Abstract
Cold-reactive lymphocytotoxin (CLT) was studied in 40 patients (active sarcoidosis 34 cases, resolved sarcoidosis 6 cases) and 12 patients with systemic lupus erythematosus (SLE). In this paper we discussed (i) the difference in the incidence, (ii) the difference of the cytotoxic cellular specificities and (iii) the immunological and in vivo significance of CLT between sarcoidosis and SLE. In addition we tried to purify the CLT from the sera of patients with sarcoidosis and SLE by applying fractionation and chromatography.
Peripheralblood lymphocytes (PBL) obtained from normal healthy volunteers were isolated by centrifugation on a Ficoll-Paque gradient. E-rosette forming cells were used as enriched T-lymphocytes. Cells which did not form E-rosettes after two attempts were used as enriched B-lymphocytes. By the methods of salt fractionation, Sephadex G-200gel filtration, DEAE-cellulose chromatography and IgM-affinity chromatography lymphocytotoxin in the sarcoidosis sera were purified. The lymphocytotoxicity was investigated by a complement-dependent 51Cr-release cytotoxicity assay according to the method of Brünner et al. Lymphocytotoxic activity was expressed as a percentage by the formula
experimental 51Cr release-spontaneous release/maximal release-spontaneous release ×100
CLT activity was present in 19 out of 34 active sarcoidosis patients (55%) and in 12 out of 12 SLE patients (100%), but was absent in all 6 resolved sarcoidosis patients and in 19 out of 20 healthy control subjects (5%). In sarcoidosis, CLT activity did not correlate with age or sex, degree of lymphopenia, presence of anergy, or with other clinical parameters, but was correlated with the activity of the disease. In addition, CLT activity was greater at 15°C and negligible at 4°C and 37°C. To clarify the cellular specificity of CLT to T- and B-lymphocyte subpopulation, the sera from 10 sarcoidosis patients, 3 SLE patients and 4 healthy control subjects were examined. All sera from sarcoidosis were cytotoxic to B-lymphocytes, although there were certain sera that were reactive to B-lymphocytes and to a minor proportion of T-lymphocytes. All sera from SLE were cytotoxic to both T- and B-lymphocytes, but these sera had higher reactivity to T-lymphocytes than to B-lymphocytes. None of the sera from healthy control subjects were reactive to either T- or B-lymphocytes. The CLT purified from pooled sarcoidosis sera (10 patients with high CLT activity) by applying salt fractionation, Sephadex G-200gel filtration, DEAE-cellulose chromatography and IgM-affinity chromatography was identical with the antibody of IgM class and was distinct from IgG molecules. These purified fractions were cytotoxic to B-lymphocytes but not cytotoxic to T-lymphocytes. The CLT purified from pooled SLE sera (10 patients with high CLT activity) by applying Sephadex G-200 gel filtration and DEAE-cellulose chromatography was IgM antibody and had higher reactivity of T-lymphocytes than to B-lymphocytes.
