Summary
In 135 patients with systemic scleroderma, we compared three different methods to determine antinuclear autoantibody (ANA) specificity: indirect immunofluorescence, double immunodiffusion, and, employing recombinant antigens, immunoblotting using both marker autoantigens of this disease. A characteristic Scl-70 antibody pattern was found on HEp-2 cells in 83.8% of the patients, double immunodiffusion was positive for the Scl-70 antibodies in 81.9%, and immunoblot with the recombinant topoisomerase I (Topo I) was positive in 71% of the patients. For the centromere autoantibodies we found a high concordance between the anticentromere antibody (ACA) pattern on HEp-2 cells (27 patients positive) and the detection of recombinant kinetochore in immunoblotting (26 patients positive). The three testing techniques gave comparable results, except that the Topo I recombinant antigen used in immunoblotting reacted strongly with fewer than expected of the known Scl-70-positive sera. However, a method using recombinant antigens expressing all epitopes (rather than one of the epitopes of Topo I) will undoubtedly become the method of choice for detecting antibodies in systemic scleroderma. Using the immunoblotting technique with the recombinant antigens we detected in four patients antibodies against both Topo I and kinetochore. More severe symptoms of systemic scleroderma were found in patients who had both antibodies. The combined presence of both marker autoantibodies is therefore not as rare as previously reported and may predict severe disease.
Similar content being viewed by others
References
Alderuccio F, Barnett AJ, Campbell JH, Pedersen JS, Toh HH (1986) Scl-95/100: doublet of endothelial marker autoantigens in progressive systemic sclerosis. Clin Exp Immunol 64:94–100
Bernstein RM, Steigerwald JC, Tan EM (1982) Association of antinuclear and antinucleolar antibodies in progressive systemic sclerosis. Clin Exp Immunol 48:43–51
Bernstein RM, Callender ME, Neuberger JM, Hughes CRV, Williams R (1982) Anticentromere antibody in primary biliary cirrhosis. Ann Rheum Dis 41:612–614
Beutner EH, Krasny SA, Chorzelski TP, Rodnan G, Jabłońska S, Kumar V (1985) Evaluation of methods for detection of anticentromere antibodies and other anti-nuclear antibodies. J Am Acad Dermatol 12:289–295
Black CM, Welsh KI, Maddison PL, Jayson MIV, Bernstein RM (1984) HLA autoantibodies and clinical subsets in scleroderma. Br J Rheumatol 23:267–271
Cattoggio LJ, Bernstein RM, Black CM, Hughes GRV, Maddison PJ (1983) Serological markers in progressive systemic sclerosis: clinical correlations. Ann Rheum Dis 42:23–27
Chen ZY, Fedrick AJ, Pandey JP, Silver R, Maricq HR, Fudenberg HH, Dobson RL, Ainsworth SK (1985) Anticentromere antibody and immunoglobulin allotypes in scleroderma. Arch Dermatol 121:339–344
Chorzelski TP, Jabłońska S, Beutner EH, Blaszczyk M, Jarzabek-Chorzelska M, Kencka D, Krasny S, Kumar V, Tchorzewska A (1985) Anticentromere antibody: an immunological marker of a subset of systemic sclerosis. Br J Dermatol 113:381–389
Cox JV, Schenk EA, Olmsted JB (1983) Human anticentromere antibodies: distribution, characterization of antigens, and effect of microtubule organization. Cell 35:331–339
Douvas AS, Achten M, Tan EM (1979) Identification of a nuclear protein (Scl-70) as a unique target of human anti-nuclear antibodies in scleroderma. J Biol Chem 254:10514–10522
Earnshaw WC, Rothfield N (1985) Identification of a family of human centromere proteins using autoimmune sera from patients with scleroderma. Chromosoma 91:313–321
Earnshaw WC, Halligan N, Cooke C, Rothfield N (1984) The kinetochore is part of that metaphase chromosome scaffold. J Cell Biol 96:352–357
