Alpha-defensins Research Articles

Association between tissue human neutrophil peptide 1-3 levels and cardiovascular phenotype: a prospective, longitudinal cohort study.

ObjectiveInflammation is associated with atherogenesis. Although a higher neutrophil count is associated with the plaque burden, the role of neutrophil activation is unclear. Human neutrophil peptides 1–3 (HNP1–3) are a risk factor for atherogenesis in bench models and are elevated in human atheromas. This study aimed to examine the association between skin HNP1–3 deposition and the severity of coronary artery disease (CAD), including long-term outcomes.MethodsHNP1–3 levels were immunohistochemically quantified in skin biopsies, which were prospectively taken from 599 consecutive patients before clinically indicated coronary angiography. Established cardiovascular risk factors and blood markers for atheroinflammation were obtained. CAD severity and the incidence of repeat revascularization and mortality at 48 months of follow-up were assessed in relation to HNP1–3 levels.ResultsThe risk of CAD was independently associated with age and HNP1–3 in the entire cohort (F = 0.71 and F = 7.4, respectively). Additionally, HNP1–3 levels were significantly associated with myocardial necrosis (R = 0.26). At the follow-up, high HNP1–3 levels negatively affected mortality (19.54%) and recurrent revascularization (8.05%).ConclusionHNP1–3 tissue deposition is positively associated with the severity of CAD, myonecrosis, and long-term sequelae. HNP1–3 levels may be suppressed using colchicine.

The capsule of Bacillus anthracis protects it from the bactericidal activity of human defensins and other cationic antimicrobial peptides.

During infection, Bacillus anthracis bacilli encounter potent antimicrobial peptides (AMPs) such as defensins. We examined the role that B. anthracis capsule plays in protecting bacilli from defensins and other cationic AMPs by comparing their effects on a fully virulent encapsulated wild type (WT) strain and an isogenic capsule-deficient capA mutant strain. We identified several human defensins and non-human AMPs that were capable of killing B. anthracis. The human alpha defensins 1–6 (HNP-1-4, HD-5-6), the human beta defensins 1–4 (HBD-1-4), and the non-human AMPs, protegrin, gramicidin D, polymyxin B, nisin, and melittin were all capable of killing both encapsulated WT and non-encapsulated capA mutant B. anthracis. However, non-encapsulated capA mutant bacilli were significantly more susceptible than encapsulated WT bacilli to killing by nearly all of the AMPs tested. We demonstrated that purified capsule bound HBD-2, HBD-3, and HNP-1 in an electrophoretic mobility shift assay. Furthermore, we determined that the capsule layer enveloping WT bacilli bound and trapped HBD-3, substantially reducing the amount reaching the cell wall. To assess whether released capsule might also play a protective role, we pre-incubated HBD-2, HBD-3, or HNP-1 with purified capsule before their addition to non-encapsulated capA mutant bacilli. We found that free capsule completely rescued the capA mutant bacilli from killing by HBD-2 and -3 while killing by HNP-1 was reduced to the level observed with WT bacilli. Together, these results suggest an immune evasion mechanism by which the capsule, both that enveloping the bacilli and released fragments, contributes to virulence by binding to and inhibiting the antimicrobial activity of cationic AMPs.

Could AMPs and B-cells be the missing link in understanding periodontitis?

Periodontal diseases are common inflammatory conditions characterized by bone loss in response to simultaneous bacterial aggression and host defenses. The etiology of such diseases is still not completely understood, however. It has been shown that specific pathogens involved in the build-up of dysbiotic biofilms participate actively in the establishment of periodontitis. This multifactorial pathology also depends on environmental factors and host characteristics, especially defenses. The immune response to the pathogens seems to be critical in preventing the disease from starting but also contributes to tissue damage. It is known that small molecules known as antimicrobial peptides (AMPs) are key actors in the innate immune response. They not only target microbes, but also act as immuno-modulators. They can help to recruit or activate cells such as neutrophils, monocytes, dendritic cells, or lymphocytes. AMPs have already been described in the periodontium, and their expression seems to be connected to disease activity. Alpha and beta defensins and LL37 are the AMPs most frequently linked to periodontitis. Additionally, leukocyte infiltrates, especially B-cells, have also been linked to the severity of periodontitis. Indeed, the particular subpopulations of B-cells in these infiltrates have been linked to inflammation and bone resorption. A link between B-cells and AMP could be relevant to understanding B-cells’ action. Some AMP receptors, such as chemokines receptors, toll-like receptors, or purinergic receptors, have been shown to be expressed by B-cells. Consequently, the action of AMPs on B—cell subpopulations could participate to B-cell recruitment, their differentiation, and their implication in both periodontal defense and destruction.

