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Host defence peptides (HDPs) are considered to be excellent candidates for the development of novel therapeutic agents. Recently, it was demonstrated that the peptide C1-15, an N-terminal segment of chicken HDP cathelicidin-2, exhibits potent antibacterial activity while lacking cytotoxicity towards eukaryotic cells. In the present study, we report that C1-15 is active against bacteria such as Bacillus anthracis and Yersinia pestis that may potentially be used by bioterrorists. Substitution of single and multiple phenylalanine (Phe) residues to tryptophan (Trp) in C1-15 resulted in variants with improved antibacterial activity against B. anthracis and Y. pestis as well as decreased salt sensitivity. In addition, these peptides exhibited enhanced neutralisation of lipopolysaccharide (LPS)-induced release of pro-inflammatory cytokines in human peripheral blood mononuclear cells (PBMCs). The antibacterial and LPS-neutralising activities of these C1-15-derived peptides are exerted at concentrations far below the concentrations that are toxic to human PBMCs. Taken together, we show that Phe → Trp substitutions in C1-15 variants enhances the antibacterial and LPS-neutralising activities against pathogenic bacteria, including those that may potentially be used as biological warfare agents.Keywords: Host defence peptide; Cathelicidin; Antibacterial activity; Cytotoxicity; Immunomodulatory activity; Biological warfare agents

Antimicrobial peptides are multifunctional in innate immunity and wound repair of multicellular organisms. We were the first to discover that histatins, a family of salivary antimicrobial peptides, enhance epithelial cell migration, suggesting a role in oral wound healing. It is unknown whether histatins display innate-immunity activities, similar to other antimicrobial peptides such as LL-37. Therefore, we compared the effect of Histatin-2 and LL-37 on several activities within the context of wound healing and innate immunity. We found that Histatin-2 enhances fibroblast migration, but only weakly induces proliferation. LL-37 enhances both fibroblast migration and proliferation, but only at a narrow concentration optimum (approximately 1 μm). At higher concentrations LL-37 causes cell death, whereas Histatin-2 is not cytotoxic. Both peptides do not alter fibroblast-to-myofibroblast differentiation. Histatin-2 does not alter interleukin-8 (IL-8) expression and lipopolysaccharide (LPS)-elevated cytokine and chemokine expression. In contrast, LL-37 induces IL-8 expression, but dampens the LPS-induced immune response. Neither Histatin-2 nor LL-37 affects human-neutrophil migration. Histatins are, unlike other antimicrobial peptides, not cytotoxic or proinflammatory. It seems that they are important for the initial stage of wound healing in which fast wound coverage is important for healing without infection, inflammation, or fibrosis development. Interestingly, these characteristics are more typical for the mouth than for skin.

Salivary agglutinin plays a vital biological role modulating the protective effect in the oral cavity by interacting with a broad range of oral pathogens. Here, we describe the first characterization of the O-linked oligosaccharides of salivary agglutinin identified by negative ion liquid chromatography-mass spectrometry. The dominating structures were neutral or monosialylated core 1 (Gal beta 1-3GalNAc alpha 1-Ser/Thr) and core 2 (Gal beta 1-3(GlcNAc beta 1-6)GalNAc alpha 1-Ser/Thr) structures extended by fucosylated oligo-N-acetyllactosamine units. Oligosaccharides detected as [M-H](-) or [M-2H](2-) ions ranged from the disaccharide Gal beta 1-3GalNAcol up to structures of almost 4000 Da, corresponding to core 1/2 structures with five N-acetyllactosamine units and 11 fucoses. Fucose was found either as terminal or internal blood group H structures in type 1 (Gal beta 1-3GlcNAc beta 1-R), type 2 (Gal beta 1-4GlcNAc beta 1-R) and type 3 (Gal beta 1-3GalNAc alpha 1-Ser/Thr) units, where the chains also could be fucosylated on GlcNAc yielding repeated Lewis a/b or Lewis x/y structures. Sialylation was located either at the non-reducing end of the N-acetyllactosamine chains as sialyl-Lewis x or as sialyl-T (NeuAc alpha 2-3Gal beta 1-3GalNAc alpha 1-Ser/Thr) type structures with or without further extension of the C-6 branch of GalNAc with neutral fucosylated N-acetyllactosamine chains. The data indicated that sialylation, fucosylation and type 1 N-acetyllactosamine termination are important regulatory elements for controlling the oligosaccharide chain length. Furthermore, it was shown that these regulatory oligosaccharide elements could be utilized by the pathogen Helicobacter pylori to colonize the oral cavity, reside in dental plaque and serve as a reservoir for reinfection after successful clearance of H. pylori gastric infection.

Increased prevalence of antibiotic-resistant bacteria has become a major threat to the health sector worldwide due to their virulence, limited therapeutic options and distribution in both hospital and community settings. Discovery and development of new agents to combat antibiotic-resistant bacteria is thus needed. This study therefore aimed to evaluate the ability of bovine lactoferrin (LF), peptides from two antimicrobial domains lactoferricin B (LFcin17-30) and lactoferrampin (LFampin265-284) and a chimeric construct (LFchimera) containing both peptides, as potential bactericidal agents against clinical isolates of antibiotic-resistant Staphylococcus aureus and Escherichia coli. Results in kinetics of growth show that LF chimera and peptides inhibited the growth of both bacterial species. By confocal microscopy and flow cytometry it was observed that LF and FITC-labeled peptides are able to interact with these bacteria and cause membrane permeabilization, as monitored by propidium iodide staining, these effects were decreased by preincubation with lipopolysaccharide in E. coli. By electron microscopy, a clear cellular damage was observed in bacteria after treatments with LFchimera and peptides, suggesting that interaction and membrane disruption are probably involved as a mechanism of action. In conclusion, results show that LFchimera, LF and peptides have potential as bactericidal agents in the antibiotic-resistant strains of S. aureus and E. coli and also the work strongly suggest that LFcin17-30 and LFampin265-284 acts synergistically with antibiotics against multidrug resistant EPEC and MRSA in vitro.

Entamoeba histolytica is a parasitic protozoan that produces amoebiasis, an intestinal disease characterized by ulcerative colitis and dysentery. In some cases, trophozoites can travel to the liver leading to hepatic abscesses and death. Recently, lactoferrin and lactoferricin B have been shown to be amoebicidal in axenic cultures. The aim of this work was to determine whether the lactoferrin-peptides lactoferricin amino acids 17-30, lactoferrampin amino acids 265-284, and lactoferrin chimera which is a fusion product of the two peptides, are capable of producing a microbicidal effect to trophozoites of E. histolytica. We evaluated the killing effect of these peptides in growth kinetics carried out in axenic culture medium to which different concentrations of peptides were added. At 50 muM of peptide concentration, lactoferricin and lactoferrampin had a moderate amoebicidal effect, since a 45-50% of trophozoites remained viable at 24 h culture. However, at 50 microM of the lactoferrin chimera 75% amoeba were killed whereas at 100 microM all cells died. These data indicate that of lactoferrin-peptides mainly the chimera have amoebicidal activity in a time- and concentration-dependent manner. The lactoferrin-peptides might be useful as therapeutic agents against amoebiasis and thereby diminish the use of metronidazole, which is extremely toxic for the host.

