Abstract
BackgroundHepatocellular 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).MethodsCell 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.ResultsViability 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.ConclusionThese 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.
Highlights
Hepatocellular carcinoma (HCC) is the third leading cause of cancer deaths reported worldwide
IC50 values of TD in Human hepatocellular carcinoma cell line (Huh7) cells after 24 and 48 h of exposure were determined as 100 μg/ml, which is used as an effective concentration for subsequent assays
The results indicate that exposure of Huh7 cells to TD triggers the pathway of apoptosis leading to decrease in viability of Huh7 cells
Summary
Hepatocellular carcinoma (HCC) is the third leading cause of cancer deaths reported worldwide. A life-threatening health problem, is one of the leading causes of death world-wide [1]. Hepatocellular Carcinoma (HCC) is the third most common cause of cancer-related deaths reported world-wide. Activation of caspase 9 is associated with changes in the permeability of the outer mitochondrial membrane and the collapse of membrane potential. The latter results in release of cytochrome c and activation of downstream effector caspases such as caspase-3 [8,9]. Cytochrome c is thereby released into the cytoplasm This event triggers a biochemical cascade resulting in the activation of caspase proteases which leads to apoptotic cell death
Sign-in/Register to view highlights & summary 
Published Version (
Free)
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call