Proteomic Analysis of Follicular Fluid from Patients with Endometriosis Submitted to Controlled Ovarian Hyperstimulation for In Vitro Fertilization.

Endometriosis, a chronic gynecological disease characterized by the presence of functional endometrial tissue outside the uterine cavity, is associated to infertility. Many studies report that pregnancy rates are lower in women with endometriosis than in normal controls, but the specific mechanisms that may account for this difference are poorly understood. Endometriosis affects 10-32% of women of reproductive age and can result in pelvic pain and infertility. There are numerous manners to diagnose this disease, but the current gold standard for diagnosis for endometriosis is laparoscopy. In recent years the development of high-throughput technologies such as proteomics has led to early diagnosis of the disease, by comparing the protein composition in diseased and normal tissues. Aside from accelerating diagnosis, proteomics may lead to a better understanding of the physiopathology of disease development by identifying proteins involved during its different stages. Thus, we set out to evaluate protein profiles of women with endometriosis achieving pregnancy and not achieving pregnancy, and to verify differentially expressed proteins. Follicular fluid samples were obtained from 12 women with endometriosis grade III and IV (study group) (age 32.16 ± 2.62 years, mean ± SD). Endometriosis diagnosis was performed by videolaparoscopy. These women were divided into two groups--E.P and E.NP--patients with endometriosis who achieved pregnancy (n=4) and who did not achieve pregnancy (n=8), respectively. Follicular fluid samples from different patients were pooled according to group, and each pool was run in triplicates in order to increase specificity of the test. Proteins were separated based on their isoelectric point and molecular weight by two-dimensional electrophoresis, in which each spot represents a protein. Spots were compared after densitometry using, excised, and submitted to matrix-assisted laser desorption/ionization time-of-flight (MALDI-ToF) mass spectrometry. A total of 27 spots were different between groups E.P and E.NP. Two spots were more intense in group E.P, 3 spots were only visualized in group E.P, 16 spots were more intense in group E.NP, and 6 spots were visualized only in group E.NP. Of all the identified proteins, 6 are related to reproduction: (i) apolipoprotein A-IV (APOA 4), (ii) alpha-I antitrypsin (SERPINA 1), (iii) vesicle transport through interaction with t-SNAREs (VTI1A), (iv) Putative zinc finger protein 818 (ZNF818P), (v) haptoglobin, and (vi) Tektin-2. Table 1 lists all the proteins determined. APOA 4, SERPINA 1, and VTI1A all participate in lipid metabolism. The fact that that their content was higher in women with endometriosis who did not achieve pregnancy may indicate that a possible pathway through which endometriosis effects decreased pregnancy rates is through alteration in lipid and cholesterol pathways within the follicle. This is in accordance with hormonal imbalances commonly observed in these women. In conclusion, endometriosis protein profiles have been presented, and patients who are able to achieve pregnancy present at least 27 well-defined distinct spots, of which 6 identified proteins have a known function related to reproduction. These proteins may be potential targets for therapeutic intervention in women with infertility due to endometriosis, as well as assist in comprehending the physiopathologic mechanisms underlying endometriosis. (platform)