Alpha-defensins Research Articles

Increased Lipocalin 2 detected by RNA sequencing regulates apoptosis and ferroptosis in COPD

BackgroundChronic obstructive pulmonary disease (COPD) is a complex respiratory condition influenced by environmental and genetic factors. Using next-generation sequencing, we aimed to identify dysregulated genes and potential therapeutic targets for COPD.MethodsPeripheral blood leukocyte RNA profiles from COPD patients and healthy controls were analyzed using next-generation sequencing. Key genes involved in COPD pathogenesis were identified through protein–protein interaction network analysis. In vitro, bronchial epithelial cells treated with cigarette smoke extract (CSE) were used to study the effects on gene expression, cell viability, apoptosis, and ferroptosis. Additionally, Lipocalin 2 (LCN2) inhibition experiments were conducted to elucidate its role in COPD-related cellular processes.ResultsAnalysis of RNA profiles revealed consistent downregulation of 17 genes and upregulation of 21 genes across all COPD groups. Among these, Cathelicidin Antimicrobial Peptide(CAMP), Defensin Alpha 4(DEFA4), Neutrophil Elastase(ELANE), LCN2 and Lactotransferrin(LTF) were identified as potentially important players in COPD pathogenesis. Particularly, LCN2 exhibited a close association with COPD and was found to be involved in cellular processes. In vitro experiments demonstrated that CSE treatment significantly increased LCN2 expression in bronchial epithelial cells in a concentration-dependent manner. Moreover, CSE-induced apoptosis and ferroptosis were observed, along with alterations in cell viability, Glutathione content, Fe2 + accumulation, ROS: Reactive Oxygen Species and Malondialdehyde levels, Lactate Dehydrogenase(LDH) release and Glutathione Peroxidase 4(GPX4) expression. Inhibition of LCN2 expression partially reversed these effects, indicating the pivotal role of LCN2 in COPD-related cellular processes.ConclusionOur study identified six candidate genes: CAMP, DEFA4, ELANE, LCN2, and LTF were upregulated, HSPA1B was downregulated. Notably, LCN2 emerges as a significant biomarker in COPD pathogenesis, exerting its effects by promoting apoptosis and ferroptosis in bronchial epithelial cells.

Mechanisms of AAV2 neutralization by human alpha-defensins.

AAVs are commonly used as gene therapy vectors due to their broad tropism and lack of disease association; however, host innate immune factors, such as human alpha-defensin antimicrobial peptides, can hinder gene delivery. Although it is becoming increasingly evident that human alpha-defensins can block infection by a wide range of nonenveloped viruses, including AAVs, their mechanism of action remains poorly understood. In this study, we describe for the first time how two types of abundant human alpha-defensins neutralize a specific AAV serotype, AAV2. We found that one defensin prevents AAV2 from binding to cells, the first step in infection, while both defensins block a critical later step in AAV2 entry. Our findings support the emerging idea that defensins use a common strategy to block infection by DNA viruses that replicate in the nucleus. Through understanding how innate immune effectors interact with and impede AAV infection, vectors can be developed to bypass these interventions and allow more efficient gene delivery.

Enhanced Antimicrobial Peptide Response Following Bacillus Calmette-Guerin Vaccination in Elderly Individuals.

Antimicrobial peptides are an important component of host defense against Mycobacterium tuberculosis. However, the ability of BCG to induce AMPs as part of its mechanism of action has not been investigated in detail. We investigated the impact of Bacillus Calmette-Guerin (BCG) vaccination on circulating plasma levels and TB-antigen stimulated plasma levels of AMPs in a healthy elderly population. We assessed the association of AMPs, including Human Beta Defensin 2 (HBD-2), Human Neutrophil Peptide 1-3 (HNP1-3), Granulysin, and Cathelicidin (LL37), in circulating plasma and TB-antigen stimulated plasma (using IGRA supernatants) at baseline (pre-vaccination) and at Month 1 and Month 6 post vaccination. Post BCG vaccination, both circulating plasma levels and TB-antigen stimulated plasma levels of AMPs significantly increased at Month 1 and Month 6 compared to pre-vaccination levels in the elderly population. However, the association of AMP levels with latent TB (LTB) status did not exhibit statistical significance. Our findings indicate that BCG vaccination is linked to heightened circulating levels of AMPs in the elderly population, which are also TB-antigen-specific. This suggests a potential mechanism underlying the immune effects of BCG in enhancing host defense against TB.

