Marine Invertebrate Antimicrobial Peptides and Their Potential as Novel Peptide Antibiotics.

Marine invertebrates constantly interact with a wide range of microorganisms in their aquatic environment and possess an effective defense system that has enabled their existence for millions of years. Their lack of acquired immunity sets marine invertebrates apart from other marine animals. Invertebrates could rely on their innate immunity, providing the first line of defense, survival, and thriving. The innate immune system of marine invertebrates includes various biologically active compounds, and specifically, antimicrobial peptides. Nowadays, there is a revive of interest in these peptides due to the urgent need to discover novel drugs against antibiotic-resistant bacterial strains, a pressing global concern in modern healthcare. Modern technologies offer extensive possibilities for the development of innovative drugs based on these compounds, which can act against bacteria, fungi, protozoa, and viruses. This review focuses on structural peculiarities, biological functions, gene expression, biosynthesis, mechanisms of antimicrobial action, regulatory activities, and prospects for the therapeutic use of antimicrobial peptides derived from marine invertebrates.

Immunomodulation, Bioavailability and Safety of Bacteriocins.

The rise of antibiotic-resistant bacteria and the emergence of new pathogens have created a need for new strategies to fight against infectious diseases. One promising approach is the use of antimicrobial peptides produced by a certain species of bacteria, known as bacteriocins, which are active against other strains of the same or related species. Bacteriocins can help in the treatment and prevention of infectious diseases. Moreover, bacteriocins can be obtained in prokaryotic organisms, and contribute s to their widespread use. While the use of bacteriocins is currently limited to the food industry (for example, nisin is used as a preservative, E234), a large number of studies on their microbicidal properties suggest that their use in medicine may increase in the foreseeable future. However, for the successful use of bacteriocins in medicine, it is necessary to understand their effect on the immune system, especially in cases where immunity is weakened due to infectious processes, oncological, allergic, or autoimmune diseases. Studies on the immuno-modulatory activity of bacteriocins in animal models and human cells have revealed their ability to induce both pro-inflammatory and anti-inflammatory factors involved in the implementation of innate immunity. The influence of bacteriocins on acquired immunity is revealed by an increase in the number of T-lymphocytes with a simultaneous decrease in B-lymphocyte levels, which makes them attractive substances for reducing inflammation. The widespread use of bacteriocins in the food industry, their low toxicity, and their broad and narrow specificity are reasons for researchers to pay attention to their immunomodulatory properties and explore their medical applications. Inflammation regulation by bacteriocins can be used in the treatment of various pathologies. The aim of the review was to analyze scientific publications on the immunomodulatory activity, bioavailability, and safety of bacteriocins in order to use the data obtained to organize preclinical and clinical studies.

Inflammation Regulation by Bacterial Molecular Patterns.

Stimulation of innate immunity by bacterial molecular patterns can induce an enhanced cellular immune response to pathogens that are associated with innate immune memory shaped by epigenetic changes. Immunological memory can be expressed in the acceleration/intensification of inflammation, as well as in the exact opposite-to maintain tolerance and non-response to a repeated stimulus. Tolerance is one of the central concepts of immunity and is ensured by the consistency of all parts of the immune response. The severe consequences of inflammation force researchers to study in detail all stages of the downstream pathways that are activated after exposure to a stimulus, while the formation of non-response to a pro-inflammatory stimulus has not yet received a detailed description. Elucidation of the mechanism of tolerance is an urgent task for the prevention and treatment of inflammatory diseases. The aim of this investigation was to study the dynamic changes in the gene expression of A20 and ATF3, the inflammation suppressors, against the background of the expression of the genes of the innate immunity receptors TLR4 and NOD2 and the pro-inflammatory cytokine TNF-α under the influence of TLR4 and NOD2 agonists, lipopolysaccharide (LPS) and glucosaminylmuramyl dipeptide (GMDP). The mechanism of inflammation regulation by bioregulators of bacterial origin-LPS and GMDP-was evaluated in vitro in human peripheral blood mononuclear cells and in vivo after i.p. administration of LPS and GMDP to mice. Gene expression was assessed by RT-PCR. Innate immune receptors and the pro-inflammatory cytokine TNF-α were found to develop early in response to LPS and GMDP, both in vitro and in vivo. Genes of cytosolic proteins controlling inflammation (A20 and ATF3) were expressed later. Prior exposure of the innate immune system to LPS and muramyl peptides may modulate host defense against acute inflammation. As a result of the study, new data were obtained on dynamic changes in deubiquitinase A20 and the transcription factor ATF3, which are involved in the limitation and suppression of inflammatory reactions caused by fragments of bacterial cell walls-LPS and GMDP. Thus, bioregulators of bacterial origin LPS and GMDP, along with pro-inflammatory factors, activate the expression of genes that suppress inflammation, which should be considered when analyzing data from studies of the pro-inflammatory properties of LPS and GMDP and when developing drugs based on them.

