Type I interferon and neutrophil transcripts in lupus nephritis renal biopsies: clinical and histopathological associations.

OBJECTIVES: To investigate the expression of type I IFN (IFN-I) and neutrophil transcripts in kidney tissue from patients with different classes of LN and their association with distinct clinical and histopathological features. METHODS: Quantitation of IFN-I, defensin-α3 and formyl peptide receptor-like 1 (FPRL-1) transcripts was performed in kidney biopsy tissue from 24 patients with various classes of LN (6 class III, 14 class IV, 4 class V) and 3 control samples. Patient demographics, glomerular filtration rate (eGFR) and histopathological characteristics, including activity and chronicity indices, were analysed. RESULTS: IFNα2 and IFNß transcripts were overexpressed in renal tissues from patients with proliferative forms of LN (III/IV) compared with patients with membranous nephritis and control kidneys. Patients with LN and impaired renal function, attested by eGFR, displayed higher relative expression of IFNα2 transcripts in renal tissues compared with those with normal renal function (23.0 ± 16.2 vs 12.0 ± 14.8, P = 0.04). Defensin-α3, but not FPRL-1, transcripts were overexpressed in LN tissues, particularly those with segmental necrotizing lesions, and were correlated with higher renal pathological activity indices (r = 0.61, P = 0.02), urinary protein levels (r = 0.44, P = 0.048) and IFNα2 expression (r = 0.50, P = 0.01). CONCLUSION: IFN-I transcripts are expressed locally in kidneys from patients with proliferative LN and are associated with impaired renal function. Elevated defensin-α3 transcripts, a neutrophil product associated with neutrophil extracellular traps, may identify a driver of local IFN-I expression. These findings provide insights into the mechanisms of proliferative LN and may inform therapeutic decisions regarding selection of IFN-I pathway inhibitors.

Defensin-lipid interactions in membrane targeting: mechanisms of action and opportunities for the development of antimicrobial and anticancer therapeutics.

Defensins are a class of host defence peptides (HDPs) that often harbour antimicrobial and anticancer activities, making them attractive candidates as novel therapeutics. In comparison with current antimicrobial and cancer treatments, defensins uniquely target specific membrane lipids via mechanisms distinct from other HDPs. Therefore, defensins could be potentially developed as therapeutics with increased selectivity and reduced susceptibility to the resistance mechanisms of tumour cells and infectious pathogens. In this review, we highlight recent advances in defensin research with a particular focus on membrane lipid-targeting in cancer and infection settings. In doing so, we discuss strategies to harness lipid-binding defensins for anticancer and anti-infective therapies.

In Silico Identification of Potential Insect Peptides against Biofilm-Producing Staphylococcus aureus.

Biofilm-producing Staphylococcus aureus (SA) strains are frequently found in medical environments, from surgical/ wound sites, medical devices. These biofilms reduce the efficacy of applied antibiotics during the treatment of several infections, such as cystic fibrosis, endocarditis, or urinary tract infections. Thus, the development of potential therapeutic agents to destroy the extra protective biofilm layers or to inhibit the biofilm-producing enzymes is urgently needed. Advanced and cost-effective bioinformatics tools are advantageous in locating and speeding up the selection of antibiofilm candidates. Based on the potential drug characteristics, we have selected one-hundred thirty-three antibacterial peptides derived from insects to assess for their antibiofilm potency via molecular docking against five putative biofilm formation and regulated target enzymes: the staphylococcal accessory regulator A or SarA (PDB ID: 2FRH), 4,4'-diapophytoene synthase or CrtM (PDB ID: 2ZCQ), clumping factor A or ClfA (PDB ID: 1N67) and serine-aspartate repeat protein C or SdrC (PDB ID: 6LXH) and sortase A or SrtA (PDB ID: 1T2W) of SA bacterium. In this study, molecular docking was performed using HPEPDOCK and HDOCK servers, and molecular interactions were examined using BIOVIA Discovery Studio Visualizer-2019. The docking score (kcal/mol) range of five promising antibiofilm peptides against five targets was recorded as follows: diptericin A (-215.52 to -303.31), defensin (-201.11 to -301.92), imcroporin (-212.08 to -287.64), mucroporin (-228.72 to -286.76), apidaecin II (-203.90 to -280.20). Among these five, imcroporin and mucroporin were 13 % each, while defensin contained only 1 % of positive net charged residues (Arg+Lys) projected through ProtParam and NetWheels tools. Similarly, imcroporin, mucroporin and apidaecin II were 50 %, while defensin carried 21.05 % of hydrophobic residues predicted by the tool PEPTIDE. 2.0. Most of the peptides exhibited potential characteristics to inhibit S. aureus-biofilm formation via disrupting the cell membrane and cytoplasmic integrity. In summary, the proposed hypothesis can be considered a cost-effective platform for selecting the most promising bioactive drug candidates within a limited timeframe with a greater chance of success in experimental and clinical studies.

