The effect of of clerodane diterpenoid 16-hydroxycleroda-3,13-dien-15,16-olide on colorectal cancer cell lines
Polyalthia longifolia is a lofty evergreen tree found in India and Sri Lanka. Today, P. longifolia var. pendula is in large-scale cultivation in south Taiwan as a landscape plant. A new compound, clerodane diterpenoid 16-hydroxycleroda-3,13-dien-15, 16 -olide¡]CD¡^, was isolated from the bark of Polyalthia longifolia var. pendula. The compound was shown with cell growth inhibitory property at first screen in KMU¡]Kaohsiung Medical University¡^. In our study, antiproliferative activity of CD on human colorectal adenocarcinoma cells¡]HT29 and HCT116¡^were tested and the inhibitory concertration of 50¢Mcell viability¡]IC50¡^is 5 £gg/ml¡]HCT116¡^ and 10 £gg/ml¡]HT29¡^determined by MTT(3-[4, 5-dimethylthiazol-2-yl]-2, 5-diphenyltetrazolium bromide) assay. The caspase 3 and PARP cleavage experiment results indicated that CD induced apoptosis is dose dependent manner. We found that CD induced sub-G1 accumulation and reactive oxygen species(ROS) release by flow cytometry analysis. Pretreatment of N-Acetyl-L-cystine(NAC), an antioxidant agent, can reverse the anti-proliferation effect by CD. We found that CD induced loss of mitochondria membrane potentiol¡]£G£Zm¡^. We also analysis differentially expressed proteins by 2-D electrophoresis and find drug response proteins, e.g. HSP10, Profilin-1, Peroxindoxin-1. RT-PCR and western blot was performed to confirm the protein expression changes. It is interesting to reveal the role of these proteins in the colorectal cancinogenesis and anti-tumor drug response.
Advisor:Hurng-Wern Huang; Yow-Ling Shiue; Chao-Neng Tseng
School:National Sun Yat-Sen University
School Location:China - Taiwan
Source Type:Master's Thesis
Keywords:two dimensional electrophoresis colorectal cancer apoptosis
Date of Publication:07/15/2008