• The expression of nicotinic acetylcholine receptor subunits of skeletal muscle during myogenic differentiation of rat adipose-derived mesenchymal stem cells
  • Zeinab Shafiei Seifabadi,1,* Dian Dayer,2 Vahid Bayati,3 Seyed Saeed Azandeh,4 Mohammad Rashno,5
    1. Cellular and molecular research center, Medical Basic Sciences Research Institute, Faculty of Medicine, Ahvaz Jundishapur University of Medical Sciences, Ahvaz, Iran
    2. Cellular and molecular research center, Medical Basic Sciences Research Institute, Faculty of Medicine, Ahvaz Jundishapur University of Medical Sciences, Ahvaz, Iran
    3. Cellular and molecular research center, Medical Basic Sciences Research Institute, Faculty of Medicine, Ahvaz Jundishapur University of Medical Sciences, Ahvaz, Iran
    4. Cellular and molecular research center, Medical Basic Sciences Research Institute, Faculty of Medicine, Ahvaz Jundishapur University of Medical Sciences, Ahvaz, Iran
    5. Cellular and molecular research center, Medical Basic Sciences Research Institute, Faculty of Medicine, Ahvaz Jundishapur University of Medical Sciences, Ahvaz, Iran


  • Introduction: A neuromuscular junction (NMJ) is a chemical synapse between motor neurons and skeletal muscles. One of its proteins is the nicotinic acetylcholine receptor (nAChR) which is located on the membrane of skeletal muscle cells. Nicotinic acetylcholine receptors are cationic pentameric membrane proteins that are sensitive to endogenous acetylcholine (ACh). nAChRs are composed of 17 different subunits α1-α10, β1-β4, γ, δ and ε. Muscle receptors and neuroreceptors are two major groups of nAChRs. The hetero-pentameric muscle nAChRs comprise two α1 plus a β1, δ, and γ (fetal) or ε (adult) subunits. After innervation, the newly synthesized nAChR contains ε subunits instead of γ subunits. Adipose-derived stem cells (ADSCs) have been validated for their low immunogenicity and ability to self-renew, to differentiate into various tissue progenitors. Given this, adipose-derived stem cells (ADSCs) are emerging as a clinically relevant cell source for establishing human NMJs to study synaptic growth and maturation as well as disease modeling and drug discovery. This study aimed to investigate the expression of subunits of the nAChR during the myogenesis of rat ADSCs.
  • Methods: For this purpose, stem cells were extracted from rat fat, and myogenic differentiation was performed using 5-aza (3μg/ml) for 24 hours and then the culture medium contained 5% horse serum. On the 3rd, 7th, 14th, 21st, and 28th days of differentiation, the Expression of subunits of nicotinic acetylcholine receptor was investigated compared to undifferentiated ADSCs (control group) using Real-time PCR and western blot.
  • Results: The results showed that ADSCs differentiated with 5-aza and horse serum could express nicotinic acetylcholine receptor α1, β1, δ, γ, and ε subunits (P<0.001). The expression of the epsilon subunit, which represents the mature subunit, is higher than the gamma subunit, which represents the embryonic subunit. In the comparison of days, the highest protein expression of the gamma subunit showed on the 28th day(P<0.0001), and the highest amount of protein of the epsilon subunit was on the 21st day(P<0.0001). The protein and gene expression of all subunits increased with the increase in the differentiation day.
  • Conclusion: The findings can be used in the study of skeletal muscle evolution. Additionally, data may be employed to develop innovative procedures to treat skeletal muscle disorders.
  • Keywords: nicotinic acetylcholine receptor, nAChRs , neuromuscular junction, NMJ, rat adipose-derived stem cel