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88 Binding of serum amyloid P component (SAP) to immobilized DNA: A study in patients with systemic lupus erythematosus
160
85
86
Abstracts
ANALYSIS AND CLINICAL CORRELATION OF SERUM INTERLEUKIN-4 LEVELS IN PROGRESSIVE SYSTEMIC Paul S. Lindner, MD; Marianne Frieri, SCLEROSIS. MD., PhD., East Meadow, New York Progressive systemic sclerosis (PSS) is a multisystemic disorder characterized by widespread fibrosis and degenerative changes of the skin, synovium, parenchyma and blood vessels. The etiology of PSS remains unknown. Cellular and humoral abnormalities have been described. Serum interleukin (IL)-2 levels and IL-2 receptors have recently been shown to be elevated in PSS. There are increased numbers of mast ccl Is in the papillary and reticular dermis, with enhanced activation in rapidly progressive disease. IL-4 is known to stimulate the growth of mucosal and connective tissue mast cells and fibroblasts. We analyzed serum IL-4 in patients with PSS to determine if it plays a role in this disease. Serum from 15 patients with PSS, 1 with morphea, 1 with sclerodactyly and 17 disease free controls was obtained and assayed for IL-4 by a highly sensitive ELISA using monoclonal antibodies. Clinical data on symptoms, disease interval, systemic involvement and a detailed skin score was obtained. There was a statistically significant correlation between IL-4 levels and skin score (r=.68, ~~005). An inverse correlation of IL-4 levels and length of disease was shown which did not reach statistical significance (r=.54). Patients with PSS were found to have a statistically significant elevation of serum IL-4 level compared to the controls (520.3~3.2 vs 10.8f1.5ng/ml, ~~01). The results of our study suggest a possible role of IL-4 in the pathogenesis of PSS, acting through its effects on mast cells, fibroblasts and T-cells.
MAST CELL EFFECTS ON CYTOKINE PRODUCTION IN SCLERODERMA PATIENTS. Marianne Friqri, Ph.D..M.D. Kushal Agarwal, M.D.,East Meadow New York Mast cell hyperplasia has bee; demonstrated in the skin and other organs in scleroderma (PSS) and irmunoregulatory cellular IL-2 defects have been reported (Frieri, M. et al. J Allergy Clin lmmunol 81:196a, 1988; Manuscript accepted by Arthritis Rheum, 1990). IL-4 can increase IgE production and stimulate mast cells and fibroblasts. Many of the effects of IL-4 are downregulated by TIFN. We evaluated 17 PSS patients and controls using systemic and skin scores along with (3H) Tdr IL-2 driven T ccl 1 pro1 iferation (TCP) with 10 ug/ml: of Con A, human type I collagen and heparin: 10'6 M histamine with antiIL-2/IL-4 monoclonal antibodies (MoAb) . A sensitive MoAb Elisa was used to determine‘OIFN levels in stimulated PBMC supernatants. Histamine and heparin along with collagen increased T cell blastogenesis in 11-16 patients at various stages of their disease compared to normals (!? SI in PSS = 2.3 + 0.28, 2.2 * 0.29, 1.8 t 0.2 vs K SI in normals = 0.5 f 0.005, 0.9 *O.OOl, 0.8 iO.01). Anti-IL-Z/IL-4 Moab decreased responses in 50% of patients. In contrast,fIFN levels in PBMC from 7 patients stimulated by histamine were significantly decreased (1.56 ko.26 ng/ml)vs 10 normals (3.10 fO.60 rig/ml), p =a1.05. The role of defective cytokine generation via IL-2, IL-4 andXlFN may contribute to significant advances in the diagnosis and treatment for patients with scleroderma.
87 Systemic lupus -e&hematosus ISLE) :-: associated with-autoant%bodies which- t&get 1 limited and oredictable set of antieens and these may p^artly be defined by 'bistinct structural properties. A rigorous statisticai method was used to analyze the sequenced SLEassociated autoantigens (n=33) far chargcrelated amino acid patterns which might be itmnunologically important. method The distinguishes significant long run 5 , local concentrations (clusters),and patterns of charge (every second or third residue). Significant
mixed charge runs often within clusters wer.? present in 12 (45.5%), and mixed charge cluster9
in another
(27%).
