Lactobacillus helveticus Prevents Periodontitis Induced by Aggregatibacter actinomycetemcomitans in Rats by Regulating ß-Defensins.

Objective: To study the preventive effect of Lactobacillus helveticus (L. helveticus) on periodontitis induced by Aggregatibacter actinomycetemcomitans (A. actinomycetemcomitans) in rats. Methods: Eighteen 8-week-old female rats were randomly divided into three groups: Sham group, Trehalose group, and L. helveticus SBT2171 (LH2171) group. We measured the distance of the cementoenamel junction-alveolar bone crest (CEJ-ABC) to evaluate alveolar bone resorption. Hematoxylin-eosin staining was used to observe the histopathological changes of rat hemimaxillary tissues. We detected the expression of ß-defensins, tumor necrosis factor-α (TNF-α), interleukin- (IL-) 1ß, and IL-6 and the number of A. actinomycetemcomitans in rat gingival tissues by quantitative reverse transcriptase polymerase chain reaction. The levels of IL-1ß, IL-6, and TNF-α in rat gingival tissues were also measured by enzyme-linked immunosorbent assay. Results: Compared with the Trehalose group, the distance of CEJ-ABC was prominently reduced and alveolar bone resorption was notably improved in the LH2171 group. And the infiltration of inflammatory cells in the hemimaxillary tissue decreased obviously, periodontal fibers were arranged neatly, connective tissue small blood vessels proliferated, and the number of A. actinomycetemcomitans reduced significantly in the LH2171 group. In addition, the mRNA expression and release of inflammatory factors in the gingival tissues in the LH2171 group were notably lower than those in the Trehalose group. On the 21st and 36th day, the expression of ß-defensins in the gingival tissue of the LH2171 group increased significantly. Conclusion: L. helveticus improves alveolar bone resorption and increases the expression of ß-defensins thereby inhibiting the number of A. actinomycetemcomitans and thus prevents periodontitis.

Human ß-defensin 1 Functions as a Tumor Suppressor via ER Stress-triggered JNK pathway in Hepatocellular Carcinoma.

PURPOSE: Human ß-defensin 1 (DEFB1) belongs to defensins family that contribute to innate immune responses and was recently found to downregulate a variety of cancers, including renal, prostatic, and oral squamous cell carcinoma, and therefore is considered as a potential tumor suppressor. However, the role of DEFB1 in hepatocellular carcinoma (HCC) still needs to be elucidated. METHODS: Quantitative PCR and Western blot were used to measure the expression levels of interested proteins. CCK-8 and colony formation assays were performed to determine the ability of cell proliferation. Tumor formation experiments in nude mice were used to examine the tumor growth. RESULTS: The expression level of DEFB1 was dramatically downregulated in human HCC. Quantitative PCR and Western blot results also showed a pronounced decrease of DEFB1 expression in the liver cancer cell lines. Rescuing the expression of DEFB1 in Huh7 cells effectively suppressed cell proliferation and reduced the colony forming ability, probably by inducing cell apoptosis and cell cycle arrest. Moreover, tumor formation experiments in nude mice also showed inhibition of tumor growth by DEFB1 expression in vivo. Furthermore, induction of DEFB1 expression induced degraded protein increase and endoplasmic reticulum (ER) stress, which subsequently activated JNK pathway. Pharmacologic inhibition of ER stress by 4-phenylbutyrate, a compound to alleviate ER stress, effectively eliminated DEFB1-induction inhibition of cell proliferation and migration. CONCLUSION: DEFB1 functions as a tumor suppressor in HCC through activating ER stress and JNK pathway, which may provide a potential strategy for HCC treatment.

Functional Insights From the Evolutionary Diversification of Big Defensins.

