SERUM EFFECTS ON MITOGENIC REACTIVITY IN SUBJECTS WITH SYSTEMIC LUPUS-ERYTHEMATOSUS, RHEUMATOID-ARTHRITIS AND SCLERODERMA - TECHNICAL CONSIDERATIONS AND LACK OF CORRELATION WITH ANTI-LYMPHOCYTE ANTIBODIES

Abstract

Methodological problems which affect the assessment of humoral effects on mitogenic reactivity include the source and concentration of serum used to support cell cultures, the day-to-day variability of inhibitory effects and the specific activity of [3H]thymidine added to the culture. These problems were alleviated by addition of half concentration (7.5%) of pooled normal human serum to all cultures, the introduction of anti-lymphocyte serum as a suitable internal control for monitoring the suppressability of lymphocytes and a reduction of specific activity of the [3H]thymidine to 1.3 Ci/mmol. Inhibitory factors were loosely bound to the lymphocyte surface and eluted off after incubation at 37.degree. C for 1 h. Cells from 25 subjects and paired controls were cultured simultaneously in medium containing 15% normal human serum (NHS) or 7.5% patient and 7.5% NHS. The cells were stimulated with various dilutions of phytohemagglutinin (PHA), Con A [concanavalin A] or pokeweed mitogen. Lupus sera suppressed the reactivity of autologous lymphocytes to PHA and pokeweed mitogen. Sera from subjects with RA [rheumatoid arthritis] and scleroderma did not significantly inhibit blastogenesis of autologous lymphocytes. One-half of the lupus sera significantly inhibited the reactivity of homologous lymphocytes to 2 of 3 mitogens. Only 1 of 8 scleroderma sera and none of 12 RA sera had this effect. All patient sera were examined for anti-lymphocyte antibodies by microcytotoxicity and immunofluorescent techniques. These antibodies were usually found in SLE and were often observed in subjects with rheumatoid arthritis but not scleroderma. A firm relationship between serum suppressors of lymphocyte blastogenesis and anti-lymphocyte antibodies was not found.