Proteomic Analysis of Human Follicular Fluid Using an Alternative Bottom-Up Approach

Abstract

Human follicular fluid (hFF) is the in vivo environment of oocytes during follicular maturation in the ovaries. It contains a huge variety of compounds such as, e.g., proteins that might play an important role in follicular development and oocyte growth. Previous proteomic studies on follicular fluid have isolated and already identified a certain number of proteins. Nevertheless, only a small part of proteins present in follicular fluid have been covered so far and a large number have still not been identified. Therefore, the need for new, more resolving, and sensitive approaches in proteome research is evident. We utilized a proteomic setup based on in solution isoelectric focusing (IEF) and reversed-phase nanoliquid chromatography coupled to matrix-assisted laser desorption/ionization time-of-flight tandem mass spectrometry (nano-LC MALDI TOF/TOF MS) for in depth protein analysis of human follicular fluid samples of patients undergoing controlled ovarian hyper stimulation (COH) for in vitro fertilization therapy (IVF). This approach led to the significant identification of 69 proteins, where 32 have not been reported before to be found in human follicular fluid with proteomic methods. Among these findings, at least two relevant compounds essentially involved in hormone secretion regulation during the folliculogenetic process were identified: sex hormone binding globulin (SHBG) and inhibin A (INHA). To confirm these results, both proteins were further validated by immunoassays.

Preliminary proteomic analysis on the alterations in follicular fluid proteins from women undergoing natural cycles or controlled ovarian hyperstimulation

“…On the other hand, a high follicular level of ZAG was associated with failed pregnancy of the women undergoing IVF-ET [14]. ZAG is one of the most abundant proteins in human FF [30], and plays a role in immune modulation during the follicular development. The levels of ZAG in FF were correlated with the concentrations of steroids in FF [14].…”

A proteomic analysis of human follicular fluid: comparison between fertilized oocytes and non-fertilized oocytes in the same patient

“…Hanrieder et al identified 69 proteins in the follicular fluids of seven females obtained during IVF using isoelectric focusing and reversed-phase nanoliquid chromatography coupled to matrix-assisted laser desorption/ionization time-of-flight tandem mass spectrometry [16]. Jarkovska et al [11].…”

Complement C3 and anti-testosterone antibody in the 150 kDa Bali cattle (Bos sondaicus/javanicus) subordinate follicular fluid peptide observed using MALDI-TOF/TOF-MS

“…Complement C3 is a plasma protein that is abundantly synthesized by hepatocytes (Zhou et al, 2016), the presence of this protein in the follicular fluid mostly through the diffusion of the protein over the bloodfollicle barrier, which its permeability is raised during follicle maturation (Hanrieder et al, 2008). C3 complement has also been reported to be synthesized and secreted by malignant ovarian epithelial cells (Cho et al, 2014), resulting in the activation of complement cascade in the tumor microenvironment and increases cancer cell proliferation, and metastasis.…”