Davide Signori, Aurora Magliocca, Kei Hayashida, Jan A. Graw, Rajeev Malhotra, Giacomo Bellani, Lorenzo Berra, Emanuele Rezoagli
6 min
Nitric oxide (NO) transitioned from being viewed merely as an environmental pollutant and toxic gas in the 1980s to being recognized as a fundamental signaling and protective molecule in mammalian biology. Endogenous NO is produced by the enzymatic oxidation of L-arginine to L-citrulline, a reaction catalyzed by three distinct isoforms of nitric oxide synthase (NOS): neuronal NOS (nNOS), inducible NOS (iNOS), and endothelial NOS (eNOS). Under physiological conditions, eNOS is the primary contributor to vascular NO, maintaining vascular tone, inhibiting platelet aggregation, and suppressing smooth muscle proliferation. However, factors such as cofactor depletion can lead to "eNOS uncoupling," shifting the enzyme to produce superoxide instead of NO, which drives endothelial dysfunction and pathological vascular states.
Most cardiovascular effects of NO are mediated through the activation of soluble guanylate cyclase (sGC), which converts guanosine triphosphate into cyclic guanosine monophosphate (cGMP). This second messenger activates protein kinase G, culminating in smooth muscle relaxation and vasodilation. Beyond the sGC pathway, NO exerts biological effects via post-translational modifications such as protein S-nitrosylation. A notable example is S-nitroso-hemoglobin, which preserves NO bioactivity in the circulation and releases NO during microvascular deoxygenation to match local blood flow with tissue oxygen demand. In pathological settings involving inflammation, iNOS is upregulated, allowing immune cells to utilize NO and its reactive derivatives as antimicrobial and immunomodulatory agents.
Because systemic NO donors often cause systemic hypotension due to non-selective vasodilation, inhaled nitric oxide (iNO) provides a targeted approach primarily affecting pulmonary and cardiopulmonary circuits. Preclinical and clinical evaluations indicate that iNO effectively treats persistent pulmonary hypertension of the newborn and improves oxygenation in acute respiratory distress syndrome. Furthermore, iNO has demonstrated protective effects in preclinical models of cardiac arrest, myocardial infarction, stroke, subarachnoid hemorrhage, traumatic brain injury, and organ transplantation by mitigating ischemia-reperfusion injury, dampening inflammation, and preserving microvascular perfusion.
The therapeutic window of iNO is bounded by several potential toxicological effects. Inhaled NO can react with oxygen to form nitrogen dioxide, a potent pulmonary irritant that can impair surfactant function. High doses can also lead to the accumulation of methemoglobin and cause systemic side effects such as prolonged bleeding times due to platelet inhibition. Additionally, excessive or unmitigated NO signaling can contribute to antiproliferative effects on smooth muscle cells and cellular damage via the generation of peroxynitrite, a reactive oxidant species capable of inducing lipid peroxidation and mitochondrial dysfunction.
Nitric oxide (NO) is a key molecule in the biology of human life. NO is involved in the physiology of organ viability and in the pathophysiology of organ dysfunction, respectively. In this narrative review, we aimed at elucidating the mechanisms behind the role of NO in the respiratory and cardio-cerebrovascular systems, in the presence of a healthy or dysfunctional endothelium. NO is a key player in maintaining multiorgan viability with adequate organ blood perfusion. We report on its physiological endogenous production and effects in the circulation and within the lungs, as well as the pathophysiological implication of its disturbances related to NO depletion and excess. The review covers from preclinical information about endogenous NO produced by nitric oxide synthase (NOS) to the potential therapeutic role of exogenous NO (inhaled nitric oxide, iNO). Moreover, the importance of NO in several clinical conditions in critically ill patients such as hypoxemia, pulmonary hypertension, hemolysis, cerebrovascular events and ischemia-reperfusion syndrome is evaluated in preclinical and clinical settings. Accordingly, the mechanism behind the beneficial iNO treatment in hypoxemia and pulmonary hypertension is investigated. Furthermore, investigating the pathophysiology of brain injury, cardiopulmonary bypass, and red blood cell and artificial hemoglobin transfusion provides a focus on the potential role of NO as a protective molecule in multiorgan dysfunction. Finally, the preclinical toxicology of iNO and the antimicrobial role of NO-including its recent investigation on its role against the Sars-CoV2 infection during the COVID-19 pandemic-are described.
Alex: And standard drugs don't help with that?
Sam: They actually make it worse. Most drugs that relax blood vessels do so everywhere at once, which drops blood pressure throughout the whole body—dangerous when a patient is already critically ill. Inhaled nitric oxide solves this more precisely. It only reaches the parts of the lung that are actually being ventilated, so it selectively relaxes vessels there and redirects blood toward the working regions. The moment it enters the bloodstream, red blood cells deactivate it almost instantly, so it never reaches the rest of the body.
Alex: So the lung itself acts as a filter, keeping the therapy local by design.
Sam: Exactly. And there's a related situation where that same principle matters—when red blood cells physically break apart inside the body. That happens in certain diseases, and also when blood is pumped through a bypass machine during heart surgery. The oxygen-carrying protein inside those cells spills into the liquid part of the blood, and that free-floating protein is extremely good at grabbing nitric oxide—essentially mopping it up before it can do its job.
Alex: So the vessels lose their signal to stay relaxed.
Sam: Right. They constrict, blood flow to organs like the kidneys drops, and you see measurable organ stress. The same problem occurs with older stored blood from blood banks—the longer blood sits in storage, the more breakdown occurs, and transfusing it can trigger the same vessel-narrowing effect. Providing supplementary inhaled nitric oxide can outcompete that rogue protein and restore normal vessel tone.
Alex: Are there risks to giving patients extra nitric oxide?
Sam: At high concentrations, the gas can convert normal hemoglobin—the molecule that carries oxygen in red blood cells—into a form that can no longer do that job effectively. There's also a risk of reactive byproducts forming that damage cell membranes. At standard therapeutic doses these risks are manageable, but they require careful monitoring.
Alex: Given all that, why haven't large clinical trials shown consistent survival benefits?
Sam: This is the honest tension the review identifies. Most large trials group together patients who share a diagnosis label but have very different underlying biology. Patients with ARDS, for example, may have arrived there through sepsis, trauma, or pneumonia—each with distinct biological drivers. Sex differences and the timing of treatment add further variability. When you average across all of that, meaningful effects in specific subgroups can disappear in the overall numbers.
Alex: So the therapy might be working well for some patients, but the trial design can't see it.
Sam: That's the concern the review raises. The proposed path forward is identifying biological subgroups before treatment rather than after. The review also points toward real-time biosensors that could adjust the dose dynamically based on each patient's specific vascular state—moving from a broad rescue therapy toward something more individually calibrated.
Alex: It's a good illustration of how understanding a mechanism at the molecular level doesn't automatically translate into a clean clinical answer.
Sam: Precisely. The preclinical science is clear: restoring this signaling gas protects organ function. The clinical challenge is matching the right dose, to the right patient, at the right moment. That work is still ongoing.
Alex: Thanks for walking us through it, and thanks to our listeners for joining us on ResearchPod.