The aims of this study were: (i) to characterise the global changes in the composition of the uterine luminal fluid (ULF) from pregnant heifers during pregnancy recognition (Day 16) using nano-LC MS/MS; (ii) to describe quantitative changes in selected proteins in the ULF from Day 10, 13, 16 and 19 by Isobaric tags for Relative and Absolute Quantification (iTRAQ) analysis; and (iii) to determine whether these proteins are of endometrial or conceptus origin, by examining the expression profiles of the associated transcripts by RNA sequencing. On Day 16, 1652 peptides were identified in the ULF by nano-LC MS/MS. Of the most abundant proteins present, iTRAQ analysis revealed RPB4, TIMP2 and GC had the same expression pattern as IFNT, while the abundance of IDH1, CST6 and GDI2 decreased on either Day 16 or Day 19. ALDOA, CO3, GSN, HSP90A1, SERPINA31 and VCN proteins decreased on Day 13 compared to Day 10, but subsequently increased on Day 16 (P<0.05). PNP and HSPA8 decreased on Day 13, increased on Day 16 and decreased and increased on Day 19 (P<0.05). The abundance of CATD, CO3, CST6, GDA, GELS, IDHC, PNPH and TIMP2,mRNAs was greater (P<0.001) in the endometrium than conceptus. In contrast, the abundance of ACTB, ALDOA, ALDR CAP1, CATB, CATG, GD1B, HSP7C, HSP90A, RET4 and TERA was greater (P<0.05) in the conceptus compared with the endometrium. In conclusion, significant changes in the protein content of the ULF during the pre-implantation period of pregnancy reflecting the morphological changes that occur in the conceptus.
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