De bipolaire stoornis is een stemmingsstoornis die wordt gekenmerkt door het recidiverende optreden van manische, depressieve en gemengde episoden met daartussen kortere of langere, relatief symptoomvrije perioden. In Nederland is de prevalentie 1,9%. Bij de behandeling wordt meestal een combinatie van psycho-educatie, zelfmanagement en farmacotherapie toegepast. Zowel de bipolaire stoornis als de toegepaste medicatie heeft negatieve effecten op de mondgezondheid. Patiënten hebben onder andere een groter risico op cariës, xerostomie, smaakafwijkingen en bruxisme. Uitgebreide instructie van de patiënt in mondverzorging, ondersteund door frequente professionele mondzorg, is daarom essentieel. Gezien de mogelijke interacties tussen geneesmiddelen dient de mondzorgverlener ‘non-steroid anti-inflammatory drugs’ alleen in overleg met de behandelende psychiater voor te schrijven.

BackgroundHelicobacter pylori is a Gram-negative microorganism which is able to colonize the gastric mucosa and is associated with peptic ulcer, gastric carcinoma, and gastric mucosa-associated lymphoid tissue lymphoma. Several studies have detected this bacterium in the oral cavity, suggesting it as a potential reservoir. The aim of this study was to investigate the presence of H. pylori in the oral cavity of individuals with periodontal disease and gastric diseases.Methods115 individuals, with mean age 49.6 (±5.8) years, were divided in 4 groups: (A) with gastric diseases and periodontal disease; (B) with gastric diseases and no periodontal disease; (C) without gastric diseases and without periodontal disease, (D) without gastric diseases and with periodontal disease. Supra and subgingival plaque samples were collected from posterior teeth of the individuals with sterile paper points, and prepared for Polymerase Chain Reaction analysis. Fisher's exact test was used for detecting statistical differences between groups (p < 0.05).ResultsH. pylori was detected in supragingival plaque of 9/36 (25%) of group A, 1/31 (0.3%) of group B, 0 (0%) of group C and 3/36 (8.3%) of group D. No subgingival samples were positive for H. pylori. There was a statistically higher prevalence of H. pylori in groups A and D when compared to B and C (p < 0.05).ConclusionH. pylori was detected in the supragingival plaque, but not in the subgingival plaque, of individuals with periodontal disease and upper gastric diseases. There was an association between the supragingival colonization of H. pylori and oral hygiene parameters such as the presence of plaque and gingival bleeding.

Objective: Many students have paid employment while studying. In the Netherlands, the Individual Health Care Professions Act (IHCP Act) allows dental hygiene students to work under certain conditions in a dental practice. The aim of the study was to determine how many dental hygiene students have part-time job employment in dental practice and which professional tasks they carry out. We also asked the dental hygiene students their opinion of the IHCP Act.Methods: All the enrolled dental hygiene students (n = 341) at a School of Health in the Netherlands received a questionnaire by email.Results: The response was 52% (176 students). Of the responding students, 75% had paid employment in addition to their study. A proportion of the students (35%) worked in a dental practice. The median number of hours worked per week was eight. Study year, age and prior education were positively related to working part-time in dental practice. Activities frequently performed were giving oral hygiene instruction, fluoride applications, scaling and root planning, providing chair side assistance and giving local anaesthesia. Although the self-reported knowledge about the IHCP Act was high, almost half of the students expressed the need for more detailed legal information.Conclusions: Many dental hygiene students work in a dental practice, taking over a number of tasks usually performed by the dentist. More information in the dental hygiene curriculum about the requirements of the IHCP Act seems desirable.

Antimicrobial peptides (AMPs) are essential host defence molecules found in a wide variety of speciesand are promising antibacterial therapeutic candidates. Focusing on the human cathelicidin peptide LL-37, the aim of the present study was to explore the mechanisms of action and antimicrobial activitiesof a library of LL-37 fragments using Burkholderia thailandensis E264 as a model. The results revealedthat IG-19 was the shortest fragment within LL-37 that exhibited antibacterial activity. LL-31, missingsix residues at the C-terminus of LL-37, exhibited the strongest killing effect. Freeze-fracture electronmicroscopy of bacterial cells treated with either LL-37 or LL-31 revealed irregular bacterial surfaces withbleb projections, indicating that these peptides disrupted the integrity of the membrane. In addition,these peptides induced leakage of cell components, including nucleotides and even proteins. Altogether,the results obtained indicate the potential of using LL-31 as a new AMP to combat Burkholderia spp.

Burkholderia pseudomallei, the causative agent of melioidosis, is intrinsically resistant to many antibiotics, resulting in high mortality rates of 19% in Australia and even 50% in Thailand. Antimicrobial peptides (AMPs) possess potent broad-spectrum bactericidal activities and are regarded as promising therapeutic alternatives in the fight against resistant microorganisms. Moreover, these peptides may also affect inflammation, immune activation and wound healing. In this study, the in vitro activities of 10 AMPs, including histatin 5 and histatin variants, human cathelicidin peptide LL-37 and lactoferrin peptides, against 24 isolates of B. pseudomallei were investigated. The results showed that the antibacterial activities of the individual peptides depended on peptide dose and bacterial isolate. Among the 10 peptides tested, LL-37 exhibited the most effective killing activity. The smooth type A lipopolysaccharide (LPS) phenotype B. pseudomallei appeared to be more susceptible than those expressing the smooth type B LPS and the rough type LPS. Four isolates of B. pseudomallei shown to be resistant to ceftazidime and trimethoprim/sulfamethoxazole were also highly susceptible to LL-37. These data indicate that LL-37 possesses antimicrobial activity against all isolates independent of the LPS phenotype and is therefore a promising peptide to combat B. pseudomallei infections.

Burkholderia pseudomallei is a gram-negative bacillus that is the causative agent of melioidosis. We evaluated host-pathogen interaction at different levels using three separate B. pseudomallei mutants generated by insertional inactivation. One of these mutants is defective in the production of the polysaccharide side chains associated with lipopolysaccharide; one does not produce the capsular polysaccharide with the structure -3)-2-O-acetyl-6-deoxy-beta-d-manno-heptopyranose-(1-; and the third mutant does not produce flagellin. We compared the in vivo virulence in BALB/c mice, the in vitro fate of intracellular survival inside human polymorphonuclear cells (PMNs) and macrophages (Mphis) and the susceptibility to killing by 30% normal human serum, reactive nitrogen and oxygen intermediates and antimicrobial peptides with that of their wild-type counterpart. The lipopolysaccharide and capsule mutants demonstrated a marked reduction in virulence for BALB/c mice, but the flagellin mutant was only slightly less virulent than the parent strain. The results from the BALB/c mice experiments correlated with survival in Mphis. The lipopolysaccharide and capsule mutants were also more susceptible to killing by antimicrobial agents. All bacteria were equally susceptible to killing by PMNs. Altogether, the data suggest that lipopolysaccharide and capsule and, to a much lesser extent, flagella, are most likely associated with the virulence of this bacterium and highlight the importance of intracellular killing by PMNs and Mphis in disease pathogenesis.

