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

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.

Testicular torsion and reperfusion: evidences for biochemical and molecular alterations.

This study was done in order to determine the molecular and biochemical alterations following testicular torsion (TT) and torsion-reperfusion (TR). For this purpose, 54 male Wistar rats were divided into five groups as control group (n = 6) and experimental group subjected to 1, 2, 4, and 8 h unilateral left torsion induction (n = 12 in each group). After induction of TT, testicular samples were collected from each group (n = 6), and the other six rats of each group underwent the same period of reperfusion after TT and then were sampled. Histological changes, the mRNA and protein expression of heat shock protein-70 (Hsp70), and caspase-3 were examined using reverse transcriptase-PCR (RT-PCR) and immunohistochemistry, respectively. Testicular total antioxidant capacity (TAC), glutathione peroxidase (GSH-px), and malondialdehyde (MDA) levels were evaluated. The mRNA damage and DNA fragmentation were assessed. The TT and TR significantly reduced differentiation and spermiogenesis indices (p < 0.05). The TT- and TR-induced groups exhibited a severe reduction in Hsp70 expression as well as remarkable enhancement in caspase-3 expression. The TAC and GSH-px levels were decreased and the MDA content was increased in TT- and TR-induced groups. Finally, the TT and TR enhanced mRNA damage and DNA fragmentation. The TT- and TR-induced damaging oxidative stress, diminished Hsp70 expression, and enhanced caspase-3 mRNA and protein levels result in apoptosis following 1, 2, and 4 h. Whereas, following 8 h, TT and TR initiate the necrosis by inducing energy depletion as well as severe mRNA damage.