Hepatoma-derived growth factor Protein quantification in uterine fluid, gene expression in endometrial-cell culture and effects on in vitro embryo development, pregnancy and birth
Gómez, Enrique | Carrocera, S. | Martín, David | Sánchez-Calabuig, M. J. | Gutiérrez-Adán, Alfonso | Murillo, Antonio | Muñoz Muñoz, María
Hepatoma-derived growth factor (HDGF) is present in the endometrium of cows and other mammals. Recombinant HDGF (rHDGF) improves bovine blastocyst development in vitro. However, specific culture conditions and essential aspects of HDGF uterine physiology are yet unknown. In this work we quantified total HDGF protein in uterine fluid (UF) by multiple reaction monitoring (MRM), and analyzed effects of rHDGF on specific embryonic stages with Day-6 bovine embryos cultured in vitro with and without BSA, and on pregnancy viability and calf phenotypes after embryo transfer to recipients. In addition, mRNA abundance of HDGF in endometrial cells co-cultured with one male or one female embryo was quantified. In the presence of BSA, rHDGF had no effect on blastocyst development; however, in BSA-free culture rHDGF mainly promoted development of early blastocysts in contrast with morulae. As the presence of HDGF contained in commercial BSA replacements was suspected, western blot confirmed HDGF identification in BSA both with and without fatty acids. Total HDGF quantified by MRM tended to increase in UF without vs. UF with embryos (P = 0.083). Pregnancy and birth rates, birth weight and calf measurements did not differ between embryos cultured with rHDGF and controls without rHDGF. However, HDGF abundance in cultured epithelial, endometrial cells tended to increase (P < 0.08) in culture with one male embryo. rHDGF acts selectively on specific embryonic stages, but care should be taken with specific macromolecular supplements in culture. The endometrial expression of HDGF can be regulated by the embryonic sex. The use of rHDGF is compatible with pregnancy and birth of normal calves.
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