hBD-2 Concentrations Research Articles

Human Beta Defensin-2 mRNA and Proteasome Subunit β Type 8 mRNA Analysis, Useful in Differentiating Skin Biopsies from Atopic Dermatitis and Psoriasis Vulgaris Patients.

(1): Atopic dermatitis and psoriasis vulgaris are chronic, inflammatory diseases. Clinical presentation usually leads to a proper diagnosis, but sometimes neither clinical examination nor histopathological evaluation can be conclusive. Therefore, we aimed to build up a novel diagnostic tool and check it for accuracy. The main objective of our work was to differentiate between healthy skin (C), atopic dermatitis (AD) and psoriasis vulgaris (PV) biopsies on the base of involucrin (IVL) and human β-defensin-2 (hBD-2) concentrations and their mRNA, as well as mRNA for TPP2 and PSMB8. (2): ELISA for IVL and hBD-2 proteins and Real-time PCR for the relative expression of mRNA for: IVL (IVL mRNA), hBD-2 (hBD-2 mRNA), PSMB8 (PSMB8 mRNA) and TPP2 (TPP2 mRNA), isolated from skin biopsies taken from AD and PV patients and healthy volunteers were performed. (3): hBD-2 mRNA and PSMB8 mRNA correlated with some parameters of clinical assessment of inflammatory disease severity. hBD-2 mRNA expression, exclusively, was sufficient to distinguish inflammatory skin biopsies from the healthy ones. (4): hBD-2 mRNA and PSMB8 mRNA analysis were the most valuable parameters in differentiating AD and PV biopsies.

Infant Feeding, Gut Permeability, and Gut Inflammation Markers.

Increased gut permeability and gut inflammation have been linked to the development of type 1 diabetes. Little is known on whether and how intake of different foods is linked to these mechanisms in infancy. We investigated whether the amount of breast milk and intake of other foods are associated with gut inflammation marker concentrations and permeability. Seventy-three infants were followed from birth to 12 months of age. Their diet was assessed with structured questionnaires and 3-day weighed food records at the age of 3, 6, 9, and 12 months. Gut permeability was assessed with the lactulose/mannitol test and fecal calprotectin and human β-defensin-2 (HBD-2) concentrations were analyzed from stool samples at the age of 3, 6, 9, and 12 months. The associations between foods and gut inflammation marker concentrations and permeability were analyzed using generalized estimating equations. Gut permeability and gut inflammation marker concentrations decreased during the first year of life. Intake of hydrolyzed infant formula ( P = 0.003) and intake of fruits and juices ( P = 0.001) were associated with lower intestinal permeability. Intake of fruits and juices ( P < 0.001), vegetables ( P < 0.001), and oats ( P = 0.003) were associated with lower concentrations of HBD-2. Higher intake of breast milk was associated with higher fecal calprotectin concentrations ( P < 0.001), while intake of fruits and juices ( P < 0.001), vegetables ( P < 0.001), and potatoes ( P = 0.007) were associated with lower calprotectin concentrations. Higher intake of breast milk may contribute to higher calprotectin concentration, whereas several complementary foods may decrease gut permeability and concentrations of calprotectin and HBD-2 in infant gut.

Correlations between antimicrobial peptides and spectrophotometric skin color parameters in patients with basal cell carcinoma

BackgroundAntimicrobial peptides (AMPs) are active molecules in the human innate immune system, that participate in host defense and regulate the inflammation process. Previous reports have confirmed that antimicrobial peptides play a critical role in carcinogenesis.ObjectiveThe present study aimed to evaluate the correlations between plasma concentrations of AMPs and spectrophotometric parameters of skin color in patients with basal cell carcinoma and compare the results with those of healthy controls.MethodsThe plasma concentrations of cathelicidin and beta-defensin-2 in 100 patients (50 with skin cancer and 50 healthy control subjects) were measured, and skin color parameters were tested using a DermaSpectrophotometer.ResultsIn patients with basal cell carcinoma, the concentrations of cathelicidin and beta-defensin-2 were significantly higher than those in healthy controls. In healthy control patients, when erythema increases, the levels of cathelicidin and beta-defensin-2 also increase. The direction of the relationship is opposite in people with basal cell carcinoma—the concentration of antimicrobial peptides decreases and the level of erythema increases.ConclusionA significantly higher level of plasma concentrations of cathelicidin and HBD-2 are correspondent to the presence of basal cell carcinoma. Skin cancer modifies the relationship between intensity of skin erythema and the levels of cathelicidin and HBD-2. This can be related to inadequate immunological response in patients with skin cancers. New direction of research may be pioneered in searching for cytokine or mast cells disorders.