The role of human saliva in oral wound-healing has never been fully elucidated. We previously demonstrated that parotid-salivary histatins enhance in vitro wound closure. The question remains whether other salivary-gland secretions enhance wound closure, and also the effects of histatins on primary and non-oral cells. Since the presence of histatins is not limited to parotid saliva, we expected to observe wound-closure activity of other salivary-gland secretions. However, here we show that non-parotid saliva does not stimulate wound closure, most probably due to the presence of mucins, since the addition of MUC5B to parotid saliva abolished its effect. Furthermore, we found that histatins stimulated wound closure of (primary) cells of both oral and non-oral origin. This suggests that the cellular receptor of histatins is widely expressed and not confined to cells derived from the oral cavity. These findings encourage the future therapeutic application of histatins in the treatment of all kinds of wounds.

The aim of this study was to detect the presence of Helicobacter pylori and its virulent cagA genes in the oral cavity of individuals with upper gastric diseases. Sixty-two individuals (42 ± 2.3 years) with dispepsy symptoms, referred for gastroscopy and who were H. pylori positive in the gastric biopsy, were recruited and separated in two groups: case group—individuals with gastric disease (n = 30); control group—individuals with no gastric disease (n = 32); saliva, dental plaque and biopsy samples were collected from all individuals. Oral and biopsy samples were analyzed by PCR using specific primers for H. pylori 16S ribosomal and cagA genes. PCR products were sequenced for DNA homology confirmation. H. pylori was detected neither in dental plaque nor in saliva in the control group. In the case group H. pylori DNA was detected in 16/30 (53.3%) saliva samples and in 11/30 (36.6%) dental plaque samples. The cagA gene was detected in 13/30 (43.3%) gastric biopsies, in 7/16 (43.8%) saliva samples, and in 3/11 (27.3%) dental plaque samples. Eighteen (60.0%) individuals in the case group were H. pylori positive both in oral and biopsy samples, and 8 (26.6%) of those were positive for cagA-H. pylori DNA. H. pylori and its virulent clone showed a higher prevalence in the oral cavity of individuals in the case group than in the control group (p < 0.05). Our results suggest that dental plaque and saliva may serve as temporary reservoir for H. pylori and its virulent cagA variant in individuals with gastric disease.

Candida albicans, the major human fungal pathogen, undergoes a reversible morphological transition from single yeast cells to pseudohyphae and hyphae filaments. The hyphae form is considered the most invasive form of the fungus. The purpose of this study is to investigate the effect of saliva on hyphae growth of C. albicans. Candida albicans hyphae were inoculated in Roswell Park Memorial Institute medium with whole saliva, parotid saliva or buffer mimicking the saliva ion composition, and cultured for 18 h at 37 degrees C under aerobic conditions with 5% CO(2). Whole saliva and parotid saliva induced transition to yeast growth, whereas the culture with buffer remained in the hyphae form. Parotid saliva was fractionated on a reverse-phase C8 column and each fraction was tested for inducing transition to yeast growth. By immunoblotting, the salivary component in the active fraction was identified as statherin, a phosphoprotein of 43 amino acids that has been implicated in remineralization of the teeth. Synthetically made statherin induced transition of hyphae to yeast. By deletion of five amino acids at the negatively charged N-terminal site (DpSpSEE), yeast-inducing activity and binding to C. albicans were increased. In conclusion, statherin induces transition to yeast of C. albicans hyphae and may thus contribute to the oral defense against candidiasis.

Human lactoferrampin is a novel antimicrobial peptide found in the cationic N-terminal lobe of the iron-binding human lactoferrin protein. The amino acid sequence that directly corresponds to the previously characterized bovine lactoferrin-derived lactoferrampin peptide is inactive on its own (WNLLRQAQEKFGKDKSP, residues 269-285). However, by increasing the net positive charge near the C-terminal end of human lactoferrampin, a significant increase in its antibacterial and Candidacidal activity was obtained. Conversely, the addition of an N-terminal helix cap (sequence DAI) did not have any appreciable effect on the antibacterial or antifungal activity of human lactoferrampin peptides, even though it markedly influenced that of bovine lactoferrampin. The solution structure of five human lactoferrampin variants was determined in SDS micelles and all of the structures display a well-defined amphipathic N-terminal helix and a flexible cationic C-terminus. Differential scanning calorimetry studies indicate that this peptide is capable of inserting into the hydrophobic core of a membrane, while fluorescence spectroscopy results suggest that a hydrophobic patch encompassing the single Trp and Phe residues as well as Leu, Ile and Ala side chains mediates the interaction between the peptide and the hydrophobic core of a phospholipid bilayer.

Objective: A survey of European dental schools was conducted in 2006 to determine the curricular structure, techniques and materials used in local anaesthesia teaching to dental students.Materials and methods: A questionnaire was designed to collect information about local anaesthesia education. The questionnaires were sent to the Dean of each dental school in Europe and Israel; 49 returned the completed survey, resulting in a response rate of 18.4%.Results: Results from this survey show that dental schools are managing local anaesthesia education in different ways. At most schools, theoretical teaching begins during the first half of the third year (41%), half a year before the practical instruction (43%). In 37% of the dental schools, students use non-human objects to practice before they inject an anaesthetic in humans. The first injection in humans, usually a fellow student (61%), is mostly supervised by an oral and maxillofacial surgeon (65%). The number of injections under supervision usually depends on the individual capabilities of the student (41%). Ten per cent of the schools need permission of a medical ethics committee for the practical instruction on fellow students. All dental curricula include teaching of mandibular block anaesthesia. The majority also include instruction of infiltration anaesthesia of the upper (98%) and lower (92%) jaws in addition to infra-orbital block anaesthesia (57%). Although 82% of the schools are satisfied with the current curriculum with regard to local anaesthesia, 43% are planning changes, frequently the introduction of preclinical training models.Conclusion: Local anaesthesia teaching programmes show considerable variation across the surveyed European dental schools.

Genital coinfections increase an individual's risk of becoming infected with HIV-1 by sexual contact. Several mechanisms have been proposed to explain this, such as the presence of ulceration and bleeding caused by the coinfecting pathogen. Here we demonstrate that Langerhans cells (LCs) are involved in the increased susceptibility to HIV-1 in the presence of genital coinfections. Although LCs are a target for HIV-1 infection in genital tissues, we found that immature LCs did not efficiently mediate HIV-1 transmission in an ex vivo human skin explant model. However, the inflammatory stimuli TNF-alpha and Pam3CysSerLys4 (Pam3CSK4), the ligand for the TLR1/TLR2 heterodimer, strongly increased HIV-1 transmission by LCs through distinct mechanisms. TNF-alpha enhanced transmission by increasing HIV-1 replication in LCs, whereas Pam3CSK4 acted by increasing LC capture of HIV-1 and subsequent trans-infection of T cells. Genital infections such as Candida albicans and Neisseria gonorrhea not only triggered TLRs but also induced TNF-alpha production in vaginal and skin explants. Thus, during coinfection, LCs could be directly activated by pathogenic structures and indirectly activated by inflammatory factors, thereby increasing the risk of acquiring HIV-1. Our data demonstrate a decisive role for LCs in HIV-1 transmission during genital coinfections and suggest antiinflammatory therapies as potential strategies to prevent HIV-1 transmission.

