Figure 4.

 Tetracycline (TET) inhibits IL-1β production in vitro via selective inhibition of caspase-1 (Casp-1). Murine bone marrow–derived macrophages (BMDM) or immortalized murine BMDM overexpressing a fluorescently tagged ASC (apoptosis-associated speck-like protein containing a CARD domain)-mCerulean fusion protein were stimulated with either LPS (30 ng/ml) alone or in combination with nigericin (10 mM) or Poly (dA:dT) (0.5 μg/ml) and then treated with TET. (A and B) IL-1β, (C) TNF-α, and (D) LDH concentrations were measured in supernatants with ELISA or LDH assay. ASC speck formation was measured by fluorescence microscopy. (E) Six different images were taken of each condition at a magnification of ×40. (F) Immunoblots of lysates (Lys.) and supernatants (Sup.) of wild-type BMDM. Representative blots from four independent experiments. (G) Active recombinant Casp-1 was incubated with its substrate and TET. Reduction of recombinant Casp-1 activity was measured in relation to control without TET. Median with interquartile range of three or more independent experiments. *P < 0.05; Mann-Whitney test.