Zhang Qing-qing, Wang Wen-juan, Li Jun, Yang Neng, Chen Gang, Wang Zhong, Liang Zhong-qin
Department of Pharmacology, College of Pharmaceutical Sciences, Soochow University, Suzhou 215000, China.
Department of Neurosurgery, The First Affiliated Hospital of Soochow University, Suzhou 215000, China.
Acta Pharmacol Sin. 2015 Sep;36(9):1113-25. doi: 10.1038/aps.2015.36. Epub 2015 Jun 22.
Cathepsin L is a lysosomal cysteine protease that plays important roles in cancer tumorigenesis, proliferation and chemotherapy resistance. The aim of this study was to determine how cathepsin L regulated the radiosensitivity of human glioma cells in vitro.
Human glioma U251 cells (harboring the mutant type p53 gene) and U87 cells (harboring the wide type p53 gene) were irradiated with X-rays. The expression of cathepsin L was analyzed using Western blot and immunofluorescence assays. Cell survival and DNA damage were evaluated using clonogenic and comet assays, respectively. Flow cytometry was used to detect the cell cycle distribution. Apoptotic cells were observed using Hoechst 33258 staining and fluorescence microscopy.
Irradiation significantly increased the cytoplasmic and nuclear levels of cathepsin L in U251 cells but not in U87 cells. Treatment with the specific cathepsin L inhibitor Z-FY-CHO (10 μmol/L) or transfection with cathepsin L shRNA significantly increased the radiosensitivity of U251 cells. Both suppression and knockdown of cathepsin L in U251 cells increased irradiation-induced DNA damage and G2/M phase cell cycle arrest. Both suppression and knockdown of cathepsin L in U251 cells also increased irradiation-induced apoptosis, as shown by the increased levels of Bax and decreased levels of Bcl-2.
Cathepsin L is involved in modulation of radiosensitivity in human glioma U251 cells (harboring the mutant type p53 gene) in vitro.
组织蛋白酶L是一种溶酶体半胱氨酸蛋白酶，在癌症肿瘤发生、增殖和化疗耐药中发挥重要作用。本研究旨在确定组织蛋白酶L如何在体外调节人胶质瘤细胞的放射敏感性。
用人胶质瘤U251细胞（携带p53基因突变型）和U87细胞（携带p53基因野生型）进行X射线照射。采用蛋白质免疫印迹法和免疫荧光分析法分析组织蛋白酶L的表达。分别采用克隆形成试验和彗星试验评估细胞存活和DNA损伤情况。采用流式细胞术检测细胞周期分布。使用Hoechst 33258染色和荧光显微镜观察凋亡细胞。
照射显著增加了U251细胞而非U87细胞中组织蛋白酶L的细胞质和细胞核水平。用特异性组织蛋白酶L抑制剂Z-FY-CHO（10 μmol/L）处理或转染组织蛋白酶L短发夹RNA可显著提高U251细胞的放射敏感性。U251细胞中组织蛋白酶L的抑制和敲低均增加了辐射诱导的DNA损伤和G2/M期细胞周期阻滞。U251细胞中组织蛋白酶L的抑制和敲低也增加了辐射诱导的细胞凋亡，表现为Bax水平升高和Bcl-2水平降低。
组织蛋白酶L参与体外调节人胶质瘤U251细胞（携带p53基因突变型）的放射敏感性。