Predictive value of baseline alpha defensin level in patients with stable coronary artery disease: A retrospective single center study

BackgroundInflammation plays a central role in atherogenesis. The major neutrophilic peptide alpha-defensin is a promising evolving risk factor for atherosclerosis. The aim of the present study was to examine the role of alpha-defensin in predicting future major adverse cardiovascular events (MACE) occurrence in fully revascularized patients with stable CAD under optimal medical therapy. Methods and resultsWe retrospectively examined the prognostic value of baseline plasma alpha-defensin levels in predicting MACE occurrence in 174 fully revascularized patients for stable CAD between March 2016 and January 2017. Alpha-defensin levels were found 20 % higher among demised patients (10,859 pg/ml, IQR [6,920 to 23,320] vs. 9,020 pg/ml, IQR [5,540 to 16,180] pg/ml, P = 0.15). The absolute increase in mortality risk in patients with alpha-defensin levels greater than the median values was 72.5 % (P = 0.33). Log-rank analysis proved both recurrent PCI for de novo lesions (14.9 % and 2.3 %) and the composite of mortality and recurrent PCI for de novo lesions (27.6 % vs. 9.2 %) were significantly related to alpha-defensin values greater than the median (>9200 pg/ml). ConclusionBaseline plasma alpha-defensin is an independent predictor of mortality and recurrent PCI among patients with stable CAD. Alpha-defensin may evolve as a promising factor in cardiovascular risk assessment beyond traditional risk factors. Targeting alpha-defensin to ameliorate MACE occurrence should be addressed in future studies.

Synovial fluid alpha-defensins in Lyme arthritis-a useful marker.

Lyme arthritis, one of the possible late manifestations of Lyme borreliosis, predominantly affects the supporting joints and in adults most often occurs in the form of monoarthritis of the knee. Early diagnosis is based on clinical findings and serology. PCR detection of Borrelia in synovial fluid has become an integral part of the laboratory testing algorithm. The clinical presentation and inflammatory markers in Lyme arthritis can resemble septic arthritis. Determining the levels of alpha-defensins (human neutrophil peptide (HNP 1-3)) in synovial fluid by liquid chromatography is a highly sensitive method revealing the presence of inflammatory process. Between 2020 and 2022, we examined eleven patients with Lyme arthritis of the knee. We measured levels of HNP 1-3 from synovial fluid by HPLC in patients, and we compared it with the corresponding C-reactive protein (CRP) levels in paired serum samples. In patients diagnosed with Lyme arthritis, HNP 1-3 levels in synovial fluid ranged from 2.5 to 261mg/L, with a median of 46.5mg/L. Average serum CRP was 43mg/L. The results show that elevated HNP 1-3 can be consistent with not only septic arthritis or systemic disease, but also with Lyme arthritis, especially in patients with negative culture and 16S PCR from synovial fluid. Final diagnosis must be verified by examination for anti-Borrelia antibodies from serum and synovial fluid. The aim of this work is to introduce an HPLC method for the determination of alpha-defensins as one of the possible diagnostic markers.

Genetically Engineered Goats as Efficient Mammary Gland Bioreactors for Production of Recombinant Human Neutrophil Peptide 1 Using CRISPR/Cas9.

Mammary gland bioreactors are promising methods for recombinant protein production. Human neutrophil peptide 1 (HNP1) exhibits antibacterial and immune-modulating properties. This study aims to establish a method to generate goats secreting HNP1 using the mammary gland as bioreactors. HNP1 transgenic goats were generated by using CRISPR/Cas9 technology to knock-in (KI) the HNP1 sequence into exon 7 of the goat β-casein (CSN2) gene under the control of the CSN2 promoter. One-cell stage embryos were cytoplasmically injected with a mixture of Cas9 mRNA, sgRNA, and a homologous plasmid including the T2A-HNP1 sequences, followed by transfer to recipient goats. A total of 22 live offspring goats were delivered, and 21 of these goats (95.45%) exhibited targeted edits at the CSN2 locus, and 2 female goats (9.09%) demonstrated successful HNP1 integration. Western blot and ELISA analyses confirmed the presence of HNP1 protein at high levels in the milk of these HNP1-positive goats, with mean concentrations of 22.10 µg/mL and 0.0092 µg/mL during the initial 60 days of lactation. Furthermore, milk from these transgenic goats exhibited notable antibacterial activity against Escherichia coli and Staphylococcus aureus, demonstrating the functionality of the expressed HNP1 protein. In conclusion, we established an efficient method for developing new transgenic goat lines as a mammary gland bioreactor, and the bioactive HNP1 protein secreted by the transgenic goat has the potential to combat microbial resistance.

