Clinical significance of autoantibodies to the pericentromeric heterochromatin protein 1a protein.

OBJECTIVE: The objective of the study is to determine the frequency and the clinical significance of autoantibodies to the pericentromeric heterochromatin protein 1 (HP1). So far this antinuclear antibody specificity has been mainly reported in patients with the CREST syndrome. METHODS: We screened the sera of 199 individuals, including patients suffering from various autoimmune disorders (Group I, n=145) and non autoimmune diseases (Group II, n=44 patients) as well as healthy individuals (Group III, n=30). The sera were systematically tested by Western blot and ELISA using a GST-HP1α fusion protein as an antigen. RESULTS: Anti-HP1 antibodies were detected in 32% of patients in Group I, 11.3% in Group II and 3.3% of individuals in Group III. They could be detected in sera containing or not antinuclear antibodies detectable by indirect immunofluorescence. Anti-HP1 antibodies were mostly associated with the CREST and Sjogren's syndromes (70% and 44.4%, respectively). They could also be detected in 22.2% of patients suffering from various other autoimmune diseases. However, their negative predictive value was 94% in the CREST syndrome. CONCLUSION: Anti-HP1 autoantibodies are associated with a large spectrum of disorders. However, they have a diagnostic value in the CREST syndrome.

Anti-myelin-associated glycoprotein polyneuropathy coexistent with CREST syndrome.

Clinical involvement of the peripheral nervous system in the calcinosis cutis, raynaud's phenomenon, esophageal dismotility, sclerodactyly and telangiectasia (CREST) variant of systemic sclerosis occurs infrequently and is characterized by axonal degeneration due to necrotizing vasculitis. We report a female patient with a known history of CREST syndrome, which developed a slowly progressive, distal symmetric demyelinating sensorimotor polyneuropathy (PN), with tremor and ataxia as prominent features, compatible with anti-myelin associated glycoprotein (MAG) PN. The diagnosis of PN was established by the presence of monoclonal immunoglobulin M anti-MAG antibodies (Thin-Layer Chromatography, Western Blot and enzyme-linked immunoabsorbent assay). Given the evidence that in CREST activation of T-helper cells is observed and that anti-MAG antibodies, despite the fact that they are T-cell-independent, may be influenced by an increase in T-helper function, the coexistence of these two rare autoimmune disorders in the same patient may not be incidental but related to the underlying immunological mechanisms involved.

Evaluation of the reactivity of sera from patients with systemic lupus erythematosus against the human MCP1.

This study evaluates metaphase chromosome protein 1 (MCP1), a nuclear antigen, as a diagnostic marker for systemic lupus erythematosus (SLE). Reactivity of sera from 114 Portuguese patients with autoimmune rheumatic disease or from healthy blood donors (HBD), against MCP1, produced in bacteria (bact-MCP1) or in its native form (native-MCP1), was determined by immunoblotting. Predictive and discriminative power of MCP1 reactivity for SLE diagnosis in disease-control groups was evaluated by logistic regression, its diagnostic value determined by receiver-operating characteristic analysis and compared with similar analysis of antinuclear antibody and double-stranded DNA (dsDNA). We demonstrated that native-MCP1, in contrast to bact-MCP1, reacts with SLE sera with significant predictive and discriminative power versus other autoimmune diseases (odds ratio [OR] ≤3.537 and ≥3.265; area under the receiver-operating characteristic curve [AUC] ≤0.643 and ≥0.636) or versus HBD (OR = 5.006; AUC = 0.671), showing a good diagnostic power with high specificity (82.1% versus HBD) and low sensitivity for SLE, similar to those of dsDNA. The reactivity of SLE sera with native-MCP1 was shown to be dependent on the presence of phosphorylated residues. Native-MCP1 was shown to have diagnostic value as a specific marker for SLE diagnosis and, therefore, is a suitable substrate for a new antibody test. The widely reported importance of phosphorylated epitopes as targets for autoantibodies in SLE could also be confirmed for native-MCP1.

[A case of anti-AQP4 antibody-positive recurrent myelitis overlapped with autoimmune disorders including incomplete CREST syndrome revealed multiple discontinuous cord lesions].

