Generation of a lymphocyte growth factor by treatment of human cells with neuraminidase and galactose oxidase.

Supernates of neuraminidase and galactose oxidase (NAGO)-treated lymphocytes induce blastogenesis in nonproliferating cells harvested 7--14 d after treatment with mitogen or alloantigen and in cells incubated with mitogen for 7--14 d but not in freshly isolated peripheral blood lymphocytes9 Virtually all the growth factor is produced by NAGO-treated cells during the first 24 h of incubation, and no increase in factor activity is detected upon further cell culture. Serum is not required for growth factor production. NAGO-primed medium induces generation of specific cytotoxic T cells from mixed lymphocyte culture (MLC) memory cells to approximately the same extent as that induced by allogeneic cells (stimulating cells in the primary MLC). NAGO-primed medium provides a useful reagent for isolation and characterization of lymphocyte growth factors and other lymphokines.

Biosynthesis of gamma globulin: studies in a cell-free system.

Incorporation of C(14)-amino acids into high-molecular-weight material precipitable by trichloroacetic acid indicates that microsomal cell-free systems, derived from spleens of immunized rabbits, are active in protein synthesis. Protein was made soluble by ultrasonic irradiation of the cell-free incubation mixtures, and low-molecular-weight materials were removed by dialysis and gel filtration. Chromatography and radio-immunoelectrophoresis of this soluble protein fraction reveal C(14)-labeled protein having several characteristics of gamma globulin.

Collagen-derived membrane: corneal implantation.

Behavior of membranes derived from collagen was investigated in rabbit corneas. Disks of the membrane were placed between the lamellae of corneas which were then examined grossly, biomicroscopically, and histologically. The membranes remained clear, and almost no reaction or evidence of reabsorption was seen during an observation period of 6 months. These characteristics make the material potentially useful for heterotransplantation in the cornea.

Collagen gels: design for a vitreous replace- ment.

Clear, stable gels have been prepared from purified tropocollagen from calf skin; the collagen was solubilized with a proteolytic enzyme (Proctase) and stabilized by ultraviolet irradiation under nitrogen. These gels are clear, possess altered immunologic reactivity, and have properties of an ideal vitreous replacement. Implantations in rabbit and monkey eyes appear to be well tolerated, remain clear, and gradually disappear in about 2 months.

Induction of cytolytic activity by anti-T3 monoclonal antibody. Activation of alloimmune memory cells and natural killer cells from normal and immunodeficient individuals.

The ability of the monoclonal antibody directed at the T3 antigen (anti-T3) to induce cytolytic activity was investigated since several agents that can activate T cells induce the acquisition of cytolytic activity in a variety of test systems. Pretreatment of human alloimmune memory cells, generated in primary long-term mixed lymphocyte cultures, with anti-T3 resulted in the induction of statistically significant specific secondary cytolytic activity and natural killer (NK) cell-like activity. No such augmentation or induction of cytolytic activity was found with anti-T3 pretreatment when syngeneic cells or inappropriate allogeneic cells (HLA-A, B antigens different from the original priming stimulus) were used as target cells and pretreatment of memory cells with anti-T4 or anti-T8 did not induce cytolytic activity to allogeneic or syngeneic target cells. Differential effects were observed when anti-T3 was added to the cytotoxicity assay in which anti-T3 pretreated alloimmune memory cells were effectors. The addition of anti-T3 to the assay prior to the introduction of target cells resulted in 39 +/- 8% inhibition of specific secondary cytolytic activity and only 5 +/- 8% inhibition of NK cell activity. NK cell activity mediated by large granular lymphocyte-enriched fraction of peripheral blood mononuclear cells (PBM) obtained from normal individuals was significantly augmented by anti-T3 when NK-sensitive cell lines MOLT-4 or K-562 were used as target cells. This augmentation in NK cell activity was not associated with nonspecific cytotoxicity to syngeneic or allogeneic PBM, and anti-T3 failed to activate the LGL fraction depleted of T cells. The monoclonal antibodies, anti-T4 or anti-T8, did not increase NK cell activity. NK cell activity mediated by PBM from eight immunodeficient individuals (four with acquired immunodeficiency syndrome and four with renal allografts) was also significantly augmented by anti-T3 pretreatment. Our findings, in addition to providing a rationale for the frequent occurrence of re-rejection episodes in renal graft recipients treated with anti-T3, suggest that anti-T3 might be utilized to enhance the cytotoxic armamentarium of immunodeficient patients.

