Identification of human host factors required for beta-defensin-2 expression in intestinal epithelial cells upon a bacterial challenge.

The human intestinal tract is colonized with microorganisms, which present a diverse array of immunological challenges. A number of antimicrobial mechanisms have evolved to cope with these challenges. A key defense mechanism is the expression of inducible antimicrobial peptides (AMPs), such as beta-defensins, which rapidly inactivate microorganisms. We currently have a limited knowledge of mechanisms regulating the inducible expression of AMP genes, especially factors from the host required in these regulatory mechanisms. To identify the host factors required for expression of the beta-defensin-2 gene (HBD2) in intestinal epithelial cells upon a bacterial challenge, we performed a RNAi screen using a siRNA library spanning the whole human genome. The screening was performed in duplicate to select the strongest 79 and 110 hit genes whose silencing promoted or inhibited HBD2 expression, respectively. A set of 57 hits selected among the two groups of genes was subjected to a counter-screening and a subset was subsequently validated for its impact onto HBD2 expression. Among the 57 confirmed hits, we brought out the TLR5-MYD88 signaling pathway, but above all new signaling proteins, epigenetic regulators and transcription factors so far unrevealed in the HBD2 regulatory circuits, like the GATA6 transcription factor involved in inflammatory bowel diseases. This study represents a significant step toward unveiling the key molecular requirements to promote AMP expression in human intestinal epithelial cells, and revealing new potential targets for the development of an innovative therapeutic strategy aiming at stimulating the host AMP expression, at the era of antimicrobial resistance.

The Antimicrobial Activity of Human Defensins at Physiological Non-Permeabilizing Concentrations Is Caused by the Inhibition of the Plasma Membrane H+-ATPases.

Human defensins are cysteine-rich peptides (Cys-rich peptides) of the innate immune system. Defensins contain an ancestral structural motif (i.e., γ-core motif) associated with the antimicrobial activity of natural Cys-rich peptides. In this study, low concentrations of human α- and ß-defensins showed microbicidal activity that was not associated with cell membrane permeabilization. The cell death pathway was similar to that previously described for human lactoferrin, also an immunoprotein containing a γ-core motif. The common features were (1) cell death not related to plasma membrane (PM) disruption, (2) the inhibition of microbicidal activity via extracellular potassium, (3) the influence of cellular respiration on microbicidal activity, and (4) the influence of intracellular pH on bactericidal activity. In addition, in yeast, we also observed (1) partial K+-efflux mediated via Tok1p K+-channels, (2) the essential role of mitochondrial ATP synthase in cell death, (3) the increment of intracellular ATP, (4) plasma membrane depolarization, and (5) the inhibition of external acidification mediated via PM Pma1p H+-ATPase. Similar features were also observed with BM2, an antifungal peptide that inhibits Pma1p H+-ATPase, showing that the above coincident characteristics were a consequence of PM H+-ATPase inhibition. These findings suggest, for the first time, that human defensins inhibit PM H+-ATPases at physiological concentrations, and that the subsequent cytosolic acidification is responsible for the in vitro microbicidal activity. This mechanism of action is shared with human lactoferrin and probably other antimicrobial peptides containing γ-core motifs.

Evaluation of Kynu, Defb2, Camp, and Penk Expression Levels as Psoriasis Marker in the Imiquimod-Induced Psoriasis Model.

