African-American Race and Antibodies to Topoisomerase I Are Associated With Increased Severity of Scleroderma Lung Disease

Study objectives: To determine whether African-American race is independently associated with lung disease in scleroderma.Design: Retrospective review.Setting: University medical center in Baltimore.Patients: One hundred one patients with diffuse cutaneous scleroderma with available serum samples.Measurements: Patients underwent lung function testing as part of their routine clinical care. Percent predicted values adjusted for race were calculated for FVC, single-breath carbon monoxide diffusing capacity (DCO), and FEV1. Serum samples were assayed for the presence of antibodies to topoisomerase I and RNA polymerase II.Results: Scleroderma patients of African-American race had lower percent predicted values than white patients for FVC (p16 for the development of pulmonary fibrosis.8Briggs DC Vaughan RW Welsh KI et al.Immunogenetic prediction of pulmonary fibrosis in systemic sclerosis..Lancet. 1991; 338: 661-662Abstract PubMed Scopus (126) Google Scholar Furthermore, antibodies to the ER4 epitope of topoisomerase I confer an increased risk of progressive decline in FVC.9Kuwana M Kaburaki J Mimori T et al.Autoantigenic epitopes on DNA topoisomerase I: clinical and immunogenetic associations in systemic sclerosis..Arthritis Rheum. 1993; 10: 1406-1413Crossref Scopus (46) Google Scholar Prior reports implicate African-American race as a possible risk factor for scleroderma lung disease, but this has not been as intensively studied. Peters-Golden et al10Peters-Golden M Wise RA Schneider P et al.Clinical and demographic predictors of loss of pulmonary function in systemic sclerosis..Medicine. 1984; 63: 221-231Crossref PubMed Scopus (72) Google Scholar found that mean values of pulmonary function tests (PFTs) were significantly lower for African-American than white patients with scleroderma, but the PFT predicted values had not been adjusted to account for the fact that normal PFT values for African-Americans are lower than those for whites.11American Thoracic Society.Lung function testing: selection of reference values and interpretative strategies..Am Rev Respir Dis. 1991; 144: 1202-1218Crossref PubMed Scopus (2575) Google Scholar,12Jacobs Jr, DR Nelson ET Dontas AS et al.Are race and sex differences in lung function explained by frame size? the CARDIA study..Am Rev Respir Dis. 1992; 146: 644-649Crossref PubMed Scopus (39) Google Scholar An investigation limited to patients with severe lung disease found that African-Americans were overrepresented in this group,13Steen VD Conte C Owens GR et al.Severe restrictive lung disease in systemic sclerosis..Arthritis Rheum. 1994; 37: 1283-1289Crossref PubMed Scopus (454) Google Scholar but data linking race and lung disease in the larger population of scleroderma patients were not presented. Some of the features associated with poor lung function in scleroderma cluster with each other. Topoisomerase I antibodies are more common in patients with diffuse cutaneous scleroderma.14Steen VD Powell DL Medsger Jr, TA Clinical correlations and prognosis based on serum autoantibodies in patients with systemic sclerosis..Arthritis Rheum. 1988; 31: 196-203Crossref PubMed Scopus (553) Google Scholar In addition, race differences are associated with altered prevalence of diffuse skin involvement and autoantibody expression.15McNeilage LJ Youngchaiyud U Whittingham S Racial differences in antinuclear antibody patterns and clinical manifestations of scleroderma..Arthritis Rheum. 1989; 32: 54-60Crossref PubMed Scopus (94) Google Scholar In particular, African-Americans have a higher prevalence than whites of diffuse skin involvement16Laing TJ Gillespie BW Toth MB et al.Racial differences in scleroderma among women in Michigan..Arthritis Rheum. 1997; 40: 734-742Crossref PubMed Scopus (51) Google Scholar and antibodies to topoisomerase I.17Reveille JD Arnett FC Frequencies of scleroderma-related autoantibodies in patients meeting the American College of Rheumatology criteria for systemic sclerosis: reply letter..Arthritis Rheum. 1993; 36: 1333Crossref Scopus (14) Google Scholar To our knowledge, however, no study has previously investigated whether race and topoisomerase autoantibody expression represent independent risk factors for lung disease in scleroderma. The current study, therefore, examines whether differences in PFT results between African-Americans and whites with scleroderma persist when accounting for racial differences in autoantibody status. All patients seen at the Johns Hopkins and University of Maryland Scleroderma Center (JHUMSC) between January 1990 and June 1996, who met American College of Rheumatology criteria for the diagnosis of scleroderma,18Masi AT Rodnan GP Medsger Jr, TA et al.Preliminary criteria for the classification of systemic sclerosis (scleroderma)..Arthritis Rheum. 