A 65-year-old woman presenting with multiple autoimmune disorders including incomplete CREST overlapping with aquaporin 4 (AQP4) antibody-positive recurrent myelitis was reported. She also clinically suffered from Sjogren syndrome and primary biliary cirrhosis (PBC). She had dysesthesia below C4 level, mild motor weakness and hyperreflexia without pathological reflexes on bilateral lower extremities. A T2-weighted MRI indicated multiple discontinuous spinal cord lesions at C1-5 and T7/8. A visual evoked potential study disclosed bilateral prolonged latency of P100. She clinically manifested not only incomplete CREST syndrome (facial teleangiectasia, sclerodactyly in bilateral fingers, and Raynaud's phenomenon), but also Sjögren (sicca syndrome) and PBC (jaundice). Immunoserological study showed that she was positive for anti-nuclear, anti-centromere, and anti-AQP4 (= NMO-IgG) antibodies. A combination therapy with corticosteroid and plasmapheresis was effective for all clinical symptoms. Therefore, this case stresses on the relevance of anti-AQP 4 antibody to the other overlapping autoimmune disorders, such as CREST syndrome, when recurrent myelitis is clinically diagnosed.

Increased prevalence of anti-third generation cyclic citrullinated peptide antibodies in patients with rheumatoid arthritis and CREST syndrome.

To investigate the prevalence of anti-third generation cyclic citrullinated peptide antibodies (anti-CCP3) in patients with systemic connective tissue diseases, we assembled a training set consisting of 115 patients with rheumatoid arthritis (RA), 52 with Calcinosis, Raynaud's phenomenon, oesophageal dysmotility, sclerodactyly, telangiectasia (CREST) syndrome, 21 with scleroderma, 20 with ankylosing spondylitis, 18 with reactive arthritis, 25 with juvenile rheumatoid arthritis (RA), 51 with osteoarthritis, 26 with mixed connective tissue disease, 23 with primary Sjogren's syndrome, 74 with systemic lupus erythematosus, 49 with Polymyalgia rheumatica, and 39 with polymyositis/dermatomyositis. The commercial enzyme-linked immunosorbent assay (ELISA) was used to detect anti-CCP antibodies, including anti-CCP2 (regular, second generation of CCP antigen) and anti-CCP3 (third generation of CCP antigen) in disease-related specimens and normal controls. These serum samples were also evaluated for anti-centomere antibodies by anti-centromere ELISA kit. The higher frequencies of anti-CCP3 and anti-CCP2 were detected in 75.6 and 70.4% patients with RA, respectively. At the same time, anti-CCP3 (not anti-CCP2) was significantly increased in samples isolated from patients with CREST syndrome. The clinical sensitivity of IgG anti-CCP3 for the patients with CREST syndrome was 29% (15 of 52) and the specificity was 96% (384 of 397), with the exception of the RA group. The anti-centromere antibodies were significantly higher in patients with CREST only. The results of our study suggest that compared to anti-CCP2 assay, the new anti-CCP3 assay can enhance the clinical sensitivity for diagnosis of RA and, as an associate marker combined with anticentromere, can distinguish CREST syndrome from other systemic connective tissue diseases, especially RA. The clinical specificity of anti-CCP3 was lower than anti-CCP2 assay in diagnosis of RA because of the crossreaction to the patients with CREST syndrome.

Primary biliary cirrhosis (PBC)-CREST overlap syndrome with coexistence of Sjögren's syndrome and thyroid dysfunction.

Calcinosis, Raynaud's phenomenon, oesophageal dysmotility, sclerodactyly, telangiectasia (CREST) syndrome, a limited form of systemic sclerosis, is sometimes complicated by primary biliary cirrhosis (PBC). A 52- and 61-year-old Japanese woman with PBC-CREST overlap syndrome accompanied by Sjögren's syndrome, and Hashimoto's thyroiditis, and Graves' disease, respectively, are reported. They had suffered from Raynaud's phenomena, sclerodactyly, morning stiffness, arthralgia, and sicca symptoms during these several years. They exhibited an increased level of alkaline phosphatase, gamma-glutamyl transpeptidase, positive antibodies against mitochondria and centromere, and hyperglobulinemia without any cholestatic symptoms. Histological findings from liver biopsy specimens were consistent with those of PBC. Clinically, they were diagnosed as both asymptomatic PBC and incomplete CREST syndrome. Their human leukocyte antigen typing showed both DR4 and DR8 positive. The association of the four autoimmune conditions is clinically and etiologically important. Although a combination of these diseases is rare, it is of importance to keep in mind that various autoimmune diseases could occur simultaneously. Of critical importance is that an active diagnostic attitude towards them is admirable, and that early diagnosis and therapy are needed.