Background: Psoriasis is a noncontagious auto-inflammatory chronic skin disease. So far, some of the inflammatory genes were upregulated in mouse model of psoriasis. This study examined changes in skin mRNA expression of L-kynureninase (Kynu), cathelicidin antimicrobial peptide (Camp), beta-defensin 2 (Defb2), and proenkephalin (Penk) in a mouse model of imiquimod-induced psoriasis. Materials and Methods: Tree groups of C57BL/6 female mice were allocated. The imiquimod (IMQ) cream was administered to the mice dorsal skin of the two groups to induce psoriatic inflammation. In the treatment group, IMQ was administered 10 min after hydrogel-containing M7 anti-IL-17A aptamer treatment. Vaseline (Vas) was administered to the negative control group. The psoriatic skin lesions were evaluated based on the psoriasis area severity index (PASI) score, histopathology, and mRNA expression levels of Kynu, Camp, Defb2, and Penk using real-time PCR. In order to assess the systemic response, the spleen and lymph node indexes were also evaluated. Results: The PASI and epidermal thickness scores were 6.01 and 1.96, respectively, in the IMQ group, and they significantly decreased after aptamer administration to 1.15 and 0.90, respectively (P < 0.05). Spleen and lymph node indexes showed an increase in the IMQ group, followed by a slight decrease after aptamer treatment (P > 0.05). Additionally, the mRNA expression levels of Kynu, Defb2, Camp, and Penk genes in the IMQ-treated region showed a significant 2.70, 4.56, 3.29, and 2.61-fold increase relative to the Vas mice, respectively (P < 0.05). The aptamer-treated region exhibited a significant decrease in these gene expression levels (P < 0.05). A positive correlation was found between Kynu, Penk, and Camp expression levels and erythema, as well as Camp expression with PASI, scaling, and thickness (P < 0.05). Conclusion: According to our results, it seems that Kynu, Camp, and Penk can be considered appropriate markers for the evaluation of psoriasis in IMQ-induced psoriasis. Also, the anti-IL-17 aptamer downregulated these important genes in this mouse model.

Metagenomic Analysis of Rhizospheric Bacterial Community of Citrus Trees Expressing Phloem-Directed Antimicrobials.

Huanglongbing, also known as citrus greening, is currently the most devastating citrus disease with limited success in prevention and mitigation. A promising strategy for Huanglongbing control is the use of antimicrobials fused to a carrier protein (phloem protein of 16 kDa or PP16) that targets vascular tissues. This study investigated the effects of genetically modified citrus trees expressing Citrus sinensis PP16 (CsPP16) fused to human lysozyme and ß-defensin-2 on the soil microbiome diversity using 16S amplicon analysis. The results indicated that there were no significant alterations in alpha diversity, beta diversity, phylogenetic diversity, differential abundance, or functional prediction between the antimicrobial phloem-overexpressing plants and the control group, suggesting minimal impact on microbial community structure. However, microbiota diversity analysis revealed distinct bacterial assemblages between the rhizosphere soil and root environments. This study helps to understand the ecological implications of crops expressing phloem-targeted antimicrobials for vascular disease management, with minimal impact on soil microbiota.

Effect of milking time on yield, composition, and antimicrobial components of milk in lactating goats.

Various studies have attempted to improve the milk yield and composition in dairy animals. However, no study has examined the effects of milking at different times on milk yield and composition. Therefore, this study aimed to compare the yield, composition, and antimicrobial components of milk obtained from milking at different times in lactating goats. Eight goats were milked once daily at different times for three consecutive weeks (first week: 06:00 h; second week: 09:00 h; and third week: 12:00 h). The light ranged from 06:30 to 19:00 h. Milk and blood samples were collected once a day during milking time. Milking at 09:00 h resulted in a significantly higher milk yield than that obtained after milking at 06:00 and 12:00 h. Prolactin levels in plasma and the fat, Na+, ß-defensin, and S100A7 (antimicrobial component) levels in milk were the lowest in the 09:00 h milking. These results indicate that milk yield, composition, and antimicrobial components can be affected by milking time, which may be related to the altered concentration of prolactin in the blood. These findings provide a rational basis for achieving maximal milk production with strong immunity by changing to a more effective milking time.

Histone demethylase JMJD2D protects against enteric bacterial infection via up-regulating colonic IL-17F to induce ß-defensin expression.