1980; 23: 581-590Crossref PubMed Scopus (4939) Google Scholar were evaluated for this study. For studies of the association of autoantibodies with scleroderma lung disease, all patients were included who had the following: (1) diffuse skin involvement (scleroderma skin changes proximal to the elbows or knees); (2) complete PFTs (including spirometry, lung volumes, and single-breath carbon monoxide diffusing capacity DCO) performed at JHUMSC; and (3) available serum samples for research analysis. All scleroderma patients who had at least two sets of complete PFTs at JHUMSC with at least a 1-year interval between tests were included in the longitudinal analysis. All patients were evaluated (by either F.M.W. or B.W.) and had clinical data recorded prospectively beginning at the time of presentation to JHUMSC. These data included age, sex, race, smoking history, date of onset of Raynaud's phenomenon, date of onset of non-Raynaud's symptoms of scleroderma, and extent of cutaneous disease. Diffuse cutaneous disease was coded if skin involvement was present on the trunk or proximal to the elbows or knees. Limited cutaneous disease was coded if skin involvement was confined to the face, the neck, and the extremities distal to the elbows and knees. Primary study outcome variables to be evaluated were defined as the percent predicted FVC, percent predicted DCO, and percent predicted FEV1. Predicted values for FVC and FEV1 were calculated according to Knudson, et al19Knudson RJ Lebowitz MD Holberg CJ Changes in the normal maximal expiratory flow-volume curve with growth and aging..Am Rev Respir Dis. 1983; 127: 725-734PubMed Google Scholar and predicted DCO was calculated according to Miller et al.20Miller A Thornton JC Warshaw R Single breath diffusing capacity in a representative sample of the population of Michigan, a large industrial state: predicted values, lower limits of normal, and frequencies of abnormality by smoking history..Am Rev Respir Dis. 1983; 127: 270-277PubMed Google Scholar Adjustment of predicted values for race was performed following American Thoracic Society recommendations.11American Thoracic Society.Lung function testing: selection of reference values and interpretative strategies..Am Rev Respir Dis. 1991; 144: 1202-1218Crossref PubMed Scopus (2575) Google Scholar The first set of PFTs performed at JHUMSC was used for analysis for each patient. When analyzing the change in percent predicted values over time, the first and last available PFTs performed at JHUMSC were used. The rate of change of PFT values was determined by subtracting the initial percent predicted values from the most recent percent predicted values, then dividing by the length of time (in years) between the initial and the most recent PFT. Patients were classified as positive for antibodies to topoisomerase I or RNA polymerase II if the serum sample provided for the patient was positive for these antibodies by immunoblot analysis.21Casciola-Rosen L Wigley F Rosen A Scleroderma autoantigens are uniquely fragmented by metal-catalyzed oxidation reactions: implications for pathogenesis..J Exp Med. 1997; 185: 71-79Crossref PubMed Scopus (189) Google Scholar Serum samples collected for immunoblotting were stored at −70°C and assayed in batches for this study, using well-characterized standard serum samples as internal controls for assay reproducibility. A trained investigator (A.R.), blinded to the identity and clinical status of each patient, interpreted each immunoblot. Immunoblots were judged as topoisomerase I positive and RNA polymerase II positive, respectively, if the blinded observer noted the presence of a visible band comigrating with the topoisomerase I band imaged using a topoisomerase I-positive control serum, or comigrating with the RNA polymerase II band pattern imaged using an RNA polymerase II positive control serum on the same gel. Antibodies to centromere were determined by immunofluorescent staining of Hep-2 cells, using standard techniques.6Weiner ES Earnshaw WC Senecal J-L et al.Clinical associations of anticentromere antibodies and antibodies to topoisomerase I: a study of 355 patients..Arthritis Rheum. 1988; 31: 378-385Crossref PubMed Scopus (198) Google Scholar Confluent monolayers of HeLa cells grown in standard culture conditions were washed, lysed, and suspended in sodium dodecyl sulfate sample buffer. Fifty-microgram aliquots of lysate were subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and transferred to nitrocellulose. Lanes of HeLa lysate were immunoblotted as previously described,22Casciola-Rosen LA Miller DK Anhalt GJ et al.Specific cleavage of the 70-kDa protein component of the U1 small nuclear ribonucleoprotein is a characteristic biochemical feature of apoptotic cell death..J Biol Chem. 1994; 269: 7-60Google Scholar using patient serum samples diluted to 1:5,000 as primary antibodies. Blots were preblocked with 3% bovine serum albumin, exposed to primary antibody, incubated with horseradish peroxidase-linked goat antihuman secondary antibody (Southern Biotech; Birmingham, Ala), and imaged with chemiluminescence (Pierce; Rockford, Ill) and 45-s film exposure (Kodak; Rochester, NY). All chemicals and reagents used were analytical grade. The Mann-Whitney U test on independent groups was used for bivariate analysis. Multivariate models were tested using multiple linear regression and analysis of variance. Population proportions were compared using Fisher's Exact Test. The standard for statistical significance was taken as p<0.05. The Bonferroni for multiple was differences in PFT were judged to be for were calculated using statistical for 1990 and 1996, patients with diffuse cutaneous scleroderma American College of Rheumatology criteria were seen at least at AT Rodnan GP Medsger Jr, TA et al.Preliminary criteria for the classification of systemic sclerosis (scleroderma)..Arthritis Rheum. 