Nonspecific esterase activity in human lymphocytes: Histochemical characterization and distribution among major lymphocyte subclasses

Abstract

In a histochemical assay 75±8% of human blood lymphocytes hydrolyzed α-naphthyl acetate after an 8-hr exposition to the substrate. The brownish-red reaction product was easily distinguished in counterstained cytocentrifuged cell smears in lymphocyte cytoplasm. The reaction had a broad temperature optimum of 20°C, it was optimal at a pH of 5.8, and nearly all lymphocytes with histochemically demonstrable esterase activity were stained within 6 hr of exposure. Lipase activity was ruled out, as β-naphthyl acetate could also be used as substrate. Cholinesterases were also ruled out, as the reaction could not be inhibited by a 10 μ M concentration of eserine. As the reaction was inhibited in 10 m M but not in 10 μ M E-600, and could not be reactivated after E-600 inhibition with a 100 μ M concentration of PCMB, the enzyme responsible for the reaction was defined as a nonspecific A-esterase. The α-naphthyl acetate esterase (ANAE) marker-carrying lymphocytes were enriched and depleted by fractionation procedures which enriched and depleted the SRBC rosette-forming lymphocytes. The number of ANAE marker-carrying lymphocytes closely followed the distribution of SRBC rosette-forming lymphocytes in spleen, tonsil, bone marrow, and blood and was inversely related to the number of surface Ig-carrying lymphocytes in these organs. Furthermore, visualization of the ANAE marker simultaneously with the surface marker demonstrated that, while more than 80% of the SRBC rosette-forming lymphocytes carried the ANAE marker, only 10%, at most, of the lymphocytes expressing large amounts of surface Ig were ANAE positive. Discordant findings were observed in the thymus: 72% of the thymocytes formed rosettes with SRBC, whereas only 13% of the thymocytes carried the ANAE marker. The results, therefore, suggest the presence of histochemically demonstrable nonspecific esterase activity predominantly in resting, mature T lymphocytes.