Quantitative analysis of surface marker densities after exposure of T-cells to concanavalin A (Con A): a sensitive early index of cellular activation.
The kinetics of expression of activation-linked T-cell surface markers were analyzed in T-cells obtained from normal donors. The cells were cocultured in triplicate for 0, 1, 24, 48, and 72 hr in the presence of Con A in RPMI 1640. The density of HLA-DR, interleukin 2, and transferrin receptors (IL-2-R and TR, respectively) on the surface of CD3- and CD8-positive cells was computed by a mathematical model based on fluorescence intensity vector analysis, adjusted for cell size, utilizing two-color flow cytometry. The results were compared with controls obtained at the same time with control cells cultured in RPMI 1640 alone. There was a significant increase in the mean density of HLA-DR on the surface of CD3- and CD8-positive cells as early as 1 hr after exposure to Con A when compared with controls (250 and 300%, respectively; P less than 0.0001). The mean density of IL-2-R and TR on CD3+ cells increased by 265 and 208%, respectively; P less than 0.06 and P = n.s., respectively, when compared with control cells. The mean density of Class II MHC products on CD3+ and CD8+ cells treated with RPMI alone increased by 202 and 468%, 234 and 540%, and 1375 and 2442%, respectively, at 24, 48, and 72 hr of culture. In contrast, the mean cell surface density of these markers in cells treated with Con A increased by 614 and 1962%, 3304 and 7231%, and 8665 and 22,619%, respectively (P less than 0.00001) at the corresponding times following exposure to Con A. The density of IL-2-R and TR on CD3+ cells exposed to Con A also increased significantly at 24, 48, and 72 hr (P less than 0.0001). At the same times, the relative percentage of cell subsets bearing these particular markers increased by 78, 138, and 175% at 24, 48, and 72 hr, respectively. The data suggest that objective quantitative evidence of lymphocyte activation after exposure to Con A may be obtained as early as 1 hr after antigen stimulation, and before significant changes in cell numbers occur. Measurement of cell surface marker densities may provide a useful index for the detection and quantitation of cell activation in the early phase of antigenic stimulation.
Shabtai
M
M
Department of Surgery (Transplantation Service), State University of New York, Stony Brook 11794-8192.
Malinowski
K
K
Waltzer
W C
WC
Pullis
C
C
Raisbeck
A P
AP
Rapaport
F T
FT
D000945:Antigens, Differentiation, T-Lymphocyte / Q000032:analysis; D000954:Antigens, Surface / Q000032:analysis*; D017252:CD3 Complex; D016827:CD8 Antigens; D003208:Concanavalin A / Q000494:pharmacology*; D006684:HLA-DR Antigens / Q000032:analysis; D006801:Humans; D008213:Lymphocyte Activation; D011948:Receptors, Antigen, T-Cell / Q000032:analysis; D015375:Receptors, Interleukin-2 / Q000032:analysis; D011990:Receptors, Transferrin / Q000032:analysis
D000945:Antigens, Differentiation, T-Lymphocyte; D000954:Antigens, Surface; D017252:CD3 Complex; D016827:CD8 Antigens; D006684:HLA-DR Antigens; D011948:Receptors, Antigen, T-Cell; D015375:Receptors, Interleukin-2; D011990:Receptors, Transferrin; D003208:Concanavalin A