https://staff.blog.ui.ac.id/hsuryadi/files/2012/02/ROS_RNS.pdf

Free radicals and antioxidants in normal physiological functions and human disease

At high concentrations, free radicals and radical-derived, nonradical reactive species are hazardous for living organisms and damage all major cellular constituents. At moderate concentrations, how...

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Free Radicals in the Physiological Control of Cell Function

The discovery that mammalian cells generate nitric oxide, a gas previously considered to be merely an atmospheric pollutant, is providing important information about many biologic processes. Nitric oxide is synthesized from the amino acid L-arginine by a family of enzymes, the nitric oxide synthases, through a hitherto unrecognized metabolic route -- namely, the L-arginine-nitric oxide pathway1–8. The synthesis of nitric oxide by vascular endothelium is responsible for the vasodilator tone that is essential for the regulation of blood pressure. In the central nervous system nitric oxide is a neurotransmitter that underpins several functions, including the formation of memory. . . .

The L-Arginine-Nitric Oxide Pathway

The metabolic syndrome

The discovery that mammalian cells have the ability to synthesize the free radical nitric oxide (NO) has stimulated an extraordinary impetus for scientific research in all the fields of biology and medicine. Since its early description as an endothelial-derived relaxing factor, NO has emerged as a fundamental signaling device regulating virtually every critical cellular function, as well as a potent mediator of cellular damage in a wide range of conditions. Recent evidence indicates that most of the cytotoxicity attributed to NO is rather due to peroxynitrite, produced from the diffusion-controlled reaction between NO and another free radical, the superoxide anion. Peroxynitrite interacts with lipids, DNA, and proteins via direct oxidative reactions or via indirect, radical-mediated mechanisms. These reactions trigger cellular responses ranging from subtle modulations of cell signaling to overwhelming oxidative injury, committing cells to necrosis or apoptosis. In vivo, peroxynitrite generation represents a crucial pathogenic mechanism in conditions such as stroke, myocardial infarction, chronic heart failure, diabetes, circulatory shock, chronic inflammatory diseases, cancer, and neurodegenerative disorders. Hence, novel pharmacological strategies aimed at removing peroxynitrite might represent powerful therapeutic tools in the future. Evidence supporting these novel roles of NO and peroxynitrite is presented in detail in this review.

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Nitric Oxide and Peroxynitrite in Health and Disease