Dental caries and associated salivary biomarkers in patients with cystic fibrosis.

To evaluate the caries status of the Cystic fibrosis (CF) children and adolescents with the comparation of some biochemical markers, secretory-immunoglobulin-A (sIgA), and antimicrobial peptides in the saliva. In this cross-sectional descriptive study, the approval Ethics Board was obtained. Unstimulated saliva samples were collected from CF and healthy control children (non-CF) patients. Both groups underwent the same dental and periodontal evaluation scheme of the assessment. Human beta defensin (HBD1), human alpha defensin (HNP-1), cathelicidin (LL-37), sIgA in saliva were evaluated by enzyme-linked immunoassay method. A general biochemical analysis was performed. Statistical analysis was performed by using Statistical Package for the Social Sciences Version 20.0 (SPSS Inc.). A total of 21 (9 male, 12 female) CF and 23 (11 male, 12 female) control patients were participated with the mean age of 10.17 ± 3.38 and 9.52 ± 2.15 years, respectively. In control children, DMFT/S (decayed-missing-filled-tooth/surface-in-permanent-dentition), dmft/s (decayed-missing-filled-tooth/surface-in-primary-dentition) values were higher; DT (decayed-tooth in permanent dentition), ft (filled-tooth in primary dentition) and plaque index values were statistically significantly higher (p = 0.042, p = 0.005, p = 0.038, respectively) than CF patients. Bicarbonate was higher in control group; sodium, chloride, and total protein were higher in CF group; magnesium, calcium and phosphate levels were similar in each group (p > 0.05). Alpha and beta defensin-1 levels in control group was statistically significantly higher (p = 0.037 and p = 0.020, respectively), while LL37 and sIgA were not statistically significantly higher (p > 0.05) than CF group. Children with CF had lower caries in permanent teeth, filling in primary teeth, and an altered salivary biomarker profile, especially in HNB1, HNP1. Therefore, it is important to conduct periodic oral-dental controls among CF patients during their childhood.

Electroacupuncture promoted intestinal defensins and rescued the dysbiotic cecal microbiota of high-fat diet-induced obese mice

Obesity is currently one of the most important challenges to public health worldwide. Acupuncture has been widely used to treat obesity. However, whether acupuncture regulates intestinal innate immunity via intestinal microbiota against obesity remains to be elucidated. In this study, electroacupuncture (EA) effectively reduced body weight and fat accumulation in obese mice persistently fed a high-fat diet. Full-length 16S rDNA sequencing showed dysbiotic microbiota in the cecum of obese mice. The composition and function of the cecal microbiota of obese mice were markedly restored after EA treatment. After 21 d of EA intervention, the expression of defensin alpha 5 (Defa5) was restored to healthy controls, whereas fat digestion and absorption genes including fabp1 were markedly decreased in the jejunum of obese mice. The Defa5 levels were positively correlated with the family Lachnospiraceae and negatively correlated with obesity indexes. EA also reduced tissue inflammation, ameliorated misaligned glucose tolerance, and inhibited key genes for intestinal lipid absorption. In summary, EA exerted an anti-obesity effect by promoting intestinal defensins, rescuing dysbiotic cecal microbiota, and reducing lipid absorption in a synergistic mode. We present for the first time the key role of alpha defensins in the relationship between gut microbiota and disease during electroacupuncture treatment of obesity. The mucosal innate immunity seems to have a stronger ability to shape the microbiota than dietary factors.