Insufficient blood supply during acute infarction and chronic ischemia leads to tissue hypoxia which can significantly alter gene expression patterns in the heart. In contrast to most mammals, some teleost fishes are able to adapt to extremely low oxygen levels. We describe here that chronic constant hypoxia (CCH) leads to a smaller ventricular outflow tract, reduced lacunae within the central ventricular cavity and around the trabeculae and an increase in the number of cardiac myocyte nuclei per area in the hearts of two teleost species, zebrafish (Danio rerio) and cichlids (Haplochromis piceatus). In order to identify the molecular basis for the adaptations to CCH, we profiled the gene expression changes in the hearts of adult zebrafish. We have analyzed over 15,000 different transcripts and found 376 differentially regulated genes, of which 260 genes showed increased and 116 genes decreased expression levels. Two notch receptors (notch-2 and notch-3) as well as regulatory genes linked to cell proliferation were transcriptionally upregulated in hypoxic hearts. We observed a simultaneous increase in expression of IGF-2 and IGFbp1 and upregulation of several genes important for the protection against reactive oxygen species (ROS). We have identified here many novel genes involved in the response to CCH in the heart, which may have potential clinical implications in the future. Electronic supplementary material The online version of this article (doi:10.1007/s00360-007-0201-4) contains supplementary material, which is available to authorized users.

In the UK almost one million individuals use cocaine on a regular basis, implying that dentists are likely to encounter individuals that use cocaine. Regular use of this drug may have several orofacial effects, such as perforation of the nasal septum and palate, gingival lesions and erosion of tooth surfaces. In addition, recent use of cocaine increases the risk of a medical emergency during dental treatment, especially when epinephrine-containing local anaesthetics or retraction cords are used. Therefore, dental treatment should be postponed for 6 to 24 hours after the use of cocaine.

Objectives: MUC5B contains sulfated and sialylated oligosaccharides that sequester water required for moisturising the oral mucosa. Xerostomia, in patients with Sjögren syndrome, is generally associated with reduced quantities, rather than altered properties, of saliva. Here, we determined the amount of MUC5B (mRNA and protein) as well as sulfation levels in salivary glands of patients with normal or altered unstimulated salivary flow. Localisation of MUC5B and sulfated MUC5B, as well as total levels sulfated groups were determined and compared with acini basal lamina disorganisation.Patients and methods: In all, 18 patients with normal or altered unstimulated salivary flow and 16 controls were studied. MUC5B mRNA and protein were evaluated in salivary glands by semiquantitative RT-PCR and Western blot analysis. MUC5B sulfation was determined by Western blotting. MUC5B and sulfo-Lewisa antigen localisation were assessed by immunohistochemistry. The total amount of sulfated oligosaccharides was determined microdensitometrically.Results: No significant differences were detected in MUC5B mRNA and protein levels between controls and patients, while sulfo-Lewisa antigen levels were lower in patients. The number of sulfo-Lewisa positive mucous acini was reduced in patients but no correlation was observed between lower levels of sulfation and unstimulated salivary flow. Microdensitometric data confirmed the presence of reduced sulfated oligosaccharides levels in mucous acini from patients with highly disorganised basal lamina.Conclusion: Disorganisation of the basal lamina observed in patients with Sjögren syndrome may lead to dedifferentiation of acinar mucous cells and, as a consequence, alter sulfation of MUC5B. These changes are suggested to represent a novel mechanism that may explain xerostomia in these patients.

3,4-methylenedioxymethamphetamine (MDMA), more commonly known as 'ecstasy' or XTC, is frequently used by young adults in the major cities. Therefore, it is likely that dentists might be confronted with individuals who use ecstasy. This review describes systemic and oral effects of ecstasy. Life-threatening complications include hyperthermia, hyponatraemia and liver failure. In addition, psychotic episodes, depression, panic disorders and impulsive behaviour have been reported. Oral effects include xerostomia, bruxism, and an increased risk of developing dental erosion. Mucosal changes have also been reported. Recent use of ecstasy may interfere with dental treatment. Finally, the potential use of saliva for non-invasive detection of ecstasy is discussed.

Homodimerization of histatin-derived peptides generally led to improved bactericidal activity against Staphylococcus aureus in vitro. In vivo, monomers and dimers were equally active in killing bacteria in mice with a soft tissue infection. Altogether, these peptides are promising compounds for the development of novel therapeutics against infections with drug-resistant bacteria.

In addition to saliva, other oral components such as gingival crevicular fluid, epithelial cells, bacteria, breath, and dental plaque have diagnostic potential. For oral diseases such as caries and periodontal disease, visual diagnosis is usually adequate, but objective diagnostic tests with predictive value are desired. Therefore, prediction models like the Cariogram have been developed that also include oral aspects such as saliva secretion, buffering capacity, and Streptococcus mutans counts for the prediction of caries. Correlation studies on salivary components and caries have not been conclusive, but correlation studies on functional aspects, such as saliva-induced bacterial aggregation and caries, look promising. Modern proteomic techniques make it possible to study simultaneously the many salivary components involved in these functions.

Patients with familial adenomatous polyposis (FAP) develop multiple premalignant colorectal adenomas. Untreated, one or more of these polyps will progress to colorectal carcinoma in middle-aged adults. Extra-intestinal manifestations of FAP are frequently observed and this combination has been called Gardner's syndrome. Oral and maxillofacial symptoms of FAP include an increased risk of jaw osteomas, odontomas and supernumerary or unerupted teeth. Early diagnosis of FAP is crucial and may be life saving. As oral signs usually precede gastrointestinal symptoms, the dentist may play an important role in the diagnosis of FAP.