Immunologic responses of graft recipients to antilymphocyte globulin: effect of prior treatment with aggregate-free gamma globulin.

The immunologic response of graft recipients to antilymphocyte globulin has been studied. The clearance from the serum of (125)I-labeled antilymphocyte globulin was studied in 15 graft recipients previously treated with antilymphocyte globulin and in 4 control patients not previously treated with antilymphocyte globulin. The mean serum half-life of antilymphocyte globulin was 7.2 days in control patients, 3.8 days in 13 renal graft recipients, and 22 hr in 2 heart graft recipients. All but one of the antilymphocyte globulin-treated patients had rapid clearance. Patients treated with equine antilymphocyte globulin had rapid clearance of rabbit and goat antilymphocyte globulin as well as horse antilymphocyte globulin. All patients with rapid clearance of antilymphocyte globulin had circulating antibodies to xenogeneic gamma globulin. Two patients with rapid clearance of antilymphocyte globulin had circulating complexes of antilymphocyte globulin. Five renal graft recipients were treated with aggregate-free equine gamma globulin before antilymphocyte globulin therapy in an attempt to induce tolerance to xenogeneic gamma globulin. In these five patients neither rapid clearance of antilymphocyte globulin nor significant titers of circulating antibody to xenogeneic gamma globulin developed. The induction of tolerance to xenogenic gamma globulin may benefit patients treated with antilymphocyte globulin.

Protein A-independent tumoricidal responses in dogs after extracorporeal perfusion of plasma over Staphylococcus aureus.

Protein A-positive or -negative Staphylococcus aureus preparations were used in an extracorporeal system to treat dogs with spontaneously occurring cancers. Tumor regression was seen in 4 of 7 dogs treated by reinfusion of plasma that had been incubated with protein A-positive S. aureus Cowan I strain (SAC). Therapy was associated with fever, liver enzyme abnormalities, and hypocomplementemia. Tumor response and toxicity could be diminished by more extensive washing of the SAC preparation. Tumor regression was also seen in 2 of 2 animals treated with protein A-negative S. aureus Wood strain 46. In addition, tumors regressed in 3 of 4 dogs treated with infusions of protein A-free saline extracts from S. aureus. These results suggest that the release of a non-protein A bacterial product contributes to tumor regression following incubation of plasma with S. aureus.

Suppression of transformation by and growth adaptation to low concentrations of glutamine in NIH-3T3 cells.

NIH-3T3 cells, commonly used as targets for oncogene-mediated neoplastic transformation, undergo high rates of spontaneous transformation. When the glutamine concentration in the medium was reduced from 5 to 1 mM or less, the transformation rate was reduced. This effect was not dependent upon a reduction in the growth rate, which remained unaffected by reduction of glutamine even to 0.6 mM. Upon trypsinization and transfer to 5 mM glutamine-containing medium, cells exposed to 0.2 mM glutamine for as little as 4 days formed fewer foci than control cells exposed over a similar period to 5 mM glutamine. This indicates that short term changes in the supply of this polyfunctional metabolite have heritable consequences in later cell generations. If populations containing highly transformed cells were passaged weekly for 1-3 weeks in 0.2 mM glutamine, resultant populations were better adapted to grow in low-glutamine medium and formed fewer transformed foci upon re-transfer to 5 mM glutamine medium, suggesting that the transformed state is at least partially reversible. If similar cell populations were exposed to low-glutamine medium but were not passaged, growth adaptation occurred but there was no reduction in focus formation, indicating that maintenance of a moderate rate of cell division may be required in addition to the lowered glutamine for reversal of transformation. Transformed and non-transformed cells originating from foci and from nonfocal areas of the same culture dishes multiplied at the same reduced rate in 0.2 mM glutamine. This indicates that suppression of spontaneous transformation in low-glutamine medium was not the result of selecting pre-existing variants but was itself an adaptive response of the population.

Differential regulation by retinoic acid and calcium of transglutaminases in cultured neoplastic and normal human keratinocytes.

