The effect of cyclosporine on ornithine decarboxylase induction with mitogens, antigens, and lymphokines.

Ornithine decarboxylase (ODC) is the initial enzyme in polyamine synthesis. An increase in ODC activity is associated with increased RNA, DNA, and protein synthesis. We have used the induction of ODC by mitogens and alloantigens in human peripheral blood lymphocytes as an intracellular marker of protein synthesis and lymphocyte activation. The immunosuppressive agent cyclosporine was found to inhibit both the mitogen and alloantigen stimulated induction of ODC in lymphocytes in a manner that parallels inhibition of subsequent 3H-thymidine incorporation. When purified T lymphocytes were stimulated with mitogen alone, minimal ODC activity was detected. The addition of 5% monocytes, human Interleukin-1 (IL-1), or T cell growth factor (IL-2) enhanced mitogen-induced ODC activity in T lymphocytes 4-10-fold. Cyclosporine inhibited the induction of ODC when T lymphocytes were combined with monocytes or growth factors. We conclude that (1) the induction of ODC in human lymphocytes by mitogen and alloantigen is inhibited in the presence of cyclosporine; (2) the induction of ODC activity in purified T lymphocytes requires the presence of both mitogen and monocytes or their products; (3) IL-1 and IL-2 can supplement for monocytes and augment the phytohemagglutinin induction of ODC in T lymphocytes; and (4) cyclosporine inhibits ODC induction in T lymphocytes stimulated with mitogen in the added presence of monocytes, IL-1, or IL-2. The inhibition of ODC induction and polyamine synthesis by cyclosporine adds insight into its mode of action on the mechanisms involved in early T cell activation.

The role of mitogens and lymphokines in the induction of ornithine decarboxylase (ODC) in T lymphocytes.

Polyamine synthesis occurs early in lymphocyte activation after stimulation with antigen or mitogen. Ornithine decarboxylase (ODC) is the primary enzyme in the polyamine cascade. We have examined the induction of ODC by mitogens and/or lymphokines in human peripheral blood T lymphocytes. When isolated populations of monocytes and T lymphocytes were stimulated with phytohemagglutinin (PHA) there was little or no change in ODC activity. The combination of T lymphocytes and monocytes enhanced mitogen-induced ODC activity 10-fold. Several interleukin 1 (IL 1)-containing supernatants and fractionated human IL 1 were capable of substituting for monocytes in supporting PHA induction of ODC in T lymphocytes. Interleukin 2 (IL 2) and IL 2-containing supernatants were also capable of increasing ODC activity in T lymphocytes in the absence of monocytes. Lymphokines alone in the absence of PHA could not induce ODC. We conclude that both mitogens and monocytes are required for the induction of polyamine synthesis in T lymphocytes, and that supernatants containing IL 1 or IL 1 and IL 2 can substitute for monocytes in the induction of ODC in mitogen-stimulated T lymphocytes.

Changes in the compositions of the null cell compartment with murine autoimmunity.

The subpopulations that comprise the null cell compartment were examined sequentially in various strains of autoimmune-prone mice. Different patterns emerged that were consistent within strains but differed from strain to strain. Abnormalities appear earlier in life in short-lived mice, such as male BXSB and MRL/l mice, than in relatively long-lived strains, such as female BXSB and NZB mice. The accumulation of T cells in MRL/l mice was accompanied by null cell changes that contrasted with those that developed in AKR/J mice after their spleens were infiltrated with leukemic T cells. It would seem that lymphocyte perturbations with murine autoimmunity also involve their precursor cells and that these precursor cell changes vary in different strains, perhaps in relation to different genetic factors.

A comparison of the regulatory effects of human monocytes, pulmonary alveolar macrophages (PAMs) and spleen macrophages upon lymphocyte responses.

Human monocytes, pulmonary alveolar macrophages (PAMs) and spleen macrophages were concentrated by immobilization on cold insoluble globulins. These cell preparations were 90 +/- 3%, 95 +/- 1% and 83 +/- 3% esterase rich, respectively, 87 +/- 4%, 95 +/- 3% and 66 +/- 11% phagocytic and 78 +/- 3%, 79 +/- 9% and 68 +/- 5% reactive with OKM1 monoclonal antibody. Spleen macrophages differed from the other two cell preparations in that significantly fewer reacted with 61D3 or 63D2 monoclonal antibodies. Monocytes and PAMs promoted the mixed leucocyte response by autologous lymphocytes when added at low concentrations, but suppressed this response at high concentrations. Spleen macrophages only promoted the mixed leucocyte reaction but were required in much higher numbers than either monocytes or PAMs for optimal promotion. Likewise, the added presence of monocytes or PAMs in high numbers suppressed Ig synthesis stimulated with pokeweed mitogen, while spleen macrophages were not suppressive in this system. This study shows that the distribution of macrophages that differ in their regulatory effects upon lymphocyte responses varies in different tissues. The human spleen is deficient in macrophage related suppression.

