concanavalin-a and Scleroderma--Systemic

Systemic sclerosis is an autoimmune disease that is associated with excessive fibroblast proliferation and collagen deposition in various tissues. Interleukin-6 (IL-6) is produced by fibroblasts, activated T and B lymphocytes, which maybe involved in the pathogenesis of systemic sclerosis.. This study was performed in order to determine whether IL-6 could be detected specifically in collagen-stimulated peripheral blood mononuclear cells from patients with systemic sclerosis.. We clinically evaluated seven patients with systemic sclerosis for disease duration and organ involvement and analyzed in vitro the ability of their peripheral blood mononuclear cells and those of disease-free controls, in the presence of concanavalin A, human type I collagen, and the mast cell mediator, heparin to secrete IL-6 spontaneously by a sensitive ELISA.. Interleukin-6 production by nonspecific stimulation with concanavalin A did not differ between patients with systemic sclerosis and controls; however, collagen stimulation significantly increased IL-6 production in patients with systemic sclerosis; mean 1728 pg/mL versus a mean of 386 pg/mL in controls P = < .05). Collagen-stimulated IL-6 levels > 2000 pg/mL were obtained in 86% of patients with systemic sclerosis compared with none in the controls. In patients with systemic sclerosis with a shorter disease duration, greater spontaneous as well as collagen- and heparin-stimulated IL-6 production was observed, whereas decreased IL-6 levels were noted with longer disease duration (> 21 years).. The results of this study suggest that peripheral blood mononuclear cells from patients with systemic sclerosis are specifically sensitized to human type I collagen to produce increased levels of IL-6, which may play a role in the pathogenesis in this fibrotic disorder.

Topics: Adult; Aged; Cells, Cultured; Collagen; Concanavalin A; Female; Humans; Interleukin-6; Leukocytes, Mononuclear; Male; Middle Aged; Scleroderma, Systemic

We previously reported on an altered immune-endocrine feedback loop via the hypothalamo-pituitary-adrenal (HPA) axis in Obese strain (OS) chickens afflicted with spontaneous autoimmune thyroiditis. These animals are deficient in plasma corticosterone increase after antigenic challenge or application of cytokine-containing conditioned medium of mitogen-stimulated spleen cells (CM). To investigate whether the impaired ability to respond to cytokines with glucocorticoid-increasing factor (GIF) activity, e.g. interleukin 1 (IL 1), is restricted to OS chickens as a model for an organ-specific autoimmune disease, we extended our experiments to another autoimmune-prone animal strain, the chickens of the University of California at Davis line 200 (UCD-200). These animals develop an inherited inflammatory fibrotic disease that closely resembles human progressive systemic sclerosis (scleroderma). Application of GIF-containing CM to UCD-200 chickens leads to a transient increase in glucocorticoid serum levels within 1-2 hours comparable to that of controls. But, while corticosterone levels in the latter returned to normal baseline levels after 4 hours, they were still elevated in autoimmune chickens. Although the peak of the glucocorticoid hormone serum concentrations was equal to that of controls, UCD-200 had to secrete twice as much adrenocorticotropic hormone to achieve this corticosterone serum level due to an apparent hyporesponsiveness of the adrenal gland to this secretagogue. The altered cytokine-induced glucocorticoid secretion is found in early as well as in chronic, sclerotic stages of the disease. Cellular alterations in the peripheral blood of UCD-200 chickens during the prolonged elevated corticosterone section, i.e. between 2-4 hours after CM application, are characterized by a significant decrease in the percentage of CD4+ and CD8+ cells. Furthermore, a significant increase in B cells up to 24 hours with a maximum after 1 hour was found. The proliferative response to the mitogen concanavalin A of peripheral mononuclear cells was inversely correlated to the serum corticosterone level, showing a permanent decrease of 80-90% after 1-4 hours in autoimmune animals. This functional alteration in UCD-200 was accompanied by an 80% decrease in serum interleukin 2 (sIL 2) activity 4 hours after CM application. Twenty-four hours later an eight-fold increase in sIL 2 rebound activity was found, indicating that the inhibitory effect of corticosterone in UCD-200 c

Topics: Adrenocorticotropic Hormone; Animals; Autoimmune Diseases; Biological Factors; Cells, Cultured; Chickens; Concanavalin A; Connective Tissue Diseases; Corticosterone; Culture Media, Conditioned; Disease Models, Animal; Feedback; Fibrosis; Immunologic Factors; Interleukin-2; Leukocyte Count; Lymphocyte Activation; Lymphocyte Subsets; Pituitary-Adrenal System; Scleroderma, Systemic; Spleen

