Sera from patients with scleroderma contained several autoantibodies to nuclear antigens which were distinguished by different patterns of nuclear immunofluorescence staining. One of these autoantibodies reacted with centromeric regions of chromosomes. In chromosome spreads, the staining appeared as two small spheres at the centromere, resembling kinetochores. The antigenic determinant appeared to be a protein or polypeptide tightly bound to DNA. The autoantibody was reactive with centromeres of cells derived from humans, mice, and Chinese hamsters. The autoantibody was present in high frequency in the calcinosis/Raynaud's phenomenon/ esophageal dysmotility/sclerodactyly/telangiectasia variant (CREST) of scleroderma.Progressive systemic sclerosis, also known as scleroderma, is a chronic systemic rheumatic disease that can affect many organ systems including the skin and subcutaneous tissues, the gastrointestinal tract, the heart, the lungs, and the kidneys. The etiology of the disease is unknown but much information has been accumulated on the clinical, pathogenetic, and serological abnormalities associated with it. Sera of patients with scleroderma have been shown to contain autoantibodies to nucleolar and other intranuclear components. Generally, these autoantibodies have been detected by the immunofluorescence technique. By using tissue sections as substrate, autoantibodies have demonstrated different patterns of nuclear staining which have been described variously as nucleolar, speckled, and homogeneous (1, 2).Some of the intranuclear antigens reacting with autoantibodies in scleroderma sera have been elucidated. There is a 4S-6S RNA, isolated from liver nucleoli, that specifically precipitates with serum autoantibody (3). Recently, another nuclear antigen has been identified and termed . This was shown to be a nonhistone nuclear protein of approximately 70,000 daltons. By polyacrylamide gel electrophoresis, Scl-70 was identified as a distinct protein band closely associated with but clearly separable from histone fraction H1.In this report, we describe an autoantibody in scleroderma sera which reacts with the centromere (kinetochore) of chromosomes. This autoantibody was initially observed to give speckled nuclear staining on substrates consisting of organ sections. When tissue culture cells of different origins were used as substrates, the speckled staining was clearly observed to be associated with the centromeres of chromosomes. Further analysis suggested that the centromere antigen might be protein or polypeptide components tightly bound to the centromeric DNA of chromosomes.MATERIALS AND METHODS Serum. Sera from 32 patients with scleroderma (25 females and 7 males) were studied. Twenty-three healthy persons (15 females and 8 males) were used as controls. All sera were heat-inactivated at 56°C for 30 min prior to use.Immunofluorescence Studies. The indirect fluorescent antibody technique (5) was used to determine the nuclear staining patterns produced by the sera under study. The substrates cons...
Patterns of nuclear staining included diffuse fine speckles, large coarse speckles, nucleolar and centromere staining. When organ sections such as mouse kidney were used as substrate for the detection of antinuclear antibodies, nucleolar staining and centromere staining were the two patterns most frequently overlooked. Three types of antibodies appeared to be highly specific for scleroderma: antibody to Scl-70 antigen, antibody to centromere, and antinucleolar antibody. The anti-centromere antibody appeared to be highly selective for the CREST variant of progressive systemic sclerosis.
Antigens associated with mammalian centromeres were localized at the light and electron microscopic levels using the peroxidase-labeled antibody method. The antibody used was of a type naturally occurring in the sera of patients with scleroderma . At the light microscopic level, it reacts specifically with the centromere regions of chromosomes in a variety of mammalian species and strains in discrete foci in interphase nuclei . We find that the number of foci approximates the number of chromosomes present in the various cell types . At the ultrastructural level, the antigenic foci are confirmed to lie in the kinetochore regions of each chromosome . In interphase nuclei, the antigenic foci were usually associated either with the inner surfaces of the nuclear envelope or with the nucleoli. These observations indicate that the centromere regions of the chromosomes in interphase are not randomly distributed within the nucleus but are usually fixed either to the inner surface of the nuclear envelope or to nucleoli .
Antinuclear antibodies were studied by indirect immunofluorescence and double immunodiffusion in 21 patients with systemic sclerosis and 35 of their relatives. When HEp-2 cells were used as the substrate, the frequency of antinuclear antibodies in the patients' sera was 100% and that in the relatives was 26%. When rat liver sections were used, the values were 86% and 17%, respectively. Anticentromere antibody was detected in the serum from the mother of one patient whose serum had anti-Scl-70 antibody. Antibody to n-RNP was positive in the sera from the brother and daughter of another patient whose serum was positive for anti-n-RNP and anti-Scl-70 antibodies. The high frequency of antinuclear antibodies in the sera from the relatives of systemic sclerosis patients suggests that immunological abnormalities play a part in the pathogenesis of this condition.
This study aims to evaluate the validity and reliability of a Japanese version of the Arthritis Impact Measurement Scales, version 2 (AIMS2) for patients with rheumatoid arthritis (RA). The Japanese version of the AIMS2 questionnaire was administered to 1643 patients with classical or definite RA at 11 hospitals nationwide in Japan. Reliability was assessed by a test-retest procedure, 4 weeks apart, using 75 randomly selected patients. Internal consistency was measured by Cronbach's α, and factor analysis was used to obtain the proportion of variance explained by the first factor in principal component analysis. The validity of the AIMS2 scales was assessed by internal standards. Internal consistency (α coefficients, 0.84-0.94), test-retest reliability (intraclass correlation coefficients, 0.75-0.93), and factor analysis (0.62-0.85) of the AIMS2 health status scales proved that they are highly reliable in the Japanese version. Validity, as measured by the relationships among the scores on the questionnaire items, was also sufficiently secured. The validity and reliability of the Japanese version of the AIMS2 are sufficient for all practical purposes when compared with the original and with other translated versions of the questionnaire.
A cloned complementary DNA, termed pS2, was isolated from a human fibroblast cDNA library in the bacteriophage expression vector lambda gt11 after screening with a patient's serum containing a high titer of anti-ribonucleoprotein (RNP) antibodies. A reasonable amount of cro-beta-galactosidase fusion protein (pS2EX) was obtained through subcloning of the pS2 insert into a plasmid expression vector pEX-2. Antibody against pS2EX (anti-pS2EX) was purified from this patient's serum by Sepharose 4B conjugated with pS2EX. Immunofluorescent staining of HeLa cells with anti-pS2EX antibody exhibited a typical speckled pattern in the interphase nuclei. In the immunoblot analysis, the anti-pS2EX antibody recognized the 22 kDa protein. Using immunoprecipitation of cell lysate and subsequent RNA analysis, anti-pS2EX antibody was shown to precipitate U1 RNP only. The reactivities of various anti-RNP sera to pS2EX correlated well with the positive reaction to C polypeptide in the immunoblot. These findings indicate that pS2 is a cDNA for C polypeptide of U1 snRNP. In the Northern blot using human RNA and radiolabeled pS2, a single band about 800 base was observed. The nucleotide sequence of pS2 showed no significant homologies to known proteins.
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