Antifungal activity of human antimicrobial peptides targeting apoptosis in Candida auris.

Introduction. Innovative antifungal therapies are of crucial importance to combat the potentially life-threatening infections linked to the multidrug-resistant fungal pathogen Candida auris. Induction of regulated cell death, apoptosis, could provide an outline for future therapeutics. Human antimicrobial peptides (AMPs), well-known antifungal compounds, have shown the ability to induce apoptosis in pathogenic fungi.Hypothesis/Gap Statement . Although it is known that AMPs possess antifungal activity against C. auris, their ability to induce apoptosis requires further investigations.Aim. This study evaluated the effects of AMPs on the induction of apoptosis in C. auris.Methods. Human neutrophil peptide-1 (HNP-1), human β-Defensins-3 (hBD-3) and human salivary histatin 5 (His 5) were assessed against two clinical C. auris isolates. Apoptosis hallmarks were examined using FITC-Annexin V/PI double labelling assay and terminal deoxynucleotidyl transferase deoxynucleotidyl transferase nick-end labelling (TUNEL) to detect phosphatidylserine externalization and DNA fragmentation, respectively. Then, several intracellular triggers were studied using JC-10 staining, spectrophotometric assay and 2',7'-dichlorofluorescin diacetate staining to measure the mitochondrial membrane potential, cytochrome-c release and reactive oxygen species (ROS) production, respectively.Results and conclusion. FITC-Annexin V/PI staining and TUNEL analysis revealed that exposure of C. auris cells to HNP-1 and hBD-3 triggered both early and late apoptosis, while His 5 caused significant necrosis. Furthermore, HNP-1 and hBD-3 induced significant mitochondrial membrane depolarization, which resulted in substantial cytochrome c release. In contrast to His 5, which showed minimal mitochondrial depolarization and no cytochrome c release. At last, all peptides significantly increased ROS production, which is related to both types of cell death. Therefore, these peptides represent promising and effective antifungal agents for treating invasive infections caused by multidrug-resistant C. auris.

Characterisation of defensins across the marsupial family tree

Defensins are antimicrobial peptides involved in innate immunity, and gene number differs amongst eutherian mammals. Few studies have investigated defensins in marsupials, despite their potential involvement in immunological protection of altricial young. Here we use recently sequenced marsupial genomes and transcriptomes to annotate defensins in nine species across the marsupial family tree. We characterised 35 alpha and 286 beta defensins; gene number differed between species, although Dasyuromorphs had the largest repertoire. Defensins were encoded in three gene clusters within the genome, syntenic to eutherians, and were expressed in the pouch and mammary gland. Marsupial beta defensins were closely related to eutherians, however marsupial alpha defensins were more divergent. We identified marsupial orthologs of human DEFB3 and 6, and several marsupial-specific beta defensin lineages which may have novel functions. Marsupial predicted mature peptides were highly variable in length and sequence composition. We propose candidate peptides for future testing to elucidate the function of marsupial defensins.

Salivary Levels of Human Neutrophil Peptide (HNP) 1-3 in Patients with Recurrent Aphthous Stomatitis and Behçet’s Disease: A Cross-sectional Study in Iran

Background: Recurrent oral ulcers are the most common complaint of patients with Behçet’s disease (BD) and recurrent aphthous stomatitis (RAS). Enhanced innate immune response and neutrophilic activity might be a possible etiopathogenesis of BD. This study aimed to determine the significance of salivary human neutrophil peptide (HNP 1-3) in BD and RAS patients and detect their correlation with different clinical presentations, disease activity, and characteristics of oral ulcers. Methods: This cross-sectional study included 25 BD patients and 25 RAS patients as well as 25 healthy participants. Five cubic centimeters of unstimulated saliva was collected and levels of HNP 1-3 were measured by ELISA. Other data were obtained through interviews, examinations, and reviews of medical records. Finally, data analysis was performed using SPSS 25.0 software. Results: Salivary HNP 1-3 levels were not significantly different between the study groups (P=0.282). Duration of oral ulcers did not correlate with HNP 1-3 levels in RAS and BD patients (P > 0.05). Also, BD patients with involvements other than oral ulcers were not found to have different levels of HNP 1-3 compared to those who did not manifest these conditions. Conclusion: The validity of HNP 1-3 to be used as a probable biological marker for evaluation, diagnosis, and estimation of disease activity in patients with BD and RAS is still questionable according to our results.

