Harmine induces apoptosis in HepG2 cells via mitochondrial signaling pathway

摘要

BACKGROUND: Harmine  has  antitumor  and  antinociceptive effects,  and  inhibits  human  DNA  topoisomerase.  However, no detailed data are available on the mechanisms of action of harmine  in  hepatocellular  carcinoma.  This  study  aimed  to investigate the effects of harmine on proliferation and apoptosis, and  the  underlying  mechanisms  in  the  human  hepatocellular carcinoma cell line HepG2. METHODS: The proliferation of HepG2 cells was determined by  the  cell  counting  kit-8  (CCK-8)  assay  and  the  clone formation test. The morphology of HepG2 cells was examined using  fluorescence  microscopy  after  Hoechst  33258  staining. Annexin  V/propidium  iodide  (PI)  was  used  to  analyze apoptosis  and  PI  to  analyze  the  cell  cycle.  Western  blotting was  used  to  assess  expression  of  the  apoptosis-regulated genes  Bcl-2,  Bax,  Bcl-xl,  Mcl-1,  caspase-3,  and  caspase-9. Mitochondrial transmembrane potential (Ψm) was determined using JC-1. RESULTS: Harmine inhibited the proliferation of HepG2 cells in a dose-dependent manner. Hoechst 33258 staining revealed nuclear  fragmentation  and  chromosomal  condensation,  cell shrinkage,  and  attachment  loss  in  HepG2  cells  treated  with harmine. The percentage of the sub/G1 fraction was increased in  a  concentration-dependent  manner,  indicating  apoptotic cell death. PI staining showed that harmine changed the cell cycle  distribution,  by  decreasing  the  proportion  of  cells  in G0/G1 and increasing the proportion in S and G2/M. Harmine induced apoptosis in a concentration-dependent manner, with rates of 20.0%, 32.7% and 64.9%, respectively. JC-1 revealed a decrease in Ψm. Apoptosis of HepG2 cells was associated with caspase-3 and caspase-9 activation, down-regulation of Bcl-2, Mcl-1, and Bcl-xl, and no change in Bax. CONCLUSIONS: Harmine  had  an  anti-proliferative  effect in  HepG2  cells  by  inducing  apoptosis.  Mitochondrial  signal pathways  were  involved  in  the  apoptosis.  The  cancer-specific selectivity  shown  in  this  study  suggested  that  harmine  is  a promising novel drug for human hepatocellular carcinoma.