Long patterns
were-present
ix
Significant ruw andior another 2 antigens. clusters were present in i6% of a control set trf proteins. The mean % charge of the antigens wai; 32 vs 26 for controls (pX.001) suggesting that.
lacking locally high charge concentrated charge might be Siwificant folded uroteins. charge runs are present' in other co&xtivetissue disease autoantigens notably CENP-R and Topisomerase T of scleroderma, but are not. in
antigens
configurations
annarent
in the
prevalent in autoantigens
configurations
set a (Z/11). are
of
prevalent
organ-specific
Simila:charge and in viral
cellular transcription factors and surface proteins. Highly charged surfaces may render these viraland h&t proteins -strongly
inxnunoeenic and antinuclear antibodies resoonses to might - result from chronic exposur'e intracellular contents, possibly in conjunction with cross-reactive viral proteins.
88
BINDING
OF SERUM AMYLOID
(SAP) TO TIENTS Arturo
M.D., SAP
IM%lOBILIZED
DNA:
P COMPONENT A STUDY IN PA-
WITH SYSTEMIC LUPUS ERYTlJP!MATOSUS. M.D. and Anita Gewurz, 3. Bonnin, Chicaso, Illinois
is a-nbrmal human serum (40 + 10 and tissue glycoprotein, also Like C-reacfound in amyloid deposits. tive protein with which it is closely homologous, SAP binds to chromatin and nucleosomes with high affinity and is the major DNA binding protein in normal serum, yet its function(s) and role in disIn order to evaluate ease are unknown. DNA binding by SAP in patients with SLE and high titers of anti-DNA antibodies, we compared the quantity and DNA binding activity of SAP from sera of SLE patients with serum and purified SAP from normal donors. Protein levels of SAP were determined by electroimmunoassay using rabBinding of SAP to immobibit antiserum. lized DNA was performed by ELISA using either serum or purified SAP and a monoclonal antibody to detect SAP. The binding of purified SAP to DNA was calcium-dependent and constant over a wide pH range. SAP protein levels in SLE patient and normal sera were compasable. the binding to immobilized DNA However, by SAP in patient sera was reduced, compared to SAP in normal serum or purified SAP. Our data suggest that decreased SAP binding to immobilized DNA may be a factor associated with elevation of antiDNA antibodies in SLE patient sera. ug/ml)
85
86
Abstracts
ANALYSIS AND CLINICAL CORRELATION OF SERUM INTERLEUKIN-4 LEVELS IN PROGRESSIVE SYSTEMIC Paul S. Lindner, MD; Marianne Frieri, SCLEROSIS. MD., PhD., East Meadow, New York Progressive systemic sclerosis (PSS) is a multisystemic disorder characterized by widespread fibrosis and degenerative changes of the skin, synovium, parenchyma and blood vessels. The etiology of PSS remains unknown. Cellular and humoral abnormalities have been described. Serum interleukin (IL)-2 levels and IL-2 receptors have recently been shown to be elevated in PSS. There are increased numbers of mast ccl Is in the papillary and reticular dermis, with enhanced activation in rapidly progressive disease. IL-4 is known to stimulate the growth of mucosal and connective tissue mast cells and fibroblasts. We analyzed serum IL-4 in patients with PSS to determine if it plays a role in this disease. Serum from 15 patients with PSS, 1 with morphea, 1 with sclerodactyly and 17 disease free controls was obtained and assayed for IL-4 by a highly sensitive ELISA using monoclonal antibodies. Clinical data on symptoms, disease interval, systemic involvement and a detailed skin score was obtained. There was a statistically significant correlation between IL-4 levels and skin score (r=.68, ~~005). An inverse correlation of IL-4 levels and length of disease was shown which did not reach statistical significance (r=.54). Patients with PSS were found to have a statistically significant elevation of serum IL-4 level compared to the controls (520.3~3.2 vs 10.8f1.5ng/ml, ~~01). The results of our study suggest a possible role of IL-4 in the pathogenesis of PSS, acting through its effects on mast cells, fibroblasts and T-cells.
