S. C. Scolari

Manipulation of the periovulatory sex steroidal milieu affects endometrial but not luteal gene expression in early diestrus Nelore cows

In beef cattle, the ability to conceive has been associated positively with size of the preovulatory follicle (POF). Proestrus estradiol and subsequent progesterone concentrations can regulate the endometrium to affect receptivity and fertility. The aim of the present study was to verify the effect of the size of the POF on luteal and endometrial gene expression during subsequent early diestrus in beef cattle. Eighty-three multiparous, nonlactating, presynchronized Nelore cows received a progesterone-releasing device and estradiol benzoate on Day-10 (D-10). Animals received cloprostenol (large follicle-large CL group; LF-LCL; N = 42) or not (small follicle-small CL group; SF-SCL; N = 41) on D-10. Progesterone devices were withdrawn and cloprostenol administered 42 to 60 hours (LF-LCL) or 30 to 36 hours (SF-SCL) before GnRH treatment (D0). Tissues were collected at slaughter on D7. The LF-LCL group had larger (P < 0.0001) POF (13.24 ± 0.33 mm vs. 10.76 ± 0.29 mm), greater (P < 0.0007) estradiol concentrations on D0 (2.94 ± 0.28 pg/mL vs. 1.27 ± 0.20 pg/mL), and greater (P < 0.01) progesterone concentrations on D7 (3.71 ± 0.25 ng/mL vs. 2.62 ± 0.26 ng/mL) compared with the SF-SCL group. Luteal gene expression of vascular endothelial growth factor A, kinase insert domain receptor, fms-related tyrosine kinase 1, steroidogenic acute regulatory protein, cytochrome P450, family 11, subfamily A, polypeptide 1, and hydroxy-delta-5-steroid dehydrogenase, 3 beta- and steroid delta-isomerase 7 was similar between groups. Endometrial gene expression of oxytocin receptor and peptidase inhibitor 3, skin-derived was reduced, and estrogen receptor alpha 2, aldo-keto reductase family 1, member C4, and lipoprotein lipase expression was increased in LF-LCL versus SF-SCL. Results support the hypothesis that the size of the POF alters the periovulatory endocrine milieu (i.e., proestrus estradiol and diestrus progesterone concentrations) and acts on the uterus to alter endometrial gene expression. It is proposed that the uterine environment and receptivity might also be modulated. Additionally, it is suggested that increased progesterone secretion of cows ovulating larger follicles is likely due to increased CL size rather than increased luteal expression of steroidogenic genes.

The Transcriptome Signature of the Receptive Bovine Uterus Determined at Early Gestation

Pregnancy success is critical to the profitability of cattle operations. However, the molecular events driving the uterine tissue towards embryo receptivity are poorly understood. This study aimed to characterize the uterine transcriptome profiles of pregnant (P) versus non-pregnant (NP) cows during early pregnancy and attempted to define a potential set of marker genes that can be valuable for predicting pregnancy outcome. Therefore, beef cows were synchronized (n=51) and artificially inseminated (n=36) at detected estrus. Six days after AI (D6), jugular blood samples and a biopsy from the uterine horn contralateral to the ovary containing the corpus luteum were collected. Based on pregnancy outcome on D30, samples were retrospectively allocated to the following groups: P (n=6) and NP (n=5). Both groups had similar plasma progesterone concentrations on D6. Uterine biopsies were submitted to RNA-Seq analysis in a Illumina platform. The 272,685,768 million filtered reads were mapped to the Bos Taurus reference genome and 14,654 genes were analyzed for differential expression between groups. Transcriptome data showed that 216 genes are differently expressed when comparing NP versus P uterine tissue (Padj≤0.1). More specifically, 36 genes were up-regulated in P cows and 180 are up-regulated in NP cows. Functional enrichment and pathway analyses revealed enriched expression of genes associated with extracellular matrix remodeling in the NP cows and nucleotide binding, microsome and vesicular fraction in the P cows. From the 40 top-ranked genes, the transcript levels of nine genes were re-evaluated using qRT-PCR. In conclusion, this study characterized a unique set of genes, expressed in the uterus 6 days after insemination, that indicate a receptive state leading to pregnancy success. Furthermore, expression of such genes can be used as potential markers to efficiently predict pregnancy success.

Gene expression profiling by high throughput sequencing to determine signatures for the bovine receptive uterus at early gestation

The uterus plays a central role among the reproductive tissues in the context of early embryo-maternal communication and a successful pregnancy depends on a complex series of endometrial molecular and cellular events. The factors responsible for the initial interaction between maternal and embryonic tissues, leading to the establishment of pregnancy, remain poorly understood. In this context, Illuminas next-generation sequencing technology has been used to discover the uterine transcriptome signature that is favourable for ongoing pregnancy. More specifically, the present report documents on a retrospective in vivo study in which data on pregnancy outcome were linked to uterine gene expression signatures on day 6 (bovine model). Using the RNA-Seq method, 14.654 reference genes were effectively analysed for differential expression between pregnant and non-pregnant uterine tissue. Transcriptome data revealed that 216 genes were differently expressed when comparing uterine tissue from pregnant and non-pregnant cows. All read sequences were deposited in the Sequence Read Archive (SRA) of the NCBI (http://www.ncbi.nlm.nih.gov/sra). An overview of the gene expression data has been deposited in NCBIs Gene Expression Omnibus (GEO) and is accessible through GEO Series accession number GSE65117. This allows the research community to enhance reproducibility and allows for new discoveries by comparing datasets of signatures linked to receptivity and/or pregnancy success. The resulting information can serve as tool to identify valuable and urgently needed biomarkers for scoring maternal receptivity and even for accurate detection of early pregnancy, which is a matter of cross-species interest. Beyond gene expression analysis as a marker tool, the RNA-Seq information on pregnant uterine tissue can be used to gain novel mechanistic insights, such as by identifying alternative splicing events, allele-specific expression, and rare and novel transcripts that might be involved in the onset of maternal receptivity. This concept is unique and provides a new approach towards strategies that are highly needed to improve efficiency of fertility treatments.

Oxylipidomics of the cow uterus: effects of the peri-ovulatory endocrine milieu of the estrous cycle

Univ Estadual Paulista, Dept Clin Surg & Anim Reprod, Fac Vet Med, UNESP, Sao Paulo, BrazilUNESP, Sch Vet Med & Anim Sci, Dept Anim Reprod, Lab Adv Reprod & Cell Therapy LAN A, Botucatu, SP, Brazil

The peri-ovulatory endocrine millieu regulates the expression of polyamine synthetic pathway in the bovine endometrium

Univ Estadual Paulista, Dept Clin Surg & Anim Reprod, Fac Vet Med, UNESP, Sao Paulo, BrazilUNESP, Sch Vet Med & Anim Sci, Dept Anim Reprod, Lab Adv Reprod & Cell Therapy LAN A, Botucatu, SP, Brazil

Gene expression of glucose transporters in cattle endometrium in response to manipulations of the pre-ovulatory follicle size

Univ Estadual Paulista, Dept Clin Surg & Anim Reprod, Fac Vet Med, UNESP, Sao Paulo, BrazilUNESP, Sch Vet Med & Anim Sci, Dept Anim Reprod, Lab Adv Reprod & Cell Therapy LAN A, Botucatu, SP, Brazil