Can Salivary Biomarkers Be Used as Predictors of Dental Caries in Young Adolescents?

BACKGROUND Identifying caries predictors in the subpopulation at risk is one of the preconditions for developing effective caries prevention measures. The present exploratory study aimed to examine the significance of socio-demographic characteristics, dietary-hygiene habits, salivary pH, and salivary antimicrobial HNP-1, hBD-2, and LL-37 peptides as potential caries risk predictors in children ages 11-13 years. MATERIAL AND METHODS This prospective 1-year study enrolled 213 children ages 11-13 years. The subjects underwent a dental examination and their mothers were interviewed. Unstimulated saliva was collected from the subjects to determine its pH value, as well as the salivary levels of HNP-1, hBD-2, and LL-37 peptides in 85 of the subjects. After 12 months, the 1-year caries incidence rate was recorded. Logistic regression analysis was used to estimate the ability of selected variables to predict caries risk. RESULTS The univariable logistic regression analysis determined that the most significant independent caries risk predictors were: sex (female) (OR=2.132, p=0.007), mothers' education (OR=1.986, p=0.020), salivary pH (OR=0.270, p=0.043), oral hygiene index (OR=1.886, p=0.015), and daily tooth brushing frequency (OR=0.565, p=0.042). The multivariable model showed that sex and oral hygiene-related variables were the most important caries predictors. CONCLUSIONS Salivary HNP-1, hBD-2, and LL-37 peptides were not found to have a significant predictive value. Therefore, socio-demographic and oral hygiene variables remain important caries predictors in early adolescents, suggesting the importance of the mechanical control of biofilm as the key measure for preventing caries. However, there is still a need for effective caries risk biomarkers, and additional research is needed in this area of caries risk prediction.

Reduced induction of human ß-defensins is involved in the pathological mechanism of cutaneous adverse effects caused by epidermal growth factor receptor monoclonal antibodies.

Epidermal growth factor receptor inhibitors (EGFRIs) frequently cause cutaneous adverse effects such as papulopustular eruptions. However, the mechanism of the reactions remains unclear. To assess the pathological mechanism of cutaneous adverse reactions caused by EGFRIs, we investigated whether EGFRIs have an influence on the innate immune response of the skin. Levels of human ß-defensins (hBDs), which serve as the first line of defence against infection by pathogenic microorganisms, in the stratum corneum samples of patients treated with EGFR. monoclonal antibodies were measured before and after starting therapy. There were no obvious trends in hBD production in patients without eruptions, whereas a significant decrease in hBD1 and hBD3 production and a nonsignficant decrease in hBD2 production were observed in patients who developed papulopustular eruptions. Our results suggest that a reduction in hBD contributes to the increased incidence of papulopustular eruptions.

Microbiological characterisation of the colonisation by Candida sp in patients with orthodontic fixed appliances and evaluation of host responses in saliva.

OBJECTIVES: We investigated the colonisation by Candida spp in patients using orthodontic fixed appliances by characterising the isolated Candida strains and by evaluating the host oral mucosa response through the measure of human ß-defensins 3 (HBD-3) expression and Interleukin-1ß/IL-10. METHODS: Ninety patients were enrolled after signing an informed consent. Prevalence, susceptibility to fluconazole, genotyping and oral fungal burden of Candida sp. isolated were determined. Host responses were evaluated by measuring HBD-3 expression as well as IL-1ß and IL-10 in saliva. RESULTS: The colonisation rate reached 6.7% (6/90), and 5 patients were colonised with C. albicans strains and one with one with C. tropicalis. The fluconazole MIC90/susceptibility of C. albicans strains ranged 1/0.25-1 µg/mL. However, isolated strains did not present different genotype (SAB>0.9), C. albicans colonisation seems to be influenced by the duration of treatment and by level expression of HBD3 that were higher in colonised patients (not statistically different). A negative correlation between the fungal burden and IL-1ß levels was found in colonised patients but not for IL-10. CONCLUSIONS: Our study revealed that patients with orthodontic fixed appliances were mainly colonised by C. albicans, which was related to a decrease in HBD-3 expression and IL-1ß levels.