Anti-centromere antibodies in patients with systemic lupus erythematosus.

BACKGROUND: Anti-centromere autoantibodies (ACA) are frequently detected in systemic sclerosis (SScl), especially in the calcinosis, Raynaud phenomenon, esophageal dysmotility, sclerodactyly, telangiectasia (CREST) syndrome, in which a prevalence of 55% has been reported. The presence of ACA in systemic lupus erythematosus (SLE) is so rare that its detection can raise serious doubts about the validity of the diagnosis. OBJECTIVE: To determine the frequency of ACA positive subjects from a wide monocentric cohort of SLE patients and analyse the clinical and biological characteristics of this group. METHODS: Five hundred and sixty consecutive SLE patients were systematically analysed for the presence of ACA and other autoantibodies using indirect immunofluorescence, counter-immunoelectrophoresis, double immunodiffusion, enzyme-linked immunosorbent assay (ELISA), and Western-blot. RESULTS: ACA were detected in 11 SLE patients (1.9%); all of them were women. The CENP-B-specific ELISA was positive in all patients. The main clinical features of scleroderma (cutaneous sclerosis, sclerodactylia, digital ulcers, or pulmonary fibrosis) were not present in these patients, who did not differ clinically from the whole SLE group. CONCLUSIONS: ACA can be detected in patients with genuine SLE without concurrent scleroderma. Therefore, the presence of this antibody does not preclude the possibility of the diagnosis of SLE. In addition, SLE patients with ACA do not represent a different clinical subgroup.

The presence of dominant T-cell clones in peripheral blood of patients with collagen vascular disorders: a prospective study of 97 cases.

BACKGROUND: T-lymphocyte dysfunction has been seldom investigated in collagen vascular disorders. The search for dominant T-cell clones has been scarcely reported, although the presence of such clones might be expected in disorders showing immune responses directed against a variety of autoantigens. OBJECTIVES: We conducted a systematic search for dominant T-cell clones in peripheral blood in patients with collagen vascular disorders. Patients and methods Ninety-seven patients with collagen vascular disorders were studied (7 cutaneous and 38 systemic lupus erythematosus; 8 multiple morphea; 12 regional scleroderma; 32 systemic sclerosis of the CREST type). A dominant T-cell clone was searched for in peripheral blood by polymerase chain reaction targeting the T-cell receptor gamma chain followed by a size analysis of amplified fragments. Peripheral blood from patients with nonlymphocyte-dependent disorders and matched by age and sex was assessed in the same conditions. Results in both groups were compared using nonparametric statistical tests. RESULTS: Overall, a circulating dominant T-cell clone was found in 52% of patients compared with 16.9% in controls. More precisely, such a dominant clone was present in 43% and 37% of cutaneous and systemic lupus erythematosus, respectively, in 75% of multiple morphea, 75% of regional scleroderma and 60% of CREST syndrome patients. The percentages in all subsets of patients were significantly higher than in the control group. CONCLUSIONS: The presence of a dominant T-cell clone in peripheral blood is significantly more frequent in collagen vascular disorders than in controls, especially in patients with scleroderma, whatever the clinical subset, which suggests T-cell involvement in the immune response dysfunction in these diseases classically characterized by disturbances of B lymphocytes. The relevance of such a dominant clone regarding diagnosis, pathomechanisms, long-term outcome and visceral prognosis of these diseases as well as therapeutic decisions remains to be evaluated.

Incidence and clinical correlation of anticentromere antibody in Thai patients.

Anticentromere antibodies (ACA) are useful in assessing and classifying patients with mild variant of systemic sclerosis called calcinosis, Raynaud's phenomenon, esophageal dysmotility, sclerodactyly, telangiectasias (CREST) syndrome. From their prognostic significance, we are interested in the prevalence and disease correlation in Thai patients. A total of 3,233 serum samples of patients with any musculoskeletal symptoms were sent for antinuclear antibody determination at Ramathibodi Immunology Laboratory Service between the years 1998 and 2001. Forty sera (1.23%) were ACA positive. These sera were from 27 patients with autoimmune diseases and 13 with nonautoimmune diseases. Among autoimmune group, scleroderma was the most common diagnosis (33.3%) with limited sclerosis being the most frequent variant. The percentages of autoimmune disease were almost the same among the low-titer (1:40) and the high-titer (1:640) groups. The study suggests that the prevalence of ACA in Thai patients is low. The presence of ACA detected in patients with vague musculoskeletal symptoms does not suggest a diagnosis of CREST syndrome. Even high-titer ACA can be found in nonautoimmune diseases.

