Induction of Inducible Nitric Oxide (NO) Synthase mRNA and NO Production in Macrophages Infected with Influenza A/PR/8 Virus and Stimulated with Its Ether‐Split Product

Abstract

We investigated the inductive activity of infective influenza A/PR/8/34 (PR8) virus and its ether‐split product (ESP) on the expression of inducible nitric oxide (NO) synthase (iNOS) and NO production in RAW264.7 (RAW) cells, a murine macrophage (MΦ) cell line, and thioglycolate‐elicited peritoneal MΦ (TPM). In both cells, PR8 virus infection induced iNOS mRNA between 4 hr and 24 hr, attaining a peak value at 12 hr. In correlation with induction of iNOS mRNA, NO amounts increased significantly from 12 to 24 hr. Moreover, this study demonstrated that ESP with the same hemagglutination titer as PR8 virus could induce iNOS mRNA and NO production, although the inductive activity of ESP was weaker than that of PR8 virus. Considering the dual role (beneficial and detrimental roles) of NO on certain inflammatory disorders and virus infections, the inductive activity of influenza virus on the iNOS‐mediated NO production independent of its infectivity might contribute to a modification of influenza virus infection.

Negative feedback regulation of IL‐32 production by iNOS activation in response to dsRNA or influenza virus infection

“…Increased IL-32 and iNOS expression stimulated by viral infections has been reported in previous investigations [1,5,6]. However, the function of IL-32 in the pro-inflammatory network is still unclear.…”

“…NO is synthesized from L-arginine by NOS in numerous mammalian cells and tissues and plays a critical role in signal modulation during immune responses, chronic inflammation and carcinogenesis. Three isoforms of NOS have been identified until now: two constitutively expressed and calciumdependent isoforms endothelial NOS and neuronal NOS; one inducible and calcium-independent isoform (iNOS), which catalyzes the production of high amount of NO in response to diverse stimuli [4].Increased IL-32 and iNOS expression stimulated by viral infections has been reported in previous investigations [1,5,6]. However, the function of IL-32 in the pro-inflammatory network is still unclear.…”

CEL-I, an N-Acetylgalactosamine (GalNAc)-Specific C-Type Lectin, Induces Nitric Oxide Production in RAW264.7 Mouse Macrophage Cell Line

“…After the incubation for the indicated periods of time, the cells were washed three times with ice cold PBS, and were lysed with 100 l extraction buffer (10 mM HEPES, 150 mM NaCl, 1 mM EGTA, 1% CHAPS, and 1% Triton X-100) containing protease inhibitor cocktail (Nakarai tesque Co., Kyoto, Japan). After shaking for 30 min at 4℃, the cytosolic fraction was obtained from the supernatant after centrifugation at 15,000 rpm for 10 min, and the protein concentrations were determined with the BCA assay kit respectively, were synthesized as described by Imanishi et al (27). Each PCR reaction (25 µl) was run on 2% agarose gels containing 0.1 g/ml ethidium bromide, and the amplified products (231 bp for iNOS) were observed by Light capture (ATTO Co., Tokyo, Japan).…”

Comparative analysis of the production of nitric oxide (NO) and tumor necrosis factor-α (TNF-α) from macrophages exposed to high virulent and low virulent strains of Edwardsiella tarda

“…After the incubation for the indicated time periods, total RNA was isolated from the cells with Trizol reagent Master Mix (Promega), 0.5 mL of forward and reverse primers (1 mM each), 0.5 mL of 1st strand cDNA, and 11 mL of nuclease-free water. The primer sets for iNOS and b-actin were synthesized as described previously [20,21]. Each PCR reaction (25 mL) was run on 2% agarose gels containing 0.1 mg/mL ethidium bromide.…”