Serum, saliva, and gingival tissue human β-defensin levels in relation to retinoic acid use.

Retinoic acid is an active derivative of vitamin A and regulates the differentiation, proliferation, and antimicrobial peptide expression profiles of human cells. The aim of the present study was to analyze the effect of systemic retinoic acid use on serum, saliva, and gingival tissue levels of human β-defensin (hBD)-1, hBD-2, and hBD-3. A total of 69 participants (34 systemic retinoic acid users and 35 healthy controls) were enrolled in this study. Plaque index, probing pocket depth, bleeding on probing (BOP), and clinical attachment loss were measured. Saliva and serum hBD-1, hBD-2, and hBD-3 levels were quantified by enzyme-linked immunosorbent assay. Gingival tissue hBD-1, hBD-2, and hBD-3 levels were determined by immunohistochemistry. A univariate general linear model was used in adjusted comparisons of hBD1, hBD-2, and hBD-3. P values of < 0.05 were considered statistically significant. Reduced salivary levels of hBD-2 (P = 0.042), but not hBD-1 or hBD-3, were detected in systemic retinoic acid users compared to non-user controls. There was a significant difference in the adjusted (for BOP%) salivary hBD-2 concentrations between retinoic acid and control groups (P = 0.031). No difference was observed in serum or tissue levels of hBD-1, hBD-2, or hBD-3 between the two study groups. Systemic retinoic acid use was associated with suppressed salivary hBD-2 level, which was independent of gingival inflammation.

The Levels of the Human-β-Defensin-2 and LL-37 in the Sputum of Children with Cystic Fibrosis: A Case–control Study and Literature Review

BACKGROUND: Cystic fibrosis (CF) is a genetic disorder with an autosomal-recessive type of inheritance. Based on their host-defending and pro-inflammatory functions, antimicrobial peptides (AMPs) likely have one of the central roles in the pathogenesis of lung disease in CF. AIM: The purpose of the study was to measure the concentration of AMPs in the sputum of children with CF and evaluate any correlation with a bacterial profile of the lungs. METHODS: Lung colonization was evaluated using a culture-dependent method, sputum was utilized. A sandwich-ELISA was used to measure hBD-2 and hCAP-18/LL-37 in the sputum. RESULTS: There were 27 children enrolled in the study group, median age of inclusion was 11.4 (8.5; 14.8) years old. The control group consisted of 14 children, 11.6 (8.6; 12.6) years old. The concentration of AMPs was not correlating with participants` age (rs = −0.286, p = 0.148 – defensin hDB-2; rs = −0.084, p = 0.676 – cathelicidin hCAP-18/LL-37). The concentration of hBD-2 was from 64.01 to 813.61 pg/mL. The concentration of hCAP-18/LL-37 was from 3.24 to 35.98 ng/mL. There were significant differences in the content of AMPs on respiratory samples between study and control group (U = 976.5, p = 0.001 – for hBD-2; U = 1080.5, p &lt; 0.001). The correlation between current infection Pseudomonas aeruginosa and concentration of hBD-2 (rs = 0.167; p = 0.406) was not found. However, the presence of P. aeruginosa correlated with density of neutrophilic infiltration (rs = 0.622; p = 0.001). The concentration of hBD-2 showed direct medium correlation with total cells count (rs = 0.881, p &lt; 0.001). Correlation between current infection P. aeruginosa and concentration of hCAP-18/LL-37 (rs = 0.788; p &lt; 0.001) was observed. With increases in total cell count and relative neutrophils count, the concentration of hCAP-18/LL-37 was increased and the power of the association was medium (rs = 0.453; p = 0,018; rs = 0,592; p = 0,001). The correlation between concentrations of hBD-2 and hCAP-18/LL-37 (rs = 0.316, p &gt; 0.1) was not found. CONCLUSIONS: Measured AMPs correlated with cellular inflammatory markers and, probably, their overexpression is dedicated to stimulating a cellular component of innate immune response; there was no correlation between bacterial colonization of lungs and levels of hBD-2, so our findings sustain that P. aeruginosa is a leading but non-single contributor to persistent local inflammation in polymicrobial lungs.

Serum Relmβ combined with abdominal signs may predict surgical timing in neonates with NEC: A cohort study.