Big defensins are antimicrobial polypeptides believed to be the ancestors of ß-defensins, the most evolutionary conserved family of host defense peptides (HDPs) in vertebrates. Nevertheless, big defensins underwent several independent gene loss events during animal evolution, being only retained in a limited number of phylogenetically distant invertebrates. Here, we explore the evolutionary history of this fascinating HDP family and investigate its patchy distribution in extant metazoans. We highlight the presence of big defensins in various classes of lophotrochozoans, as well as in a few arthropods and basal chordates (amphioxus), mostly adapted to life in marine environments. Bivalve mollusks often display an expanded repertoire of big defensin sequences, which appear to be the product of independent lineage-specific gene tandem duplications, followed by a rapid molecular diversification of newly acquired gene copies. This ongoing evolutionary process could underpin the simultaneous presence of canonical big defensins and non-canonical (ß-defensin-like) sequences in some species. The big defensin genes of mussels and oysters, two species target of in-depth studies, are subjected to gene presence/absence variation (PAV), i.e., they can be present or absent in the genomes of different individuals. Moreover, big defensins follow different patterns of gene expression within a given species and respond differently to microbial challenges, suggesting functional divergence. Consistently, current structural data show that big defensin sequence diversity affects the 3D structure and biophysical properties of these polypeptides. We discuss here the role of the N-terminal hydrophobic domain, lost during evolution toward ß-defensins, in the big defensin stability to high salt concentrations and its mechanism of action. Finally, we discuss the potential of big defensins as markers for animal health and for the nature-based design of novel therapeutics active at high salt concentrations.

Human ß-Defensin-2 Improves Hyperoxia-Induced Lung Structural and Functional Injury in Neonatal Rats.

BACKGROUND Bronchopulmonary dysplasia (BPD) is a major complication of extreme prematurity, characterized by alveolar simplification and pulmonary malfunction. Hyperoxia-induced lung injury in neonatal rats has been used as a model of BPD, as indicated by lung architectural change and alveolar simplification that resembles clinical feature of BPD. ß-defensin-2 (BD2) plays an important role in lung diseases by inhibiting inflammation response. However, little is known about its role in BPD. The aim of this study was to determine the effect of human BD2 (hBD2) gene on hyperoxia-induced animal model of BPD. MATERIAL AND METHODS The neonatal rats were exposed to 90% oxygen (O2) continuously for 14 days to mimic the BPD-like lung injury. These rats were then randomly assigned to the following four groups: in room air (air), in 90% O2, in 90% O2 with null adenovirus vector infection (O2+Ad), and in 90% O2 with gene therapy through adenovirus transfected hBD2 (O2+Ad-hBD2). Morphology of lungs, pulmonary function and expression of inflammatory cytokines on P7, P10, P14, and P21 were documented and compared across the 4 groups. RESULTS The overexpression of hBD2 mediated by the adenovirus vector was successfully constructed. hBD2 gene therapy increased hBD2 mRNA expression, increased radial alveolar count (RAC), lung volume and compliance, decreased mean linear intercept (MLI), tissue damping, and elastance. Furthermore, pro-inflammatory cytokines IL-1ß, IL-6, and TNF-alpha were inhibited and anti-inflammatory cytokines IL-10 was increased in the lungs of rats in O2+Ad-hBD2 group. CONCLUSIONS In hyperoxia-induced rat models of BPD, hBD2 promotes alveolarization and improves pulmonary function. The mechanism may contribute in alleviating inflammation response and inhibiting pro-inflammatory factors including IL-1ß, IL-6, and TNF-alpha.

Bladder-Drained Pancreas Transplantation: Urothelial Innate Defenses and Urinary Track Infection Susceptibility.