Innate Immunity Mechanisms in Marine Multicellular Organisms.

The innate immune system provides an adequate response to stress factors and pathogens through pattern recognition receptors (PRRs), located on the surface of cell membranes and in the cytoplasm. Generally, the structures of PRRs are formed by several domains that are evolutionarily conserved, with a fairly high degree of homology in representatives of different species. The orthologs of TLRs, NLRs, RLRs and CLRs are widely represented, not only in marine chordates, but also in invertebrates. Study of the interactions of the most ancient marine multicellular organisms with microorganisms gives us an idea of the evolution of molecular mechanisms of protection against pathogens and reveals new functions of already known proteins in ensuring the body's homeostasis. The review discusses innate immunity mechanisms of protection of marine invertebrate organisms against infections, using the examples of ancient multicellular hydroids, tunicates, echinoderms, and marine worms in the context of searching for analogies with vertebrate innate immunity. Due to the fact that mucous membranes first arose in marine invertebrates that have existed for several hundred million years, study of their innate immune system is both of fundamental importance in terms of understanding molecular mechanisms of host defense, and of practical application, including the search of new antimicrobial agents for subsequent use in medicine, veterinary and biotechnology.

Regulation of Immune Homeostasis via Muramyl Peptides-Low Molecular Weight Bioregulators of Bacterial Origin.

Metabolites and fragments of bacterial cells play an important role in the formation of immune homeostasis. Formed in the course of evolution, symbiotic relationships between microorganisms and a macroorganism are manifested, in particular, in the regulation of numerous physiological functions of the human body by the innate immunity receptors. Low molecular weight bioregulators of bacterial origin have recently attracted more and more attention as drugs in the prevention and composition of complex therapy for a wide range of diseases of bacterial and viral etiology. Signaling networks show cascades of causal relationships of deterministic phenomena that support the homeostasis of multicellular organisms at different levels. To create networks, data from numerous biomedical and clinical research databases were used to prepare expert systems for use in pharmacological and biomedical research with an emphasis on muramyl dipeptides. Muramyl peptides are the fragments of the cell wall of Gram-positive and Gram-negative bacteria. Binding of muramyl peptides with intracellular NOD2 receptors is crucial for an immune response on pathogens. Depending on the microenvironment and duration of action, muramyl peptides possess positive or negative regulation of inflammation. Other factors, such as genetic, pollutions, method of application and stress also contribute and should be taken into account. A system biology approach should be used in order to systemize all experimental data for rigorous analysis, with the aim of understanding intrinsic pathways of homeostasis, in order to define precise medicine therapy and drug design.

How Do Pollen Allergens Sensitize?

Plant pollen is one of the main sources of allergens causing allergic diseases such as allergic rhinitis and asthma. Several allergens in plant pollen are panallergens which are also present in other allergen sources. As a result, sensitized individuals may also experience food allergies. The mechanism of sensitization and development of allergic inflammation is a consequence of the interaction of allergens with a large number of molecular factors that often are acting in a complex with other compounds, for example low-molecular-mass ligands, which contribute to the induction a type 2-driven response of immune system. In this review, special attention is paid not only to properties of allergens but also to an important role of their interaction with lipids and other hydrophobic molecules in pollen sensitization. The reactions of epithelial cells lining the nasal and bronchial mucosa and of other immunocompetent cells will also be considered, in particular the mechanisms of the activation of B and T lymphocytes and the formation of allergen-specific antibody responses.

Immunomodulatory and Allergenic Properties of Antimicrobial Peptides.