Udgivelsesdato: 2007-Nov BACKGROUND & AIMS: Impaired mucosal defense plays an important role in the pathogenesis of Crohn's disease (CD), one of the main subtypes of inflammatory bowel disease (IBD). Deleted in malignant brain tumors 1 (DMBT1) is a secreted scavenger receptor cysteine-rich protein with predominant expression in the intestine and has been proposed to exert possible functions in regenerative processes and pathogen defense. Here, we aimed at analyzing the role of DMBT1 in IBD. METHODS: We studied DMBT1 expression in IBD and normal tissues by quantitative reverse transcription-polymerase chain reaction, immunohistochemistry, and mRNA in situ hybridization. Genetic polymorphisms within DMBT1 were analyzed in an Italian IBD case-control sample. Dmbt1(-/-) mice were generated, characterized, and analyzed for their susceptibility to dextran sulfate sodium-induced colitis. RESULTS: DMBT1 levels correlate with disease activity in inflamed IBD tissues. A highly significant fraction of the patients with IBD displayed up-regulation of DMBT1 specifically in the intestinal epithelial surface cells and Paneth cells. A deletion allele of DMBT1 with a reduced number of scavenger receptor cysteine-rich domain coding exons is associated with an increased risk of CD (P = .00056; odds ratio, 1.75) but not for ulcerative colitis. Dmbt1(-/-) mice display enhanced susceptibility to dextran sulfate sodium-induced colitis and elevated Tnf, Il6, and Nod2 expression levels during inflammation. CONCLUSIONS: DMBT1 may play a role in intestinal mucosal protection and prevention of inflammation. Impaired DMBT1 function may contribute to the pathogenesis of CD

OBJECTIVE: The aim of this study was to compare the oral health status of chronic renal failure (CRF) patients on renal replacement therapy with a matched reference population. DESIGN: Cross-sectional study. SUBJECTS: Forty-two dentate CRF patients - aged 25-52 years old - were matched with a reference group of 808 dentate subjects. METHODS: The oral health was assessed using decayed missing filled (DMF) indices, simplified oral hygiene index and periodontal status. An oral health questionnaire was used to assess self-reported dental problems. Student t-tests and chi-square tests were performed to compare the CRF patients with the controls. RESULTS: All index-scores in the CRF patients were comparable with the controls except for number of teeth covered with calculus that was significantly higher (P < 0.05) in CRF patients (4.1 ± 2.6) than in controls (3.0 ± 2.9). The self-reported oral health questionnaire revealed a trend for increased temporomandibular complaints in CRF patients (16.7%vs 5.7% in controls; P = 0.06) as well as bad taste (31.0%vs 6.8% in controls, P = 0.08). CONCLUSIONS: For most dental aspects, the oral health of CRF patients is comparable with controls. © 2006 Blackwell Munksgaard. All rights reserved.

Antimicrobial peptides have been found throughout living nature, yet antimicrobial sequences may still lie hidden within a wide variety of proteins. A rational strategy was developed to select interesting domains, based on the presumed common features of antimicrobial peptides, and to release these from accessible and safe proteins. In silico proteolysis simulations of bovine lactoferrin (bLF) with selected endoproteinases predicted the liberation of peptides that encompasses a cationic amphipathic alpha-helix. Three predicted peptides were synthesized and tested for their biological activity, demonstrating that one single enzyme was sufficient to obtain an antimicrobial peptide. The proof of principle demonstrated that a 32-mer fragment isolated from the endoproteinase AspN digestion of bLF possessed strong antimicrobial activity. Moreover, desalted crude digest had improved activity over native bLF. Hence, selective digestion of bLF increases its antimicrobial activity by release of antimicrobial stretches.

Millions of individuals in Europe and the United States use cocaine regularly. Use of cocaine may have several orofacial effects such as perforation of nasal septum and palate, gingival lesions and erosion of tooth surfaces. Recent use of cocaine may also increase medical risks during dental treatment, especially when local anaesthetics with epinephrine or epinephrine-impregnated retraction cords are used. Therefore, it is recommended to postpone dental treatment at least 6 to 24 hours after the use of cocaine.

Porphyria is a disease of specific enzymes in the biosynthesis of heme, caused by either genetic defects or environmental factors. This review of the literature presents the different types of porphyria and describes their possible causes, clinical signs, diagnosis and therapy. In addition, oral findings observed in patients with porphyria and the potential implications of the disease for dental treatment are discussed.

Many patients on haemodialysis (HD) therapy suffer from a dry mouth and xerostomia. This can be relieved by mechanical and gustatory stimulation or palliative care. The aim of this crossover study was to investigate the effect and preferences of a sugar-free chewing gum (Freedent WhiteTM) and a xanthan gum-based artificial saliva (XialineTM) in the management of xerostomia in chronic HD patients. Sixty-five HD patients participated in a 6-week crossover trial. The artificial saliva was rated significantly lower than the chewing gum for effectiveness, taste and a global assessment. No preference differences were found for gender and age, although older subjects rated the artificial saliva with a higher mark. Thirty-nine subjects (60%) preferred chewing gum, 15% ( n = 10) preferred the artificial saliva. Therefore, both chewing gum and artificial saliva could play an important role in the palliative care of xerostomia in HD patients.

The objective of this study was to investigate the release mechanism and kinetics of the antimicrobial peptide, Dhvar-5, both alone and in combination with gentamicin, from a standard commercial polymethyl methacrylate (PMMA) bone cement. Different amounts of Dhvar-5 were mixed with the bone cement powders of Osteopal and the gentamicin-containing Osteopal G bone cement and their release kinetics from the polymerized cement were investigated. Additionally, the internal structure of the bone cements were analysed by scanning electron microscopy (SEM) of the fracture surfaces. Secondly, porosity was investigated with the mercury intrusion method and related to the observed release profiles. In order to obtain an insight into the mechanical characteristics of the bone cement mixtures, the compressive strength of Osteopal and Osteopal G with Dhvar-5 was also investigated. The total Dhvar-5 release reached 96% in the 100 mg Dhvar-5/g Osteopal cement, whereas total gentamicin release from Osteopal G reached only 18%. Total gentamicin release increased significantly to 67% with the addition of 50 mg Dhvar-5/g, but the Dhvar-5 release was not influenced. SEM showed an increase of dissolved gentamicin crystals with the addition of Dhvar-5. The mercury intrusion results suggested an increase of small pores (

The antimicrobial activity of bovine lactoferrin (bLF) is attributed to lactoferricin, which is situated in the N1-domain of bLF. Recently, another antimicrobial domain consisting of residues 268–284, designated lactoferrampin (LFampin), has been identified in the N1-domain of bLF, which exhibited antimicrobial activity againstCandida albicansand several bacteria. In the present study, the candidacidal activity of a series of peptides spanning this antimicrobial domain was investigated in relation to the charge and the capacity to form a helical conformation in hydrophobic environments. C-Terminal truncation of LFampin resulted in a drastic decrease in candidacidal activity. Positively charged residues clustered at the C-terminal side of the LFampin domain appeared to be crucial for the candidacidal activity. The ability to adopt helical conformations did not change when LFampin was truncated at the C-terminal side. N-Terminally truncated LFampin peptides, truncated up to the sequence 270–284, were more reluctant to adopt a helical conformation. Therefore, we conclude that the C-terminal part of LFampin 265–284, which is the most active peptide, is crucial for its candidacidal activity, due to the presence of clustered positive charges, and that the N-terminal part is essential for activity as it facilitates helix formation.

