Muramyl peptide enhancement of slow-wave sleep

Sleep-promoting Factor S, derived from brain or urine, has been characterized as a muramyl peptide. In rabbits, one pmol of Factor S induced excess slow-wave sleep (SWS) for several hours following cerebral intraventricular infusion. Factor S also induced increases in EEG slow-wave (1/2-4 Hz) amplitudes similar to those observed during the deep sleep that follows sleep deprivation. Sleep following Factor S treatment remained normal in that it was episodic and waking behaviours such as eating, drinking and grooming persisted. The structure of Factor S is similar to those of naturally occurring monomers of bacterial cell wall peptidoglycans. Other muramyl peptides such as muramyl dipeptide (MDP) were also found to be somnogenic. Structure-activity studies of various MDP derivatives indicated that somnogenic activity is dependent on precise structural requirements. Muramyl peptides also are immunomodulators and pyrogens. These activities could be separated from somnogenic activity by using various MDP derivatives or by pharmacologic intervention. Muramyl peptides may induce their somnogenic effects through the release of the cytokine interleukin-1; interleukin-1 also has the capacity to induce excess SWS. Although muramyl peptides have many biological activities in mammals, the role(s) they may play in mammalian physiology remains unknown. It is possible that mammalian tissue may store and/or modify muramyl peptides in conjunction with their use as modulators of sleep, temperature and immune responses.     

New developments in the field of synthetic muramyl peptides, especially as adjuvants for synthetic vaccines

MDP (N-acetylmuramyl-L-alanyl-D-isoglutamine), the first synthetic peptidoglycan derivative capable of replacing whole mycobacteria in Freund's complete adjuvant, is highly active in stimulating antibody production. It also produces delayed hypersensitivity and stimulates non-specific resistance. MDP itself is pyrogenic but murabutide (N-acetyl-muramyl-L-alanyl-D-glutamine-n-butyl ester) is not pyrogenic and is undergoing clinical trials. MDP and its derivatives will prove useful as adjuvants, especially for the "new generation" of synthetic vaccines, which will be discussed in detail. They are also interesting for increasing host resistance against various bacterial and parasitic infections and in experimental tumour immunotherapy. MDP and its derivatives, such as murabutide, will also prove useful in combination with chemotherapy, especially for immunodepressed and elderly patients. Some muramyl peptides are also active in prolonging slow wave sleep in rabbits, cats and monkeys. Muramyl peptides are typical bacterial metabolites, which are apparently essential for establishing a normal immune status and slow wave sleep; they thus represent a new category of vitamins.     

Inhibition of Nod2 signaling and target gene expression by curcumin

Nod2 is an intracellular pattern recognition receptor that detects a conserved moiety of bacterial peptidoglycan and subsequently activates proinflammatory signaling pathways. Mutations in Nod2 have been implicated to be linked to inflammatory granulomatous disorders, such as Crohn's disease and Blau syndrome. Many phytochemicals possess anti-inflammatory properties. However, it is not known whether any of these phytochemicals might modulate Nod2-mediated immune responses and thus might be of therapeutic value for the intervention of these inflammatory diseases. In this report, we demonstrate that curcumin, a polyphenol found in the plant Curcuma longa, and parthenolide, a sesquiterpene lactone, suppress both ligand-induced and lauric acid-induced Nod2 signaling, leading to the suppression of nuclear factor-kappaB activation and target gene interleukin-8 expression. We provide molecular and biochemical evidence that the suppression is mediated through the inhibition of Nod2 oligomerization and subsequent inhibition of downstream signaling. These results demonstrate for the first time that curcumin and parthenolide can directly inhibit Nod2-mediated signaling pathways at the receptor level and suggest that Nod2-mediated inflammatory responses can be modulated by these phytochemicals. It remains to be determined whether these phytochemicals possess protective or therapeutic efficacy against Nod2-mediated inflammatory disorders.     

The protein Nod2: An innate receptor more complex than previously assumed

For almost 10 years, Nod2 has been known as a cytosolic innate receptor able to sense peptidoglycan from Gram-positive and -negative bacteria and to trigger RIP2- and NF-κB-mediated pro-inflammatory and antibacterial response. Mutations in the gene encoding Nod2 in humans have been associated with Crohn's disease (CD). Mechanisms by which Nod2 variants can lead to CD development are still under investigation. The most admitted hypothesis suggests that the impaired function of Nod2 variants in intestinal epithelial and phagocytic cells results in deficiencies in epithelial-barrier function which subsequently lead to increased bacterial invasion and inflammation at intestinal sites. Very recent results have just reinforced this hypothesis by demonstrating that Nod2 wild-type (unlike Nod2 variants) could mediate autophagy, allowing an efficient bacterial clearance and adaptative immune response. Other recent data have attributed new roles to Nod2. Indeed, Nod2 has been shown to activate antiviral innate immune responses involving IRF3-dependent IFN-β production after viral ssRNA recognition through a RIP2-independent mechanism requiring the mitochondrial adaptor protein MAVS. Recently, Nod2 has been also shown to be exquisitely tuned to detect mycobacterial infections and mount a protective immunity against these pathogens.     

