Abstract
Simple SummaryWe compared the thermotolerance of cow and sheep fibroblasts after exposure to acute hyperthermia (45 °C for 4 h). The primary culture, first passage, and cryopreserved cow fibroblasts resisted acute hyperthermia in terms of cell viability, proliferation, and migration to close cell scratch, in addition to increased expression of heat shock protein (HSP70 and HSP90) mRNA transcripts.This study was conducted to compare the effects of acute hyperthermia (45 °C for 4 h) on the viability, proliferation, and migratory activity through wound-healing assays of cow and sheep fibroblasts. The study examined the effects on primary cultures and first passage skin-derived fibroblasts. Relative quantification of HSP70, HSP90, P53, BAX, BCL2, and BECN1 was investigated after normalization to housekeeping genes GAPDH and beta-actin. The results revealed that cultured cow primary fibroblasts exhibited increased viability and reinitiated cell migration to close the cell monolayer scratch earlier than sheep cells. Similar patterns were observed in the first passage fibroblasts, with severe effects on sheep cells. Both cow and sheep cells exhibited decreased cell viability and failed to regain migratory activity after re-exposure of recovered heat-shocked cells. Effects of hyperthermia on sheep cells were potentiated by cell cryopreservation. The qPCR results showed that cow cells significantly increased HSP70 and HSP90 expression, which decreased the elevation of P53, and ameliorated the effects of the increased BAX/BCL2 ratio. The results provide a paradigm to compare thermotolerance among different animal species and revealed that trypsin could be an additional stress, which potentiates the effects of heat shock in in vitro experiments.
Highlights
Heat stress (HS) has major effects on milk production and fertility in cows and sheep [1,2,3,4,5,6].Genetic variations in susceptibility to hyperthermia have been reported among different species [7].Acclimation or adaptation to increased temperatures basically depends on physical mechanisms, such as heat dissipation through conduction, convection, radiation, and evaporation by means of sweatingAnimals 2020, 10, 545; doi:10.3390/ani10040545 www.mdpi.com/journal/animalsAnimals 2020, 10, 545 or panting [8,9,10]
We showed that camel cells better tolerated chronic heat shock (45 ◦ C for 20 h) than porcine cells [16], and the results of this study revealed that cow cells were more tolerant to acute heat shock than sheep cells on the primary and first passage level, as well as for cryopreserved fibroblasts
The BAX/BCL2 ratio was elevated in cow fibroblasts, the P53 transcript expression was significantly elevated in both the primary culture and first passage fibroblasts of sheep cells as compared with that in cow cells
Summary
Heat stress (HS) has major effects on milk production and fertility in cows and sheep [1,2,3,4,5,6].Genetic variations in susceptibility to hyperthermia have been reported among different species [7].Acclimation or adaptation to increased temperatures basically depends on physical mechanisms, such as heat dissipation through conduction, convection, radiation, and evaporation by means of sweatingAnimals 2020, 10, 545; doi:10.3390/ani10040545 www.mdpi.com/journal/animalsAnimals 2020, 10, 545 or panting [8,9,10]. In cattle and sheep cells, thermotolerance is higher in breeds adapted to warm climates than in animals with temperate origins [11,12,13,14] This resistance could be mediated by heat shock protein (HSP)-related mechanisms [15]. HSPs are a family of proteins with diverse molecular weights (100, 90, 70, and 60 kDa) and act to prevent thermal damage to proteins [16,17] This acute response plays a pivotal role in the cellular thermal acclimation mechanism to high temperatures and could be used as a marker for the adaptation capabilities of animals to thermal stress [8,16,18,19]
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