# Expression of MHC I Isoforms in Bovine Placentomes: Impact of Cloning

## TL;DR

This study shows that cloned cow pregnancies have altered immune-related protein expression in the placenta, which may explain higher rates of pregnancy loss.

## Contribution

The study reveals differences in MHC-Ib protein and gene expression patterns between cloned and non-cloned bovine pregnancies.

## Key findings

- Qa-2 protein expression in cloned pregnancies was intense at both early and late stages, unlike in controls.
- ILA-88 labeling was weak in early cloned pregnancies but increased near term, contrasting with controls.
- BoLA-NC3 gene was significantly upregulated in cloned pregnancies during early and near-term gestation.

## Abstract

The immune system protects the body from infections and distinguishes between self and foreign components. During pregnancy, the mother’s immune system recognizes and tolerates the foreign paternal proteins present in the fetus. When there is a maladjustment of the maternal immune system during pregnancy, it may result in maternal rejection and pregnancy loss. The fetus also helps regulate the maternal immune system by producing placenta-specific proteins called non-classical major histocompatibility complexes (MHC-Ib). This study examined how MHC-Ib proteins differ between non-cloned (control) and cloned cow pregnancies since cloned pregnancies have more complications and losses. Using tissue staining and gene expression analyses, we found that in control pregnancies, early expression of a specific MHC-Ib (Qa-2) was barely present but increased near calving, whereas Qa-2 had an opposite pattern in clones. Another MHC-Ib protein (ILA-88) was strongly present throughout the control pregnancies and weaker in early cloned pregnancies, with staining increasing near calving. Additionally, a gene linked to MHC-Ib (BoLA-NC3) showed an opposite expression pattern in cloned pregnancies compared to controls. These changes in MHC-Ib protein levels and gene activity suggest that clones may struggle with proper immune regulation, contributing to the higher rates of pregnancy loss and complications.

Major histocompatibility complex class I (MHC-I) gene expression in the placenta is modulated to tailor the maternal immune response to fetal antigens during pregnancy. This study evaluated MHC-I expression through immunohistochemistry (IHC) using an anti-mouse preimplantation embryo development (PED) clone Qa-2 and anti-bovine leukocyte antigen I (BoLA) monoclonal antibody clone IL-A88 (n = 23), as well as RT-qPCR (n = 17) for classical and non-classical (BoLA-NC) genes in control and cloned bovine placentomes during early and near-term gestation. Control samples showed minimal Qa-2 protein expression in early gestation, with intense labeling in trophoblasts and the maternal uterine epithelium near term. In contrast, cloned samples exhibited intense Qa-2 labeling in both maternal and trophoblastic epithelia at both stages, while trophoblast giant cells (TGCs), located apposed to the maternal epithelium, showed no labeling. Control samples exhibited intense IL-A88 labeling in the maternal epithelium at both stages. In cloned samples, weak to no labeling was observed in early gestation, with intense labeling in maternal and fetal epithelium near term. RT-qPCR revealed significant upregulation of BoLA-NC3 in early gestation, with sustained elevated expression in cloned samples in the near term. These findings suggest that altered BoLA protein expression and gene regulation in cloned pregnancies may contribute to pregnancy complications and increased losses.