Clinical significance of anti-Ro52 (TRIM21) antibodies non-associated with anti-SSA 60kDa antibodies: results of a multicentric study.

Ro52 antigen has recently been identified as TRIM21 protein, but the clinical significance of anti-Ro52/TRIM21 antibodies remains controversial. The aim of this multicentric study was to investigate the significance of anti-Ro52 antibodies without anti-SSA/Ro60 antibodies in various connective diseases. Sera were selected by each laboratory using its own method (ELISA, immunodot or Luminex technology), and then performed with ANA Screen BioPlex™ reagent (BIO-RAD). Among the 247 screened sera, 155/247 (63%) were confirmed as anti-Ro52 positive and anti-SSA/Ro60 negative. These sera were analyzed for the detection of other antibodies in relation with clinical settings. Isolated anti-Ro52 antibodies were detected in 89/155 (57%) sera. For the remaining sera (66/155), the main antibodies associations were Sm/SmRNP or Chromatin (n=38; 57%), Jo1 (n=17; 26%) and CenpB (n=9; 14%). Clinical data from the 155 patients showed high prevalence in autoimmune diseases (73%) including myositis or dermatomyositis (n=30), lupus (n=23); Sjögren and/or sicca syndrome (n=27); CREST or Systemic sclerosis (n=11) and autoimmune hepatitis (n=11). We found that pulmonary manifestations were often associated with the presence of anti-Ro52 antibodies (n=34, 22%), in addition with anti-tRNA synthetases, anti-SRP or anti-Ku antibodies (18/34) or isolated in half of cases (16/34). Separate detection of anti-Ro52 antibodies might be useful in related antisynthetase syndrome diagnosis. The presence of anti-Ro52 antibodies should probably precede development of autoimmune disease and must induce sequential follow-up of positive patients, particularly in interstitial lung disease progression.

Evaluation of the LIAISON ANA screen assay for antinuclear antibody testing in autoimmune diseases.

Antinuclear antibodies (ANA) are widely detected by immunofluorescence on HEp-2 cells in patients with connective tissue diseases and other pathological conditions. We evaluated the first-automated chemiluminescence immunoassay for the detection of ANA (LIAISON ANA screen, DiaSorin). This study was carried out simultaneously in two laboratories by testing 327 patient samples with clinically defined connective diseases, 273 routine samples for ANA screening, and 300 blood donors. A total of 268 out of 337 IIF-positive sera were positive with LIAISON ANA screen (79.5% of agreement) and 240 out of 263 IIF-negative sera were negative with LIAISON ANA screen (91.2% of agreement). After resolution of discrepant results, the concordance reached, respectively, 94.9% and 98.8%. The specificity was 99.3% and the sensitivity was 94%. Unlike results obtained by other ANA screening assays, we observed acceptable sensitivity and specificity. Despite the presence of HEp-2 cell extract, we failed to detect some antibodies as antinucleolar, antinuclear envelope, and antiproliferating cell nuclear antigen. This automated assay allows quick process to results and exhibits satisfactory sensitivity for the detection of the main ANA specificities of connective tissue diseases.

Detection of antibodies to dsDNA: an overview of laboratory assays.

This brief review is focused on different methodologies available for detection of anti-dsDNA antibodies with respect to the best adequacy between biological results of laboratory and clinical significance. A large array of assays has been developed for the measurement of anti-dsDNA. New assays continually introduced have reflected not only technical innovations to avoid difficulties of some assays, but even more with hope to correlate better with systemic lupus erythematosus (SLE), particularly with its clinical course and exacerbations. Finally, with the development of micro arrays technology new insights into the pathophysiology of autoimmune disease processes should be revealed.

[Chronic urticaria, thyroiditis and autologous serum test]. / Urticaire chronique, thyroïdite auto-immune et test au sérum autologue.

INTRODUCTION: The autoimmune nature of certain forms of chronic urticaria remains debatable. Aim of the study. To find a correlation in terms of autoimmune pattern between chronic urticaria and thyroiditis using the autologous serum test and the search for anti-thyroid antibodies. PATIENTS AND METHODS: Study in a single center of 59 patients having consulted for urticaria that had progressed for more than six weeks and without precise etiology. An autologous serum test was conducted and anti-thyroid autoantibodies were searched for in the serum stored after the test. RESULTS: Out of 59 patients, 51 had an interpretable autologous serum test and 57 underwent research for anti-thyroid antibodies. Twelve patients out 57 (21 p.cent) exhibited one or more types of positive (n=10) or borderline (n=2) anti-thyroid antibodies, and eight out of 12 (66.6 p.cent) had a known thyroid disease before the onset of urticaria. The systematic blood test permitted the discovery of 4 cases of unknown anti-thyroid antibodies. The symptoms regressed in two of these patients without any specific treatment, one patient was treated with L thyroxin for hyperthyroidism without any improvement in the symptoms and one other patient had normal thyroid function. Among these 12 patients, 11 underwent autologous serum tests, but only 3 of them exhibited clearly positive results and one was doubtful. CONCLUSIONS: Within the limits of this study, we found a 21 p.cent rate of patients with chronic urticaria exhibiting one or more anti-thyroid antibodies. The positivity of the autologous serum test does not appear to be related with autoimmune thyroid disorders.

