Cell Requirements for Optimum Proliferative Responses of Rat Spleen Cells

Abstract

Spleen cells from New Zealand black rats (NBR) were separated by nylon wool (NW) and by density gradient centrifugation (Percoll) to identify certain physical, chemical, morphological and functional attributes of mononuclear cells. Density distribution profiles of nylon wool nonadherent (NA) and adherent (ADH) cells indicated that at least two different populations of spleen cells existed (NA = 1.070–1.075 g/ml; ADH = 1.064 g/ml). 38% of all NA cells and 13 % of all ADH cells are T-cells (mean density = 1.080 g/ml). Nonspecific esterase staining indicated gradient enrichment of ADH cells at densities of 1.064 g/ml (mononuclears) and 1.080 g/ml (polymorphonuclears). NA cells contained 3 % esterase positive cells. Plastic adherent cells had a mean density distribution of 1.064 g/ml. Mitogen binding cells were identified by function and density. ADH cells (1.064 g/ml) demonstrate higher specific binding of PHA-I 125 compared to NA cells (1.072 g/ml). Unlike the PHA binding cells, Con-A binding ADH and NA cells share the same mean densities (1.055–1.064 g/ml). As with the PHA responses, ADH cells have greater specific binding activities for Con-A than NA cells. Mitogen binding cells have lower mean densities than cells which incorporate tritiated thymidine at 5 days post-cultivation with PHA or Con-A. Adherent cells enriched for T cells and polymorphonuclears produced the highest PHA responses. PHA responsive NA cells were not enriched by gradient separation. NA cells produced a suppressive effect when mixed with cells which produced optimum PHA responses. ADH cells (density 1.073 g/ml) produced a significant PHA response helper effect when mixed with gradient separated cells. A helper effect was not produced by this fraction in the presence of Con-A responsive cells. Unlike PHA responses (where optimum binding and proliferation occurred by ADH cells), enrichment for Con-A responders occurred in NA gradient fractions (1.073–1.080 g/ml).