1980; 23: 581-590Crossref PubMed Scopus (4939) Google Scholar had complete PFTs performed at JHUMSC and available serum samples for research were no differences between the study patients and the patients with diffuse scleroderma the following: of of of mean age at time of initial or mean of non-Raynaud's symptoms at time of of Study Population and Patients Diffuse to topoisomerase I by to RNA polymerase II by PFT performed at JHUMSC; as percent of or as values determined by Fisher's Exact or Mann-Whitney U on independent Patients Patients at to topoisomerase I by to RNA polymerase II by PFT performed at JHUMSC; as percent of or as values determined by Fisher's Exact or Mann-Whitney U on independent in a In the study of the patients were was a than to to and mean age of onset was of the study patients were African-American African-Americans were younger than whites of of non-Raynaud's symptoms of scleroderma of Patients Diffuse Scleroderma by Racial of at For of in a percent of the patients had antibodies to topoisomerase I by immunoblot with the rate of topoisomerase I antibody in scleroderma patients at and anticentromere antibodies in the Rev 1994; Google Scholar Antibodies to RNA polymerase an autoantibody associated with increased mortality and diffuse skin disease in M Kaburaki J Mimori T et with of RNA in from patients with systemic sclerosis..J 1993; PubMed Scopus Google Scholar were present in of the study of had anticentromere antibodies. These patients were included in the analyses analyses the anticentromere positive patients A higher of African-American patients were positive for antibodies to topoisomerase I of Antibodies to RNA polymerase II were present in of African-Americans and of whites Antibodies to RNA polymerase II were associated with increased prevalence of antibodies to topoisomerase I of patients with antibodies to RNA polymerase II had antibodies to topoisomerase I, of patients RNA polymerase II antibodies had I antibodies of Patients Diffuse by of at at For of in a differences between topoisomerase I or RNA polymerase II and patients were found in the of of age at or of scleroderma symptoms African-American patients had lower percent predicted values compared with white patients for FVC DCO and The presence of antibodies to topoisomerase I was associated with percent predicted FVC compared with patients antibodies to topoisomerase I lower DCO and FEV1 in patients with topoisomerase I antibodies were RNA polymerase II antibody was not significantly associated with changes in FVC, DCO, or FEV1. of PFT values for the African-American and white are by topoisomerase I and RNA polymerase II antibody status of on of values determined by Mann-Whitney U on independent predicted for for multiple predicted predicted predicted predicted predicted For of values determined by Mann-Whitney U on independent for for multiple in a of percent predicted values for FVC, DCO, and FEV1 by race and autoantibody status. Patients with diffuse cutaneous scleroderma were classified based on race and autoantibody status have antibodies to topoisomerase I, not have antibodies to topoisomerase I, have antibodies to RNA polymerase not have antibodies to RNA polymerase status was determined by immunoblot of 1:5,000 of patient serum samples HeLa Percent predicted values for FVC, DCO, and FEV1 were calculated and for each patient. standard of the mean for each linear regression models race, topoisomerase I antibody status, RNA polymerase II antibody status, smoking history, of non-Raynaud's scleroderma symptoms, and age were for FVC, DCO, and models no between race and autoantibody status. FVC from race and topoisomerase I antibody status, with no The of the of race and of topoisomerase I antibody status on FVC was African-American race predicted a of in the percent of predicted FVC, the presence of antibodies to topoisomerase I predicted an independent of The regression for DCO for race and also for sex but the topoisomerase I antibody status was not race a to the regression for FEV1. RNA polymerase II antibody status was not a in of the models for Percent Predicted PFT in Patients Diffuse Scleroderma and of For of in a To the differences in lung function between African-American and white patients over time, scleroderma patients with a mean of years of non-Raynaud's symptoms at were for an of African-American race was independently associated with lower FVC, DCO, and FEV1 on initial for age, sex, smoking status, of disease, of and extent of cutaneous disease not the time the rate of change in lung function was and was not significantly between African-Americans and whites for DCO and FEV1 not African-Americans a of percent of predicted FVC whites had a of of predicted FVC The of this study is that African-American race and topoisomerase I antibody were independently associated with worse lung function in scleroderma by et VD Conte C Owens GR et al.Severe restrictive lung disease in systemic sclerosis..Arthritis Rheum. 