No abstract

Variant forms of human a-1-proteinase inhibitor (a-l-PI), obtained by the treatment of human Hep G2 cells with specific inhibitors of glycosylation were tested for both inhibitory activity and heat stability. All were found to have the same second-order association rate with human neutrophil elastase, indicating a lack of importance. of the carbohydrate moiety. In contrast, incompletely glycosylated forms of a-l-PI were found to be heat sensitive relative to the mature protein, suggesting a role for carbohydrate in protein stabilization.a-1-Proteinase inhibitor; Inhibitor of glycosylation; Hep G 2 cell

We have previously described inhibition of the synthesis of three acute-phase inflammatory cytokines in human and rat macrophages by acetate esters of rooperol, a dicatechol of plant origin. Analysing the mechanism of anticytokine activity of rooperol, we compared levels of TNFalpha, IL-1beta and IL-6 mRNAs in the human promonocytic U937 cell line pretreated with phorbol myristate acetate (PMA) and incubated with rooperol tetraacetate (RTA) alone or in combination with LPS (500 ng/ml). It was found that 10 microM RTA decreased the levels of cytokine mRNAs both in the presence and absence of LPS, suggesting pretranslational inhibition of cytokine synthesis. Electrophoretic mobility shift analysis (EMSA) showed that RTA may influence cytokine mRNA expression by decreasing the binding activity of transcription factors NF-kappaB and AP-1.

Luminol-enhanced chemiluminescence was measured in fresh whole human blood, or human neutrophils isolated from heparinized blood, human alveolar macrophages and rat alveolar macrophages stimulated with bacterial endotoxin (LPS). Tetraacetate esters of rooperol, a dicatechol showing anticytokine activity, added to cells simultaneously with LPS inhibited the respiratory burst. The effective concentrations of rooperol were in the range of 1-10 μM depending on cell type and corresponded well with inhibition of nitric oxide production by rat alveolar macrophages. Thus rooperol may reduce some effects of excessive phagocytic activity and inflammatory reaction but by quenching free radicals production may also diminish the resistance to bacterial infections.