calpain and quinone

Chronic exposure of humans of benzene affects hematopoietic stem and progenitor cells and leads to aplastic anemia. The stromal macrophage, a target of benzene toxicity, secretes interleukin-1 (IL-1), which induces the stromal fibroblast to synthesize hematopoietic colony-stimulating factors. In a mouse model, benzene causes an acute marrow hypocellularity that can be prevented by the concomitant administration of IL-1 alpha. The ability of benzene to interfere with the production and secretion of IL-1 alpha was tested. Stromal macrophages from benzene-treated mice were capable of the transcription to the IL-1 alpha gene and the translation of the message but showed an inability to process the 34-kDa pre-IL-1 alpha precursor to the 17-kDa biologically active cytokine. Treatment of normal murine stromal macrophages in culture with hydroquinone (HQ) also showed an inhibition in processing of pre-IL-1 alpha. Hydroquinone is oxidized by a peroxidase-mediated reaction in the stromal macrophage to p-benzoquinone, which interacts with the sulfhydryl (SH) groups of proteins and was shown to completely inhibit the activity of calpain, the SH-dependent protease that cleaves pre-IL-1 alpha. In a similar manner, HQ, via peroxidase oxidation to p-benzoquinone, was capable of preventing the IL-1 beta autocrine stimulation of growth of human B1 myeloid tumor cells by preventing the processing of pre-IL-1 beta to mature cytokine. Benzoquinone was also shown to completely inhibit the ability of the SH-dependent IL-1 beta converting enzyme. Thus benzene-induced bone marrow hypocellularity may result from apoptosis of hematopoietic progenitor cells brought about by lack of essential cytokines and deficient IL-1 alpha production subsequent to the inhibition of calpain by p-benzoquinone and the prevention of pre-IL-1 processing.

Topics: Animals; Benzene; Benzoquinones; Bone Marrow; Bone Marrow Cells; Calpain; Caspase 1; Cell Line; Cysteine Endopeptidases; Enzyme Inhibitors; Humans; Indomethacin; Interleukin-1; Mice; Protein Precursors; Protein Processing, Post-Translational; Recombinant Proteins

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-1 alpha 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-1 alpha when stimulated in culture with endotoxin, but cannot convert the precursor to interleukin-1 alpha. 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 IC50 of 3 microM. Hydroquinone also inhibits the processing of pre-interleukin-1 beta by interleukin-1 beta convertase. The addition of 2 microM hydroquinone to B1 cells that undergo autocrine stimulation by interleukin-1 beta resulted in the cessation of autocrine cell growth and interleukin-1 beta secretion into the culture medium, as determined by Western immunoblots of the culture supernatants. Purified converting enzyme treated with 3 microM benzoquinone was incapable of converting 31 kDa recombinant pre-interleukin-1 beta 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-1 alpha subsequent to an inhibition by benzoquinone of calpain, the protease required for converting pre-interleukin-1 alpha to active cytokine. The results may provide a basis for studying benzene-induced aplastic anemia in a mouse model.

Topics: Benzene; Benzoquinones; Blotting, Western; Calpain; Caspase 1; Cell Division; Cysteine Endopeptidases; Cysteine Proteinase Inhibitors; Humans; Hydroquinones; Indomethacin; Interleukin-1; Macrophages; Protein Precursors; Recombinant Proteins; Tumor Cells, Cultured