AimsTo examine the predictive value of serum biomarkers combined with other indicators for necrotizing enterocolitis (NEC) surgery decision-making.MethodsClinical data, including baseline information, clinical features, imaging presentation and serum assessment, of the infants enrolled were collected, and the serum concentrations of HBD2, HMGB-1, Claudin-3 and Relmβ were determined. Student's t test, the Mann–Whitney U test, the chi-square test and logistic regression analysis were used. Receiver operating characteristic (ROC) curves were also generated.ResultsForty-nine infants were enrolled, with 23 in the surgical NEC group and 26 in the medical NEC group. There were no differences in the baseline clinical information, including birth weight, gestational age, admission age and risk factors, during pregnancy and before enrollment (P > 0.05). Peritonitis, intestinal adhesion and sepsis were more common in the surgical group (P < 0.05). The incidences of abdominal distention, abdominal wall tenseness, abdominal tenderness and absent bowel sounds in the surgical group were significantly higher when NEC occurred (P < 0.05). There were no differences between the two groups in the imaging presentation (P > 0.05). The concentration of Relmβ {[8.66 (4.29, 19.28) vs. 20.65 (9.51, 44.65)]} in the surgical group was significantly higher (P < 0.05). Abdominal wall tenseness, abdominal tenderness and a Relmβ concentration > 19.7 μmol/L were included in the predictive model, and the AUC of the predictive score was 0.943 (95% CI: 0.891–1.000) (P < 0.05).ConclusionSerum Relmβ concentration combined with abdominal wall tenseness and abdominal tenderness may be useful in determining surgical timing in neonates with NEC.

Novel human skin surface antimicrobial peptide quantification method using a skin patch test chamber: A pilot study.

Antimicrobial peptides (AMPs) on the skin surface are related to the innate immunity of the skin in preventing external infection. Skin rinsing and tape stripping (TS) are acceptable methods for analyzing AMPs on the skin surface but have limitations, such as causing skin damage. In this study, we proposed a noninvasive method to measure AMPs on the skin surface with minimal skin damage. Using the patch test assay, we aimed to analyze the skin surface human β-defensin (hBDs) levels without damaging the skin barrier. The concentrations of hBDs on the skin surface were evaluated through the skin patch testing of 13 healthy subjects, and hBD-1 concentrations were compared with those obtained using the TS method in this proof-of-concept study. In addition, changes in skin physiology and concentration of hBDs under 1% sodium lauryl sulfate stimulation were monitored in 14 healthy subjects (8 young and 6 elderly subjects) for 150 h. The correlation between the two methods had a Pearson's coefficient of 0.640, and skin patch analysis led to a relatively less impaired barrier with no significant increase in transepidermal water loss after analysis. Age-specific comparisons suggested that higher skin surface hBD-2 concentrations were present in the young group as compared with the elderly group. Skin surface expression of hBD-2 after skin barrier disruption was also higher in the young group. Our findings show that skin patch analysis is a convenient method to analyze hBDs on the skin surface. hBDs are factors of innate immunity that can be used as an index to predict a decreased chemical immune response of skin due to aging.

Toll-like receptor-1, -2, and -6 genotypes in relation to salivary human beta-defensin-1, -2, -3 and human neutrophilic peptide-1.

To examine whether functional gene polymorphisms of toll-like receptor (TLR)1, TLR2, and TLR6 are related to the salivary concentrations of human beta-defensins (hBDs)-1, -2, -3, and human neutrophilic peptide (HNP)-1. Polymorphisms of TLR1 (rs5743618), TLR2 (rs5743708), and TLR6 (rs5743810) were genotyped by PCR-based pyrosequencing from the salivary samples of 230 adults. Salivary hBD-1, -2, -3, and HNP-1 concentrations were measured using enzyme-linked immunosorbent assay. General and periodontal health examinations, including panoramic radiography, were available for all participants. The genotype frequencies for wild types and variant types were as follows: 66.5% and 33.5% for TLR1, 95.5% and 4.5% for TLR2, and 25.1% and 74.9% for TLR6, respectively. The TLR2 heterozygote variant group exhibited higher salivary hBD-2 concentrations than the TLR2 wild-type group (p= .038). On the contrary, elevated hBD-2 concentrations were detected in the TLR6 wild-type group compared with the TLR6 heterozygote and homozygote variant group (p= .028). The associations between TLR6 genotypes and salivary hBD-2 concentrations remained significant after adjusting them for periodontal status, age, and smoking. hBD-2 concentrations in saliva are related to TLR2 and TLR6 polymorphisms, but only the TLR6 genotype seems to exhibit an independent association with the salivary hBD-2 concentrations.