Major endothelial damage markers identified from hemadsorption filters derived from treated patients with septic shock - endoplasmic reticulum stress and bikunin may play a role.

In septic patients the damage of the endothelial barrier is decisive leading to circulatory septic shock with disseminated vascular coagulation, edema and multiorgan failure. Hemadsorption therapy leads to rapid resolution of clinical symptoms. We propose that the isolation of proteins adsorbed to hemadsorption devices contributes to the identification of mediators responsible for endothelial barrier dysfunction. Plasma materials enriched to hemadsorption filters (CytoSorb®) after therapy of patients in septic shock were fractionated and functionally characterized for their effect on cell integrity, viability, proliferation and ROS formation by human endothelial cells. Fractions were further studied for their contents of oxidized nucleic acids as well as peptides and proteins by mass spectrometry. Individual fractions exhibited a strong effect on endothelial cell viability, the endothelial layer morphology, and ROS formation. Fractions with high amounts of DNA and oxidized DNA correlated with ROS formation in the target endothelium. In addition, defined proteins such as defensins (HNP-1), SAA1, CXCL7, and the peptide bikunin were linked to the strongest additive effects in endothelial damage. Our results indicate that hemadsorption is efficient to transiently remove strong endothelial damage mediators from the blood of patients with septic shock, which explains a rapid clinical improvement of inflammation and endothelial function. The current work indicates that a combination of stressors leads to the most detrimental effects. Oxidized ssDNA, likely derived from mitochondria, SAA1, the chemokine CXCL7 and the human neutrophil peptide alpha-defensin 1 (HNP-1) were unique for their significant negative effect on endothelial cell viability. However, the strongest damage effect occurred, when, bikunin - cleaved off from alpha-1-microglobulin was present in high relative amounts (>65%) of protein contents in the most active fraction. Thus, a relevant combination of stressors appears to be removed by hemadsorption therapy which results in fulminant and rapid, though only transient, clinical restitution.

Human Neutrophil Alpha-Defensins Promote NETosis and Liver Injury in Alcohol-Related Liver Cirrhosis: Potential Therapeutic Agents.

Background: Neutrophils are thought to play a pivotal role in the pathogenesis of many inflammatory diseases, such as hepatitis, liver cirrhosis, etc. Activated human neutrophils release human neutrophil peptides (HNP1-3) or alpha-defensins that are antimicrobial peptides in azurophil granules. Furthermore, HNP1-3 build a scaffold of neutrophil extracellular traps (NETs) and promote the process of programmed cell death called NETosis. Our study aimed to investigate the role of alpha-defensins in the pathogenesis of alcohol-related liver cirrhosis (ALC). Methods: The concentrations of alpha-defensins in the plasma of 62 patients with ALC and 24 healthy subjects were measured by ELISA. The patients with ALC were prospectively recruited based on the severity of liver dysfunction according to the Child-Pugh and Model of End-Stage Liver Disease-Natrium (MELD-Na) scores, modified Maddrey's Discriminant Function (mDF), and the presence of ALC complications. Results: The concentrations of alpha-defensins in plasma were significantly higher in the ALC patients than in the controls. The plasma levels of HNP1-3 correlated with the MELD and mDF scores. ALC subgroups with MELD > 20 and mDF > 32 displayed significantly higher HNP1-3 concentrations. The plasma levels of HNP1-3 revealed a good predictive AUC for hepatic encephalopathy and ascites development (0.81 and 0.74, respectively) and for patient survival (0.87) in those over 40 years of age. Conclusion: These findings suggest that alpha-defensins play an important role in the assessment of ALC.

Genes and Pathways Underpinning Klinefelter Syndrome at Bulk and Single-Cell Levels.