Prevention of experimental septic shock by pretreatment of mice with muramyl peptides

Muramyl peptides, immunostimulators with macrophage as a main target cell, are used for protecting mice from LPS-lethality (the experimental model of septic shock). Different protocols of pretreatment mice by muramyl peptides lead to opposite results. LPS and glycopeptides act synergistically in the induction of lethal shock, when mice receive peptides 1 day prior to lethal dose of LPS. However, extension of the period between the peptide and LPS injections to 6 days cancels the effect of synergism. Moreover, a 14-day interval between the same injections leads to protection of 70-90% animals from the toxic effect of LPS. Lipophilic analogs require 10-100 lower concentrations to protect the animals than the parent highly hydrophilic glycopeptides. Production of TNF, IL-1 and phagocytosis by macrophages was studied within the periods corresponding to "synergism" and LPS-resistance. High level of macrophage activity was observed during the "synergism" period. Low TNF production and reduced macrophage phagocyte activity corresponded to LPS-resistant state. These results partly explain the LPS-unresponsiveness in mice after their pretreatment by muramyl peptides.     

Muramyl dipeptide and its derivatives: peptide adjuvant in immunological disorders and cancer therapy

Muramyl dipeptide (MDP) is a synthetic immunoreactive peptide consisting of N-acetyl muramic acid attached to a short amino acid chain of L-Ala-D-isoGln. It was first identified in bacterial cell wall peptidoglycan as an active component in Freund's complete adjuvant. In the cell, MDP is detected by NOD2, a cytoplasmic receptor belonging to the human innate immune system. NOD2 mutations are frequently observed in patients with Crohn's disease, an autoimmune disorder, suggesting the significance of the MDP-NOD2 pathway in activating immunity. For this reason, structural modifications of MDP and its derivatives have been extensively studied in an attempt to increase adjuvant activity and boost the immune response effectively for clinical use in the treatment of cancer and other diseases. This review summarizes the synthetic chemistry of MDP and its derivatives and discusses their pharmacological action and stereoselective synthesis.     

New conjugates of muramyl dipeptide and nor-muramyl dipeptide linked to tuftsin and retro-tuftsin derivatives significantly influence their biological activity

The synthesis and biological activity of new conjugates of muramyl dipeptide (MDP) and nor-muramyl dipeptide (nor-MDP) with tuftsin and retro-tuftsin derivatives containing isopeptide bond between ε-amino group of lysine and carboxyl group of simple amino acids such as Ala, Gly and Val are presented. We presumed, based on the cytokine profile, that the examined conjugates of tuftsin and MDP were capable of activating antibacterial mechanisms by switching on Th1 immune response. The most active were compounds 11, 14 and 19-23.     

Protéines de reconnaissance intra-cellulaire : les voies Nod

Peptidoglycan is an essential cell wall component of bacteria. The host exploits the peptidoglycan particular composition and uniqueness to bacteria for specific bacterial recognition. Peptidoglycan recognition is accomplished by receptors like the peptidoglycan recognition proteins (PGRPs) and the intracellular “nucleotide oligomerization domain” (Nod) proteins. These proteins are required for an adequate immune response to different pathogens. Inversely, pathogens have developed sophisticated strategies to modulate the host response.     

The phosphatidylinositol 3-kinase/Akt pathway negatively regulates Nod2-mediated NF-kappaB pathway

Nucleotide-binding oligomerization domain containing proteins (Nods) are intracellular pattern recognition receptors (PRRs) that recognize conserved moieties of bacterial peptidoglycan and activate downstream signaling pathways, including NF-kappaB pathway. Here, we show that Nod2 agonist muramyldipeptide (MDP) induces Akt phosphorylation in time and dose-dependent manner. The pharmacological inhibitor of phosphatidylinositol 3-kinase (PI3K) (wortmannin) and dominant-negative forms of p85 (the regulatory subunit of PI3K) or Akt enhance, while constitutive active forms of p110 (the catalytic subunit of PI3K) or Akt inhibit, NF-kappaB activation and the target gene interleukin (IL)-8 induced by MDP. In addition, the pharmacological inhibitors of PI3K (wortmannin and LY294002) enhance phosphorylation of NF-kappaB p65 on Ser529 and Ser536 residues, which result in enhanced p65 transactivation activity. Furthermore, we show that the inhibition of PI3K by the pharmacological inhibitors prevent the inactivation of glycogen synthase kinase (GSK)-3beta, suggesting that the negative regulation of PI3K/Akt on MDP-induced NF-kappaB activation is at least in part mediated through inactivation of GSK-3beta. Taken together, our results demonstrate that PI3K/Akt pathway is activated by Nod2 agonist MDP and negatively regulates NF-kappaB pathway downstream of Nod2 activation. Our results suggest that PI3K/Akt pathway may involve in the resolution of inflammatory responses induced by Nod2 activation.     