Decreased beta-defensin-2 level in the gingival crevicular fluid as a potential predictor for developing inflammatory periodontal diseases

-defensin-2 (HBD-2) is a peptide of innate immunity that provides the first defenсe line in the oral mucosa against invading pathobionts. Under inflammatory conditions, epithelial cells and gingival fibroblasts produce HBD-2. The defective defensin secretion may play a crucial role in the development of inflammatory periodontal diseases. The study was aimed at comparing HBD-2 levels in the gingival fluid and/or periodontal pockets in patients with dental plaque-induced gingivitis (PG), aggressive periodontitis (AgP), chronic generalized periodontitis (CP) and in the periodontally healthy subjects (Control). We examined 142 patients (45.01.03 years) residing in Moscow, including 11 patients with PG (35.73.69 years), 43 patients with AgP (35.40.84 years), 71 patients with CP (54.40.86 years) and 17 controls (36.12.92 years). We assessed the periodontal tissue condition in all patients during the periodontal and X-ray examination. The samples of the gingival crevicular fluid and periodontal pocket contents were collected from the gingival sulcus and periodontal pockets at 8 teeth of both jaws by paper points. The concentration (C) of -defensin-2 was determined by enzyme immunoassay (ELISA Kit for Defensin Beta 2, Cloud-Clone Corp., USA). MannWhitney U-test (U), the KruskalWallis test (H) and the DwassSteelCritchlowFligner post hoc test (W) analyzed a difference significance between the parameters. We estimated the parameter relationship and its power by using the Spearmanʼs rank correlation coefficient (rS). The critical significance level was p 0.05. The current study showed that the progression of the periodontal inflammation is accompanied by sharply decreased HBD-2 concentration in patient samples (H = 42.8, df = 3, р 0.001). Thus, the concentration of HBD-2 in the gingival crevicular fluid of the periodontally healthy subjects (control group) ranged from 225 to 1720 pg/ml (C = 738 [477; 1114] pg/ml). In patients with PG, the median value of peptide concentration was 242 [42.5; 610] pg/ml (Cmin = 19 pg/ml, Cmax = 1000 pg/ml). In patients with periodontitis, it declined to critically low levels: CAgP = 54 [3; 195] pg/ml (Cmin = 0, Cmax = 478 pg/ml) and ССP = 25.5 [0; 125] pg/ml (Cmin = 0, Cmax = 298 pg/ml). Thus, we can consider the level of HBD-2 in the gingival crevicular fluid as a potential predictor for developing inflammatory periodontal diseases.

Increased fecal human beta-defensin-2 expression in preterm infants is associated with allergic disease development in early childhood

BackgroundThis study aimed to investigate whether fecal human beta-defensins (HBD)-2 and eosinophil cationic protein (ECP) expression in preterm infants are associated with allergic disease development by age 2 years. MethodsPreterm infants' stool samples were collected at the age of 6 and 12 months postnatally. Information regarding medication exposure histories (antibiotics, antipyretics, probiotics) and physician-diagnosed allergic diseases was obtained using age-specific questionnaires and medical records. We compared the 6-month and 12-month fecal HBD-2 and ECP concentrations between the medication exposure and non-exposure group, respectively, and between children who developed allergic diseases and those who did not by 2 years of age. Univariate and multivariable logistic regression analyses were performed to investigate independent variables related to physician-diagnosed allergic diseases by 2 years of age. ResultsSeventy-four preterm infants (gestational age, 31–36 weeks) were included. Fecal HBD-2 levels were significantly increased at 12 months of age among children who developed allergic diseases compared to those who did not (37.18 ± 11.80 ng/g vs. 8.56 ± 4.33 ng/g, P = 0.011). This association was more apparent among allergic children given antibiotics (50.23 ± 16.15 ng/g vs. 9.75 ± 7.16 ng/g, P = 0.008) or antipyretics (46.12 ± 14.22 ng/g vs. 10.82 ± 6.81 ng/g, P = 0.018) during the first year, whereas among allergic children who were previously not exposed to antibiotics or antipyretics, the differences were not significant. Results of the multivariable logistic regression analysis indicated that HBD-2 concentration in 12-month stools was an independent indicator associated with physician-diagnosed allergic diseases by 2 years of age (adjusted odds ratio: 1.03 [95% confidence interval: 1.00–1.05], P = 0.036). Our data revealed a lack of association between fecal ECP and allergic diseases. ConclusionsWe found that preterm infants who expressed high fecal HBD-2 at 12 months of age were associated with physician-diagnosed allergic diseases by the age of 2 years. Further studies are needed to determine the role of fecal HBD-2 in the development of allergic diseases.

Salivary levels of hBDs in children and adolescents with type 1 diabetes mellitus and gingivitis.

