Effect of differentiation, de novo innervation, and electrical pulse stimulation on mRNA and protein expression of Na+,K+-ATPase, FXYD1, and FXYD5 in cultured human skeletal muscle cells

• DOI: 10.1371/journal.pone.0247377
• Publication Date: 2021-02-26T18:36:55Z
• Authors: Vid Jan; Katarina Miš; Natasa Nikolic; Klemen Dolinar; Metka Petrič; Andraž Bone; G. Hege Thoresen; Arild C. Rustan; Tomaž Marš; Alexander V. Chib...; Sergej Pirkmajer
• Keywords: 0301 basic medicine; 570; Science; Muscle Fibers, Skeletal; 610; Ion Channels; Cell Line; 03 medical and health sciences; Animals; Humans; Muscle, Skeletal; Cells, Cultured; Q; Microfilament Proteins; R; Membrane Proteins; Cell Differentiation; Phosphoproteins; Coculture Techniques; Electric Stimulation; Rats; Gene Expression Regulation; Medicine; Sodium-Potassium-Exchanging ATPase; Research Article; Muscle Contraction

Denervation reduces the abundance of Na+,K+-ATPase (NKA) in skeletal muscle, while reinnervation increases it. Primary human muscle cells, most widely used model to study vitro, are usually cultured as myoblasts or myotubes without neurons and typically do not contract spontaneously, which might affect their ability express regulate NKA. We determined how differentiation, de novo innervation, electrical pulse stimulation expression NKA (α β) subunits regulators FXYD1 (phospholemman) FXYD5 (dysadherin). Differentiation into under low serum conditions increased myogenic markers CD56 (NCAM1), desmin, myosin heavy chains, dihydropyridine receptor subunit α1S, SERCA2 well NKAα2 FXYD1, it decreased mRNA. Myotubes, were innervated by motor co-culture with embryonic rat spinal cord explants, started spontaneously within 7-10 days. A short-term (10-11 days) promoted mRNA myokines, such IL-6, IL-7, IL-8, IL-15, but did NKA, FXYDs, musclin, cathepsin B, meteorin-like protein, SPARC. long-term (21 protein NKAα1, NKAα2, phospho-FXYD1Ser68 attendant changes levels. Suppression neuromuscular transmission α-bungarotoxin tubocurarine for 24 h alter FXYD expression. Electrical (48 h) non-innervated NKAβ2, NKAβ3, FXYD5. In conclusion, concentration promotes expression, innervation is essential upregulation myotubes. Finally, although EPS both stimulate contractions myotubes, they exert distinct effects on FXYDs.