Using backbone-cyclized Cys-rich polypeptides as molecular scaffolds to target protein-protein interactions.

The use of disulfide-rich backbone-cyclized polypeptides, as molecular scaffolds to design a new generation of bioimaging tools and drugs that are potent and specific, and thus might have fewer side effects than traditional small-molecule drugs, is gaining increasing interest among the scientific and in the pharmaceutical industries. Highly constrained macrocyclic polypeptides are exceptionally more stable to chemical, thermal and biological degradation and show better biological activity when compared with their linear counterparts. Many of these relatively new scaffolds have been also found to be highly tolerant to sequence variability, aside from the conserved residues forming the disulfide bonds, able to cross cellular membranes and modulate intracellular protein-protein interactions both in vitro and in vivo These properties make them ideal tools for many biotechnological applications. The present study provides an overview of the new developments on the use of several disulfide-rich backbone-cyclized polypeptides, including cyclotides, θ-defensins and sunflower trypsin inhibitor peptides, in the development of novel bioimaging reagents and therapeutic leads.

Plant Defensins from a Structural Perspective.

Plant defensins form a family of proteins with a broad spectrum of protective activities against fungi, bacteria, and insects. Furthermore, some plant defensins have revealed anticancer activity. In general, plant defensins are non-toxic to plant and mammalian cells, and interest in using them for biotechnological and medicinal purposes is growing. Recent studies provided significant insights into the mechanisms of action of plant defensins. In this review, we focus on structural and dynamics aspects and discuss structure-dynamics-function relations of plant defensins.

Rhesus θ-Defensin-1 Attenuates Endotoxin-induced Acute Lung Injury by Inhibiting Proinflammatory Cytokines and Neutrophil Recruitment.

Acute lung injury (ALI) is a clinical syndrome characterized by acute respiratory failure and is associated with substantial morbidity and mortality. Rhesus θ-defensin (RTD)-1 is an antimicrobial peptide with immunomodulatory activity. As airway inflammation and neutrophil recruitment and activation are hallmarks of ALI, we evaluated the therapeutic efficacy of RTD-1 in preclinical models of the disease. We investigated the effect of RTD-1 on neutrophil chemotaxis and macrophage-driven pulmonary inflammation with human peripheral neutrophils and LPS-stimulated murine alveolar macrophage (denoted MH-S) cells. Treatment and prophylactic single escalating doses were administered subcutaneously in a well-established murine model of direct endotoxin-induced ALI. We assessed lung injury by histopathology, pulmonary edema, inflammatory cell recruitment, and inflammatory cytokines/chemokines in the BAL fluid. In vitro studies demonstrated that RTD-1 suppressed CXCL8-induced neutrophil chemotaxis, TNF-mediated neutrophil-endothelial cell adhesion, and proinflammatory cytokine release in activated murine alveolar immortalized macrophages (MH-S) cells. Treatment with RTD-1 significantly inhibited in vivo LPS-induced ALI by reducing pulmonary edema and histopathological changes. Treatment was associated with dose- and time-dependent inhibition of proinflammatory cytokines (TNF, IL-1ß, and IL-6), peroxidase activity, and neutrophil recruitment into the airways. Antiinflammatory effects were demonstrated in animals receiving RTD-1 up to 12 hours after LPS challenge. Notably, subcutaneously administered RTD-1 demonstrates good peptide stability as demonstrated by the long in vivo half-life. Taken together, these studies demonstrate that RTD-1 is efficacious in an experimental model of ALI through inhibition of neutrophil chemotaxis and adhesion, and the attenuation of proinflammatory cytokines and gene expression from alveolar macrophages.

The potential for immunoglobulins and host defense peptides (HDPs) to reduce the use of antibiotics in animal production.