[Effect of Bifidobacterium on the expression of ß-defensin-2 in intestinal tissue of neonatal rats with necrotizing enterocolitis].

OBJECTIVE: To study the effect of Bifidobacterium on the expression of ß-defensin-2 (BD-2) in intestinal tissue of neonatal rats with necrotizing enterocolitis (NEC). METHODS: A total of 40 rats were randomly divided into four groups: normal control, Bifidobacterium control, NEC model, and Bifidobacterium treatment, with 10 rats in each group. A rat model of NEC was induced by hypoxia, cold stimulation, and artificial feeding. The rats in the Bifidobacterium control and Bifidobacterium treatment groups were given Bifidobacterium via the gastric tube after cold stimulation once a day for three consecutive days. The morphological changes of the terminal ileum were observed under a light microscope and the intestinal injury score was determined. Immunohistochemistry and qRT-PCR were used to measure the protein and mRNA expression of BD-2 in the ileal mucosal tissue. RESULTS: The NEC model group had a significantly higher intestinal injury score than the normal control, Bifidobacterium control, and Bifidobacterium treatment groups (P<0.05). The Bifidobacterium treatment group had a significantly higher intestinal injury score than the normal control and Bifidobacterium control groups (P<0.05). The mRNA and protein expression of BD-2 in the normal control group was significantly lower than in the Bifidobacterium control, NEC model, and Bifidobacterium treatment groups (P<0.05). The Bifidobacterium control group had significantly higher mRNA and protein expression of BD-2 than the NEC model and Bifidobacterium treatment groups (P<0.05). The Bifidobacterium treatment group had significantly higher mRNA and protein expression of BD-2 than the NEC model group (P<0.05). CONCLUSIONS: Bifidobacterium can induce the expression of BD-2 in intestinal tissue of rats and reduce inflammatory response by increasing the expression of BD-2. This provides a protective effect on neonatal rats with NEC.

Effect of steroid hormones on the innate immune response induced by Staphylococcus aureus in the goat mammary gland.

The objective of this study was to compare the dynamics of innate immune components after intramammary infusion of Staphylococcus aureus (SA) under conditions of high oestrogen and high progesterone in goats. In one group ("E-group"), controlled internal drug release (CIDR) devices were inserted intravaginally from days -11 to -4. Prostaglandin F2α was administered immediately after removal of the CIDR device at day -3, and then oestradiol benzoate (E) was injected intramuscularly once a day from days -2 to 3. Heat-inactivated SA was then administered via intramammary infusion to the left udder at day 0, whilst only saline was infused to the right udder as a control. In a second group ("P-group"), CIDR devices were inserted intravaginally from days -3 to 7 and SA was infused at day 0 in the same way as in the E-group. The milk yield and the concentration of innate immune components (somatic cell count (SCC), lactoferrin (LF), S100A7 and goat ß-defensin 1 (GBD-1)) in the milk were measured. Milk yield decreased drastically in both SA and control udders in the E-group, whereas the P-group exhibited increased milk yield in both SA and control udders. SCC increased after SA infusion in both E- and P-groups, although it was higher in the E-group than in the P-group. There was no significant change in LF concentration in the E-group, but a decrease was observed in the P-group. Concentrations of S100A and GBD-1 were significantly increased after SA infusion in the E-group but not in the P-group. These results suggest that E enhances the innate immune response induced by SA in the goat mammary gland. This effect may be due to the reduction in milk yield and upregulation of innate immune components.

Cervicovaginal Levels of Human ß-Defensin 1, 2, 3, and 4 of Reproductive-Aged Women With Chlamydia trachomatis Infection.

