Silymarin inhibited proliferation and induced apoptosis in hepatic cancer cells.
OBJECTIVE
The aim of this study was to investigate mechanisms involved in the growth inhibitory effect of silymarin, in humanhepatocellular carcinoma.


METHODS
The human hepatocellular carcinoma cell line HepG2 was utilized and the MTT assay was performed to study the antiproliferative effect of silymarin. Dual staining was undertaken for ethidium bromide/acridine orange, propidium iodide staining and DNA fragmentation studies were executed to confirm the presence of apoptosis. Cell-cycle analysis was revealed by flow cytometry and mitochondrial transmembrane potential was measured by uptake of the mitochondrial-specific lipophilic cationic dye rhodamine 123. Western blotting analysis for cytochrome c, p53, Bax, Bcl-2, APAF-1, caspase-3, survivin, beta-catenin, cyclin D1, c-Myc and PCNA was carried out.


RESULTS
Silymarin inhibited population growth of the hepatocellular carcinoma cells in a dose-dependent manner, and the percentage of apoptotic cells was increased after treatment with 50 and 75 microg/ml silymarin for 24 h. Silymarin treatment increased the proportion of cells with reduced DNA content (sub-G(0)/G(1) or A(0) peak), indicative of apoptosis with loss of cells in the G(1) phase. Silymarin also decreased mitochondrial transmembrane potential of the cells, thereby increasing levels of cytosolic cytochrome c while up-regulating expression of pro-apoptotic proteins (such as p53, Bax, APAF-1 and caspase-3) with concomitant decrease in anti-apoptotic proteins (Bcl-2 and survivin) and proliferation-associated proteins (beta-catenin, cyclin D1, c-Myc and PCNA).


CONCLUSIONS
Our results demonstrate that silymarin treatment inhibited proliferation and induced apoptosis in the human hepatocellular carcinoma cell line HepG2.
Ramakrishnan
G
G
Department of Biochemistry, University of Madras, Guindy Campus, Chennai, Tamil Nadu, India.
Lo Muzio
L
L
Elinos-Báez
C M
CM
Jagan
S
S
Augustine
T A
TA
Kamaraj
S
S
Anandakumar
P
P
Devaki
T
T