(168) Cytokines and Products of Fructolysis in the Ovine Uterus During the Peri-Implantation Period of Pregnancy

Elizabeth A. Moore¹, Maddison A. Olivarez¹, Gregory A. Johnson², Karl Clark¹, Gunter Wagner¹, and Fuller W. Bazer¹

Departments of Animal Sciences¹ and Veterinary Integrative Biosciences², Texas A&M

University, College Station, TX, USA 77843

Abstract Text:

Lactate, a metabolic product of glycolysis and fructoylsis, is often considered a waste product inspite of its abundance in the uterine lumen during the pre-implantation period of pregnancy. Ovine conceptuses (embryo and extra-embryonic membranes) produce high levels of lactate in an environment similar to that under hypoxic stress. Hypoxia inducible factor 1 (HIF1A), expressed in a hypoxic environment, upregulates expression of genes required for angiogenesis, such as vascular endothelial growth factor (VEGF), to increase blood flow to tissues, genes for cell survival, and genes for metabolism via fructolysis such as ketohexokinase (KHK). This study characterized changes in amounts of glucose, pyruvate and lactate with primary interest in lactate that acts via hydroxycarbolic acid receptor 1 (HCAR1) to induce and/or stabilize expression of HIF1A during the peri-implantation period of gestation. A second aim was to characterize expression of interferon gamma (IFNG) and interleukin 1-beta (IL1B) in uterine flushings that may be required for elongation of conceptuses and resolution of inflammation during the per-implantation period of pregnancy. Ewes were synchronized to estrus by placing a CIDR in the vagina for 12 days and administering 20 mg Lutalyse the day of CIDR removal. Ewes were bred to fertile rams at onset of estrus and 12 and 24 h later and assigned randomly to be euthanized and hysterectomized on either Day 13, 14, 15, 16, or 18 (N=3 ewes/day of gestation). Endometrium, whole uterine sections, and conceptus tissues were fixed in 4% paraformaldehyde and also frozen in OCT gel for histological analyses. Endometrium and conceptus tissues were snap frozen for analyses to determine expression of mRNAs and protein. Uterine flushings were collected and stored at -20°C until analyzed for concentrations of lactate, pyruvate, glucose, IFNG and IL1B. Concentrations of lactate tended to be greater on Day 18 than Day 13 (P=0.056) and were significantly greater that concentrations of pyruvate. Concentrations of pyruvate and glucose were not affected by day of pregnancy (P< 0.05). IFNG was detected in uterine flushings at approximately 0.718 ng/ml versus published values of over 1,000 ng/ml for interferon tau, but there was no effect of day of gestation (P>0.05).  Interleukin 1 beta (IL1B) was also detected in uterine flushings with concentrations of 0.382 ng/ml being lower on Day 13 than Days 14, 15, 16, and 18 of gestation (P< 0.05). High concentrations of lactate in uterine fluid may stabilize or increase expression of HIF1A and downstream genes for angiogenesis to increase blood flow to the uterus and within the conceptus. IL1B may have a role in stimulating elongation of the conceptus, while IFNG may have a role in reducing inflammation in the uterus. This research was supported in part by the Hagler Institute for Advanced Studies at Texas A&M University Fellowship program and Agriculture and Food Research Initiative Competitive Grant no. 2018-67015-28093 from the USDA NIFA.