Cold-inducible RNA-binding proteinan RNA chaperone with potential roles in animal reproductive physiology

Abstract

Vitrification has replaced the slow freezing method for cryopreservation of oocytes in several species. However, the optimal protocol for bovine oocyte cryopreservation remains to be established. Several strategies for improving bovine oocyte vitrification have been developed. In this regard, little attention has been given to sublethal mild hypothermia as an inductor of cryotolerance in bovine oocytes. The cold-inducible RNA-binding protein (CIRBP) belongs to the cold-inducible proteins (CIPs), a group of peptides induced by mild hypothermia and other stressors. CIRBP is a potential candidate to improve bovine oocyte cryotolerance due to its function in cell survival. In addition, CIPs play crucial roles in RNA transcript control, raising questions about the link between these proteins and the physiological changes in the female reproductive tract. Accordingly, the present thesis was conducted with the general objective to study CIRBP as a potential molecule to improve bovine oocyte vitrification and its modulation during physiological changes in the female reproductive tract. We first evaluated the effects of cold temperatures on the maturation of bovine cumulus-oocyte complexes. Additionally, we determined CIRBP protein expression by Western blot. Sublethal mild hypothermia was demonstrated to increase the expression of CIRBP. However, as expected, we reported detrimental effects induced by the sublethal stress. After this first approach, we aimed to find other strategies to increase the expression of CIRBP, avoiding the damaging effects of sub-physiological temperature variations. For this purpose, we tested different concentrations of exogenous CIRBP and a well-defined hypothermia mimetic small molecule (zr17-2), to supplement the in vitro maturation media of bovine cumulus-oocyte complexes. Minor changes in mRNA CIRBP expression and other CIPs were linked to this supplementation. Additionally, our results revealed that mild hypothermia and vitrification increased the mRNA expression of CIRBP and other CIPs. Finally, the female reproductive tract studies revealed that CIRBP is physiologically modulated by the cyclic hormonal changes and by the male-female interaction. Overall, this thesis offers new insight into CIRBP reproductive physiology, increasing the knowledge between CIRBP and the female reproductive tract and its potential use in improving bovine oocyte vitrification. A better understanding of the reproductive complex process may serve as novel clinical biomarkers, potential diagnostic tools for reproductive evaluation, and the improvement of animal production.