Innate defense mechanisms are aimed at quickly containing and removing infectious microorganisms and involve local stromal and immune cell activation, neutrophil recruitment and activation and the induction of host defense peptides (defensins and cathelicidins), acute phase proteins and complement activation. As an alternative to antibiotics, innate immune mechanisms are highly relevant as they offer rapid general ways to, at least partially, protect against infections and enable the build-up of a sufficient adaptive immune response. This review describes two classes of promising alternatives to antibiotics based on components of the innate host defense. First we describe immunoglobulins applied to mimic the way in which they work in the newborn as locally acting broadly active defense molecules enforcing innate immunity barriers. Secondly, the potential of host defense peptides with different modes of action, used directly, induced in situ or used as vaccine adjuvants is described.

Efficacy of Rhesus Theta-Defensin-1 in Experimental Models of Pseudomonas aeruginosa Lung Infection and Inflammation.

Chronic airway infection and inflammation contribute to the progressive loss of lung function and shortened survival of patients with cystic fibrosis (CF). Rhesus theta defensin-1 (RTD-1) is a macrocyclic host defense peptide with antimicrobial and immunomodulatory activities. Combined with favorable preclinical safety and peptide stability data, RTD-1 warrants investigation to determine its therapeutic potential for treatment of CF lung disease. We sought to evaluate the therapeutic potential of RTD-1 for CF airway infection and inflammation using in vitro, ex vivo, and in vivo models. We evaluated RTD-1's effects on basal and Pseudomonas aeruginosa-induced inflammation in CF sputum leukocytes and CF bronchial epithelial cells. Peptide stability was evaluated by incubation with CF sputum. Airway pharmacokinetics, safety, and tolerance studies were performed in naive mice. Aerosolized RTD-1 treatment effects were assessed by analyzing lung bacterial burdens and airway inflammation using an established model of chronic P. aeruginosa endobronchial infection in CF (ΔF508) mice. RTD-1 directly reduces metalloprotease activity, as well as inflammatory cytokine secretion from CF airway leukocyte and bronchial epithelial cells. Intrapulmonary safety, tolerability, and stability data support the aerosol administration route. RTD-1 reduced the bacterial lung burden, airway neutrophils, and inflammatory cytokines in CF mice with chronic P. aeruginosa lung infection. Collectively, these studies support further development of RTD-1 for treatment of CF airway disease.

Immuno-Stimulatory Peptides as a Potential Adjunct Therapy against Intra-Macrophagic Pathogens.

The treatment of infectious diseases is increasingly prone to failure due to the rapid spread of antibiotic-resistant pathogens. Antimicrobial peptides (AMPs) are natural components of the innate immune system of most living organisms. Their capacity to kill microbes through multiple mechanisms makes the development of bacterial resistance less likely. Additionally, AMPs have important immunomodulatory effects, which critically contribute to their role in host defense. In this paper, we review the most recent evidence for the importance of AMPs in host defense against intracellular pathogens, particularly intra-macrophagic pathogens, such as mycobacteria. Cathelicidins and defensins are reviewed in more detail, due to the abundance of studies on these molecules. The cell-intrinsic as well as the systemic immune-related effects of the different AMPs are discussed. In the face of the strong potential emerging from the reviewed studies, the prospects for future use of AMPs as part of the therapeutic armamentarium against infectious diseases are presented.

[Alternatives for antibiotics - antimicrobial peptides and phages].

The constant increase in the number of bacteria resistant to antibiotics poses a substantial problem for the therapy of infectious diseases of different etiologies. The growing insensitivity of pathogens on the classical methods of treatment is associated mainly with multiple mechanisms of resistance created by bacteria. Furthermore, no proper antibiotic treatment causes the appearance of resistant strains even at the last line drugs. Therefore, there are still being sought alternatives in the treatment of difficult to eradicate pathogens. The antimicrobial peptides including cathelicidins, defensins, lysozyme, lactoferrin, histatins and bacteriocins arouse huge interest as potential therapeutics. They exhibit a broad spectrum of activity against many Gram-positive and Gram-negative bacteria, but also against fungi. Moreover, they are considered much safer than antibiotics, due to the fact that they are present in all eukaryotic organisms, in which they are an essential element of the immune system. In addition, phage therapy is also strongly recommended as alternative antibacterial approach. In this review we highlight the potential uses of antimicrobial peptides and bacteriophages in the treatment of infections of various etiologies.

θ-Defensin RTD-1 improves insulin action and normalizes plasma glucose and FFA levels in diet-induced obese rats.