With the growing problem of the emergence of antibiotic-resistant bacteria, the search for alternative ways to combat bacterial infections is extremely urgent. While analyzing the effect of antimicrobial peptides (AMPs) on immunocompetent cells, their effect on all parts of the immune system, and on humoral and cellular immunity, is revealed. AMPs have direct effects on neutrophils, monocytes, dendritic cells, T-lymphocytes, and mast cells, participating in innate immunity. They act on B-lymphocytes indirectly, enhancing the induction of antigen-specific immunity, which ultimately leads to the activation of adaptive immunity. The adjuvant activity of AMPs in relation to bacterial and viral antigens was the reason for their inclusion in vaccines and made it possible to formulate the concept of a "defensin vaccine" as an innovative basis for constructing vaccines. The immunomodulatory function of AMPs involves their influence on cells in the nearest microenvironment, recruitment and activation of other cells, supporting the response to pathogenic microorganisms and completing the inflammatory process, thus exhibiting a systemic effect. For the successful use of AMPs in medical practice, it is necessary to study their immunomodulatory activity in detail, taking into account their pleiotropy. The degree of maturity of the immune system and microenvironment can contribute to the prevention of complications and increase the effectiveness of therapy, since AMPs can suppress inflammation in some circumstances, but aggravate the response and damage of organism in others. It should also be taken into account that the real functions of one or another AMP depend on the types of total regulatory effects on the target cell, and not only on properties of an individual peptide. A wide spectrum of biological activity, including direct effects on pathogens, inactivation of bacterial toxins and influence on immunocompetent cells, has attracted the attention of researchers, however, the cytostatic activity of AMPs against normal cells, as well as their allergenic properties and low stability to host proteases, are serious limitations for the medical use of AMPs. In this connection, the tasks of searching for compounds that selectively affect the target and development of an appropriate method of application become critically important. The scope of this review is to summarize the current concepts and newest advances in research of the immunomodulatory activity of natural and synthetic AMPs, and to examine the prospects and limitations of their medical use.

Dual Effect of Low-Molecular-Weight Bioregulators of Bacterial Origin in Experimental Model of Asthma.

Asthma is one of the most common noncommunicable diseases, affecting over 200 million people. A large number of drugs control asthma attacks, but there is no effective therapy. Identification of reasons for asthma and preventing this disease is a relevant task. The influence of bacterial components is necessary for the normal development of the immune system and the formation of an adequate immune response to antigens. In the absence of microorganisms or their insufficient exposure, the prerequisites are formed for excessive reactivity to harmless antigens. In the present study, we analyzed cellular and humoral factors in a standard mouse model of OVA-induced asthma modified by 5-fold intraperitoneal injection of bacterial cell wall fragments of glucosaminylmuramyl dipeptide (GMDP) 5 µg/animal or 1 µg lipopolysaccharide (LPS) per animal for 5 days before sensitization by ovalbumin (OVA). Preliminary administration of LPS or GMDP to animals significantly reduced goblet cells as well as the number of neutrophils, lymphocytes, and eosinophils in bronchoalveolar lavage, wherein GMDP corrected neutrophilia to a 2-fold degree, and LPS reduced the severity of eosinophilia by 1.9 times. With OVA administration of GMDP or LPS at the sensitization stage, an increase in the total number of bronchoalveolar lavage cells due to neutrophils, macrophages, lymphocytes, and eosinophils in relation to the group with asthma without GMDP or LPS was observed. The administration of GMDP or LPS to normal mice without asthma for 5 days had no statistically significant effect on the change in the number and population composition of cells in bronchoalveolar lavage in comparison with the control group receiving PBS. As a result of a study in a mouse model of asthma, a dual effect of LPS and GMDP was established: the introduction of LPS or GMDP before sensitization reduces neutrophilia and eosinophilia, while the introduction of LPS or GMDP together with an allergen significantly increases neutrophilia and eosinophilia. The study of the immunoglobulin status shows that in normal-asthma mice, GMDP and LPS slightly increase IgA in bronchoalveolar lavage; at the same time, in the asthma model, injections of GMDP or LPS before sensitization contribute to a significant decrease in IgA (2.6 times and 2.1 times, respectively) in BALF and IgE (2.2 times and 2.0 times, respectively) in blood serum. In an experimental model of asthma, the effect of GMDP and LPS was multidirectional: when they are repeatedly administered before sensitization, the bacterial components significantly reduce the severity of the allergic process, while in the case of a joint injection with an allergen, they increase the influx of macrophages, lymphocytes, and neutrophils into the lungs, which can aggravate the course of pathological process. Thus, the insufficient effect of antigens of a bacterial nature, in particular, with prolonged use of antibiotics can be compensated for by substances based on low-molecular-weight bioregulators of bacterial origin to establish the missing signals for innate immunity receptors, whose constant activation at a certain level is necessary to maintain homeostasis.

Strategies for Using Muramyl Peptides - Modulators of Innate Immunity of Bacterial Origin - in Medicine.

