Proteomics & Glyco-Proteomic Analysis of Follicular Fluid
Proteomics & Glyco-Proteomic Analysis of Follicular Fluid
To the best of the investigators knowledge, exhaustive characterization of the low and high
 abundant proteins and glyco-proteins of the Follicular Fluid (FF) has not yet been achieved.
 Such an analysis may provide critical molecular data on the role of the FF in oocyte
 maturation and may identify specific changes in the FF proteome of patients with gynecologic
 problems, such as Polycystic Ovary Syndrome (PCOS).

 Specific Aims

 1. To perform a comprehensive analysis of normal human FF using sensitive mass spectrometry
 in combination with conventional approaches for proteomic evaluation and using HPLC and
 Western blot for glyco-proteomic analysis.

 2. Characterize differential proteomic and glyco-proteomic patterns of the FF in normal
 women compared to lean and obese women with PCOS.

 3. To supplement the differential proteomic and glyco-proteomic analysis with steroid
 hormone analysis in all FF samples.
In this study, we plan to utilize ultrasensitive mass spectrometry (MS) and other
 conventional proteomic approaches to identify the low and high abundant proteins present in
 human FF. Additionally, we plan to use high-performance liquid chromatography (HPLC) and
 Western blot techniques to evaluate the Neu5Gc and glycan array based ELISA techniques to
 detect anti-Neu5Gc antibody profile in human FF. This analysis will be performed on FF
 samples obtained from normal women undergoing In-Vitro Fertilization and Embryo Transfers
 (IVF-ET) for a male factor alone and oocyte donors from our 3rd Party Reproduction Program
 and from lean and obese women with PCOS. This study will provide information on protein,
 glycoprotein, and steroid hormone expression during normal folliculogenesis and during the
 pathologic condition of PCOS, which should also provide basic scientific information on
 normal and abnormal oocyte development.

 Human FF bathes the developing oocyte. Previous studies indicate that the FF contains
 cytokines, steroidal and protein hormones, and growth factors. The presence of proteins with
 such significant biological properties implies a paracrine and autocrine role for the FF in
 promoting normal oocyte development. Furthermore, the presence of any antigenic sialic acid
 Neu5Gc and the presence of antibodies targeting these antigenic glycoconjugates (glycolipid
 and glycoproteins decorated with sialic acid) may interfere with oocyte development, hormonal
 expression, fertilization, and possibly implantation. Here we hypothesize that an exhaustive
 proteomic and glyco-proteomic characterization of human FF is essential for a thorough
 understanding of its biological significance. We also hypothesize that PCOS may have
 differential expression of the FF protein and glyco-protein milieu, and that the expression
 may differ further between lean and obese women with PCOS. PCOS represents a heterogeneous
 disorder. The severity of hyperandrogensim, metabolic and menstrual disturbance, and obesity
 is variable with up to 40% not clinically expressing signs of classic hyperandrogenism. On
 the other hand, these atypical, often lean, PCOS women can have impaired glucose tolerance
 and diabetes. Reports suggest that these lean PCOS women have altered serum IGFBP-1, a
 characteristic endocrine feature of patients with obese PCOS, and related to hyperinsulinemia
 and/or obesity. The lean phenotype of PCOS and its significance is unclear but may represent
 a cryptic or unexpressed form of PCOS or may be a prelude to individuals who will later
 manifest clinical signs of obese/overweight PCOS. Changes in expression may be expected
 because of the different amounts of steroidal hormones and inflammatory markers in the FF
 derived from women with PCOS.
Inclusion criteria: Inclusion Criteria All

 1. Female patients undergoing controlled ovarian hyperstimulation (COH), transvaginal
 oocyte aspiration (TVA), and Saline Infused Sonography (SIS) with UL collection

 2. Age <35 y/o at time of in vitro fertilization (IVF) cycle

 3. Normal ovarian function defined Day 3 Follicular Stimulating Hormone (FSH) <8 pg/ml or
 Anti-Mullerian Hormone (≥ 1.0 ng/ml)

 Inclusion Criteria Controls:

 1. Female patients undergoing COH and TVA donating her oocytes

 2. Female patients undergoing COH and TVA for male factor infertility only (i.e. no
 female causes of infertility)

 3. Normal menstrual cycles

 Inclusion Criteria Lean PCOS:

 1. Diagnosis of PCOS by Rotterdam Criteria

 2. BMI ≤ 25 kg/m2 Inclusion Criteria Classic PCOS

 1. Diagnosis of PCOS by Rotterdam Criteria 2. BMI > 30 kg/m2

 Exclusion criteria:

 1. Age ≥ 35 y/o

 2. Female partners with infertility associated diagnosis (i.e. tubal factor, cervical
 factor, endometriosis)

 3. Unexplained infertility