Peptides of 2-aminopimelic acid: antibacterial agents that inhibit diaminopimelic acid biosynthesis

Succinyl-CoA:tetrahydrodipicolinate-N-succinyltransferase is a key enzyme in the biosynthesis of diaminopimelic acid (DAP), a component of the cell wall peptidoglycan of nearly all bacteria. This enzyme converts the cyclic precursor tetrahydrodipicolinic acid (THDPA) to a succinylated acyclic product. L-2-Aminopimelic acid (L-1), an acyclic analogue of THDPA, was found to be a good substrate for this enzyme and was shown to cause a buildup of THDPA in a cell-free enzyme system but was devoid of antibacterial activity. Incorporation of 1 into a di- or tripeptide yielded derivatives that exhibited antibacterial activity against a range of Gram-negative organisms. Of the five peptide derivatives tested, (L-2-aminopimelyl)-L-alanine (6) was the most potent. These peptides were shown to inhibit DAP production in intact resting cells. High levels (30 mM) of 2-aminopimelic acid were achieved in the cytoplasm of bacteria as a result of efficient uptake of the peptide derivatives through specific peptide transport systems followed, presumably, by cleavage by intracellular peptidases. Finally, the antibacterial activity of these peptides could be reversed by DAP or a DAP-containing peptide. These results demonstrate that the peptides containing L-2-aminopimelic acid exert their antibacterial action by inhibition of diaminopimelic acid biosynthesis.     

Evaluation in rats of the somnogenic,pyrogenic, and central nervous system depressant effects of muramyl dipeptide

Muramyl dipeptide increased sleep during the dark-phase, but not the light-phase of the rats' sleep-awake cycle. This circadian variation may be due to the inability of MDP to increase sleep over the high baseline levels of sleep that occur during the light-phase. However, MDP was pyrogenic during the light-phase, indicating it was pharmacologically, active. In the dark-phase, MDP was not pyrogenic, but when compared to concurrent vehicletreated rats, rats treated with MDP did not demonstrate as great a fall in body temperature. At approximately equisomnogenic doses, MDP produced less potentiation of ethanol-induced loss of righting reflex than triazolam, indicating it produces less non-specific central nervous system depressant effects. These data indicate the possibility of a new generation of hypnotic agents derived from muramyl peptides.     

Host defense cathelicidins in cattle: types,production, bioactive functions and potential therapeutic and diagnostic applications

Cathelicidins are a primitive class of host defense peptides and are known for their broad–spectrum antimicrobial activity against bacteria, fungi, and enveloped viruses. These small, cationic, proteolytically–activated peptides are diverse in structure, encompassing a wide range of activities on host immune and inflammatory cell responses. The dual capacity of cathelicidins to directly control infection and regulate host defenses highlights the potential use of these peptides as alternatives to antibiotics and immunomodulators. Cathelicidins are found in many mammalian species; this review focuses on bovine cathelicidins. Eight naturally and two synthetically occurring bovine cathelicidins are described in detail, with a focus on recent advances in their expression, location and biological roles. This review also presents an overview of the bioactive functions of cathelicidins in bovine mastitis, a disease causing economic losses in cattle dairy production. Comparison of the structural, antimicrobial, cytotoxic and mechanistic properties of bovine cathelicidins advances the knowledge needed for the development of these peptides as potential identifiers of infectious diseases (e.g., bovine mastitis) and as novel therapeutic alternatives to antibiotics.     

The multifaceted activities of mammalian defensins

Defensins are an important family of cationic and cysteine-rich host defense peptides that are widely distributed in plants, fungi, and animals. In mammals, defensins exert potent antimicrobial and immunomodulatory activities linking the innate and adaptive immune responses. These peptides play critical roles in health and disease as defects in their production are associated with abnormal host responses to infection, chronic inflammatory diseases, and cancer. There is much interest in elucidating the structure-function relation and modes of action of the defensins to better understand how these peptides kill microbes and regulate the host immune responses. Such knowledge is expected to help in the design of novel defensin-based therapeutics. This review focuses on the multifaceted antimicrobial and immunomodulatory activities of human and murine defensins.     

