Quantitative proteomic analysis of seminal plasma and sperm and its association with sperm functional aspects and lipid peroxidation levels of seminal plasma.

• Grant number: 11/14631-7
• Support Opportunities: Scholarships in Brazil - Master
• Start date: May 01, 2012
• End date: April 30, 2014
• Field of knowledge: Health Sciences - Medicine
• Principal Investigator: Ricardo Pimenta Bertolla
• Grantee: Paula Intasqui Lopes
• Host Institution: Escola Paulista de Medicina (EPM). Universidade Federal de São Paulo (UNIFESP). Campus São Paulo. São Paulo , SP, Brazil

Abstract

The main mechanisms involved in male infertility are changes in the functional quality of spermatozoa and oxidative stress, which can be evaluated in the laboratory in order to obtain further information on male reproductive potential. However, functional tests have no clinical impact, since they show limited accuracy for clinical purposes. Thus, molecular studies of sperm and seminal plasma associated with functional changes of spermatozoa are important from a clinical perspective.This project proposes the proteomic study of the seminal plasma and sperm in association with lipid peroxidation levels of seminal plasma and sperm mitochondrial, acrosomal, and DNA integrity. Initially, the following sperm functional tests will be conducted: analysis of mitochondrial activity, by the DAB (3,3'-diaminobenzidine) staining method; evaluation of acrosomal integrity, using PNA (Peannut Agglutinin) probes; analysis of DNA fragmentation using the alkaline Comet assay. Then, seminal plasma will be separated from the cell fraction counterpart, and the latter will be processed for selection of viable sperm. Also, an aliquot of seminal plasma will be used to evaluate lipid peroxidation levels by the TBARS (Thiobarbituric acid reactive substances) assay. Then, samples will be distributed in experimental groups, according to sperm functional quality and level of lipid peroxidation. Seminal plasma and sperm samples from each group will be used for proteomics studies, which will be performed by nanoUPLC tandem nanoESI-MSE.This project aims to identify possible protein markers for sperm functional quality, which may be used in the future for diagnosis and treatment of male infertility and for selection of undamaged sperm for injection into the oocyte.