Nicotinamide adenine dinucleotides permeate through mitochondrial membranes in human Epstein-Barr virus-transformed lymphocytes

Human cultured cells are widely used for the investigation of respiratory chain disorders. Oxidative properties are generally investigated by means of polarographic studies carried out on detergent-permeabilized cells. By studying the oxidative properties of Epstein-Barr virus-transformed B lymphocytes, we found that the respiration was significantly decreased after 3-4 days of cell culture. Simultaneously, we observed decreased NAD(+)-dependent oxidations (malate, glutamate, pyruvate) that became dependent upon the addition of exogenous NAD(+). The effect of NAD(+) was shown to be related to an influx of catalytic amount of NAD(+) into the mitochondrial matrix. A full ability to oxidize NAD(+)-dependent substrates was restored less than 2 h after a change of the culture medium. These observations suggested: (a) the occurrence of fluxes of catalytic amounts of NAD(+) through the mitochondrial inner membrane in human cells; (b) an early control of mitochondrial metabolism by matrix NAD(+) content in cells grown under limiting growth conditions; (c) the possible confusion between complex I deficiency and a decrease content of matrix NAD(+) when using human cultured cells.