Inflammation is implicated in metabolic abnormalities in obesity and type 2 diabetes. Because θ-defensins have anti-inflammatory activities, we tested whether RTD-1, a θ-defensin, improves metabolic conditions in diet-induced obesity (DIO). DIO was induced by high-fat feeding in obese-prone CD rats from 4 wk of age. Starting at age 10 wk, the DIO rats were treated with saline or RTD-1 for 4 or 8 wk. DIO rats gained more weight than low-fat-fed controls. RTD-1 treatment did not alter body weight or calorie intake in DIO rats. Plasma glucose, FFA, triglyceride (TG), and insulin levels increased in DIO rats; RTD-1 normalized plasma glucose and FFA levels and showed tendencies to lower plasma insulin and TG levels. Hepatic and skeletal muscle TG contents increased in DIO rats; RTD-1 decreased muscle, but not hepatic, TG content. Insulin sensitivity, estimated using homeostasis model assessment of insulin resistance and the glucose clamp technique, decreased in DIO rats, but this change was markedly reversed by RTD-1. RTD-1 had no significant effects on plasma cytokine/chemokine levels or IL-1ß and TNF-α expression in liver or adipose tissues. RTD-1 treatment decreased hepatic expression of phosphoenolpyruvate carboxykinase and glucose-6-phosphatase, suggesting that the effect of RTD-1 on plasma glucose (or insulin action) might be mediated by its effect to decrease hepatic gluconeogenesis. Thus, RTD-1 ameliorated insulin resistance and normalized plasma glucose and FFA levels in DIO rats, supporting the potential of RTD-1 as a novel therapeutic agent for insulin resistance, metabolic syndrome, or type 2 diabetes.

Diverse functions of defensins and other antimicrobial peptides in periodontal tissues.

Defensins are antimicrobial peptides that exhibit direct microbicidal activity as well as mediator-like functions by, for example, activating immature dendritic cells. This review focuses on defensins and other antimicrobial peptides that are present in periodontal tissues. Their antimicrobial capacity against periodontal microorganisms, their regulation and their expression profiles during periodontal diseases is discussed. As antimicrobial peptides may possess great potential for new diagnostic, preventive and therapeutic strategies, a better understanding of how antimicrobial peptides are regulated as part of the innate host immune response is crucial.

Defensin-based anti-infective strategies.

Cationic and amphiphilic peptides are widely distributed in eukaryotic organisms and constitute a first line of host defense against invading pathogens. Some of these host defense peptides (HDPs) combine specific antibiotic activities with modulation of immune responses. Moreover, they are active against bacteria resistant to conventional antibiotics and show only modest resistance development under in vitro selection pressure. Based on these features, HDPs and particularly defensins are considered a promising source of novel anti-infective agents. This review summarizes the current knowledge about defensins from different kingdoms and discusses their potential for therapeutic application.

Defensins: A novel weapon against Mycobacterium tuberculosis?

Tuberculosis (TB) is a serious airborne communicable disease caused by organisms of the Mycobacterium tuberculosis (Mtb) complex. Although the standard treatment antimicrobials, including isoniazid, rifampicin, pyrazinamide, and ethambutol, have made great progress in the treatment of TB, problems including the rising incidence of multidrug-resistant tuberculosis (MDR-TB) and extensively drug-resistant tuberculosis (XDR-TB), the severe toxicity and side effects of antimicrobials, and the low immunity of TB patients have become the bottlenecks of the current TB treatments. Therefore, both safe and effective new strategies to prevent and treat TB have become a top priority. As a subfamily of cationic antimicrobial peptides, defensins are rich in cysteine and play a vital role in resisting the invasion of microorganisms and regulating the immune response. Inspired by studies on the roles of defensins in host defence, we describe their research history and then review their structural features and antimicrobial mechanisms, specifically for fighting Mtb in detail. Finally, we discuss the clinical relevance, therapeutic potential, and potential challenges of defensins in anti-TB therapy. We further debate the possible solutions of the current application of defensins to provide new insights for eliminating Mtb.

Etiology of viral induced acute liver failure and defensins as potential therapeutic agents in ALF treatment.

