Nitric Oxide and Septic Vascular Dysfunction (2000)

Septic shock is characterized by a systemic inflammatory response syndrome, hypotension with vasopressor-resistant systemic vasodilation, and development of multiple organ hypoperfusion and dysfunction (1). Organ failure in sepsis is thought to be mediated by a combination of direct parenchymal cytotoxicity and locally ineffective delivery or use of oxygen and nutrients (2). Sepsis is the leading cause of death in noncoronary intensive care units (1), while infectious disease, in general, is the third leading cause of death (after cardiovascular disease and cancer) in the United States (3). The mortality of patients with septic shock is approximately 50% (4,5), and it is an important cause of early (< 3 days) death in patients with severe sepsis (5). Sepsis leads to elaboration of a self-amplifying cascade of pro- or antiinflammatory cytokines and mediators, including a number of vasoactive substances (prostaglandins, nitric oxide [NO], endothelins, platelet activating factor [PAF], leukotrienes) (1). NO appears to be an important mediator of impaired vascular responsiveness to vasoconstrictor agents in sepsis. Inhibition of NO synthesis improves vasopressor-responsiveness and increases blood pressure (BP) in most septic animal models and in humans; however, animal studies reveal numerous adverse effects of NO synthase (NOS) inhibition (6–10). This article reviews the role of NO in the pathogenesis of septic vascular dysfunction, focusing on the physiologic role of endothelial NO production in regulating blood flow distribution and tissue perfusion and the adverse effects of sepsis and of NOS inhibitors on endothelial function. Contributions of NO to the physiologic regulation of renal function are reviewed, and the renal effects of sepsis and NOS inhibition are discussed in detail. The effects of sepsis and NOS inhibition in the pulmonary and splanchnic circulations are likewise reviewed. We discuss promising data suggesting that selective inhibition of the inducible NOS (iNOS) isoform favorably affects global and regional hemodynamics in animal experiments and improves short-term survival when compared with either standard (catecholamine) or isoform-non-selective NOS inhibitor support. Finally, we conclude that the balance between beneficial and deleterious effects of NO in human sepsis will ultimately determine whether NOS inhibitors become standard components of septic shock therapy. Physiologic Roles of NO The discovery of an endothelium-derived relaxing factor in 1980 and its identification as NO in 1987 together led to widespread interest in NO biology, culminating in the award of the 1998 Nobel Prize in Physiology and Medicine to Drs. Furchgott, Ignarro, and Murad (see references for a detailed review of these experiments) (11–14). NO serves multiple physiologic roles, including neurotransmission (central and peripheral), regulation of tissue perfusion, vascular tone and responsivity, platelet responsivity, renal volume control, and antimicrobial defense (11,14–19). NO is synthesized from the amino acid L-arginine by enzymes of the NOS family. Three NOS isoforms have been identified: neuronal NOS (nNOS), endothelial NOS (eNOS), and iNOS (20). The former two isozymes are constitutively expressed and have physiologic roles; the last is usually present only after the induction by inflammatory stimuli. Most NO in the central nervous system and peripheral nerves is produced by nNOS, mediating the “nitridergic” component of peripheral nonadrenergic, noncholinergic neurotransmission, as well as contributing to central regulation of autonomic outflow to the cardiovascular system (11,15). nNOS is also present in the kidney, predominantly in the macula densa (see below) (21). Tight physiological regulation of eNOS modulates vascular tone, by releasing NO from the endothelium to counterbalance vasoconstrictor stimuli and accommodate changes in blood flow. Accordingly, eNOS “knockout” mice are routinely hypertensive (22). Multiple vasodilator stimuli (shear stress, vasodilator agonists) stimulate eNOS activity, resulting in the synthesis of NO from L-arginine; NO then diffuses to the adjacent smooth muscle layer and causes vasorelaxation (11,14) (Figure 1). The constitutive, physiologic nature of eNOS activity is most commonly inferred from effects of NOS inhibition in humans or laboratory animals, generally with guanidino-substituted analogues of L-arginine; inhibition leads to both systemic and pulmonary hypertension (23,24). NO, like prostacyclin, appears to be a major endothelium-derived regulator of vascular tone and tissue perfusion.Figure 1: The physiologic and pathophysiologic roles of nitric oxide (NO). Endothelial NO production is stimulated by numerous vasodilator agonists or shear stress. NO produced by constititutive endothelial NO synthase (eNOS) causes relaxation of adjacent vascular smooth muscle and vasodilation. NO also inhibits platelet aggregation