Juan M. Guerrero

A review of the multiple actions of melatonin on the immune system.

This review summarizes the numerous observations published in recent years which have shown that one of the most significant of melatonin’s pleiotropic effects is the regulation of the immune system. The overview summarizes the immune effects of pinealectomy and the association between rhythmic melatonin production and adjustments in the immune system as markers of melatonin’s immunomodulatory actions. The effects of both in vivo and in vitro melatonin administration on non-specific, humoral, and cellular immune responses as well as on cellular proliferation and immune mediator production are presented. One of the main features that distinguishes melatonin from the classical hormones is its synthesis by a number of nonendocrine extrapineal organs, including the immune system. Herein, we summarize the presence of immune system-synthesized melatonin, its direct immunomodulatory effects on cytokine production, and its masking effects on exogenous melatonin action. The mechanisms of action of melatonin in the immune system are also discussed, focusing attention on the presence of membrane and nuclear receptors and the characterization of several physiological roles mediated by some receptor analogs in immune cells. The review focuses on melatonin’s actions in several immune pathologies including infection, inflammation, and autoimmunity together with the relation between melatonin, immunity, and cancer.

Melatonin-Immune System Relationships

In this paper we review the historical milestones that first highlighted the existence of a relationship between melatonin and the immune system and we summarize data from experiments which correlate the rhythmic production of melatonin with the rhythmic activity of the immune system. The effects of pinealectomy and in vivo administration of melatonin on a variety of immune parameters, including specific and non-specific immunity are considered and we also present contradictory data concerning the effect of melatonin in cultured immunocompetent cells and a possible scheme of how melatonin regulates the production of a number of cytokines. Finally, the mechanism of action of melatonin in the immune system is discussed. Many data suggest the existence of both nuclear and membrane receptors for melatonin in the immune system. Both of these appear to be clearly identified but their specific physiological role is still under discussion. In summary, although there is overwhelming information demonstrating the immunoenhancing properties of melatonin, many questions related to the cytokines involved and the mechanisms of action of the indoleamine require answers.

Evidence of melatonin synthesis by human lymphocytes and its physiological significance: possible role as intracrine, autocrine, and/or paracrine substance

It has been historically assumed that the pineal gland is the major source of melatonin (N‐acetyl‐ 5‐methoxytryptamine) in vertebrates. Melatonin plays a central role in fine‐tuning circadian rhythms in vertebrate physiology. In addition, melatonin shows a remarkable functional versatility exhibiting antioxidant, oncostatic, antiaging, and immunomodulatory properties. Melatonin has been identified in a wide range of organisms from bacteria to human beings. Its biosynthesis from tryptophan involves four well‐defined intracellular steps catalyzed by tryptophan hydroxylase, aromatic amino acid decarboxylase, serotonin‐N‐acetyltransferase, and hydroxyndole‐O‐methyltransferase. Here, for the first time, we document that both resting and phytohemagglutinin‐stimulated human lymphocytes synthesize and release large amounts of melatonin, with the melatonin concentration in the medium increasing up to five times the nocturnal physiological levels in human serum. Moreover, we show that the necessary machinery to synthesize melatonin is present in human lymphocytes. Furthermore, melatonin released to the culture medium is synthesized in the cells, because blocking the enzymes required for its biosynthesis or inhibiting protein synthesis in general produced a significant reduction in melatonin release. Moreover, this inhibition caused a decrease in IL‐2 production, which was restored by adding exogenous melatonin. These findings indicate that in addition to pineal gland, human lymphoid cells are an important physiological source of melatonin and that this melatonin could be involved in the regulation of the human immune system, possibly by acting as an intracrine, autocrine, and/or paracrine substance

Melatonin prevents changes in microsomal membrane fluidity during induced lipid peroxidation

We tested the effect of melatonin on membrane fluidity in microsomes of a rat liver model in which lipid peroxidation was induced by the addition of FeCl3, ADP and NADPH. Membrane fluidity was monitored using fluorescence spectroscopy and lipid peroxidation was estimated by quantifying malonaldehyde (MDA)+4‐hydroxyalkenals (4‐HDA) concentrations following the induction of lipid peroxidation with and without pre‐incubation with melatonin (1 μM–3 mM). Membrane rigidity increased during induced lipid peroxidation while melatonin reduced in a concentration‐dependent manner both membrane rigidity and MDA+4‐HDA generation. Melatonins protective effect may relate to its known ability to scavenge free radicals and function as an antioxidant.

Physiological concentrations of melatonin inhibit nitric oxide synthase in rat cerebellum.

In the present paper we show the inhibitory effect of melatonin on rat cerebellar nitric oxide synthase (NOS) activity. NO production was monitored by the stoichiometric conversion of L-arginine to L-citrulline. The inhibitory effect of melatonin was dose-dependent, with an IC50 value of about 0.1 mM. However, a significant inhibition of enzyme activity (> 22%) was observed at 1 nM melatonin which is in the range of the physiological serum concentration of the hormone at night. The inhibitory effect of melatonin was observed exclusively in the presence of Ca++. Results suggest a new and important role of the pineal hormone melatonin on central nervous system processes, i.e., by modulating NO production.

