Induction of cell cycle arrest, DNA damage, and apoptosis by nimbolide in human renal cell carcinoma cells.
Nimbolide is a tetranortriterpenoid isolated from the leaves and flowers of Azadirachta indica which has been shown to exhibit anticancer, antioxidant, anti-inflammatory, and anti-invasive properties in a variety of cancer cells. However, the anti-tumor effect on human renal cell carcinoma (RCC) cells is unknown. In this study, we found that nimbolide treatment had a cytotoxic effect on 786-O and A-498 RCC cells in a dose-dependent manner. According to flow cytometric analysis, nimbolide treatment resulted in G2/M arrest in 786-O and A-498 cells accompanied with an increase in the phosphorylation status of p53, cdc2, cdc25c, and decreased expressions of cyclin A, cyclin B, cdc2, and cdc25c. Nimbolide also caused DNA damage in a dose-dependent manner as determined by comet assay and measurement of γ-H2AX. In addition, apoptotic cells were observed in an Annexin V-FITC/propidium iodide double-stained assay. The activities of caspase-3, -9, and poly ADP-ribose polymerase (PARP) were increased, and the expression of pro-caspase-8 was decreased in nimbolide-treated 786-O and A-498 cells. Western blot analysis revealed that the levels of intrinsic-related apoptotic proteins Bax and extrinsic-related proteins (DR5, CHOP) were significantly increased in nimbolide-treated 786-O and A-498 cells. In addition, the expressions of Bcl-2 and Mcl-1 were decreased in 786-O and A-498 cells after nimbolide treatment. We conclude that nimbolide can inhibit the growth of human RCC cells by inducing G2/M phase arrest by modulating cell cycle-related proteins and cell apoptosis by regulating intrinsic and extrinsic caspase signaling pathways. Nimbolide may be a promising therapeutic strategy for the treatment of RCC.
Department of Biochemistry, School of Medicine, Chung Shan Medical University, Taichung, Taiwan.|Clinical laboratory, Chung Shan Medical University Hospital, Taichung, Taiwan.
Yi-Hsien
YH
Hsieh
Division of Pediatric Surgery, Changhua Christian Hospital, Changhua, Taiwan.|Graduate Institute of Medical Sciences, Chang Jung Christian University, Tainan, Taiwan.
Chien-Hsing
CH
Lee
Institute of Biochemistry, Microbiology and Immunology, Chung Shan Medical University, Taichung, Taiwan.
Hsiao-Yun
HY
Chen
Institute of Medicine, Chung Shan Medical University, Taichung, Taiwan.
Shu-Ching
SC
Hsieh
Institute of Biochemistry, Microbiology and Immunology, Chung Shan Medical University, Taichung, Taiwan.
Chia-Liang
CL
Lin
School of Medicine, Tzu Chi University, Hualien, Taiwan. abi0918@outlook.com.|Department of Nephrology, Buddhist Dalin Tzu Chi General Hospital, No 2, Minsheng Road, Dalin Township, Chiayi, Taiwan. abi0918@outlook.com.
Jen-Pi
JP
Tsai
D017209:Apoptosis / Q000187:drug effects*; D051017:Apoptosis Regulatory Proteins / Q000378:metabolism; D002292:Carcinoma, Renal Cell / Q000188:drug therapy* / Q000378:metabolism; D059447:Cell Cycle Checkpoints / Q000187:drug effects*; D018797:Cell Cycle Proteins / Q000378:metabolism; D045744:Cell Line, Tumor; D004249:DNA Damage / Q000187:drug effects*; D059565:G2 Phase Cell Cycle Checkpoints / Q000187:drug effects; D006801:Humans; D007680:Kidney Neoplasms / Q000188:drug therapy* / Q000378:metabolism; D036701:Limonins / Q000494:pharmacology*; D015398:Signal Transduction / Q000187:drug effects
D051017:Apoptosis Regulatory Proteins; D018797:Cell Cycle Proteins; D036701:Limonins; C042198:nimbolide
Apoptosis; Cell cycle; DNA damage; Nimbolide; Renal cell carcinoma