NET INFLAMMATORY CAPACITY OF HUMAN SEPTIC SHOCK PLASMA EVALUATED BY AMONOCYTE-BASED TARGET-CELL ASSAY - IDENTIFICATION OF INTERLEUKIN-10 AS A MAJOR FUNCTIONAL DEACTIVATOR OF HUMAN MONOCYTES

We have developed a functional assay to study the inflammatory capacit y of plasma collected from patients with severe gram-negative sep:ic s hock. In this assay, elutriation-purified, cryopreserved human monocyt es from one healthy donor are combined with plasma from patients with severe persistent septic shock for 5 h. Subsequently, the plasma is re moved, medium added, and procoagulant activity (PCA) and secretion of tumor necrosis factor or (TNF-alpha) and interleukin 6 (IL-6) measured after 18-h incubation. Plasma from 10 patients (6 died) infected with Neisseria meningitidis previously shown to contain high levels of nat ive lipopolysaccharide (LPS) (median 2,700 pg/ml), TNF-alpha, IL-6, IL -8, and complement activation products, had a low net spontaneous infl ammatory capacity on the monocytes. The median levels of PCA, TNF-alph a, and IL-6 were 5, 0, and 4%; respectively, of the monocyte activitie s induced by normal plasma boosted with purified N. meningitidis (Nm)- LPS (2,500 pg/ml; net LPS-boosted capacity, 100%). The levels of PCA, TNF-alpha, and IL-6 obtained with plasma from shock patients were not different from those induced by plasma from 10 meningococcal patients without shock or with plasma from healthy persons. Boosting shock plas ma with 2,500 pg/ml Nm-LPS had little effect on the monocyte activitie s since the median values of PCA, TNF-alpha, and IL-6 revealed a minim al increase from 5, 0, and 4% to 9, 2, and 6%, respectively. The shock plasmas revealed a strong LPS-inhibitory capacity that was largely ab sent in plasmas from 10 meningococcal patients without shock since the median levels of PCA, TNF-alpha, and IL-6 increased from 5, 0, and 0% to 135, 51, and 73%, respectively, after boosting with 2,500 pg/ml Nm -LPS. The LPS-inhibitory capacity was closely associated with the leve ls of IL-10. The median levels of IL-10 were 19,000 pg/ml in nine shoc k patients vs. 22 pg/ml in nine nonshock patients with systemic mening ococcal disease. Removal of native IL-10 by immunoprecipitation restor ed the capacity of plasmas to induce monocyte activation either by nat ive LPS or by boosting with Nm-LPS. IL-4 and TGF-beta were not detecte d in shock plasmas. In 24 patients with detectable meningococcal LPS ( > 10 pg/ml, 0.1 endotoxin units/ml, the levels of IL-10 were correlat ed to the levels of LPS (r = 0.79, P < 0.001). IL-10 declined from ini tiation of antibiotic therapy and paralleled the levels of native LPS. Decreasing levels of IL-10 in serially collected shock plasmas were d irectly related to increasing monocyte responsiveness after Nm-LPS boo sting. These results suggest that IL-10 plays a major role in containi ng activation of monocytes and possibly other LPS-responsive cells dur ing overwhelming meningococcemia.