Discovery may provide a new target to treat sepsis

Part of the immune system's pro-inflammatory response to bacterial invasion is to increase nitric oxide levels with an enzyme called inducible nitric oxide synthase.

In a study published in the Journal of Biological Chemistry, scientists report that the predominantly anti-inflammatory enzyme, endothelial nitric oxide synthase, is also involved in nitric oxide production in response to infection. This discovery may eventually provide a new target to treat sepsis, which is caused by overproduction of nitric oxide.

The research appears as the "Paper of the Week" in the March 18 issue of the Journal of Biological Chemistry, an American Society for Biochemistry and Molecular Biology journal.

When immune cells are exposed to pro-inflammatory cytokines or bacterial endotoxin (part of the bacterial cell wall) they start to produce inducible nitric oxide synthase (iNOS), an enzyme responsible for the manufacture of nitric oxide (NO). This results in an increase in cellular NO which contributes to inflammation and host defense.

"NO acts as a cytotoxic/cytostatic effector molecule released (predominantly) by immune cells," explains Dr. Adrian J. Hobbs of University College London. "It kills pathogens via a variety of mechanisms, mostly related to inhibition of metabolic enzymes and destruction of DNA."

However, too much NO can be a bad thing. Sustained overproduction of NO can cause septic shock (sepsis). "In sepsis, which is a systemic bacterial infection, the body expresses iNOS which generates relatively high concentrations of NO," says Dr. Hobbs. "This aids host defense by killing the invading organism, but in excessive quantities starts to lead to host-damage. In sepsis, this is manifested predominantly as a profound hypotension, inadequate tissue perfusion and organ failure. This often results in death."

Previously, Dr. Hobbs and colleagues demonstrated in vitro that endothelial nitric oxide synthase (eNOS) also plays a pro-inflammatory role by facilitating iNOS expression. "eNOS is found almost exclusively in the vascular endothelium and the NO that it synthesizes plays a key role in regulation of blood pressure, platelet aggregation, the reactivity of immune cells and growth of vascular smooth muscle cells," explains Dr. Hobbs. "iNOS is not expressed under normal physiological conditions, but is up-regulated for host-defense purposes."

Now, the researchers have validated their hypothesis in vivo using mice that do not produce eNOS. These mutant mice had a marked reduction in iNOS production in response to bacterial endotoxin, as well as lower plasma levels of NO2- and NO3- and less mortality than normal mice. The scientists also showed that endotoxin activates eNOS in macrophages and that this effect is an essential trigger for the induction of iNOS.

"eNOS has until recently been thought to act principally in an anti-inflammatory manner," notes Dr. Hobbs. "The results of our study show clearly that eNOS can also act in a pro-inflammatory manner and accelerate host-defense in response to pathogenic stimuli."

This discovery may eventually lead to new treatments for septic shock and other inflammatory diseases. "Pharmaceutical companies have been developing iNOS inhibitors to treat sepsis," explains Dr. Hobbs. "However, it now appears as if these are ineffective in reducing the mortality associated with the disease. The identification of a pro-inflammatory role for eNOS-derived NO may provide the stimulus for further research in this area and thereby identify novel targets for treatment of inflammatory diseases."