## Involvement of Nitrogen Species

NO reacts rapidly with ferrous iron, and at physiological concentrations, NO also binds to soluble guanylate cyclase and to another hemoprotein, cytochrome c oxidase (Complex IV), the terminal enzyme of the mitochondrial respiratory chain. NO can therefore control cellular functions via the reversible inhibition of respiration. There are a number of reactive NO species, such as N2O3 and ONOO- that can also alter critical cellular components. During the first hours after injury, iNOS-mediated NO production is upregulated, producing a burst of NO that far exceeds basal levels. This overabundance of NO produces significant cellular injury via several mechanisms.

- (1) NO may directly promote overwhelming peripheral vasodilation, resulting in vascular decomposition;
- (2) NO may upregulate the transcription NF-kB initiating an inflammatory signaling pathway that, in turn, triggers numerous inflammatory cytokines.
- (3) NO also interacts with the O-2 to yield ONOO-, a highly reactive compound that exacerbates the injury produced by either O-2 alone or NO alone.

The ONOO- generation which occurs during fluid resuscitation in the injured subject produces cellular death by enhancing DNA single strand breakage, activates the nuclear enzyme polyADP ribose synthetase (PARS), leading to cellular energy depletion and cellular necrosis. The detrimental effects of ONOO- in shock and resuscitation have been attributed to oxidation of sulfhydryl groups, the nitration of tyrosine, tryptophane, and guanine, as well as inhibition of the membrane sodium-potassium adenosine triphosphatase. PARS activation depletes NAD and thus alters electron transport, ATP synthesis, and glycolysis; and leads to DNA fragmentation and cellular apoptosis. The activation of monocytes, macrophages and endothelial cells by LPS results in the expression of iNOS, and consequently increases the transformation of L-arginine to NO, which can combine with O2- to form ONOO-, causing tissue injury during shock, inflammation and ischemia reperfusion. NO stimulates H2O2 and O-2 production by mitochondria, increasing leakage of electrons from the respiratory chain. H2O2, in turn, participates in the upregulation of iNOS expression via NFkB activation. ONOO- has been shown to stimulate H2O2 production by isolated mitochondria. On the other hand, NO can decrease ROS- produced damage that occurs at physiological levels of NO. The high reactivity of NO with radicals might be beneficial in vivo by scavenging peroxyl radicals and inhibiting peroxidation. ONOO- may also be a signal transmitter and can mediate vasorelaxation, similarly to NO. Local generation of RNS contributes to tissue injury. Recent studies have demonstrated that activation of the nuclear enzyme poly(ADP-ribose) polymerase-1 by RNS-mediated DNA damage is an important pathway of tissue injury in conditions associated with oxidative stress. Increased formation of RNS in response to endotoxin challenge is organ specific. In sepsis, NO may exert direct and indirect effects on cardiac function. Sustained generation of NO occurs in systemic inflammatory reactions, such as septic shock with involvement in circulatory failure.