Effects of temperature on proliferation of myoblasts from donor piglets with different thermoregulatory maturities.

Background: Climate change and the associated risk for the occurrence of extreme temperature events or permanent changes in ambient temperature are important in the husbandry of farm animals. The aim of our study was to investigate the effects of permanent cultivation temperatures below (35 °C) and above (39 °C, 41 °C) the standard cultivation temperature (37 °C) on porcine muscle development. Therefore, we used our porcine primary muscle cell culture derived from satellite cells as an in vitro model. Neonatal piglets have limited thermoregulatory stability, and several days after birth are required to maintain their body temperature. To consider this developmental step, we used myoblasts originating from thermolabile (five days of age) and thermostable piglets (twenty days of age).

Results: The efficiency of myoblast proliferation using real-time monitoring via electrical impedance was comparable at all temperatures with no difference in the cell index, slope or doubling time. Both temperatures of 37 °C and 39 °C led to similar biochemical growth properties and cell viability. Only differences in the mRNA expression of myogenesis-associated genes were found at 39 °C compared to 37 °C with less MYF5, MYOD and MSTN and more MYH3 mRNA. Myoblasts grown at 35 °C are smaller, exhibit higher DNA synthesis and express higher amounts of the satellite cell marker PAX7, muscle growth inhibitor MSTN and metabolic coactivator PPARGC1A. Only permanent cultivation at 41 °C resulted in higher HSP expression at the mRNA and protein levels. Interactions between the temperature and donor age showed that MYOD, MYOG, MYH3 and SMPX mRNAs were temperature-dependently expressed in myoblasts of thermolabile but not thermostable piglets.

Conclusions: We conclude that 37 °C to 39 °C is the best physiological temperature range for adequate porcine myoblast development. Corresponding to the body temperatures of piglets, it is therefore possible to culture primary muscle cells at 39 °C. Only the highest temperature of 41 °C acts as a thermal stressor for myoblasts with increased HSP expression, but it also accelerates myogenic development. Cultivation at 35 °C, however, leads to less differentiated myoblasts with distinct thermogenetic activity. The adaptive behavior of derived primary muscle cells to different cultivation temperatures seems to be determined by the thermoregulatory stability of the donor piglets.