Effects of insulin-like growth factor-1 on the proliferation and apoptosis of stallion testicular cells under normal and heat stress culture conditions

Abstract

This study investigated the effect of heat stress on stallion testicular cells (TCs) and the effect of insulin-like growth factor (IGF)−1 on TC viability, proliferation, and apoptosis, including different stages of germ cells. TCs were divided into control or treatment groups with 0.01, 0.1, 1, 10, and 100 ng/mL of recombinant human IGF-1 (rhIGF-1) for 24 h at 34 °C and 37 °C. The population and viability were measured before and after treatment. The effects of rhIGF-1 on TC viability, proliferation, and apoptosis were determined using RT-qPCR. Proliferating cell nuclear antigen (PCNA) and marker of proliferation Ki-67 (MKI-67) were used as proliferation markers. Myeloid leukemia-1 (MCL-1) was used as an antiapoptotic marker. BCL2 antagonist/killer-1 (BAK-1) was used as a proapoptotic marker. The relative abundance of mRNA transcript of undifferentiated cell transcription factor 1 (UTF-1), protein gene product 9.5 (PGP9.5), and deleted in azoospermia-like (DAZL), was measured for spermatogenesis progression. TCs treated with 1 ng/mL rhIGF-1 at 34 °C exhibited the highest viability. Significant upregulation of the relative abundance of mRNA transcript of PCNA, MKI-67, and MCL-1 was observed in treated TCs compared with untreated TCs; however, BAK-1 was significantly downregulated in treated TCs. Germ cells treated with 1 ng/mL rhIGF-1 exhibited the highest relative abundance of mRNA transcript of UTF-1 and DAZL, whereas TCs exposed to 0.1 ng/mL showed the highest PGP9.5 level. These data confirm that heat stress in stallions decreases TC viability. These findings may help identify a basal IGF-1 level for TC proliferation and apoptosis during heat stress-induced testicular degeneration in stallions.