Assessment of cytotoxicity in vitro. The murine hepatoma cell line Hepa1c1c7 purchased from American Type Culture Collection (Manassas, VA) was used to assess cytotoxicity in vitro. Hepa1c1c7 cells were maintained in Dulbecco's modified Eagle's medium (Invitrogen, Carlsbad, CA) with 1% antibiotic-antimycotic (Invitrogen) and 10% heat-inactivated fetal bovine serum (SAFC Biosciences, Lenexa, KS) in 75-cm 2 tissue culture flasks at 37°C in a humidified atmosphere of 95% air and 5% CO 2 . Cells were plated in 96well plates at 15,000 cells per well and allowed to attach for 8 h before medium was replaced. Various concentrations of AMD, DEA, and/or TNF were added to designated wells, and cells were incubated under maintenance conditions. Twenty-four hours later, lactate dehydrogenase (LDH) activity released into the culture medium was measured using the Cytotox-One Homogeneous Membrane Integrity Assay (Promega, Madison, WI). The percent LDH release was calculated as LDH in supernatant/(LDH in supernatant þ LDH in cell lysate). Lysate LDH was determined after addition of Triton to lyse the cells.