Annona Diterpenoid Compounds That Inhibit Liver Cancer Proliferation And Its Mechanism

BackgroundHepatocellular cancer(HCC) is a fatal malignant tumor doing great harm to human's health. There is a high occurrence rate of HCC in China. The mortality is the 3rd highest of digestive malignant tumor following to gastric and esophageal cancer. Since there is no effective treatment to HCC at middle and late stage till now, TCM treatment with its particular effect is still an important part of integrate therapy of HCC. It is a hot research topic to find effective anticancer component and extract from natural products. The researches on the anticancer function of Annona glabra Linn had obtained good performance recently. The research group led by Pr.Zhang Yong-hong had fulfilled a series of experiments on the anticancer function of Annona glabra Linn.The results shows, the extract of Annona glabra Linn can inhibit the growth of the implanted sarcoma of the mice with the highest inhibitory rate of 41% at the dosage of 50mg/Kg×7d. It is also shown that Annona glabra Linn can inhibit the proliferation of human hepatocellular cancer cell lines SMMC-7721 and induce apoptosis by increasing the expression of bax gene and decreasing the expression of bcl-2 gene.ObjectiveTo select the most active anticancer monomer from 5 diterpenoid compounds and evaluate its anticancer action through experiments in vivo and vitro. To establish a fundamental base for putting Annona glabra Linn into a new anticancer drug by disclosing its anticancer mechanism in molecular biologic method.MethodsIn vitro1 Selection of monomerObserve the inhibitory action of 5 diterpenoid compounds on human hepatocellular cancer cell line HepG2 and SMMC-7721 by MTT assay and find out the correlation of dosage-effect and time-effect.2 Anticancer mechanism①Inverted phase-contrast microscope was used to observe the morphological changes of cells.②Fluorescent microscope was used to observe the change of chromatin.③Transmission electron microscope was used to observe the change of the ultramicro-structure of the cells, e.g., budding, apoptotic bodies, plasma membrane blebbing, nuclear condensation, collapse, etc.④Agarose gel electrophoresis was used to observe the DNA ladder and analyze the biochemical change of the cells.⑤FCM(flow cytometer) was used to observe the apoptotic peak and analyze the cellular cycle and calculate the apoptotic rate.⑥LCSM(laser confocal scanning microscopy) was used to observe the affection of Annona glabra Linn on the MP(cellular membrane potention), ΔΨ_m (mitochondria...