calcimycin and diacetyldichlorofluorescein

Nitric oxide (NO) is responsible for cytochrome P450 (CYP450) loss during isolation and cytokine treatment of primary rat hepatocytes. As P450s mediate the metabolism of toxic chemicals, their inhibition could compromise the cells competence to eliminate toxins, a condition potentially relevant in neurological diseases involving constitutive activation of nitric oxide synthase (NOS) and NO over-production. Here, we have investigated the correlation between NO accumulation and CYP1A2 down-regulation during maturation of mouse cerebellar granule cells (CGC). As neurons matured in culture, the inducible levels of CYP1A2 protein and catalytic activity decreased to almost undetectable values. In parallel, a significant increase in NO concentration was observed. Neuronal NOS remained constitutively active during maturation, thus contributing to NO accumulation. The NOS inhibitor l-NAME, restored CYP1A2 catalytic activity up to 9 days in vitro, supporting a role for NO in the inhibition process. Maturation was also followed by increased NMDA receptor activity and intracellular Ca2+ concentration. We suggest that maintained NOS activity during CGC maturation could lead to NO accumulation and to decreased CYP1A2 inducibility. Increased NMDA receptor activity and Ca2+ entry could contribute to this process. Thus, neurodegeneration could diminish the induction of specific P450s and impair the metabolism of foreign and/or endogenous chemicals in the CNS.

Topics: Aging; Aniline Compounds; Animals; Animals, Newborn; Calcimycin; Calcium; Cells, Cultured; Cellular Senescence; Cerebellum; Citrulline; Copper Sulfate; Cytochrome P-450 CYP1A2; Dizocilpine Maleate; DNA-Binding Proteins; Down-Regulation; Enzyme Induction; Enzyme Inhibitors; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Fluoresceins; Ionophores; Mice; Mice, Inbred C57BL; N-Methylaspartate; Neurons; NG-Nitroarginine Methyl Ester; Nitric Oxide; Nitric Oxide Synthase; Repressor Proteins; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Time Factors; Tritium; Xanthenes

Defective phagocytic cell function may partially account for the morbidity and mortality associated with thermal injury. In experimental thermal injury in the rat, small circulating blood volumes increase the difficulty in obtaining significant data. Furthermore, purification and or elicitation procedures have the potential for altering the cell surface characteristics and/or the functional response of the cell in question. We have examined the circulating neutrophils and pulmonary alveolar macrophages of anesthetized rats following a 16-20% body surface area scald injury to the shaved back. The circulating neutrophils of thermally injured rats were examined by flow cytometry following stimulation with phorbol myristate acetate (PMA) (100 ng/ml) in terms of the change in fluorescence intensity of the potentiometric cyanine dye, dipentyloxocarboxyanine and the formation of the oxidized product of 2',7'-dichlorofluorescin diacetate-loaded cells. The alveolar macrophages were examined after stimulation with PMA (100 ng/ml) in terms of the change in fluorescence intensity of the potentiometric dye, dipropylthiodicarbocyanine and the generation of superoxide production, as assessed by the superoxide dismutase inhibitable reduction of cytochrome c. Both cells exhibited a profound inhibition of cell function 4 hours after the insult, with partial return toward control values at later time points. Furthermore, the plasma of thermally injured rats, 4 hours after the burn was inhibitory to normal rat neutrophils. Fluorescent compounds suggestive of in vivo lipid peroxidation were maximally detectable at this time point. Further research is needed to establish the role of these products in the induction of phagocytic cell dysfunction.

Topics: Animals; Burns; Calcimycin; Carbocyanines; Flow Cytometry; Fluoresceins; In Vitro Techniques; Leukocyte Count; Macrophages; Neutrophils; Oxygen; Phagocytes; Plasma; Rats; Rats, Inbred Strains; Superoxides; Tetradecanoylphorbol Acetate