Klinefelter syndrome (KS) is the most frequent genetic anomaly in infertile men. Given its unclear mechanism, we aim to investigate critical genes and pathways in the pathogenesis of KS based on three bulk and one single-cell transcriptome data sets from Gene Expression Omnibus. We merged two data sets (GSE42331 and GSE47584) with human KS whole blood samples. When comparing the control and KS samples, five hub genes, including defensin alpha 4 (DEFA4), bactericidal permeability increasing protein (BPI), myeloperoxidase (MPO), intelectin 1 (ITLN1), and Xg Glycoprotein (XG), were identified. Besides, infiltrated degree of certain immune cells such as CD56bright NK cell were positively associated with the expression of ITLN1 and XG. Kyoto Encyclopedia of Genes and Genomes analysis identified upregulated phosphatidylinositol 3-kinase (PI3K)/AKT pathway in KS. Gene set enrichment analysis followed by gene set variation analysis confirmed the upregulation of G2M checkpoint and heme metabolism in KS. Thereafter, the GSE200680 data set was used for external validation of the expression variation of hub genes from healthy to KS testicular samples, and each hub gene yielded excellent discriminatory capability for KS without exception. At the single-cell level, the GSE136353 data set was utilized to evaluate intercellular communication between different cell types in KS patient, and strong correlations were detected between macrophages/ dendritic cells/ NK cells and the other cell types. Collectively, we provided hub genes, pathways, immune cell infiltration degree, and cell-cell communication in KS, warranting novel insights into the pathogenesis of this disease.

HIGH COPY NUMBER POLYMORPHISMS OF ALPHA DEFENSIN 1-3 ARE ASSOCIATED WITH THERAPEUTIC RESPONSIVENESS IN INFLAMMATORY BOWEL DISEASE

Abstract BACKGROUND AND PURPOSE Prednisolone therapy is the first choice for the treatment of acute severe ulcerative colitis (ASUC), however, the refractory case is sometimes observed. Alpha defensin 1-3 (DEFA1A3) stored in neutrophils acts as an effector factor of innate immunity. It has been reported that the copy number variation of the DEFA1A3 gene is associated with the severity of ulcerative colitis (UC). However, the impact of DEFA1A3 in response to treatment have not been fully investigated. Therefore, we aimed to clarify the relationship between the number of DEFA1A3 copies and UC prednisolone treatment response. SUBJECTS AND METHODS Patients with UC who had attended our department and related facilities and had the number of DEFA1A3 copies measured were included between 2020 and 2022. The number of DEFA1A3 copies were measured using droplet digital PCR after DNA extraction from peripheral blood mononuclear cells. Mayo score was used to determine the response to prednisolone and severity of UC. ASUC was defined as presence of more than 6 bloody stools/d along with any one of the following: pulse rate> 90 bpm, temperature > 37.8 °C, hemoglobin < 10.5 g/dL, and/or CRP > 30mg/L. Clinical background and various parameters were extracted from medical record information. RESULTS The 47 ASUC patients were included in the study. The median DEFA1A3 copy number of the patients was 9, the median DEFA1A3 copy number in ASUC was higher than that in healthy subjects. The DEFA1A3 copy number was significantly higher in the non-remission group than in the remission group after treatment with prednisolone.There were no differences in other parameters such as CRP and WBC between the two groups. CONCLUSION DEFA1A3 copy number may be a predictive marker of prednisolone treatment response.

Serum proteomics analysis of drug-naïve patients with generalised anxiety disorder: Tandem mass tags and multiple reaction monitoring

Objectives The prevalence of generalised anxiety disorder (GAD) is high. However, the underlying mechanisms remain elusive. Proteomics techniques can be employed to assess the pathological mechanisms involved in GAD. Methods Twenty-two drug-naive GAD patients were recruited, their serum samples were used for protein quantification and identified using Tandem Mass Tag and Multiple Reaction Monitoring (MRM). Machine learning models were employed to construct predictive models for disease occurrence by using clinical scores and target proteins as input variables. Results A total of 991 proteins were differentially expressed between GAD and healthy participants. Gene Ontology analysis revealed that these proteins were significantly associated with stress response and biological regulation, suggesting a significant implication in anxiety disorders. MRM validation revealed evident disparities in 12 specific proteins. The machine learning model found a set of five proteins accurately predicting the occurrence of the disease at a rate of 87.5%, such as alpha 1B-glycoprotein, complement component 4 A, transferrin, V3-3, and defensin alpha 1. These proteins had a functional association with immune inflammation. Conclusions The development of generalised anxiety disorder might be closely linked to the immune inflammatory stress response.

