Melatonin Ameliorates Bisphenol-a Induced DNA Damage In The Germ Cells Of Adult Male Rats

Objective: Bisphenol A (BPA) is a well-known endocrine-disruptingchemical (EDC), and it is a monomer of polycarbonate plastic and aconstituent of epoxy and polystyrene resins. Due to have the function ofachromatic, hyaline and time-proof in the plastic production, it is widelyused as a baby’s bottle, food packing, plastic eating utensils and medicalequipments. Its widespread distribution in our daily life. Due to its chemicalsimilarity to diethylstilbestrol, which is a carcinogen in humans, the possiblegenotoxicity of BPA has already largely been evaluated. However, the resultsare still inconclusive and controversial. In essence, whether BPA exposurecould exert genotoxicity in male germ cells is still unclear. In our research,we choose the BPA as a study object and establish BPA sub-acute exposureanimal model aims to investigate the genotoxic effects of BPA in rat germcells and the potential protective action of melatonin against these effects.Besides, the underlying mechanism was also be explored.Method: The SPF adult male Sprague-Dawley (SD) rats aged8weekswere randomly divied into four groups, with10rats in each group. There are (1) solvent control group;(2) melatonin treated group, melatonin wasintraperitoneally injected (i.p.) at the dose of10mg/kg body weight/day;(3)BPA treated group, BPA was gavage at the dose of200mg/kg bodyweight/day,(4) melatonin+BPA combined treated group, receivedmelatonin at10mg/kg body weight along with200mg/kg body weight ofBPA per day, melatonin was injected i.p.30minutes before the BPA gavageadministration, all of the treatments continued for10consecutive days. At24h after the last treatment, the method of thiobarbituric acid was used tomeasure the levels of malondialdehyde (MDA), and the superoxidedismutase (SOD) activity was determined on the basis of the xanthineoxidase. The DNA damage to isolated spermatocytes was measured byalkaline comet assay and meiotic chromosomal spread immunostainingγH2AX to detect the γH2AX positive foci on the autosomes of pachytenespermatocytes. Flow cytometric analysis was employed to examine thenumber of the different DNA content subpopulations in the testicular cellpopulations. TUNEL fluorescence and HE staining were performed toevaluate the germ cells apoptosis and the testis histopathology.Results:①The dose of200mg/kg body weight/day BPA exposurehave not observed the effect of growth suppression in rats. The body weightgain， the weight of reproductive organs or sperm counts were notsignificantly change（P＞0.05）.②200mg/kg body weight/day BPA consecutively treatment10days caused oxidative damage, results in a significant increase in the MDA levelsand a notable decrease in the concentrations of SOD activity（P＜0.01）.③BPA exposure induced DNA damage in rat spermatocytes. Thecomet parameters TL, Tail DNA%, TM and OTM were significant increasewhen compared with the control animals (P＜0.01); compared with thecontrol animals, the number of γH2AX foci on the autosomes of pachytenespermatocytes was significant increase (P＜0.01).④BPA administration significantly decrease the proportion of4C-DNA content cells in the testicular cell population（sP＜0.05）. However,BPA failed to induce the apoptosis in germ cells and gross morphologicalchanges in the testes.⑤Compared with the BPA treatment group,10mg/kg bodyweight/day melatonin pretreatment alleviated the oxidative damage,effectively ameliorated the MDA accumulation and restored the SODactivity in the testis（P＜0.05）.⑥The group that was pretreated with melatonin showed a protectiveeffect of DNA damage induced by BPA. Significantly reduced the DNAdamage level （P＜0.01）and the number of γH2AX foci on the autosomesof pachytene spermatocytes（P＜0.05）. Moreover, resulted in a4C-DNAcontent cells increase, compared with the BPA alone treatment group, thedifference have statistical significance（P＜0.05）.Conclusion: BPA at the dose of200mg/kg body weight/day sub-acute consecutively10days treatment can lead to a significant increase in theDNA damage of spermatocytes and can alter the proportion of germ cells.However, the BPA-induced effects may not have been potent enough toevoke a more severe cellular response to irreparable DNA damage, such ascell cycle arrest, apoptosis in the germ cell and the gross morphologicalchanges in the testes. The BPA-induced DNA damage was associated withelevated MDA levels and SOD activity depletion. These alterations wereeffectively protected by10mg/kg body weight/day melatonin pretreatment.In view of these data, oxidative stress is probably one of the mechanisms ofBPA-induced DNA damage in in the germ cells of male rats. Moreover,melatonin may be a promising pharmacological candidate for preventing thepotential genotoxicity of BPA following occupational or environmentalexposure.