and is thought to cause vasorelaxation primarily through guanylyl cyclase stimulation, cyclic guanosine monophosphate (cGMP) production, and decreased myocyte intracellular calcium availability and/or contractile apparatus calcium sensitivity (i.e., myosin light chain (MLC) phosphorylation). NO also causes vasorelaxation by potassium (K + )-channel activation, membrane hyperpolarization, and decreased intracellular calcium availability. Also depicted is the putative septic pathophysiologic mechanism whereby NO (and its oxidative breakdown product peroxynitrite, ONOO−), derived from inducible NO synthase (iNOS), directly causes vasorelaxation by effects on vascular smooth muscle (independent of the endothelium).The Effects of Pathophysiologic NO Production on Vascular Contractility The hemodynamic profile of septic (or “distributive”) shock is characterized by hypotension primarily caused by decreased systemic vascular resistance, accompanied (after volume resuscitation) by a relatively elevated cardiac output (in contrast to hypovolemic or cardiogenic shock) (1,6,11,14). Impaired venous return, which mandates aggressive initial volume resuscitation to unmask a hyperdynamic shock profile, is caused by a combination of increased capacitance (septic venodilation) (25,26) and extravascular volume loss as a result of increased capillary permeability (a manifestation of septic endothelial dysfunction) (27–29). Septic endothelial injury and microvascular leakage is ameliorated by PAF antagonism, inhibition of leukotriene synthesis (27,28), and prevention of iNOS induction or action (27,29), suggesting a multifactorial pathogenesis. Several candidate mediators may contribute to septic vascular contractile dysfunction; one of these is NO, produced primarily by iNOS. Sepsis-induced myocardial dysfunction also contributes to septic hypotension and shock (30–32). The circulating septic “myocardial depressant factor” has yet to be identified; various inflammatory cytokines, iNOS-derived NO, and other mediators, such as PAF, may contribute to myocardial dysfunction (30–32). Several lines of evidence suggest that sepsis-induced NO plays a pivotal role in the pathogenesis of septic vascular hypocontractility and shock. iNOS is expressed in tissues of the immune system (leukocytes, macrophages), vasculature (myocytes, endothelium), kidney (mesangium, tubules), and other sites (pancreas, liver, enterocytes, airway, pneumocytes) after exposure to inflammatory stimuli such as endotoxemia or bacteremia (11,14,20,21,33). Plasma and urinary concentrations of NO breakdown products (primarily nitrite and nitrate) are elevated in septic animals and patients (6,11,33,34). Inhibitors of NOS or of guanylyl cyclase (e.g., methylene blue) augment vasopressor-responsiveness and limit hypotension in septic animals and in a small series of human subjects (6–11); in contrast, supplemental L-arginine exacerbates septic hypotension (34). Diminished norepinephrine contractions in isolated omental vessels from septic patients are restored by Nω-nitro-L-arginine-methyl ester (L-NAME) or methylene blue (35). Accordingly, pathologic NO synthesis is absent and hypotension is less severe after experimental sepsis in iNOS knockout mice (36). Such data, and analogy with exogenous nitrovasodilators, suggest that NO impairs vascular contraction, primarily by activating guanylyl cyclase and augmenting cyclic guanosine monophosphate (cGMP) levels (though other data suggest a role of potassium channel activation), leading to vascular hyperpolarization (37). It is then assumed that cGMP, acting through a specific protein kinase (or NO, acting in a cGMP-independent manner), impairs vascular contraction by decreasing myocyte free intracellular calcium concentration and/or by diminishing the calcium sensitivity of the contractile apparatus (12). Work from our laboratory (38,39) and by others (40) suggests that the latter mechanism is the more important of the two. It should be noted that there are only limited clinical observations supporting the role of iNOS activation leading to septic vasodilation in humans; its role is largely inferred from animal studies. Some evidence suggests a lesser role for NO in humans (33,41). Concerns include evidence of lesser NO production in septic humans compared with animals, the lack of a direct demonstration of iNOS induction in hypocontractile septic human vessels, a number of reports in which NOS inhibition raised BP without altering plasma nitrite-nitrate concentrations, and the continuing identification of NO-independent pathways contributing to septic vascular hypocontractility (33,41). NO-independent mediators, which have been shown in animals or humans to play a causal role in septic vascular hypocontractility, include activation of vascular potassium (K+) -channels (leading to vascular smooth muscle hyperpolarization and relaxation) (42), vasopressin deficiency (43,44), vascular carbon monoxide production (45), and