BACKGROUND: Pancreas transplantation restores insulin secretion in type 1 diabetes mellitus. The graft also produces exocrine secretions that can be drained enterically (enteric drainage [ED]) or via the bladder (bladder drainage [BD]). We suggest that in BD transplants, such secretions destroy bladder innate immunity, specifically host defense peptides/proteins (HDPs), which increases patient susceptibility to recurrent urinary tract infections (rUTIs). MATERIALS AND METHODS: BD and ED patient records were reviewed retrospectively for UTIs. Urine samples from ED and BD transplant recipients were analyzed for pH, the HDPs ß-defensin 2 (HBD2) and lipocalin-2, and amylase concentrations. In vitro, bacterial growth curves and antimicrobial assays were used to evaluate the effects of pH, HBD2, and HBD2 + pancreatic digestive enzymes (pancreatin) on uropathogenic Escherichia coli (UPEC) survival and growth. RESULTS: Urinalysis revealed a significant difference in pH between the BD and ED cohorts (7.2 ± 0.8 versus 6.7 ± 0.8; P = 0.012). Urinary HDPs were measured and BD, but not ED, lipocalin-2 concentrations were significantly decreased compared with those of diabetics awaiting transplant (P < 0.05). In vitro, an alkaline environment, pH 8.0, concomitant with the urine of the patient who underwent BD transplantation, significantly reduced UPEC growth (P < 0.05); addition of pancreatin to the growth medium was associated with a significant increase (P < 0.001) in growth rate. Antimicrobial data suggested significant UPEC killing in the presence of HBD2 (P < 0.01), but not in the presence of HBD2 + pancreatin (>12,500 amylase units). CONCLUSIONS: These in vivo and in vitro data suggest that BD pancreatic exocrine secretions inactivate the bladder innate defenses, which facilitate UPEC growth and underpins the increased susceptibility of patients who underwent BD pancreas transplantation to rUTIs.

[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.

CRISPR/Cas9-mediated genome editing reveals the synergistic effects of ß-defensin family members on sperm maturation in rat epididymis.

The epididymis is a male reproductive organ involved in posttesticular sperm maturation and storage, but the mechanism underlying sperm maturation remains unclear. ß-Defensins (Defbs) belong to a family of small, cysteine-rich, cationic peptides that are antimicrobial and modulate the immune response. A large number of Defb genes are expressed abundantly in the male reproductive tract, especially in the epididymis. We and other groups have shown the involvement of several Defb genes in regulation of sperm function. In this study, we found that Defb23, Defb26, and Defb42 were highly expressed in specific regions of the epididymis. Rats with CRISPR/Cas9-mediated single-gene disruption of Defb23, Defb26, or Defb42 had no obvious fertility phenotypes. Those with the deletion of Defb23/ 26 or Defb23/ 26/ 42 became subfertile, and sperm isolated from the epididymal cauda of multiple-mutant rats were demonstrated decreased motility. Meanwhile, the sperm showed precocious capacitation and increased spontaneous acrosome reaction. Consistent with premature capacitation and acrosome reaction, sperm from multiple-gene-knockout rats had significantly increased intracellular calcium. These results suggest that Defb family members affect sperm maturation by a synergistic pattern in the epididymis.-Zhang, C., Zhou, Y., Xie, S., Yin, Q., Tang, C., Ni, Z., Fei, J., Zhang, Y. CRISPR/Cas9-mediated genome editing reveals the synergistic effects of ß-defensin family members on sperm maturation in rat epididymis.

Control mechanisms for producing antimicrobial factors in ruminant mammary gland.

Mastitis, a symptom of inflammation in mammary tissue by infection with various kinds of bacteria, causes huge economic losses in the milk industry. One of the popular methods for treatment of mastitis is antibiotics, although this prohibits milk shipping and sometimes causes resistant microbes. Therefore, a new strategy to treat mastitis without antibiotics is eagerly required around the world. Antimicrobial factors belong to innate immunity and can start their function extremely early after bacterial stimulation. These factors have antimicrobial activity for a broad spectrum of bacteria. Elucidation of causal mechanisms and functions of antimicrobial factors in the mammary gland is thought to result in suitable methods for prevention and treatment of mastitis. Therefore, this review introduces traits of some antimicrobial factors and the mechanisms for expressing, producing and secreting them in the mammary gland. For antimicrobial factors, lingual antimicrobial peptide (LAP), S100A7, cathelicidin and lactoferrin are controlled in different sites and different time courses, suggesting that antimicrobial factors play different roles for local defense against bacterial infection in the mammary gland. These findings will contribute to the development of prevention and treatment methods for mastitis.