The effects of antimicrobial peptides on artificial membranes have been well-documented; however, reports on the ultrastructural effects on the membranes of micro-organisms are relatively scarce. We compared the effects of histatin 5 and LL-37, two antimicrobial peptides present in human saliva, on the functional and morphological properties of the Candida albicans cell membrane. Fluorescence microscopy and immunogold transmission electron microscopy revealed that LL-37 remained associated with the cell wall and cell membrane, whereas histatin 5 transmigrated over the membrane and accumulated intracellularly. Freeze-fracture electron microscopy revealed that LL-37 severely affected the membrane morphology, resulting in the disintegration of the membrane bilayer into discrete vesicles, and an instantaneous efflux of small molecules such as ATP as well as larger molecules such as proteins with molecular masses up to 40 kDa. The effects of histatin 5 on the membrane morphology were less pronounced, but still resulted in the efflux of nucleotides. As the morphological defects induced by histatin 5 are much smaller than those induced by LL-37, but the efflux of nucleotides is similar at comparable candidacidal concentrations, we suggest that the loss of nucleotides plays an important role in the killing process.

The peptide domain of salivary agglutinin responsible for its interaction with cell surface protein antigen (PAc) of Streptococcus mutans or bovine lactoferrin was found in the same peptide, scavenger receptor cysteine-rich domain peptide 2 (SRCRP2). Inhibition studies suggest that PAc and lactoferrin, of which residues 480 to 492 seem important, competitively bind to the SRCRP2 domain of salivary agglutinin.

MS is a chronic, immune-mediated inflammatory and neurodegenerative disease of the central nervous system (CNS), with an etiology that is not yet fully understood. The prevalence of MS is highest where environmental supplies of vitamin D are lowest. It is well recognized that the active hormonal form of vitamin D, 1,25-dihydroxyvitamin D (1,25-(OH)(2)D), is a natural immunoregulator with anti-inflammatory action. The mechanism by which vitamin D nutrition is thought to influence MS involves paracrine or autocrine metabolism of 25OHD by cells expressing the enzyme 1 alpha-OHase in peripheral tissues involved in immune and neural function. Administration of the active metabolite 1,25-(OH)(2)D in mice and rats with experimental allergic encephalomyelitis (EAE, an animal model of MS) not only prevented, but also reduced disease activity. 1,25-(OH)(2)D alters dendritic cell and T-cell function and regulates macrophages in EAE. Interestingly, 1,25-(OH)(2)D is thought to be operating on CNS constituent cells as well. Vitamin D deficiency is caused by insufficient sunlight exposure or low dietary vitamin D(3) intake. Subtle defects in vitamin D metabolism, including genetic polymorphisms related to vitamin D, might possibly be involved as well. Optimal 25OHD serum concentrations, throughout the year, may be beneficial for patients with MS, both to obtain immune-mediated suppression of disease activity, and also to decrease disease-related complications, including increased bone resorption, fractures, and muscle weakness.

Udgivelsesdato: 2004-Nov-12 The scavenger receptor cysteine-rich (SRCR) proteins form an archaic group of metazoan proteins characterized by the presence of SRCR domains. These proteins are classified in group A and B based on the number of conserved cysteine residues in their SRCR domains, i.e. six for group A and eight for group B. The protein DMBT1 (deleted in malignant brain tumors 1), which is identical to salivary agglutinin and lung gp-340, belongs to the group B SRCR proteins and is considered to be involved in tumor suppression and host defense by pathogen binding. In a previous study we used nonoverlapping synthetic peptides covering the SRCR consensus sequence to identify a 16-amino acid bacteria-binding protein loop (peptide SRCRP2; QGRVEVLYRGSWGTVC) within the SRCR domains. In this study, using overlapping peptides, we pinpointed the minimal bacteria-binding site on SRCRP2, and thus DMBT1, to an 11-amino acid motif (DMBT1 pathogen-binding site 1 or DMBT1pbs1; GRVEVLYRGSW). An alanine substitution scan revealed that VEVL and Trp are critical residues in this motif. Bacteria binding by DMBT1pbs1 was different from the bacteria binding by the macrophage receptor MARCO in which an RXR motif was critical. In addition, the homologous consensus sequences of a number of SRCR proteins were synthesized and tested for bacteria binding. Only consensus sequences of DMBT1 orthologues bound bacteria by this motif.

The antimicrobial activity of bovine lactoferrin is attributed to lactoferricin, situated in the N1-domain. Based on common features of antimicrobial peptides, a second putative antimicrobial domain was identified in the N1-domain of lactoferrin, designated lactoferrampin. This novel peptide exhibited candidacidal activity, which was substantially higher than the activity of lactoferrin. Furthermore, lactoferrampin was active against Bacillus subtilis, Escherichia coli, and Pseudomonas aeruginosa, but not against the fermenting bacteria Actinomyces naeslundii, Porphyromonas gingivalis, Streptococcus mutans and Streptococcus sanguis. Notably, lactoferrampin is located in the N1-domain in close proximity to lactoferricin, which plays a crucial role in membrane-mediated activities of lactoferrin.

The mechanism of action of antimicrobial peptides is still a matter of debate. The formation of ROS (reactive oxygen species) has been suggested to be the crucial step in the fungicidal mechanism of a number of antimicrobial peptides, including histatin 5 and lactoferrin-derived peptides. In the present study we have investigated the effects of histatin 5 and of a more amphipathic synthetic derivative, dhvar4, on the generation of ROS in the yeast Candida albicans, using dihydroethidium as an indicator for ROS. With both peptides, a substantial enhancement of fluorescence was observed. However, TEMPO (2,2,6,6-tetramethylpiperidine-N-oxyl), a cell-permeant ROS scavenger, did not have an inhibitory effect on killing or on the enhancement of fluorescence. Furthermore, antimycin and azide, which have been reported to induce ROS in vitro, were not able to enhance the dihydroethidium fluorescence, while chlorhexidine, a non-specific antiseptic agent, enhanced dihydroethidium fluorescence to the same extent as did the peptides. Fluorescence microscopy showed the fluorescence enhancement to be a consequence of the release of unbound preformed ethidium from the mitochondrial matrix within the cell. It is concluded that ROS do not play a role in the histatin 5-mediated killing of C. albicans.

SAG (salivary agglutinin), which is identical to gp-340 (glycoprotein-340) from the lung, is encoded by DMBT1 (deleted in malignant brain tumours 1). It is a member of the SRCR (scavenger receptor cysteine-rich) superfamily and contains 14 SRCR domains, 13 of which are highly similar. SAG in saliva is partially complexed with IgA, which may be necessary for bacterial binding. The goal of the present study was to characterize the binding of purified SAG to IgA. SAG binds to a variety of proteins, including serum and secretory IgA, alkaline phosphatase-conjugated IgGs originating from rabbit, goat, swine and mouse, and lactoferrin and albumin. Binding of IgA to SAG is calcium dependent and is inhibited by 0.5 M KCl, suggesting that electrostatic interactions are involved. Binding of IgA was destroyed after reduction of SAG, suggesting that the protein moiety is involved in binding. To pinpoint further the binding domain for IgA on SAG, a number of consensus-based peptides of the SRCR domains and SRCR interspersed domains were designed and synthesized. ELISA binding studies with IgA indicated that only one of the peptides tested, comprising amino acids 18–33 (QGRVEVLYRGSWGTVC) of the 109-amino-acid SRCR domain, exhibited binding to IgA. This domain is identical to the domain of SAG that is involved in binding to bacteria. Despite this similar binding site, IgA did not inhibit binding of Streptococcus mutans to SAG or peptide. These results show that the binding of IgA to SAG is specifically mediated by a peptide sequence on the SRCR domains.