Toll-like receptor 4-mediated endoplasmic reticulum stress induces intestinal paneth cell damage in mice following CLP-induced sepsis

A marked elevation of TLR4 was observed in various organs of septic mice. The mechanism of TLR4 in intestinal epithelial cell damage in sepsis remains unclear. CLP mice models were used to assess the role of TLR4 in intestinal Paneth cell damage by histological, polymerase chain reaction, western-blot analyses. The ileal expression of TLR4 was increased by more than five-fold after CLP. CLP significantly increased 7-day mortality and was associated with a higher murine sepsis score (MSS), closely related with increased TLR4 expression. Histological staining revealed that a reduced number of Paneth cells, accompanied by reduced lysozyme and defensin alpha 5(DEF-5) expression as detected by PCR. Of note, the expression levels of ATF6, XBP1 and CHOP increased in the ileal of the sepsis group. Meanwhile, the uncleaved p90 ATF6 was markedly reduced and cleaved p50 ATF6 was increased in the sepsis group. Intriguingly, The TAK-242 had improved intestinal mucosal injury, reduced the expression of ATF6, XBP1 and CHOP and relieved the cleavage of ATF6. We found that increased the expression level of TLR4 in the ileal of CLP mice promoted the depletion of Paneth cell and reduced LYZ and DEF-5 expression. Furthermore, our findings suggested that TLR4-mediated the hyperactivation of ER stress, via activating the ATF6/CHOP pathway, might be one of the mechanisms associated with Paneth cells loss and dysfunction during intestinal barrier impairment of sepsis.

Human Neutrophil Defensins Disrupt Liver Interendothelial Junctions and Aggravate Sepsis

Human neutrophil peptides 1-3 (HNP1-3), also known as human α-defensins, are the most abundant neutrophil granule proteins. The genes that encode HNP1-3, DEFA1/DEFA3, exhibit extensive copy number variations, which correlate well with their protein levels. Human and mouse studies have shown that increased copy numbers of DEFA1/DEFA3 worsen sepsis outcomes. Additionally, high concentrations of HNP1-3 in body fluids have been reported in patients with sepsis. However, direct evidence for the pathogenic role of HNP1-3 proteins during sepsis progression is lacking. In current study, sepsis was induced by means of cecal puncture and ligation. Various doses of HNP-1 (low dose with 0.5 mg/kg body weight and high dose with 10 mg/kg body weight) or phosphate buffer saline were intraperitoneally administered to mice at six hours after sepsis onset. Survival rate was monitored, and vascular permeability, endothelial cell pyroptosis, and immunofluorescence of endothelial adherens junction protein vascular endothelial-cadherin were evaluated. The administration of a high dose of HNP-1 after sepsis onset led to increased mortality, more severe liver injury, and increased vascular permeability in the liver and mesentery. The injection of high dose of HNP-1 did not directly induce liver endothelial cell death but destroyed interendothelial junctions in the liver. Moreover, genetic deficiency of nucleotide-binding oligomerization domain-like receptor protein-3 or caspase-1 abrogated the high mortality and disrupted liver interendothelial junctions caused by high dose of HNP-1 during sepsis. This study directly demonstrates that neutrophil defensins play a key role in regulating endothelial stability during sepsis development.

PPBP gene as a biomarker for coronary heart disease risk in postmenopausal Thai women.

BackgroundEstrogen is an important ovarian hormone with anti-atherogenic and cardioprotective effects. Postmenopausal women have lower estrogen levels, associated with significantly higher risks of coronary heart disease (CHD) and CHD-related death. Effective biomarkers for the diagnosis, prediction, and treatment of CHD are needed to address this problem and thus reduce the mortality due to CHD in postmenopausal women. We recently reported that the PPBP and DEFA1/DEFA3 genes may be feasible synergistic biomarkers for CHD risk in Thai men with hyperlipidemia. The PPBP gene encodes pro-platelet basic protein (PPBP) from activated platelets, and DEFA1/DEFA3 encodes human neutrophil peptides (HNP) 1–3, mainly produced by activated neutrophils. Both platelets and neutrophils are involved in chronic inflammation during the development of atherogenesis and CHD. This study investigated the potential roles of PPBP and DEFA1/DEFA3 and their proteins as biomarkers for CHD risk in postmenopausal Thai women.MethodsThis cross-sectional study enrolled 90 postmenopausal Thai women, including 12 healthy controls (N), 18 patients with hyperlipidemia (H), and 21 patients diagnosed with CHD. The remaining 39 women were receiving cholesterol-lowering drugs for hyperlipidemia (HD) were excluded from the study. All CHD patients underwent coronary bypass grafting or coronary angioplasty. PPBP and DEFA1/DEFA3 mRNA expression levels in peripheral blood mononuclear cells isolated from heparinized blood were determined by quantitative reverse-transcription polymerase chain reaction. Levels of PPBP and HNP-1–3 proteins in corresponding plasma samples were assessed by enzyme-linked immunosorbent assay. Differences in parameters were compared among groups and correlations between parameters and clinical manifestations were analyzed.ResultsPPBP mRNA and protein levels were significantly increased in the CHD group compared with the N and H groups. In contrast, DEFA1/DEFA3 mRNA and HNP-1–3 protein levels did not differ significantly among the groups. None of the levels were associated with any of the clinical parameters analyzed in this study.ConclusionThe results indicate that gene and protein expression levels of PPBP, but not DEFA1/DEFA3, and HNP-1–3, may be feasible biomarkers for assessing CHD risk in postmenopausal Thai women with hyperlipidemia.