Characterization of metabolic alterations in the lean metabolically unhealthy alpha defensin transgenic mice

Inflammation is consistently linked to dysmetabolism. In transgenic mice (Def+/+) model the neutrophilic peptide, alpha defensin, proved atherogenic. This phenotype occurred despite favorable cholesterol and glucose levels, and lower body weight and blood pressure. In this study, integration of metabolic&behavioral phenotyping system, endocrine, biochemical and mitochondrial assessment, pathological and immunohistochemical tests, and multiple challenge tests was established to explore the metabolic impact of alpha defensin. Compared to the control group, Def+/+ mice exhibited lower total energy expenditure and carbohydrate utilization, and higher fat oxidation. Their ACTH-cortisol and thyroid profiles were intact. Intriguingly, they had low levels of glucagon, with high ammonia, uric acid, triglyceride, and lactate. Mitochondrial evaluations were normal. Overall, defensin-induced hypoglucagonemia is associated with lipolysis, restricted glucose oxidation, and enhanced wasting. Def+/+ mice may be a useful model for studying the category of lean, apparently metabolically healthy, and atherosclerotic phenotype, with insight into a potential inflammatory-metabolic link.

Antimicrobial Peptide Recognition Motif of the Substrate Binding Protein SapA from Nontypeable Haemophilus influenzae.

Nontypeable Haemophilus influenzae (NTHi) is an opportunistic pathogen associated with respiratory diseases, including otitis media and exacerbations of chronic obstructive pulmonary disease. NTHi exhibits resistance to killing by host antimicrobial peptides (AMPs) mediated by SapA, the substrate binding protein of the sensitivity to antimicrobial peptides (Sap) transporter. However, the specific mechanisms by which SapA selectively binds various AMPs such as defensins and cathelicidin are unknown. In this study, we report mutational analyses of both defensin AMPs and the SapA binding pocket to define the specificity of AMP recognition. Bactericidal assays revealed that NTHi lacking SapA are more susceptible to human beta defensins and LL-37, while remaining highly resistant to a human alpha defensin. In contrast to homologues, our research underscores the distinct specificity of NTHi SapA, which selectively recognizes and binds to peptides containing the charged-hydrophobic motif PKE and RRY. These findings provide valuable insight into the divergence of SapA among bacterial species and NTHi SapA's ability to selectively interact with specific AMPs to mediate resistance.

Acute and chronic rhinosinusitis age characteristics

Children's and adults' rhinosinusitis are two diseases that have both similarities and differences in anatomy, epidemiology, causes, pathogenesis, diagnosis and treatment. At the same rhinosinusitis is one of the most common in otorhinolaryngology's practice, both in children and adults. The of adults paranasal sinuses (PNS) anatomy differs from children's PNS anatomy. Although ostiomeatal complex occlusion is recognized as a major cause of poor ventilation and drainage of the adult paranasal sinuses, it does not have a strong effect on pediatric rhinosinusitis, but adenoids play a key role. Adenoids are bacteria and biofilms reservoirs that cause chronic refractory rhinosinusitis regardless of pharyngeal tonsil size. The prevalence of chronic rhinosinusitis (CRS) is lower in children than in adults. Diagnosis of children's rhinosinusitis is more difficult because nasal cavity endoscopic examination is performed rarely due to the occasional need of general anesthesia during the procedure. Moreover, it's necessary to take into account prevailing etiological role of viruses in ARS at children's age and chronic adenoiditis often accompanies pediatric CRS, which requires attention prescribing medical therapy as the basis of rhinosinusitis treatment. The DysheLORz based on Pelargonium sidoides roots is highly effective and safe for children's and adults ARS and CRS treatment, both as monotherapy and in combination with topical steroids and antibiotics. This herbal medicine immunomodulatory effect is mediated mainly by stimulating the production of TNF-α, IL-1, IL-12 and IFN-γ. It activates macrophages and improves their phagocytic activity. IL-12, together with TNF-α, enhances NK and cytotoxic CD8+ lymphocytes' activity against infected cells. IL-12 effect on Th1 lymphocytes maturation provides a link between innate and adaptive immunity. This is also increasing MCP-1, IP-10 and MIP-1β chemokines synthesis and decreasing MIP-1α, ENA-78, GROα and IL-8 production in PNS and nasal mucosa. This leads to decrease of neutrophils chemotaxis to the inflammation site, and decline of serine proteases concentration (neutrophils main enzymes), that increases mucous membrane epithelial barrier permeability, reducing bacterial infections risk. Additionally, Pelargonium sidoides increases epithelial cells beating cilia frequency and inhibits hemagglutinin and neuraminidase present on influenza virus surface. The drug increases antimicrobial peptides production as defensins, human neutrophil peptides (HNP) and bactericidal permeability-increasing protein (BPI), which is also important for rapid inflammation regression in rhinosinusitis. It causes bacterial adhesion to epithelial cells inhibition, phagocytosis stimulation, nitric oxide (NO) release and oxidative burst. The medicine had a direct effect on Streptococcus pneumoniae, Staphylococcus aureus, Neisseria, Moraxella catarrhalis and Haemophilus influenza. Based on these data, it is possible to explain the high effectiveness and safety of the drugs based on Pelargonium sidoides in ENT organs inflammation treatment, for both adults and children over 1 year old.