OBJECTIVE: This study included women attending primary health care units in Botucatu, São Paulo, Brazil, to assess the cervicovaginal levels of human ß-defensin (hBD) 1, 2, 3, and 4 during Chlamydia trachomatis infection. PATIENTS AND METHODS: Cervicovaginal samples were collected for Pap testing and assessing the presence of infection by C. trachomatis, human papillomavirus, Neisseria gonorrhoeae, and Trichomonas vaginalis. Vaginal smears were taken to evaluate local microbiota. Human ß-defensin levels were determined using enzyme-linked immunosorbent assay in cervicovaginal fluid samples. Seventy-four women with normal vaginal microbiota and no evidence of infection were included in hBD quantification assays; 37 tested positive for C. trachomatis and 37 were negative. Statistical analysis was performed using Mann-Whitney U test. RESULTS: Women positive for C. trachomatis had significantly lower cervicovaginal hBD-1, hBD-2, and hBD-3 compared with those who tested negative (hBD-1: 0 pg/mL [0-2.1] vs 1.6 pg/mL [0-2.4], p < .0001; hBD-2: 0 pg/mL [0-3.9] vs 0.61 pg/mL [0-8.9], p = .0097; and hBD-3: 0 pg/mL [0-4.3] vs 0.28 pg/mL [0-8.4], p = .0076). Human ß-defensin 4 was not detected. CONCLUSIONS: Lower levels of hBD-1, hBD-2, and hBD-3 in cervicovaginal fluid were detected in the presence of C. trachomatis infection.

Ratio of T-Helper Type 1 (Th1) to Th17 Cytokines in Whole Blood Is Associated With Human ß-Defensin 3 Expression in Skin and Persistent Staphylococcus aureus Nasal Carriage.

BACKGROUND: Nasal colonization has gained attention as an effect modifier in Staphylococcus aureus vaccine trials, suggesting interference of carriage with T-cell immunity. Likewise, T-cell signals may be involved in regulating effectors of epithelial innate defense. METHODS: Whole blood from 43 persistent carriers and 49 noncarriers was stimulated with viable S. aureus T-helper type 1 (Th1), Th2, and Th17 cytokine expression was measured, compared between carrier groups, and linked with data on human ß-defensin 3 (hBD-3) messenger RNA (mRNA) in skin while adjusting for transcriptionally relevant promoter haplotypes. RESULTS: Compared with carriers, stimulated whole blood from noncarriers contained on average 60% more interferon γ mRNA (P = .031) and 19% less interleukin 17A (IL-17A) mRNA (P = .11), resulting in, on average, a 90% higher IFN-γ to IL-17A mRNA ratio (P = .003). In a multivariable model, per duplication of the mRNA template, the risk of being a carrier increased by 93% for IL-17A (odds ratio [OR], 1.93; 95% confidence interval [CI], 1.10-3.41; P = .023) and decreased by 35% for IFN-γ (OR, 0.65; 95% CI, 0.47-0.91; P = .01). Independent of carriage and DEFB promotor haplotype, a 1-unit increase in the IFN-γ to IL-17A mRNA ratio (mean ± SD, 5.93 ± 1.60) led to a 24% increase in hBD-3 transcription in experimentally wounded human skin (P = .003). CONCLUSIONS: A low Th1 to Th17 mRNA ratio increases the propensity for persistent S. aureus nasal colonization, with downregulated hBD-3 transcription providing a potential link.

Human beta-defensin 1, 2 and 3 production by amniotic epithelial cells with respect to human papillomavirus (HPV) infection, HPV oncogenic potential and the mode of delivery.

BACKGROUND: Human beta-defensins (HBD) produced by human amniotic epithelial cells (HAEC) co-create an innate antiviral immune response in the materno-placento-fetal unit. Oncogenic potential of HPV may reflect its ability to avoid immune recognition. In this study we assessed the risk of HAEC infection with human papillomavirus (HPV) in relation to the type of labor and the impact of the oncogenic potential of HPV on HBD production in HAEC. METHODS: A comparative analysis [HPV(+) vs. HPV(-)HAEC] of the production of HBD were performed. HAEC were isolated from placentas of 116 HPV(+) and 36 HPV(-) parturients (groups I and II, respectively) using trypsin-based method. The cases of premature rupture of membranes (PROM), natural labors (NL) and cesarean sections (CS) were analysed in respective subgroups. High-risk (HR-HPV) and low-risk (LR-HPV) genotypes of HPV in cervical smears and HAEC were identified using the Roche Linear Array(®) HPV Genotyping Test. HBD-1,-2,-3 concentrations in the HAEC culture supernatant were assessed using ELISA. RESULTS: The highest percentage (42.1%) of HPV transmission to HAEC occurred in PROM, an intermediate value was observed after NL (38.5%), and the lowest (25.6%) after CS. The mean concentrations of HBD-2 and HBD-3 in group I were up to 3.1- and 2.8-fold higher (p < 0.05), respectively. The mean concentration of HBD-2 was higher (p < 0.05) in LR-HPV infection compared with HR-HPV. CONCLUSIONS: The course of labor and the mode of delivery influence the risk of HPV transmission to the HAEC. HPV infection upregulates HBD-2 and HBD-3 production in HAEC. Smaller increases in HBD-2 level after HR-HPV infection as compared to LR-HPV may affect cancerogenesis. Therapeutic potential of HBD-2 for HR-HPV infection should be assessed in future studies.

