Cytotoxicity of Low Dose Epirubicin-HCI Combined with Lymphokine Activated Killer Cells Against Hepatocellular Carcinoma Cell Line Hepatoma G2

Authors: AYSUN ÖZKAN, KAYAHAN FIŞKIN

Abstract: Recent evidance supports the concept that Epirubicin-HCI and Lymphokine-activated killer cells (LAK) cytotoxicity may be mediated by free radical and oxyradical generation. We tested this hypothesis further by exposing Hep G2 cells to Epirubicin-HCI and LAK cells and subsequently monitored cell viability as a measure of cytotoxicity. The cytotoxicity of LAK alone and LAK together with low dose epirubicin-HCI (IC50 1/10) which is measured using the MTT cytotoxicity test on viability of Hep G2 cells and sensitization of target cells to effector cells was investigated. The cytotoxicity of LAK (IC50 value of 5LAK/Hep G2 in 24 hours) and LAK+epirubicin-HCI (IC50 value of 2.5LAK+epirubicin-HCI/Hep G2 cell in 24 hours) appeared to involve a free radical species production type of mechanism since free radicals scavenger enzymes activity, Mn-SOD, Cu,Zn-SOD, Se-dependent GPx and catalase, were increased (p<0.01). Preincubation of Hep G2 cell with SOD before adding LAK and LAK+epirubicin-HCI prevented increasing enzymes activity and cytotoxicity. Also, after LAK and LAK+epirubicin-HCI treatment, increasing expression of NADPH-dependent Cytochrome P450 reductase supported cytotoxicity results depending on free radical production. Increasing activity of Mn-SOD, Cu,Zn-SOD, Se-dependent GPx and catalase are higher in Hep G2 cell treated with combining epirubicin-HCI with LAK than LAK alone (p<0.01). The combining treatment made the Hep G2 cell more sensitive to free radical production and cytotoxicity than LAK treatment alone. SOD, catalase, glutathione peroxidase and NADPH-dependent Cytochrome P450 reductase must be considered as part of the intracellular antioxidant defense mechanism of Hep G2 cells against to single electron reducing quinone-containing anticancer antibiotics and free radical production as result of excess amount of NO synthesis.

Keywords: Hepatoma, epirubicin-HCI, cytotoxicity, enzymes

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