Differentiation Of Muscle Cells In Culture Research Articles

SNARK is a Novel Regulator of Muscle Mass and Myocyte Apoptosis

The maintenance of skeletal muscle mass is critical for sustaining health; yet the mechanisms responsible for muscle loss with aging and chronic diseases such as diabetes and obesity are poorly understood. The Snf1-AMPK-related kinase (SNARK) belongs to the family of LKB1 substrates known as AMPK-related kinases. SNARK is expressed in skeletal muscle; however, little is known about SNARK function in this tissue. We found that SNARK expression was increased by muscle cell differentiation in culture. Muscle SNARK expression was also elevated with high-fat feeding in mice and with aging in humans (78±5 vs 22±1 yr), suggesting that SNARK may function in the regulation of muscle mass. In line with this hypothesis, decreasing endogenous SNARK expression (siRNA) in cultured muscle cells exposed to metabolic stress (250 µM palmitate) caused a 50% reduction in cell viability together with increased caspase activation compared to control conditions, demonstrating that SNARK plays a protective, anti-apoptotic role in myocyte survival. Likewise, muscle-specific transgenic animals expressing a SNARK dominant-negative inactive mutant (SDN) had increased expression and activation of catabolic and apoptotic mediators in skeletal muscles. Moreover, SDN had severe, age-accelerated muscle atrophy and increased adiposity, consistent with sarcopenic obesity. These data indicate that SNARK protects against age-related muscle atrophy by inhibiting stress-induced apoptosis. This investigation identifies a novel role for SNARK in myocyte survival and the maintenance of muscle mass with age. Funding: NIH R01AR42238 (LJG); APS postdoctoral fellowship in Physiological Genomics (SJL).

Acetylcholinesterase and molecular interactions at the neuromuscular junction

The efficiency and the tight control of neurotransmission require the accumulation of synaptic proteins in discrete domains. In neuromuscular junctions, the main form of acetylcholinesterase (AChE) is a hetero-oligomer in which the catalytic subunits are associated to a specific collagen, ColQ. This structural protein is responsible for the insertion and the accumulation of AChE in the synaptic basal lamina. We have analyzed the time-course of acetylcholinesterase and acetylcholine receptors (AChR) mRNAs during mouse muscle cell differentiation in culture. In parallel, we have visualized the formation of AChE and AChR aggregates. We show that AChR clusters form first which correlates with high γ-subunit mRNA levels. Then, AChE clusters appear with the onset of contraction and correlate with a dramatic increase in AChE, ColQ1 and ColQ1A mRNA levels in muscle cells. At that stage, AChR γ-subunit levels drop while the expression level of ɛ-subunits increase. AChE aggregates are organized by a ternary complex, which involves direct interactions between ColQ, perlecan and MuSK.

Use of a Novel Collagen Matrix with Oriented Pore Structure for Muscle Cell Differentiation in Culture and in Grafts

Use of a Novel Collagen Matrix with Oriented Pore Structure for Muscle Cell Differentiation in Culture and in Grafts

Changes in expression level of V-1, a novel protein containing ankyrin repeats during muscle cell differentiation in culture

Mn-Cu alloys with a high manganese content are traditional damping alloys. In this study, the effects of cyclic annealing on the spinodal decomposition of the 82.2Mn-15.8Cu-2Al(at%) alloy and the martensitic transformation and reverse martensitic transformation of the manganese-rich phase were investigated using differential scanning calorimetry (DSC) analysis, scanning electron microscopy, energy dispersive X-ray spectroscopy, internal friction analysis, and X-ray diffraction analysis. The apparent activation energy and the logarithm of the pre-exponential factor of the spinodal decomposition of the alloy and reverse martensitic transformation were calculated by the Friedman’s and Ozawa–Flynn–Wall methods. The results showed that the cyclic annealing temperature (Ta) and the cyclic annealing number (C) affect the degree of spinodal decomposition. When Ta and C increased, the heat hysteresis of the alloy became narrow. The DSC results showed that the heat hysteresis was only 0.8 K when Ta =693 K for C = 9.

Down-regulation of vimentin gene expression during myogenesis is controlled by a 5′-flanking sequence

During myogenesis, the intermediate filament proteins vimentin and desmin are differentially expressed. While desmin levels increase dramatically, vimentin mRNA levels decrease substantially. Here, we show that transfected whole- and mini-vimentin-coding genes ( Vim) are expressed in fibroblasts (mouse L cells) and down-regulated during muscle cell differentiation in culture. Functional assays with 5 ' -end Vim ::cat constructs demonstrate that this repression is controlled by a 5′-element (nt −321 to −160). This region is distinct from Vim promoter elements (nt −160 to + 71) which do not contribute to vimentin's down-regulation during myogenesis.

Phospholipid biosynthesis during normal and dystrophic avian muscle cell differentiation in culture

The biosynthesis of phospholipids during differentiation of normal and dystrophic muscle cells was studied using [32Pi] added to primary culture media. Labeled phosphatides were separated by two-dimensional thin-layer chromatography, exposing the plates to HCl vapors between solvents to cleave the alk-1-enyl bonds. Phosphatidalethanolamine labeling did not vary during myotube formation. Although lower labeling of phosphatidalcholine was observed in dystrophic myoblasts at the single cell stage, no differences were observed between normal and dystrophic cultures after cell contact and myotube formation. A sharp decrease in phosphoinositide content was observed as myoblasts first achieved cell contact and continued to decline slowly as myotubes developed. During myogenesis, phosphatidylcholine content gradually increased, from 18.4% of the total labeled phospholipid in dividing myoblasts to 48.6% of the labeled phospholipids in fully differentiated myotubes.

Developmental changes in glycosaminoglycans during skeletal muscle cell differentiation in culture

Tissue specificity and developmental changes in glycosaminoglycans were studied in chick embryo skeletal muscle cells differentiating in vitro. Myogenic cells appear to be able to synthesize three main kinds of glycosaminoglycans, namely hyaluronic acid, heparan sulfate and chondroitin sulfate. The patterns of glycosaminoglycans synthesized by terminally differentiated multinucleated myotubes are clearly distinguishable from those synthesized by duplicating mononucleated myogenic cells and by sparse or confluent fibroblasts. These multinucleated myotubes, in particular, exhibit a specific increase of heparan sulfate and a specific decrease of hyaluronic acid in their cell layer-associated fraction of the total synthesized glycosaminoglycans.