iProXProteomicsZhenGao SunHomo Sapienshttp://www.iprox.org/page/project.html?id=IPX0003461000sunzhengao77@126.comThe First Affiliated Hospital of Shandong University of traditional Chinese MedicineiProX<h4>Background</h4>Endometriosis (EM) leads to a decline in fertility, which is characterized by a decrease in the number and quality of follicles, and thus has a negative impact on in vitro fertilization (IVF) outcomes. However, the mechanism of how EM affects oocytes and leads to infertility remains unclear. As a potentially available sample directly related to oocyte growth, follicular fluid (FF) has important research value. Evaluating the association of FF content and EM-associated infertility through proteomics may helpful to explore the possible pathogenesis of EM-associated infertility.<h4>Methods</h4>In the present experimental study, from August 2019 to June 2020, FF samples were obtained as control group (CON-G; n = 10) from women with no one female factor of infertility and were undergoing IVF due to other reasons, 20 women with EM-associated infertility undergoing IVF with no other female factors were distributed into the EM group according to the time for IVF: (i) EM-group 1 (EM-G1, Stage I to Stage III, n = 10); (ii) EM-group 2 (EM-G2, Stage I to Stage III, n = 10). label-free quantitative proteomics (LFQP) technology and parallel reaction monitoring (PRM) approach were combined to aid in identifying and validating FF protein biomarkers for EM-associated infertility. In PRM analysis, another 20 subjects were enrolled as EM-associated infertility group (EM,Stage I to Stage III, n = 10) and controls (CON, n = 10) within the same time and inclusion criteria are the same as previously described. Finally, a potential protein biomarker panel of FF differential expressed proteins to EM-associated infertility was also evaluated by t-test and receiver operating characteristic (ROC) curve and binary Logistic regression models.<h4>Results</h4>7 significant differential expressed proteins which closely related to EM-associated infertility were found by LFQP technology, among which immunoglobulin lambda variable 7-46 (IGLV7-46), Immunoglobulin heavy constant gamma 2 (IGHG2), glia-derived nexin (GDN) and Inter-alpha-trypsin inhibitor heavy chain H3 (ITIH3) were significantly up-regulated (p < 0.05), while corticosteroid-binding globulin (CBG), angiotensinogen (AGT) and Fetuin-B (FETUB) were significantly down regulated (p < 0.05). Additionally, GDN and AGT was identified as a potential protein biomarker by further PRM analysis for EM-associated infertility according to ROC curve analysis and t-test (p < 0.05), the area under the curve (AUC) for GDN and AGT was 0.78 and 0.69 with optimum sensitivity of 50%, 70% and specificity of 100%, 90%, respectively. According to binary logistic regression and evaluated ROC analysis, the AUC for the combination of GDN and AGT was 0.80.<h4>Conclusions</h4>To the best of our knowledge, this is the first time that elevated GDN protein levels have been found in the FF of patients with EM-associated infertility. Combining LFQP technology and PRM method we found the abnormal of GDN and AGT in FF may be the potential cause of EM-associated infertility which may help to better understand the physiological and pathological mechanism of EM-associated infertility. Further experimental studies are required to confirm their mechanism in EM-associated infertility. The results of this study are also consistent with the previous conclusion that EM is a chronic inflammatory disease.<h4>Significance</h4>To the best of our knowledge, this is the first time that elevated GDN protein levels have been found in the follicular fluid of patients with EM-associated infertility. Combining LFQP technology and PRM methods we found the abnormal of GDN and AGT protein in FF may be the potential cause of EM-associated infertility which may help to better understand the physiological and pathological mechanism of EM-associated infertility. Clinically, it has been recognized that EM is related to infertility, but the mechanism remains unclear. Our study combines label-free quantitative proteomics technology and parallel reaction monitoring methods to identify and verify the FF protein biomarkers of EM-associated infertility, which provides a good research method for follow-up research.Quantitative label-free proteomic analysis of human follicle fluid to identify novel candidate protein biomarker for endometriosis-associated infertility.Cao Xian-Ling XL, Song Jing-Yan JY, Sun Zhen-Gao ZGfalseQuantitative label-free proteomic analysis of follicle fluid to identify novel candidate protein biomarker for endometriosis-associated infertilityBackground: Gold standard for endometriosis (EM) diagnosis tends to be invasive and histopathological exams following surgery are required. EM lack precise non-invasive biomarkers and current biomarkers do not provide detailed knowledge and are therefore deficient in precision that prevents early diagnosis and treatment of EM. Methods: In the present retrospective study, label-free quantitative proteomics(LFQP) technology was used to analyze the follicular fluid (FF) of infertile patients with EM in order to recognize FF biomarkers for EM. The participants were distributed into the EM group according to the severity of EM: (i) EM-group 1 (EM-G1, stage I to Stage II, n=10); (ii) EM-group 2 (EM-G2, Stage III to Stage IV, n=10) and the control-group (CON-G, n=10, infertility due to male factors) from August 2019 to June 2020. A potential biomarker panel of FF differential protein to EM-associated infertility was also evaluated by receiver operating characteristic (ROC) curve and binary Logistic regression models. Results: A total of 237 and 236 proteins were identified in the EM-associated infertility and control groups, respectively; After untargeted proteomic analysis, 6 significant differential proteins were found in both EM-G1 VS. CON-G and EM-G2 VS. CON-G. Through bioinformatics analysis, we identified a several groups of proteins that may be closely related to EM, among which immunoglobulin lambda variable 7-46 (IGLV7-46), immunoglobulin heavy constant gamma 2 (IGHG2), glia-derived nexin (GDN) and Inter-alpha-trypsin inhibitor heavy chain H3 (ITIH3) were significantly up-regulated, while corticosteroid-binding globulin (CBG) and angiotensinogen (AGT) were significantly down regulated. According to ROC curve analysis, the area under the curve (AUC) for IGLV7-46, IGHG2, GDN, ITIH3, AGT and CBG was 0.87, 0.82, 0.78, 0.82, 0.77 and 0.81 with optimum sensitivity of 78%, 80%, 100%, 80%, 70%, 100% and specificity of 93%, 80%, 60%, 80%, 80% and 60% respectively. According to binary logistic regression and evaluated ROC analysis, the AUC for the combination of IGLV7-46, IGHG2 and ITIH3 was 0.926. Conclusions: The research use LFQP to study the FF proteome of EM-associated infertility and totally scan out 6 differentially expressed proteins. These proteins participate primarily in the inflammation and immune response phase. Additionally, ROC analysis suggests that the IGLV7-46, IGHG2 and ITIH3 are likely strong diagnostic biomarkers of EM-associated infertility, which may be a way of thinking for identifying possible minimally invasive diagnostic indicators and therapeutic goals of EM-associated infertility.Thu Sep 02 00:00:00 GMT+01:00 2021PXD028199960635811008