In five lines of cultured human squamous carcinoma cells, transglutaminase activity and envelope competence were highly sensitive to retinoic acid and calcium levels in the growth medium. In cells grown in low calcium medium, these measures of keratinocyte differentiation were reduced. Retinoic acid suppressed envelope competence but total transglutaminase activity was markedly reduced, slightly affected, or greatly stimulated depending upon the cell line and whether the cells were grown in low calcium or 1.8 mM calcium-containing medium. Examination by anion exchange chromatography of the transglutaminase activity in SCC-12B2 cultures showed that expression of the particulate form (type I) of the enzyme was greatly stimulated by calcium. The increase in this activity to high levels that occurs at confluence could be almost completely suppressed by retinoic acid in the medium. The soluble form (type II) in the SCC-12B2 cells was induced in growing or confluent cultures by retinoic acid independent of the calcium concentration in the medium, but the 50% effective concentration (100 nM) for its stimulation was approximately 50-fold higher than the 50% effective concentration for suppression of the type I enzyme (2 nM). Thus, these enzymes appear to be distinct and independently regulated. This conclusion is supported by the finding that SCC-4 and SCC-9 almost exclusively expressed types II and I forms, respectively. In contrast to the results with neoplastic cells, in cultured normal epidermal cells type I enzyme comprised the overwhelming majority of activity and was only partially (75-90%) suppressible by retinoic acid, while type II enzyme seemed poorly if at all stimulable. Thus, the SCC lines appear appropriate for studying biochemical mechanisms of action of certain physiological agents, the molecular basis for altered regulation of differentiated function in neoplastic cells, and the origin of diversity within tumors.

Progressive state selection of cells in low serum promotes high density growth and neoplastic transformation in NIH 3T3 cells.

A subline of NIH 3T3 cells maintained by frequent passage (every 2 to 3 days) in 10% calf serum (CS) at low population density reached a low saturation density in 2% CS and produced no transformed foci on prolonged incubation at confluency in 2% CS. Within 3 frequent low density passages in 2% CS, the saturation density and focus-forming capacity in that serum concentration began an increase which was continued in subsequent passages. The saturation density and focus-forming capacities of the cells in both 2% and 1% CS were further enhanced by passage in 1% CS. The cells could then be passaged in 0.5% CS and then in 0.25% CS, which would support no multiplication of cells previously passaged only in 10% CS. The cells passaged in 0.25% CS gradually increased their saturation density and focus-forming capacity in that extremely low serum concentration during 24 low density passages, although their initial growth rate did not increase. They also attained a colony-forming efficiency in 0.25% CS of about 30%, as compared to less than 1% for cells passaged in 10% CS. Cells passaged, cloned, and passaged again in 2% CS yielded clonal populations which differed from one another in saturation density and focus-forming capacity in 2% CS. We conclude that NIH 3T3 cells diversify phenotypically at a high rate in their capacity to multiply and produce foci in limiting concentrations of serum, and we propose that progressive selection of these heterogeneous states accounts for the acquired capacity to function effectively in low concentrations of serum growth factors. Since lymph and presumably extracellular fluid in vivo contain low concentrations of growth factors which govern the multiplication of normal cells, the adaptation we observe in vitro may be related to tumor production in the animal.

Inhibition of phorbol ester-mediated interleukin-2 production by cellular differentiating agents.

Lymphocyte proliferation induced by the tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA) is inhibited by agents known to induce differentiation in murine erythroleukemia cells and other cell lines. In the present study, we determined the cellular targets for the action of TPA among murine thymocyte subpopulations, the phase of blastogenesis that is activated by the tumor promoter, and the phase that is inhibited by the differentiating agents. Mouse thymocytes were fractionated into populations bearing receptors for peanut agglutinin (PNA; PNA-positive cells) and populations lacking such receptors (PNA-negative cells). TPA is comitogenic for lectin-treated, unfractionated thymocytes and PNA-negative thymocytes but not for PNA-positive thymocytes. PNA-negative cells, a minor population in unfractionated thymocytes, are therefore the cellular targets for the comitogenic activity of TPA. TPA induces the production of interleukin-2 (IL-2) in lectin-treated PNA-negative populations but not in PNA-positive cells. The differentiating agents inhibit TPA-mediated proliferation of unfractionated and PNA-negative, lectin-treated thymocytes. In contrast, IL-2-mediated proliferation of lectin-treated thymocyte subpopulations is resistant to inhibition by these agents. Inhibition appears to be related to decreased production of IL-2, since the differentiating agents inhibit IL-2 production by both PNA-negative thymocytes and by a human leukemic cell line.

The insect brain (Na+ + K+)-ATPase. Binding of ouabain in the hawk moth, Manduca sexta.