Histone demethylase JMJD2D (also known as KDM4D) can specifically demethylate H3K9me2/3 to activate its target gene expression. Our previous study has demonstrated that JMJD2D can protect intestine from dextran sulfate sodium (DSS)-induced colitis by activating Hedgehog signaling; however, its involvement in host defense against enteric attaching and effacing bacterial infection remains unclear. The present study was aimed to investigate the role of JMJD2D in host defense against enteric bacteria and its underlying mechanisms. The enteric pathogen Citrobacter rodentium (C. rodentium) model was used to mimic clinical colonic infection. The responses of wild-type and JMJD2D-/- mice to oral infection of C. rodentium were investigated. Bone marrow chimeric mice were infected with C. rodentium. JMJD2D expression was knocked down in CMT93 cells by using small hairpin RNAs, and Western blot and real-time PCR assays were performed in these cells. The relationship between JMJD2D and STAT3 was studied by co-immunoprecipitation and chromatin immunoprecipitation. JMJD2D was significantly up-regulated in colonic epithelial cells of mice in response to Citrobacter rodentium infection. JMJD2D-/- mice displayed an impaired clearance of C. rodentium, more body weight loss, and more severe colonic tissue pathology compared with wild-type mice. JMJD2D-/- mice exhibited an impaired expression of IL-17F in the colonic epithelial cells, which restricts C. rodentium infection by inducing the expression of antimicrobial peptides. Accordingly, JMJD2D-/- mice showed a decreased expression of ß-defensin-1, ß-defensin-3, and ß-defensin-4 in the colonic epithelial cells. Mechanistically, JMJD2D activated STAT3 signaling by inducing STAT3 phosphorylation and cooperated with STAT3 to induce IL-17F expression by interacting with STAT3 and been recruited to the IL-17F promoter to demethylate H3K9me3. Our study demonstrates that JMJD2D contributes to host defense against enteric bacteria through up-regulating IL-17F to induce ß-defensin expression.

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 ß-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.

Gut Inflammation Markers, Diet, and Risk of Islet Autoimmunity in Finnish Children - A Nested Case-Control Study.

BACKGROUND: Gut dysbiosis and increased intestinal permeability have been reported to precede type 1 diabetes-related autoimmunity. The role of gut inflammation in autoimmunity is not understood. OBJECTIVES: This study aimed to assess whether gut inflammation markers are associated with risk of islet autoimmunity and whether diet is associated with gut inflammation markers. METHODS: A nested case-control sample of 75 case children with islet autoimmunity and 88 control children was acquired from the Finnish Type 1 Diabetes Prediction and Prevention cohort. Diet was assessed with 3-d food records, and calprotectin and human ß-defensin-2 (HBD-2) were analyzed from stool samples at 6 and 12 mo of age. Conditional logistic regression analysis was used in a matched case-control setting to assess risk of autoimmunity. Analysis of variance, independent samples t test, and a general linear model were used in secondary analyses to test associations of background characteristics and dietary factors with inflammation markers. RESULTS: In unadjusted analyses, calprotectin was not associated with risk of islet autoimmunity, whereas HBD-2 in the middle (odds ratio [OR]: 3.23; 95% confidence interval [CI]: 1.03, 10.08) or highest tertile (OR: 3.02; 95% CI: 1.05, 8.69) in comparison to the lowest at 12 mo of age showed borderline association (P-trend = 0.063) with higher risk of islet autoimmunity. Excluding children with cow milk allergy in sensitivity analyses strengthened the association of HBD-2 with islet autoimmunity, whereas adjusting for dietary factors and maternal education weakened it. At age 12 mo, higher fat intake was associated with higher HBD-2 (ß: 0.219; 95% CI: 0.110, 0.328) and higher intake of dietary fiber (ß: -0.294; 95% CI: -0.510, -0.078), magnesium (ß: -0.036; 95% CI: -0.059, -0.014), and potassium (ß: -0.003; 95% CI: -0.005, -0.001) with lower HBD-2. CONCLUSIONS: Higher HBD-2 in infancy may be associated with higher risk of islet autoimmunity. Dietary factors play a role in gut inflammatory status.

Supplementation with organic yeast-derived selenium provides immune protection against experimental necrotic enteritis in broiler chickens.

