Endometrial mRNA expression of selected pro-inflammatory factors and mucins in repeat breeder cows with and without subclinical endometritis

Abstract

Repeat breeder cows (RBC) are defined as cyclic cows without clinical abnormalities that fail to conceive after at least three subsequent inseminations. Previous studies have elucidated cellular defence mechanisms in the bovine uterus but detailed information on inflammatory events of endometrial cells in RBC is still lacking. Thus, the objective of this study was to analyse endometrial mRNA expression of selected transcripts associated with uterine inflammatory processes. Cytobrush samples from 91 RBC and 11 synchronised heifers with no history of gynaecological abnormalities (controls, CON) were collected. The proportion of polymorphonuclear neutrophils in these samples was used for the diagnosis of subclinical endometritis (SE). Ultrasonography and progesterone blood concentrations were used to determine ovarian activity and the stage of the oestrous cycle. Total RNA was isolated from the cytobrush samples and subjected to reverse transcription-quantitative PCR for interleukins (IL) 1A, IL1B, IL6, IL8, chemokine CXL ligand (CXCL) 3, CXCL5, prostaglandin-endoperoxide synthase 2 (PTGS2), tracheal antimicrobial peptide (TAP) and mucin (MUC) 4, MUC5, MUC6, MUC12 and MUC16. CXCL3 mRNA was higher (2-fold) and PTGS2 mRNA lower (6-fold) expressed in RBC compared with CON (P < 0.05). After subdivision of RBC in animals with (RBC-SE) and without SE (RBC-noSE), these differences remained significant between RBC-noSE and CON. Higher mRNA abundances of IL1A and IL1B were found in RBC-SE compared with RBC-noSE (3- and 4-fold; P < 0.05). No differences in the mRNA expression of IL6, IL8, CXCL5 and TAP were observed between RBC-SE, RBC-noSE and CON. MUC4 and MUC12 mRNA was more highly expressed in RBC than in CON (P < 0.05). In RBC-noSE, a 5- and 14-fold higher MUC4 and MUC12 mRNA expression was noticed compared with CON (P < 0.05). A significantly lower mRNA expression of MUC5 and MUC16 (7- and 4-fold) was detected in RBC in the luteal phase compared with RBC in the follicular phase, whereas such a down-regulation was not observed for MUC4 and MUC12. In conclusion, we demonstrated different PTGS2 and CXCL3 mRNA expression between RBC and control heifers, which might be related to subfertility in RBC. Further studies are required to confirm that an unregulated MUC4 and MUC12 mRNA expression may contribute to subfertility of RBC. These findings provide a valid basis for further research on regulatory mechanisms of mRNA expression in subfertile cows.