Abstract:

Objective To study influence of andrographolide（AD）on cell growth、apoptosis and proliferating cell nuclear antigen(PCNA) protein expression in human skin carcinoma A431 cells. Methods Acid phosphatase assay (APA) was used to detect
inhibition of cell proliferation. Cell morphology was observed by fluorescence microscopy. Annexin V-FITC/PI double staining method was used to detect A431 cell apoptosis by the flow cytometry (FCM). Rh123 staining detected mitochondrial membrane potential by FCM. PCFNA protein expression on A431 cells was detected by immunocytochemistry. Results AD inhibited A431 cell proliferation by time and dose dependent. There was a significant difference between 50mg/L, 100mg/L AD group and vehicle control group (P<0.05) on the same time. A part of nuclear chromatin appeared typical apoptosis morphological changes after AD group acted A431 cells for 24 hours.The early apoptosis ,late apoptosis and necrosis rates were increased with various AD concentrations treatment for 24 hours. There was a significant inhibition rate difference between 50mg/L, 100mg/L and vehicle control group(P<0.05). PCNA protein expression intensity was decreased gradually with AD treatment for 24 hours.
Mitochondrial membrane potential decreased significantly after AD acted on A431 cells for 24 hours; 50mg/L and 100mg/L AD groups had significant difference compared to vehicle control group（P＜0.05）. Conclusion AD significantly inhibits A431 cells proliferation and AD inhibiting cells proliferation may be associated with PCNA protein down-regulation.AD induces A431 cells apoptosis and AD inducing apoptosis mechanism may be related to decreasing mitochondrial membrane potential.

Key words: Andrographolide, A431 cells, Mitochondrial membrane potential, PCNA protein, Flow cytometry, Acid phosphatase assay, Human