Ca²⁺ Selectively Enhances Doxorubicin-induced Apoptosis In Human Hepatoma Cells And Its Mechanism

Hepatocellular carcinoma （HCC）, a primary hepatic tumor with aggressive malignanceand high prevalence, is one of the most common malignancies in the world. So far, theoverall survival of patients with HCC is not optimistic due to the poor selectivity of thechemotherapy drugs. Our early research indicated that Ca²⁺can selectively enhancedoxorubicin-induced cell growth inhibition in HepG2cells. Many chemotherapy drugs playthe anti-cancer effect by the mechanism of inhibiting cell proliferation and promotingapoptosis. Mitochondrial pathway plays a very important role in the process of apoptosis. Inthis paper, we study the influence of calcium on apoptosis of HepG2and L02cellsdoxorubicin induced and study the mechanism of involvement of mitochondrial pathway.Part I The different influences of calcium on Adriamycin induced HepG2andL02cells apoptosisObjective: To investigate whether Ca²⁺can selectively enhance ADM-induced apoptosis inHepG2.Methods: Cells were treated with ADM、Ca²⁺or ADM+Ca²⁺(Ca²⁺9mM, ADM0.1μg/mL,Ca²⁺9mM+ADM0.1μg/mL) for48hours. FCM was used to measure apoptosis of HepG2and L02cells.Results: For single ADM group, the apoptosis rates of L02cells were clearly higher thanthat of HepG2cells （P<0.05）. In combination tests, the apoptosis rates of HepG2cells weregreatly increased than that of signal ADM group （P<0.05） and higher than that of L02cells（P<0.05）, whereas no increases were observed in L02cells （P>0.05）.Conclusions: Ca²⁺can enhance ADM-induced apoptosis in HepG2cells, whereas Ca²⁺doesn’t affect ADM-induced apoptosis in LO2cells. Part II The mechanism of the different influences of calcium on Adriamycininduced HepG2and L02cells apoptosisObjective: To investigate the mechanism of Ca²⁺selectively enhances ADM-inducedapoptosis.Methods: Cells were treated with ADM、Ca²⁺or ADM+Ca²⁺(Ca²⁺9mM, ADM0.1μg/mL,Ca²⁺9mM+ADM0.1μg/mL) for48hours. Apoptosis-related proteins expressions weremeasured by using western-blot. Microplate reader was used to measure the enzyme activity ofcaspase-3.Results: For HepG2cells, compared with single ADM group, bax protein expressions andthe enzyme activity of caspase-3were increased（P<0.05） and bcl-2、procaspsase-3proteinsexpressions were decreased in combination tests（P<0.05）, but there were no changes in theexpressions of bax、bcl-2and procaspsase-3and the enzyme activity of caspase-3（P>0.05） inL02cells.Conclusions: The mechanism that Ca²⁺can enhance ADM-induced apoptosis in HepG2cells may be Ca²⁺increase bax expressions and decrease bcl-2protein expressions followedby switching from procaspase-3to caspase-3, which executed apoptosis.