In Vitro and In Vivo Anticancer Activity of Human ß-Defensin-3 and Its Mouse Homolog.

BACKGROUND/AIM: Defensins comprise a family of mammalian cationic antimicrobial peptides. We investigated the anticancer effects of human ß-defensin-3 (hBD3) and its mouse homolog, Defb14, on lung cancer cells. MATERIALS AND METHODS: We stained lung cancer cells cultured after treatment with the defensin peptide using propidium iodide and Hoechst 33342. In vivo, Defb14 peptide or vehicle was continuously infused near subcutaneous Lewis lung carcinoma cell tumor in mice. After 9-day infusion, the weights of excised tumors were determined. RESULTS: A 10-min treatment with hBD3 (70 µg/ml) induced propidium iodide uptake in lung cancer cells. The anticancer activity of hBD3 was significantly more potent than the activity of other defensin isoforms. Continuous infusion of Defb14 peptide showed significant tumor-growth suppression in Lewis lung carcinoma cells in mice. CONCLUSION: Our study demonstrated the suppression of tumor growth by Defb14 peptide in an animal model.

Structure-function analysis of Avian ß-defensin-6 and ß-defensin-12: role of charge and disulfide bridges.

BACKGROUND: Avian beta-defensins (AvBD) are small, cationic, antimicrobial peptides. The potential application of AvBDs as alternatives to antibiotics has been the subject of interest. However, the mechanisms of action remain to be fully understood. The present study characterized the structure-function relationship of AvBD-6 and AvBD-12, two peptides with different net positive charges, similar hydrophobicity and distinct tissue expression profiles. RESULTS: AvBD-6 was more potent than AvBD-12 against E. coli, S. Typhimurium, and S. aureus as well as clinical isolates of extended spectrum beta lactamase (ESBL)-positive E. coli and K. pneumoniae. AvBD-6 was more effective than AvBD-12 in neutralizing LPS and interacting with bacterial genomic DNA. Increasing bacterial concentration from 10(5) CFU/ml to 10(9) CFU/ml abolished AvBDs' antimicrobial activity. Increasing NaCl concentration significantly inhibited AvBDs' antimicrobial activity, but not the LPS-neutralizing function. Both AvBDs were mildly chemotactic for chicken macrophages and strongly chemotactic for CHO-K1 cells expressing chicken chemokine receptor 2 (CCR2). AvBD-12 at higher concentrations also induced chemotactic migration of murine immature dendritic cells (DCs). Disruption of disulfide bridges abolished AvBDs' chemotactic activity. Neither AvBDs was toxic to CHO-K1, macrophages, or DCs. CONCLUSIONS: AvBDs are potent antimicrobial peptides under low-salt conditions, effective LPS-neutralizing agents, and broad-spectrum chemoattractant peptides. Their antimicrobial activity is positively correlated with the peptides' net positive charges, inversely correlated with NaCl concentration and bacterial concentration, and minimally dependent on intramolecular disulfide bridges. In contrast, their chemotactic property requires the presence of intramolecular disulfide bridges. Data from the present study provide a theoretical basis for the design of AvBD-based therapeutic and immunomodulatory agents.

Targeted inactivation of the mouse epididymal beta-defensin 41 alters sperm flagellar beat pattern and zona pellucida binding.

