Whole-Transcriptome Profiling of Ovarian Tissues in Gilts with Normal Estrus and Follicular Cyst-Associated Anestrus

We explored the potential functions of differentially expressed miRNAs, mRNAs, lncRNAs, and circRNAs identified in the ovaries of gilts with normal estrus (NE) and follicular cyst-associated anestrus (AE), as well as their putative competing endogenous RNA (ceRNA) regulatory networks. Ovarian morphology was assessed via ultrasonography, and serum concentrations of Follicle-stimulating hormone (FSH), (Estradiol) E2, and Progesterone(P4) were measured. Ovarian tissues were harvested after slaughter for whole-transcriptome sequencing. Bioinformatic tools were used to screen differentially expressed RNAs(DERNAs) between NE and AE gilts. We further predicted target interactions among these transcripts, conducted functional enrichment analysis on target genes, and constructed candidate ceRNA regulatory networks potentially associated with gilt estrus. Phenotypic verification confirmed that ovarian ultrasonographic characteristics, histological morphology, and serum reproductive hormone levels were consistent with the physiological status of NE and AE gilts. Under the screening thresholds of p < 0.05 and |log2FC| ≥ 1, we identified 22 lncRNAs that may interact with 21 mRNAs via 50 miRNAs, alongside 39 circRNAs predicted to regulate 26 mRNAs through 72 miRNAs. Functional enrichment analysis indicated that target genes of these differentially expressed transcripts were predominantly enriched in the lysosome pathway, PPAR signaling pathway, chemokine signaling pathway, cholesterol metabolism and NOD-like receptor signaling pathway. Hub molecules including FGF1, GHR, TLR2, ssc-miR-370, and miR-21-5p were shared in lncRNA/circRNA-miRNA-mRNA regulatory networks; these molecules have been reported to participate in progesterone synthesis, estrus modulation, and endocrine homeostasis. Of particular interest, two non-coding RNAs, MSTRG.1285.1 and novel_circ_056113, were predicted to act as candidate ceRNAs that may sponge ssc-miR-370 and miR-21-5p, which could in turn modulate the expression of estrus-associated mRNAs including FGF1, GHR, and TLR2. The expression trends of MSTRG.1285.1, novel_circ_056113, miR-370, miR-21-5p, GHR, TLR2, and FGF1 were validated by qRT-PCR, and the quantification results agreed with transcriptome sequencing data. Collectively, this study constructed a predicted ceRNA regulatory network of ovarian transcripts comparing NE and AE gilts and uncovered multiple RNA molecules potentially involved in estrus regulation. These findings offer preliminary theoretical clues for exploring the onset of puberty in gilts.