Acute liver failure (ALF) is a rare and severe disease, which, despite continuous advances in medicine, is still characterized by high mortality (65-85%). Very often, a liver transplant is the only effective treatment for ALF. Despite the implementation of prophylactic vaccinations in the world, the viral background of ALF is still a problem and leads to many deaths. Depending on the cause of ALF, it is sometimes possible to reverse this condition with appropriate therapies, which is why the search for effective antiviral agents seems to be a very desirable direction of research. Defensins, which are our natural antimicrobial peptides, have a very high potential to be used as therapeutic agents for infectious liver diseases. Previous studies on the expression of human defensins have shown that increased expression of human α and ß-defensins in HCV and HBV infections is associated with a better response to treatment. Unfortunately, conducting clinical trials for ALF is very difficult due to the severity of the disease and the low incidence, therefore animal models are important for the development of new therapeutic strategies. One of the best animal models that has real reference to research on acute liver failure (ALF) is rabbit hemorrhagic disease in rabbits caused by the Lagovirus europaeus virus. So far, there have been no studies on the potential of defensins in rabbits infected with Lagovirus europaeus virus.

Alternative approaches to antifungal therapies.

The expansive use of immunosuppressive medications in fields such as transplantational medicine and oncology, the higher frequency of invasive procedures in an ageing population and the HIV/AIDS pandemic have increased the frequency of systemic fungal infections. At the same time, increased resistance of pathogenic fungi to classical antifungal agents has led to sustained research efforts targeting alternative antifungal strategies. In this review, we focus on two promising approaches: cationic peptides and the targeting of fungal virulence factors. Cationic peptides are small, predominantly positively charged protein fragments that exert direct and indirect antifungal activities, one mechanism of action being the permeabilization of the fungal membrane. They include lysozyme, defensins and cathelicidins as well as novel synthetic peptides. Among fungal virulence factors, the targeting of candidal secreted aspartic proteinases seems to be a particularly promising approach.

Adaptively evolved human oral actinomyces-sourced defensins show therapeutic potential.

The development of eukaryote-derived antimicrobial peptides as systemically administered drugs has proven a challenging task. Here, we report the first human oral actinomyces-sourced defensin-actinomycesin-that shows promise for systemic therapy. Actinomycesin and its homologs are only present in actinobacteria and myxobacteria, and share similarity with a group of ancient invertebrate-type defensins reported in fungi and invertebrates. Signatures of natural selection were detected in defensins from the actinomyces colonized in human oral cavity and ruminant rumen and dental plaque, highlighting their role in adaptation to complex multispecies bacterial communities. Consistently, actinomycesin exhibited potent antibacterial activity against oral bacteria and clinical isolates of Staphylococcus and synergized with two classes of human salivary antibacterial factors. Actinomycesin specifically inhibited bacterial peptidoglycan synthesis and displayed weak immunomodulatory activity and low toxicity on human and mammalian cells and ion channels in the heart and central nervous system. Actinomycesin was highly efficient in mice infected with Streptococcus pneumoniae and mice with MRSA-induced experimental peritoneal infection. This work identifies human oral bacteria as a new source of systemic anti-infective drugs.

Humanized theta-defensins (retrocyclins) enhance macrophage performance and protect mice from experimental anthrax infections.

Retrocyclins are humanized versions of the -defensin peptides expressed by the leukocytes of several nonhuman primates. Previous studies, performed in serum-free media, determined that retrocyclins 1 (RC1) and RC2 could prevent successful germination of Bacillus anthracis spores, kill vegetative B. anthracis cells, and inactivate anthrax lethal factor. We now report that retrocyclins are extensively bound by components of native mouse, human, and fetal calf sera, that heat-inactivated sera show greatly enhanced retrocyclin binding, and that native and (especially) heat-inactivated sera greatly reduce the direct activities of retrocyclins against spores and vegetative cells of B. anthracis. Nevertheless, we also found that retrocyclins protected mice challenged in vivo by subcutaneous, intraperitoneal, or intranasal instillation of B. anthracis spores. Retrocyclin 1 bound extensively to B. anthracis spores and enhanced their phagocytosis and killing by murine RAW264.7 cells. Based on the assumption that spore-bound RC1 enters phagosomes by "piggyback phagocytosis," model calculations showed that the intraphagosomal concentration of RC1 would greatly exceed its extracellular concentration. Murine alveolar macrophages took up fluorescently labeled retrocyclin, suggesting that macrophages may also acquire extracellular RC1 directly. Overall, these data demonstrate that retrocyclins are effective in vivo against experimental murine anthrax infections and suggest that enhanced macrophage function contributes to this property.