Necrotic enteritis (NE) is a potentially fatal poultry disease that causes enormous economic losses in the poultry industry worldwide. The study aimed to evaluate the effects of dietary organic yeast-derived selenium (Se) on immune protection against experimental necrotic enteritis (NE) in commercial broilers. Chickens were fed basal diets supplemented with different Se levels (0.25, 0.50, and 1.00 Se mg/kg). To induce NE, Clostridium perfringens (C. perfringens) was orally administered at 14 days of age post hatch. The results showed that birds fed 0.25 Se mg/kg exhibited significantly increased body weight gain compared with the non-supplemented/infected birds. There were no significant differences in gut lesions between the Se-supplemented groups and the non-supplemented group. The antibody levels against α-toxin and NetB toxin increased with the increase between 0.25 Se mg/kg and 0.50 Se mg/kg. In the jejunal scrapings and spleen, the Se-supplementation groups up-regulated the transcripts for pro-inflammatory cytokines IL-1ß, IL-6, IL-8, iNOS, and LITAF and avian ß-defensin 6, 8, and 13 (AvBD6, 8 and 13). In conclusion, supplementation with organic yeast-derived Se alleviates the negative consequences and provides beneficial protection against experimental NE.

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.

Single-cell and spatial transcriptomic investigation reveals the spatiotemporal specificity of the beta-defensin gene family during mouse sperm maturation.

Low sperm motility is a significant contributor to male infertility. beta-defensins have been implicated in host defence and the acquisition of sperm motility; however, the regulatory mechanisms governing their gene expression patterns and functions remain poorly understood. In this study, we performed single-cell RNA and spatial transcriptome sequencing to investigate the cellular composition of testicular and epididymal tissues and examined their gene expression characteristics. In the epididymis, we found that epididymal epithelial cells display a region specificity of gene expression in different epididymal segments, including the beta-defensin family genes. In particular, Defb15, Defb18, Defb20, Defb25 and Defb48 are specific to the caput; Defb22, Defb23 and Defb26 to the corpus; Defb2 and Defb9 to the cauda of the epididymis. To confirm this, we performed mRNA fluorescence in situ hybridisation (FISH) targeting certain exon region of beta-defensin genes, and found some of their expression matched the sequencing results and displayed a close connection with epididimosome marker gene Cd63. In addition, we paid attention to the Sertoli cells and Leydig cells in the testis, along with fibroblasts and smooth muscle cells in the epididymis, by demonstrating their gene expression profile and spatial information. Our study provides a single-cell and spatial landscape for analysing the gene expression characteristics of testicular and epididymal environments and has important implications for the study of spermatogenesis and sperm maturation.

Dietary tryptophan improves growth and intestinal health by promoting the secretion of intestinal ß-defensins against enterotoxigenic Escherichia coli F4 in weaned piglets.

Adequate dietary L-tryptophan (Trp) governs intestinal homeostasis in piglets. However, the defensive role of Trp in the diet against enterotoxigenic Escherichia coli F4 (K88) in pigs is still poorly understood. Here, sixty (6.15 ± 1.52 kg, 24-day-old, Duroc × Landrace × Yorkshire) weaned piglets were used for an E. coli F4 attack test in a 2 × 2 factorial design. The growth (ADG, ADFI, GH), immune factors (IL-10, IgA, IgG, IgM), Trp metabolite 5-HT, intestinal morphology (jejunal and colonic VH), mRNA expression of ß-defensins (jejunal BD-127, BD-119, ileal BD-1, BD-127), and abundance of beneficial microorganisms in the colon (Prevotella 9, Lactobacillus, Phascolarctobacterium, Faecalibacterium) were higher in the piglets in the HT (High Trp) and HTK (High Trp, K88) groups than in the LT (Low Trp) and LTK (Low Trp, K88) groups (P<.05), while FCR, diarrhea rate, diarrhea index, serum Trp, Kyn, IDO, D-LA, ET, and abundance of harmful microorganisms in the colon (Spirochaetes, Fusobacteria, Prevotella, Christensenellaceae R7) were lower in the HT and HTK groups than in the LT and LTK groups (P<.05). High Trp reduced the expression of virulence genes (K88 and LT) after E. coli F4 attack (P<.05). The IL-6, TNF-α was lower in the HTK group than in the LT, LTK group (P<.05). In short, a diet containing 0.35% Trp protected piglets from enterotoxigenic E. coli F4 (K88) via Trp metabolism promoting BD expression in the intestinal mucosa, which improved growth and intestinal health.

Possible Impact of Human ß-defensin 1 on sperm motility in infertile men with abnormal sperm parameters.

