Inhibition of ASM activity ameliorates DSS-induced colitis in mice.

Acid sphingomyelinase (ASM) is a membrane lipid hydrolase, acting to generate ceramide and regulate cell functions and inflammatory responses.The roles of ASM in mediating T cell functions are postulated whereas its function in regulation of macrophages remains uncertain. The study was performed to explore ASM activity in control of macrophage functions. RAW 264.7 cells were pretreated with desipramine, an ASM inhibitor, prior to LPS challenge in vitro. LPS initiated ASM activity in RAW 264.7 cells. Conversely, inhibition of ASM activity by desipramine diminished LPS induced ASM activities and TNF production of RAW 264.7 cells. The DSS colitis in mice was induced, and desipramine was administered to the mice two days post induction of colitis. Murine colitis was characterized by elevation of ASM activities in colon tissues. Desipramine administration overrode ASM activities in colon, and ameliorated DSS-induced colitis evidenced with the reduced disease activities and the decreased cytokine levels. Together, our data show a crucial role of ASM activity in regulation of macrophage functions and responses, and suggest that ASM represents a novel therapeutic approach for the management of immune diseases.

Inhibition of NADPH oxidase activities ameliorates DSS-induced colitis.

NADPH oxidases (NOX) act to generate reactive oxygen species (ROS) and exhibit microbicidal bioactivity, whereas their roles in mediating immune responses of inflammation in intestine remain to be further elucidated. The study was performed to explore the effects of NOX activity on regulation of macrophage functions. Macrophage responses were induced by lipopolysaccharides (LPS) in RAW 264.7 cells (in vitro) or dextran sulfate sodium (DSS) in BALB/c mice (in vivo) respectively. LPS induced NOX2 expression and initiated NOX activities in RAW 264.7 cells. Conversely, inhibition of NOX activity by DPI and VAS2870 diminished LPS induced NOX activities and the downstream signaling in RAW 264.7 cells. Murine colitis was characterized by macrophage accumulation and elevation of NOX activities in colon tissues. DPI and VAS2870 administration overrode NOX activities and ROS productions in colon tissues, and ameliorated DSS-induced colitis evidenced with the reduced disease activities and the decreased cytokine levels. Intriguingly, NOX2 expression levels were elevated in colon tissues of patients with active inflammatory bowel disease. Together, our data show a crucial role of NOX activity in regulation of macrophage functions and responses, and suggest that NOX represents a novel therapeutic approach for the management of immune diseases.

Reactions of peroxynitrite and nitrite with organic molecules and hemoglobin.

In this work, the reactions of nitrite (NO2-) and peroxynitrite (ONOO-) with organic molecules as well as with hemoglobin (Hb) were examined and the potential interference with the detection of hydrogen peroxide and Hb was investigated. ONOO- at low concentrations (35-140 microM) induced a concentration-dependent oxidation of o-phenylenediamine and guaiacol, and this process can be improved by the addition of Hb in a concentration-dependent manner. This enhancing effect of Hb was possibly due to the formation of such highly reactive species as ferrylHb during the reaction of ONOO- and Hb. NO2- also oxidized the aromatic amine o-phenylenediamine, but its efficiency was much lower than that of ONOO-. A 300-fold excess of NO2- over hydrogen peroxide inhibited the oxidation of Pyrogallol Red mediated by hydrogen peroxide and Hb, which was due in part to the reaction of NO2- with Hb ferryl species compound I and compound II and the phenoxyl radical. These data suggest that ONOO- and NO2- can interfere with the detection of hydrogen peroxide. The overestimation or underestimation of the hydrogen peroxide detected is dependent upon the organic molecule utilized for detection and the relative rate of NO2-, superoxide, and ONOO- generation.