Isolation and characterization of myogenic precursor cells from human cremaster muscle

Neia Naldaiz-Gastesi,Pedro M Baptista,Adolfo López De Munain,Ander Izeta,María Fernanda Lara,Resi De Luna-Diaz,M Alejandro Fernández,Virginia Pérez-López,Isabel M-ª Aragón,María Goicoechea,Bernardo Herrera-Imbroda,Sandra Fuertes-Alvarez

doi:10.1038/s41598-019-40042-6

Abstract

Human myogenic precursor cells have been isolated and expanded from a number of skeletal muscles, but alternative donor biopsy sites must be sought after in diseases where muscle damage is widespread. Biopsy sites must be relatively accessible, and the biopsied muscle dispensable. Here, we aimed to histologically characterize the cremaster muscle with regard number of satellite cells and regenerative fibres, and to isolate and characterize human cremaster muscle-derived stem/precursor cells in adult male donors with the objective of characterizing this muscle as a novel source of myogenic precursor cells. Cremaster muscle biopsies (or adjacent non-muscle tissue for negative controls; N = 19) were taken from male patients undergoing routine surgery for urogenital pathology. Myosphere cultures were derived and tested for their in vitro and in vivo myogenic differentiation and muscle regeneration capacities. Cremaster-derived myogenic precursor cells were maintained by myosphere culture and efficiently differentiated to myotubes in adhesion culture. Upon transplantation to an immunocompromised mouse model of cardiotoxin-induced acute muscle damage, human cremaster-derived myogenic precursor cells survived to the transplants and contributed to muscle regeneration. These precursors are a good candidate for cell therapy approaches of skeletal muscle. Due to their location and developmental origin, we propose that they might be best suited for regeneration of the rhabdosphincter in patients undergoing stress urinary incontinence after radical prostatectomy.

Highlights

In striated muscle, adult myogenic stem cells are known as satellite cells, due to their superficial position on muscle fibres[1]
Despite the important recent advances in the purification and characterization of human satellite cells, they are still isolated in small numbers out of muscle biopsies of a limited size, and the stem cells present restricted expansion capacities in vitro[22,23]
Human myosphere cultures present at least a CD31−CD34− CD45−CD56−CD117−CD29+CD73+CD90+CD105+ “mesenchymal” stem cell population, which possibly is non-myogenic[40] and could be equivalent to fibro-adipogenic precursor cells (FAPs), and a CD34−CD45−PAX7+CD56+ALDH1+ myogenic precursor cell population that possibly corresponds to satellite cells[41]

Summary

Introduction

Adult myogenic stem cells are known as satellite cells, due to their superficial position on muscle fibres[1]. Despite the important recent advances in the purification and characterization of human satellite cells, they are still isolated in small numbers out of muscle biopsies of a limited size (typically of 50–100 mg; there are between 500–1,000 satellite cells per mm3 20), and the stem cells present restricted expansion capacities in vitro[22,23] For these reasons, myoblasts (a heterogeneous mixture of semi-purified CD56+ cells24,25) have been mostly used in clinical trials to date, since they expand well in vitro[26,27]. We propose human cremaster muscle as a convenient source of muscle-derived stem/precursor cells in male donors This is a striated muscle not inserted through a tendon, and which contains a variable number of smooth muscle fibres[42]. Since alternative donor biopsy sites must be identified in diseases where muscle affection is widespread, we here aimed to histologically characterize the cremaster muscle with regard number of satellite cells and regenerative fibres, and to isolate and characterize human cremaster muscle-derived stem/precursor cells in adult male donors to evaluate this muscle as a novel source of myogenic precursor cells

Objectives

Results

Discussion

Conclusion