Induction of apoptosis and necrosis in human peripheral blood mononuclear cells by fatty acid ethyl esters

Background: Fatty acid ethyl esters (FAEE), nonoxidative products of ethanol metabolism, are formed by the esterification of fatty acids and ethanol. Alcoholic subjects have high levels of FAEE in the circulation as well as in organs and tissues, especially those most often damaged by ethanol abuse. Our previous studies showed a significant synthesis of FAEE by human mononuclear cells within seconds of exposure to physiologic doses of ethanol. In addition, FAEE inhibited phytohemagglutinin (PHA)-stimulated interleukin-2 production and calcium (Ca²⁺) influx into human mononuclear cells. FAEE also caused a rapid increase in the intracellular cAMP. The mechanism by which alcohol suppresses the immune system remains undetermined. Objectives: To evaluate the morphological and physiological effects of FAEE on human mononuclear cells and to study the impact of FAEE on cell viability. Methods: Mononuclear cell fractions of human white blood cells (WBC) were incubated with physiological doses (25 and 50 μM) of ethyl oleate, a representative FAEE, for 15, 30, 60, 120 or 180 minutes. Morphological changes were evaluated by light and transmission electron microscopy (TEM). Lactate dehydrogenase (LDH) release was measured as a physiological indicator of necrosis. Physiological changes were also evaluated by western blots performed on whole-cell lysates of treated and untreated cells and by DNA electrophoresis. Results: Significant morphological changes were detected in cells exposed to FAEE by both light and TEM. Concentration and time-dependent increases in the rates of apoptosis and necrosis were found by light microscopy and by LDH release, respectively, following 60 minutes exposure to 25 or 50 μM FAEE. One-hour 50 μM FAEE exposure caused activation of the caspase cascade, as demonstrated by Poly (ADP-ribose) Polymerase (PARP) cleavage, and significant DNA damage as a result of necrosis in human mononuclear cells. Conclusions: These studies provide evidence to support the toxic effects of FAEE on intact human mononuclear cells. The results from our studies also show that both apoptosis and necrosis are modes of cell death in FAEE-treated human mononuclear cells. This may be an important mechanism in alcohol-induced immunosuppression.