Interleukin-1 (IL-1) production by peripheral blood mononuclear cells (PBMC) from patients with scleroderma and healthy controls was studied. Supernatants from unstimulated PBMC cultures from 10 of 13 patients with progressive systemic sclerosis (PSS) had significantly less IL-1 activity as measured by thymocyte proliferation than controls. IL-1 activity per monocyte/macrophage in both patients and controls was 10 times greater when PBMC were cultured at 10(5) cells/ml compared to 10(6) cells/ml. Five-fold dilution of supernatants from PBMC cultured at 10(6) cells/ml revealed more IL-1 activity than undiluted supernatant and addition of indomethacin increased IL-1 activity primarily of the undiluted supernatant. The results show that IL-1 activity from crude PBMC supernatants from PSS patients is low and may be regulated by non-dialysable inhibitors produced by PBMC and/or cell interactions.

Topics: Blood Cell Count; Cells, Cultured; Concanavalin A; Humans; Indomethacin; Interleukin-1; Interleukin-2; Leukocytes; Mitosis; Scleroderma, Systemic; T-Lymphocytes

Dermal fibroblast cultures from patients with progressive systemic sclerosis (PSS) synthesize up to 5 times more glycosaminoglycan (GAG) than normal cultures. In an in vitro model of fibroblast-lymphocyte interactions, we show that the supernatants of activated mononuclear cells (MNC) modulate GAG synthesis, as measured by the incorporation of 3H-glucosamine into GAG following incubation of the confluent fibroblast monolayers with active supernatant preparations. GAG accumulation was selectively increased up to 18 times in normal dermal fibroblast cultures. Cell viability was not affected, and 3H-thymidine uptake and cell numbers were depressed in cultures treated with the supernatants. In contrast to normal dermal fibroblast cultures, PSS fibroblasts responded to MNC supernatants by only a 1-2-fold increase in GAG. Supernatants of concanavalin A-activated PSS MNC had higher stimulatory activity than those of normal MNC. Supernatants made with MNC that had been depleted of monocytes on Sephadex G-10 columns were only minimally stimulatory. The GAG-stimulatory supernatants modulated the synthesis, but not the degradation of GAG. Gel filtration on a calibrated Sephadex G-100 column indicated the presence of stimulatory activity in both the 50,000 and 15,000 molecular weight fractions. These activities were trypsin-sensitive, but had different susceptibilities to heat. The active column fractions also contained interleukin-1 activity, as shown in an assay measuring proliferation of mouse thymocytes. Like our factors, interleukin-1 preparations increased GAG in normal and PSS dermal fibroblasts. Products of activated MNC may modulate normal and pathologic processes in human skin.

Topics: Cells, Cultured; Concanavalin A; Fibroblasts; Glycosaminoglycans; Humans; Lymphocyte Activation; Lymphokines; Monokines; Proteins; Scleroderma, Systemic; Skin

Supernatants of human mononuclear cells activated with concanavalin A contain soluble factors that modulate accumulation of collagen in human dermal fibroblasts in culture. The supernatants decreased collagen accumulation by 67% in normal fibroblast lines and by greater than 80% in lines established from skin of patients with scleroderma. Collagen was measured by incorporation of 3H-proline into collagenase-sensitive protein. The inhibitory activity of collagen was optimal after a 24-hour incubation of confluent fibroblast monolayers with unfractionated mononuclear cell supernatants. Kinetic studies of this response showed a delay in accumulation of collagen-sensitive protein in supernatant-treated cultures. The 3H-thymidine uptake and fibroblast cell count and viability were only minimally altered. Noncollagen protein was inhibited by 30% to 40%. The presence of serum in fibroblast cultures did not affect this activity. The inhibitory factors were produced by purified T-lymphocyte subpopulations. Supernatant inhibitory activity was present after removal of monocytes from mononuclear cell cultures. By gel filtration on Sephadex G-100, active supernatants fractionated into at least three peaks of activity with molecular weight of 50,000, 30,000, and 15,000 daltons. Interleukin 1 might be one of the factors in mononuclear cell supernatants that modulates production of collagenase-sensitive protein in human dermal fibroblasts. Factors modulating collagen-sensitive protein are important in processes leading to excessive accumulation of the connective tissue and fibrosis in certain diseases.

