Metabolism of acetate by human leukocytes
Pastore, Edward Joseph
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The purpose of this investigation was to study the metabolism of normal human leukocytes. Leukocytes were incubated in vitro with c14-labeled acetate, and the fate of the radioactive carbon was determined by fractionation and analysis of the major, cell components. Leukocytes were obtained from whole blood by fibrinogen sedimentation and differential centrifugation. Optimal conditions for isolation and incubation of viable cells were developed and assessed using phase microscopy for direct observation of their morphological integrity and by their oxidative metabolism. Control experiments to determine the possible effect of erythrocyte utilization of acetate were run using twice the number of cells ordinarily found in leukocyte suspensions. No utilization of acetate by red blood cells was observed. Respiration studies were performed using standard Warburg manometry. Otherwise, incubations were carried out in modified Erlenmeyer flasks equipped with center wells for C02 collection and stoppered with serum bottle caps. Flasks were equilibrated and various additions were made to the suspensions using hypodermic needles. Total cells per flask varied from 1 to 5 X 108 with concentrations ranging from 60 to 80 X 106 cells per ml. of suspension. Response of respiration and combustion of acetate were directly proportional to cell number and no detrimental effects due to cell crowding were detectable within this range. [TRUNCATED]
Thesis (Ph.D.)--Boston University
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