Circulating immune complexes and tumor cell cytotoxins as prognostic indicators in malignant melanoma: a prospective study of 53 patients.

To evaluate the relationship between tumor burden and circulating immune complexes (IC) in malignant melanoma, we tested sera collected serially from 15 normal donors and 53 patients. Forty-eight of these had Stage III or IV disease at the outset of the study. The median survival time (MST) of ten patients with Stage IV disease whose sera contained C1q-binding IC at the outset of the study was 4.7 months; the MST of the 25 Stage IV patients whose sera were initially free of IC by this test was 8.65 months (p less than 0.02). C1q-binding IC were not found in the initial serum samples from 13 patients with Stage III or 5 patients with Stage I disease. Abnormal C1q binding tests were measured in 4 of 67 sera (6%) from 13 patients who remained free of evident tumor for up to 41 months. IC were detected in 13 of 39 sera (33%) from 19 patients with progressively growing tumors and in 21 of 68 sera (31%) from 21 patients who were initially free of disease but developed recurrences later, or who had significant remissions of variable duration during follow-up. The MST of 31 patients whose serial serum samples remained free of C1q-binding IC was 15.8 months. Twelve patients whose sera were initially free of circulating IC later developed abnormal serum C1q-binding levels. Their MST was 10.3 months. The MST of ten patients with persistently abnormal serum IC levels was 4.7 months. C1q-binding IC were reciprocally related to the presence of complement-dependent antibodies, cytotoxic for cultured allogeneic malignant melanoma cells in sera from 29 of these patients (r = -0.491;p = 0.003). These results suggest that the appearance of circulating C1q-binding IC is pathophysiologically important in malignant melanoma. Measurement of C1q-binding IC may be useful in assigning prognosis in this disease.

Suppression of lymphocyte responses by monocytes with untreated and treated multiple myeloma.

Studies were performed on 15 untreated and 14 treated patients with multiple myeloma. The monocyte content was normal in blood but elevated in mononuclear leukocytes (MNL) from treated but not untreated patients (p less than 0.001). This correlated with the severity of lymphopenia in blood (p less than 0.01). Three patterns of immunoglobulin(Ig) synthesis emerged. (1) Most untreated patients showed normal polyclonal responses to pokeweek mitogen. (2) Of 12 treated patients, the 8 whose MNL included greater than 30% monocytes had subnormal Ig responses to pokeweek mitogen. Ig synthesis increased when adherent cells that suppressed Ig synthesis were depleted. Suppression in vitro bore no relationship to polyclonal immunoglobulin levels in serum. (3) Three patients had early blood invasion by plasmacytoid cells. Their MNL spontaneously released large amounts of the Ig class of their serum gammopathies. Proliferative responses to phytohemagglutinin by MNL from all patients were reduced, in part due to monocytoid cell suppression and in part to intrinsic T-cell hyporesponsiveness. B- and T-cell responses in vitro are sometimes suppressed with myeloma. This is related to elevated monocyte percentages in MNL preparations. This excess of monocytes is a function of lymphopenia secondary to therapy, rather than the primary malignant process itself. No evidence was found that suppression by monocytes is qualitatively altered by myeloma or its treatment.

Null cell senescence and its potential significance to the immunobiology of aging.

The null cell compartments of human bone marrow and mouse spleen were arbitrarily divided into three subpopulations based upon the ability of cells to acquire T or B cell membrane markers when incubated with poly A:U or ubiquitin. There was an accumulation of T cell precursors with congenital absence of the thymus. In contrast, T cell precursors were reduced and there was an accumulation of uninduced null cells with old age. These observations suggest that there is an intrinsic defect of null cell differentiation with a drift towards more differentiated precursors in T cell differentiation with aging. This could result in a diminution in the range of responses by their progeny, mature T lymphocytes.