Human ß-defensins and interleukins may be auxiliary in sperm maturation. This cross-sectional study aimed to evaluate the expression of Human ß-defensins 1 and 2, interleukins (ILs)- 10 and -18 genes in sperm, as well as seminal plasma levels of these two cytokines in subfertile men with different types of sperm abnormalities compared to those with normozoospermic men. Participants were separated into two experimental groups: the control group (n = 25) and the group with sperm abnormalities (SA) (n = 45). SA participants were further subdivided into the following groups with n = 15 individuals each: Teratozoospermia (T), Asthenoteratozoospermia (AT), and Oligoasthenoteratozoospermia (OAT) groups. The quantitative real-time polymerase chain reaction was used to quantify the mRNA levels of hBDs 1 and 2, IL-10, and IL-18 in sperm. The seminal plasma concentrations of IL-10 and IL-18 were measured by using the enzyme-linked immunosorbent assay technique. The mRNA expression of hBD-1 and IL-10 showed a significant decrease in the OAT compared to the controls (P < 0.0001 and P = 0.02, respectively). The lowest seminal plasma concentration of IL-10 belonged to the OAT (P = 0.04). ROC curve analysis showed a sensitivity, specificity, and cutoff value of 82.35%, 86.67%, and 0.63 for hBD-1 levels, respectively. A positive and significant correlation was found between hBD-1 expression and sperm motility and IL-10 expression rate and normal sperm morphology.Therefore, hBD-1 could be considered as the alternative biomaterial to pre-treatments of infertile men with abnormal sperm parameters, specifically OAT men, which led to improving the assisted reproduction success rate.

Change in Tissue Microbiome and Related Human Beta Defensin Levels Induced by Antibiotic Use in Bladder Carcinoma.

It is now generally accepted that the success of antitumor therapy can be impaired by concurrent antibiotic therapy, the presence of certain bacteria, and elevated defensin levels around the tumor tissue. The aim of our current investigation was to identify the underlying changes in microbiome and defensin levels in the tumor tissue induced by different antibiotics, as well as the duration of this modification. The microbiome of the tumor tissues was significantly different from that of healthy volunteers. Comparing only the tumor samples, no significant difference was confirmed between the untreated group and the group treated with antibiotics more than 3 months earlier. However, antibiotic treatment within 3 months of analysis resulted in a significantly modified microbiome composition. Irrespective of whether Fosfomycin, Fluoroquinolone or Beta-lactam treatment was used, the abundance of Bacteroides decreased, and Staphylococcus abundance increased. Large amounts of the genus Acinetobacter were observed in the Fluoroquinolone-treated group. Regardless of the antibiotic treatment, hBD1 expression of the tumor cells consistently doubled. The increase in hBD2 and hBD3 expression was the highest in the Beta-lactam treated group. Apparently, antibiotic treatment within 3 months of sample analysis induced microbiome changes and defensin expression levels, depending on the identity of the applied antibiotic.

Drug repositioning for immunotherapy in breast cancer using single-cell analysis.

Immunomodulatory peptides, while exhibiting potential antimicrobial, antifungal, and/or antiviral properties, can play a role in stimulating or suppressing the immune system, especially in pathological conditions like breast cancer (BC). Thus, deregulation of these peptides may serve as an immunotherapeutic strategy to enhance the immune response. In this meta-analysis, we utilized single-cell RNA sequencing data and known therapeutic peptides to investigate the deregulation of these peptides in malignant versus normal human breast epithelial cells. We corroborated our findings at the chromatin level using ATAC-seq. Additionally, we assessed the protein levels in various BC cell lines. Moreover, our in-house drug repositioning approach was employed to identify potential drugs that could positively impact the relapse-free survival of BC patients. Considering significantly deregulated therapeutic peptides and their role in BC pathology, our approach aims to downregulate B2M and SLPI, while upregulating PIGR, DEFB1, LTF, CLU, S100A7, and SCGB2A1 in BC epithelial cells through our drug repositioning pipeline. Leveraging the LINCS L1000 database, we propose BRD-A06641369 for B2M downregulation and ST-4070043 and BRD-K97926541 for SLPI downregulation without negatively affecting the MHC complex as a significantly correlated pathway with these two genes. Furthermore, we have compiled a comprehensive list of drugs for the upregulation of other selected immunomodulatory peptides. Employing an immunotherapeutic approach by integrating our drug repositioning pipeline with single-cell analysis, we proposed potential drugs and drug targets to fortify the immune system against BC.

