Nitric oxide is a highly reactive molecule that has been implicated in host defense and tissue injury. In the present studies, we determined whether rat type II alveolar epithelial cells have the capacity to produce this mediator. We found that type II cells synthesize significant quantities of nitric oxide after treatment with the inflammatory cytokines, interferon-gamma (IFN-gamma) and/or interleukin-1 beta (IL-1 beta), or with the combination of IFN-gamma and tumor necrosis factor-alpha. In contrast to rat alveolar macrophages, type II cells were unresponsive to lipopolysaccharide. Production of nitric oxide by type II cells in response to IFN-gamma was dose dependent, reaching a maximum at 100 U/ml, and blocked by NG-monomethyl-L-arginine (L-NMA), a nitric oxide synthase inhibitor. Northern blot analysis demonstrated that nitric oxide production by type II cells was due to expression of mRNA for an inducible form of nitric oxide synthase (iNOS). Following brief exposure of rats to irritant-inducing doses of ozone (2 ppm, 3 h), type II cells were found to produce significantly more nitric oxide than were cells from control animals. This was due to increased expression of iNOS mRNA. Cells from ozone-treated rats were also sensitized to produce more nitric oxide in response to IFN-gamma and IL-1 beta. This was associated with a marked increase in expression of iNOS mRNA and enzyme protein in the cells. We also found that ozone inhalation caused enhanced production of hydrogen peroxide, as well as spontaneous and IFN-gamma-induced cytostasis of type II cells toward P815 mouse mastocytoma cells.(ABSTRACT TRUNCATED AT 250 WORDS)

Benzene is a widely used industrial solvent known to cause bone marrow depression. This is associated with increased production of reactive oxygen metabolites and nitric oxide by bone marrow phagocytes, which have been implicated in hematotoxicity. Benzene metabolism to phenolic intermediates appears to be an important factor in bone marrow toxicity. In the present studies, we compared the effects of benzene and several of its metabolites on nitric oxide production by murine bone marrow leukocytes. Bone marrow cells readily produced nitric oxide in response to the inflammatory mediators lipopolysaccharide (LPS) and interferon-gamma (IFN-gamma). Treatment of mice with benzene (800 mg/kg), or its metabolites hydroquinone (100 mg/kg), 1,2,4-benzenetriol (25 mg/kg), or p-benzoquinone (2 mg/kg), at doses that impair hematopoiesis, sensitized bone marrow leukocytes to produce increased amounts of nitric oxide in response to LPS and IFN-gamma. Granulocyte-macrophage colony-stimulating factor (GM-CSF) and macrophage colony-stimulating factor (M-CSF) augmented bone marrow leukocyte production of nitric oxide induced by inflammatory mediators. Benzene, as well as its metabolites, markedly increased the sensitivity of the cells to both GM-CSF and M-CSF. Cells from hydroquinone- or 1,2,4-benzenetriol-treated mice were significantly more responsive to the inflammatory cytokines and growth factors than cells isolated from benzene- or p-benzoquinone-treated mice, suggesting that the phenolic metabolites of benzene are important biological reactive intermediates. Because nitric oxide suppresses cell growth and can be metabolized to mutagens and carcinogens, the ability of benzene and its metabolites to modulates its production in the bone marrow may be important in their mechanism of action.

The human monoblast leukemia line, U937, is growth-inhibited and induced to develop markers of mature monocytes by lymphokine preparations. Lymphokine is cytostatic and induces expression of Fc receptors in U937 and in myelomonocytic leukemic lines RC-2A and KG-1, but does not have these effects on T- and B-lymphocytic lines. In addition to previously described properties, including complement receptors, phagocytosis, and antibody-dependent cellular cytotoxicity (ADCC), Mac-1 and Mac-3 surface antigens defined by monoclonal antibodies are induced on U937 cells by lymphokine and phorbol ester. The Mac-1 surface component appears to have a regulatory role in differentiation of the monocyte lineage line, since antibodies to this antigen block the induction of Mac-3 antigen. The lymphokine activity was concentrated by salt precipitation and characterized by ion- exchange and size chromatography. Fractions of about 40,000 daltons were responsible for growth inhibition and induction of Fc receptors and Mac-1 antigen in U937 cells. However, ADCC was not induced in U937 by individual fractions of lymphokine, suggesting that this cytotoxic capacity may be regulated by a lymphokine of a different size, which is only effective after initial maturation steps. Since gamma-interferon is present on the 40K size range of lymphokine, the possibility that interferon is a differentiation modulator for the monoblast cells was investigated. Highly purified gamma-interferon (10(7) U/mg protein) at 10–300 U/ml inhibited growth and induced Fc receptors in U937 similar to the effect of lymphokine. The Fc-receptor-inducing activity of lymphokine was inhibited by a neutralizing monoclonal antibody to gamma- interferon, suggesting that this differentiation factor in lymphokine is gamma-interferon.