Alterations in Immune-Related Defensin Alpha 4 (DEFA4) Gene Expression in Health and Disease.

Defensin Alpha 4 (DEFA4) is the fourth member of the Alpha Defensins family known as a part of antimicrobial peptides in the innate immune system. DEFA4 has a strong preference to kill Gram-negative bacteria more than Gram-positive bacteria. In addition, DEFA4 exhibits antiviral activity against human immunodeficiency virus type 1 (HIV-1) in vitro. Moreover, DEFA4 can act as an inhibitor of corticosterone production (Corticostatin). On the other hand, alternations in DEFA4 gene expression have been reported in different disorders such as diseases related to inflammation and immunity dysfunction, brain-related disorders, and various cancers. The up-regulation of DEFA4 appears to be involved in the malignant transformation or aggressive form of cancer. Interestingly, the modified version of DEFA4 fragment (1–11) was potent and efficient against antibiotic-resistant bacteria. This review provides a general background abSaudi Arabia out DEFA4 and sheds light on changes in DEFA4 gene expression in different diseases. The paper also discusses other aspects related to DEFA4 as an antimicrobial and antiviral agent. The research was conducted based on available articles obtained from databases starting from 1988 to the present.

Expression of antimicrobial peptides in the amniotic fluid of women with cervical insufficiency.

Cervical insufficiency (CI) is associated with intra-amniotic infection or inflammation. Antimicrobial peptides (AMPs) in amniotic fluid may protect the fetus against microbial invasion, giving their broad-spectrum microbiocidal properties. We evaluated changes in amniotic fluid AMP expression in women with CI and assessed whether these changes are related to their pregnancy outcomes. We evaluated amniotic fluid human neutrophil peptide 3 (HNP-3), human β-defensin-2 (hBD-2), and LL-37 levels in 66 women with CI and 25 normal controls at 16-24 weeks of gestation. The CI group was divided into short cervix and cervical dilation groups, and the cervical dilation group was further divided into preterm and full-term delivery groups according to the pregnancy outcomes, and AMP expression was analyzed in each group. HNP-3 and hBD-2 levels were higher in women with CI than in normal controls and in the cervical dilation as compared to the short cervix group. Among women with cervical dilation, 22 delivered at full-term, and 23 had spontaneous preterm births. The hBD-2 level in amniotic fluid mid-pregnancy was higher in the full-term delivery than in the preterm delivery groups. However, LL-37 levels in amniotic fluid were low in women with CI and normal controls. Amniotic fluid HNP-3 and hBD-2 levels increased in women with CI compared with normal controls. Moreover, increased amniotic fluid hBD-2 levels mid-pregnancy were associated with favorable pregnancy outcomes in women with CI. AMPs in the amniotic fluid may participate in host defense against ascending infection in women with CI.

Are Leukocyte Esterase (LE) strip and Alpha-Defensin kit reliable enough to diagnose peri-prosthetic joint infection, intra-operatively?