Reduced levels of genital tract immune biomarkers in postmenopausal women: implications for HIV acquisition.

BACKGROUND: Rates of HIV infections are increasing in older adults. Although it is known that the HIV/AIDS epidemics affects women disproportionately, little is known regarding immune functions in the genital tract of postmenopausal women, as relevant to HIV susceptibility. OBJECTIVE: The objective of the study was to compare levels of female reproductive tract immune mediators that are important for HIV-associated immune responses as well as intrinsic anti-HIV activity in the cervical vaginal lavages collected from HIV-negative pre- and postmenopausal women. STUDY DESIGN: Cervical vaginal lavage from 20 premenopausal and 20 postmenopausal women were assayed for interleukin-6, interleukin-8, tumor necrosis factor-α, secretory leukocyte protease inhibitor, elafin, human ß-defensin-2, and macrophage inflammatory protein-3α using standard enzyme-linked immunosorbent assays. Anti-HIV activity of cervical-vaginal lavage was measured using TZM-bl indicator cells against HIV-1 IIIB and BaL. Whereas each postmenopausal woman provided only 1 sample, each premenopausal woman provided 3 samples, during proliferative, ovulatory, and secretory stages, based on menstrual dates. RESULTS: We observed significantly lower levels of tumor necrosis factor-α, MIP-3α, secretory leukocyte protease inhibitor, elafin, and human ß-defensin-2 in cervical vaginal lavage from postmenopausal women compared with premenopausal women. Inhibition of HIV-1 infection was observed for both pre- and postmenopausal women, but cervical vaginal lavage from postmenopausal women showed significantly higher inhibition against HIV-1 BaL after adjusting for total protein concentration, genital pH, and reproductive tract infections. No change in mediators or HIV inhibition was observed through the stages of menstrual cycle. In addition, we observed that postmenopausal women with reproductive tract infections had significantly higher levels of tumor necrosis factor-α and significantly lower levels of interleukin-8, which were not observed in premenopausal women. CONCLUSION: Our findings suggest that female reproductive tract immune microenvironment is distinct in HIV-negative postmenopausal women. Further studies are needed to assess the risk of HIV acquisition/transmission in this population.

Expression of Antimicrobial Peptides in Human Monocytic Cells and Neutrophils in Response to Dengue Virus Type 2.

BACKGROUND/AIMS: The innate immune response is remarkably important for controlling infections. Information about the participation of antimicrobial peptides (AMPs) in response to dengue virus (DENV) is scarce. The aim of this study was to examine the AMP response to DENV-2 in human THP-1 cells and neutrophils. METHODS: Protein and mRNA levels of two AMPs - hBD-1 and cathelicidin LL-37 - were assessed in DENV-infected macrophage-like THP-1 cells using qRT-PCR and indirect immunofluorescence. Also, mRNA levels of α-defensins (hDEFAs) and LL-37 were examined by qRT-PCR in human neutrophils taken from peripheral blood and treated with DENV-2. RESULTS: mRNA expression of hBD-1 rose in THP-1 cells at 24-72 h, while protein expression increased later, from 48 to 72 h after infection. Cathelicidin LL-37 mRNA expression of DENV-infected THP-1 cells was observed at 6-48 h after infection, while protein levels increased importantly up to 72 h after infection. Regarding neutrophils, the mRNA expression of hDEFAs and LL-37 increased slightly at 2 and 5 h after the contact with DENV-2. CONCLUSION: THP-1 cells and human neutrophils strongly respond to DENV by producing AMPs: hBD-1 and LL-37 for the THP-1 cells and hDEFAs and LL-37 for neutrophils. However, the direct effect of these molecules on DENV particles remains unclear.