Nitric oxide (NO) is a unique signaling molecule in the mammalian species. NO is produced by a variety of cell types to elicit distinct physiological actions. In the vascular system, NO is produced by the endothelium, a single layer of cells forming the inner lining of all blood vessels. Endothelium-derived NO has several different functions, one of which is vascular smooth muscle relaxation, resulting in vasodilation and a consequent decrease in blood pressure and increase in local blood flow. In the erectile tissue, NO is released as a neurotransmitter from the nerves innervating the corpus cavernosum during sexual stimulation, and causes profound smooth muscle relaxation and increased blood flow to the erectile tissue. This results in engorgement with blood and consequent penile erection.The uniqueness of NO as a signaling molecule derives, at least in part, by the fact that it is a gaseous molecule in its native state. However, despite being a gas, NO, like oxygen (O2), elicits its pharmacological effects as a solute in aqueous solution. Another unique characteristic of NO is its fleeting action because of its highly unstable chemical nature and reactivity. Unlike many other signaling molecules, NO elicits its wise array of physiological effects by distinct mechanisms. For example, vascular and nonvascular smooth muscle relaxation, and inhibition of platelet function are mediated by intracellular cyclic GMP (cyclic 3’, 5’-guanosine monophosphate). NO elicits many cyclic GMP-independent effects as well. For example, nitric oxide is a reactive free radical that can covalently modify protein function. One good example is protein S-nitrosylation, which can result in both regulatory and aberrant effects. By this and a variety of other mechanisms, NO also reacts with other molecules, such as reactive oxygen species, in invading cells such as bacteria, parasites and viruses to kill them or inhibit their replication or spread.The first pharmacological action of nitric oxide, demonstrated several years before it’s production in mammals was actually discovered, was vascular and nonvascular smooth muscle relaxation. One of many examples of the latter is the smooth muscle enveloping the sinusoidal cavities within the corpus cavernosum. Another important example is the airway smooth muscle in the trachea and bronchioles of the lungs. Indeed, inhalation of NO gas causes bronchodilation and increased delivery of air into the lungs. However, perhaps more significant than the bronchodilator effect of inhaled NO is its vasodilator effect. In fact, advantage was taken of the vasodilator action of NO in the lungs by Warren Zapol, MD, from the Massachusetts General Hospital in Boston, who discovered that inhalation of very small amounts of NO gas by newborn babies with life-threatening, persistent pulmonary hypertension (PPHN) results in a dramatic and permanent reversal of pulmonary vasoconstriction. Inhaled NO (INO) literally turned blue babies into pink babies. Without INO, most babies would have died while others would have required highly invasive procedures (extracorporeal membrane oxygenation; ECMO) to oxygenate their lungs, and may not have survived.Regarding its antiviral action, NO has been shown to increase the survival rate of mammalian cells infected with SARS-CoV (Severe Acute Respiratory Syndrome caused by coronavirus). In an in vitrostudy, NO donors (i.e., S-nitroso-N-acetylpenicillamine) greatly increased the survival rate of SARS-CoV-infected eukaryotic cells, suggesting direct antiviral effects of NO (1). In this study, NO significantly inhibited the replication cycle of SARS CoV in a concentration-dependent manner. NO also inhibited viral protein and RNA synthesis. Furthermore, NO generated by inducible nitric oxide synthase inhibited the SARS CoV replication cycle. The coronavirus responsible for SARS-CoV shares most of the genome of COVID- 19 indicating potential effectiveness of inhaled NO therapy in these patients.In 2004, during the SARS-CoV outbreak in China, the administration of inhaled NO reversed pulmonary hypertension, improved severe hypoxia and shortened the length of ventilatory support as compared to matched control patients with SARS-CoV (2). The mechanism of action was thought to be pulmonary vasodilation and consequent improved oxygenation in the blood of the lungs, thereby killing the virus, which does not do well in a high oxygen environment. In addition, however, I would offer the opinion that the NO also interacts directly with the virus to kill it and/or inhibit its replication, as shown in a prior study (1).Although studies have not yet been reported with COVID-19, NO has been shown to have an antiviral effect on several DNA and RNA virus families (3). The NO-mediated S-nitrosylation of viral molecules might be an intriguing general mechanism for the control of the virus life cycle. In this regard, it is conceivable that NO could nitrosylate cysteine-containing enzymes and proteins, including nucleocapsid proteins and glycoproteins, present in the coronavirus.In view of the knowledge gained by treating SARS-CoV patients with INO, it follows that INO might be effective in patients with the current SARS CoV-2 (COVID-19) infection. Indeed, a clinical trial of inhaled nitric oxide in patients with moderate to severe COVID-19 with pneumonia and under assisted ventilatory support recently received IRB (Institutional Review Board) approval at the Massachusetts General Hospital. Warren Zapol is director of this project. This trial has now been expanded to include at least two additional hospitals in the U.S. In the successful treatment of persistent pulmonary hypertension in newborns, the amount of NO inhaled is generally one ppm (part per million). In the clinical trial using COVID-19 patients, the amount of NO will be approximately 100-fold higher, about 100 ppm. This is a safe dose of INO, which could prove to be effective in killing the virus and allowing recovery of the patient. The effective use of INO would also lessen the need for oxygen, ventilators, and beds in the ICU.One thing I urge everyone to practice during this coronavirus pandemic is to breathe or inhale through your NOSE and exhale through your mouth. Swedish investigators at the Karolinska Institute in Stockholm have shown that the cells and tissues in the nasal sinusoids, but not the mouth, constantly and continuously produce nitric oxide, which is a gas, and can be easily detected in the exhaled breath. The physiological significance of this is that nasally-derived NO, when inhaled through the nose, improves oxygen delivery into the lungs by causing bronchodilation. This physiological action of inhaled NO is well-known by competitive athletes, especially runners. Moreover, when inhaling through the nose, your nasal nitric oxide is inhaled into your lungs where it stands a chance of meeting up with the coronavirus particles and killing them or inhibiting their replication. Inhaling through your mouth will NOT accomplish this. By the same token, exhaling through your nose is highly wasteful in that you would be expelling the NO away from the lungs, where it is needed most.“INHALE THROUGH YOUR NOSE, AND EXHALE THROUGH YOUR MOUTH!”

