Cross-link between mitochondrial-dependent apoptosis and cell cycle checkpoint proteins after experimental torsion and detorsion in rats

Abstract

The current study assessed the cross-link between mitochondria-related apoptosis and cell cycle machinery systems during ischemia and reperfusion in a rat model of testicular torsion and detorsion. The Wistar male rats were divided into control, 1 h, 2 h, 4 h and 8 h testicular torsion-induced, and 1 h, 2 h, 4 h and 8 h testicular detorsion-induced groups. The Johnson's score was analyzed. The mRNA and protein contents of Bcl-2, Bax, Caspase-3, Cyclin D1, Cdk4, P21 and P53 were investigated by sqRT-PCR and immunohistochemical staining, respectively. The apoptosis index was analyzed by TUNEL staining. The mRNA levels of bax, p53, p21 and cyclin D1 were increased, and the mRNA levels of bcl-2 and cdk4 were decreased in torsion and reperfusion-induced groups, time-dependently. The caspase-3 mRNA was increased in torsion-induced and diminished in detorsion-induced groups. A time-dependent reduction in Bcl-2⁺, Caspase-3⁺, Cyclin D1⁺, Cdk4⁺ and P53⁺ and increment in P21⁺ cells distribution per mm² of tissue were revealed after torsion and detorsion. The apoptosis index was increased after torsion and decreased after detorsion. In conclusion, torsion-induced severe DNA damage stimulates the cyclin D1, p53 and p21 mRNA expression while more than 8 h is needed to reveal them as protein content in testicular tissue. About detorsion, decreased Cyclin D1 and Cdk4 proteins and the P53-induced transcriptional effect on p21 expression, stimulates the p21 bind to cdk4 and consequent failure in Cyclin D1/Cdk4 complex formation. This situation in association with apoptotic genes results in spermatogenesis failure.

Keywords: Apoptosis; Cell cycle; Detorsion; Spermatogenesis; Torsion.