N OCIETY' - the Society for Reproductive Biology

THE EFFECTS OF WHOLE BODY HEAT-STRESS OF MALE MICEONASPECTSOFPATERNALFERTaITY· The Role of Mitochondriain Ischaemic/ Re-Perfusion Injury in Rat Testes·F. A. Ahmed l , J. Whelan l , A.M. Jequier2, and 1. M. Cummins 3lDepartment ofBioc~emistr;, ~~~ersity of ':'estemAus~alia,~edla~ds 6907 WA, 2perthAndrology, WesternAustralIa and DIVISIOn ofVetennary and BIOmedIcal SCIence, Murdoch UniversityIntroductionThe aim ofthis study was to gain further insight intothe mechanism underlying ischemic/re-perfusiondamage in the testis. It has been proposed that themitochondria is the target for reactive oxygen speciesmediated damage (1). A rat model for male infertilitywas developed, using torsion followed by variousperiods of recovery. The mitochondrial "commondeletion" was measured in these testes as an indicatorofgeneral mitochondrial damage. We also hypothesizedthat this type ofdamage leads to the induction ofcertainfactors that playa vital role in apoptosis and protectionof the suviving cells, hence we carried out an RNAsubtractive hybridization between the control and thetorsioned testes to identify differentially expressedgenes.Materials & MethodsThe study was comprised ofeight groups of3 rats each.Groups 1-3 were controls, groups 4-8 subjected tounilateral testicular torsion and allowed to recover foreither 0, 1 hour, I day, 1 wk or 4 weeks respectively.The mitochondrial common deletion was estimatedusing a competitive quantitative PCR assay that wasspecifically developed for this study (2). The effect oftorsion on the testes was detennined by the testicularbiopsy score count on representative tissues.Differentially expressed genes were isolated usingClontech PCR-select cDNA subtraction kit.ResultsBased on the biopsy scores it was clear that theexperimental model produced testicular damage thatwas more severe in the longer recovery groups (Tablel).However this did not correlated to the extent of the"common deletion " since no significant differenceswere seen between the control and torsioned testes (Fig1).Table 1. Testicular biopsy score counts in rat testestaking by counting 30 tubules in three rat testes. C isthe control testes and T is the torsioned testes. Scoresbelow 10 are considered subfertile. The values of theshort and long recovery groups were averaged.Groupcontrolnon-recovery-Cnon-recovery-Tshort recovery-Cshort recovery-Tlong recovery··Clong recovery-T60Average Score9.9 ± 0.189.9 ± 0.189.8 ± 0.379.8 ± 0.378.3 ± 1.99.7 ± 0.52.4 ± 1.2918 clones were found to be differentially expressed bytorsioned testes. Northern blots and sequecing are nowbeing undertaken to confirm which ones are truelydifferentially expressed and detennine their identity.0.05