Expression of defensins in gingiva and their role in periodontal health and disease

Oral epithelium is a stratified squamous epithelium that functions as the barrier between the outside environment and the host. In the oral cavity, epithelial tissues are constantly exposed to a variety of bacteria, but most individuals maintain healthy homeostasis. Epithelial cells contribute to the innate host response, and antimicrobial peptide expression in all human epithelia, including oral epithelia, is an important part of this epithelial function. These antimicrobial peptides have a broad spectrum of activity against both Gram-negative and Gram-positive bacteria as well as against yeast and viruses. In humans these antimicrobial peptides include defensins and a cathelicidin family member LL-37 in skin and oral mucosa and other epithelia. The human defensins include the alpha-defensins of intestinal and neutrophil origin, and the beta-defensins of skin and oral mucosa and other epithelia. Present studies have identified specific signaling routes that pathogens and commensals take in stimulating these innate immune responses, and this may open the way for development of new therapeutic agents for periodontal diseases.     

Probes for vasopressin receptors Attachment of affinity and fluorescent groups in vasopressin

Fluorescent, photoreactive, and biotinylated analogs of vasopressin have been prepared in which one of these three groups has been attached to a reactive amino group in either position 4 or position 7. Using solid phase methodology, we have synthesized two active parent compounds, [l-desamino,4-lysine,7-hydroxy-prolinelarginine vasopressin and [l-desamino,7-aminoproline]arginine vasopressin, and acylated them to obtain biotinyl, azidobenzoyl, and fluoresceinyl derivatives. We have also prepared analogs in which a “spacer arm” was inserted between lysine in position 4 and the marker group. Some of these derivatives have good antidiuretic activity and could be valuable probes in studying hormone-receptor interaction and in receptor visualization and purification.     

Microbes and their products--physiological effects upon mammalian mucosa

A dynamic array of interactions occurs between pathogens and host mucosal surfaces. The signature molecules unique to microbial pathogens allow the mammalian immune system to recognize them as non-self. This recognition, mediated by the toll-like receptor proteins, results in innate immune responses targeted against the invading organism. Pathogens also elaborate a variety of proteins that actively engage host signaling pathways and subvert them to facilitate their growth and dispersal. These interactions, developed over a long evolutionary period, have been specialized to exquisite detail. These proteins and toxins are either secreted into the medium or directly delivered into host cells by specialized secretion systems. An array of host function alterations is mediated by microbial pathogens including inflammatory responses, secretory responses, alteration of host cytoskeleton, disruption of epithelial tight junctions and apoptosis. The signaling axes involved in these interactions are potential targets for therapeutic strategies against infectious microbes.     

Immunomodulatory properties of novel nucleotide oligomerization domain 2 (nod2) agonistic desmuramyldipeptides

There is a pressing need for the development of novel adjuvants for human use. The minimal bioactive structure of bacterial peptidoglycan (PGN), muramyldipeptide (MDP), and its derivative murabutide (MB) have long been known for their adjuvant activities. For this reason, a series of novel desmuramyldipeptides have been designed and synthesized as part of our search for therapeutically useful MDP analogues. Since nucleotide oligomerization domain 2 (Nod2) is a putative receptor for MDP, we used engineered HEK293 cells overexpressing Nod2 to screen and validate our compounds for their Nod2-agonist activity. Their immunomodulatory properties were subsequently assessed in vitro by evaluating their effect on proinflammatory cytokine production of phorbol 12-myristate 13-acetate (PMA)/ionomycin-stimulated human peripheral blood mononuclear cells (PBMCs). Herein, we present novel desmuramyldipeptides, the most active of them possessing immunoenhancing properties as a result of their potent Nod2-agonistic effect.     

Aerosol delivery of muramyl dipeptide to rodent lungs

Tuberculosis is the single most serious infectious disease worldwide. The respiratory tract is the primary site of infection by Mycobacterium tuberculosis (MTB). A number of immunogenic components of the cell wall of MTB, if delivered to the lungs as aerosols, can be used to study the local immune response. The site of deposition of these aerosols can be employed to control their residence time in the lungs. Muramyl dipeptide (MDP) aerosols were delivered to alveolar macrophages in the lungs of rodents. Guinea pig macrophages harvested by bronchoalveolar lavage were examined by differential interference contrast microscopy for morphological changes indicative of activation. Bronchoalveolar lavage fluid was analyzed for the presence of alkaline phosphatase, lactate dehydrogenase, N-acetyl-glucosaminidase (NAG), and total protein content. Rat alveolar macrophages were studied for the production of nitric oxide, by induction of nitric oxide synthase. Twenty-four hours following exposure to an aerosol of MDP, alveolar macrophages exhibited morphological characterstics (spreading and pseudopodia), enzyme activity (NAG 50% above control), and production of the reactive intermediate nitric oxide. Rat macrophages subjected to aerosol exposure to MDP when challenged with a second dose of MDP or lipopolysaccharide exhibited a linear dose response as measured by nitric oxide production. These studies indicate that the topical delivery of an MTB bacterial cell wall component. muramyl dipeptide, results in activation of alveolar macrophages. This approach may be useful in elucidating elements of the immune response to MTB.