Type 1 diabetes mellitus (T1DM), a chronic autoimmune disease characterized by insulin deficiency, is related to periodontal diseases in children and adolescents. Our aim was to profile salivary human beta-defensin (hBD)-2 and hBD-3 concentrations in relation to periodontal and T1DM status in children and adolescent populations. Unstimulated saliva samples were collected from 66 participants including periodontally healthy T1DM patients (T1DM + C; n = 18), T1DM patients with gingivitis (T1DM + G; n = 20), systemically and periodontally healthy individuals (SH + C: n = 15), and systemically healthy gingivitis patients (SH + G; n = 13). Full mouth plaque index (PI), bleeding on probing (BOP), probing pocket depth (PPD), and clinical attachment level (CAL) were recorded. Salivary hBD-2 and hBD-3 concentrations were evaluated by sandwich ELISA method. A p value of < 0.05 was considered statistically significant. Salivary hBD-3 concentrations were lower in T1DM groups in comparison to systemically healthy counterparts (SH + G vs. T1DM + G; p < 0.001 and SH + C vs. T1DM + C; p < 0.001). Salivary hBD-2 levels did not differ between related groups. The difference in hBD-3 concentrations between T1DM and control groups was still significant (p = 0.008) after being adjusted for PI%, BOP%, and age. In the limits of study, T1DM patients were found to have decreased salivary hBD-3 concentrations, regardless of their gingival inflammatory status. Altered salivary hBD-3 concentration can partly explain why diabetic children are more prone to periodontal diseases.

До питання вітамін-D-залежної регуляції локальних механізмів неспецифічного захисту в дітей із дисплазією сполучної тканини

Актуальність. Особливе значення в дітей при дисплазії сполучної тканини (ДСТ) може мати вплив активних метаболітів вітаміну D (ВD) на реакцію захисних неспецифічних механізмів слизових оболонок. Мета. Визначити концентрацію людського â-дефензину (HBD-2) і лізоциму в локальних секретах у дітей iз ДСТ з урахуванням забезпеченості організму ВD. Матеріали та методи. Обстежено 127 дітей 11–17 років iз фенотиповими проявами ДСТ з урахуванням забезпеченості ВD. Визначено 4 групи дітей: 1-ша — здорові діти з фізіологічним рівнем 25ОНD, 2-га — діти з помірними та вираженими ступенями ДСТ та з фізіологічними концентраціями ВD (75–100 нмоль/л), 3-тя — діти з ДСТ та недостатністю 25ОНD (50–75 нмоль/л), 4-та група — діти з ДСТ та дефіцитом вітаміну D (ДВD) (нижче 50 нмоль/л). В усіх дітей умiст HBD-2 визначали методом імуноферментного аналізу та лізоциму — за методикою з використанням сухого порошку одноденної культури Micrococcus lyzodeiticus у локальних секретах (слина, копрофільтрат (КФ)). Результати. При дослідженні HBD-2 у слині найбільш високі його концентрації виявлені в дітей 1-ї групи — 4,52 ± 0,06 нг/мл. У дітей 2-ї та 3-ї групи зареєстровані більш низькі концентрації HBD-2, а в дітей iз ДСТ та ДВD показники були найменшими — 3,88 ± 0,08 нг/мл. Найбільш високі рiвнi HBD-2 у КФ виявлені в 1-й групі — 81,14 ± 5,13 нг/мл. У групах дітей iз диспластичними проявами відмічається достовірна різниця даних (р ≤ 0,05) залежно від концентрації 25ОНD, при цьому найменші концентрації виявлені в групі ДВD — 52,63 ± 3,01 нг/мл. Найвищі показники лізоциму в КФ мали діти 1-ї (4,68 ± 0,10 мг/л) та 2-ї (4,41 ± 0,09 мг/л) груп. Проте найбільш низька концентрація лізоциму виявлена в дітей iз ДСТ та ДВD — 4,09 ± 0,08 мг/л. Визначається пряма залежність між концентраціями вітаміну D та лізоциму (r = 0,65). Висновки. Отримані результати свідчать про вітамін D-залежну регуляцію продукції антимікробних пептидів в епітеліоцитах слизової оболонки кишечника при дисплазії сполучної тканини. Вираженість порушень продукції HBD-2 та лізоциму в дітей iз ДСТ та ДВD обумовлює доцільність проведення корекції вітаміну D у цієї категорії пацієнтів.

Expression of epidermal antimicrobial peptides is increased in tinea pedis.

