Effects of anti-CD4 antibodies on the release of IL-6 and TNF-alpha in whole blood samples from patients with systemic lupus erythematosus.

Anti-CD4 antibodies have been recently introduced into the therapy of various autoimmune diseases, among them systemic lupus erythematosus (SLE). Their modes of action are not yet fully understood. Interference with cytokine release may be one possible mechanism. Therefore, the effects of anti-CD4 antibodies on the cytokine release of IL-6 (interleukin-6) and TNF-alpha (tumor necrosis factor alpha) were investigated in a whole blood culture system. Basal and phytohemagglutin/lipopolysaccharide (PHA/LPS)-stimulated cytokine patterns were compared to cytokine release after the addition of anti-CD4 antibodies (MAX.16H5) or methylprednisolone in short time whole blood cell culture systems from 12 patients with active SLE, 23 patients with inactive SLE and 12 healthy volunteers. TNF-alpha and IL-6 concentrations were determined in the supernatants by ELISA. High disease activity correlated with an increased production of proinflammatory cytokines. Cell cultures of patients with inactive SLE showed a diminished capacity to respond to mitogenic stimulation. Anti-CD4 antibodies added in vitro suppressed significantly the unstimulated production of IL-6 (P<0.02) in the cell cultures of patients with active SLE and in the PHA/LPS-stimulated cell cultures from both groups of SLE patients (both P<0.001) and healthy volunteers (P<0.01). However, MAX.16H5 did not affect the release of TNF-alpha. In control samples methylprednisolone considerably reduced stimulated and unstimulated IL-6 and TNF-alpha production in all SLE patients, irrespective of the disease state, and in all healthy controls. These data indicate that the proinflammatory cytokines are involved in the pathogenesis of SLE. It is assumed that anti-CD4 antibodies, which can be effective in the treatment of highly active lupus patients, may act via their influence on cytokine release. The decrease of the proinflammatory cytokines IL-6 under therapy with MAX.16H5 could explain the observations of clinical trials and animal studies which showed a reduction of inflammatory parameters and diminished production of autoantibodies following treatment with anti-CD4 antibodies.

A novel epitope on the C-terminus of SmD1 is recognized by the majority of sera from patients with systemic lupus erythematosus.

The SmD1 protein is a specific target for the autoantibody response in SLE. To further analyze this reactivity epitope, mapping was performed with cellulose-bound 13-mer peptides overlapping 10 amino acids (aa). In this initial approach, 4 out of 15 SLE sera recognized more than five overlapping peptides of the SmD1 C-terminus. Therefore, longer oligopeptides of up to 37 aa of this region were generated and probed for as antigens by ELISA. For the SmD1 aa 83-119 polypeptide, there was a striking increase of reactivity with 70.0% positive reactions out of 167 SLE sera. In contrast, 105 healthy control sera were negative, and only 8.3% of sera from patients with other inflammatory diseases (n = 267) exhibited a response, which was of low level only. The anti-SmD183-119 reactivity was significantly higher in anti-dsDNA antibody positive vs. negative sera (P < 0.001) and correlated with disease activity. Four of five human monoclonal anti-dsDNA antibodies also reacted with SmD183-119. The specificity for SmD1 was demonstrated by inhibition experiments and immunization of rabbits with SmD183-119 inducing SmD1-specific antibodies. In conclusion, the SmD183-119 peptide was demonstrated to be an important and highly specific target of the autoimmune response in SLE. The high sensitivity of this ELISA probably depends on a conformational epitope, which appears not to be accessible in the full-size SmD1 protein.

Serologic characteristics in primary biliary cirrhosis associated with sicca syndrome.

BACKGROUND: Patients with primary biliary cirrhosis (PBC) may have coincidental sicca syndrome (SS). The frequencies of SS and the occurrence of the Sjögren's-associated anti-Ro and -La antibodies in PBC patients have been reported at widely varying prevalences. This study investigated whether distinctive serologic characteristics are associated with SS in PBC. METHODS: Forty PBC patients and thirty patients with other types of liver cirrhosis were tested for SS and associated autoantibodies (ANA, AMA 2, anti-Ro, anti-La, anti-U1RNP-A, -C, -68 kD, and rheumatoid factors). RESULTS: Fourteen PBC patients (35%) complained of sicca symptoms, of whom 10 (25%) had a positive Schirmer-I test, and 7 (17.5%) had serologic characteristics similar to those of Sjögren's syndrome. Anti-52-kD Ro antibodies were positive in seven PBC/SS cases (p < 0.025). There was no anti-Ro positive PBC patient without SS. Three patients with PBC/SS with anti-52-kD Ro and anti-smooth-muscle antibodies developed lung fibrosis. No patient in the other cirrhosis group had SS or its characteristic autoantibody findings. CONCLUSIONS: It was suggested that PBC and SS are frequently associated. Anti-52-kD Ro antibodies seem to be a characteristic serologic finding for SS in PBC, suggesting their pathogenic role in autoimmune sialadenitis.

Detection of autoantibodies to Ro(SS-A), La(SS-B) and U1RNP in different congenital heart rhythm disorders using immunoblot and enzyme immunoassay.

The aim of the study was to determine if autoantibodies have distinctive qualitative or quantitative characteristics in children with congenital heart rhythm disorders compared to unaffected infants. Sera of fifty-three infants with different heart rhythm disorders and eight unaffected children born to SLE mothers were tested for antibodies to Ro(SS-A), La(SS-B) and U1RNP (including recombinant A-, C-, 68 kD-proteins) using enzyme immunoassays and immunoblot. 21 sera from affected and 8 sera from healthy children reacted in counter-immunoelectrophoresis with Ro(SS-A). 8/8 infants with third degree atrioventricular (AV)-block, 4/8 with first degree AV-block, one with sinusbradyarrhythmia (SB) and another with SB/first degree AV-block were anti-52 kD Ro(SS-A) positive compared to 7/8 healthy infants. Infants with third degree AV-block had significantly higher anti-52 kD Ro(SS-A) levels than those with the milder heart rhythm disorders (P < 0.01) and the healthy group (P < 0.002). Anti-La(SS-B) antibodies were detected significantly (P < 0.01) and in significantly higher titres (P < 0.01) in AV III than in other types of congenital heart block. Anti-U1RNP-68 kD antibodies were positive in 3/8 healthy, 2/8 first degree AV-block and in 1/8 third degree AV-block cases. Third degree AV-block was more frequently found in female infants (P < 0.05). It was concluded that the influence of antibody levels to the 52 kD Ro(SS-A) and La(SS-B) components and the association of anti-52 kD Ro(SS-A), La(SS-B) and -60 kD Ro(SS-A) antibodies, as well as the sex, on the severity of congenital AV-conduction defects suggests involvement of these factors in the pathogenesis of these disorders.