Chloroquine inhibits production of TNF-alpha, IL-1beta and IL-6 from lipopolysaccharide-stimulated human monocytes/macrophages by different modes.
OBJECTIVE
TNF-alpha, IL-1 and IL-6 are known to have primary roles in the pathogenesis of rheumatoid arthritis and other inflammatory diseases. The anti-rheumatic drug chloroquine has been shown to inhibit TNF-alpha, IL-1 and IL-6 production from mononuclear phagocytes. We examined the underlying mechanisms involved in the chloroquine-induced inhibition of cytokine production.


METHODS
Human peripheral blood mononuclear cells and monocytes/macrophages and monocytic U-937 and THP-1 cells were stimulated with lipopolysaccharide, and TNF-alpha, IL-1beta and IL-6 production was measured by ELISA. Levels of mRNA were measured by northern blotting and reverse transcription-polymerase chain reaction. Synthesis of 26-kDa TNF-alpha precursor was measured by metabolic labelling and immunoprecipitation analysis. Transcription rate was determined by nuclear run-on assay.


RESULTS
TNF-alpha release from the cells was inhibited by chloroquine, whereas the steady-state level of TNF-alpha mRNA and synthesis of 26-kDa TNF-alpha precursor were not changed by chloroquine. In contrast, chloroquine-induced inhibition of IL-1beta and IL-6 release was accompanied by a decrease in their steady-state mRNA levels. The transcription rates of the IL-1beta and IL-6 genes were not changed by chloroquine, whereas the stability of IL-1beta and IL-6 mRNA was decreased by chloroquine. Weak-base amines such as methylamine and ammonium chloride had no effect on the production of TNF-alpha, whereas they partially blocked the production of IL-1beta and IL-6.


CONCLUSIONS
Our results indicate that chloroquine-mediated inhibition of TNF-alpha, IL-1beta and IL-6 synthesis occurs through different modes in lipopolysaccharide-stimulated human monocytes/macrophages: it blocks the conversion of cell-associated TNF-alpha precursor to mature soluble protein, whereas it reduces the levels of IL-1beta and IL-6 mRNA, at least in part, by decreasing their stability and by a pH-dependent mechanism.
Jang
C-H
CH
Department of Biochemistry, College of Medicine, Catholic University of Korea, 505 Banpo-dong, Seocho-gu, Seoul 137-701, South Korea.
Choi
J-H
JH
Byun
M-S
MS
Jue
D-M
DM
D018501:Antirheumatic Agents / Q000494:pharmacology*; D002478:Cells, Cultured; D002738:Chloroquine / Q000494:pharmacology*; D016207:Cytokines / Q000096:biosynthesis* / Q000235:genetics; D004305:Dose-Response Relationship, Drug; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D007375:Interleukin-1 / Q000096:biosynthesis / Q000235:genetics; D015850:Interleukin-6 / Q000096:biosynthesis / Q000235:genetics; D008070:Lipopolysaccharides / Q000276:immunology; D008264:Macrophages / Q000187:drug effects / Q000276:immunology; D009000:Monocytes / Q000187:drug effects* / Q000276:immunology; D012333:RNA, Messenger / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D014407:Tumor Cells, Cultured; D014409:Tumor Necrosis Factor-alpha / Q000096:biosynthesis / Q000235:genetics
D018501:Antirheumatic Agents; D016207:Cytokines; D007375:Interleukin-1; D015850:Interleukin-6; D008070:Lipopolysaccharides; D012333:RNA, Messenger; D014409:Tumor Necrosis Factor-alpha; D002738:Chloroquine