Number Of Myosin Heavy Chain Research Articles

Reduction of senescent fibro-adipogenic progenitors in progeria-aged muscle by senolytics rescues the function of muscle stem cells.

Fibro-adipogenic progenitors (FAPs) in the muscles have been found to interact closely with muscle progenitor/stem cells (MPCs) and facilitate muscle regeneration at normal conditions. However, it is not clear how FAPs may interact with MPCs in aged muscles. Senolytics have been demonstrated to selectively eliminate senescent cells and generate therapeutic benefits on ageing and multiple age-related disease models. By studying the muscles and primary cells of age matched WT mice and Zmpste24-/- (Z24-/- ) mice, an accelerated ageing model for Hutchinson-Gilford progeria syndrome (HGPS), we examined the interaction between FAPs and MPCs in progeria-aged muscle, and the potential effect of senolytic drug fisetin in removing senescent FAPs and improving the function of MPCs. We observed that, compared with muscles of WT mice, muscles of Z24-/- mice contained a significantly increased number of FAPs (2.4-fold; n>=6, P<0.05) and decreased number of MPCs (2.8-fold; n>=6, P<0.05). FAPs isolated from Z24-/- muscle contained about 44% SA-β-gal+ senescent cells, in contrast to about 3.5% senescent cells in FAPs isolated from WT muscle (n>=6, P<0.001). The co-culture of Z24-/- FAPs with WT MPCs resulted in impaired proliferation and myogenesis potential of WT MPCs, with the number of BrdU positive proliferative cells being reduced for 3.3 times (n>=6, P<0.001) and the number of myosin heavy chain (MHC)-positive myotubes being reduced for 4.5 times (n>=6, P<0.001). The treatment of the in vitro co-culture system of Z24-/- FAPs and WT MPCs with the senolytic drug fisetin led to increased apoptosis of Z24-/- FAPs (14.5-fold; n>=6, P<0.001) and rescued the impaired function of MPCs by increasing the number of MHC-positive myotubes for 3.1 times (n>=6, P<0.001). Treatment of Z24-/- mice with fisetin in vivo was effective in reducing the number of senescent FAPs (2.2-fold, n>=6, P<0.05) and restoring the number of muscle stem cells (2.6-fold, n>=6, P<0.05), leading to improved muscle pathology in Z24-/- mice. These results indicate that the application of senolytics in the progeria-aged muscles can be an efficient strategy to remove senescent cells, including senescent FAPs, which results in improved function of muscle progenitor/stem cells. The senescent FAPs can be a potential novel target for therapeutic treatment of progeria ageing related muscle diseases.

Strength Training Effects on Muscular Regeneration after ACL Reconstruction.

Protracted quadriceps muscle atrophy is observed after anterior cruciate ligament reconstruction (ACL-R). The aim of this study was to assess if quadriceps strength training with eccentric overload (CON/ECC) is more efficient to induce muscle regeneration after ACL-R than conventional concentric/eccentric (CON/ECC) strength training. Biopsies from the vastus lateralis muscle were obtained from 37 recreational athletes after 12 wk of regular rehabilitation after ACL-R and again after 12 wk with twice a week of either conventional CON/ECC (n = 16) or CON/ECC (n = 21) one-legged supervised leg-press training. Immunohistochemical analyses were used to determine satellite cell (SC) number (Pax7); activated SC number (Pax7/MyoD); fibers expressing myosin heavy-chain (MHC) I and II, MHC neonatal, and fiber cross-sectional area. Magnetic resonance imaging was performed to measure quadriceps cross-sectional area and isokinetic testing for the measurement of quadriceps strength. CON/ECC induced a significantly (P = 0.002) greater increase in quadriceps cross-sectional area than did CON/ECC. There also was a significant increase in the fiber cross-sectional areas of all fiber types and in quadriceps strength, but without significant difference between training groups. Only CON/ECC training led to a significant (P < 0.05) increase in percent type I fibers. After training, the number of MHC I/MHCneo fibers was significantly (P < 0.05) greater in the CON/ECC than after in the CON/ECC group. The proportion of hybrid fibers tended to decrease in both groups; percent type II fibers, SC number, and activated SC number remained unchanged. CON/ECC leads to significantly greater muscle hypertrophy compared with CON/ECC, but without the hypothesized enhancing effect on SC activation. At the same time, CON/ECC+ induces a less favorable slower muscle phenotype for strong and fast movements.

Diacylglycerol kinase δ controls down-regulation of cyclin D1 for C2C12 myogenic differentiation

Diacylglycerol kinase (DGK) is a lipid-metabolizing enzyme that phosphorylates diacylglycerol (DG) to produce phosphatidic acid (PA). DGKδ is highly expressed in the skeletal muscle, and a decrease in DGKδ expression increases the severity of type 2 diabetes. However, the role of DGKδ in myogenic differentiation is still unknown. The present study demonstrated that DGKδ expression was down-regulated in the early stage of C2C12 myogenic differentiation almost concurrently with a decrease in cyclin D1 expression. The knockdown of DGKδ by DGKδ-specific siRNAs significantly increased the levels of cyclin D1 expression at 48 h after C2C12 myogenic differentiation. In contrast, at the same time, the knockdown of DGKδ decreased the levels of myogenin expression and the number of myosin heavy chain (MHC)-positive cells. These results indicate that DGKδ regulates the early differentiation of C2C12 myoblasts via controlling the down-regulation of cyclin D1 expression. Moreover, the suppression of DGKδ expression increased the phosphorylation levels of conventional and novel protein kinase Cs (cnPKCs). Furthermore, DGKδ suppression increased the levels of cyclin D1 and phospho-cnPKCs even at the first 24 h of myogenic differentiation. These results suggest that DGKδ controls the down-regulation of cyclin D1 expression by attenuating the PKC signaling pathway for C2C12 myogenic differentiation.