Cigarette Smoke Modulates NOD1 Signal Pathway and Human ß Defensins Expression in Human Oral Mucosa.

BACKGROUND/AIMS: Nucleotide binding oligomerization domain 1 (NOD1) signal pathway and human ß defensins (hBDs) play crucial roles in innate immune. Cigarette smoke has been confirmed to dampen innate immune in some human tissues, such as oral mucosa. The aim of this study was to evaluate potential effects of smoking on NOD1 signaling and hBDs expression in oral mucosa. METHODS: Tissue specimens of normal oral mucosa were collected from donors undergoing routine surgical treatment. All 20 participants were classified equally as two groups: non-smokers and smokers. By using Western blotting and immunohistochemistry, we investigated differential expression of crucial molecules in NOD1 signal pathway, hBD-1, -2, and -3 in oral mucosa tissues between non-smokers and smokers. Immortalized human oral mucosal epithelial (Leuk-1) cells were treated with various concentrations of cigarette smoke extract (CSE) for 24h. Western blotting and immunofluorescence assays were performed to study CSE-induced alteration of protein expression. Leuk-1 cells were treated with 4% CSE, iE-DAP (NOD1 agonist), CSE + iE-DAP, BAY 11-7082 (NF-κB inhibitor), 4% CSE + BAY 11-7082, respectively. Real-time PCR and ELISA were performed to detect the mRNA levels and secretion of hBD-1, -2, and -3, respectively. RESULTS: The levels of NOD1, NF-κB, hBD-1 and hBD-3 significantly reduced in oral mucosa tissues of smokers compared with non-smokers. The levels of RIP2 (receptor-interacting protein 2), phospho-NF-κB (P-NF-κB) and hBD-2 remarkably enhanced in oral mucosal tissues of smokers. CSE treatment suppressed NOD1 and NF-κB expression and activated RIP2 and P-NF-κB expression in Leuk-1 cells. The mRNA and secretory levels of hBD-1 and -3 were down-regulated by CSE, while the mRNA and secretory level of hBD-2 were up-regulated by CSE. The iE-DAP or BAY 11-7082 treatment reversed the regulatory effects of CSE on levels of hBDs. CONCLUSION: The present study indicated that cigarette smoke could potentially modulate the expression of crucial molecules of NOD1 signal pathway and hBDs in human oral mucosal epithelium. NOD1 signal pathway could play an important role in the regulatory effects of CSE on hBDs levels in oral mucosal epithelial cells.

Relative quantification of human ß-defensins by a proteomics approach based on selected reaction monitoring.

RATIONALE: A targeted proteomics method based on selected reaction monitoring (SRM) is a relevant approach for the analysis of multiple analytes in biological samples. Defensins are phylogenetically conserved small antimicrobial peptides contributing to innate host defense and exhibiting low immunogenicity, resistance to proteolysis and a broad range of antimicrobial activities. The goal of the present study was to develop and optimize SRM-based targeted proteomics methods for the detection of human ß-defensins 1-4 in various biological fluids. METHODS: An SRM-based targeted proteomics method was developed and validated for the detection of human ß-defensins 1-4. The supernatants of resting and IL-1ß-stimulated Caco2, HT-29 and SW-1116 colonic epithelial cells (CEC), cell lysates of CECs and tear samples of human healthy individuals were analyzed and the feasibility of the developed method was validated by ELISA and dot-blot analysis complemented by RT-qPCR. RESULTS: Our results demonstrate that the developed SRM method offers an alternative approach for the cost-effective and rapid analysis of human ß-defensins in samples with biological relevance. CONCLUSIONS: A semi-quantitative targeted mass spectrometry method was developed and validated for the relative quantification of ß-defensins 1-4 in cell culture supernatants and body fluid analyses.

Relationship between expression of human gingival beta-defensins and levels of periodontopathogens in subgingival plaque.

