Luteal function in cyclic and gestational canine diestrus: a cellular approach in genic and metabolic viewpoint

• Grant number: 13/15358-8
• Support Opportunities: Scholarships in Brazil - Post-Doctoral
• Start date: December 01, 2013
• End date: November 26, 2017
• Field of knowledge: Agronomical Sciences - Veterinary Medicine - Animal Reproduction
• Principal Investigator: Maria Denise Lopes
• Grantee: Ana Augusta Pagnano Derussi
• Host Institution: Faculdade de Medicina Veterinária e Zootecnia (FMVZ). Universidade Estadual Paulista (UNESP). Campus de Botucatu. Botucatu , SP, Brazil
• Associated scholarship(s): 15/23450-7 - Evaluation the mechanism of luteolysis in cyclic and gestational canine corpora lutea, BE.EP.PD

Abstract

The similarity between on cyclic and pregnancy canine luteal phase is one of the most intriguing aspect in the reproductive physiology of this species and despite the many studies and progress in relation this issue, these are still very inconclusive. In order to investigate the variations between cyclic and gestational corpus luteum (CL) gene profile, we evaluate the principal mitogenic, angiogenic, immunologic and apoptotic factors and hormone receptors existent in this phase, and analyze metabolic pathways and hormonal production of these luteal cells in culture. We will perform the monitoring of the estrous cycle and ovariohysterectomy in canine female on cycling (n = 16) and gestational diestrus (n = 16). The corpus luteum (CLs) will be collected between 7-11, 21-25, 40-44 and 61-64 days after the LH pre-ovulatory surge in all females. For gene profile analysis, the CLs of right ovary will subjected to RNAseq technical and thereafter will be selected among the 3 principal genes between factors and hormone receptors differentially expressed to be validated by qRT-PCR and immunohistochemistry. . The CLs of the left ovary will be subjected to cell culture technique and the medium will be collected every 06, 12 and 24 hours for metabolic analysis production (ATP, ADP, glucose, lactate) and hormonal (estrogen and progesterone) with a commercial kit . The analysis of RNA-seq will be conducted through the Galaxy platform and CASAVA v.1.8.2, TopHat v.2.0.1, Bowtie v. 0.12.8 programs for mapping and differential expression analysis. Gene expression Validation will be evaluated normality and variances, can be used ANOVA or Kruskal Wallis. Data will be analyzed using GraphPad Prism 5. Cellular metabolism and hormonal profile will be performed by ANOVA. Will be considered the significance level of p <0.05.