The thesis is dedicated to the study of the influence of exogenous factors on the processes of spermatogenesis and embryogenesis of animals in vitro and in vivo and the improvement of biotechnological methods for their regulation and correction. In our investigation the most attention we have paid to the peculiarities of mammalian gametogenesis during the action of exogenous factors. It’s allows to find out the causes of the emergence and development of reproductive function disorders and to develop preventive measures for their prevention.
We also set a goal to test in model systems in vitro and in vivo the mammalian gametogenesis to study at all levels the reproductive system from cellular response mechanisms, gametes development to implantation capacity, enhanced embryo-uterine interactions and early embryogenesis. It was established that reduction of reproductive ability of females, early embryonic mortality are connected with the nature and degree of violations of male and female reproductive function of mammals by environmental factors, violation of endocrine and metabolic profiles, imbalance of microelement status of an organism.
A methodology of cultivating of uterine cells on surfaces modified by various grafted polymeric biocompatible nanolayers, while creating a 3D cell culture system for cultivation of oocytes and embryos in vitro, was developed. It was found that the maturation of rabbit oocyte-cumulus complexes onto 3D cell culture system with endometrial cells created on biogel was higher as compared to other experimental groups. It was established that after 24 hours of cultivation on biogel, 76,7 % oocytes of the experimental group recovered meiosis and reached the metaphase stage II, while the number of oocytes with the polar body in the control group was 63.3%. The culturing of 2-cell mouse embryos onto volumetric culture system with endometrial cells created on the biogel promoted the development of pre-implantation embryos in vitro, stimulated the blastomerization and blastocyst formation and reduced the number of embryos with delayed development.
It has been proved that co-culturing of cryopreserved preimplantation mice embryos on the monolayer of oviduct rabbit cells increases the number of embryos reaching the blastocyst stage by 6.38-6.55%.
While studying the influence of stress factors on the spermatogenesis of mice, a difference in response to the action of hyperthermia in somatic and germ cells was revealed. It has been shown that the effects of heat shock on mice testicles alter the activity of the mDazl gene (azoospermia factor) and increase the expression level of methyltransferase Suv39h2, Hdac4 histone deacetylase and Mdm1 gene.
The pattern of the Cideb gene expression in mice tissues under the influence of local hyperthermia was analyzed. High expression levels of Cideb genes were observed in liver, intestine and kidneys, with low levels thereof found in the stomach and testis tissues of mice after a heat shock. However, a difference in the Cideb gene expression in the testes and spermatocytes depending on the degree of hyperthermia was detected. The expression of the gene does not change at 38°C, whereas in the case of a moderate hyperthermia of 42°C, the Cideb gene expression was observed in testes at 2, 6 and 24 hours after the heat stress. An alternative splice variant of the Cideb gene in the testes at the hyperthermia of 42°C was found.
Our studies indicated that the administration of the "Lipoorganic" liposomal preparation had a beneficial effect on the correction of metabolic processes, intensification of reproductive function, stimulation of embryo-uterine interactions and reduction of pre- and post-implantation losses during pregnancy. The liposomal preparation administered to female rabbits during insemination resulted in an increased number of corpora lutea of pregnancy and an elevated number of implantations, as well as allowed to reduce by half the number of fetus resorptions and to diminish post-implantation losses by 2.3 times, while the use of the preparation 14 days prior to insemination reduced the percentage of pre-implantation loss of fetuses, as compared to similar indices of the control group. A corrective effect of the preparation on the activity of some serum enzymes, the content of structural and reserve lipids, metabolism intensity, as well as on the maintenance of antioxidant-prooxidant balance in reproductive organs improving fertilization and rabbits embryo implantation, was found in female rabbits in the early period of pregnancy.