BACKGROUND AND OBJECTIVE: Human beta-defensins (hBDs) are a group of antimicrobial peptides important in epithelial innate immunity, and their differential expression is associated with periodontal diseases. The aim of this study was to explore relationships among hBDs, total subgingival bacteria and periodontopathogens in healthy subjects and in patients with chronic periodontitis. MATERIAL AND METHODS: The periodontal clinical parameters of 29 healthy subjects and 25 patients with chronic periodontitis were recorded. The relative expression of hBD1, hBD2 and hBD3 genes in gingival biopsies was measured using real-time PCR. The numbers of total bacteria and of Treponema denticola, Aggregatibacter actinomycetemcomitans, Porphyromonas gingivalis, Fusobacterium nucleatum and Tannerella forsythia in subgingival plaque were quantified by real-time PCR. Data were analyzed using the Mann-Whitney U-test and Spearman's rank correlation test. RESULTS: No significant differences in expression of the hBD genes were found between the group of healthy subjects and the group of patients with chronic periodontitis. Total bacteria and T. denticola were detected in all participants. F. nucleatum and T. forsythia were detected in all patients with chronic periodontitis and in 86.21% and 51.72%, respectively, of healthy volunteers. P. gingivalis and A. actinomycetemcomitans were detected in 24.14% and 17.24%, respectively, of the healthy group and in 84.00% and 12.00%, respectively, of the chronic periodontitis group. The prevalence of all bacteria, except A. actinomycetemcomitans, was significantly higher in the group of patients with chronic periodontitis than in the group of healthy subjects (p < 0.05). A significant, negative correlation was observed between total bacteria and hBD-2 (r = -0.384, p = 0.011). Upon analyzing the data in different groups, total bacteria and hBD-2 were significantly correlated (r = -0.492, p = 0.026) only in the group of healthy subjects. CONCLUSION: The negative correlations between hBD-2 and total bacteria, especially in the group of healthy subjects, indicate that hBDs may play an important role by limiting an increase of bacterial load at the initial stage of periodontitis.

Periodontopathogens and human ß-defensin-2 expression in gingival crevicular fluid from patients with periodontal disease in Guangxi, China.

BACKGROUND AND OBJECTIVE: Periodontal diseases are often induced by periodontopathogens, which are always exposed to certain innate immune factors in gingival crevicular fluid, including human ß-defensin-2 (hBD-2). This study aims to investigate the relationship among periodontopathogens, clinical parameters and hBD-2 expression. MATERIAL AND METHODS: Thirty-two healthy controls, 42 patients with chronic gingivitis and 95 patients with chronic periodontitis were recruited in Guangxi, China. Bleeding index, probing depth and clinical attachment level were measured for all teeth including mesiobuccal, buccal, disobuccal, mesiolingual, lingual, disolingual six sites of all patient. Gingival crevicular fluid samples were collected from the study sites. The prevalence and copy numbers (CN) of Aggregatibacter actinomycetemcomitans, Porphyromonas gingivalis, Prevotella intermedia, Treponema denticola, Tannerella forsythia and total bacteria in gingival crevicular fluid were quantified by real-time PCR. The hBD-2 concentration in gingival crevicular fluid was measured by ELISA. RESULTS: Both the prevalence and the CN of A. actinomycetemcomitans, P. gingivalis, T. denticola and T. forsythia were higher in patients with chronic periodontitis than in healthy controls and patients with chronic gingivitis; however, there was no significant difference in the prevalence of P. intermedia among the three study groups, and the highest CN was found in patients with chronic gingivitis, rather than in patients with chronic periodontitis. The loads of P. gingivalis, P. intermedia, T. denticola and total bacteria were positively related to probing depth, bleeding index and clinical attachment level. The concentration of hBD-2 in gingival crevicular fluid was higher in patients with chronic gingivitis and in patients with chronic periodontitis than in healthy controls. In addition, the hBD-2 concentration was positively related to the CN of P. gingivalis, T. forsythia and total bacteria, as well as to bleeding index and probing depth. CONCLUSION: The prevalence, composition and CN of periodontopathogens were closely related to the severity of periodontal disease, and the red complex was related to the severity of clinical symptoms of periodontal diseases. The concentration of hBD-2 in gingival crevicular fluid from periodontal disease sites was higher than that in gingival crevicular fluid from healthy sites, which suggests that hBD-2 expression might be up-regulated by periodontopathogens.

