Stimulation of NO Synthase Activity in the Immune-Competent LepidopteranEstigmene acraeaHemocyte Line

Abstract

In contrast to the vertebrate immune system, nearly nothing is known about the immunological role of nitric oxide (NO) in invertebrates. This study provides evidence of the presence of a NO synthase (NOS) activity in an immune-competent, macrophage-like insect hemocyte line, previously established from larvae of the lepidopteran insectEstigmene acraea.As proven by photometric determination of nitroblue tetrazolium reduction after cell fixation, theE. acraeacells possess NADPH diaphorase (NADPHd) activity. This NADPH diaphorase activity was NADPH dependent, not inhibitable by superoxide dismutase, influenced by extracellular addition ofl-arginine, and inhibited in a dose-dependent manner by the specific NOS inhibitorNω-monomethyl-l-arginine. Furthermore, the NADPH diaphorase activity was stimulated within 30 min by the addition of insect pathogenic bacteria (Bacillus thuringiensisvar.kurstaki, Photorhabdus luminescens), bacterial lipopolysaccharide, and silica beads. In activatedE. acraeacell suspensions strongly increased amounts ofl-citrulline and enhanced levels of total nitrite/nitrate (as NO derivates) can be determined. This is the first report on stimulable NOS activity in insect hemocytes.