MSACL 2019 EU Abstract

Prognostic Potential of Combined Omics Analysis for Diagnostics of Azoospermia

A.G. Brzhozovzkiy (1), N.L. Starodubtseva (1), A.S. Kononikhin (1,2), A.E. Bugrova (1), V.V. Chagovets (1), A. Tambiev (1), S.I. Gamidov (1), V.E. Frankevich (1), E.N. Nikolaev (2,3), G.T. Sukhikh (1)
(1) V. I. Kulakov National Medical Research Center for Obstetrics, Gynecology and Perinatology of Ministry of Healthcare of Russian Federation, Moscow, Russia; (2) V.L. Talrose Institute for Energy Problems of Chemical Physics, Russian Academy of Sciences, Moscow, Russia; (3) Skolkovo Institute of Science and Technology, Skolkovo, Russia.

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Alexnder Brzhozovskiy (Presenter) National Medical Research Center for Obstetrics

Presenter Bio: I graduate biomedical faculty of Pirogov Russian National Research Medical University at 2014 year. Being a second year student I start my work as a laboratory assistant at laboratory of microbiology and bacteriology of my university. At the end of my education I wrote my diploma work called «Species Diversity of Bifidobacteria in the Intestinal Microbiota Studied Using MALDI-TOF Mass-Spectrometry». At 2014 I started my PHD work at molecular biology using proteomic methods such as mass-spectrometry. From 2017 year I work at Research Center of Obstetrics, Gynecology and Perinatology, Ministry of Health of Russia as a junior researcher. Do not include CV. Do not include publications.

Relevant Financial Disclosures (within past 24 months) No relevant financial relationship(s) to disclose. Conflict mitigation NOT REQUIRED.

Background: The male factor contributes to more than 50% of couples with conceive failure. Seminal plasma seems to be a promising noninvasive informative source for male infertility biomarkers. The possibility combined omics analysis of seminal plasma composition in defining the residual spermatogenesis during azoospermia was tested by the supervised OPLS-DA. Quantitative parameters of the model obtained turned out to be satisfactory to differentiate patients with residual spermatogenesis.
Methods: Seminal plasma samples were obtained from 36 male patients from 21 to 45 years (33.3 ± 3.9 years), examined and treated for azoospermia. Clinical examination includes clinical and anamnestic data collection. A testicular sperm micro extraction method was used for the residual spermatogenes.
For proteomic analysis the in-solution samples preparation method was used followed by trypsin digestion. Tryptic peptide mixtures were separated on a nano-HPLC Agilent 1100 system (Agilent Technologies) combined with 7 T LTQ-FT Ultra (Thermo Electron, Bremen, Germany) using a nanospray ion source. For label-free semi-quantitative analysis MS files were processed with MaxQuant software (version 1.6.2) against SwissProt database.
Lipids were extracted from supernatant by modified Folch method. LC-MS/MS analysis was performed via liquid chromatography Dionex Ultima 3000 (Thermo Fisher Scientific) combined with a hybrid time-of-flight mass spectrometer Maxis Impact (Bruker Daltonics) operating in the positive-ion ESI mode and negative-ion ESI mode.
Results: To assess the changes in seminal plasma proteome composition during azoospermia Welch’s t-test (p-value p<0.01) was performed and resulted in 18 significantly changed proteins. Previously, these proteins were shown to be involved in spermatogenesis. Thus, the most altered processes in the presence/absence of residual spermatogenesis in the ovaries (testicles) are immune system process (11 proteins), wound response (8) and in homeostatic processes (5).To assess the changes in seminal plasma lipids composition during azoospermia Manna-Whitney test (p-value<0.05) was performed and resulted in 69 significantly changed lipids. Analysis of lipid’s level shows that in azoospermic fluid levels of lysophosphatidylcholines and sphingomyelins are lower and levels of phosphatidylcholines, phosphatidylethanolamines and phosphatidylserines are higher.
Novel aspect: The possibility analysis in samples differentiation was tested by the supervised OPLS-DA. The score plot demonstrated good clustering of dots corresponding to distinct group of patients with azoospermia (with and without residual spermatogenesis).