Background: Despite improvement in sterilization techniques, peri-prosthetic joint infection (PJI) is the most fearsome complication after hip and knee arthroplasties. Various strategies have been devised from time to time to diagnose and treat PJI. Recently, Leukocyte esterase and alpha-defensin are identified as markers of active infection in synovial fluid. Therefore, kits are designed to detect these two markers during surgery. If found reliable, these tests will increase the confidence of surgeons in situations, where the diagnosis of PJI is not established. Material and methods: This study was conducted on 132 patients in Lahore General Hospital; from August 2020 to December 2021. Leukocyte Esterase strip and Alpha-Defensin kit were used to detect infection in synovial samples taken just before performing arthrotomy, intra-operatively. Patients were divided into 2 groups, Group-A had 31 patients with peri-prosthetic joint infection (PJI) diagnosed as per Musculoskeletal Infection Society (MSIS) criteria, whereas in Group B 101 patients were present in whom PJI was not present. Synovial tissue and fluid samples were also sent to a laboratory for culture and histopathology; so that sensitivity and specificity of LE strips and Alpha-Defensin Kits could find out. Results: The mean age of patients in our study was 59.6 SD 11.90 years with a male to female ratio of 85:47. 31 patients were diagnosed as PJI after primary hip or knee arthroplasty based on serological investigations and culture of joint aspiration. The sensitivity and specificity of LE strips were 90.32% and 95.04%, whereas that of Alpha defensin was 93.54% and 100% respectively. The correlation coefficient between the LE strip test and synovial fluid polymorphonuclear neutrophils (PMN) counts was 0.811 and it was even higher when the Alpha-Defensin kit was used (0.845). Conclusion: Both LE and Alpha-Defensin kits are highly specific and sensitive in diagnosing PJI. Though Alpha-defensin is more accurate the cost-effectiveness of the LE strip makes it a more feasible option in diagnosing PJI, intra-operatively.

Which Minor Criteria is the Most Accurate Predictor for the Diagnosis of Hip and Knee Periprosthetic Joint Infection in the Asian Population?

BackgroundThe aim of this study was to evaluate the diagnostic performance of minor criteria from the 2018 International Consensus Meeting (ICM) for the diagnosis of chronic periprosthetic joint infection (PJI) in an Asian population. MethodsWe retrospectively reviewed 76 patients who underwent a revision knee or hip arthroplasty at an academic institution between September 2018 and December 2019. All major and minor 2018 ICM criteria were available for all patients included. Cases with at least 1 major criterion or score ≥6 in minor criteria were considered as infected. The diagnostic performance was evaluated by a receiver operative characteristic curve analysis and area under the curve (AUC) for each minor criterion. An AUC value of more than 0.9 was considered outstanding and 0.8-0.9 as excellent. ResultsWhen using 2018 ICM threshold, the diagnostic performance ranked based on AUC was the following: alpha defensin (0.92), positive histology (0.83), leukocyte esterase (0.82), synovial white blood cell (0.81), serum erythrocyte sedimentation rate (0.78), synovial polymorphonuclear neutrophils (0.77), serum C-reactive protein (0.74), D-dimer (0.59), single positive culture (0.53), and positive intraoperative purulence (0.51). Alpha defensin was considered as an outstanding test among the 2018 ICM minor criteria. Positive histology, leukocyte esterase, and synovial white blood cell were considered as excellent tests. ConclusionBased on our findings, alpha-defensin has the best diagnostic performance in Asian population among the minor criteria of 2018 ICM.

Dietary Indole‐3‐Carbinol Supplementation Restores Epithelial Abnormalities in SAMP1/YitFc Mouse Model of Ileitis

