Immunological and Endometrial Changes of Early Pregnancy in Dairy Cattle
Restricted (Penn State Only)
- Author:
- Da Silva, Maria
- Graduate Program:
- Integrative and Biomedical Physiology
- Degree:
- Doctor of Philosophy
- Document Type:
- Dissertation
- Date of Defense:
- August 02, 2023
- Committee Members:
- Joy Pate, Major Field Member
Gary Perdew, Outside Unit & Field Member
Troy Ott, Chair & Dissertation Advisor
Alison Gernand, Major Field Member
Gregory Shearer, Program Head/Chair - Keywords:
- immune cells
endometrium
blood
transcriptome
angiogenesis
AHR
HIF1A
progesterone receptors
pregnancy
cattle - Abstract:
- Research in bovine reproduction is crucial for enhancing fertility, efficiency, and sustainability in dairy production. Despite progress in breeding practices and herd health management, a significant number of bovine embryos do not survive past the first month of pregnancy. Factors like the immune system play a critical role in protecting against pathogens and tolerating the semiallogeneic embryo. Immune cells also mediate endometrial tissue remodeling and homeostasis during successful reproduction. However, little is known about immunological and endometrial changes during early pregnancy. To support pregnancy, the early conceptus secretes crucial proteins, including interferon tau (IFNT) that maintains the corpus luteum. Pregnancy-associated glycoproteins (PAGs) are another group of conceptus molecules abundantly secreted from the time of conceptus attachment until parturition, but their specific function remains unknown. In our first study (Chapter 2), we analyzed the transcriptome of the endometrium and blood leukocytes in cyclic heifers treated in vivo with intrauterine infusions of IFNT and PAG. IFNT activated interferon response, immune activation, inflammation, and cell death pathways in the endometrium, while downregulating genes related to vesicle transport and extracellular matrix (ECM) remodeling. Prolonged IFNT altered genes linked to cell invasion, retinoic acid (RA) signaling, and embryo implantation. In peripheral blood, PAG stimulated genes involved in leukocyte differentiation and TGFB signaling but had limited effects on endometrial genes. These findings shed light on how conceptus secretions impact endometrial and immune function during maternal recognition of pregnancy and embryo attachment/implantation. Shortly after attachment/implantation, the endometrium undergoes vascular remodeling to establish the maternal placenta. Adequate blood flow and vascular development are essential for successful embryo implantation, nutrient exchange, and overall fetal development. In the second study (Chapter 3), expression of HIF1A, AHR, and angiogenic factors were characterized in the cyclic and pregnant endometrium at the time of attachment/implantation. Contrary to our hypothesis, the expression of HIF1A, AHR, and angiogenic factors remains low during the attachment/implantation period. Moreover, our results suggest that IFNT may exert a suppressive effect on the expression of molecules of HIF signaling. Understanding the timing and mechanisms governing blood vessel growth, structure, and function in the placenta is crucial for the prevention of placental disorders that impact embryonic health and survival. Although the uterine environment undergoes major immunological adaptations to accommodate the developing embryo, the peripheral immune system is not isolated from pregnancy changes. Hormonal and metabolic signals associated with pregnancy have systemic effects on the immune response in the blood and extrauterine organs. It is generally accepted that systemic immunity must balance tolerance to the developing embryo while also remaining vigilant in eliminating pathogens and threats to pregnancy health. Understanding this delicate immunological balance can support efforts in the prevention of pregnancy loss and complications. In the third study (Chapter 4), we characterized changes in circulating leukocytes during early pregnancy and investigated peripheral immunity differences between lactating cows and heifers. In the first experiments, heifers and cows were studied separately, revealing both similarities and differences in the phenotype of blood leukocytes in response to early pregnancy. In the final experiment, we simultaneously compared samples from heifers and cows, and the results indicated that cows exhibited greater expression of proinflammatory cytokines and molecules associated with leukocyte migration and phagocytosis compared to heifers. Moreover, animals that failed to become pregnant showed altered expression of anti-inflammatory molecules compared to pregnant and cyclic animals. Overall, these findings support the notion that immune responses to early pregnancy differ in heifers and cows, which may contribute to the reduced fertility observed in lactating cows. During our experiments with PBL, we observed that some heifers experienced luteolysis between days 16 and 17 of the estrous cycle, while cows experienced it after day 18. Progesterone primarily regulates cell function via the nuclear progesterone receptor (PGR) and can also signal through membrane-associated receptors of the PAQR and PGRMC family. In cattle, progesterone secretion remains elevated throughout pregnancy, but there is conflicting evidence regarding the presence of PGR in leukocytes of both cattle and humans. In our last study (Chapter 5), we aimed to determine the expression of progesterone and estradiol receptors in bovine peripheral blood leukocytes (PBL) and if their expression was affected by reproductive status, parity, or the day of pregnancy. We also investigated the correlation between steroid hormone concentrations in plasma and the phenotype of blood leukocytes. The results revealed that bovine leukocytes lack PGR during early and mid-pregnancy but express nuclear ESR and membrane PGR (mPGR). Additionally, there was a positive correlation between plasma estradiol and a negative correlation of plasma progesterone with changes in the proportion of myeloid cells and the expression of molecules involved in leukocyte migration during proestrus. Contrary to some published literature, these findings suggest that bovine PBL do not express PGR, and instead, mPGR of the PAQR and PGRMC family may mediate progesterone effects on leukocyte function. In summary, the findings presented in this dissertation have expanded our understanding in several areas: (1) the effects of IFNT and PAG on gene expression in the endometrium and circulating leukocytes, (2) the expression of angiogenic factors in the uterus, (3) the peripheral immune response to early pregnancy in heifers and lactating cows, and (4) the expression of progesterone receptors in immune cells. Our results provide support for the hypothesis that uterine and peripheral immunity is regulated by conceptus and maternal signals to facilitate the establishment of successful pregnancies. The insights gained from this research can help identify immunological factors associated with embryo loss and offer valuable perspectives on the application of strategies to enhance fertility in dairy cattle, other domestic species, and potentially even in humans.