Clinicopathological studies on ulcerative lymphangitis in cattle: Alterations in serum inflammatory cytokines, anti-microbial, organs functions, and oxidative stress-related biomarkers.

Background: Affection with Corynebacterium pseudotuberculosis (C. pseudotuberculosis) and development of cellulitis and/or abscess formation with cutaneous lymphangitis in cattle is rare to some extent, so literature about the biochemical changes that would accompany this infection is rare. Aim: In this context, the present study was designed to screen the effect of the infection with C. pseudotuberculosis cutaneous lymphangitis on the release of some immune molecules, organ functions, and redox state in Baladi cows. Methods: Fourteen Baladi cows from a small dairy farm in El-Behira, Egypt, were selected to complete this study. After bacteriological culture confirmation, seven of them were found suffering from cutaneous lesions due to infection with C. pseudotuberculosis (Diseased group), while the others were healthy (Healthy group). Serum samples were obtained to evaluate the presumptive changes in some clinicopathological parameters. Results: Serum analysis revealed a significant decrease in the levels of interferon-gamma and interleukin-17 as well as a significant decrement in the concentration of beta-defensin (ß-defensin) and lipocalin-2. While serum level of interleukin-10 recorded a significant increase in these animals when compared to healthy control animals. Concurrently, the affected animals recorded a significant elevation in serum levels of hepato-cardiac enzymes, urea, and creatinine in addition to disturbance in the serum redox state. Conclusion: In conclusion, infection with C. pseudotuberculosis cattle may disturb the defensive immune state, body organ function, and redox state of the animals.

Values of serum intestinal fatty acid-binding protein, fecal calprotectin, and fecal human ß-defensin 2 for predicting necrotizing enterocolitis.

BACKGROUND: This study aimed to assess the diagnostic potential of serum intestinal fatty acid-binding protein (I-FABP), fecal calprotectin (FC), and fecal human ß-defensin 2 (hBD2) in predicting necrotizing enterocolitis (NEC) in preterm infants. METHODS: A prospective cohort of neonates with a gestational age < 32 weeks, suspected of NEC, was enrolled between June 2021 and December 2022. Serum I-FABP, FC, and fecal hBD2 levels were measured upon NEC suspicion, and diagnosis was confirmed through radiological examination or surgical intervention. Diagnostic precision of serum I-FABP, FC, and fecal hBD2 was assessed using a logistic regression model with multiple variables. RESULTS: The study included 70 neonates (45 males, 25 females), with 30 developing NEC (40% Stage III, n = 12; 60% Stage II, n = 18) and 40 in the control group. NEC patients exhibited significantly higher serum I-FABP and FC levels (4.76 ng/mL and 521.56 µg/g feces, respectively) than those with other diagnoses (1.38 ng/mL and 213.34 µg/g feces, respectively; p ˂ 0.05 for both biomarkers). Stage II NEC neonates showed elevated fecal hBD2 levels (376.44 ng/g feces) than Stage III NEC neonates and controls (336.87 ng/g and 339.86 ng/g feces, respectively; p ˂ 0.05). No such increase was observed in infants progressing to Stage III NEC. Using a serum I-FABP threshold of > 2.54 ng/mL yielded 76.7% sensitivity, 87.5% specificity, 82.1% positive predictive value (PPV), and 83.3% negative predictive value (NPV). For FC (cutoff > 428.99 µg/g feces), corresponding values were 76.7% sensitivity, 67.5% specificity, 63.9% PPV, and 79.4% NPV. CONCLUSION: Serum I-FABP and FC levels are valuable for early NEC detection and provide insights into disease severity. Low fecal hBD2 levels suggest an inadequate response to luminal bacteria, potentially rendering these infants more susceptible to NEC development or exacerbation.

Effect of transporting Oreochromis niloticus in water with and without sodium chloride on skin morphology and some immunity-related genes expression.