Tinea pedis is often chronic or recurrent, but not all individuals are equally susceptible to this infection. Dermatophytes are able to induce the expression of antimicrobial peptides and proteins (AMPs) in human keratinocytes and certain AMPs can inhibit the growth of dermatophytes. The focus of this study was to analyse the secretion of relevant AMPs, especially RNase 7, human beta-defensin-2 (hBD-2) and the S-100 protein psoriasin (S100A7), in patients with confirmed tinea pedis. To verify the diagnosis, skin scales were obtained from all patients (n=13) and the dermatophytes were identified by potassium hydroxide mount, culture and molecular analysis. To determine the AMP concentrations, the lesional skin area of the foot was rinsed with a buffer that was subsequently analysed by ELISA. The corresponding area of the other unaffected foot as well as defined healthy skin areas of the forearm and forehead and samples from age and gender-matched healthy volunteers served as controls. In tinea pedis patients the AMP concentrations were higher in lesional skin than in non-lesional skin and in healthy skin of controls. In particular, concentrations of hBD-2 and psoriasin were significantly elevated. The induction of AMPs in tinea pedis might be triggered directly by the dermatophytes; furthermore, attendant inflammation and/or differentiation processes may play a role. Our results indicate that there is no defect in the constitutive expression and induction of the analysed AMPs by dermatophytes in the epidermis of affected patients. However, it is not known why the elevated AMP concentrations fail to efficiently combat dermatophyte growth.

Gingival crevicular fluid levels of Interleukin-22 (IL-22) and human β Defensin-2 (hBD-2) in periodontal health and disease : A correlative study

BackgroundInterleukin 22(IL-22), a recently identified, Th-22 associated cytokine has a key role in the production of human β defensin-2(hBD-2) and hence an indirect role in innate, nonspecific immunity. Hence this study was conducted as preliminary research to quantify and correlate the levels of IL-22 and hBD-2 in periodontal disease. Materials and methodsGingival crevicular fluid from subjects with chronic periodontitis (n = 27), gingivitis (n = 25) and healthy controls (n = 27) were obtained for the study. The periodontal status of each subject was assessed by criteria based on plaque index, gingival index, bleeding on probing, probing depth and clinical attachment loss. The levels of IL-22 and hBD-2 were measured in GCF samples with an enzyme-linked immunosorbent assay. ResultPeriodontal parameters were described and reported as mean values. The distribution and comparison of clinical parameters and gingival crevicular fluid IL-22, hBD-2 levels in healthy controls, gingivitis and chronic periodontitis groups were assessed using one way ANOVA test. Post Hoc Bonferroni test was used for intergroup comparisons. Association among the IL-22 , hBD-2 and clinical parameters in all three groups were examined using Pearson's correlation test.The IL-22 level was significantly greater in chronic periodontitis group than healthy controls (P < 0.001) and gingivitis group (P < 0.001). The hBD-2 level was significantly higher in the chronic periodontitis group compared to gingivitis (P = 0.003) and healthy controls (P < 0.001). The results also showed a statistically significant correlation (P = 0.002) between IL-22 and hBD-2 concentration in chronic periodontitis group. ConclusionThese results indicate the role of IL- 22 and hBD-2 in the innate immune response during periodontitis.

Serum vitamin D concentration is associated with antimicrobial peptide level in periodontal diseases

ObjectiveThe aim of the study was to assess whether or not Vitamin D deficiency was associated with the GCF and gingival tissue antimicrobial peptides (AMP), namely, human beta defensin-2 (hBD-2) and cathelicidin (LL-37) level in chronic periodontitis (CP) and gingivitis patients. DesignA total of 80 volunteers were included in this study. Forty was classified as Vitamin D deficient (25(OH)D <20 ng/mL), and 40 Vitamin D sufficient patients (25(OH)D ≥ 20 ng/mL). Of these, 20 of them were affected by gingivitis and 20 by CP. Following sampling, the hBD-2 and LL-37 concentration in gingival tissues and GCF were determined by the ELISA method. ResultsThe hBD-2 and LL-37 levels were higher in periodontitis compared to gingivitis patients within Vitamin D sufficient and deficient groups. The AMP levels of GCF and gingival tissue in the vitamin D deficient group was lower compared to sufficient serum 25(OH)D within gingivitis and CP groups. Additionally, a non-parametric regression model known as the generalized additive model was used to identify the contribution of diagnosis, Vitamin D status, and other potential clinical variables on the local levels of AMPs. Regression analysis showed that the periodontal disease status, serum vitamin D concentration were independent predictors for elevated GCF AMP levels. Finally, a positive correlation between GCF and tissue levels of both hBD-2 (r = 0.82; <0.0001) and LL-37 (r = 0.65; <0.0001) was detected. ConclusionThis study shows that serum 25(OH)D deficiency is associated with decreased hBD-2 and LL-37 expression of GCF and gingival tissues in both gingivitis and CP patients.

