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Autophagy Induction in the Pig Follicular Stage Ovary during Heat Stress
Heat stress (HS) occurs when heat dissipation mechanisms are overwhelmed by heat accumulation processes. HS in pigs is associated with seasonal infertility, a major source of production losses in the swine industry. We have previously shown that HS interferes with molecular pathways contributing to follicle recruitment and development in pigs. Autophagy is the process by which somatic cells degrade and recycle cellular components and is activated by a variety of stressors. During autophagy the cytoplasm is sequestered into a double-membraned cytosolic vesicle, the autophagosome, which fuses with a lysosome to form an autolysosome enabling degradation by lysosomal hydrolases. The Bcl-2 family member protein, Bcl-xL, regulates autophagy through its interaction with Beclin 1 (BECN1), and BECN1 increases membrane isolation after initial induction. During autophagosome membrane extension microtubule-associated protein 1 light chain 3 beta (LC3B) is cleaved to form LC3B-II, which associates with the autophagosomal membrane. An increase in LC3B-II is considered to directly correlate with the abundance of autophagosomes. The objective of this study was to characterize autophagy-related proteins following cyclical HS experienced during the follicular phase preceding ovulation. To accomplish this, twelve gilts were synchronized using Matrix® administered orally for 14 days and subjected to cyclical HS (n = 6) or thermal neutral (TN; n = 6) conditions for 5 days immediately following Matrix® withdrawal. The ambient temperature in the TN room was 20.3°C ± 0.1 °C, and the TN room humidity was 33 ± 13%. The cyclical HS room temperature was 26-32°C and the HS room humidity was 24 ± 6%. During the maximal HS load for each day, the HS pigs had increased (P = 0.001) average rectal temperatures (39.8°C ± 0.2°) compared to the TN pigs (38.8°C ± 0.2°). Ovarian protein abundance of BECN1, LC3B-II, and Bcl-xL were elevated as a result of HS (P < 0.05). LC3B was localized to the granulosa and theca cells of growing follicles, though HS had no effect on LC3B cell specificity. Bcl-xL was more abundant in the oocytes of growing follicles (P = 0.008) due to HS and was qualitatively increased in primary follicles due to HS. These results suggest that autophagy is induced in the ovary due to HS, though future investigations are required to determine if autophagy is a mechanism which enables the ovary to mitigate the deleterious effects of HS on the follicle and maturing oocyte. This project was supported by the Iowa Pork Producers Association.
Keywords: heat stress, ovary, autophagy