SAMP1/YitFC (SAMP) mice exhibit spontaneous ileitis as they age with morphological and functional epithelial abnormalities mimicking human Crohn’s disease, a chronic disease characterized by ileal inflammation. Indole‐3‐carbinole (I3C), found in broccoli and Brussel sprouts, plays a crucial role in the anti‐inflammatory processes by activating the aryl hydrocarbon receptor (AhR), a transcription factor vital for epithelial and immune cell functions. The impact of I3C diet on disease progression and epithelial integrity in SAMP model of ileitis is not known.Methods6‐week‐old male and female (n=8 per group) SAMP and control AKR/J mice (Jackson) were fed for 14 weeks either a standard chow diet or a purified diet supplemented with 200 ppm with indole‐3‐carbinole (+I3C diet), or purified diet depleted of AhR ligands (‐I3C diet). Ileum samples were collected for qPCR and histological analyses. Statistical analysis was performed by Tukey’s multiple comparison test for all three diets. To detect changes between +I3C vs. ‐I3C, delta values (Δ +/‐I3C = SAMP‐AKR) were analyzed by unpaired T‐test.ResultsAs expected, H&E staining for the ileum of SAMP mice fed chow diet, showed thickening of the muscle layer, neutrophil infiltration, and epithelial tufting vs. AKR mice. Severity of inflammation in SAMP mice, as assessed by fecal calprotectin ELISA, mRNA expression of cytokines, and histological analysis was not significantly impacted by dietary depletion or supplementation of I3C. With respect to epithelial abnormalities, Periodic Acid Schiff staining showed increased number of goblet cells per villi (~47‐75%) in SAMP mice on all three diets. While mRNA levels of the major goblet cell secretory proteins, mucin2 (Muc2) and intestinal trefoil factor (ITF), were dramatically decreased in SAMP mice vs. AKR, supplementation of I3C attenuated this decrease in Muc2 and ITF by ~35% and 51% (p<.001), respectively, indicating defective synthesis or secretion. Similarly, I3C diet significantly (p<.01) attenuated the decrease in Paneth cell markers lysozyme C‐1 (LYZ1) alpha defensin 5 (Def5α) and regenerating islet‐derived protein 3 alpha (Reg3α) in SAMP mice by ~65%, 60%, and 24%, respectively. A decrease in fibroblast growth factor 15 (FGF15) and the epithelial ion transporters serotonin transporter (SERT) and organic solute transporter (OST) were observed in ileal mucosa of SAMP mice vs. AKR, and the supplementation of I3C mitigated these decreases by 30‐58% (p<.01). There were no significant changes in the expression of tryptophan hydroxylase 1 (tph‐1), C‐X‐C motif chemokine ligand 2 (CXCL2), or AhR by I3C diet in SAMP mice. In summary, I3C supplementation significantly attenuated the goblet cell, Paneth cell, and epithelial‐specific abnormalities seen in SAMP mice. These data imply that dietary supplementation of AhR ligands may be a viable therapeutic strategy for Crohn’s patients to restore epithelial integrity.

The diversity of beta defensins in lungfish (Dipnoi)

Abstract Mucosal surfaces are the first line of defense against pathogens. At these surfaces, mucus containing innate and adaptive immune molecules is constantly secreted to protect the animal host. African lungfish are the extant relatives to all tetrapods and have a dual mode of living, swimming in freshwater during the wet seasons and aestivating during the dry seasons. During terrestrialization, African lungfish produce large amounts of mucus that form a dry cocoon to prevent water loss. We recently reported that the lungfish cocoon acts as a bacterial trapping structure and it is rich in antimicrobial peptides (AMPs), specifically beta defensins (BD). As many AMPs, BDs are small amphipathic molecules and mainly cationic. The goal of this study is to provide a full characterization of BD genes from transcriptomes and genomes of different lungfish species. Using the canonical six cysteine motif from vertebrate BD molecules, we searched for all BD genes Protopterus dolloi transcriptomes, the genomes of P. annectens and Australian lungfish (a non-aestivating lungfish, Neoceratodus forsteri). BLASTs searches identified 9 BD genes in P. dolloi transcriptomes with predicted highly cationic characteristics except for BD-7. Sequence alignment and phylogenetic trees indicate that three P. dolloi BD genes may be an intermediate form of AMP between beta and alpha defensins. Genomic analyses indicate that P. annectens BD genes contain 3 exons and 2 introns similar to all defensin genes. Future studies will determine the tissue distribution and antimicrobial function of Protopterus spp. BD molecules and determine their potential application as alternatives to antibiotics. Supported by National Science Foundation Integral Organismal Systems (IOS) award # 1938816

Are Positive Urine Cultures Directly Correlated with Elevated Levels of Novel Biomarkers for Childhood Urinary Tract Infection?