Activation of Sirtuin 1 (SIRT1) Signaling by Resveratrol Increases Human Beta‐Defensins‐2 and ‐3 (HBD2, HBD3) Production in Response to Lipopolysaccharide (LPS) in Human Amniotic Epithelial Cells (HAEC): Pregnancy Complicated by Diabetes (PCD) vs. Normoglycemic Pregnancy (NP)

Human beta defensins (HBDs) are small cationic peptides with chemoattractant activities. HBD2 and HBD3 are generally expressed at low levels in physiologic conditions and are induced in response to microbial challenge (e.g. LPS), whereas HBD1 low expression is constitutive, with little regulation in response to infection or other stimuli. HBDs produced by HAEC co‐create an innate antimicrobial immune response in the materno‐placento‐fetal unit. PCD increases the risk of infection. According to recent studies, PCD activates the innate immune response through TLR5 or TLR1/2 on neonatal monocyte, but the data about HBDs are still lacking. The aim of this study was to examine comparatively the effect of resveratrol on SIRT1 level and HBD2 and HBD3 production in HAEC isolated after PCD (White’s class C) vs. NP. HAEC were isolated from the amnion immediately after cesarean section delivery terminating PCD (N = 24, group I) and NP (N = 24, group II) using trypsin‐based method and cultured in vitro under normoxia at 37 degrees C. Upon reaching confluence, HAEC were cultured in media supplemented with 0.5 μM resveratrol and SIRT1 levels were determined in HAEC culture lysates obtained once a day for 7 consecutive days. After pretreatment with LPS (1 μg/mL), HBD2 and HBD3 concentrations in the HAEC culture supernatant, obtained in the days 1,3 and 7 of the culture with resveratrol, were assessed in both goups using ELISA. Nonparametric Wilcoxon rank‐sum test was used and the differences between the groups were considered statistically significant if p &lt; 0.05. In both groups resveratrol increased significantly SIRT1 levels in consecutive days. The response was stronger in PCD as compared to NP. Despite of this, observed increase in both HBD2 and HBD3 concentrations were significantly lower in PCD, amounting to 32.8 / 37.7, 45.5 / 42.9 and 55.3% / 49.5% (HBD2 / HBD3 days 1,3 and 7, respectively) of the values obtained in group II (NP). Importantly, the mean viability of cells in both groups did not differ significantly at the consecutive time points. Thus, susceptibility to infections in diabetes may be to some extent caused by compromised HBDs‐dependent antimicrobial immune respose. The nature of impaired SIRT1 – HBDs interaction is still under investigation.Support or Funding InformationWUM grant: 2M2‐W2‐18/2M2‐W1‐19

Influence of chewing on expression level of human beta-defensin 2 and secretory immunoglobulin A in the epithelium

PurposeThis study aimed to clarify the influence of chewing on human β-defensin 2 (hBD-2) and secretory immunoglobulin A (SIgA) expression levels. MethodsWe included 15 healthy males with no missing teeth (mean age, 25.5±2.5years). Subjects were instructed to chew a piece of gum for 30min. Saliva and skin-extraction samples were collected before and after chewing for 15 and 30min. hBD-2 and SIgA concentrations in the samples were determined using enzyme-linked immunosorbent assay (ELISA). hBD-2 and SIgA expression levels before and after chewing were analyzed using the Mann–Whitney U test, following the Friedman test. The significance level was 0.05. ResultsThe hBD-2 level in skin-extraction samples was significantly different before (99.4±17.3pg/mL) and after chewing for 30min (142±23.0pg/mL). The SIgA level in skin-extraction samples was also significantly different before (2.39±0.25μg/mL) and after chewing for 30min (3.61±0.33μg/mL). No significant difference was noted in either hBD-2 or SIgA secretion rate in saliva between before and after chewing. ConclusionsChewing gum for 30min increased hBD-2 and SIgA expression levels in skin. Moreover, chewing gum could influence the secretion pattern of these two biomolecules on skin, but not in saliva.

Salivary human beta-defensins affected by oral Candida status in Chinese HIV/AIDS patients undergoing ART.

To observe relationships between oral Candida status and salivary human beta-defensin 2 and 3 (hBD-2 and hBD-3) levels in HIV/AIDS patients of Guangxi, China during the first year of antiretroviral therapy (ART) dynamically, and to understand the influence of ART on oral Candida status and salivary hBDs expressions. A prospective self-controlled study was carried to observe the dynamic changes of CD4+ T cell counts, oral Candida carriages and salivary hBD-2,3 expressions in HIV/AIDS patients during the first year of ART. A total of 90 HIV/AIDS patients were enrolled and were examined at the baseline, 3rd, 6th, 12th month of ART. Thirty healthy individuals were enrolled as control. Peripheral blood, oral rinse sample, and unstimulated whole saliva were collected to test CD4+ T cell counts, oral Candida carriages, and hBD-2,3 expressions. In the first year of ART, CD4+ T cell counts increased significantly. However, oral Candida carriages and oral candidiasis decreased significantly, and salivary hBD-2 expressions in HIV/AIDS patients decreased gradually, salivary hBD-3 levels were highly variable. Salivary hBD-2 concentrations were positively related to oral Candida carriages. The incidence of oral candidiasis among HIV/AIDS patients gradually decreased due to the immune reconstruction of ART. Salivary defensins might play an important role in Candida-host interaction in HIV/AIDS patients.

