Proteome changes of porcine follicular fluid during follicle development

Ovarian follicular fluid influences follicle and oocyte growth, but the fluctuation of its protein content during folliculogenesis has not been comprehensively analyzed. Here we used a shotgun approach and bioinformatics analyses to investigate and compare the proteomes of porcine follicular fluid (pFF) obtained from small (< 4 mm), medium (4–6 mm) and large (> 6–12 mm) follicles. Follicular fluid samples containing highest estrogen levels were selected as non-atretic from small (SNA: 26.1 ± 15 ng/mL), medium (MNA: 162 ± 54 ng/mL), and large (LNA: 290 ± 37 ng/mL) follicles for proteomic analyses. We detected 1627, 1699, and 1756 proteins in SNA, MNA, and LNA samples, respectively. Nearly 60–63% of total proteins were specific to each sample, 11–13% were shared in pairwise comparisons, and 247 proteins were shared among all samples. Functional categorization indicated comparable gene ontology (GO) terms distribution per cellular component, molecular function, and biological process categories across samples; however, the ranking of highly significantly enriched GO terms per category revealed differences between samples. The patterns of protein-to-protein interactions varied throughout follicle development, and proteins such as serine protease inhibitor, clade E (SERPINE); plasminogen activator, urokinase (PLAU); and plasminogen activator, urokinase receptor (PLAUR) appeared stage-specific to SNA, MNA, and LNA, respectively. The “complement and coagulation cascades” was the common major pathway. Besides, properdin and fibulin-1 were abundant proteins that appeared absent in LNA samples. This study provides extensive and functional analyses of the pFF proteome changes during folliculogenesis and offers the potential for novel biomarker discovery in pFF for oocyte quality assessment.