Effect of porcine Akirin2 on skeletal myosin heavy chain isoform expression.

Akirin2 plays an important role in skeletal myogenesis. In this study, we found that porcine Akirin2 (pAkirin2) mRNA level was significantly higher in fast extensor digitorum longus (EDL) and longissimus lumborum (LL) muscles than in slow soleus (SOL) muscle of pigs. Overexpression of pAkirin2 increased the number of myosin heavy chain (MHC)-positive cells, indicating that pAkirin2 promoted myoblast differentiation. We also found that overexpression of pAkirin2 increased the mRNA expressions of MHCI and MHCIIa and decreased the mRNA expression of MHCIIb. Myocyte enhancer factor 2 (MEF2) and nuclear factor of activated T cells (NFAT) are the major downstream effectors of calcineurin. Here we also observed that the mRNA expressions of MEF2C and NFATc1 were notably elevated by pAkirin2 overexpression. Together, our data indicate that the role of pAkirin2 in modulating MHCI and MHCIIa expressions may be achieved through calcineurin/NFATc1 signaling pathway.

Role of microRNA‐27a in myoblast differentiation

MicroRNAs (miRNAs) are a class of endogenous non-coding RNAs that are critically involved in roles in various aspects of skeletal myogenesis. microRNA miR-27a promotes myoblast proliferation by targeting myostatin, a critical inhibitor of skeletal muscle development, but its mode of action in myoblast differentiation remains unclear. We have found that expression of miR-27a and myostatin were upregulated and downregulated, respectively, during myoblast differentiation. Overexpression of miR-27a increased the number of myosin heavy chain (MHC)-positive cells and upregulated mRNA and protein of MyoD and myogenin. These findings indicate that miR-27a plays a role in enhancing myoblast differentiation.

A Molecular Mechanism for Variations in Muscle Function in Rainbow Trout

Salmonids undergo a developmental transition from parr to smolt that involves a number of physiological and morphological changes. In recent years, my laboratory has studied shifts in red muscle function at this parr-smolt transformation (PST) in rainbow trout, Oncorhynchus mykiss. Parr red muscle has faster contraction kinetics than smolts, including faster rates of activation and relaxation and a faster maximum shortening velocity. At PST, a transition in swimming behavior is also observed, with lower tailbeat frequencies and longer EMG duty cycles in the older smolts. Lastly, there is molecular correlate to changes in kinetics and behavior. During PST, there is a developmental reduction in the number of myosin heavy chain (MHC) isoforms in the red muscle of rainbow trout. Since MHC composition of muscle can determine contractile properties, these molecular results suggest a mechanism for the transition in red muscle kinetics and steady swimming. The red muscle of parr is more likely to contain the fast-twitch or white isoform of MHC, resulting in faster contractile properties of that muscle and higher tailbeat frequencies during steady swimming. Lastly, experimental work supports the conclusion that the shift in kinetics causes the observed shift in swimming behavior.

Radiation retards muscle differentiation but does not affect osteoblastic differentiation induced by bone morphogenetic protein-2 in C2C12 myoblasts

Purpose : Effects of radiation on the differentiation of C2C12 myoblasts were studied using cell cultures in which the cells differentiated into either muscle cells or osteoblasts. Materials and methods : A mouse skeletal myoblast cell line, C2C12, was cultured and irradiated at confluence. Muscle differentiation was estimated by counting the number of myosin heavy chain (MHC)-positive multinucleated myotubes. Osteoblast differentiation was analysed by measuring alkaline phosphatase (ALP) activity and osteocalcin production in C2C12 cells treated with recombinant human bone morphogenetic protein-2 (rhBMP-2). Results : Myotube formation in C2C12 cells was inhibited by 2 and 4 Gy irradiation, but recovered to non-irradiated control levels following further culture. In irradiated cultures, the number of MHC-positive mononucleated cells increased significantly. Irradiation did not, however, alter the induction of osteoblastic phenotypes induced by rhBMP-2 in C2C12. Irradiation also did not affect the expression of MyoD mRNA and myogenin mRNA in myogenic differentiation and induction of osteocalcin mRNA in osteoblastic differentiation. Conclusions : During differentiation of C2C12 cells, myotube formation was delayed after irradiation, while induction of osteoblasts with BMP-2 was unaffected.

Sub-Cellular Distribution of the Cytoplasmic Myosin Heavy Chain mRNA During Myogenesis

In the light of earlier work [1] which demonstrated the presence of a large number of myosin heavy chain (MHC) transcripts in chick myoblasts prior to cell fusion and the burst of MHC synthesis it was of great interest to determine the subcellular localization of the still inactive transcripts. It has been determined in differentiating muscle cells in culture. Two populations of cells were examined -- monucleated myoblasts just prior to cell fusion and myotubes where at least 80% of the cells were fused. Utilizing a myosin complementary DNA (cDNA) probe [2] it is observed that just prior to cell fusion, when the "burst" of myosin synthesis has not yet occurred, the vast majority of cytoplasmic myosin mRNA transcripts are found in a stored messenger RNA protein complex with a minimal amount found in the heavy polysome fraction. In differentiated myotube cultures, when myosin synthesis is progressing at a high rate, the reverse is found, i.e, the amount of stored myosin messenger RNA (mRNA) is minimal while the largest amount of myosin mRNA transcripts are localized in the polysome fraction. The number of total cytoplasmic myosin transcripts is found to decrease after cell fusion at a time when myosin synthesis is maximal suggesting that the efficiency of translation of myosin mRNA increases during terminal differentiation.