Maintaining bovine satellite cells stemness through p38 pathway

Shijie Ding,Chunbao Li,Mark J Post,Daniël G M Molin,G N M Swennen,Guanghong Zhou,Tobias Messmer,Mick Gagliardi

doi:10.1038/s41598-018-28746-7

Shijie Ding, Chunbao Li + Show 6 more

Open Access

https://doi.org/10.1038/s41598-018-28746-7

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Abstract

Isolating and maintaining the appropriate stem cell for large scale cell culture is essential in tissue engineering or food production. For bovine satellite cells an optimized isolation and purification protocol is lacking and there is also no detailed understanding on the factors that maintain stemness of these cells. Here, we set up a fluorescence-activated cell sorting strategy to enrich bovine satellite cells. We found that p38-MAPK signalling is activated and PAX7 expression is gradually lost during satellite cell proliferation. The p38 inhibitor (SB203580) treatment maintained PAX7 expression but inhibited the fusion of satellite cells in a concentration-dependent way in short-term incubation. The mechanism of p38 inhibition was confirmed by inhibiting canonical p38 signalling, i.e. HSP27. Long-term culture with an appropriate concentration of p38i enhanced the proliferation and PAX7 expression, while the differentiation capacity recovered and was enhanced compared to vehicle control. These studies indicate that bovine satellite cells maintenance depends on cell purity and p38 MAPK signalling. Inhibition of p38 MAPK signaling is a promising strategy to facilitate large scale cell expansion of primary cells for tissue engineering and cultured meat purposes.

Highlights

IntroductionInitially identified by Mauro[1] in 1961, are the bona fide muscle stem cells
Satellite cells, initially identified by Mauro[1] in 1961, are the bona fide muscle stem cells
In a purified bovine satellite cell population, we investigated if bovine satellite cells showed up-regulated p38 MAPK signaling accompanied by a loss of differentiation ability during long-term culturing in vitro and if p38 inhibition can rescue stemness of satellite cells

Summary

Introduction

Initially identified by Mauro[1] in 1961, are the bona fide muscle stem cells. Purified mouse[16], human[15,17] and pig[18] satellite cell populations can be obtained by fluorescence-activated cell sorting (FACS) using CD34, β7-integrin, CD56 or CD2915,16,18,19. Expression of these markers is species specific. We first set out to purify the initial population of bovine satellite cells, based on marker expression. In the other daughter cell p38α/β MAPK signaling is not activated and MyoD is not induced, renewing the quiescent satellite cell to maintain the stem cell pool[23]. Inhibition of p38α/β MAPK signaling and culture on soft hydrogel substrates rejuvenated older satellite cells’ potential for regeneration[24]

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