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Effect of sex sorted sperm on transcriptome and proteome of oviduct cells

Grant number: 13/12089-6
Support type:Scholarships in Brazil - Post-Doctorate
Effective date (Start): October 01, 2013
Effective date (End): January 03, 2016
Field of knowledge:Agronomical Sciences - Veterinary Medicine - Animal Reproduction
Principal researcher:Roberto Sartori Filho
Grantee:José de Oliveira Carvalho Neto
Home Institution: Escola Superior de Agricultura Luiz de Queiroz (ESALQ). Universidade de São Paulo (USP). Piracicaba , SP, Brazil

Abstract

Although in vivo studies in mouse have identified that the binding of sperm on oviduct cells change the transcription and protein secreted by oviduct cells, there are no in vivo studies in bovine showing this effect of sperm on oviduct cells. Moreover, due to the difference between X and Y sperm or sexed and non sexed sperm in sperm oviduct binding, is possible that the transcription and protein secretion be modified, compromising the formation of oviduct environment when sexed sperm are used on artificial insemination (AI). Therefore, the aim of this study are assess the effect of sexed or non sexed sperm on transcription of the oviduct cells and proteome of the oviduct fluid, and optimize a in vitro model for assessed this modifications in oviduct cells. For this, ejaculate of Nelore bulls (n=4) will be collected and separated into three fractions: non-sexed (NS), sexed for X-sperm (X), and sexed for Y-sperm (Y). A fourth group will be formed by pooling X and Y samples (XY). This semen will be used to AI 75 heifers, separated in 3 replicates with 5 heifers per group in each replicate. Eighteen hours after AI, each heifer will be felled to recovery the oviducts. After dissection, each oviduct will be flushed to recovery the epithelial oviduct cells and oviduct fluid. The cells will be used to assess the transcriptome of the oviduct cells using RNA-Seq. The fluid will be used to assess the proteic profile of the oviduct fluid using 2D nanoUPLC-HDMSE. From dates obtained in transcriptome, will be selected sixteen genes differentially expressed between the control group and the inseminated with non sexed sperm. This genes will be used to optimize the in vitro model for studies about the changes in oviduct cells due to sperm interaction. Oviduct collected in slaughterhouse will be inseminated in vitro with different concentrations of sperm, and will be incubated for different times. The oviduct cells will be recovered and the gene expression will be assessed by real time PCR. This study can provide information about the physiology characteristics necessary in vivo fecundation, and information related to differences in oviduct environment when sexed sperm are used.

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