Circulating microRNA as candidates for early embryonic viability in cattle.

Blood-borne extracellular vesicles (i.e., exosomes and microvesicles) carrying microRNAs (miRNAs) could make excellent biomarkers of disease and different physiologic states, including pregnancy status. We tested the hypothesis that circulating extracellular vesicle-derived miRNAs might differentiate the pregnancy status of cows that had maintained pregnancy to Day 30 from non-pregnant cows or from those that exhibited embryonic mortality between Days 17 and 30 of gestation. Cows were randomly assigned for artificial insemination with fertile semen (n = 36) or dead semen (n = 8; control group) on Day 0 (day of estrus). Blood was collected from all animals on Day 0 and on Days 17 and 24 after artificial insemination. Cows receiving live sperm were retrospectively classified as pregnant on Day 30 (n = 17) or exhibiting embryonic mortality between Days 17 and 30 (n = 19). Extracellular vesicles from Day 17 and 24 samples were isolated from serum using ultra-centrifugation, and their presence was confirmed by nanoparticle tracking and Western blot analyses (for CD81) prior to RNA extraction. MicroRNA sequencing was performed on pregnant, embryonic-mortality, and control cows (n = 4 per day), for a total of 24 independent reactions. In total, 214 miRNAs were identified in serum, 40 of which were novel. Based on differential abundance parameters, we identified 32 differentially abundant loci, representing 27 differentially abundant mature miRNA. At Days 17 and 24, specific miRNAs (e.g., miR-25, -16b, and -3596) were identified that differentiated the pregnancy status. In summary, we identified several circulating extracellular vesicles derived miRNAs that differ in abundance between embryonic mortality and pregnant cows.

Estrogenicity of captive southern white rhinoceros diets and their association with fertility.

The captive southern white rhinoceros (SWR) population is not currently self-sustaining, primarily due to poor or absent reproduction of captive-born (F1+) females. In this study, we investigate the role of dietary phytoestrogens in this reproductive phenomenon by characterizing activation of SWR estrogen receptors (ESRs) 1 and 2 by diet items from nine North American institutions and comparing female SWR fertility to total diet estrogenicity. Of the diet items tested, alfalfa hay and soy and alfalfa-based commercial pellets were found to be the most potent activators of SWR ESRs. In contrast, most grass hays tested were not estrogenic. The estrogenicity of total diets varied across the institutions surveyed and the degree of diet estrogenicity was positively associated with the percentage of the total diet comprised by pellets. Comparisons of fertility records of the institutions surveyed showed no significant relationship between diet estrogenicity and fertility for female SWR conceived or born in the wild (F0). However, for F1+ females, there was a significant negative relationship between institutional diet estrogenicity and fertility. Taken together, these data suggest that developmental exposure to phytoestrogens may be the cause of poor fertility in captive-born female SWR. Whether the low fertility of the current population of captive-born female SWR is permanent or can be reversed by removing phytoestrogens from the diet remains unclear. However, our findings suggest that in order for the SWR population to become self-sustaining, the development and feeding of low phytoestrogen diets should be strongly considered.