vitro. Satellite cell cultures derived from four different individuals were used in this study. To analyze the expression of PTPIP51, myoblasts were cultured under conditions promoting either proliferation or differentiation. In addition, further differentiation of already-differentiated myobtubes was inhibited by resubmitting the cells to co ...[more]

PMID: 19124842

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Project description:BackgroundSkeletal muscle in livestock develops into meat, an important source of protein and other nutrients for human consumption. The muscle is largely composed of a fixed number of multinucleated myofibers determined during late gestation and remains constant postnatally. A population of postnatal muscle stem cells, called satellite cells, gives rise to myoblast cells that can fuse with the existing myofibers, thus increasing their size. This requires a delicate balance of transcription and growth factors and specific microRNA (miRNA) expressed by satellite cells and their supporting cells from the muscle stem cell niche. The role of transcription and growth factors in bovine myogenesis is well-characterized; however, very little is known about the miRNA activity during this process. We have hypothesized that the expression of miRNA can vary between primary cultures of skeletal muscle cells isolated from the semitendinosus muscles of different cattle breeds and subjected to myogenic differentiation.ResultsAfter a 6-day myogenic differentiation of cells isolated from the muscles of the examined cattle breeds, we found statistically significant differences in the number of myotubes between Hereford (HER)/Limousine (LIM) beef breeds and the Holstein-Friesian (HF) dairy breed (p ≤ 0.001). The microarray analysis revealed differences in the expression of 23 miRNA among the aforementioned primary cultures. On the basis of a functional analysis, we assigned 9 miRNA as molecules responsible for differentiation progression (miR-1, -128a, -133a, -133b, -139, -206, -222, -486, and -503). The target gene prediction and functional analysis revealed 59 miRNA-related genes belonging to the muscle organ development process.ConclusionThe number of myotubes and the miRNA expression in the primary cultures of skeletal muscle cells derived from the semitendinosus muscles of the HER/LIM beef cattle breeds and the HF dairy breed vary when cells are subjected to myogenic differentiation. The net effect of the identified miRNA and their target gene action should be considered the result of the breed-dependent activity of satellite cells and muscle stem cell niche cells and their mutual interactions, which putatively can be engaged in the formation of a larger number of myotubes in beef cattle-related cells (HER/LIM) during in vitro myogenesis.

Project description:Heterotopic ossification (HO) is defined as the formation of ectopic bone in soft tissue outside the skeletal tissue. HO is thought to result from aberrant differentiation of osteogenic progenitors within skeletal muscle. However, the precise origin of HO is still unclear. Skeletal muscle contains two kinds of progenitor cells, myogenic progenitors and mesenchymal progenitors. Myogenic and mesenchymal progenitors in human skeletal muscle can be identified as CD56(+) and PDGFR?(+) cells, respectively. The purpose of this study was to investigate the osteogenic differentiation potential of human skeletal muscle-derived progenitors. Both CD56(+) cells and PDGFR?(+) cells showed comparable osteogenic differentiation potential in vitro. However, in an in vivo ectopic bone formation model, PDGFR?(+) cells formed bone-like tissue and showed successful engraftment, while CD56(+) cells did not form bone-like tissue and did not adapt to an osteogenic environment. Immunohistological analysis of human HO sample revealed that many PDGFR?(+) cells were localized in proximity to ectopic bone formed in skeletal muscle. MicroRNAs (miRNAs) are known to regulate many biological processes including osteogenic differentiation. We investigated the participation of miRNAs in the osteogenic differentiation of PDGFR?(+) cells by using microarray. We identified miRNAs that had not been known to be involved in osteogenesis but showed dramatic changes during osteogenic differentiation of PDGFR?(+) cells. Upregulation of miR-146b-5p and -424 and downregulation of miR-7 during osteogenic differentiation of PDGFR?(+) cells were confirmed by quantitative real-time RT-PCR. Inhibition of upregulated miRNAs, miR-146b-5p and -424, resulted in the suppression of osteocyte maturation, suggesting that these two miRNAs have the positive role in the osteogenesis of PDGFR?(+) cells. Our results suggest that PDGFR?(+) cells may be the major source of HO and that the newly identified miRNAs may regulate osteogenic differentiation process of PDGFR?(+) cells.

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Differentiation-dependent PTPIP51 expression in human skeletal muscle cell culture.

Publications

Differentiation-dependent PTPIP51 expression in human skeletal muscle cell culture.

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