A number of cationic antimicrobial peptides, among which are histatin 5 and the derived peptides dhvar4 and dhvar5, enter their target cells and interact with internal organelles. There still are questions about the mechanisms by which antimicrobial peptides translocate across the membrane. We used a liposome model to study membrane binding, translocation and membrane-perturbing capacities of histatin 5, dhvar4 and dhvar5. Despite the differences in amphipathic characters of these peptides, they bound equally well to liposomes, whereas their membrane activities differed remarkably: dhvar4 translocated at the fastest rate, followed by dhvar5, whereas the histatin 5 translocation rate was much lower. The same pattern was seen for the extent of calcein release: highest with dhvar4, less with dhvar5 and almost none with histatin 5. The translocation and disruptive actions of dhvar5 did not seem to be coupled, because translocation occurred on a much longer timescale than calcein release, which ended within a few minutes. We conclude that peptide translocation can occur through peptide–phospholipid interactions, and that this is a possible mechanism by which antimicrobial peptides enter cells. However, the translocation rate was much lower in this model membrane system than that seen in yeast cells. Thus it is likely that, at least for some peptides, additional features promoting the translocation across biological membranes are involved as well.

The efficacy of prophylactic treatment with human lactoferrin 1-11 (hLF1-11), a broad-spectrum antimicrobial peptide, was studied in a rabbit model of femur infection.Calcium phosphate cement with 50 mg/g hLF1-11 or gentamicin was injected into the femoral canal, after inoculation with Staphylococcus aureus. Three weeks later, slices of the proximal femora were sawn for quantitative bacterial culture and histology.Treatment with hLF1-11 (P0.038) or gentamicin (P0.008) caused a reduction of cfu compared with the untreated control rabbits. The number of sterile cultures was higher in hLF1-11- (3/7) and gentamicin- (5/6) treated animals than in controls (1/7). Radiological and histological analysis showed early bone ingrowth into the cement cracks, and only moderate pathological changes in rabbits with positive cultures.Local prophylaxis with hLF1-11 effectively reduced development of osteomyelitis in a rabbit model, but gentamicin resulted in a larger number of sterile femora.

Osteoclastic bone resorption depends on the activity of various proteolytic enzymes, in particular those belonging to the group of cysteine proteinases. Biochemical studies have shown that cystatins, naturally occurring inhibitors of these enzymes, inhibit bone matrix degradation. Since the mechanism by which cystatins exert this inhibitory effect is not completely resolved yet, we studied the effect of cystatins on bone resorption microscopically and by Ca-release measurements. Calvarial bone explants were cultured in the presence or absence of family 2 cystatins and processed for light and electron microscopic analysis, and the culture media were analyzed for calcium release. Both egg white cystatin and human cystatin C decreased calcium release into the medium significantly. Microscopic analyses of the bone explants demonstrated that in the presence of either inhibitor, a high percentage of osteoclasts was associated with demineralized non-degraded bone matrix. Following a 24-h incubation in the presence of cystatin C, 41% of the cells were adjacent to areas of demineralized non-degraded bone matrix, whereas in controls, this was only 6%. If bone explants were cultured with both PTH and cystatin C, 60% of the osteoclasts were associated with demineralized non-degraded bone matrix, compared to 27% for bones treated with PTH only (P < 0.01). Our study provides evidence that cystatins, the naturally occurring inhibitors of cysteine proteinases, reversibly inhibit bone matrix degradation in the resorption lacunae adjacent to osteoclasts. These findings suggest the involvement of cystatins in the modulation of osteoclastic bone degradation.

Histidine-free variants of salivary histatin 5 have a broad antimicrobial activity against various bacteria. In relation to a possible therapeutic application, we were interested in the susceptibility of these small peptides (14 amino acids long) to microbial proteinases and whether this affects their antimicrobial activity. Analyses by SDS-PAGE of supernatants of peptide-bacteria incubation showed a reduction in protein bands within 15 minutes’ incubation, as a result of cellular internalization. Degradation products of dhvar1 and dhvar2 appeared within one hour in the supernatants of Streptococcus mutans and Staphylococcus aureus. In contrast, the variants dhvar3 and dhvar4 were more resistant to degradation under the same conditions. MALDI-TOF analyses identified cleavage of dhvar1 and dhvar2 at Glu6. The N-terminal peptide part (1–6) of dhvar1 and 2 showed no bactericidal activity, while peptide fragment (7–14) showed a highly reduced bactericidal activity.

Objective:: The mucosal secretory proteins, such as the salivary proteins, play a key role in the acquisition and regulation of the mucosal microflora. Most notably, some microorganisms utilize the host's secretory proteins to adhere to the mucosa; a first step in colonization and infection. The secretory proteins also influence colonization by affecting the binding among microorganisms, a process denoted as coadherence. Previously we reported that acute stressors cause specific changes in saliva composition. The present study investigated to what extent these changes influence saliva-mediated microbial adherence and coadherence (ex vivo). Methods:: Thirty-two male undergraduates provided unstimulated saliva before and during a control condition and two stressors: A memory test and a surgery video presentation. We used saliva-coated microplates to test the adherence of bacteria for which the oral cavity is either a natural reservoir (eg, viridans streptococci) or a portal of entry (eg, Helicobacter pylori). We also tested the saliva-mediated co-adherence between Streptococcus gordonii and the yeast Candida albicans. Correlation analyses were performed to determine the relationships between changes in microbial adherence and the concentrations of potential salivary ligands, viz. cystatin S, the mucins MUC5B and MUC7, S-IgA, lactoferrin, [alpha]-amylase, and total salivary protein. Results:: During the memory test, saliva-mediated adhesion of Streptococcus sanguis, Streptococcus gordonii, and H. pylori increased, whereas the coadherence of C. albicans with S. gordonii decreased. During the surgical video presentation the saliva-mediated adherence of H. pylori, S. sanguis, and Streptococcus mitis increased. These changes were independent of salivary flow rate, but correlated with specific changes in salivary protein composition. Conclusion:: The results show that even moderate stressors, by altering the activity of the mucosal secretory glands, may affect microbial colonization processes such as adherence and coadherence. This study hereby presents a mechanism by which stress may affect the mucosal microflora and susceptibility to infectious disease.

