Abundant Synthesis of Netrin-1 in Satellite Cell-Derived Myoblasts Isolated from EDL Rather Than Soleus Muscle Regulates Fast-Type Myotube Formation.

Takahiro Suzuki,Yuriko Nishi,Rio Arimatsu,Ken Kobayashi,Ryuichi Tatsumi,Kouga Hisaeda,Aika Mori,Takahiro Maeno,Takanori Nishimura,Koichi Ojima,Emi Ichimura,Yuki Yamaya,Mako Nakamura

doi:10.3390/ijms22094499

Abstract

Resident myogenic stem cells (satellite cells) are attracting attention for their novel roles in myofiber type regulation. In the myogenic differentiation phase, satellite cells from soleus muscle (slow fiber-abundant) synthesize and secrete higher levels of semaphorin 3A (Sema3A, a multifunctional modulator) than those derived from extensor digitorum longus (EDL; fast fiber-abundant), suggesting the role of Sema3A in forming slow-twitch myofibers. However, the regulatory mechanisms underlying fast-twitch myotube commitment remain unclear. Herein, we focused on netrin family members (netrin-1, -3, and -4) that compete with Sema3A in neurogenesis and osteogenesis. We examined whether netrins affect fast-twitch myotube generation by evaluating their expression in primary satellite cell cultures. Initially, netrins are upregulated during myogenic differentiation. Next, we compared the expression levels of netrins and their cell membrane receptors between soleus- and EDL-derived satellite cells; only netrin-1 showed higher expression in EDL-derived satellite cells than in soleus-derived satellite cells. We also performed netrin-1 knockdown experiments and additional experiments with recombinant netrin-1 in differentiated satellite cell-derived myoblasts. Netrin-1 knockdown in myoblasts substantially reduced fast-type myosin heavy chain (MyHC) expression; exogenous netrin-1 upregulated fast-type MyHC in satellite cells. Thus, netrin-1 synthesized in EDL-derived satellite cells may promote myofiber type commitment of fast muscles.

Highlights

We have previously reported that satellite cells synthesize and secrete the multipotent modulator semaphorin 3A (Sema3A), a class 3 vertebrate-secreted semaphorin originally identified as a neural chemorepellent, during early myogenic differentiation in response to in vivo injury, as well as in primary cultures in vitro [11,12,13,14]
Exogenous netrin-1 upregulated the expression levels of fast-twitch myofiber markers, and strongly downregulated the slow-twitch markers. These results suggest that satellite cells isolated from extensor digitorum longus (EDL) muscle autonomously promote fast-twitch myotube formation by overexpressing netrin-1 than those isolated from soleus muscle
MRNA would be upregulated by netrin-1, as predicted from the results of the knockdown experiment above, but no significant differences were observed in each netrin-1 addition culture. These results suggest that netrin-1 synthesized in satellite cells during myogenic differentiation promotes myotube formation

Summary

Introduction

Mammalian skeletal muscle can be classified into different myofiber types based on their contractility (fast- and slow-twitch), metabolism (glycolytic and oxidative), fatigue resistance (low and high), and morphological features (e.g., colors, thickness, etc.). The specific myofiber type and its maintenance system are generally affected by the frequency of motor nerve impulse [1]. Motor neurons are categorized as slow and fast type as well as myofiber type [2]. The intracellular mechanisms underlying the myofiber type regulation has been revealed; a transcriptional circuitry element centered around the nuclear receptor peroxisome proliferator-activated receptor (PPAR)-delta and the co-activator PGC1alpha complex, the signaling of calcineurin-dependent nuclear factor of activated T-cells (NFAT)

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Conclusion