Serum exosomal miR-192 serves as a potential detective biomarker for early pregnancy screening in sows

• Published in: Animal Bioscience Vol. 36; no. 9; pp. 1336 - 1349
• Main Authors: Gao, Ruonan, Li, Qingchun, Qiu, Meiyu, Xie, Su, Sun, Xiaomei, Huang, Tao
• Format: Journal Article
• Language: English
• Published: Korea (South) Asian - Australasian Association of Animal Production Societies 01.09.2023; Animal Bioscience; Asian-Australasian Association of Animal Production Societies; 아세아·태평양축산학회
• Subjects: Biological markers; biomarkers; Cell organelles; early pregnancy; exosomal mirnas; Genetic aspects; Health aspects; itga4; MicroRNA; mir-192; Swine; 축산학; China; Biomarkers; Early pregnancy; Exosomal miRNAs; ITGA4; miR-192
• ISSN: 2765-0189; 2765-0235
• DOI: 10.5713/ab.22.0422

The study was conducted to screen differentially expressed miRNAs in sows at early pregnancy by high-throughput sequencing and explore its mechanism of action on embryo implantation. The blood serum of pregnant and non-pregnant Landrace×Yorkshire sows were collected 14 days after artificial insemination, and exosomal miRNAs were purified for high throughput miRNA sequencing. The expression patterns of 10 differentially expressed (DE) miRNAs were validated by qRT-PCR. Quantitative reverse transcription-polymerase chain reaction (qRT-PCR) quantified the abundance of serum exosomal miR-192 in pregnant and control sows, and the diagnostic power was assessed by receiver operating characteristic (ROC) analysis. The target genes of DE miRNAs were predicted with bioinformatics software, and the functional and pathway enrichment analysis was performed on Gene Ontology (GO) and the Kyoto Encyclopedia of Genes and Genomes (KEGG) terms. Furthermore, a luciferase reporter system was used to identify the target relation between miR-192 and integrin alpha 4 (ITGA4), a gene influencing embryo implantation in pigs. Finally, the expression levels of miRNAs and the target gene ITGA4 were analyzed by qRT-PCR, and western blot, with the proliferation of BeWo cells detected by CCK-8. A total of 221 known miRNAs were detected in the libraries of the pregnant and non-pregnant sows, of which 55 were up-regulated and 67 were down-regulated in the pregnant individuals compared with the non-pregnant controls. From these, the expression patterns of 10 DE miRNAs were validated. The qRT-PCR analysis further confirmed a significantly higher expression of miR-192 in the serum exosomes extracted from pregnant sows, when compared to controls. ROC analysis revealed that miR-192 provided excellent diagnostic accuracy for pregnancy (area under the ROC curve [AUC] = 0.843; P>0.001). The dual-luciferase reporter assay indicated that miR-192 directly targeted ITGA4. The protein expression of ITGA4 was reduced in cells that overexpressed miR-192. Overexpression of miR-192 resulted in the decreased proliferation of BeWo cells and regulated the expression of cell cycle-related genes. Serum exosomal miR-192 could serve as a potential biomarker for early pregnancy in pigs. miR-192 targeted ITGA4 gene directly, and miR-192 can regulate cellular proliferation.