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Profile of germinative, epigenetic and pluripotency markers in male and female germ cells during embryonic and adult development in swine

Grant number: 20/10503-3
Support type:Scholarships in Brazil - Scientific Initiation
Effective date (Start): September 01, 2021
Effective date (End): August 31, 2022
Field of knowledge:Agronomical Sciences - Veterinary Medicine - Animal Reproduction
Principal researcher:Fabiana Fernandes Bressan
Grantee:Amanda Soares Jorge
Home Institution: Faculdade de Zootecnia e Engenharia de Alimentos (FZEA). Universidade de São Paulo (USP). Pirassununga , SP, Brazil
Associated research grant:15/26818-5 - Investigation of cellular and molecular mechanisms on in vitro induced toti- and pluripotency acquisition - a translational approach, AP.JP

Abstract

The primordial germ cells (PGCs), are the precursor cells of the male and female gametes, for this reason they are the only cells in the body capable of generating a new individual. These cells in most animals and even in pigs appear close to the epiblastic region and from this place they move towards the primitive intestine of the embryo, by means of morphogenic movements and extrinsic signals, following towards the posterior body wall and settling in the gonadal mesenchyme, where they induce the differentiation of adjacent cells that will form future gonads. In domestic and wild animals just as in pigs, few researches were found regarding the profile of PGCs in vivo and in vitro of the species. In these animals, recent studies show the expression of several markers during the initial period of development (9-15.5 days) and the analysis of the PGCs methylation levels during the 24th and 31st days of pregnancy, however little is known about the phenotypic profile and epigenetic changes of PGCs in later gestational periods and adulthood, whether in males or females. The porcine is considered a translational model in several areas of medicine, as it is a species with metabolism, physiology, and anatomy similar to that of humans, and is therefore widely used as a model for human diseases, in addition to having a short gestation period, which makes the species even more favorable to research with embryos and PGCs. Therefore, given the importance and complexity of primordial germ cells for the formation and reproduction of an individual, understanding the mechanisms and pathways of migration and differentiation of PGCs, as well as understanding the differences between males and females becomes essential. In such a way, this study aims to analyze the expression of epigenetic markers and the various proteins associated with the development and differentiation of PGCs in female and male pigs, from the beginning of embryonic development to adulthood, in order to understand the different gene expressions between the sexes. For this, samples will be collected in different gestational periods (from 22 to 40 days after fertilization), testicles, and ovaries of adult pigs. All samples will be analyzed for the expression of germline, pluripotent and epigenetic markers (POU5F1, DDX4, DAZL, STRA8, STELLA, PLZF, SOX2, 5hmC, 5mC, H3K27me3, and H3K9me2), through histology, immunofluorescence, and RT-qPQ. The results obtained in this study may help in understanding the reprogramming pathways suffered by pig PGCs during the development and reproductive maturation, as well as adding knowledge about the in vivo PGCs of males and females, they can be transposed to the generation of mature gametes in vitro and viable, which can be used in the biotechnology of the reproduction of domestic animals and as a model of studies in humans.

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