Inhibition of the conversion of pre-interleukins-1α and 1β to mature cytokines by p-benzoquinone, a metabolite of benzene

Abstract

Chronic exposure of humans to benzene causes severe bone marrow cell depression leading to aplastic anemia. Marrow stromal macrophage dysfunction and deficient interleukin-1 production has been reported for patients with severe aplastic anemia. The stromal macrophage, a target of benzene toxicity, is involved in hematopoietic regulation through the synthesis of several cytokines including interleukin-1, which is required for production by stromal fibroblasts of a number of cytokines required for the survival of hematopoietic progenitor cells. We have previously demonstrated that hydroquinone, a major toxic metabolite of benzene in marrow, prevents the proteolytic conversion of 31 kDa pre-interleukin-1α to the 17 kDa cytokine by calpain in purified murine stromal macrophages. Furthermore, stromal macrophages from benzene-treated mice produce the 31 kDa pre-interleukin-1α when stimulated in culture with endotoxin, but cannot convert the precursor to interleukin-1α. In this report, we show that 1,4-benzoquinone, the oxidation product of hydroquinone in the cell, causes a concentration-dependent inhibition of highly purified human platelet calpain with an IC 50 of 3 μM. Hydroquinone also inhibits the processing of pre-interleukin-1β by interleukin-1β convertase. The addition of 2 μM hydroquinone to B1 cells that undergo autocrine stimulation by interleukin-1β resulted in the cessation of autocrine cell growth and interleukin-1β secretion into the culture medium, as determined by Western immunoblots of the culture supernatants. Purified converting enzyme treated with 3 μM benzoquinone was incapable of converting 31 kDa recombinant pre-interleukin-1β to the 17 kDa mature cytokine as analyzed by polyacrylamide gel electrophoresis and Western immunoblotting. These findings support our observations in a mouse model that benzene-induced bone marrow cell depression results from a lack of interleukin-1α subsequent to an inhibition by benzoquinone of calpain, the protease required for converting pre-interleukin-1α to active cytokine. The results may provide a basis for studying benzene-induced aplastic anemia in a mouse model.