Epigenetic insights into fertility: involvement of immune cell methylation in dairy cows reproduction

Infertility and post-partum reproductive diseases present significant clinical and economic challenges, particularly in cattle farming. Successful gestation is largely contingent upon the ability of the maternal immune system to recognize and tolerate the embryo. The role of immune cells in promoting fetal-maternal immune tolerance is critical, underlining their importance in post-partum fertility. In this context, DNA methylation in hematopoietic cells may play a pivotal role in determining the susceptibility of the animal to post-partum fertility problems. Identifying epigenetic changes linked to infertility is therefore crucial for advancing sustainable animal production. Characterizing the methylome of immune cells in relation to fertility will enable early phenotyping, setting the stage for innovative methods to detect potential subfertility in animals. In our study, whole epigenome sequencing and enzymatic methyl-seq (EM-seq) were employed to analyze the DNA methylation patterns in total blood collected from twelve Holstein cows raised on the same farm before any disease appeared. This approach helped to map the epigenetic modification profiles of genes associated with the varying fertility phenotypes observed the following year, revealing significant differences between fertile and subfertile cows. We identified 216,990 differentially methylated cytosines (DMCs), with three genes, Interferon tau-3 (IFNT3), KIAA0825, and RAS-Related Protein 2A (RAP2A), showing high significance. Additionally, 23 key genes affected by DMCs across five regions (transcription start sites-TSS shores, introns, exons, distal intergenic areas, and downstream of genes) were identified. Notably, Interferon tau-3 emerged as especially crucial due to its role in early embryonic development and the establishment of gestation in cattle, with seven DMCs in its TSS shores in subfertile cows. Similarly, the KLRA1 gene (Ly49), analogous to KIRs in humans and primarily expressed on Natural Killer (NK) cells, was highlighted for its potential impact on fertility. Past research underscored that a balance between activating and inhibiting KIR receptors is essential for embryo implantation and proper placental development. Moreover, interleukin genes, such as IL-6, IL15, IL22, IL-36G, which are vital for conception, contained multiple DMCs, reinforcing the hypothesis of the involvement of the immune system in bovine fertility. These findings illustrate the potential control that immune cell epigenetics exert on cattle post-partum fertility. Additionally, this study suggests that the risk of developing subfertility could potentially be estimated with as few as 220 methylation biomarkers, paving the way for enhanced animal health management and improved fertility treatments. Overall design: Blood from cows aged between three and eight years and raised under the same conditions at a Vancouver farm, was collected from November 2020 to January 2021. A year later, we selected six cows that were culled during the year due to complications related to reproduction that were not identified at the time of sampling, thus forming the Subfertile Group. We also selected a second group of six cows that were still in the herd and had no fertility issues. This second group represented the Fertile Group.