EFFECT OF LONG-TERM EXPOSURE TO LOW DOSE OF LEAD ACETATE ON FERTILITY POTENTIAL OF SPERMS IN IVF AND EMBRYONIC DEVELOPMENT OF MICE

Male reproductive function in general population had attracted increasing attention, due to reports suggesting that the occurrence of several biological problems affecting male genital tract, have increased during the last 50 years, including apparent decline in semen quality in different regions of the world. Such events have been linked to wide spread use of different types of chemicals. Lead, the heavy metal, is one of the major environmental pollutants, and is considered as one of the most significant reproductive toxicants.
This study was conducted to detect the adverse effects of exposure to lower doses of lead than the allowable range, on the male reproductive organs and semen quality, including DNA integrity and it's correlation with the early embryonic development , pregnancy outcome and it's later effects on the sex ratio of their offspring . Adult male mice were used as a representative example for human. All experiments were performed on 180 males and 120 females of Swiss Webester mature mice, with average body weights of 20-24gm, and an age range of 6-8weeks, obtained from the colony of the animal house of the High Institute of Infertility and Assisted Reproductive Technology/ Alnahrain University. Animals were kept in a temperature controlled room of (18-24) C°, and housed in plastic cages, supported on ventilated racks. Mice were fed with standard, balanced pellets which was presented together with the distilled water ad libitum.The study included three separate experiments. For each one, the males were divided into three equal groups (20mice/group): G1, G2, and G3. Both G1and G2were treated with either 50ug or 100ug/kg.bw/day, respectively, of lead acetate dissolved in their drinking water, for 16 weeks, while G3 was considered as a control group, and received distilled water (free of lead) for the same period. Female mice were used for mating and fertilization their oocytes by the treated and control males, which also were divided into three equal groups (40female/ group). Each group was used for mating with the parallel experimental group. Body weights for all males were recorded at the beginning of the study and monthly till the end of the experiments.
After 16 weeks, blood lead level (BLL) in all males were determined, using atomic absorption spectrophotometer. In experiment 1, the adverse effects of lead on sperm parameters were detected, including: concentration, motility, percent of grade of activity, morphology, vitality and agglutination. In experiment 2, the effect of lead on sperm DNA fragmentation ,was determined, using the modified sperm chromatin dispersion technique(SCD).Testes and epididymis of these animals were fixed and histological sections of them were prepared to study the structural changes appeared after lead treatment. The other 60 male mice were used for experiment 3, in which males of all groups were mated with the healthy, intact females. Implantation rate (for half number of the pregnant females) were detected after 5 days of successful mating, using Chicago blue stain to localize the implantation sites in the uterus, while the other half was left to complete the gestation period. At birth the number, body weights and gender type of the offspring were recorded. Results showed no significant differences were noticed in body weights of both treated groups of male mice as compared to control group, at the end of treatment period. BLL detected in G1 was 23.5ug/dl and in G2 it was 28ug/dl, both doses caused significant reduction in sperm concentration, motility, and morphology, with highly significant increase in dead sperms and agglutination, with the higher level of BL. Sperm chromatin integrity showed a highly significant increase in sperm DNA fragmentation percent in G2only and result was reflected in the testicular and epididymal histological sections. Highly significant reduction in implantation rate in females mated with males of G2, while those from G1, didn't show significant difference from that of the control group. No significant changes appeared in body weights of male offspring, but females of G1, showed a significant increase in comparison with control group, while number of males offspring were highly significantly reduced in G2, leading to a clear reduction in the sex ratio of males to females, which also revealed by offspring belong to G1. It is concluded that the very low BLL (23.5ug/dl ), severely affect the sperm parameters, while sperm DNA integrity had shown highly significant reduction, together with adverse structural changes in the testes and epididymis, with the higher BLL (28ug/dl) only, and similar results was found with the implantation rate, birth rate and sex ratio in offspring belong to females being mated with males having the higher level of blood lead.