Adenylate Cyclase in Thymus-Derived and Bone Marrow-Derived Lymphocytes from Normal Donors and Patients with Chronic Lymphocytic Leukemia

Abstract

Lymphocytes were purified from peripheral blood of normal donors and patients with chronic lymphocytic leukemia (CLL) by Ficoll-Hypaque centrifugation. Adenylate cyclase activity, expressed as picomoles [³²P]cyclic AMP generated per milligram protein per minute, was 57±4 in normals and 26±4 in CLL patients. Enzyme activity, expressed as picomoles [³²P]cyclic AMP generated per 10⁶ lymphocytes per minute, was 2.09±0.19 for normal lymphocytes and 1.10±0.16 for CLL lymphocytes. The differences between normal and CLL peripheral lymphocytes are highly significant (P < 0.001) with either method of calculating activity. Cyclic AMP levels (picomoles per 10⁶ lymphocytes) also differed significantly: 1.38±0.29 for normals and 0.45±0.08 for CLL lymphocytes. Adenylate cyclase was assayed in lymphocytes enriched for bone marrow-derived (B) cells by removing E-rosetted thymus-derived (T) cells, and enriched for T cells by harvesting E-rosetted lymphocytes or by removing B cells with nylon wool absorption. Solutions to simultaneous equations gave the following calculated enzyme activities for pure B- and T-cell subpopulations (in picomoles [³²P]cyclic AMP generated per milligram mg protein per minute): normal B, 196±22; normal T, 30±10; CLL B, 34±6; CLL T, 19±4. Thus. normal B-lymphocyte adenylate cyclase exceeds normal T-lymphocyte activity by more than sixfold, whereas in the case of CLL the enzyme activity in B lymphocytes is markedly reduced to levels comparable to T lymphocytes. The responses of lymphocytes to stimulation with the hormones prostaglandin E₁ and isoproterenol, and with NaF, were assessed. Compared with normal lymphocytes, enzyme activities were reduced in CLL lymphocytes incubated with these agents, but to a degree paralleling the reduced basal activities. Thus, the ratios between stimulated and basal adenylate cyclase levels in Ficoll-Hypaque-purified, normal lymphocytes were 2.3±0.1 after incubation with 10 μM isoproterenol, and 3.9±0.2 with 10 mM NaF, values which did not differ significantly from those obtained with CLL lymphocytes. When the enzyme activities calculated for purified T- and B-lymphocyte subpopulations were used to derive the stimulation ratios, the responses of normal and CLL T and B cells to these agents were also indistinguishable. The simplest explanation for these findings is a reduced number of normally responsive enzyme sites on the surface membranes of CLL lymphocytes, although alternative explanations are possible.