Experimental Study On Apoptosis Of Human Lung Cancer Cell Line SPC-A-1 Induced By Hedyotis Diffusa Extract

Objective: hedyotis diffusa is a herb plant of one year of Rubia cordifolia species which has many function such as clearing up heat and detoxicating,activating blood circulation to dissipate blood stases,urinating and so on.modern study indicated that the main chemical composition of hedyotis diffusa have ursolic acid,immunopolysaccharide and hedyotis diffusain.they can strengthen immune activity and resist tumor activity.clinic observation fund out hedyotis diffusa has prominent efficacy to lung cancer.Proto-oncogene bcl-2 is the most important gene of inhibiting apoptosis,its gene expression production is a sort of protein which relative molecular weight is 26x10.It consists in mitochondrion membrane,nuclear membrane and endoplasmic reticulum membrane.bcl-2 can inhibit cell apoptosis,make cell live very longer, urge some oncogene to active and produce clone of malignant cell. In the present report, we want to study the effect of hedyotis diffusa extract on human lung cancer cell line and its possible effect mechanism. Method: 1. Effect of hedyotis diffusa extract on the growth of SPC-A-1 cell line was measured by means of SPC-A-1 cell proliferation experiments. 2. Invert microscope was used to detect the morphological changes of SPC-A-1 cells which were dealed with by hedyotis diffusa extract and DDP for 48h. 3. Fluorescenece staining microscope was used to to detect the morphological changes of dyed SPC-A-1 cell nucleus which were treated by hedyotis diffusa extract and DDP for 48h. 4. To detect DNA ladder Agarose gel electrophoresis was applied. 5. To detect the cell apoptosis rate FCM was performed. 6. Bcl-2 gene expression was evaluated with immunohistochemical analysis in SPC-A-1 cells which were treated by 12.5,25,50 mg/ml hedyotis diffusa extract and DDP for 48h.. Results: 1. Hedyotis diffusa extract could inhibit SPC-A-1 cell growth,its effect is related with the concentration and action time of hedyotis diffusa extract. 2. SPC-A-1 cells treated with hedyotis diffusa extract for 48h showed early changes of apoptosis. 3. Apoptotic cells demonstrated condensation and fragmentation of chromatin. 4. Agarose gel electrophoreris took on typical DNA ladder. 5. After SPC-A-1 cells were treated by 25mg/ml,50mg/ml hedyotis diffusa extract and DDP for 48h,FCM showed apoptosis mountain and apoptosis rate depended on drug concentration. 6. Immunohistochemical analysis confirmed that bcl-2 expression was decreased by hedyotis diffusa extract and decreased with the increase of drug concentration. Conclusion:a certain concentration of hedyotis diffusa extract can inhibit the growth of SPC-A-1 cells and can induce apoptosis of SPC-A-1 cells.Its action mechanism is related with bcl-2 gene.