Urinary tract infection (UTI) in children is one of the most common bacterial infections that propels inappropriate antibiotic use. Long-term, potentially fatal complications can occur if not properly treated. Prompt investigation and appropriate treatment would prevent these complications. Although urine culture remains the gold standard investigation for UTI, its process is cumbersome and requires time (24–72 hours). Hence, there has been growing interest in the use of urinary biomarkers. However, some conventional urinary biomarkers detected on urinalysis have poor sensitivity values when used singly as a screening tool. Thus, the searchlight has shifted to the role of novel biomarkers in UTI diagnosis. This narrative review aimed to determine if elevated levels of these biomarkers directly correlate with positive urine cultures. A positive correlation may imply that these biomarkers could serve as novel UTI diagnostics and thus augment urine culture requests. Established and recent serum and urinary biomarkers show disparate predictive abilities for UTI and its related complications. Some have elevated differential levels in upper and lower UTI or febrile and non-febrile UTI. All studies that investigated these biomarkers established culture-positive UTI, highlighting a direct correlation between positive urine cultures and increased concentrations of the biomarkers in body fluids. Because certain uropathogens were less likely to be associated with pyuria, the sensitivities of some neutrophil-related novel biomarkers (such as urine neutrophil gelatinase-associated lipocalin and human neutrophil peptides 1–3) were reduced in cases of UTI caused by these bacteria. While levels of these novel biomarkers directly correlate with positive urine cultures, it appears that there is yet no standalone biomarker with the optimal sensitivity and specificity for UTI. Although these novel biomarkers are promising, translating their measurements into clinical practice with specific clinical utilities will take time. Novel methods interrogating high-throughput serum (and urine) metabolome data with positive urine cultures in a platform-agnostic manner (metabolome-wide approach) will help confirm and identify novel biomarkers that might capture specific aetiologic agents or shared pathways of related agents. The authors recommend that future research on UTI diagnostics should specifically focus on identifying highly sensitive and specific standalone novel biomarkers that can be easily applied as a point-of-care investigation.

Concentration of alpha-defensins in the blood plasma of children with Helicobacter pylori – associated peptic ulcer of the duodenum

Introduction. Various gastrointestinal and extragastric diseases are associated with H. pylori in children and adolescents, but the strongest testing and treatment guidelines are only for children and adolescents with peptic ulcer. The most promising developments now are the study of the antibacterial effect of endogenous antimicrobial peptides (AP), among which the most important are defenses 1-3 (human neutrophil peptides, HNPs1-3). Objective. To investigate the concentration of HNPs 1-3 in children with H. pylori-associated peptic ulcer of the duodenum (DU). Methods. A study of 65 children with DU (47 children with H. pylori-associated DU, 18 children with H. pylori-negative DU) and 25 healthy children aged 7-18 years with the determination of HNPs 1-3 levels in plasma. Results. More common was H. pylori in the group of children with III grade of inflammation than in the group of children with II grade (χ2:p<0.0001). A significant association was found between mucosal inflammation activity and the presence of H. pylori (r=0.66, p<0.0001). The mean level of HNP1-3 in healthy children was 13.67±0.96 ng/ml, in children with DU it was probably higher and was 104.88±11.5 ng/ml (p=0.005). There is a direct correlation between the level of HNP1-3 and the severity of DU in children (r=0.67, p>0.05), as well as between HNP1-3 and the activity of the inflammatory process (r=0.73, p>0.01). In children with H. pylori-associated DU, HNP1-3 values were significantly higher (107.34±16.18 [98.45-119.22]) ng/ml than those in children with DU not associated with H. pylori infection (66.70±11.31 [59.54-73.29]) ng/ml and in healthy children (13.67±0.96 [7.27-18.91] ng/ml). Conclusion. Elevated plasma HNP1-3 levels in children with DU in the presence of H. pylori are likely to be a protective response to limit infection that can be used as a potential biomarker of adverse events.

Diagnostic and Therapeutic Potential for HNP-1, HBD-1 and HBD-4 in Pregnant Women with COVID-19.

Pregnancy is characterized by significant immunological changes and a cytokine profile, as well as vitamin deficiencies that can cause problems for the correct development of a fetus. Defensins are small antimicrobial peptides that are part of the innate immune system and are involved in several biological activities. Following that, this study aims to compare the levels of various cytokines and to investigate the role of defensins between pregnant women with confirmed COVID-19 infection and pregnant women without any defined risk factor. TNF-α, TGF-β, IL-2 and IL-10, β-defensins, have been evaluated by gene expression in our population. At the same time, by ELISA assay IL-6, IL-8, defensin alpha 1, defensin beta 1 and defensin beta 4 have been measured. The data obtained show that mothers affected by COVID-19 have an increase in pro-inflammatory factors (TNF-α, TGF-β, IL-2, IL-6, IL-8) compared to controls; this increase could generate a sort of “protection of the fetus” from virus attacks. Contemporarily, we have an increase in the anti-inflammatory cytokine IL-10 and an increase in AMPs, which highlights how the mother’s body is responding to the viral attack. These results allow us to hypothesize a mechanism of “trafficking” of antimicrobial peptides from the mother to the fetus that would help the fetus to protect itself from the infection in progress.

