Effect of sequential TRU-CUT needle biopsies on testicular gene expression

• Grant number: 21/11898-4
• Support Opportunities: Scholarships in Brazil - Scientific Initiation
• Start date: September 01, 2022
• End date: May 14, 2023
• Field of knowledge: Agronomical Sciences - Veterinary Medicine - Animal Reproduction
• Principal Investigator: João Carlos Pinheiro Ferreira
• Grantee: Paula Zanin Rattes
• Host Institution: Faculdade de Medicina Veterinária e Zootecnia (FMVZ). Universidade Estadual Paulista (UNESP). Campus de Botucatu. Botucatu , SP, Brazil

Abstract

The effects of heat stress (ET) on testicular gene expression have been attracting more and more interest, however, to obtain testicular samples, it is almost always necessary to castrate the males, thus making it impossible to collect samples and limiting the understanding of the phenomena being studied. Testicular biopsies using Punch, Split and Tru-cut needles have been used to obtain testicular tissue samples for the diagnostic purpose of infertility or subfertility. This procedure is also used in testicular physiological and pathophysiological studies. However, little is known about the consequences of serial harvesting, as they are potentially harmful to tissue integrity and trigger inflammation, events known to be deleterious to testicular physiology and spermatogenesis. In order to investigate the effects of sequential testicular biopsies with TRU-CUT needle on testicular gene expression, eight crossbred lambs (Dorper x Santa Inês), aged ~18 months will be housed in stalls, with free access to trough and water, on a diet sized for maintenance. After the adaptation period (15 days), sequential testicular biopsies will be taken using a 16 G TRU-CUT needle at 0 (time of the first collection), 6, 9, 24, and 45 hours, alternating the testicle at each collection. The samples will be stored in cryotubes and kept at -80ºC until further use. By RT-PCR the relative abundance of mRNA transcripts of some early genes (EgR1, Jun, Fos) and genes that have been shown to have their expression altered during inflammatory episodes (PLA2 and COX) or ET (StAR, HSF1, Hsp70, GPX1, BAX, BCL2, P53) will be investigated. The results will be evaluated by employing the Shapiro-Wilk and Leven test. For comparison of relative mRNA abundance of genes, a linear mixed model with repeated measures in time will be applied which time, biopsy site, as well as their interaction, will be established as fixed effects (PROC MIXED).(AU)