Demonstration of nitric oxide synthase activity in crustacean hemocytes and anti-microbial activity of hemocyte-derived nitric oxide.
We determined the biochemical characteristics of nitric oxide synthase (NOS) in hemocytes of the crayfish Procambarus clarkii and investigated the roles of hemocyte-derived NO in host defense. Biochemical analysis indicated the presence of a Ca2+ -independent NOS activity, which was elevated by lipopolysaccharide (LPS) treatment. When bacteria (Staphylococcus aureus) and hemocytes were co-incubated, adhesion of bacteria to hemocytes was observed. NO donor sodium nitroprusside (SNP) significantly increased the numbers of hemocytes to which bacteria adhered. Similarly, LPS elicited bacterial adhesion and the LPS-induced adhesion was prevented by NOS inhibitor NG-monomethyl-L-arginine (L-NMMA). Finally, plate count assay demonstrated that addition of LPS to the hemocytes/bacteria co-incubation resulted in a significant decrease in bacterial colony forming unit (CFU), and that L-NMMA reversed the decreasing effect of LPS on CFU. The combined results demonstrate the presence of a Ca2+ -independent LPS-inducible NOS activity in crayfish hemocytes and suggest that hemocyte-derived NO is involved in promoting bacterial adhesion to hemocytes and enhancing bactericidal activity of hemocytes.
Yeh
Feng-Ching
FC
Department of Biology, National Changhua University of Education, Changhua, Taiwan 50058, Republic of China.
Wu
Su-Hua
SH
Lai
Chi-Yung
CY
Lee
Chi-Ying
CY
D000818:Animals; D000890:Anti-Infective Agents / Q000494:pharmacology*; D003400:Astacoidea / Q000201:enzymology*; D002118:Calcium / Q000378:metabolism; D002448:Cell Adhesion / Q000187:drug effects; D003114:Colony-Forming Units Assay; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006434:Hemocytes / Q000201:enzymology*; D008070:Lipopolysaccharides / Q000494:pharmacology; D009569:Nitric Oxide / Q000494:pharmacology*; D020030:Nitric Oxide Donors / Q000494:pharmacology; D019001:Nitric Oxide Synthase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D009599:Nitroprusside / Q000494:pharmacology; D013211:Staphylococcus aureus / Q000187:drug effects* / Q000254:growth & development; D019323:omega-N-Methylarginine / Q000494:pharmacology
D000890:Anti-Infective Agents; D004791:Enzyme Inhibitors; D008070:Lipopolysaccharides; D020030:Nitric Oxide Donors; D009599:Nitroprusside; D019323:omega-N-Methylarginine; D009569:Nitric Oxide; D019001:Nitric Oxide Synthase; D002118:Calcium