[Hashimoto's thyroiditis and silicone breast implants: 2 cases]. / Thyroïdite de hashimoto et prothèses mammaires en silicone: 2 observations.

The silicone implant controversy wavers between reassuring epidemiological studies and about 300 case reports of patients developing a definite or incomplete/atypical connective tissue disease (CTD) after receiving a silicone gel-filled breast implant (SBI). Since Hashimoto's thyroiditis (HT) is rarely reported in this context, we report here two new cases of HT associated with a history of bilateral cosmetic SBIs. The first patient was a 45-year-old white woman who had SBIs in 1976. In 1991 she developed HT, evolving to thyroid deficiency which was compensated with levothyroxine treatment. In addition, the patient complained of fatigue, arthralgia, morning stiffness and developed a sicca syndrome necessitating artificial tears. The 1995 evaluation disclosed the presence of antinuclear antibodies at a titre of 1/640, and high level anti-thyroid microsomal antibodies (1/256,000). Gamma globulins rose to 22.6%. Thyroid ultrasonography showed an enlarged thyroid gland with a diffusely hypoechogenic pattern. The implants were painful, and in 1996 they were removed. Microscope examination of the fibrous capsule surrounding the prostheses showed extremely dense connective tissue with fibrosis. The second patient was a 55-year-old white woman who had SBIs in 1984. In 1995, she developed HT with clinical pain and tenderness of the thyroid gland, with mild hyperthyroidism and positive antithyroglobulin antibodies, and was given corticosteroid treatment for 5 months. In 1996, the implants were again painful and the patient developed positive antinuclear antibodies with a titre of 1/200. Ultrasonography showed a heterogeneous thyroid gland, and implant removal was advised. Hashimoto's thyroiditis is recognized as a subset of chronic auto-immune thyroiditis, and its association with SBI is rare. In these 2 observations, an association without relation is possible, but a future survey of similar cases seems warranted.

Autoantibodies to phospholipids and to the coagulation proteins in AIDS.

In HIV-1 infection, an increased prevalence of anticardiolipin autoantibodies (aCL) and lupus anticoagulant (LA) has been described. In order to see if these antibodies are isolated or, like in autoimmune diseases, associated with hematological disorders and with antibodies to other phospholipids and to proteins of coagulation, we investigated 3 groups of patients: 1. 342 HIV-1 infected patients, 2. 145 control patients including 61 systemic lupus erythematosus (SLE) patients, 58 patients with a connective tissue disease, 15 patients with stroke, 11 patients with syphilis and 3. 100 blood donors. In HIV-1 infection antiprothrombin (aPrT) antibodies were present in 2% of patients, the prevalence of antiphosphatidylcholine antibodies (aPC) (50%) was almost as high as aCL (64%), and 39% had both antibodies. Absorption on liposomes of the latter revealed an heterogeneous mixture of aCL and aPC or cross-reacting antibodies. In contrast with SLE, anti-beta 2-glycoprotein I (4%), LA (1%), biological false positive test for syphilis (0.3%), thrombosis (p < 0.001) were uncommon. In HIV-1 infection, antiphospholipid antibodies do not associated with features linked to them in SLE or syphilis.

Advantage of using both anionic and zwitterionic phospholipid antigens for the detection of antiphospholipid antibodies.

To investigate the benefit of assaying for antiphospholipid antibodies (aPA) with different antigenic specificities, sera from 141 patients suspected of having aPA were tested by ELISA for IgG and IgM antibodies directed against the following phospholipids (PL) coated individually or together: cardiolipin, phosphatidylinositol, phosphatidylserine and phosphatidylethanolamine. Nonspecific background optical density (OD) was systematically subtracted from the test OD value. Positive reactions were defined as having an OD greater than the 97th percentile OD distribution obtained with sera from 100 healthy individuals. Although the majority of the 79 detected aPA (89% IgG and 77% IgM) were polyspecific, 11 reacted with a single PL and, moreover, belonged to only one isotype. Seven of these 11 patients presented recurrent fetal losses or thrombotic events. These results suggest that routine use of a mixture of both anionic and zwitterionic PL antigens to coat ELISA plates would better detect aPA involved in suggestive pathologies and enhance the ability to identify patients with these mono- or polyspecific antibodies directed or not against cardiolipin, the current standard.

