BACKGROUND: Hepatocellular carcinoma (HCC) is the third leading cause of cancer 
deaths reported worldwide. The incidence is higher in Asia and Africa, where 
there is greater endemic prevalence of hepatitis B and C. The devastating 
outcome of cancer can be minimized only by the use of potent therapeutic agents. 
Tridham (TD) has been acknowledged since olden days for its wide spectrum of 
biological properties and was used by traditional practitioners of Siddha and 
other indigenous systems of medicine. The present study aims at investigating 
the mechanistic action of TD by assessing the antiproliferative and 
pro-apoptotic effects on human hepatocellular carcinoma cell line (Huh7).
METHODS: Cell viability and apoptosis assay using MTT analysis and trypan blue 
staining, DAPI staining, DNA fragmentation, cell cycle analysis, mitochondrial 
membrane potential, real-time reverse transcription-polymerase chain reaction, 
western blotting and immunofluorescence staining were determined in Huh7 cells.
RESULTS: Viability studies of TD treated Huh7 cells showed an inhibition in cell 
growth in time and dose dependent manner. Chromatin condensation, DNA 
fragmentation and apoptotic bodies, which are structural changes characteristic 
of apoptosis, were found following TD treatment of Huh7 cells. DAPI staining and 
agarose gel electrophoresis confirmed the induction of apoptosis by TD. Cell 
cycle analysis of Huh7 cells treated with TD exhibited a marked accumulation of 
cells in the sub-G1 phase of the cell cycle in a dose dependent manner. 
Immunofluorescent staining for Ki-67 showed a higher level of expression in 
untreated cells as compared to TD treated cells. We observed a significant loss 
in the mitochondrial membrane potential and the release of cytochrome c into the 
cytosol in TD treated cells. Down regulation of Bcl-2, up regulation of Bax and 
Bad as well as activation of caspases-3 and 9 were also observed. The p53 gene 
expression was found to be unaltered in TD treated cells.
CONCLUSION: These results suggest that TD induces apoptosis of Huh7 cells 
through activation of Bax and triggered caspase cascade, independent of p53 
function. This study throws light on the mechanistic action of TD in triggering 
apoptosis in Huh 7 cells.