Soluble protein fraction of human seminal plasma

Human seminal plasma (SP) is a complex fluid where sperm cells are bathed. Until recently, SP was simply retained a spermatozoa transport medium with nourishing functions. Growing evidences are nowadays recognizing it as a main actor "on the stage" of reproduction: SP orchestrates the synchronized cascade of events that make spermatozoa able to fertilize and modulates male and female reproductive fitness in physiological and in pathological states. Despite all SP studies have focused on "whole" SP or on its extracellular vesicles, to obtain an in-depth functional comprehension of SP, also the merely vesicle-free SP (vf-SP) soluble fraction deserves consideration. Here we present the first vf-SP functional proteomic study applying 2-DE, MALDI-TOF MS, and cluster and pathway analyses. Our work evidenced the occurrence of few unique proteins in vf-SP and an unexpected high heterogeneity of their corresponding protein species, conceivably derived by massive co- and/or post-translational modification events. Generated nets and clusters revealed tight functional correlations among identified proteins as well as their involvement in key functions for spermatozoa support and fertilization. Moreover, since SP is released by urogenital tissues and glands, our work may pave the way to the evaluation of vf-SP expression-profiles in diagnosing pathological processes in the secreting tissues. Biological significance: Seminal plasma vesicles are emerging as rich reservoirs of biomarkers for male infertility and urogenital disorders. Our proteomic approach to human SP diverges from the general trend in SP characterization, investigating not sperm extracellular vesicles but rather the vesicle-depleted SP fraction. Here we discussed how also SP soluble fraction may be considered actively involved in spermatozoa maturation, and we suggested that vf-SP protein profile may offer a precious overview on spermatogenesis, epididymal maturation, ancillary gland functionality, and sperm quality, thus highlighting its potentiality as a biomarker source in man health and reproduction. The present work provided the first 2-DE reference map of the so far underestimated vf-SP and an innovative and comprehensive functional-overview of its proteins. Obtained data may concur to clarify biochemical and molecular processes that the in fieri and the ejaculated semen undergoes as well as how semen properties can affect fertility or may reflect genital disorders. This paper is dedicated to the memory of Dr Riccardo Focarelli, associated professor in Comparative Anatomy and Glycobiology at the Dpt. of Life Sciences, Siena University, to acknowledge his fundamental contribution to the design of this study and in recognition of his distinguished research legacy.