During epididymal maturation, sperm acquire the ability to swim progressively by interacting with proteins secreted by the epididymal epithelium. Beta-defensin proteins, expressed in the epididymis, continue to regulate sperm motility during capacitation and hyperactivation in the female reproductive tract. We characterized the mouse beta-defensin 41 (DEFB41), by generating a mouse model with iCre recombinase inserted into the first exon of the gene. The homozygous Defb41(iCre/iCre) knock-in mice lacked Defb41 expression and displayed iCre recombinase activity in the principal cells of the proximal epididymis. Heterozygous Defb41(iCre/+) mice can be used to generate epididymis specific conditional knock-out mouse models. Homozygous Defb41(iCre/iCre) sperm displayed a defect in sperm motility with the flagella primarily bending in the pro-hook conformation while capacitated wild-type sperm more often displayed the anti-hook conformation. This led to a reduced straight line motility of Defb41(iCre/iCre) sperm and weaker binding to the oocyte. Thus, DEFB41 is required for proper sperm maturation.

ß-defensins and the epididymis: contrasting influences of prenatal, postnatal, and adult scenarios.

ß-defensins are components of host defense, with antimicrobial and pleiotropic immuno-modulatory properties. Research over the last 15 years has demonstrated abundant expression of a variety of ß-defensins in the postnatal epididymis of different species. A gradient of region- and cell-specific expression of these proteins is observed in the epithelium of the postnatal epididymis. Their secretion into the luminal fluid and binding to spermatozoa as they travel along the epididymis has suggested their involvement in reproduction-specific tasks. Therefore, continuous attention has been given to various ß-defensins for their role in sperm function and fertility. Although ß-defensins are largely dependent on androgens, the underlying mechanisms regulating their expression and function in the epididymis are not well understood. Recent investigation has pointed out to a new and interesting scenario where ß-defensins emerge with a different expression pattern in the Wolffian duct, the embryonic precursor of the epididymis, as opposed to the adult epididymis, thereby redefining the concept concerning the multifunctional roles of ß-defensins in the developing epididymis. In this review, we summarize some current views of ß-defensins in the epididymis highlighting our most recent data and speculations on their role in the developing epididymis during the prenatal-to-postnatal transition, bringing attention to the many unanswered questions in this research area that may contribute to a better understanding of epididymal biology and male fertility.

Expression of Beta-Defensin 131 Promotes an Innate Immune Response in Human Prostate Epithelial Cells.

Previously, using the Illumina HumanHT-12 microarray we found that ß-defensin 131 (DEFB131), an antimicrobial peptide, is upregulated in the human prostate epithelial cell line RWPE-1 upon stimulation with lipoteichoic acid (LTA; a gram-positive bacterial component), than that in the untreated RWPE-1 cells. In the current study, we aimed to investigate the role of DEFB131 in RWPE-1 cells during bacterial infection. We examined the intracellular signaling pathways and nuclear responses in RWPE-1 cells that contribute to DEFB131 gene induction upon stimulation with LTA. Chromatin immunoprecipitation was performed to determine whether NF-κB directly binds to the DEFB131 promoter after LTA stimulation in RWPE-1 cells. We found that DEFB131 expression was induced by LTA stimulation through TLR2 and p38MAPK/NF-κB activation, which was evident in the phosphorylation of both p38MAPK and IκBα. We also found that SB203580 and Bay11-7082, inhibitors of p38MAPK and NF-κB, respectively, suppressed LTA-induced DEFB131 expression. The chromatin immunoprecipitation assay showed that NF-κB directly binds to the DEFB131 promoter, suggesting that NF-κB is a direct regulator, and is necessary for LTA-induced DEFB131 expression in RWPE-1 cells. Interestingly, with DEFB131 overexpression in RWPE-1 cells, the accumulation of mRNA and protein secretion of cytokines (IL-1α, IL-1ß, IL-6, and IL-12α) and chemokines (CCL20, CCL22, and CXCL8) were significantly enhanced. In addition, DEFB131-transfected RWPE-1 cells markedly induced chemotactic activity in THP-1 monocytes. We concluded that DEFB131 induces cytokine and chemokine upregulation through the TLR2/NF-κB signaling pathway in RWPE-1 cells during bacterial infection and promotes an innate immune response.