Achieving High Accuracy in Predicting the Probability of Periprosthetic Joint Infection From Synovial Fluid in Patients Undergoing Hip or Knee Arthroplasty: The Development and Validation of a Multivariable Machine Learning Algorithm.

Background and objective The current periprosthetic joint infection (PJI) diagnostic guidelines require clinicians to interpret and integrate multiple criteria into a complex scoring system. Also, PJI classifications are often inconclusive, failing to provide a clinical diagnosis. Machine learning (ML) models could be leveraged to reduce reliance on these complex systems and thereby reduce diagnostic uncertainty. This study aimed to develop an ML algorithm using synovial fluid (SF) test results to establish a PJI probability score. Methods We used a large clinical laboratory's dataset of SF samples, aspirated from patients withhip or knee arthroplasty as part of a PJI evaluation. Patient age and SF biomarkers [white blood cell count, neutrophil percentage (%PMN), red blood cell count, absorbance at 280 nm wavelength, C-reactive protein (CRP), alpha-defensin (AD), neutrophil elastase, and microbial antigen (MID) tests] were used for model development. Data preprocessing, principal component analysis, and unsupervised clustering (K-means) revealed four clusters of samples that naturally aggregated based on biomarker results. Analysis of the characteristics of each of these four clusters revealed three clusters (n=13,133) with samples having biomarker results typical of a PJI-negative classification and one cluster (n=4,032) with samples having biomarker results typical of a PJI-positive classification. A decision tree model, trained and tested independently of external diagnostic rules, was then developed to match the classification determined by the unsupervised clustering. The performance of the model was assessed versus a modified 2018International Consensus Meeting (ICM) criteria, in both the test cohort and an independent unlabeled validation set of 5,601 samples. TheSHAP (SHapley Additive exPlanations) method was used to explore feature importance. Results The ML model showed an area under the curveof 0.993, with a sensitivity of 98.8%, specificity of 97.3%, positive predictive value (PPV) of 92.9%, and negative predictive value (NPV) of 99.8%in predicting the modified 2018 ICM diagnosis among test set samples. The model maintained its diagnostic accuracy in the validation cohort, yielding 99.1% sensitivity, 97.1% specificity, 91.9% PPV, and 99.9% NPV. The model's inconclusive rate (diagnostic probability between 20-80%) in the validation cohort was only 1.3%,lower than that observed with the modified 2018 ICM PJI classification (7.4%; p<0.001). The SHAP analysis found that AD was the most important feature in the model, exhibiting dominance among >95% of "infected" and "not infected" diagnoses. Other important features were the sum of the MID test panel, %PMN, and SF-CRP. Conclusions Although defined methods and tools for diagnosis of PJI using multiple biomarker criteria are available, they are not consistently applied or widely implemented. There is a need for algorithmic interpretation of these biomarkers to enable consistent interpretation of the results to drive treatment decisions. The new model, using clinical parameters measured from a patient's SF sample, renders a preoperative probability score for PJI which performs well compared to a modified 2018 ICM definition. Taken together with other clinical signs, this model has the potential to increase the accuracy of clinical evaluations and reduce the rate of inconclusive classification, thereby enabling more appropriate and expedited downstream treatment decisions.

