In cattle, the oviduct plays a major role in the reproductive process; however, molecular control of oviduct receptivity to the embryo is poorly understood. A model for receptivity based on size of the pre-ovulatory follicle (POF) was used to compare oviductal morphology, cellular proliferation, and candidate transcript abundance. Growth of the POF of Nelore (Bos indicus) cows was manipulated to produce two groups: a large POF-large corpus luteum (CL) group (LF-LCL; greater receptivity) and a small POF-small CL group (SF-SCL). Samples of the ampulla and isthmus ipsilateral and contralateral to CL were collected 4 days after GnRH-induced ovulation. Tissues were either embedded in paraffin for Harris-Hematoxylin and Eosin and periodic acid-Schiff staining and KI67 immunostaining, followed by morphological analyses, or stored at -80 °C for RNA extraction, cDNA synthesis, and qPCR analyses. The effects of group (LF-LCL and SF-SCL), region (ampulla and isthmus), and side (ipsilateral and contralateral) were analyzed using three-way nested ANOVA. The ipsilateral ampulla of the LF-LCL group presented more primary mucosal folds, a greater mucosal-folding grade and luminal perimeter, and more secretory cells and proliferating cells when compared with the ampulla of the SF-SCL group and with the contralateral ampulla of both groups. There were no morphological differences in the isthmus between groups and sides. Changes in transcript abundance are suggestive of LF-LCL-stimulated secretory activity. In summary, ovulation of a larger POF generates a periovulatory endocrine milieu that modulates morphological and functional features of the bovine oviduct which may support embryo survival and development.
Information on molecular mechanisms through which sex-steroids regulate oviductal function to support early embryo development is lacking. Here, we hypothesized that the periovulatory endocrine milieu affects the miRNA processing machinery and miRNA expression in bovine oviductal tissues. Growth of the preovulatory follicle was controlled to obtain cows that ovulated a small follicle (SF) and subsequently bore a small corpus luteum (CL; SF-SCL) or a large follicle (LF) and large CL (LF-LCL). These groups differed in the periovulatory plasmatic sex-steroid’s concentrations. Ampulla and isthmus samples were collected on day four of the estrous cycle. Abundance of DROSHA, DICER1, and AGO4 transcripts was greater in the ampulla than the isthmus. In the ampulla, transcription of these genes was greater for the SF-SCL group, while the opposite was observed in the isthmus. The expression of the 88 most abundant miRNAs and 14 miRNAs in the ampulla and 34 miRNAs in isthmus were differentially expressed between LF-LCL and SF-SCL groups. Integration of transcriptomic and miRNA data and molecular pathways enrichment showed that important pathways were inhibited in the SF-SCL group due to miRNA control. In conclusion, the endocrine milieu affects the miRNA expression in the bovine oviduct in a region-specific manner.
The extracellular matrix (ECM) is a group of molecules that offer structural and biochemical support to cells and interact with them to regulate their function. Also, growth factors (GFs) stored in the ECM can be locally released during ECM remodeling. Here, we hypothesize that the balance between ECM components and remodelers is regulated according to the ovarian steroid milieu to which the oviduct is exposed during the periovulatory period. Follicular growth was manipulated to generate cows that ovulated small follicles (SF-small corpus luteum [SCL]; n = 20) or large follicles (LF-large corpus luteum [LCL]; n = 21) and possess corresponding Estradiol (E2) and Progesterone (P4) plasmatic concentrations. Ampulla and isthmus samples were collected on day 4 (day 0 = ovulation induction) and immediately frozen or fixed. The transcriptional profile (n = 3/group) was evaluated by RNA sequencing. MMP Antibody Array was used to quantify ECM remodelers' protein abundance and immunohistochemistry to quantify type I collagen. Transcriptome analysis revealed the over-representation of ECM organization and remodeling pathways in the LF-LCL group. Transcription of ECM components (collagens), remodelers (ADAMs and MMPs), and related GFs were upregulated in LF-LCL. Protein intensities for MMP3, MMP8, MMP9, MMP13, and TIMP4 were greater for the LF-LCL group. Type I collagen content in the mucosa was greater in SF-SCL group. In conclusion, that the earlier and more intense exposure to E2 and P4 during the periovulatory period in LF-LCL animals stimulates ECM remodeling. We speculate that differential ECM regulation may contribute to oviductal receptivity to the embryo.
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