The spread of infectious diseases is rampant. The emergence of new infections, the irrational use of antibiotics in medicine and their widespread use in agriculture contribute to the emergence of microorganisms that are resistant to antimicrobial drugs. By 2050, mortality from antibiotic-resistant strains of bacteria is projected to increase up to 10 million people per year, which will exceed mortality from cancer. Mutations in bacteria and viruses are occurring faster than new drugs and vaccines are being introduced to the market. In search of effective protection against infections, new strategies and approaches are being developed, one of which is the use of innate immunity activators in combination with etiotropic chemotherapy drugs. Muramyl peptides, which are part of peptidoglycan of cell walls of all known bacteria, regularly formed in the body during the breakdown of microflora and considered to be natural regulators of immunity. Their interaction with intracellular receptors launches a sequence of processes that ultimately leads to the increased expression of genes of MHC molecules, pro-inflammatory mediators, cytokines and their soluble and membrane-associated receptors. As a result, all subpopulations of immunocompetent cells are activated: macrophages and dendritic cells, neutrophils, T-, B- lymphocytes and natural killer cells for an adequate response to foreign or transformed antigens, manifested both in the regulation of the inflammatory response and in providing immunological tolerance. Muramyl peptides take part in the process of hematopoiesis, stimulating production of colony-stimulating factors, which is the basis for their use in the treatment of oncological diseases. In this review we highlight clinical trials of drugs based on muramyl peptides, as well as clinical efficacy of drugs mifamurtide, lycopid, liasten and polimuramil. Such a multifactorial effect of muramyl peptides and a well-known mechanism of activity make them promising drugs in the treatment and preventing of infectious, allergic and oncological diseases, and in the composition of vaccines.

Pathogenetic Therapy of Psoriasis by Muramyl Peptide.

Psoriasis is a multifactorial disease with a dysregulation in immune system. The aim of this study was to survey the clinical efficacy and safety of muramyl peptide-the ligand of the receptors of innate immunity (drug Licopid, AO Peptek, Moscow, Russia) in patients with psoriasis. The effect of muramyl peptide on 86 patients with different severity of plaque psoriasis was tested. The Psoriasis Area and Severity Index (PASI), cytokine status and production of nitric oxide in blood serum, and the subsequent course of psoriasis have been evaluated. Evaluation of significance of observed differences was presented by the Student's t-test. As a result of the treatment, clinical cure or improvement was detected in 98.2% of patients (p < 0.05), while 24.4% had a complete cure. Subsequent observations during 4 years showed that patients who received muramyl peptide statistically significantly increased relapse-free period. Moreover, subsequent relapses of the disease after treatment with muramyl peptide were in much more milder form in the cases of mild psoriasis. The conducted studies showed that monotherapy with muramyl peptide stopped the clinical manifestations of psoriasis, normalized the processes of cytokine-dependent [interleukin (IL)-4, IL-10, IL-12, tumor necrosis factor alpha (TNF-α)] regulation of the immune response and nonspecific resistance, expressed in a decreasing amount of serum antigens sCD54 [soluble intercellular adhesion molecule-1 (sICAM-1)] to reference values (p ≤ 0.01). Taken together, our research demonstrated the effectiveness of therapy with muramyl peptide and moreover, that elevated levels of sCD54 and MIF (p ≤ 0.01) in the serum of patients with psoriasis considered as potential biomarkers of the severityof psoriasis and control over their dynamics have prognostic significance in determining the effectiveness of the therapy.

sbv IMPROVER: Modern Approach to Systems Biology.

The increasing amount and variety of data in biosciences call for innovative methods of visualization, scientific verification, and pathway analysis. Novel approaches to biological networks and research quality control are important because of their role in development of new products, improvement, and acceleration of existing health policies and research for novel ways of solving scientific challenges. One such approach is sbv IMPROVER. It is a platform that uses crowdsourcing and verification to create biological networks with easy public access. It contains 120 networks built in Biological Expression Language (BEL) to interpret data from PubMed articles with high-quality verification available for free on the CBN database. Computable, human-readable biological networks with a structured syntax are a powerful way of representing biological information generated from high-density data. This article presents sbv IMPROVER, a crowd-verification approach for the visualization and expansion of biological networks.

Muramyl peptides activate innate immunity conjointly via YB1 and NOD2.