Clinical and serological heterogeneity in patients with anticentromere antibodies.

OBJECTIVE: To evaluate the clinical and serological heterogeneity in patients with anticentromere antibodies (ACA). METHODS: One hundred twenty patients with ACA were analyzed retrospectively. ACA were detected initially on the basis of indirect immunofluorescence on HEp-2 cells, and then antibodies to CENP-B were measured by ELISA. Antibodies to other nuclear antigens were also detected by double immunodiffusion and/or ELISA. RESULTS: Eighty-four patients (70.0%) had systemic sclerosis (SSc; scleroderma) and 36 patients (30.0%) had other rheumatic diseases or miscellaneous disorders. Among patients with SSc, 35 patients (41.7%) had SSc in overlap mostly with Sjögren's syndrome (SS), in part with rheumatoid arthritis and/or primary biliary cirrhosis (PBC). Five of 36 patients (13.9%) without SSc also had overlap syndrome of more than 2 rheumatic diseases or PBC. All CREST features (calcinosis, Raynaud's, esophageal dysmotility, sclerodactyly, telangiectasias) were found significantly more in SSc than in other diseases. A combination of RST was the most frequently seen, followed by CREST and CRST in the SSc group. In contrast, 22 of 36 patients (61.1%) without SSc had no CREST features, and the rest had only Raynaud's phenomenon and/or telangiectasia. Twenty-five of 75 patients (33.3%) with SSc and 6 of 25 patients (24.0%) with other diseases had a slight elevation of creatine phosphokinase concentration with no apparent myositis signs and/or skin lesions, suggesting a new additional sign of patients with ACA. Seventy-two patients (60.0%) had ACA alone and 48 patients (40%) had ACA mixed with other disease marker antinuclear antibodies (ANA). ACA alone occurred more frequently in patients with SSc and in the non-overlap group, whereas patients with ACA mixed with other ANA were more frequently found in the other disease and the overlap syndrome groups. Anti-CENP-B ELISA levels of the SSc group were significantly higher than those of other disease groups in all patients, in patients with ACA alone, and in patients having ACA together with other ANA. The most frequently concurrent ANA were anti-SSA/Ro antibodies; and the other ANA, including anti-SSB/La, RNP, topoisomerase-I, Jo-1, Ku, and dsDNA antibodies, were also positive alone or combined with more than 2 ANA in patients with ACA. Five patients with CREST syndrome having ACA and anti-RNP antibodies had clinical manifestations compatible with mixed connective tissue disease. SS was found in 37.0% of patients who had higher anti-CENP-B ELISA levels and higher coincidence of anti-SSA/Ro antibodies than the patients without SS. CONCLUSION: ACA were positive mostly in patients with SSc with CREST features and partly in other rheumatic disorders. The high levels of ACA may be necessary for the development of CREST features, and frequent concurrence of other disease marker ANA may contribute to the development of heterogeneous clinical characteristics, including overlap syndrome, in patients with ACA.

[Analytic study of dot blotting for the detection of anti-Jo-1, anti-M2, anti-ribosomes and anti-LKM]. / Etude analytique d'un dot blot pour la détection des auto-anticorps anti-Jo-1, anti-M2, anti-ribosomes et anti-LKM1.

The Cyto-Dot 4 HM043 kit commercialised by BMD, has replaced the Cyto-Dot HM010 kit that allowed three auto-antibodies detection (anti-Jo-1, anti-M2 and anti-ribosomal protein). Detection of anti-LKM1 auto-antibody was added. These four auto-antibodies have in common only the intracytoplasmic localisation of their respective antigen. The aim of our study was to evaluate this new kit using 104 sera and to compare our results with reference techniques (indirect immunofluorescence IF for anti-M2, anti-ribosomal protein and anti-LKM1, double immunodiffusion ID for anti-Jo-1 and anti-LKM1, western blotting WB for anti-M2) and with Cyto-Dot HM010. The one hundred and four sera were divided into five groups: Group I (n = 12) with anti-Jo-1 detected by ID; Group II (n = 28) with 26 anti-M2 positive by IF and WB, 2 anti-M2 positive only by WB; Group III (n = 10) with anti-ribosomal protein detected by IF 5 of which precipitated by ID; Group IV (n = 32) with anti-LKM1 by IF and ID divided into 18 AIH2 and 14 HCV; Group V (n = 22) consisting of 14 healthy individuals and 8 patients with hypergammaglobulinemia. Results of this study are similar to those of Cyto-Dot HM010 for the three auto-antibodies already in use. Cyto-Dot 4 is a very good anti-LKM1 confirmation method as it is ID.