[Conclusions of the first French workshop of standardization of anticardiolipin antibody determination associated with autoimmune pathology]. / Conclusions du 1er Atelier francais de standardisation du dosage des anticorps anticardiolipine associés à la pathologie autoimmune.

Anticardiolipine antibodies are a marker of the antiphospholipid syndrome. They are detected by Elisa. Despite its simplicity, the results obtained by this assay are not always reproducible despite several worldwide standardizations and the distribution of calibration sera by EN Harris. On 4 December 1992, the First French Anticardiolipin Antibodies Standardization Workshop was held in Paris. Eight coded standards were sent to 33 laboratories in France, one in Switzerland and one in Luxembourg. Some of them used several assays. Agreement between qualitative results was good for samples with high and moderate level of antibodies. But the results expressed in IgG antiphospholipid units were scattered, the interlaboratory coefficient of variation for each of the eight standards was higher than 70%, though the within-run coefficient of variation had a median value of 10%. Assays using bovine serum for the blocking buffer and for the dilution buffer seemed to give more reproducible results than assays using purified bovine serum albumin (p < 0.001). For only nine assays out of 38 which expressed results quantitatively, was a linear regression observed between the dilutions of a sample and the results expressed in IgG antiphospholipid units. Absence of linearity is an obstacle which must be overcome if we wish to compare quantitative results obtained by different laboratories.

Two-site enzyme immunoassay of CD4 and CD8 molecules on the surface of T lymphocytes from healthy subjects and HIV-1-infected patients.

A highly sensitive two-site enzyme immunoassay (Capcellia) was developed to determine the concentration of CD4 and CD8 molecules expressed on the surface of human T lymphocytes. This assay, performed in one step (20 min), involves the specific immunocapture of T lymphocytes and reaction of the CD4 or CD8 molecules with an enzyme-labeled monoclonal antibody (mAb). The results were expressed as molar concentrations of the T-cell markers on the basis of results obtained with calibrated CD4 and CD8 standards. The assay was sensitive enough to detect 0.4 pmol/L CD4 or 0.8 pmol/L CD8, which corresponded to approximately 20 x 10(6) CD4+ or CD8+ T cells per liter of blood. Mean concentrations in healthy adults were 17.2 pmol/L for CD4 and 22.1 pmol/L for CD8. The CD4 concentration was < 8 pmol/L in 50% of HIV-1-infected patients and in 95% of AIDS patients. Given the epitopic specificity of the mAb to CD4 we used, these values correspond to the concentration of CD4 molecules free of envelope glycoprotein (gp)120.

[Autoimmune abortive disease. A new entity of gynaeco-obstetrical expression]. / La maladie abortive auto-immune. Nouvelle entité à expression gynéco-obstétricale.

The relationship between abortive disease and systemic lupus erythematosus is complex as shown by data from the literature and by our 9 patients selected for presenting with abortive disease, circulating anticoagulant and biological signs of autoimmunity. The risk of transformation into a systemic disease is real, although difficult to evaluate in the absence of prospective studies, but the major problem with these patients is the severity of the obstetrical pathology. The sombre foetal prognosis, already reported in the literature, requires close supervision and sustained treatment during pregnancy. For these reasons we suggest calling autoimmune abortive disease the disease which affects women with signs of autoimmunity who have had several, often late foetal deaths associated with the presence of circulatory anticoagulant thought to interfere with placental blood flow.

[Value of the study of biological autoimmunity in gyneco-obstetrical pathology]. / Intérêt de l'exploration de l'auto-immunité biologique en pathologie gynéco-obstétricale.

The relationship between autoimmunity and obstetrical pathology is reviewed in the light of data from the literature and of the authors' own experience. The link, already demonstrated, between recurrent spontaneous abortion and the risk factor represented by antibodies directed against phospholipids (circulating anticoagulant antibodies of the lupus type, anticordiolipin antibody) is described, and the pathogenetic theories concerning these antibodies are presented, together with the treatments used for routine prevention of the foetal and maternal risk (corticosteroids, anti-platelet drugs, heparin). The results of screening for antinuclear antibodies in 354 non-pregnant women are reported. Antinuclear antibodies were present in titers of 1/100 in 4.2 percent and 1/50 in 8.7 percent of these women. In the same category of women the frequency of antithyroid antibodies was as high as 12 percent. These data should encourage routine examination for auto-antibodies before and during pregnancy in order to evaluate and prevent the obstetrical risk to which these women are exposed.

Immunohistological localization of immunoglobulins in pneumocystosis of adults.