The aim of this study was to examine the effects of salt addition on the skin gene expression of Mucin, Antimicrobial peptides, cortisol, and glucose in Oreochromis niloticus after 5-hour transportation in water. Three groups were compared: Control, post-transport without salt (PT-S), and post-transport with 5 g salt-1(PT + S), with a stocking density of 28.6 gL-1, 20 fish for each experimental group. The results showed that the PT-S group had more significant changes in gene expression than the PT + S group, suggesting that salt alleviated the stress and immune responses of O. niloticus. The PT-S group had higher expression of mucin- 2(MUC + 2) (7.58 folds) and mucin-5AC (MUC5-AC) (6.29 folds) than the PT + S group (3.30 folds and 4.16 folds, respectively). The PT-S group also had lower expression of ß-defensin-1 (Dß1) (0.42 folds), ß-defensin-2 (Dß2) (0.29 folds), and Cath1 (0.16 folds) than the PT + S group (0.82 folds, 0.69 folds, and 0.75 folds, respectively). The skin morphology of the PT-S group revealed some white patches with no goblet cell openings, while the PT + S group had better preservation of skin features with some goblet cell openings and slight white patches. This study indicates that O. niloticus can benefit from sodium chloride during transportation, as it helps to reduce stress and inflammation, balance mineral levels, enhance health and immunity, and regulate mucous secretion.

Beta-defensin index: A functional biomarker for oral cancer detection.

There is an unmet clinical need for a non-invasive and cost-effective test for oral squamous cell carcinoma (OSCC) that informs clinicians when a biopsy is warranted. Human beta-defensin 3 (hBD-3), an epithelial cell-derived anti-microbial peptide, is pro-tumorigenic and overexpressed in early-stage OSCC compared to hBD-2. We validate this expression dichotomy in carcinoma in situ and OSCC lesions using immunofluorescence microscopy and flow cytometry. The proportion of hBD-3/hBD-2 levels in non-invasively collected lesional cells compared to contralateral normal cells, obtained by ELISA, generates the beta-defensin index (BDI). Proof-of-principle and blinded discovery studies demonstrate that BDI discriminates OSCC from benign lesions. A multi-center validation study shows sensitivity and specificity values of 98.2% (95% confidence interval [CI] 90.3-99.9) and 82.6% (95% CI 68.6-92.2), respectively. A proof-of-principle study shows that BDI is adaptable to a point-of-care assay using microfluidics. We propose that BDI may fulfill a major unmet need in low-socioeconomic countries where pathology services are lacking.

Association of intestinal inflammation and permeability markers with clinical manifestations of Parkinson's disease.

INTRODUCTION: Intestinal inflammation and gut microbiota dysbiosis can stimulate degeneration of dopaminergic neurons and development of Parkinson's disease (PD) via the gut-brain axis in certain patients. METHODS: In a case-control study, fecal markers of intestinal inflammation and permeability were measured using the ELISA method in PD patients and healthy controls. Motor and nonmotor symptoms were assessed using the Movement Disorder Society (MDS) Unified PD Rating Scale, Hoehn & Yahr scale, MDS Non-Motor Symptom Scale, Scales for Outcomes in PD - Autonomic Dysfunction, PD Sleep Scale - 2, Montreal Cognitive Assessment, Beck Anxiety Inventory, and Beck Depression Inventory-II. A correlation was established between the intestinal inflammation and permeability markers and PD symptoms. RESULTS: Higher levels of beta-defensin 2, zonulin and lactoferrin were recorded in PD patients compared to controls. Calprotectin and secretory immunoglobulin A showed no significant differences. Regression analysis indicated the roles of beta-defensin 2 and lactoferrin in predicting PD likelihood. Calprotectin yielded positive correlations with disease duration, depression, motor fluctuations, and gastrointestinal symptoms; beta defensin 2 with thermoregulation; and secretory immunoglobulin A with depression. Secretory immunoglobulin A showed negative correlation with age and age at disease onset, while zonulin showed negative correlation with the MDS Unified PD Rating Scale total score. CONCLUSIONS: Fecal markers differed in PD patients compared to controls and correlated with age, disease duration, and some nonmotor symptoms. Future studies should identify the subgroups of PD patients that are likely to develop intestinal inflammation.