Immunohistological expression of human ß-defensin-1 and human ß-defensin-2 in exacerbation of acute and secondary chronic osteomyelitis of the mandible.

BACKGROUND: The majority of patients diagnosed with osteomyelitis of the jaw have severe complaints. Unfortunately, the pathogenesis still remains unclear. Human ß-defensins expressed in epithelial and bone tissues as a part of the innate immunity may be involved in disease development. In this study, we hypothesize that expression levels of human ß-defensin-1 and -2 in the acute and secondary chronic osteomyelitis may be altered in comparison with healthy bone and with bisphosphonate-associated necrosis as well as irradiation from a previous study. METHODS: Bone samples were collected during surgical debridement in a total of eight patients suffering from acute or secondary chronic osteomyelitis of the jaw. Expression levels of hBD-1 and -2 were quantified and related to non-stained cells. Ratios were compared by one-way ANOVA and multiple tests by Holm-Bonferroni. RESULTS: Multiple testing revealed no significant differences for expression levels of human ß-defensin-1 between all groups, whereas labeling index of human ß-defensin-2 was significantly different between specimens of bisphosphonate-associated osteonecrosis of the jaws and all other groups. No significant difference occurred between samples of floride osteomyelitis and healthy bone for expression of hBD-1 and -2. CONCLUSIONS: Although the affected patients showed all clinical signs of acute inflammation, expression levels in acute and secondary chronic osteomyelitis in the jaws did not reveal statistically significant differences compared with healthy bone samples. The weak immunological host response in terms of a putative genetically predisposition should be further discussed as pathogenesis factor for osteomyelitis in the future.

A study on ß-defensin-2 and histatin-5 as a diagnostic marker of early childhood caries progression.

BACKGROUND: Recently, a continuous growth of interest has been observed in antimicrobial peptides (AMPs) in the light of an alarming increase in resistance of bacteria and fungi against antibiotics. AMPs are used as biomarkers in diagnosis and monitoring of oral cavity pathologies. Therefore, the determination of specific protein profiles in children diagnosed with early childhood caries (ECC) might be a basis for effective screening tests and specialized examinations which may enable progression of disease. METHODS: The objective of the studies was to determine the role of histatin-5 and ß-defensing-2 as a diagnostic marker of early childhood caries progression. In this work, results of concentration determination of two salivary proteins (histatin-5 and ß-defensin-2) were presented. In addition, bacterial profiles from dental plaque in various stages of ECC and control were marked. The assessment of alteration in the concentration of these two proteins in a study group of children with various stages of ECC and a control group consisting of children with no symptoms was performed by enzyme-linked immunosorbent assays. RESULTS: The statistical analysis showed a significant increase in the concentration of histatin-5 and ß-defensin-2 in the study group compared to the control group and correlated with the progression of the disease. CONCLUSIONS: The confirmation of concentration changes in these proteins during the progression of dental caries may discover valuable disease progression biomarkers.

Airway ß-Defensin-1 Protein Is Elevated in COPD and Severe Asthma.

BACKGROUND: Innate immune antimicrobial peptides, including ß-defensin-1, promote the chemotaxis and activation of several immune cells. The role of ß-defensin-1 in asthma and chronic obstructive pulmonary disease (COPD) remains unclear. METHODS: Induced sputum was collected from healthy controls and individuals with asthma or COPD. ß-defensin-1 protein in sputum supernatant was quantified by ELISA. Biomarker potential was examined using receiver operating characteristic curves. ß-defensin-1 release from primary bronchial epithelial cells (pBECs) was investigated in culture with and without cigarette smoke extract (CSE). RESULTS: Airway ß-defensin-1 protein was elevated in COPD participants compared to asthma participants and healthy controls. Inflammatory phenotype had no effect on ß-defensin-1 levels in asthma or COPD. ß-defensin-1 protein was significantly higher in severe asthma compared to controlled and uncontrolled asthma. ß-defensin-1 protein could predict the presence of COPD from both healthy controls and asthma patients. Exposure of pBECs to CSE decreased ß-defensin-1 production in healthy controls; however in pBECs from COPD participants the level of ß-defensin-1 remanied unchanged. CONCLUSIONS: Elevated ß-defensin-1 protein is a feature of COPD and severe asthma regardless of inflammatory phenotype. ß-defensin-1 production is dysregulated in the epithelium of patients with COPD and may be an effective biomarker and potential therapeutic target.

