The Department of Pediatric Urology, Ministry of Education, Key Laboratory of Child Development and Disorders, Chongqing International Science and Technology Cooperation Center for Child Development and Disorders, Children's Hospital of Chongqing Medical University, No. 136, Zhongshan 2 RD, Yuzhong District, Chongqing, 400014, China.
Mol Biol Rep. 2012 Jan;39(1):493-500. doi: 10.1007/s11033-011-0763-9. Epub 2011 May 8.
The aim of this study is to explore the mechanism by which acrolein (ACR), a metabolite of cyclophosphamide (CP), induces immature Sertoli cell cytoskeletal changes. Sertoli cells obtained from rats were cultivated and treated with 50 and 100 μM ACR. XTT assays were performed to detect cell viability. Activities of superoxide dismutase (SOD), glutathione peroxidases (GSH-Px), and catalase (CAT), as well as total anti-oxidation competence (T-AOC) were examined. Superoxide anion levels were detected by a fluorescent probe. Cell ultrastructure changes were observed by transmission fluorescent microscope. Actin filament (F-actin) distribution was detected by immunofluorescence, and ERK and p38MAPK expression were detected by western blot analysis. ACR significantly decreased the viability of Sertoli cells in a dose- and time-dependent manner. T-AOC and the antioxidant activity of SOD, CAT and GSH-Px, were decreased in ACR-treated groups compared with the control group. The levels of reactive oxygen species (ROS) in ACR-treated Sertoli cells were increased. In addition, characteristics of cell apoptosis such as mitochondrial swelling, aggregated chromatin, condensed cytoplasm, nuclei splitting, and nuclei vacuolization were observed in ACR-treated cells. Furthermore, ACR-treatment also induced microfilament aggregation, marginalization and regionalization. The expression levels of ERK and p38MAPK were also increased in ACR-treated cells in a dose- and time-dependent manner. ACR, a major CP metabolite, impairs the cytoskeleton which is likely caused by induction of the oxidative stress response through up-regulation of ERK and p38MAPK expression.
本研究旨在探讨丙烯醛（ACR），环磷酰胺（CP）的代谢产物，诱导未成熟支持细胞细胞骨架变化的机制。从大鼠中培养和处理 50 和 100μM ACR 的支持细胞。通过 XTT 测定法检测细胞活力。测定超氧化物歧化酶（SOD）、谷胱甘肽过氧化物酶（GSH-Px）和过氧化氢酶（CAT）的活性以及总抗氧化能力（T-AOC）。通过荧光探针检测超氧阴离子水平。通过透射荧光显微镜观察细胞超微结构变化。通过免疫荧光检测肌动蛋白丝（F-actin）的分布，并通过 Western blot 分析检测 ERK 和 p38MAPK 的表达。ACR 显著降低 Sertoli 细胞的活力呈剂量和时间依赖性。与对照组相比，ACR 处理组的 T-AOC 和 SOD、CAT 和 GSH-Px 的抗氧化活性降低。ACR 处理的 Sertoli 细胞中活性氧（ROS）的水平增加。此外，在 ACR 处理的细胞中观察到线粒体肿胀、染色质聚集、细胞质浓缩、核分裂和核空泡化等细胞凋亡特征。此外，ACR 处理还诱导微丝聚集、边缘化和区域化。ERK 和 p38MAPK 的表达水平也呈剂量和时间依赖性增加。ACR，CP 的主要代谢物，通过上调 ERK 和 p38MAPK 的表达来诱导氧化应激反应，从而损害细胞骨架。
