Use of platelet lysate from estrus heifer for in vitro embryo production in dairy cows

Enhancing the efficiency of bovine in vitro embryo production (IVEP) requires continuous optimization of culture microenvironments. Fetal bovine serum (FBS) remains the standard biological supplement but introduces issues related to batch variability and compromised embryonic ultrastructure. Consequently, identifying alternative, standardized biological supplements is essential to enhance developmental competence.

This study evaluated the effects of supplementing in vitro culture (IVC) medium with 5% platelet lysate (PL), derived from the platelet-rich plasma (PRP) of a single heifer selected during spontaneous estrus, on bovine blastocyst development and hatching rates. A total of 78 ovaries from 39 high-yielding Holstein cows were opportunistically collected from a local abattoir across 5 independent IVEP sessions. To eliminate donor-specific confounding bias, retrieved cumulus-oocyte complexes (COCs) were completely pooled within each session prior to morphological grading and random distribution into either the PL-supplemented group or the IVC Control group (C-group). Across the 5 replicates, a cumulative total of 632 follicles were aspirated (mean: 126.4 ± 8.14 per session), yielding 550 recovered COCs (mean: 110 ± 6.91 per session) and achieving a mean recovery rate of 86.53 ± 3.56%.

Using the independent session as the experimental unit (n = 5; paired analysis), 5% PL supplementation significantly increased the proportion of hatched blastocysts on Day 8 of culture (40.33 ± 3.54% vs. 16.71 ± 1.27% in the C-group; p = 0.007), indicating accelerated hatching kinetics. The cumulative blastocyst yield on Day 8 (expanded plus hatched blastocysts relative to the allocated COCs) was numerically higher in the PL-group (42.75 ± 4.00% vs. 28.28 ± 1.97%) but only approached statistical significance (p = 0.06). Cleavage and morula formation rates, together with all expanded-blastocyst rates, were comparable between the two groups.

Supplementing bovine embryo culture medium with 5% PL derived from an estrus heifer significantly increases blastocyst hatching on Day 8, with a positive but non-significant trend for the overall blastocyst yield. These findings highlight the efficacy of estrus-specific platelet-derived factors as a viable alternative to standard biological supplements for optimizing IVEP outcomes in cattle.