Altered expression profile of glycolytic enzymes during testicular ischemia reperfusion injury is associated with the p53/TIGAR pathway: effect of fructose 1,6-diphosphate.
Altered expression profile of glycolytic enzymes during testicular ischemia reperfusion injury is associated with the p53/TIGAR pathway: effect of fructose 1,6-diphosphate.
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作者：
Al-Maghrebi M;Renno WM
背景资料。睾丸缺血再灌注损伤(TIRI)被认为是睾丸扭转和复位的病理机制。如果不进行治疗，Tiri可能会导致睾丸功能障碍，损害精子发生，并可能导致不育。在本研究中，我们旨在评估睾丸糖酵解酶(GES)的活性和表达，以及它们在Tiri过程中的可能调节。还研究了糖酵解中间体1，6-二磷酸果糖(FDP)对TIRI的影响。方法：研究方法。雄性SD大鼠分为假手术组、单侧Tiri组和Tiri+FDP(2 mg/kg)组。阻断睾丸动脉1h后再灌流4h，建立TIRI模型。再灌流前30min腹腔注射FDP。组织学和生化分析用于评估对精子发生的损害、主要GE的活性以及能量和氧化应激标志物。实时荧光定量聚合酶链式反应(Real-Time-PCR)检测GES的相对mRNA表达。采用ELISA法和免疫组织化学法检测糖酵解和细胞凋亡调节因子(TIGAR)对P53和TP53蛋白表达的影响。结果。组织学分析显示Tiri诱导生精损伤，表现为Johnsen活检评分显着降低。此外，Tiri还降低了己糖激酶1、磷酸果糖激酶-1、3-磷酸甘油醛脱氢酶和乳酸脱氢酶C的活性，但只下调了己糖激酶1、磷酸甘油酸激酶2和乳酸脱氢酶C的mRNA表达。Tiri还诱导了ATP和NADPH的耗竭，并伴随着丙二醛浓度的增加、谷胱甘肽水平的降低、超氧化物歧化酶和过氧化氢酶活性的降低。TIRI后P53和TIGAR的免疫表达明显增加。上述Tiri诱导的改变可被FDP处理减弱。讨论。我们的发现表明，Tiri诱导的生精损伤与GE活性和基因表达的失调有关，而GE活性和基因表达的失调与Tigar/P53途径的激活有关。FDP治疗对TIRI的损伤有一定的缓解作用。这项研究进一步强调了代谢调节对正常精子发生的重要性。
Background. Testicular ischemia reperfusion injury (tIRI) is considered the mechanism underlying the pathology of testicular torsion and detorsion. Left untreated, tIRI can induce testis dysfunction, damage to spermatogenesis and possible infertility. In this study, we aimed to assess the activities and expression of glycolytic enzymes (GEs) in the testis and their possible modulation during tIRI. The effect of fructose 1,6-diphosphate (FDP), a glycolytic intermediate, on tIRI was also investigated. Methods. Male Sprague-Dawley rats were divided into three groups: sham, unilateral tIRI, and tIRI + FDP (2 mg/kg). tIRI was induced by occlusion of the testicular artery for 1 h followed by 4 h of reperfusion. FDP was injected peritoneally 30 min prior to reperfusion. Histological and biochemical analyses were used to assess damage to spermatogenesis, activities of major GEs, and energy and oxidative stress markers. The relative mRNA expression of GEs was evaluated by real-time PCR. ELISA and immunohistochemistry were used to evaluate the expression of p53 and TP53-induced glycolysis and apoptosis regulator (TIGAR). Results. Histological analysis revealed tIRI-induced spermatogenic damage as represented by a significant decrease in the Johnsen biopsy score. In addition, tIRI reduced the activities of hexokinase 1, phosphofructokinase-1, glyceraldehyde 3-phosphate dehydrogenase, and lactate dehydrogenase C. However, mRNA expression downregulation was detected only for hexokinase 1, phosphoglycerate kinase 2, and lactate dehydrogenase C. ATP and NADPH depletion was also induced by tIRI and was accompanied by an increased Malondialdehyde concentration, reduced glutathione level, and reduced superoxide dismutase and catalase enzyme activities. The immunoexpression of p53 and TIGAR was markedly increased after tIRI. The above tIRI-induced alterations were attenuated by FDP treatment. Discussion. Our findings indicate that tIRI-induced spermatogenic damage is associated with dysregulation of GE activity and gene expression, which were associated with activation of the TIGAR/p53 pathway. FDP treatment had a beneficial effect on alleviating the damaging effects of tIRI. This study further emphasizes the importance of metabolic regulation for proper spermatogenesis.