Melatonin inhibits expression of the inducible NO synthase II in liver and lung and prevents endotoxemia in lipopolysaccharide-induced multiple organ dysfunction syndrome in rats

We evaluated the role of melatonin in endotoxemia caused by lipopolysaccharide (LPS) in unanesthetized rats. The expression of inducible isoform of nitric oxide synthase (iNOS) and the increase in the oxidative stress seem to be responsible for the failure of lungs, liver, and kidneys in endotoxemia. Bacterial LPS (10 mg/kg b.w) was i.v. injected 6 h before rats were killed and melatonin (10–60 mg/kg b.w.) was i.p. injected before and/or after LPS. Endotoxemia was associated with a significant rise in the serum levels of aspartate and alanine aminotransferases, γ‐glutamyl‐transferase, alkaline phosphatase, creatinine, urea, and uric acid, and hence liver and renal dysfunction. LPS also increased serum levels of cholesterol and triglycerides and reduced glucose levels. Melatonin administration counteracted these organ and metabolic alterations at doses ranging between 20 and 60 mg/kg b.w. Melatonin significantly decreased lung lipid peroxidation and counteracted the LPS‐induced NO levels in lungs and liver. Our results also show an inhibition of iNOS activity in rat lungs by melatonin in a dose‐dependent manner. Expression of iNOS mRNA in lungs and liver was significantly decreased by melatonin (60 mg/kg b.w., 58–65%). We conclude that melatonin inhibits NO production mainly by inhibition of iNOS expression. The inhibition of NO levels may account for the protection of the indoleamine against LPS‐induced endotoxemia in rats.—Crespo, E., Macías, M., Pozo, D., Escames, G., Martín, M., Vives, F., Guerrero, J. M., Acuña‐Castroviejo, D. Melatonin inhibits expression of the inducible NO synthase II in liver and lung and prevents endotoxemia in lipopolysaccharide‐induced multiple organ dysfunction syndrome in rats. FASEB J. 13, 1537–1546 (1999)

Inhibition of cerebellar nitric oxide synthase and cyclic GMP production by melatonin via complex formation with calmodulin

Constitutive rat cerebellar nitric oxide synthase (NOS) activity is shown to be inhibited by physiological concentrations of the pineal hormone melatonin. The inhibition was dose‐dependent and was coupled to an inhibition of the cyclic GMP production activated by L‐arginine. Results also show that calmodulin appears to be involved in this process because its presence in the incubation medium was able to prevent the effect of melatonin on both NOS activity and cyclic GMP production. Moreover, polyacrylamide gel electrophoresis studies suggest that melatonin can interact with calmodulin modifying the binding of the peptide to the synthetic NOS peptide encompassing the calmodulin‐binding domain of constitutive NOS from rat cerebellum, the natural mechanism by which calmodulin activates cerebellar NOS. J. Cell. Biochem. 65:430–442.

Melatonin: Buffering the Immune System

Melatonin modulates a wide range of physiological functions with pleiotropic effects on the immune system. Despite the large number of reports implicating melatonin as an immunomodulatory compound, it still remains unclear how melatonin regulates immunity. While some authors argue that melatonin is an immunostimulant, many studies have also described anti-inflammatory properties. The data reviewed in this paper support the idea of melatonin as an immune buffer, acting as a stimulant under basal or immunosuppressive conditions or as an anti-inflammatory compound in the presence of exacerbated immune responses, such as acute inflammation. The clinical relevance of the multiple functions of melatonin under different immune conditions, such as infection, autoimmunity, vaccination and immunosenescence, is also reviewed.

Melatonin reduces nitric oxide synthase activity in rat hypothalamus

Abstract: In this report, rat hypothalamic nitric oxide synthase (NOS) activity is shown to be partially inhibited by physiological concentrations of the pineal hormone melatonin. In vitro studies demonstrate that 1 nM melatonin, which approximates the physiological concentration of the hormone at night, significantly inhibited NOS activity. In vivo studies show that administering melatonin or collecting the hypothalamus from animals at night, when endogenous melatonin levels are elevated, results in a significant decrease of NOS activity. Results also show that calmodulin may be involved in this process since its presence in the incubation medium prevents the inhibitory effect of melatonin on NOS activity.

Reactive Oxygen Intermediates, Molecular Damage, and Aging: Relation to Melatonin

ABSTRACT: Melatonin, the chief secretory product of the pineal gland, is a direct free radical scavenger and indirect antioxidant. In terms of its scavenging activity, melatonin has been shown to quench the hydroxyl radical, superoxide anion radical, singlet oxygen, peroxyl radical, and the peroxynitrite anion. Additionally, melatonins antioxidant actions probably derive from its stimulatory effect on superoxide dismutase, glutathione peroxidase, glutathione reductase, and glucose‐6‐phosphate dehydrogenase and its inhibitory action on nitric oxide synthase. Finally, melatonin acts to stabilize cell membranes, thereby making them more resistant to oxidative attack. Melatonin is devoid of prooxidant actions.