Comparison of IL-1β, TNF-α, hBD-2, and hBD-3 Expression in the Dental Pulp of Smokers Versus Nonsmokers

IntroductionTo date, the endodontic literature lacks research on the effect of smoking on cytokine and defensin expression in the dental pulp. Therefore, the aim of this study was to investigate the expression of interleukin (IL)-1β, tumor necrosis factor (TNF)-α, human beta defensin (hBD)-2 and hBD-3 in the dental pulp of smokers and compare them with nonsmokers. We hypothesized that cytokine and defensin expression would be reduced in smokers as compared with nonsmokers. MethodsThirty-two smokers and 37 nonsmokers with endodontic pulpal diagnoses of normal, symptomatic irreversible pulpitis and asymptomatic irreversible pulpitis were included in this cross-sectional study. Samples from pulp chambers were collected and stored in phosphate-buffered saline at −80°C. Luminex was used to measure IL-1β and TNF-α levels. The levels of hBD-2 and hBD-3 were measured using enzyme-linked immunosorbent assay. Marker levels were normalized to protein concentrations and data were analyzed using Kruskal-Wallis test, Mann-Whitney U test, and 2-way analysis of variance (α = 0.05). ResultsPulpal concentrations of TNF-α and hBD-2 were significantly lower among smokers (P < .01), whereas no significant difference was observed for IL-1β, or hBD-3. Two-way analysis of covariance revealed that smoking status (P < .001), not endodontic diagnosis (pulpal status), significantly affected TNF-α and hBD-2 levels. ConclusionsThis study reported that smokers are immunologically deficient in TNF-α and hBD-2, suggesting that dental pulps of smokers possess limited defense mechanisms, affecting their endodontic prognosis and indicating a cause for their reported inferior outcome.

Impact of high-dose vitamin D3 on plasma free 25-hydroxyvitamin D concentrations and antimicrobial peptides in critically ill mechanically ventilated adults.

High-dose vitamin D3 increases plasma total 25-hydroxyvitamin D [25(OH)D] in critically ill, ventilated patients; however, to our knowledge, the effect on plasma levels of free (nonprotein-bound) 25(OH)D has not been investigated in critical illness. Moreover, the relationship of free 25(OH)D and the regulation of endogenous antimicrobial peptides (AMPs) remains unknown. The aims of this study were to determine in critically ill adults with respiratory failure the effect of previous high-dose regimens of vitamin D3 on free 25(OH)D concentrations, the relationship of free 25(OH)D with circulating cathelicidin (LL-37) and human beta-defensin-2 (hBD-2), and the associations between plasma levels of free 25(OH)D and these AMPs to alveolar macrophage phagocytosis function. In a double blind, randomized controlled trial, critically ill ventilator-dependent adults (N=30) received enteral vitamin D3 (250,000 or 500,000 IU total over 5d) or placebo. Plasma was obtained serially for concentrations of free 25(OH)D, LL-37, hBD-2, and expression of peripheral blood mononuclear cell human cationic antimicrobial protein (hCAP18) mRNA. Total 25(OH)D and LL-37 concentrations and alveolar macrophage phagocytosis were determined in bronchoalveolar lavage fluid. Plasma concentrations of free 25(OH)D over time were correlated with total 25(OH)D levels (r=0.82; P<0.001). The increase in free 25(OH)D was greater with the 500 000 IU vitamin D3 dose than with the lower dose. The percent change in mRNA expression of hCAP18 was positively associated with percent change in free 25(OH)D at days 7 and 14 (ρ=0.48; P=0.04 and ρ=0.59; P=0.03, respectively). Additionally, plasma LL-37 levels correlated with the percentage of alveolar macrophages exhibiting phagocytosis (ρ=0.51; P=0.04). The present study found a dose-related increase in plasma free-25(OH)D levels, which was associated with increasing circulating mRNA expression of hCAP18 over time. There were no correlations between changes in total and free 25(OH)D against plasma LL-37 and hBD-2 concentrations. Larger studies appear warranted to determine the impact of high-dose vitamin D3 administration on endogenous AMPs.