Bacterial cell wall muramyl dipeptide (MDP) and glucosaminyl-MDP (GMDP) are potent activators of innate immunity. Two receptor targets, NOD2 and YB1, have been reported; we investigated potential overlap of NOD2 and YB1 pathways. Separate knockdown of NOD2 and YB1 demonstrates that both contribute to GMDP induction of NF-κB expression, a marker of innate immunity, although excess YB1 led to induction in the absence of NOD2. YB1 and NOD2 co-migrated on sucrose gradient centrifugation, and GMDP addition led to the formation of higher molecular mass complexes containing both YB1 and NOD2. Co-immunoprecipitation demonstrated a direct interaction between YB1 and NOD2, a major recombinant fragment of NOD2 (NACHT-LRR) bound to YB1, and complex formation was stimulated by GMDP. We also report subcellular colocalization of NOD2 and YB1. Although YB1 may have other binding partners in addition to NOD2, maximal innate immunity activation by muramyl peptides is mediated via an interaction between YB1 and NOD2.

Community-Reviewed Biological Network Models for Toxicology and Drug Discovery Applications.

Biological network models offer a framework for understanding disease by describing the relationships between the mechanisms involved in the regulation of biological processes. Crowdsourcing can efficiently gather feedback from a wide audience with varying expertise. In the Network Verification Challenge, scientists verified and enhanced a set of 46 biological networks relevant to lung and chronic obstructive pulmonary disease. The networks were built using Biological Expression Language and contain detailed information for each node and edge, including supporting evidence from the literature. Network scoring of public transcriptomics data inferred perturbation of a subset of mechanisms and networks that matched the measured outcomes. These results, based on a computable network approach, can be used to identify novel mechanisms activated in disease, quantitatively compare different treatments and time points, and allow for assessment of data with low signal. These networks are periodically verified by the crowd to maintain an up-to-date suite of networks for toxicology and drug discovery applications.

A product of RpfB and RipA joint enzymatic action promotes the resuscitation of dormant mycobacteria.

Resuscitation-promoting factor proteins (Rpfs) are known to participate in reactivating the dormant forms of actinobacteria. Structural analysis of the Rpf catalytic domain demonstrates its similarity to lysozyme and to lytic transglycosylases - the groups of enzymes that cleave the ß-1,4-glycosidic bond between N-acetylmuramic acid (MurNAc) and GlcNAc, and concomitantly form a 1,6-anhydro ring at the MurNAc residue. Analysis of the products formed from mycobacterial peptidoglycan hydrolysis reactions containing a mixture of RpfB and resuscitation-promoting factor interacting protein (RipA) allowed us to identify the suggested product of their action - N-acetylglucosaminyl-ß(1 → 4)-N-glycolyl-1,6-anhydromuramyl-L-alanyl-D-isoglutamate. To identify the role of this resulting product in resuscitation, we used a synthetic 1,6-anhydrodisaccharide-dipeptide, and tested its ability to stimulate resuscitation by using the dormant Mycobacterium smegmatis model. It was found that the disaccharide-dipeptide was the minimal structure capable of resuscitating the dormant mycobacterial cells over the concentration range of 9-100 ng · mL(-1). The current study therefore provides the first insights into the molecular mechanism of resuscitation from dormancy involving a product of RpfB/RipA-mediated peptidoglycan cleavage.

Enhancement of COPD biological networks using a web-based collaboration interface.

The construction and application of biological network models is an approach that offers a holistic way to understand biological processes involved in disease. Chronic obstructive pulmonary disease (COPD) is a progressive inflammatory disease of the airways for which therapeutic options currently are limited after diagnosis, even in its earliest stage. COPD network models are important tools to better understand the biological components and processes underlying initial disease development. With the increasing amounts of literature that are now available, crowdsourcing approaches offer new forms of collaboration for researchers to review biological findings, which can be applied to the construction and verification of complex biological networks. We report the construction of 50 biological network models relevant to lung biology and early COPD using an integrative systems biology and collaborative crowd-verification approach. By combining traditional literature curation with a data-driven approach that predicts molecular activities from transcriptomics data, we constructed an initial COPD network model set based on a previously published non-diseased lung-relevant model set. The crowd was given the opportunity to enhance and refine the networks on a website ( https://bionet.sbvimprover.com/) and to add mechanistic detail, as well as critically review existing evidence and evidence added by other users, so as to enhance the accuracy of the biological representation of the processes captured in the networks. Finally, scientists and experts in the field discussed and refined the networks during an in-person jamboree meeting. Here, we describe examples of the changes made to three of these networks: Neutrophil Signaling, Macrophage Signaling, and Th1-Th2 Signaling. We describe an innovative approach to biological network construction that combines literature and data mining and a crowdsourcing approach to generate a comprehensive set of COPD-relevant models that can be used to help understand the mechanisms related to lung pathobiology. Registered users of the website can freely browse and download the networks.