(1) A quantitative study has been made of the binding of ouabain to the (Na+ + K+)-ATPase in homogenates prepared from brain tissue of the hawk moth, Manduca sexta. The results have been compared to those obtained in bovine brain microsomes. (2) The insect brain (Na+ + K+)-ATPase will bind ouabain either in the presence of Mg2+ and Pi, ('Mg2+, Pi' conditions) or in the presence of Na+, Mg2+, and an adenine nucleotide ('nucleotide' conditions) as is the case for the bovine brain (Na+ + K+)-ATPase. The binding conditions did not alter the total number of receptor sites measured at high ouabain concentrations in either tissue. (3) Potassium ion decreases the affinity (increases the KD) of ouabain to the M. sexta brain (Na+ + K+)-ATPase under both binding conditions. However, ouabain binding is more sensitive to K+ inhibition under the nucleotide conditions. In bovine brain ouabain binding is equally sensitive to K+ inhibition under both conditions. (4) The enzyme-ouabain complex has a rate of dissociation that is 10-fold faster in the M. sexta preparation than in the bovine brain preparation. Because of this, the M. sexta (Na+ + K+)-ATPase has a higher KD for ouabain binding and is less sensitive to inhibition by ouabain than the bovine brain enzyme. (5) This data supports the hypothesis that two different conformational states of the M. sexta (Na+ + K+)-ATPase can bind ouabain.

Collagen-induced platelet aggregation and release. I Effects of side-chain modifications and role of arginyl residues.

To investigate the mechanisms governing collagen interaction with blood platelets, the effects of side-chain modifications on collagen-induced platelet aggregation and release of serotonin were studied. Since many chemical modifications alter the ability of collagen to form fibers that, according to current theory, may complicate interpretation of data, we eliminated this possibility by using collagen stabilized in a native-type fibrillar structure by treatment with either glutaraldehyde or ultraviolet irradiation. Acetylation, methylation, succinylation, treatment with 2,4-dinitrofluorobenzene, 2,4,6-trinitrobenzene sulfonic acid or 1,2-cyclohexanedione, and deguanidination with hypobromite were used to modify collagen side-chain reactive groups: amino, carboxyl, hydroxyl and guanidino. Both unmodified monomeric dispersed and fibrillar collagen preparations initiated platelet aggregation and release, although the kinetics and magnitude of the response were different. Monomeric collagen which had been modified by deguanidination, methylation or succinylation, failed to polymerize in physiological conditions and did not induce platelet aggregation and release. However, none of the chemical modifications of stabilized native-type collagen fibers, except treatment with hypobromite or cyclohexanedione, had an effect on collagen-induced platelet aggregation and release. Both hypobromite and cyclohexanedione modified guanidino groups of arginyl residues. Results showed that the ability of a collagen sample to induce platelet aggregation and release of serotonin is dependent on the arginine content of fibrillar collagen. These data demonstrate that manipulation of amino, carboxyl and hydroxyl groups is unimportant as long as the native-type fibrillar structure is maintained, and that arginyl residues are directly involved in collagen-platelet interaction. Moreover, the data suggest that only the arginyl residues in the Y position of the tripeptide unit Gly-X-Y of collagen are responsible.

A phase II clinical trial of adoptive immunotherapy for advanced renal cell carcinoma using mitogen-activated autologous leukocytes and continuous infusion interleukin-2.

Forty patients with metastatic, recurrent, or unresectable renal cell carcinoma were entered into a study of the therapeutic efficacy of adoptive immunotherapy using periodate (IO4-) and interleukin-2 (IL2)-activated autologous leukocytes and continuous infusion low-dose IL2. Patient survival was also examined. The first 15 consecutive patients were enrolled in protocol A without an IL2 priming phase and the following 25 patients were entered in protocol B where a 5-day priming phase was initiated before leukapheresis. A maintenance regimen consisted of either 3 x 10(6) units of recombinant interferon-alpha (rIFN-alpha), three times per week only or together with leukapheresis and infusion of IO4-/IL2-activated cells and 2 days of continuous infusion IL2 per month. Thirty-four patients completed the protocol treatment. Four patients were removed from the study owing to rapid tumor progression and two patients died while receiving treatment. The clinical response rate was 22%: two patients had a complete response and five patients had a partial response. Among the 25 patients who had no clinical response, 11 patients had either a mixed response or stabilization. Neither response, response duration, nor site response correlated with the total dose of IL2 administered or the number of activated killer cells infused. Patients who received maintenance therapy had longer survival times than patients who did not receive such therapy. All toxicity and side effects associated with IL2 treatment were transient and resolved after discontinuation of the drug. Patients on maintenance therapy tolerated both rIFN-alpha and monthly infusions of activated killer cells and IL2 well. This study confirms the concept of adoptive immunotherapy as a new treatment approach for advanced renal cell carcinoma and suggests that maintenance therapy may prolong survival time.