direct (NO- and carbon monoxide-independent) activation of guanylyl cyclase (46). The source of NO contributing to septic vascular hypocontractility varies according to the duration of experimental sepsis. Endotoxin (lipopolysaccharide [LPS]) administration itself produces the systemic effects typical of the sepsis syndrome, including (as a function of dose) shock and death (6–10,47,48). Early endotoxemia is characterized by hypotension which is responsive to NOS inhibition, at a time (1–2 h) when iNOS is not yet induced (49). This early phase of septic vascular hypocontractility seems to be mediated in part by eNOS, acutely releasing endothelial NO (50). Bradykinin and other kinins are the putative intermediate stimuli for eNOS activation (PAF and endothelin are other potential contributors) (51–53). Kinin-elicited endothelial prostacyclin release may also contribute to early septic vascular hypocontractility (51). Later (after 4–6 h), inhibitors of the NO-cGMP pathway continue to improve septic vascular hypocontractility, independent of endothelium or eNOS, and raise systemic BP (Figure 2) (6–11,54–56). Early coadministration (with LPS) of dexamethasone, to prevent iNOS induction, also ameliorates septic vascular hypocontractility and systemic hypotension during this later phase of sepsis (57). Thus it appears that initial septic hypotension is mediated primarily by eNOS-derived NO, with nonendothelium-derived NO assuming a greater role in mediating septic vascular hypocontractility after iNOS induction in the vascular wall and elsewhere.Figure 2: Vasoconstrictor concentration-response curves for phenylephrine (PE) in control (triangles) and lipopolysaccharide (LPS)-treated (circles) rat aortic rings with endothelium in the and of for are a that between and + + a that between + and control + to in from compared with vessels from not the contractile the to the in from both and from Effects of in on endothelial and function in and rat of of the and the mediators of septic vascular The for experimental sepsis may the response to therapy. is less sepsis caused by and by endotoxemia the response to sepsis and its and that studies in are more to human sepsis the less of septic The of may also the pathogenesis of septic vascular dysfunction; for vasorelaxation more on a vasodilator on NO in vessels compared with The isoform of the NOS inhibitor and the of the (see may in the response to The Effects of Sepsis on Physiologic NO Impaired and Organ in Sepsis to sepsis vasorelaxation (Figure This of sepsis through of and endothelial calcium and of NO release systemic vascular resistance, in septic most in the pulmonary and renal circulations of these vascular NO appears to play an important role in tissue perfusion during primarily by sepsis-induced vasoconstrictor detailed of the of septic vascular dysfunction in the and renal circulations the in the use of inhibitors of the NO-cGMP pathway for of septic shock. effects on the and other will not be discussed in this may also be Finally, the role of NO in the pathogenesis of impaired tissue oxygen in sepsis is also from the effects of septic cardiovascular dysfunction on oxygen delivery and there is experimental evidence to suggest that NO improve tissue by oxygen release from in the also directly oxygen at the tissue We and others that NOS inhibition to splanchnic or global oxygen in the use of NOS inhibitors may have blood flow potential relaxation to after of an concentration of (i.e., is as a function of curves of renal and from to the compared with from are for that from control contrast, relaxation to directly on vascular smooth independent of is in renal and from compared with from from Endotoxin in impairs relaxation of in of of the The is to blood flow in to of oxygen when delivery is We and others have shown that capillary in is decreased during endotoxemia and that NOS inhibition splanchnic perfusion in this resulting in and/or injury with either L-arginine or exogenous NO septic splanchnic hypoperfusion other reports suggest that prevention of NO either by with NOS ameliorated injury and/or in and in may be for of the in this for early NOS inhibition microvascular leakage in later The of these data for use of NOS inhibitors in septic shock are and injury may be the role of the and the system in sepsis and multiple organ system failure caused by of and to the systemic (2). contrast to the systemic tone systemic pressure and perfusion, the pulmonary is a relatively pressure and NO a vasodilator in the pulmonary as it this in mediated through release contrast with effects in the systemic endotoxemia to in the pulmonary it vasodilator pulmonary leading to pulmonary hypertension in contrast to systemic hypotension hypertension iNOS induction in pulmonary and appears as a result of vasoconstrictor including of both and endothelin NO inhibition septic pulmonary hypertension diminishing cardiac output iNOS inhibition (with in only increased pulmonary increased pulmonary