In cancer patients, as in the general population, medication is the most common cause of xerostomia. In general, saliva flow in these patients can be stimulated by mechanical or pharmacological stimulation of the salivary glands. Painful damaged oral mucosa can be treated by softening, lubricating mouthwashes or gels. A specific group of patients are those receiving radiotherapy for malignant tumours in the head and neck region. This treatment is inevitably associated with damages to the oral tissues, including the salivary glands, resulting in salivary gland hypofunction. When residual secretory capacity is present, it is advisable to stimulate the salivary glands by mechanical or gustatory stimuli regularly in these patients as supportive oral care. Alternatively, salivary flow can be stimulated by the use of cholinergic pharmaceutical preparations, such as pilocarpine or cevimeline. After the radiation therapy is ended, a dental check-up should be done every 3 months to allow control of any incipient oral inflammation and dental decay. For daily use, a special dentifrice (e.g. children's toothpaste) is recommended, since the taste of a regular dentifrice may be too strong for these patients. Nocturnal oral dryness can be alleviated by spraying the oral surfaces with water, or by applying a small amount of dentifrice on the dental smooth surfaces. When stimulation of salivary secretion fails, patients can be given palliative oral care in the form of application of mouthwashes and saliva substitutes. The daily use of a mouthwash, e.g. Biotene, Oral Balance or Zendium, or one of the saliva substitutes is indicated. Different types of saliva substitutes are now commercially available, containing different polymers as thickening agents, e.g. carboxymethylcellulose (Oralube and Glandosane), polyacrylic acid, and xanthan gum (Xialine). Recent developments, which are, however, still in the experimental stage, are bio-active saliva substitutes and mouthwashes containing antimicrobial peptides to protect the oral tissues against microbial colonization and to suppress and to cure mucosal and gingival inflammation.

Histatins, salivary antimicrobial peptides, are susceptible to proteolytic degradation, often ascribed to host proteinases. In this study, we addressed the question whether proteolytic activity from microbial sources can contribute to this degradation. Candida albicans, an opportunistic yeast that is susceptible to the histatins, was used as target organism. The most potent histatin (histatin 5: sequence: DSHAKRHHGYKRKFHEKHHSHRGY), two histatin 5 fragments (dh-5: sequence: KRKFHEKHHSHRGY; P-113: sequence: AKRHHGYKRKFH) and an all-D isomer of the latter (P-113D) were used as model peptides. All L-peptides were susceptible to degradation by C. albicans. Cleavage was established at Lys5 and His19 of histatin 5, Lys11, Arg12, Phe14, Glu16, Lys17, His18 and Ser20 of dh-5 and Ala4 and Lys11 of P-113. In addition, it was found that secreted C. albicans enzymes are not involved in the degradation process and that blocking cell entry of the peptides greatly impedes degradation. Moreover, P-113D, which is biologically as active as P-113, was hardly susceptible to proteolysis. These data imply that proteolysis occurs mainly intracellularly and is not used as a protective mechanism against histatin activity. Together, our results suggest that, besides host proteinases, microbial enzymes play an important role in histatin degradation.

The effects of polymers applicable in saliva substitutes on the anti-Candida activity of the cationic antimicrobial peptide dhvar1 were investigated. Dhvar1 is a derivative of the 14 C-terminal amino acids of histatin 5. The effects of the following polymers were tested: uncharged hydroxyethylcellulose (HEC), negatively charged xanthan (XG) and three types of negatively charged carboxymethylcellulose (CMC) of identical mass but different degrees of carboxylic acid-group substitution (DS). The effects were tested at pH 4.0, 7.0 and 8.5 in a killing assay. HEC had no effect at any pH tested; XG and the three types of CMC caused a decrease in activity at increasing concentrations. Within the CMC group, inhibition increased slightly with increasing DS. These results suggest that the reduction in activity associated with these polymers is the result of electrostatic interaction between the positively charged peptides and the negatively charged polymers. In the absence of polymers, no effect of pH was found on the activity of dhvar1. In the presence of the charged polymers XG and CMC, lowering the pH from 7.0 to 4.0 resulted in a decrease of dhvar1 activity. It was concluded that, with respect to the retention of activity, HEC is the most appropriate polymer for use in combination with dhvar1. However, for use in saliva substitutes XG seems more suitable because of its rheological properties. If XG or CMC are to be used, their reductive effect on the anti-Candida activity of dhvar1 should be compensated for by increasing the peptide dose.

Background-The common occurrence of xerostomia in Sjogren's syndrome (SS) as well as the easy accessibility of saliva supports the use of sialometry and sialochemistry in the diagnosis of SS. Collection and analysis of whole saliva (oral fluid) is currently the routine technique for sialometry, despite the fact that it is rather inaccurate and impure.Objective-To assess the value of glandular sialometry and sialochemistry as diagnostic instruments in SS.Methods-In a group of 100 consecutive patients referred for diagnosis of SS, glandular secretory flow rates and a spectrum of salivary components (sodium, potassium, chloride, calcium, phosphate, urea, amylase, total protein) were assessed. The patients were classified as positive or negative for SS according to the revised European classification criteria.Results-Patients with SS differed clearly from those who tested negative for SS, showing lower submandibular/sublingual (SM/SL) flow rates and an appreciably changed salivary composition of parotid and SM/SL saliva. Besides changes in salivary flow rate and composition, distinct sialometric profiles were observed, characteristic of either early or late salivary manifestation of SS, or of the xerogenic side effects of medication.Conclusions-Glandular sialometry and sialochemistry are not only useful tools for differentiating SS from other salivary gland disease in clinical practice, but they also have great potential as diagnostic criteria for SS, showing distinct sialometric and sialochemical changes as well as profiles. Being simple, safe (non-invasive), and sensitive (early disease detection), they have three major advantages over other oral tests for SS.

This case report describes a 9-year-old boy with severe tooth wear as a result of drinking a single glass of soft drink per day. This soft drink was consumed over a period of one to two hours, while he was gaming intensively on his computer. As a result, a deep bite, enamel cupping, sensitivity of primary teeth and loss of fillings occurred. Therefore, dentists should be aware that in patients who are gaming intensively, the erosive potential of soft drinks can be potentiated by mechanical forces leading to excessive tooth wear.

Acid production, in particular at low pH, is thought to be an important ecological determinant in dental caries. The aim of the present study was to determine the acid producing capability at different pH levels of 47 streptococcal strains, representing 9 species, isolated from human dental plaque. The bacteria were grown until mid log–phase under anaerobic conditions and acid production was measured in a pH–stat system at pH 7.0, 6.0, 5.5 and 5.0. At all pH values, the mean velocity of acid production (Vap) by Streptococcus mutans and S. sobrinus was significantly higher (pS. mitis, S. oralis, S. gordonii, S. sanguis, S. intermedius, S. anginosus, S. constellatus, and S. vestibularis. However, the Vap of some strains of S. mitis biovar 1 and S. oralis, particularly at pH values of 7.0 and 6.0, exceeded that of some strains of S. mutans. The Vap decreased with pH for all strains, but some strains of S. mitis biovar 1 and strains of the mutans streptococci maintained a relatively high rate of acid production. The results suggest that some strains of S. mitis biovar 1 and S. oralis may play an important role in caries development by modifying the environment in dental plaque to become favourable for the succession of aciduric species. The study furthermore emphasises the need for detailed species and biovar identification of oral streptococci and for recognition of the significant physiological differences that occur within single species.