In-depth identification of pathways related to cisplatin-induced hepatotoxicity through an integrative method based on an informatics-assisted label-free protein quantitation and microarray gene expression approach.
Cisplatin is used widely for treatment of a variety of cancer diseases. Recently, however, the use of cisplatin is restricted because of its adverse effects such as hepatotoxicity. There is no study with current proteomics technology to evaluate cisplatin-induced hepatotoxicity, even if some studies have reported on the hepatotoxicity. In this study, proteomic as well as genomic analyses have been used for identification of proteins and genes that respond to cisplatin treatment in rat primary hepatocytes. To investigate the hepatotoxic effects of cisplatin, rat primary hepatocytes were treated with an IC(20) concentration for 24 h. From proteomic analysis based on label-free quantitation strategy, cisplatin induced 76 up-regulated and 19 down-regulated proteins among 325 distinct proteins. In the mRNA level, genomic analysis revealed 72 up-regulated and 385 down-regulated genes in the cisplatin-treated group. Based on these two analyses, 19 pathways were commonly altered, whereas seven pathways were identified only by proteomic analysis, and 19 pathways were identified only by genomic analysis. Overall, this study explained the mechanism of cisplatin-induced hepatotoxicity with two points of view: well known pathways including drug metabolism, fatty acid metabolism, and glycolysis/TCA cycle and little known pathways including urea cycle and inflammation metabolism, for hepatotoxicity of other toxic agents. Up-regulated proteins detected by proteomic analysis in the cisplatin-treated group: FBP1 (fructose 1,6-bisphosphatase 1), FASN (fatty acid synthase), CAT (catalase), PRDX1 (peroxiredoxin-1), HSPD1 (60-kDa heat shock protein), MDH2 (malate dehydrogenase 2), and ARG1 (arginase 1), and also down-regulated proteins in the cisplatin-treated group: TPM1 (tropomyosin 1), TPM3 (tropomyosin 3), and CTSB (cathepsin B), were confirmed by Western blot analysis. In addition, up-regulated mRNAs detected by microarray analysis in the cisplatin-treated group: GSTA2, GSTT2, YC2, TXNRD1, CYP2E1, CYP2C13, CYP2D1, ALDH17, ARG1, ARG2, and IL-6, and also down-regulated mRNAs: CYP2C12, CYP26B1, TPM1, and TPM3, were confirmed by RT-PCR analysis. In case of PRDX1, FASN, and ARG1, they were further confirmed by immunofluorescence analysis. Through the integrated proteomic and genomic approaches, the present study provides the first pathway map related to cisplatin-induced hepatotoxicity, which may provide new insight into the mechanism of hepatotoxicity.
Department of Molecular Medicine, Cell and Matrix Biology Research Institute, School of Medicine, Kyungpook National University, Daegu 700-422, Republic of Korea.
Young-Eun
YE
Cho
Department of Pharmacology, School of Medicine, Kyungpook National University, Daegu 700-422, Republic of Korea.
Thoudam S K
TSK
Singh
D & P Biotech, School of Medicine, Kyungpook National University, Daegu 700-422, Republic of Korea.
Hyun-Chul
HC
Lee
Department of Molecular Medicine, Cell and Matrix Biology Research Institute, School of Medicine, Kyungpook National University, Daegu 700-422, Republic of Korea.
Pyong-Gon
PG
Moon
Department of Molecular Medicine, Cell and Matrix Biology Research Institute, School of Medicine, Kyungpook National University, Daegu 700-422, Republic of Korea.
Jeong-Eun
JE
Lee
D & P Biotech, School of Medicine, Kyungpook National University, Daegu 700-422, Republic of Korea.
Myung-Hoon
MH
Lee
College of Pharmacy and Research Institute of Pharmaceutical Sciences, Seoul National University, Seoul 151-742, Republic of Korea.
Eung-Chil
EC
Choi
Institute of Chemistry, Academia Sinica, Taipei 11529, Taiwan.
Yu-Ju
YJ
Chen
Department of Pharmacology, School of Medicine, Kyungpook National University, Daegu 700-422, Republic of Korea.
Sang-Hyun
SH
Kim
Department of Molecular Medicine, Cell and Matrix Biology Research Institute, School of Medicine, Kyungpook National University, Daegu 700-422, Republic of Korea. Electronic address: mcbaek@knu.ac.kr.
Moon-Chang
MC
Baek
D000818:Animals; D000970:Antineoplastic Agents / Q000633:toxicity*; D002478:Cells, Cultured; D056486:Chemical and Drug Induced Liver Injury / Q000378:metabolism*; D002945:Cisplatin / Q000633:toxicity*; D020869:Gene Expression Profiling; D022781:Hepatocytes / Q000187:drug effects* / Q000378:metabolism; D008099:Liver / Q000187:drug effects / Q000378:metabolism; D020411:Oligonucleotide Array Sequence Analysis; D040901:Proteomics; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats
D000970:Antineoplastic Agents; D012333:RNA, Messenger; D002945:Cisplatin