Characterization of ß-defensin 42 expressed in principal cells at the initial segment of the rat epididymis.

ß-defensins, preferentially expressed in male reproductive tracts, particularly in the testes and epididymis with region-specific patterns, play an important role in both innate immunity and sperm fertility. Expressed in the caput region of epididymis, ß-defensins have been known to contribute to innate immunity, sperm motility initiation, and maintenance. However, ß-defensins of the initial region remain to be uncharacterized. In this study, rat ß-defensin 42 (Defb42) was revealed to be exclusively located in the principal cells at the initial segment of the rat epididymis and its sperm's acrosome. Furthermore, the expression of Defb42 was dependent on luminal testicular factors and developmental phases. The recombinant Defb42 was predominantly antimicrobial not against Candida albicans, but against Escherichia coli and Staphylococcus aureus. Based on these findings, Defb42 was suggested to play a dual role in sperm fertility and host defense in rat epididymis.

A competitive chemiluminescence enzyme immunoassay method for ß-defensin-2 detection in transgenic mice.

A competitive chemiluminescence enzyme immunoassay (CLEIA) method for porcine ß-defensin-2 (pBD-2) detection in transgenic mice was established. Several factors that affect detection, including luminol, p-iodophenol and hydrogen peroxide concentrations, as well as pH, were studied and optimized. The linear range of the proposed method for pBD-2 detection under optimal conditions was 0.05-80 ng/mL with a correlation coefficient of 0.9960. Eleven detections of a 30 ng/mL pBD-2 standard sample were performed. Reproducible results were obtained with a relative standard deviation of 3.94%. The limit of detection of the method for pBD-2 was 3.5 pg/mL (3σ). The proposed method was applied to determine pBD-2 expression levels in the tissues of pBD-2 transgenic mice, and compared with LC-MS/MS and quantitative real-time reverse-transcriptase polymerase chain reaction. This suggests that the CLEIA can be used as a valuable method to detect and quantify pBD-2.

Identification of sociodemographic and clinical factors associated with the levels of human ß-defensin-1 and human ß-defensin-2 in the human milk of Han Chinese.

Human milk provides infants with various immune molecules. The objective of the present study was to measure human ß-defensin-1 (hBD-1) and human ß-defensin-2 (hBD-2) levels in the colostrum and mature milk of healthy Han Chinese, to identify factors regulating milk hBD-1 and hBD-2 expression and to explore the potential protective effect of milk hBD-1 and hBD-2 on infants. A total of 100 mothers and their babies were recruited into the study. Sociodemographic characteristics and other factors were obtained by a questionnaire. Babies were followed up for a period of 6 months. Colostrum samples (n 100) and mature milk samples (n 82) were collected by hand expression. The hBD-1 and hBD-2 concentrations were measured by ELISA. The hBD-1 and hBD-2 levels differed in the colostrum and mature milk. In the colostrum, the concentration ranges of hBD-1 and hBD-2 were 1·04-12·81 µg/ml and 0·31-19·12 ng/ml, respectively. In mature milk, the hBD-1 and hBD-2 levels were 1·03-31·76 ng/ml and 52·65-182·29 pg/ml, respectively. Several independent factors influence their production. The multivariable analysis showed a strong association between pre-pregnancy BMI and hBD-1 levels in the colostrum (P=0·001), mode of delivery was significantly associated with hBD-2 levels in the colostrum (P=0·006) and gestational age was significantly associated with hBD-1 levels in mature milk (P= 0·010). During the first 6 months of life, the incidence rate of upper respiratory infection was found to be less in the high-colostrum hBD-1 group than in the low-colostrum hBD-1 group (χ²=4·995, P=0·025). The present study suggested that the abundance of hBD-1 in the colostrum may have a protective function against upper respiratory infection for infants younger than 6 months.