Plasma clearance and endocytosis of cytosolic malate dehydrogenase in the rat.

1. Pig heart cytosolic malate dehydrogenase was radiolabelled with O-(4-diazo-3,5-di-[125I]iodobenzoyl)sucrose and intravenously injected into rats. Enzyme activity and radioactivity were cleared from plasma identically, with first-order kinetics, with a half-life of about 30 min. 2. The tissue distribution of radioactivity was determined at 2 h after injection. All injected radioactivity was recovered from the tissues. A high percentage of the injected dose was found in liver (37%), spleen (6%) and bone including marrow (19%). 3. Radioactivity in liver and spleen increased up to 2 h after injection and subsequently declined, with a half-life of about 20 h. 4. After differential fractionation of liver, radioactivity was largely found in the mitochondrial and lysosomal fraction. 5. Liver cells were isolated 1 h after injection of labelled enzyme. We found that Kupffer cells, endothelial cells and parenchymal cells had endocytosed the enzyme at rates corresponding to 2725, 94 and 63 ml of plasma/day per g of cell protein respectively. 6. Radioautography indicated that in spleen and bone marrow the enzyme is mainly taken up by macrophages. 7. Internalization of the enzyme by liver, spleen and bone marrow was saturable. This indicates that the enzyme is taken up in these tissues by adsorptive endocytosis. 8. The present results closely resemble those obtained previously for the mitochondrial isoenzyme of malate dehydrogenase and for lactate dehydrogenase M4. Since those enzymes are positively charged at physiological pH, whereas cytosolic malate dehydrogenase is negative, net charge cannot be the major factor determining the rate of uptake of circulating enzymes by reticuloendothelial macrophages, as has been suggested in the literature [Wachsmuth & Klingmüller (1978) J. Reticuloendothel. Soc. 24, 227-241].