MAST CELL EFFECTS ON CYTOKINE PRODUCTION IN SCLERODERMA PATIENTS. Marianne Friqri, Ph.D..M.D. Kushal Agarwal, M.D.,East Meadow New York Mast cell hyperplasia has bee; demonstrated in the skin and other organs in scleroderma (PSS) and irmunoregulatory cellular IL-2 defects have been reported (Frieri, M. et al. J Allergy Clin lmmunol 81:196a, 1988; Manuscript accepted by Arthritis Rheum, 1990). IL-4 can increase IgE production and stimulate mast cells and fibroblasts. Many of the effects of IL-4 are downregulated by TIFN. We evaluated 17 PSS patients and controls using systemic and skin scores along with (3H) Tdr IL-2 driven T ccl 1 pro1 iferation (TCP) with 10 ug/ml: of Con A, human type I collagen and heparin: 10'6 M histamine with antiIL-2/IL-4 monoclonal antibodies (MoAb) . A sensitive MoAb Elisa was used to determine‘OIFN levels in stimulated PBMC supernatants. Histamine and heparin along with collagen increased T cell blastogenesis in 11-16 patients at various stages of their disease compared to normals (!? SI in PSS = 2.3 + 0.28, 2.2 * 0.29, 1.8 t 0.2 vs K SI in normals = 0.5 f 0.005, 0.9 *O.OOl, 0.8 iO.01). Anti-IL-Z/IL-4 Moab decreased responses in 50% of patients. In contrast,fIFN levels in PBMC from 7 patients stimulated by histamine were significantly decreased (1.56 ko.26 ng/ml)vs 10 normals (3.10 fO.60 rig/ml), p =a1.05. The role of defective cytokine generation via IL-2, IL-4 andXlFN may contribute to significant advances in the diagnosis and treatment for patients with scleroderma.
87 Systemic lupus -e&hematosus ISLE) :-: associated with-autoant%bodies which- t&get 1 limited and oredictable set of antieens and these may p^artly be defined by 'bistinct structural properties. A rigorous statisticai method was used to analyze the sequenced SLEassociated autoantigens (n=33) far chargcrelated amino acid patterns which might be itmnunologically important. method The distinguishes significant long run 5 , local concentrations (clusters),and patterns of charge (every second or third residue). Significant
mixed charge runs often within clusters wer.? present in 12 (45.5%), and mixed charge cluster9
in another
(27%).
Long patterns
were-present
ix
Significant ruw andior another 2 antigens. clusters were present in i6% of a control set trf proteins. The mean % charge of the antigens wai; 32 vs 26 for controls (pX.001) suggesting that.
lacking locally high charge concentrated charge might be Siwificant folded uroteins. charge runs are present' in other co&xtivetissue disease autoantigens notably CENP-R and Topisomerase T of scleroderma, but are not. in
antigens
configurations
annarent
in the
prevalent in autoantigens
configurations
set a (Z/11). are
of
prevalent
organ-specific
Simila:charge and in viral
cellular transcription factors and surface proteins. Highly charged surfaces may render these viraland h&t proteins -strongly
inxnunoeenic and antinuclear antibodies resoonses to might - result from chronic exposur'e intracellular contents, possibly in conjunction with cross-reactive viral proteins.
88
BINDING
OF SERUM AMYLOID
(SAP) TO TIENTS Arturo
M.D., SAP
IM%lOBILIZED
DNA:
P COMPONENT A STUDY IN PA-
WITH SYSTEMIC LUPUS ERYTlJP!MATOSUS. M.D. and Anita Gewurz, 3. Bonnin, Chicaso, Illinois
is a-nbrmal human serum (40 + 10 and tissue glycoprotein, also Like C-reacfound in amyloid deposits. tive protein with which it is closely homologous, SAP binds to chromatin and nucleosomes with high affinity and is the major DNA binding protein in normal serum, yet its function(s) and role in disIn order to evaluate ease are unknown. DNA binding by SAP in patients with SLE and high titers of anti-DNA antibodies, we compared the quantity and DNA binding activity of SAP from sera of SLE patients with serum and purified SAP from normal donors. Protein levels of SAP were determined by electroimmunoassay using rabBinding of SAP to immobibit antiserum. lized DNA was performed by ELISA using either serum or purified SAP and a monoclonal antibody to detect SAP. The binding of purified SAP to DNA was calcium-dependent and constant over a wide pH range. SAP protein levels in SLE patient and normal sera were compasable. the binding to immobilized DNA However, by SAP in patient sera was reduced, compared to SAP in normal serum or purified SAP. Our data suggest that decreased SAP binding to immobilized DNA may be a factor associated with elevation of antiDNA antibodies in SLE patient sera. ug/ml)