Elsevier

Reproductive Toxicology

Volume 31, Issue 2, February 2011, Pages 239-246
Reproductive Toxicology

Effect of trans-resveratrol on induced oxidative stress in human sperm and in rat germinal cells

https://doi.org/10.1016/j.reprotox.2010.11.010Get rights and content

Abstract

Resveratrol is a phytoalexin with antioxidant properties. We evaluated resveratrol toxicity in swim-up selected human sperm and in rat spermatocytes and spermatids separated by the STAPUT method. Resveratrol antioxidant activity was tested against lipid peroxidation (LPO) induced by tert-butyl hydroperoxide. LPO was evaluated using the C11-BODIPY581/591 probe and transmission electron microscopy in samples incubated with and without resveratrol. LD50 for human sperm and rat spermatids was 50 μM; spermatocytes were more sensitive to resveratrol cytotoxicity. Sperm motility increased progressively at 30 μM, 15 μM and 6 μM. 15 μM resveratrol acts against LPO, preserving sperm chromatin and plasma membranes. LPO were more marked in spermatocytes than in spermatids and the effect of resveratrol was more evident in spermatocytes. In this study, the scavenger properties of resveratrol were demonstrated in vitro in human sperm and rat germ cells, thus resveratrol could be added to the media used in assisted reproduction techniques and cryopreservation when oxidative stress is exacerbated.

Introduction

Reactive oxygen species (ROS) are products of normal cellular metabolism and they play a functional role in many cell types as second messengers. When ROS are produced at a very low concentration, they trigger cell signaling events and regulate physiological function [1], [2].

Sperm were the first type of cell reported to produce free radicals and MacLeod [3] noted that the incubation of sperm under high oxygen tension leads to a rapid loss of sperm motility. Low level production of ROS by sperm supports some main functions, such as capacitation, acrosome reaction, zona pellucida binding and oocyte fusion [4]. Nevertheless, uncontrolled ROS production can play an important role in causing sperm aberration, leading to infertility. What is usually called “oxidative stress” appears to be the result of defects in the balance between the concentration of ROS and the antioxidant scavenging system. Sperm membranes are rich in polyunsaturated fatty acid, which makes them very susceptible to oxygen-induced damage mediated by lipid peroxidation (LPO) [5]. Oxidative stress has been suggested to be an important factor in the aetiology of poor sperm function through peroxidative damage to the cell membrane, to DNA (inducing single- and double-strand DNA breaks) and to proteins [6], [7], [8].

Seminal plasma and sperm are endowed with an array of protective antioxidants, such as the glutathione peroxidase/reductase system, superoxide dismutase, catalase and low-molecular weight antioxidants, vitamin E, vitamin C, urate, and albumin, which scavenge ROS in order to prevent possible cellular damage [9]. One of the rational strategies for counteracting oxidative stress is to increase the scavenging capacity of seminal plasma. However, the use of antioxidant supplementation to reverse the effect of ROS is still being debated [2], [10].

Resveratrol (RSV) is a natural phytoalexin with antioxidant properties that is widely consumed in the Mediterranean diet in the form of peanuts, grapes and wine. Interest in the compounds present in wine has increased since epidemiological studies indicated an inverse correlation between red wine consumption and the incidence of cardiovascular disease [11]. In addition, RSV has exhibited a broad range of biological activities, including antiinflammatory, antiviral and antitumoral properties [12].

Regarding the male reproductive system, some recent in vivo studies in animal models demonstrated that RSV administration enhances sperm production in rats by stimulating the hypothalamic–pituitary–gonadal axis without inducing adverse effects [13]. Moreover, RSV may decrease germ cell apoptosis in mice and rats [14], [15] and have a positive effect by triggering penile erection and by enhancing blood testosterone levels, testicular sperm count and epididymal sperm motility, as demonstrated in rabbits [16].

A protective effect of RSV, against oxidative damages but not against the loss of motility induced by the cryopreservation of human semen, has recently been observed [17].

This study was designed to evaluate, in vitro, the effect of RSV in ejaculated human sperm in order to assess the possible toxicity of such a compound. RSV was then used to investigate its antioxidant properties against LPO induced in human sperm by tert-butyl hydroperoxide (TBHP). The oxidative damage in samples incubated with and without RSV was evaluated using the C11-BODIPY581/591 probe and transmission electron microscopy (TEM). The study was also extended to two populations of spermatocytes and round spermatids from rat testis separated with the STAPUT method to assess the analysed properties of RSV, also in cells of seminiferous epithelium.

Section snippets

Semen analysis

Semen samples were obtained from men recruited at the Interdepartmental Centre for Research and Therapy of Male Infertility, University of Siena.

Semen samples were collected by masturbation after 3–5 days of sexual abstinence and examined after liquefaction for 30 min at 37 °C. Volume, pH, concentration and motility were evaluated according to World Health Organization guidelines [18]. Patients were informed of and gave written consent for the procedures related to the study.

Sperm selection: swim-up

Motile sperm fraction

Results

We tested the effect of the possible cytotoxicity of RSV and its scavenger properties in selected human sperm and rat germ cells fractioned as a meiotic population composed of pachytene spermatocytes and a post meiotic fraction of early spermatids.

Discussion

At present, a large body of evidence from in vitro and animal studies indicates that RSV may be beneficial to many aspects of human health. Interest in this molecule has increased since it shows biological attributes, mainly antioxidant and anti-inflammatory activity, chemoprevention, immunomodulation and cardioprotective properties, and it has positive effects on several aspects of metabolism, leading to increasing lifespan in metazoan models and also in rodents [19].

This compound displays an

Conflict of interest

The authors declare that there are no conflicts of interest.

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