vascular sensitivity to either or pulmonary that NOS inhibition likewise and decreased in septic it appears that iNOS induction septic pulmonary at the of impaired and resulting in increased in the systemic also impairs pulmonary vascular in a NO-independent that selective iNOS inhibition is to and in sepsis. It is role impaired relaxation plays in the pathogenesis of septic pulmonary hypertension or the of eNOS to the adverse effects of NOS inhibition in this it is important to that selective iNOS inhibition may augment sepsis-induced pulmonary hypertension a vasodilator leading to myocardial dysfunction and systemic administration of NO ameliorated pulmonary hypertension caused by NOS inhibition in animal sepsis models NO likewise pulmonary and dysfunction as result of sepsis in animals in one the of patients with septic shock and NOS to to NO limited suggest other combination to septic pulmonary such as endothelin or antagonism, also be as with NOS NOS isoforms are expressed constitutively in the kidney The macula densa (a to the of the of is the of renal nNOS other include renal nerves and a of vascular and delivery the of in caused by a which serves to volume by when is This is by NO, locally synthesized in the macula densa such that NOS inhibition to impaired and renal failure in eNOS is expressed in and and the it modulates vasoconstrictor both in the and the renal NO is synthesized in the not during The resulting selective caused by to capillary and in of renal perfusion, such as volume and shock. The kidney is one of the sites iNOS is constitutively absent inflammatory or immune sites include of of and a vascular The physiologic role of renal iNOS of to balance and volume endotoxemia or renal iNOS is predominantly in the and renal the of this is as will be discussed NOS inhibition causes hypertension in animals (and human with (in increases in and tone, decreased plasma and relatively of increased BP and increased NOS inhibition, which the in caused by hypertension during systemic NOS inhibition, a lesser in is and (in not tone The vasoconstrictor for the in tone during systemic not NOS inhibition is not there is evidence of roles of endothelin and in these endothelin tone during systemic NOS inhibition in then the combination of endothelin and NOS inhibition to septic shock with pulmonary hypertension (see be by decreased and The of septic endothelial dysfunction to impaired organ perfusion and the potential for NOS inhibition to organ systemic hemodynamic is most in the renal has been interest in the of and sepsis-induced renal vascular resistance, renal blood flow and and the development of are of septic most in the rat endotoxemia is when NOS inhibition administration of NOS or exogenous to the combination of endotoxemia and the NOS inhibitor (Figure coadministration of such as or vascular and relaxing to prevent in This the of sepsis-induced endothelial dysfunction in the of for septic of lipopolysaccharide to in ester a NOS in of L-arginine decreased the of induced by the combination of and causes to a to The to the in shown in not from control in with decreased the in combination with restored the in combination with and with from blood a and a endothelial in to of hypotension by in the rat renal vascular resistance, which in contrast to systemic vasodilation, renal blood flow during sepsis or endotoxemia Septic renal on by the renal by mediators (e.g., and and by production of and Physiologic NO synthesis in the renal microvascular and endothelium is important in renal vasodilation and perfusion in Such NO (and endothelial vasodilator be more important to the renal which septic shock. sepsis impairs vasodilator NO release by the renal microvascular to observations in the systemic vasculature Inhibition of eNOS by NOS inhibitor when on this of an impaired and and this both the of and the of the inhibitor reports with the in the rat improves both and output in when as a a in the a and the the loss of at after administration which is less likewise not to have adverse renal effects in septic or suggesting that limited eNOS inhibition may be to renal function during NOS inhibition in sepsis. It is also that the of NOS inhibition is through prevention of vascular and tissue by iNOS-derived NO and its (see below) through hemodynamic may to the observations from studies of NOS iNOS inhibition, either or increased and not to in (Figure contrast, and inhibition with raised BP caused as data on which be in to NOS inhibition to septic shock without tissue iNOS inhibition to endothelial vasodilator function by suggesting that iNOS-derived NO (and from the systemic and renal vasculature and may be to endothelium and systemic blood pressure in in the or of ester (L-NAME) inducible NOS of to decreased an by of contrast, or restored in to control of when to blood pressure to levels to The systemic blood pressure to the data in shown in pressure from control in with increased blood pressure control in or