Normal levels of lymphotoxin secretion by freshly isolated and refrigerated human peripheral blood lymphocytes

Abstract

Lymphocytes obtained by Ficoll-Hypaque density gradient centrifugation of leukocytes from 40 blood bank donors were incubated at 10 6 cells/ml for 24 h with 5 μg phytohemagglutinin/ml serum-free RPMI 1640 medium and secreted 34 ± 6 U lymphotoxin/ml and 52 ± 29 U interferon/ml. Addition of 5% fetal bovine serum (FBS) or acid-citrate-dextrose treated autologous plasma (AP) augmented lymphotoxin secretion 2-fold and interferon secretion 6–8-fold. Lymphotoxin alone was secreted in 39% of serum free cultures. Both lymphotoxin and interferon were secreted in the presence of FBS or AP. Lymphocytes refrigerated at 4°C for 18 h secreted upon serum-free culture similar levels of lymphotoxin as fresh cells but cultures secreting lymphotoxin alone increased from 39 to 77%. Lymphotoxin secretion by refrigerated lymphocytes was similar in the presence or absence of FBS or AP. Interferon secretion by refrigerated lymphocytes in the presence of FBS or AP, however, increased 30–40-fold approaching levels produced by freshly isolated cells and indicating that the modulatory effects of refrigeration, FBS, and AP affected interferon not lymphotoxin secretion. The levels of lymphotoxin and interferon secreted by lymphocyte populations producing both lymphokines, furthermore, varied independently of one another providing additional evidence for the dissociability of their secretion yet possible dependence of interferon upon lymphotoxin secretion. The ability to reliably quantitate lymphotoxin secretion using refrigerated lymphocytes permits transport of lymphocytes to laboratories performing quantitative bioassay of the lymphokine. This should facilitate further evaluation of the presence and role of lymphotoxin in immunopathological and other disease states.