Topics: Cells, Cultured; Collagen; Concanavalin A; Fibroblasts; Humans; Interleukin-1; Kinetics; Microbial Collagenase; Monocytes; Scleroderma, Systemic; Skin

Unstimulated mononuclear cell supernatants from patients with systemic scleroderma significantly augmented the DNA and collagen biosynthesis of 3T3 cells compared to normal mononuclear cells. This increase was suppressed by either a particular glycosaminoglycan, which from previous studies, has appeared to act as one of the tissue antigens in sclerotic skin, or concanavalin A stimulation.

Topics: Adult; Aged; Cells, Cultured; Collagen; Concanavalin A; DNA; Female; Fibroblasts; Glycosaminoglycans; Humans; Lymphocytes; Male; Middle Aged; Mitogens; Scleroderma, Systemic

The primary in-vitro antibody response developed by peripheral blood mononuclear cells (PBM) towards trinitrophenyl coupled to polyacrylamide beads (TNP-PAA) was evaluated in 17 untreated patients with progressive systemic sclerosis (PSS). This response was markedly depressed as compared with that of 19 control patients and 28 normal subjects. In eight PSS patients and eight normal controls the anti-TNP response was measured before, and after, a PBM filtration on nylon wool columns. This procedure dramatically reduced the proportion of monocytes identified as mononuclear cells staining positively for peroxidases, and restored the response of PSS PBM to the level observed in normal PBM. In four experiments, plastic-adherent cells from either normal subjects of PSS patients were added to autologous nylon-passed PBM. This did not modify the response from normal PBM but inhibited the response of PSS PBM. The inhibitory effect of PSS plastic-adherent cells was insensitive to a 2,000 R X-ray irradiation. These results strongly suggest that the impaired in-vitro antibody response observed in PSS can be attributed to a suppressor monocyte. The concanavalin-A-induced suppressor cells of the antibody response were assayed in PSS. They exerted a suppressive effect to the same extent as in controls.

Topics: Adolescent; Adult; Antibody Formation; B-Lymphocytes; Cell Adhesion; Cell Separation; Concanavalin A; Female; Hemolytic Plaque Technique; Humans; Male; Middle Aged; Monocytes; Nylons; Plastics; Scleroderma, Systemic; T-Lymphocytes, Regulatory; Trinitrobenzenes

The study of T cell subpopulations and their immunoregulatory circuits in 9 patients with progressive systemic sclerosis (PSS) and 9 age/sex matched controls showed: 1. Normal postthymic precursor autologous rosette-forming T cells (Tar cells). 2. Normal T cells with receptors for the Fc portion of IgG(Ty). 3. Decreased T cells with receptor for Fc portion of IgM(Tmu). 4. Normal function of postthymic precursors. 5. Normal Concanavalin-A-induced and spontaneously-expanded suppressor cell functions. 6. Abnormally increased T helper cell function. These findings suggest that the primary immunoregulatory aberration in PSS is at the level of Tmu cells and their helper function. Relationship between this T cell disturbance and fibroblast function may explain the pathogenesis of PSS.

Topics: B-Lymphocytes; Cell Differentiation; Concanavalin A; Immunoglobulin M; Leukocyte Count; Lymphocyte Cooperation; Receptors, Fc; Rosette Formation; Scleroderma, Systemic; T-Lymphocytes; T-Lymphocytes, Regulatory

T lymphocyte subpopulations were studied in 40 patients with scleroderma (PSS), 26 of whom were studied simultaneously for lymphoproliferative responses to phytohaemagglutinin (PHA), concanavalin A (Con A) and pokeweed mitogen (PWM). PSS patients exhibited a reduction relative to 42 age- and sex-matched controls in the absolute number and percentage of early E rosettes, late E rosettes and E rosettes formed with aminoethylisothiouronium bromide (AET) treated sheep red blood cells. There was no difference between patients and controls in the proportions of B lymphocytes. PSS patients exhibited normal lymphocyte transformation responses to PHA and ConA and an augmented response to PWM. The mitogen responses did not correlate with the absolute number or percentage of lymphocytes or T and B lymphocyte subpopulations. No correlation was observed between any immunological variable studied and the extent of skin or organ involvement, disease duration or therapy.

Topics: Adult; Aged; B-Lymphocytes; Concanavalin A; Female; Humans; Immunoglobulin G; In Vitro Techniques; Lymphocyte Activation; Male; Middle Aged; Mitogens; Phytohemagglutinins; Pokeweed Mitogens; Rosette Formation; Scleroderma, Systemic; T-Lymphocytes