restored blood pressure to control in of hypotension to in with with from of NO may also be an important factor in in the of endothelial activation with and that from both and to for from the combination and not NO production by iNOS may have an important role in septic and this iNOS-derived NO may contribute to renal by systemic and cause renal cytotoxicity directly through effects on the renal endothelium and data the in and inhibitors of the system for hemodynamic in septic shock. Effects of Inhibition is Sepsis NOS Inhibition NOS inhibitors primarily guanidino-substituted of which NOS isoforms by at the to inhibition of the pathway have such as guanylyl cyclase inhibition blue) or NO and data 50% are derived from a number of experimental including NOS or for endothelial for in with blood vessels, and animal between the two most NOS inhibitors suggest that is primarily for eNOS, or nNOS It is important to data the of the however, experimental may for is an of less iNOS in activity to the the of the of endothelial activation and dysfunction in the role of eNOS in tissue perfusion in the of vasoconstrictor and the increased of this in septic it is not that the of NOS inhibition in of experimental sepsis have been global and regional (see tissue perfusion have been in and mortality has been increased in not (6–10). the early that restored BP in led to studies in which or in combination with not only to survival in this at the led to increased mortality other studies have that adverse with NOS inhibition are to that small of agents may be beneficial of a more selective may to eNOS It is whether effects or other experimental the beneficial in sepsis of a of which ineffective in endotoxemia The of NOS inhibition may also be an important of its the of to septic vascular hypocontractility may during sepsis. with more severe septic endothelial injury and dysfunction are more to adverse of NOS inhibition the of sepsis. the and of the various NOS the of adverse effects of NOS inhibition in experimental sepsis has led to the development of with the that hemodynamic effects be Inhibitors inhibitors are in development and data suggest potential the L-arginine in the of studies. Some of these NOS inhibitors are on amino other and have been shown to greater isoform nNOS and or iNOS number of NOS inhibitors are also in such as and for the relatively inhibitor and it has beneficial effects in experimental sepsis (see on data from animal inhibitors are to have more hemodynamic effects as in human septic shock. We and others have the of which causes global and regional hemodynamic effects in animal septic venous and both function and tissue Finally, inhibitors have to norepinephrine in animal septic have with function and survival in these data are direct of and selective NOS both and not function and survival in with the renal effects of selective iNOS inhibition these animal studies the that short-term h) use of iNOS inhibitors may an and to in septic and survival have not been in animal studies. Effects of NO in Sepsis it be that inhibitors will favorably global and regional perfusion in human septic it be that and mortality in these subjects will also be by the of other beneficial and effects of iNOS-derived NO and its breakdown products The of NOS inhibitor may be by beneficial effects of NO in to the sepsis with NO has both and effects and may have effects in sepsis NO also inhibits other pathways that are such as the shock response prevention of NO and cytotoxicity and tissue injury with hemodynamic may the loss of the various beneficial effects of NO in sepsis. as pulmonary or renal vascular or the infectious sepsis may also limit the of NOS inhibition with mice are to the hemodynamic and of also impaired and defense NO and a number of its products (e.g., antimicrobial activity, intracellular also more such as and Such data the of iNOS inhibition in sepsis and the potential for NOS inhibition to contribute to adverse early hemodynamic NO-cGMP Inhibition in Sepsis with inhibitors of the NO-cGMP pathway in human septic shock has been to have beneficial hemodynamic without adverse effects to patients with the of hypotension and shock. data in septic shock include a number of reports or small series of patients to standard and and a small studies of patients and/or early a clinical of NOS inhibition with in human septic shock in early 1998 when a in mortality in the compared with only in from an of this in subjects of shock and decreased in subjects with adverse effects on of or other It is to however, that pulmonary hypertension in a of patients from the human data for septic shock the of NOS inhibitors to pulmonary hypertension while systemic pressure of such potential through the use of agents (with or without combination such as NO, or of such as will in studies of NOS inhibition in septic shock. of and clinical of numerous agents in it to be whether use of inhibitors or other agents this pathway will a for of septic shock.