Therapeutic Role of Antimicrobial Peptides in Diabetes Mellitus

Antimicrobial peptides (AMPs) have recently become widely publicized because they have the potential to function in alternative therapies as “natural” antibiotics, with their main advantage being a broad spectrum of activity. The potential for antimicrobial peptides to treat diabetes mellitus (DM) has been reported. In diabetes mellitus type I (T1D), cathelicidin-related antimicrobial peptide (CRAMP), cathelicidin antimicrobial peptide (CAMP) and mouse-β- defensin 14 (mBD14) are positively affected. Decreased levels of LL-37 and human neutrophil peptide 1-3 (HNP1-3) have been reported in diabetes mellitus type II (T2D) relative to healthy patients. Moreover, AMPs from amphibians and social wasps have antidiabetic effects. In infections occurring in patients with tuberculosis-diabetes or diabetic foot, granulysin, HNP1, HNP2, HNP3, human beta-defensin 2 (HBD2), and cathelicidins are responsible for pathogen clearance. An interesting alternative is also the use of modified M13 bacteriophages containing encapsulated AMPs genes or phagemids.

The antimicrobial peptide alpha defensin correlates to type 2 diabetes via the advanced glycation end products pathway.

Diabetes is a serious health problem that results in high mortality rates worldwide. α-defensins are antimicrobial peptides of the innate immune system that contribute to inflammation. However, data on serum levels of α-defensin in patients suffering from type 2 diabetes are limited. This study aimed to assess the possible changes in α-defensin serum levels in patients suffering from type 2 diabetes and to investigate its correlation with relevant biomarkers. Analysis of serum α-defensin levels in 47 type 2 diabetics with diabetic neuropathy, 19 type 2 diabetics with no complications and 19 healthy control subjects by enzyme-linked immunosorbent assay was established. Furthermore, measurement of advanced glycation end products (AGEs) and fasting blood glucose (FBG) serum levels was performed, together with the lipid profile analysis. The serum levels of α-defensin were higher in patients with and without diabetic neuropathy in comparison to control subjects. In addition, there was a significant correlation between α-defensin serum levels and AGEs and FBG serum levels as well as with the body mass index. α-defensins are significantly elevated in serum of type II diabetics, and correlate with AGEs serum levels indicating a crosstalk that may aggravate inflammation in type 2 diabetes.

Recombinant HNP-1 Produced by Escherichia coli Triggers Bacterial Apoptosis and Exhibits Antibacterial Activity against Drug-Resistant Bacteria.

ABSTRACTHuman neutrophil peptide-1 (HNP-1) is a promising antibiotic candidate, but its clinical applications have been hampered by challenges during mass production and an inadequate understanding of its bactericidal mechanisms. In this study, we demonstrated that Escherichia coli expressing full-length preproHNP-1 secretes a soluble form of HNP-1, which can be recovered from the total cell lysate after isopropyl thio-β-d-galactoside (IPTG) induction and ultrafiltration. Label-free quantitative proteomics and co-immunoprecipitation experiments revealed that HNP-1 induces cell apoptosis in bacteria by causing DNA and membrane damage. Notably, we found that HNP-1 disrupts the DNA damage response pathway by interfering with the binding of RecA to single-stranded DNA (ssDNA). Further experiments demonstrated that HNP-1 encapsulated in liposomes inhibits the growth of methicillin-resistant Staphylococcus aureus (MRSA) and meropenem-resistant Pseudomonas aeruginosa (MRPA). These results indicated that recombinant protein expression may be a simple and cost-effective solution to produce HNP-1 and that RecA inhibition via HNP-1 may serve as an alternative strategy to counteract antibiotic resistance.IMPORTANCE Human neutrophil peptide-1 (HNP-1) is a promising antibiotic candidate, but its clinical application has been hampered by the difficulty of mass production and an inadequate understanding of its bactericidal mechanisms. In this study, we demonstrated that recombinant protein expression combined with ultrafiltration may be a simple and cost-effective solution to HNP-1 production. We further found that HNP-1 induces bacterial apoptosis and prevents its SOS repair pathway from binding to the RecA protein, which may be a new antibacterial mechanism. In addition, we showed that HNP-1 encapsulated in liposomes inhibits the growth of methicillin-resistant Staphylococcus aureus (MRSA) and meropenem-resistant Pseudomonas aeruginosa (MRPA). These results provide new insights into the production and antibacterial mechanism of HNP-1, both of which may promote its clinical application.