We examined by direct immunofluorescence open lung biopsies from seven adult patients with Pneumocystis carinii pneumonia. In all of them, immunoglobulin deposits (especially IgA) were found in alveoli that exhibited by light microscopy the classical foamy exudate infested by Pneumocystis. These immunoglobulins were eluted at an acid pH and were able to fix complement fractions in vitro. Moreover, we could reproduce this fixation by incubating previously eluted tissue sections with concentrated immunoglobulins isolated from a bronchopulmonary lavage performed in another case of pneumocystis. This immunohistological pattern suggests the presence of a humoral response in pulmonary parenchyma of patients infected with Pneumocystis carinii, despite the consistent underlying immunocompromised status. Considering these results, we conclude that humoral immune mechanisms do not constitute the main barrier against the development of Pneumocystis carinii in human lungs.

Study of the local graft versus host reaction in the rat. I. Evidences for a donor-related "rat effect".

The local graft versus host reaction (GVHR) observed in the hybrid F1 LBN rat was chosen to evaluate the immunologic effect induced by allogeneic transfusion of Brown Norway blood to the Lewis rat. Study of the kinetics of this GVHR with immunized as well as with nonimmunized cells shows an "individual rat effect" which is shown by a variability of the popliteal lymph node weight from one recipient to another. When spleen cells from different Lewis donors are pooled this effect disappears which indicates that it is related to the donor's rather than to the F1 recipient's cells. To even out variation between individual donors, cells from a number of donors should be pooled to allow better interpretation of experimental protocols using this GVHR.

Autoimmunity, fetal losses, lupus anticoagulant: beginning of systemic lupus erythematosus or new autoimmune entity with gynaeco-obstetrical expression?

Among the various autoantibodies commonly found in women with systemic lupus erythematosus, the so-called lupus anticoagulant has been described in association with fetal losses. Recently, women with repeated spontaneous abortions and lupus anticoagulant, but no apparent systemic lupus erythematosus have been described. We have studied prospectively the possible connections between fetal losses and autoimmunity in a large control study of 130 idiopathic habitual aborters. These non-pregnant patients without any antecedent autoimmune disease were explored for the presence for autoantibodies especially lupus anticoagulant. Ten percent of the patients exhibit an antithromboplastin antibody (lupus anticoagulant) and half of this group possesses a striking association of biological manifestations of autoimmunity. In conclusion, two points appear: firstly, statistical correlation is demonstrated between antithromboplastin antibody and habitual abortion; secondly, doubt remains about the significance of the association between autoimmunity, fetal losses and antithromboplastin: beginning of systemic lupus erythematosus or new autoimmune entity with gynaeco-obstetrical expression.

[Immunological disorders of coagulation in habitual abortion. Prospective study]. / Désordres immunologiques de la coagulation dans la maladie abortive. Etude prospective.

The physiopathological role of antithromboplastin-type circulating anticoagulants in habitual abortion may be envisaged since the presence of antithromboplastin has been reported in most studies on women at high risk of abortion. To avoid a possible statistical bias, we conducted a prospective study in a sufficiently large group of women with habitual abortion (n = 99) compared with a control group of women with normal fecundity (n = 50). In addition, all women were investigated for lupus symptoms. The circulating antibody was detected by the diluted thromboplastin time and activated cephalin time methods. The results were considered positive when the patient/control diluted thromboplastin time ratio was 1.2 and/or when the increase in activated cephalin time was not corrected by a control plasma. In the patients' group, 10 women (10%) had an anti-thromboplastin type circulating anticoagulant, whereas no circulating anticoagulant could be detected in the control group. Three women with circulating anticoagulant had signs of systemic lupus erythematosus. None of the patients presented with Soulier-Boffa syndrome. These data have established a significant correlation between habitual abortion and circulating anticoagulant whilst avoiding statistical bias. Our results suggest that women with idiopathic habitual abortion should be subjected to systematic immunological exploration and that a small number of them should be followed attentively.

Biological tolerance to polydioxanone absorbable clips: a comparison with metallic ligating clips.

An experiment was conducted in 96 rats to determine the tissue tolerance to a hemostatic and absorbable clip made of polydioxanone, as compared to standard metallic clips. Clinical tolerance was similar and no bleeding or death occurred because of the material. Pathological studies showed a mild histiocytic reaction with absorbable clips and a stronger inflammatory reaction with metallic clips. This experiment indicates that absorbable clips are efficient and well tolerated and we strongly recommend the use of this material.

[Experimental study of an absorbable hemostatic clip]. / Etude expérimentale d'un clip hémostatique résorbable.

A new absorbable hemostatic clip was investigated by a study in the rat. The reliability and innocuity of the clip were similar to that of the metallic clips from the clinical point of view. Its absorbability and radiotransparency qualities make it the method of choice for ensuring hemostasis and lymphostasis when long-term follow-up investigations involve modern radiological techniques such as computed tomography.