Jan 2021 PLoS One

From Bioblast
Publications in the MiPMap
Jan V, Miő K, Nikolic N, Dolinar K, Petrič M, Bone A, Thoresen GH, Rustan AC, Marő T, Chibalin AV, Pirkmajer S (2021) 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. PLoS One 16:e0247377.

Β» PMID: 33635930 Open Access

Jan Vid, Mis Katarina, Nikolic Natasa, Dolinar Klemen, Petric Metka, Bone Andraz, Thoresen G Hege, Rustan Arild C, Mars Tomaz, Chibalin Alexander V, Pirkmajer Sergej (2021) PLoS One

Abstract: Denervation reduces the abundance of Na+,K+i-ATPase (NKA) in skeletal muscle, while reinnervation increases it. Primary human skeletal muscle cells, the most widely used model to study human skeletal muscle in vitro, are usually cultured as myoblasts or myotubes without neurons and typically do not contract spontaneously, which might affect their ability to express and regulate NKA. We determined how differentiation, de novo innervation, and electrical pulse stimulation affect expression of NKA (Ξ± and Ξ²) subunits and NKA regulators FXYD1 (phospholemman) and FXYD5 (dysadherin). Differentiation of myoblasts into myotubes under low serum conditions increased expression of myogenic markers CD56 (NCAM1), desmin, myosin heavy chains, dihydropyridine receptor subunit Ξ±1S, and SERCA2 as well as NKAΞ±2 and FXYD1, while it decreased expression of FXYD5 mRNA. Myotubes, which were innervated de novo by motor neurons in co-culture with the embryonic rat spinal cord explants, started to contract spontaneously within 7-10 days. A short-term co-culture (10-11 days) promoted mRNA expression of myokines, such as IL-6, IL-7, IL-8, and IL-15, but did not affect mRNA expression of NKA, FXYDs, or myokines, such as musclin, cathepsin B, meteorin-like protein, or SPARC. A long-term co-culture (21 days) increased the protein abundance of NKAΞ±1, NKAΞ±2, FXYD1, and phospho-FXYD1Ser68 without attendant changes in mRNA levels. Suppression of neuromuscular transmission with Ξ±-bungarotoxin or tubocurarine for 24 h did not alter NKA or FXYD mRNA expression. Electrical pulse stimulation (48 h) of non-innervated myotubes promoted mRNA expression of NKAΞ²2, NKAΞ²3, FXYD1, and FXYD5. In conclusion, low serum concentration promotes NKAΞ±2 and FXYD1 expression, while de novo innervation is not essential for upregulation of NKAΞ±2 and FXYD1 mRNA in cultured myotubes. Finally, although innervation and EPS both stimulate contractions of myotubes, they exert distinct effects on the expression of NKA and FXYDs.

β€’ Bioblast editor: Plangger M



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