Alpha 2-adrenergic receptor function in depression. The cortisol response to yohimbine.

There is evidence that the abnormalities in hypothalamic-pituitary-adrenal (HPA) axis function observed in patients with depression may be related to changes in central neurotransmitter receptor function. To evaluate this possibility further, the alpha 2-adrenergic receptor antagonist yohimbine hydrochloride, which increases brain norepinephrine turnover, was administered to 40 patients with DSM-III major depression (18 melancholic, 22 nonmelancholic) and 16 healthy controls. Plasma free 3-methoxy-4-hydroxyphenylglycol (MHPG) level was measured as an index of noradrenergic function, and plasma cortisol level was used to assess the HPA response. Baseline cortisol levels were elevated in melancholic depressed patients, but not in nonmelancholic patients, when compared with healthy controls. The cortisol response to yohimbine was significantly greater in depressed patients than in controls, despite similar MHPG responses between groups. Since there is evidence that stimulation of postsynaptic alpha 2-adrenergic receptors inhibits HPA axis function, the abnormally increased cortisol response to the alpha 2-antagonist yohimbine suggests a relative subsensitivity of postsynaptic alpha 2-adrenergic receptors in depression.

Endophthalmitis following staphylococcal sepsis in renal failure patients.

Endophthalmitis occurred three months following completion of therapy for documented staphylococcal septicemia in two patients on long-term hemodialysis. The indolent course of the endophthalmitis, and its excellent response to systemic and subconjunctival antibiotics and subconjunctival and topical corticosteroid therapy, suggest the possibility that the acute fulminating clinical course of metastatic bacterial endophthalmitis may be modified in this population of patients. The reason for this modified clinical picture is probably the immune incompetence associated with uremia, which favors both the development of metastatic endophthalmitis as well as altering its clinical presentation. While funduscopic examination is suggested in all dialysis patients with eye complaints, this procedure becomes mandatory following episodes of sepsis.

Studies of plasma aldosterone in anephric people: evidence for the fundamental role of the renin system in maintaining aldosterone secretion.

When plasma aldosterone (PA) was measured in 32 predialysis samples from 18 anephric subjects more than 3 months postnephrectomy, all but two values were subnormal, 16 measurements were undetectable, i.e. below 0.35 ng/100 ml, and PA averaged only 1.7 ng/100 ml +/- 0.5 (SE) in the remaining 16. In contrast, PA averaged 29.7 +/- 5.6 ng/100 ml in 12 dialysis patients with kidneys. Plasma renin activity was undetectable in 15 assays of anephric subjects and averaged 0.5 +/- 0.1 ng/ml/h in the remaining 17. However, these latter measurements appeared to be an artifact caused by the presence of plasma prorenin. Serum potassium fell during dialysis and appeared responsible for the concurrent 75% fall in PA observed in six nephric patients (delta 14 ng/100 ml), and in three anephric patients (delta 3.3 ng/100 ml) with low but measurable PA values. Overt hyperkalemia was required for potassium to stimulate aldosterone into the normal range in anephric subjects. The angiotensin II analog, saralasin, also increased PA slightly in five studies. Renin and PA fell after nephrectomy and increased after transplantation. Decreases and increases in renin always preceded those of aldosterone. Usually it took several weeks for aldosterone to fall to undetectable levels and several days to return to normal levels after renal transplantation. These observations suggest that the baseline plasma level of angiotensin determines both the baseline level of aldosterone secretion and its capacity to respond either to more angiotensin or to other stimuli. In the absence of renin, the adrenal almost loses its capacity to generate aldosterone. The data support the view that the renin system plays a fundamental role in maintaining aldosterone secretion in man.

Keratinocyte differentiation markers: involucrin, transglutaminase, and toxicity.

