A thermochemical study of metabolic pathways in activated and triggered 2C11–12 mouse macrophage hybridoma cells

Abstract

Abstract Hybridization with lymphosarcoma cells immortalized mouse macrophages for the study of phagocytosis and cytobicidal properties. In the study of macrophage physiology, it became necessary to know more of their cellular metabolism and the changes which occurred when the cells were triggered into the respiratory burst. It was found that, in common with many other types of growing cell, activated 2C11-12 macrophage hybridomas produced considerable lactate under fully aerobic conditions, judging from the highly exothermic CR ratio and subsequent spectrophotometric analysis. Most of both the substrates (glucose and glutamine) was converted to lactate, respectively, by glycolysis and glutaminolysis, in the demand for biosythetic precursors during growth. Glucose was the more important energy source. Approximately 60% of heat production was explained in terms of enthalpy changes in glucose and glutamine metabolism. It was suspected that fatty acid oxidation from contaminants in the bovine serum albumin needed for cell culture may be important in catabolism. The respiratory burst was triggered by phorbol-12-myristate-13-acetate and recorded by greatly (5-fold) increased heat production and enhanced chemiluminescence. Oxygen consumption was very rapid and soon led to anoxia in the closed culture system. The calorimetric-respirometric (CR) ratio was less negative and analysis confirmed that there was less lactate production. Radioisotope studies indicated that glycolysis and glutaminolysis were less intensive, with respiration of glucose accounting for over 90% of the heat production. The imperative for producing NADPH and cytotoxic oxygen metabolites heavily biased catabolism, reducing the supply of biosynthetic precursors. Known sources of heat production accounted for 87% enthalpy recovery and the remainder may well be caused by fatty acid oxidation.