Why do we study animal toxins?

Venom (toxins) is an important trait evolved along the evolutionary tree of animals. Our knowledges on venoms, such as their origins and loss, the biological relevance and the coevolutionary patterns with other organisms are greatly helpful in understanding many fundamental biological questions, i.e., the environmental adaptation and survival competition, the evolution shaped development and balance of venoms, and the sophisticated correlations among venom, immunity, body power, intelligence, their genetic basis, inherent association, as well as the cost-benefit and trade-offs of biological economy. Lethal animal envenomation can be found worldwide. However, from foe to friend, toxin studies have led lots of important discoveries and exciting avenues in deciphering and fighting human diseases, including the works awarded the Nobel Prize and lots of key clinic therapeutics. According to our survey, so far, only less than 0.1% of the toxins of the venomous animals in China have been explored. We emphasize on the similarities shared by venom and immune systems, as well as the studies of toxin knowledge-based physiological toxin-like proteins/peptides (TLPs). We propose the natural pairing hypothesis. Evolution links toxins with humans. Our mission is to find out the right natural pairings and interactions of our body elements with toxins, and with endogenous toxin-like molecules. Although, in nature, toxins may endanger human lives, but from a philosophical point of view, knowing them well is an effective way to better understand ourselves. So, this is why we study toxins.

Host defense (Antimicrobial) peptide, human ß-defensin-3, improves the function of the epithelial tight-junction barrier in human keratinocytes.

Human ß-defensins (hBDs) are host defense peptides that not only exhibit microbicidal properties but also stimulate various cellular activities, including keratinocyte proliferation, migration, and wound healing. hBDs are overexpressed in the skin in cases of psoriasis but are downregulated in atopic dermatitis skin, although both diseases are associated with stratum corneum barrier defects. Because the tight-junction (TJ) barrier is also dysfunctional in both atopic dermatitis and psoriasis patients, we hypothesized that hBDs may regulate the TJ barrier function in keratinocytes. We observed that, among the hBDs tested, only hBD-3 increased the expression of several claudins and their localization along the cell-cell borders. In addition, hBD-3 elevated the transepithelial electrical resistance and reduced the paracellular permeability of keratinocyte layers, and this effect was reversed by the claudin inhibitor ochratoxin A, CCR6 antibody, and CCR6 small interfering RNA. Moreover, hBD-3 enhanced the activation of Rac1, atypical protein kinase C, glycogen synthase kinase-3, and phosphatidylinositol 3 kinase, which are required for the hBD-3-mediated regulation of the TJ barrier function, as evidenced by the effects of their respective inhibitors. Collectively, our findings provide evidence regarding the contribution of host defense peptides to the innate immunity of skin by regulating TJ barrier function, in addition to their antimicrobial and other immunomodulatory activities.

Human beta-defensin-1 suppresses tumor migration and invasion and is an independent predictor for survival of oral squamous cell carcinoma patients.

BACKGROUND: Human beta-defensin-1 (hBD-1) has recently been considered as a candidate tumor suppressor in renal and prostate cancer. The aim of this study was to investigate the role of hBD-1 in the progression of oral squamous cell carcinoma (OSCC) and its potential as diagnostic/prognostic biomarker and therapeutic target for OSCC. METHODS: HBD-1 expression in tissues at different stages of oral carcinogenesis, as well as OSCC cell lines was examined. HBD-1 was overexpressed in HSC-3, UM1, SCC-9 and SCC-25 cells and subjected to cell growth, apoptosis, migration and invasion assays. Tissue microarray constructed with tissues from 175 patients was used to examine clinicopathological significance of hBD-1 expression in OSCC. RESULTS: HBD-1 expression decreased from oral precancerous lesions to OSCC and was lower in OSCC with lymph node metastasis than those without metastasis. In vitro, the expression of hBD-1 was related to the invasive potential of OSCC cell lines. Induction of exogenous expression of hBD-1 inhibited migration and invasion of OSCC cells, probably by regulation of RhoA, RhoC and MMP-2; but had no significant effect on proliferation or apoptosis. In a cohort of patients with primary OSCC, cases with no expression of hBD-1 had more chance to be involved in lymph node metastasis. Eventually, the positive expression of hBD-1 was associated with longer survival of patients with OSCC, and multivariate analysis and ROC curve analysis confirmed hBD-1 positivity to be an independent prognostic factor of OSCC, especially OSCC at early stage. CONCLUSIONS: Overall, these data indicated that hBD-1 suppressed tumor migration and invasion of OSCC and was likely to be a prognostic biomarker and a potential target for treatment of OSCC.