Studies of three keratinocyte differentiation markers are described. First, the involucrins of several mammals are identified, facilitating use of this marker in animal models of human disease. The rapid evolution of involucrin has prevented its routine immunochemical identification beyond the primates, but its unusual solubility and its labeling by transglutaminase have permitted detection in rats, cats, and sheep. Second, the re-expression of keratinocyte transglutaminase in carcinoma cells lacking the enzyme is demonstrated. Lack of this enzyme expression has been observed previously in squamous cell carcinomas. The present finding suggests genomic hypermethylation could contribute to this phenomenon and offers an approach to analyzing transcriptional features of the enzyme regulation. Third, the sensitivity of keratinocytes to growth suppression by aflatoxin B1 is reported. The observed toxicity appears to be mediated by aryl hydrocarbon hydroxylase, a metabolic enzyme inducible in keratinocytes by environmental agents. Such expression may be relevant to carcinogenesis in tissues subject to squamous metaplasia as well as in other exposed cell types stimulated to express this biotransformation enzyme.

Pharmacokinetics of essential amino acids in chronic dialysis-patients.

Possible disorders of essential amino acid (EAA) metabolism in maintenance dialysis patients (D) were studied by measuring plasma amino acids before and sequentially after administering a mixture of 8 EAA po and iv. The EAA were in a ratio similar to that required for optimal utilization, and the total dose given was within physiological range. Ten D and six normals (N) received 150 mg/kg po as a 10.5 g/dl solution and 117 mg/kg iv as a 5.1 g/dl solution infused at a constant rate of mg/kg per min. Blood glucose and immunoreactive insulin were also measured. Both D and N were postabsorptive and at least 18 hr postdialysis. The fraction of the oral dose appearing in the systemic circulation was variable for each EAA in both N and D. Total body clearance was significantly lower in D (P less than 0.05) for theronine, phenylalanine, valine, leucine, and isoleucine, and this difference could not be explained by changes in renal excretion. The apparent volume of distribution did not differ between N and D for all EAA except for valine and phenylalanine. Blood glucose insulin varied only slightly in both N and D for all EAA except for valine and phenylalanine. Blood glucose and insulin varied only slightly in both N and D. These studies indicate that there are a variety of significant abnormalities in the metabolism of specific EAA in D. Decreased total body clearance of the branched-chain amino acids, since they are primarily metabolized total body clearance of the brnached-chain amino acids, since they are primarily metabolized by muscle, may result from a defect in muscle metabolism in D.

Decreased macrophage-mediated suppression of lymphocyte activation in chronic renal failure.

Prostaglandin-dependent adherent cell suppressor activity was assessed in patients with end-stage renal insufficiency. Proliferative responses of uremic peripheral blood mononuclear cells to optimal concentrations of phytohemagglutinin and concanavalin A were impaired. Responses to the galactosyl-directed lectins, soybean agglutinin and peanut agglutinin, were, however, normal or supranormal. The addition of 1 microgram/ml of indomethacin, to cell cultures resulted in relatively less potentiation of blastogenic responses to the galactosyl-directed lectins in cells from uremic patients (soybean agglutinin, p less than 0.02; peanut agglutinin, p less than 0.05). Similarly, depletion of adherent cells markedly enhanced blastogenesis induced by the galactosyl-directed lectins in normal cell cultures, whereas the effect was much less pronounced (soybean agglutinin, p less than 0.02; peanut agglutinin, p less than 0.02) in uremic cells. Reduced activity of the adherent cell suppressor system in patients with renal failure might be associated with altered sensitivity of uremic lymphocytes to soluble mediators of suppression. The lymphocytes of uremic patients, depleted of adherent cells, were relatively resistant to the inhibitory action of prostaglandin E1 (0.001 microgram/ml, p less than 0.05, and 0.01 microgram/ml, p less than 0.02) on galactosyl-directed, lectin-induced mitogenesis. In contrast, dibutyryl cyclic AMP (10(-4) M), 8-bromo cyclic AMP (10(-5) M), and 3-isobutyl-1-methyl xanthine (20 micrograms/ml) inhibited both control subject and patient cultures to the same extent. Prostaglandin E1 in combination with methyl isobutyl xanthine produced, in adherent-cell-depleted control subjects, levels of cyclic AMP that were significantly higher than in cells from uremic patients (p less than 0.05). Thus, depressed adherent cell suppressor activity in patients with renal failure may result in part from impaired generation of cyclic AMP by lymphocytes.