1994; 37: 1283-1289Crossref PubMed Scopus (454) Google Scholar and Peters-Golden et al10Peters-Golden M Wise RA Schneider P et al.Clinical and demographic predictors of loss of pulmonary function in systemic sclerosis..Medicine. 1984; 63: 221-231Crossref PubMed Scopus (72) Google Scholar African-American race as associated with more severe lung disease in scleroderma. the reports by that in lung and expiratory are of the scleroderma lung disease associated with African-American race, and by that the of race when for topoisomerase I antibody status. The between African-American race and lung disease in scleroderma are not The fact that white and African-American race of and the that the differences from or R The of of a Med. 1996; PubMed Scopus Google Scholar included that topoisomerase I antibodies were more in African-Americans than that antibodies to topoisomerase I were associated with FVC, and that antibodies to RNA polymerase II were not associated with worse lung disease. Antibodies to topoisomerase I have been more in African-Americans in one JD Arnett FC Frequencies of scleroderma-related autoantibodies in patients meeting the American College of Rheumatology criteria for systemic sclerosis: reply letter..Arthritis Rheum. 1993; 36: 1333Crossref Scopus (14) Google Scholar but not in two TJ Gillespie BW Toth MB et al.Racial differences in scleroderma among women in Michigan..Arthritis Rheum. 1997; 40: 734-742Crossref PubMed Scopus (51) Google M Kaburaki J et al.Racial differences in the of systemic serum antinuclear Rheum. 1994; 37: PubMed Scopus Google Scholar in the prevalence of topoisomerase I antibodies in African-Americans compared with whites differences in population or differences in the used to topoisomerase I African-American race independently predicted lower lower lung volumes, and lower expiratory pattern of changes that is with pulmonary MB et in systemic sclerosis: of lung function in to extent of disease on Rheum. 1997; 40: PubMed Google I was independently associated with lung RNA polymerase II antibodies were not associated with changes in lung function The independent association of a component of scleroderma lung disease with I antibodies the whether the in NK Glanville AR Strickland B et al.Pulmonary involvement in systemic sclerosis: detection of early changes by thin section CT scan, brochoalveolar lavage and 99mTc-DTPA clearance..Respir Med. 1989; 83: 403-414Abstract Full Text PDF PubMed Scopus (129) Google Scholar be by a topoisomerase models also that sex was independently associated with DCO, but not with changes in FVC or FEV1. is that a that associated sex with increased risk of scleroderma lung disease defined lung disease based on DCO VD Conte C Owens GR et al.Severe restrictive lung disease in systemic sclerosis..Arthritis Rheum. 1994; 37: 1283-1289Crossref PubMed Scopus (454) Google Scholar in DCO in scleroderma have previously been linked with pulmonary VD Conte C et diffusing capacity in systemic sclerosis..Arthritis Rheum. 1992; PubMed Scopus Google Scholar is whether sex is a risk factor for pulmonary hypertension in scleroderma. When a study that the of multiple variables on clinical of statistical is a study included In multivariate regression models with independent our sample predicted a of a in outcome variables of of predicted for FVC, DCO, and of of at A F a and analyses for the University of Scholar this is to clinical more associations The presence of of change in lung function between the initial and PFTs in a longitudinal patient with Tashkin DP et changes in lung function and symptoms in progressive systemic sclerosis: study..Am J Med. 83: Full Text PDF PubMed Scopus Google Wise RA et pulmonary function in systemic sclerosis..Am J Med. Full Text PDF PubMed Scopus Google Scholar that this of patients had an early progressive of their disease by the time of their first and that time had by the time of their PFT for pulmonary complications of scleroderma to Diffuse cutaneous skin is to over the first years in VD Medsger Jr, TA and of systemic Google Scholar and diffuse cutaneous scleroderma has been to and over years of P P Furst D et of skin involvement in systemic sclerosis over years in a of Rheum. 1993; 36: PubMed Scopus Google Scholar the and of lung disease early in scleroderma the same pattern of early by previously for the VD Medsger Jr, TA and of systemic Google Scholar et VD Conte C Owens GR et al.Severe restrictive lung disease in systemic sclerosis..Arthritis Rheum. 1994; 37: 1283-1289Crossref PubMed Scopus (454) Google Scholar a pattern of early decline in lung function by in patients with severe restrictive lung disease. In this study has found in a large of scleroderma patients seen at a center in African-American patients had lung volumes, and expiratory compared with white The association of African-American race with lower lung function in scleroderma patients was predicted values to lower mean lung in The of race when accounting for autoantibody status, and when patients were Antibodies to topoisomerase I were more among African-American patients than whites and were associated with lower Racial differences in scleroderma or factors that as