Suppression of antimicrobial peptide expression by ureaplasma species.

Ureaplasma species commonly colonize the adult urogenital tract and are implicated in invasive diseases of adults and neonates. Factors that permit the organisms to cause chronic colonization or infection are poorly understood. We sought to investigate whether host innate immune responses, specifically, antimicrobial peptides (AMPs), are involved in determining the outcome of Ureaplasma infections. THP-1 cells, a human monocytoid tumor line, were cocultured with Ureaplasma parvum and U. urealyticum. Gene expression levels of a variety of host defense genes were quantified by real-time PCR. In vitro antimicrobial activities of synthetic AMPs against Ureaplasma spp. were determined using a flow cytometry-based assay. Chromosomal histone modifications in host defense gene promoters were tested by chromatin immunoprecipitation (ChIP). DNA methylation status in the AMP promoter regions was also investigated. After stimulation with U. parvum and U. urealyticum, the expression of cell defense genes, including the AMP genes (DEFB1, DEFA5, DEFA6, and CAMP), was significantly downregulated compared to that of TNFA and IL-8, which were upregulated. In vitro flow cytometry-based antimicrobial assay revealed that synthetic peptides LL-37, hBD-3, and hBD-1 had activity against Ureaplasma spp. Downregulation of the AMP genes was associated with chromatin modification alterations, including the significantly decreased histone H3K9 acetylation with U. parvum infection. No DNA methylation status changes were detected upon Ureaplasma infection. In conclusion, AMPs have in vitro activity against Ureaplasma spp., and suppression of AMP expression might be important for the organisms to avoid this aspect of the host innate immune response and to establish chronic infection and colonization.

Role of urinary cathelicidin LL-37 and human ß-defensin 1 in uncomplicated Escherichia coli urinary tract infections.

Cathelicidin (LL-37) and human ß-defensin 1 (hBD-1) are important components of the innate defense in the urinary tract. The aim of this study was to characterize whether these peptides are important for developing uncomplicated Escherichia coli urinary tract infections (UTIs). This was investigated by comparing urinary peptide levels of UTI patients during and after infection to those of controls, as well as characterizing the fecal flora of participants with respect to susceptibility to LL-37 and in vivo virulence. Forty-seven UTI patients and 50 controls who had never had a UTI were included. Participants were otherwise healthy, premenopausal, adult women. LL-37 MIC levels were compared for fecal E. coli clones from patients and controls and were also compared based on phylotypes (A, B1, B2, and D). In vivo virulence was investigated in the murine UTI model by use of selected fecal isolates from patients and controls. On average, UTI patients had significantly more LL-37 in urine during infection than postinfection, and patient LL-37 levels postinfection were significantly lower than those of controls. hBD-1 showed similar urine levels for UTI patients and controls. Fecal E. coli isolates from controls had higher LL-37 susceptibility than fecal and UTI E. coli isolates from UTI patients. In vivo studies showed a high level of virulence of fecal E. coli isolates from both patients and controls and showed no difference in virulence correlated with the LL-37 MIC level. The results indicate that the concentration of LL-37 in the urinary tract and low susceptibility to LL-37 may increase the likelihood of UTI in a complex interplay between host and pathogen attributes.