Inflammatory response modulation by epinephrine and norepinephrine

Relevance. Inflammation is a defense response of an organism to a pathogen. It appears in order to maintain homeostasis and is regulated by the immune, nervous, and endocrine systems. The hormones epinephrine and norepinephrine are produced in the adrenal medulla and in the brain, and are universal messengers that trigger the transmission of nerve impulses at synapses, and also have a receptor-mediated effect on immunocompetent cells. The aim of this study was to investigate adrenergic pathway regulation of inflammation on the neutrophil granulocytes in the presence of activators of innate immunity receptors. Materials and Methods. Neutrophil granulocytes were obtained from peripheral blood of healthy volunteers in a density gradient of Histopaque 1077 and Histopaque 1119 (Sigma Aldrich, Steinheim, Germany), and cultured in the presence of LPS, GMDP, epinephrine and norepinephrine. The amount of human neutrophil peptides 1-3 (HNP1-3) was examined using an enzyme-linked immunosorbent assay; the gene expression of TLR4, NOD2, ATF3 and A20 was determined using RT-PCR. Results and Discussion. Norepinephrine (noradrenaline) was found to decrease the synthesis of human neutrophils peptides 1-3 (HNP 1-3 defensins, alone and in the combination with agonists of TLR4 and NOD2 receptors - LPS and GMDP respectively. It was found out that there was no a statistically significant effect of epinephrine (adrenaline) on the production of HNP 1-3, including when combined with LPS and GMDP. As a result of the study, an increase in the levels of expression of the genes TLR4, NOD2 and regulator of inflammatory reactions A20 both in LPS- and GMDP- induced neutrophil culture were uncovered, while ATF3 was increased only in LPS-induced neutrophil culture. Epinephrine demonstrated the absence of a statistically significant effect on the expression of the studied genes. While norepinephrine significantly increased the expression of A20 genes. Conclusion. The data obtained shows that norepinephrine can reduce the synthesis of HNP 1-3, including the one induced by LPS and GMDP. Moreover, the ability of norepinephrine to induce the expression of A20 may play a significant role in modulation of inflammation.

INNOVATIVE APPROACH TO THE TREATMENT OF PSORIASIS USING NANOTECHNOLOGIES

Psoriasis is a chronic immune-mediated genetic disease with systemic and cutaneous manifestations that can significantly impair patients' quality of life. 2-3% of the world population suffers from psoriasis, and this imposes a significant economic burden on patients. Aetiology is mainly related to genes and environmental factors. The pathophysiology of psoriasis is characterized by T cells and dendritic cells, antimicrobial peptides, genetic predisposition, lipoprotein-2, galactosin-3, fractalkine, vaspin, human neutrophil peptides, etc. in the progression of psoriasis. For patients with psoriasis, conventional treatments include corticosteroids, vitamin D3 analogs, calcineurin inhibitors, methotrexate, cyclosporine, acitretin, phototherapy, and biological agents, etc. Today, there are various standard topical therapeutic approaches that can help control the condition for months to years, however, complete recovery from psoriasis with these treatments has not been reported. Therefore, researchers around the world are mainly considering the possibility of using various nanotechnological therapies for complete recovery. New drug delivery carriers, in particular nanocarriers, can overcome certain disadvantages of conventional treatment methods, such as: dose minimization, frequency of administration, and dose-dependent side effects. Nanodermatology is a new multidisciplinary science that is gaining more and more recognition in the treatment of psoriasis. The use of nanotechnology makes it possible to select drugs to achieve dermal targeting, increase efficiency and minimize unwanted effects. Currently, these nanocarriers are becoming increasingly popular as delivery vehicles for psoriasis drugs due to their non-toxicity, natural degradability, excellent biocompatibility and biodegradability, they do not cause harmful inflammatory reactions and are easily excreted from the body. Reports of nanocarrier delivery for the treatment of psoriasis have shown improved efficacy and reduced toxicity compared to standard pharmacotherapy. To better clarify the application of nanotechnology in the treatment of psoriasis, various drugs based on nanocarriers will be summarized. This review provides a concise overview of the pathophysiology, epidemiology, clinical diagnosis, and classical pharmacotherapy of psoriasis. The review also summarizes various nanotechnological treatments for the effective treatment of psoriasis.