Angioimmunoblastic lymphadenopathy with hypogammaglobulinemia. Possible role of monocyte suppression.

A patient wit angioimmunoblastic lymphadenopathy had low serum immunoglobulin values and no antibodies to injected immunogens. This occurred despite the proliferation of polyclonal B cells. T cells were deficient in number and in lymphoproliferative responses, but their helper and suppressor functions were maintained. Ia-antigen bearing leukocytes from the patient stimulated poorly in mixed leukocyte culture. In vitro immunoglobulin synthesis by mononuclear leukocytes form the patient was severely impaired. These leukocytes actively suppressed immunoglobulin synthesis by normal cells from healthy subjects in co-culture. The responsible cell had characteristics of a monocyte. The suppression was selective for humoral immunity and was manifest despite normal numbers of monocytes. It appears that heterogeneous immunoregulatory abnormalities can underlie the syndrome of angioimmunoblastic lymphadenopathy. Furthermore, monocyte suppressor abnormalities may be implicated in clinical disease phenomena.

Spectrum of immunodeficiencies with Hodgkin's disease.

The role of six suppressor mechanisms upon T and B cell responses was studied on 17 untreated patients with Hodgkin's disease. Proliferative hyporesponsiveness to mitogen was greatly impaired in 8 of the 13 patients. 10 of these patients had an excessive degree of suppression by cells that adhered to foreign surfaces. Suppression by adherent cells correlated with impairment of proliferative responses and, in some instances, suppression was largely inhibited with indomethacin. Likewise, adherent cells suppressed immunoglobulin synthesis. A correlation was evident between suppression of T and B cell responses by adherent mononuclear leukocytes from individual patients. This suppression coincided with elevated percentages of monocytes in the patient mononuclear cell preparations. This excess of monocytes was not the result of a circulating monocytosis. The monocyte excess may have been acquired during isopyknic cell separation. A second form of suppression was observed in 5 of the 11 patients affected by a lymphocyte that neither adhered to glass wool nor required preactivation. It did not inhibit allogeneic lymphocytes, which contrasts with the suppressor abnormality of monocytoid cells.

An inhibitor of thymic hormone activity in serum from patients with lymphoblastic leukemia.

Serum from 21 patients with lymphoblastic leukemia, five with myeloblastic leukemia and 30 age matched control subjects tested for thymic hormone activity in an assay that measures the induction of T cell surface antigen. This activity was subnormal in serum from 10 of 16 patients with untreated lymphoblastic leukemia (p less than 0.001) but was within the normal range when the leukemia was in remission. Low inductive activity was associated with an inhibitor of T cell induction which was less than 30,000 daltons in molecular size and interfered with induction by purified thymopoietin plus a high concentration of ubiquitin or by normal serum alone.

A novel lymphocyte differentiating factor in serum of patients with mycosis fungoides and Sezary syndrome.

Sera from 13 patients with mycosis fungoides and 2 with Sezary syndrome were tested for activity that induces lymphocyte differentiation. Induction of Thy-1.2 antigen and surface immunoglobulin were used, respectively, to measure T- and B-cell differentiation. The indicator cells were null lymphocytes from the spleens of congenitally athymic nude mice. Normal serum induced some T-cell but no B-cell differentiation. The T-cell-inducing activity was ascribed to thymic hormone and declined with advancing age. A totally different pattern emerged with patient serum. T-cell-inducing activity was significantly more active than in normal serum (p less than 0.001). This activity did not decline with advancing age and was not inhibited by a concentration of ubiquitin, which blocks nonspecific beta-adrenergic induction. B-cell-inducing activity was also present. This novel serum factor (or factors) is a potent inducer of T- and B-lymphocyte differentiation and is associated with neoplastic lymphoproliferation of the T-cell series.

Myasthenia gravis, thymectomy and serum thymic hormone activity.

Serum thymic hormone activity was measured in 36 patients with myasthenia gravis and in 10 control subjects from each age decade. In all 25 patients under 50 years of age results were within, or close to, the normal range. Activity at levels considered normal for juveniles was detected in 10 of the 11 older patients whereas levels normally decline in older subjects. One week after thymectomy, 13 of 17 patients (76 per cent) had no demonstrable serum thymic hormone activity. However, 10 months or longer after thymectomy only five patients (30 per cent) lacked thymic hormone activity in the serum. There was a significant correlation between clinial improvement and sustained lowering of serum thymic hormone activity after thymectomy.