Inhaled Nitric Oxide and COVID-19

Mechanism of action of nitric oxide: heme-dependent activation of guanylate cyclase represents a unifying signal transduction mechanism.

Contributions to a quest.

Oxygen (O2)-dependent and O2-independent antimicrobial mechanisms are used by alveolar macrophages (AM) to maintain lung sterility, but these mechanisms are underdeveloped in neonatal AM. Nitric oxide (NO(.)), a more recently described antimicrobial and immunomodulating molecule, has not been studied in neonatal AM. Lavaged AM from 3-day-old, 10-day-old, maternal and adult rats were treated with or without lipopolysaccharide (LPS) and/or interferon-γ (IFN-γ) and NO(.) synthase activity was measured as its L-arginine metabolites: NO2(-), NO3(-), and citrulline. Superoxide anion (O2(.-)) production by suspended macrophages, initiated by either opsonized zymosan or phorbol, was used as a marker of O2-dependent antimicrobial activity. Lysozyme content of AM was measured as a component of O2-independent antimicrobial activity. Unstimulated 3-day-old macrophages generated >10-fold more NO2(-) + NO3(-) than did 10-day-old, maternal or adult AM. Twenty hours after LPS and IFN-γ stimulation, 3-day-old AM produced > 2 times more NO2(-) and NO3(-) than did the more mature macrophages. Basal and stimulated O2(.-) release was similar among 3-day-old, 10-day-old and adult AM, while lysozyme concentrations were > 4-fold higher in adult macrophages compared to AM from 3-day-old pups. Rather than having a role in NO(.)-dependent antimicrobial activity, we propose that newborn AM have amplified NO(.) production to modulate their own differentiation and replication after birth. The age-dependent differences in NO(.) synthase expression by AM may lend insight into the regulation of this important enzyme.

Amplified nitric oxide production by pulmonary alveolar macrophages of newborn rats.

The recent report by Fan et al alleged that the ProPerDP method is inadequate for the detection of protein persulfidation. Upon careful evaluation of their work, we conclude that the claim made by Fan et al is not supported by their data, rather founded in methodological shortcomings. It is understood that the ProPerDP method generates a mixture of cysteine-containing and non-cysteine-containing peptides. Instead, Fan et al suggested that the detection of non-cysteine-containing peptides indicates nonspecific alkylation at noncysteine residues. However, if true, then such peptides would not be released by reduction and therefore not appear as products in the reported workflow. Moreover, the authors' biological assessment of ProPerDP using Escherichia coli mutants was based on assumptions that have not been confirmed by other methods. We conclude that Fan et al did not rigorously assess the method and that ProPerDP remains a reliable approach for analyses of protein per/polysulfidation.

https://advances.sciencemag.org/content/advances/7/17/eabe7006.full.pdf

Louis J. Ignarro

Papers

Inhaled Nitric Oxide and COVID-19

Mechanism of action of nitric oxide: heme-dependent activation of guanylate cyclase represents a unifying signal transduction mechanism.

Contributions to a quest.

Amplified nitric oxide production by pulmonary alveolar macrophages of newborn rats.

Comment on "Evidence that the ProPerDP method is inadequate for protein persulfidation detection due to lack of specificity".