Lee Sang Hun, Heo Jung Sun, Lee Min Young, Han Ho Jae
Department of Veterinary Physiology, BK21 Biotherapy Human Resources Center, College of Veterinary Medicine, Chonnam National University, Gwangju 500-757, Korea.
J Cell Physiol. 2008 Jul;216(1):269-75. doi: 10.1002/jcp.21402.
Steroid hormones have been reported to activate various signal transducers that trigger a variety of cellular responses. Among these hormones, testosterone has been identified as an antioxidant that protects against cellular damage. Therefore, using mouse embryonic stem (ES) cells as a model system, this study evaluated the effects of dihydrotestosterone (DHT), a biologically active testosterone metabolite, on H2O2-induced apoptosis. H2O2 increased the release of lactate dehydrogenase (LDH) and DNA fragmentation but reduced the cell viability in a time-dependent manner (> or =8 h). Moreover, H2O2 decreased the level of DNA synthesis and the levels of the cell cycle regulatory proteins [cyclin D1, cyclin E, cyclin-dependent kinase (CDK) 2, and CDK 4]. These effects of H2O2 were inhibited by a pretreatment with DHT. However, a treatment with flutamide (androgen receptor inhibitor, 10(-3) M) abolished the protective effects of DHT. This result was supported by the presence of the androgen receptor in mouse ES cells. The activity of the antioxidant enzyme, catalase, was increased by the DHT treatment but not by a co-treatment with DHT and flutamide. Using CM-H(2)DCFDA (DCF-DA) for the detection of intracellular H2O2, DHT decreased the intracellular H2O2 levels but flutamide blocked this effect. H2O2 also increased the level of p38 MAPK, JNK/SAPK, and NF-kappaB phosphorylation, which were inhibited by the DHT pretreatment. Catalase inhibited the effect of H2O2 on MAPKs and NF-kappaB. However, the flutamide treatment abolished the inhibitory effects of DHT on the H2O2-induced increase in the levels of p38 MAPK, JNK/SAPK, and NF-kappaB phosphorylation. DHT inhibited the H2O2-induced increase in caspase-3 expression and decreased the level of Bcl-2 and the cellular inhibitor of apoptosis protein (cIAP)-2. These effects were abolished by the flutamide treatment. In conclusion, DHT prevents the H2O2-induced apoptotic cell death of mouse ES cells through the activation of catalase and the downregulation of p38 MAPK, JNK/SAPK, and NF-kappaB via the androgen receptor.
据报道，类固醇激素可激活各种信号转导分子，引发多种细胞反应。在这些激素中，睾酮已被确认为一种可防止细胞损伤的抗氧化剂。因此，本研究以小鼠胚胎干细胞（ES细胞）作为模型系统，评估了具有生物活性的睾酮代谢产物双氢睾酮（DHT）对H2O2诱导的细胞凋亡的影响。H2O2以时间依赖性方式（≥8小时）增加乳酸脱氢酶（LDH）的释放和DNA片段化，但降低细胞活力。此外，H2O2降低DNA合成水平以及细胞周期调节蛋白[细胞周期蛋白D1、细胞周期蛋白E、细胞周期蛋白依赖性激酶（CDK）2和CDK 4]的水平。DHT预处理可抑制H2O2的这些作用。然而，氟他胺（雄激素受体抑制剂，10⁻³ M）处理可消除DHT的保护作用。小鼠ES细胞中存在雄激素受体支持了这一结果。DHT处理可增加抗氧化酶过氧化氢酶的活性，但DHT与氟他胺共同处理则无此作用。使用CM-H₂DCFDA（DCF-DA）检测细胞内H2O2，DHT可降低细胞内H2O2水平，但氟他胺可阻断此效应。H2O2还可增加p38丝裂原活化蛋白激酶（MAPK）、JNK/SAPK和核因子κB（NF-κB）的磷酸化水平，而DHT预处理可抑制这些作用。过氧化氢酶可抑制H2O2对MAPKs和NF-κB的作用。然而，氟他胺处理可消除DHT对H2O2诱导的p38 MAPK、JNK/SAPK和NF-κB磷酸化水平升高的抑制作用。DHT可抑制H2O2诱导的半胱天冬酶-3表达增加，并降低Bcl-2水平和细胞凋亡抑制蛋白（cIAP）-2水平。氟他胺处理可消除这些作用。总之，DHT通过激活过氧化氢酶并经由雄激素受体下调p38 MAPK、JNK/SAPK和NF-κB，从而防止H2O2诱导的小鼠ES细胞凋亡性细胞死亡。