Earnshaw WC, Bordwell B, Marino C, Rothfield N (1986) Three human chromosomal autoantigens are recognized by sera from patients with anticentromere antibodies. J Clin Invest 77:426–430
Earnshaw WC, Sullivan KF, Machlin PF, Cooke CA, Kaiser DA, Pollard TD, Rothfield NF, Cleveland DW (1987) Molecular cloning of cDNA for CENP-B, the major human centromere autoantigen. J Cell Biol 104:817–829
French BT, Maul HM, Maul GG (1986) Screening cDNA expression libraries with monoclonal and polyclonal antibodies using amplified biotin-avidin-peroxidase technique. Analyt Biochem 156:417–423
Fritzler MJ, Kinsella TD (1980) The CREST syndrome: a distinct serologic entity with anticentromere antibodies. Am J Med 69:520–526
Giordano M, Valentini G, Migliaresi S, Picillo U, Vatti M (1986) Different antibody patterns and different prognoses in patients with scleroderma with various extent of skin sclerosis. J Rheumatol 13:911–916
Guldner HH, Lakomek HJ, Bautz FA (1985) Human anticentromere sera recognize a 19.5 kD nonhistone chromosomal protein from HeLa cells. Clin Exp Immunol 58:12–20
Guldner HH, Szostecki C, Vosberg HP, Lakomek HJ, Penner E, Bautz FA (1986) Scl-70 autoantibodies from scleroderma patients recognize a 95 kD protein identified as DNA topoisomerase I. Chromosoma 94:132–138
Jarzabek-Chorzelska M, Blaszczyk M, Jabłońska S, Chorzelski TP, Kumar V, Beutner EH (1986) Scl-70 antibody: a specific marker of systemic sclerosis. Br J Dermatol 115:393–401
Juarez C, Vila JL, Celpi C, Augusti M, Amengual ML, Martinez MA, Rodriguez JL (1988) Characterization of the antigen reactive with anti-Scl-70 antibodies and its application in an enzymelinked immunosorbent assay. Arthritis Rheumat 31:108–115
Kallenberg CGM, Pastoor GW, Wouda AA, The TH (1982) Antinuclear antibodies in patients with Raynaud's phenomenon: clinical significance of anticentromere antibodies. Ann Rheum Dis 41:382–387
Kumar V, Beutner EH, Dabski K, Steger R, Koelle M (1984) A standardized method of detecting antibodies to extractable nuclear antigens (RNP and Sm) by gel precipitation. J Clin Lab Immunol 15:163–166
Laemmli UK (1970) Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227:680–685
Lakomek MJ, Guldner HM, Bautz FA, Goerz G, Kind P, Mensing H, Krüskember HL (1987) Kern-Antikörper als serologische Marker der progressiv systemischen Sklerodermie (PSS). Hautarzt 38:63–69
Maddison PJ, Skinner RP, Pereira RS, Black CM, Ansell BM, Jayson MIV, Rowell NR, Welsh KI (1986) Antinuclear antibodies in the relatives and spouses of patients with systemic sclerosis. Ann Rheum Dis 45:793–799
Makinen D, Fritzler M, Davis P, Sherlock S (1983) Anticentromere antibodies in primary biliary cirrhosis. Arthritis Rheum 26:914–917
Masi AT, Rodnan GP, Medsger TA Jr, Altman RD, d'Angelo WA, Fries JF, Le Roy EC, Krisner AB, MacKenzie AH, McShane DS, Myers AR, Sharp GC (1980) Preliminary criteria for the classification of systemic sclerosis (scleroderma). Arthritis Rheum 23:581–590
Maul GG, French BT, van Venrooij WJ, Jimenez SA (1986) Topoisomerase I identified by scleroderma 70 antisera: enrichment of topoisomerase I at the centromere in mouse mitotic cells before anaphase. Proc Natl Acad Sci USA 83:5145–5149
Maul GG, Jimenez SA, Riggs E, Ziemnicka-Kotula D (1989) Determination of an epitope of the diffuse scleroderma marker antigen topoisomerase I. Sequence similarity with retroviral p30gag protein suggests a possible cause for autoimmunity in scleroderma. Proc Natl Acad Sci USA (in press)
McCarty GA, Rice RJ, Bembe ML, Barada FA (1983) Anticentromere antibody. Clinical correlation and association with favorable prognosis in patients with scleroderma variants. Arthritis Rheum 26:1–7
McNeilage LJ, Whittingham S, McHough N, Barnett AJ (1986) A highly conserved 72000 dalton centromeric antigen reactive with autoantibodies from patients with progressive systemic sclerosis. J Immunol 137:2541–2547
