Comparative proteomic network signatures in seminal plasma of infertile men as a function of reactive oxygen species.

Ashok Agarwal,Ahmet Ayaz,Mourad Assidi,Luna Samanta,Edmund Sabanegh,Adel M Abuzenadah,Rakesh Sharma

doi:10.1186/s12014-015-9094-5

Abstract

BackgroundReactive oxygen species (ROS) plays a major role in the pathology of male infertility. It is an independent biomarker of sperm function. Seminal plasma is a natural reservoir of antioxidants responsible for the nourishment, protection, capacitation, and motility of sperm within the female reproductive tract resulting in successful fertilization and implantation of the embryo. A comparative proteomic analysis of seminal plasma proteins from fertile men and infertile men with varying levels of ROS was carried out to identify signature proteins involved in ROS-mediated reproductive dysfunction.MethodsA total of 42 infertile men presenting with infertility and 17 proven fertile donors were enrolled in the study. ROS levels were measured in the seminal ejaculates by chemiluminescence assay. Infertile men were subdivided into Low ROS (0–<93 RLU/s/106 sperm; n = 11), Medium ROS (>93–500 RLU/s/106 sperm; n = 17) and High ROS (>500 RLU/s/106 sperm; n = 14) groups and compared with fertile men (4–50 RLU/s/106 sperm). 4 subjects from fertile group and 4 each from the Low, Medium and High ROS were pooled. 1D gel electrophoresis followed by in-gel digestion and LC/MS–MS in a LTQ-Orbitrap Elite hybrid mass spectrometer system was used for proteome analysis. Identification of differentially expressed proteins (DEPs), their cellular localization and involvement in different pathways were examined utilizing bioinformatics tools.ResultsThe results indicate that proteins involved in biomolecule metabolism, protein folding and protein degradation are differentially modulated in all three infertile patient groups in comparison to fertile controls. Membrane metallo-endopeptidase (MME) was uniformly overexpressed (>2 fold) in all infertile groups. Pathway involving 35 focus proteins in post-translational modification of proteins, protein folding (heat shock proteins, molecular chaperones) and developmental disorder was overexpressed in the High ROS group compared with fertile control group. MME was one of the key proteins in the pathway. FAM3D was uniquely expressed in fertile group.ConclusionWe have for the first time demonstrated the presence of 35 DEPs of a single pathway that may lead to impairment of sperm function in men with Low, Medium or High ROS levels by altering protein turn over. MME and FAM3D along with ROS levels in the seminal plasma may serve as good markers for diagnosis of male infertility.Electronic supplementary materialThe online version of this article (doi:10.1186/s12014-015-9094-5) contains supplementary material, which is available to authorized users.

Highlights

Reactive oxygen species (ROS) plays a major role in the pathology of male infertility
A common end to numerous pathways that lead to defective sperm function is attributed to reactive oxygen species (ROS), a group of molecules with incompletely reduced oxygen atom [10, 11] that are capable of reacting with almost all biomolecules leading to their altered function such as inhibition/activation of enzymes
The first report on harmful effects of ROS on spermatozoa was published over 60 years ago [15] and a large body of literature has provided growing support for the concept that abnormal semen parameters and sperm damage are consequences of excessive levels of ROS resulting in impaired sperm function and subfertility [10, 16,17,18,19]

Summary

Introduction

Reactive oxygen species (ROS) plays a major role in the pathology of male infertility. Seminal plasma is a natural reservoir of antioxidants responsible for the nourishment, protection, capacitation, and motility of sperm within the female reproductive tract resulting in successful fertilization and implantation of the embryo. Physiological levels of ROS is necessarily maintained in all aerobic cells [12] as well as in the semen for optimal sperm function such as capacitation, motility and acrosome reaction [13, 14]. While both leukocyte and spermatozoa serve as principal sources of ROS generation in semen, the spermatozoa are more susceptible to ROS—induced damage at stake against ROS due to its rich content of polyunsaturated fatty acids and poor antioxidant capacity. Consensus is growing about the clinical utility of seminal oxidative stress testing in infertility clinics [20, 21]

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