Human ninein is a centrosomal autoantigen recognized by CREST patient sera and plays a regulatory role in microtubule nucleation.

Centrosome is the major microtubule organizing center in mammalian cells that plays a critical role in a variety of cellular events by the microtubule arrays emanating from it. Despite its significance, the molecular mechanisms underlying the structure and function of the centrosome are still not clear. Herein we describe the identification of three isotypes of human ninein by expression library screening with autoimmune sera from CREST patients. All three ninein isotypes exhibit centrosomal localization throughout the cell cycle when GFP-tagged fusion proteins are expressed transiently in mammalian cells. Construction of serial deletions of GFP-tagged ninein reveals that a stretch of three leucine zippers with a flanking sequence is required and sufficient for centrosomal targeting. Overexpression of ninein results in mislocalization of gamma-tubulin, recruiting it to ectopic (noncentrosomal) ninein-containing sites which are not active in nucleating microtubules. In these cells, nucleation of microtubules from the centrosome is also inhibited. These results thus suggest a regulatory role for ninein in microtubule nucleation.

Successful long-term treatment with cyclosporin A in protein losing gastroenteropathy.

Protein-losing gastroenteropathy (PLG) can occur as a manifestation of various diseases including autoimmune disorders, and optimal therapy of these underlying diseases may be the only effective remedy for PLG. In the present report, we describe a case of a 54-year-old woman with PLG associated with an autoimmune disease, presumably CREST syndrome. She failed to respond to steroid treatment. Subsequently, cyclosporine was initiated, which resulted in a rapid recovery. The patient was successfully treated with low-dose cyclosporine for five years. There has not been, to our knowledge, any report of PLG successfully treated with cyclosporine. Cyclosporine therapy may be effective not only in inducing but also in maintaining complete remission in patients with autoimmune-associated PLG, especially refractory or intolerable to steroids and/or immunosuppressive therapies.

Induction of kinetochore-positive and kinetochore-negative micronuclei in CHO cells by ELF magnetic fields and/or X-rays.

To test the genotoxic effects of extremely low frequency (ELF) magnetic fields, the induction of micronuclei by exposure to ELF magnetic fields and/or X-rays was investigated in cultured Chinese hamster ovary (CHO) cells, using the cytokinesis block method. Micronuclei derived from acentric fragments or from whole chromosomes were evaluated by immunofluorescent staining using anti-kinetochore antibodies from the serum of scleroderma (CREST syndrome) patients. A 60 Hz ELF magnetic field at 5 mT field strength was applied, either before or after 1 Gy X-ray irradiation or without additional X-ray irradiation. No statistically significant difference in the frequency of micronuclei in CHO cells was observed between a sham exposure (no exposure to an ELF magnetic field) and a 24 h ELF magnetic field exposure. Exposure to an ELF magnetic field for 24 h before X-ray irradiation or for 18 h after X-ray irradiation did not affect the frequency of X-ray-induced micronuclei. However, the number of kinetochore-positive micronuclei was significantly increased in the cells subjected to X-ray irradiation followed by ELF magnetic field exposure, but not in the cells treated with ELF magnetic field exposure before X-ray irradiation, compared with exposure to X-rays alone. The number of spontaneous kinetochore-positive and kinetochore-negative micronuclei was not affected by exposure to an ELF magnetic field alone. Our data suggest that exposure to an ELF magnetic field has no effect on the number of spontaneous and X-ray-induced micronuclei. However, ELF magnetic field exposure after but not before X-ray irradiation may somehow accelerate X-ray-induced lagging of whole chromosomes (or centric fragments) in CHO cells.

[Polymyositis/scleroderma autoantibody-positive scleroderma with dermatomyositis (scleromyositis)]. / Polymyositis/sclerodema autoantitest pozitív scleroderma dermatomyositissel (scleromyositis).

Polymiositis/scleroderma autoantibody positive scleroderma with dermatomyositis connected to interstitial lung fibrosis is reported. The recently observed overlap syndrome is characterised by the presence of special autoantibodies, HLA-type association and benign course. A new skin symptom (mechanic's hands) is described which may indicate the syndrome and one of its important internal manifestations, the interstitial lung disease.