Miller MH, Littlejohn GO, Davidson A, Jones B, Topliss DJ (1987) The clinical significance of the anticentromere antibody. Br J Rheumatol 26:17–21
Nishikai M, Okano Y, Yamashita H, Watanabe M (1984) Characterization of centromere (kinetochore) antigen reactive with sera of patients with a scleroderma variant (CREST syndrome). Ann Rheum Dis 43:819–824
Powel FC, Winkelmann RK, Venencie-Lemarekant F, Spurbeck JL, Schroeter AL (1984) The anticentromere antibody: disease specificity and clinical significance. Proc Mayo Clinic 59:700–706
Provost T, Herrera-Esparza R, Diaz LA (1985) Nucleoprotein autoantibodies in lupus erythematosus. Invest Dermatol 85:133s-139s
Rodnan GP, Medsger TA, Buckingham RE (1975) Progressive systemic sclerosis — CREST syndrome: observations on natural history and late complications in 90 patients. Arthritis Rheum 18:423
Rodnan GP, Jablonska S, Medsger TA Jr (1979) Classification and nomenclature of progressive systemic sclerosis (scleroderma). Clin Rheum Dis 5:5–13
Ruffatti A, Caligaro A, Ferri C, Bombardieri S, Gambari PF, Todesco S (1985) Association of anticentromere and anti-Scl-70 antibodies. Report of two cases. J Clin Lab Immunol 16:227–229
Shero JH, Bordwell B, Rothfield NF, Earnshaw WC (1986) High titers of autoantibodies to topoisomerase I (Scl-70) in sera from scleroderma patients. Science 231:737–740
Sontheimer RD (1985) Immunobiological significance of the Ro/SSA antigen-antibody system. Arch Dermatol 121:327–330
Steen VD, Ziegler GL, Rodnan GP, Medsger TA Jr (1984) Clinical and laboratory association of anticentromere antibody in patients with progressive systemic sclerosis. Arthritis Rheum 27:125–131
Steen VG, Powell DL, Medsger TM Jr (1988) Clinical correlations and prognosis based on serum autoantibodies in patients with systemic sclerosis. Arthritis Rheum 31:196–203
Takehara K, Moroi Y, Ishibashi Y (1985) Antinuclear antibodies in the relatives of patients with systemic sclerosis. Br J Dermatol 112:23–33
Tan EM (1981) The biological and diagnostic significance of antinuclear antibodies. Aust N Z J Med 11:193–197
Tan EM, Rodnan GP, Garcia I, Moroi Y, Fritzler MJ, Peebles C (1980) Diversity of antinuclear antibodies in progressive systemic sclerosis. Anticentromere antibody and its relationship to CREST syndrome. Arthritis Rheum 23:617–625
Towbin H, Staehelin T, Gordon J (1979) Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci USA 76:4350–4354
Tuffanelli DL, McKeon F, Kleinsmith A, Burnham TK, Kirschner M (1983) Anticentromere and anticentriole antibodies in the scleroderma spectrum. Arch Dermatol 119:560–566
Van Venrooij WJ, Stapel SO, Hoben H, Habets WJ, Kallenberg GM, Penner E, van de Putte LB (1985) Scl-86, marker antigen for diffuse scleroderma. J Clin Invest 75:1053–1060
Whittingham S, Naselli G, McNeilage LK, Coppel RL, Sturgess AD (1987) Serological diagnosis of primary Sjögren's syndrome by means of human recombinant La (SS-B) as nuclear antigen. Lancet I:1–3
Author information
Authors and Affiliations
Additional information
Supported by grants from the CPBP Nr. 06.01. of the Polish Academy of Sciences, AR grant 38907 from the National Institute of Health, and the Scleroderma Research Foundation
Rights and permissions
About this article
Cite this article
Jarzabek-Chorzelska, M., Blaszczyk, M., Kolacinska-Strasz, Z. et al. Antikinetochore and antitopoisomerase I antibodies in systemic scleroderma: comparative study using immunoblotted recombinant antigens, immunofluorescence, and double immunodiffusion. Arch Dermatol Res 282, 76–83 (1990). https://doi.org/10.1007/BF00493462
Received:
Issue Date:
DOI: https://doi.org/10.1007/BF00493462