Qualitative and Quantitative Analysis of Proteome and Peptidome of Human Follicular Fluid Using Multiple Samples from Single Donor with LC–MS and SWATH Methodology

Abstract

Human follicular fluid (hFF) is a natural environment of oocyte maturation, and some components of hFF could be used to judge oocyte capability for fertilization and further development. In our pilot small-scale study three samples from four donors (12 samples in total) were analyzed to determine which hFF proteins/peptides could be used to differentiate individual oocytes and which are patient-specific. Ultrafiltration was used to fractionate hFF to high-molecular-weight (HMW) proteome (>10 kDa) and low-molecular-weight (LMW) peptidome (<10 kDa) fractions. HMW and LMW compositions were analyzed using LC-MS in SWATH data acquisition and processing methodology. In total we were able to identify 158 proteins, from which 59 were never reported before as hFF components. 55 (45 not reported before) proteins were found by analyzing LMW fraction, 67 (14 not reported before) were found by analyzing HMW fraction, and 36 were identified in both fractions of hFF. We were able to perform quantitative analysis for 72 proteins from HMW fraction of hFF. We found that concentrations of 11 proteins varied substantially among hFF samples from single donors, and those proteins are promising targets to identify biomarkers useful in oocyte quality assessment.

Seasonal variation in equine follicular fluid proteome

“…The present study applied strict filters (FDR < 1%, and the minimum of two unique peptides per protein) to obtain proteins with high confidence identification, which may explain the slightly lower number of detected proteins (90 vs. 113) when compared to available FF mare proteome [28]. In addition, the proteome dataset of the current study contains fewer proteins in comparison to other monovular species such as humans (158 to 1079; [24, 37–40]), and dairy cattle (113 to 219; [15, 21]). Nonetheless, the aforementioned proteomic studies were generally performed with FF samples obtained from follicles of different sizes and unknown physiological statuses (i.e., growing and regressing follicles), under different technical approaches (e.g., gel-based or gel-free) and protein call stringencies (e.g., false discovery rate and peptides).…”

Anti-Candida albicans effect of the protein-carbohydrate fraction obtained from the coelomic fluid of earthworm Dendrobaena veneta

“…The reduced and alkylated sample was divided into three vials and digested overnight individually with trypsin, chymotrypsin, and endopeptidase Lys-C, E:S 1:50 w/w [35]. The digestion was stopped by acidifying and desalting the solution according to the standard protocol [36].…”

S100‐A9 protein in exosomes derived from follicular fluid promotes inflammation via activation of NF‐κB pathway in polycystic ovary syndrome

“…Although proteome and peptidome were identificated in human FF, little was known about the proteomic analysis of exosomes in follicular fluid and their potential roles in follicular development and reproduction‐related disorders. Therefore, we used an TMT‐tagged quantitative proteomic approach to compare the proteomic profile of exosomes derived from FF of PCOS and healthy women.…”

“…[21][22][23] Recently, some studies uncovered the potential function of exosomes derived from FF as carriers of miRNAs in steroidogenesis, follicular development and other pathological conditions. [24][25][26][27] Although proteome and peptidome were identificated in human FF, [28][29][30][31][32][33] little was known about the proteomic analysis of exosomes in follicular fluid and their potential roles in follicular development and reproduction-related disorders. Therefore, we used an TMT-tagged quantitative proteomic approach to compare the proteomic profile of exosomes derived from FF of PCOS and healthy women.…”