%0 Journal Article
%~ Pubmed
%A Charadram, Nattida
%A Farahani, Ramin M
%A Harty, Derek
%A Rathsam, Catherine
%A Swain, Michael V
%A Hunter, Neil
%T Regulation of reactionary dentin formation by odontoblasts in response to polymicrobial invasion of dentin matrix.
%B Bone
%D 2012
%V 50
%N 1
%P 265-75
%@ 1873-2763
%X Odontoblast synthesis of dentin proceeds through discrete but overlapping phases characterized by formation of a patterned organic matrix followed by remodelling and active mineralization. Microbial invasion of dentin in caries triggers an adaptive response by odontoblasts, culminating in formation of a structurally altered reactionary dentin, marked by biochemical and architectonic modifications including diminished tubularity. Scanning electron microscopy of the collagen framework in reactionary dentin revealed a radically modified yet highly organized meshwork as indicated by fractal and lacunarity analyses. Immuno-gold labelling demonstrated increased density and regular spatial distribution of dentin sialoprotein (DSP) in reactionary dentin. DSP contributes putative hydroxyapatite nucleation sites on the collagen scaffold. To further dissect the formation of this altered dentin matrix, the associated enzymatic machinery was investigated. Analysis of extracted dentin matrix indicated increased activity of matrix metalloproteinase-2 (MMP-2) in the reactionary zone referenced to physiologic dentin. Likewise, gene expression analysis of micro-dissected odontoblast layer revealed up-regulation of MMP-2. Parallel up-regulation of tissue inhibitor of metalloproteinase-2 (TIMP-2) and membrane type 1- matrix metalloproteinase (MT1-MMP) was observed in response to caries. Next, modulation of odontoblastic dentinogenic enzyme repertoire was addressed. In the odontoblast layer expression of Toll-like receptors was markedly altered in response to bacterial invasion. In carious teeth TLR-2 and the gene encoding the corresponding adaptor protein MyD88 were down-regulated whereas genes encoding TLR-4 and adaptor proteins TRAM and Mal/TIRAP were up-regulated. TLR-4 signalling mediated by binding of bacterial products has been linked to up-regulation of MMP-2. Further, increased expression of genes encoding components of the TGF-?? signalling pathway, namely SMAD-2 and SMAD-4, may explain the increased synthesis of collagen by odontoblasts in caries. These findings indicate a radical adaptive response of odontoblasts to microbial invasion of dentin with resultant synthesis of modified mineralized matrix.
%Z FOR Codes: 110506



%0 Journal Article
%~ Pubmed
%A Li, Nan
%A Yun, Peter
%A Jeffries, Cy M
%A Langley, David
%A Gamsjaeger, Roland
%A Church, W Bret
%A Hunter, Neil
%A Collyer, Charles A
%T The modular structure of haemagglutinin/adhesin regions in gingipains of Porphyromonas gingivalis.
%B Molecular microbiology
%D 2011
%V 81
%N 5
%P 1358-73
%@ 1365-2958
%X High-molecular-weight arginine- and lysine-specific (Kgp) gingipains are essential virulence factors expressed by the oral pathogen Porphyromonas gingivalis. Haemagglutinin/adhesin (HA) regions of these proteases have been implicated in targeting catalytic domains to biological substrates and in other adhesive functions. We now report the crystal structure of the K3 adhesin domain/module of Kgp, which folds into the distinct ??-jelly roll sandwich topology previously observed for K2. A conserved structural feature of K3, previously observed in the Kgp K2 module, is the half-way point anchoring of the surface exposed loops via an arginine residue found in otherwise highly variable sequences. Small-angle X-ray scattering data for the recombinant construct K1K2K3 confirmed a structure comprising a tandem repeat of three homologous modules, K1, K2 and K3 while also indicating an unusual 'y'-shape arrangement of the modules connected by variable linker sequences. Only the K2 and K3 modules and a K1K2 construct were observed to be potently haemolytic. K2, K3 and the K1K2 construct showed preferential recognition of haem-albumin over albumin whereas only low affinity binding was detected for K1 and the K1K2K3 construct. The data indicate replication of some biological functions over the three adhesin domains of Kgp while other functions are restricted.
%Z FOR Codes: 601



%0 Journal Article
%~ Pubmed
%A Ye, Ping
%A Yu, Hong
%A Simonian, Mary
%A Hunter, Neil
%T Ligation of CD24 expressed by oral epithelial cells induces kinase dependent decrease in paracellular permeability mediated by tight junction proteins.
%B Biochemical and Biophysical Research Communications
%D 2011
%V 412
%N 1
%P 165-9
%@ 1090-2104
%X In previous studies we demonstrated uniform strong expression of CD24 in the epithelial attachment to the tooth and in the migrating epithelium of the periodontitis lesion. Titers of serum antibodies auto-reactive with CD24 peptide correlated with reduced severity of periodontal disease. In the present study an epithelial culture model with close correspondence for expression patterns for tight junction components in periodontal epithelia was used. Ligation of CD24 expressed by oral epithelial cells with an anti-CD24 antibody induced formation of tight junctions and live-cell imaging confirmed that paracellular diffusion of fluorochrome-labeled dextran was reduced. Expression of mRNA and protein for zona occludens-1, -2, junction adhesion molecule-A (JAM-A), occludin and claudins-1, -4, -8, -15, -18 was significantly increased following ligation of CD24 but only claudins-4 and -15, JAM-A, occludin and zona occludens-1 and -2 were increased at cell contacts. This change in expression patterns reflected that observed between the epithelium of the periodontal lesion and that of the healthy gingival attachment. In the model system, response profiles to kinase inhibitors indicated a key role of c-Src kinase activation in the development of diffusion-limiting tight junction complexes. Activation was confirmed by demonstrating concomitant phosphorylation of the kinase. Pre-incubation with antibodies against JAM-A and claudin-15 prevented barrier-enhancing effects of anti-CD24 antibodies while pre-incubation with antibody to claudin-4 was partially effective. It is concluded that antibodies to CD24 facilitate expression and location of JAM-A, claudins-4 and -15 that mediate enhanced epithelial barrier function in a protective response against bacterial products.
%Z FOR Codes: 1105



%0 Journal Article
%~ Pubmed
%A Harty, Derek W S
%A Hunter, Neil
%T Carboxypeptidase activity common to viridans group streptococci cleaves angiotensin I to angiotensin II: an activity homologous to angiotensin-converting enzyme (ACE).
%B Microbiology
%D 2011
%V 157
%N Pt 7
%P 2143-51
%@ 1465-2080
%X We have found that Streptococcus gordonii FSS2, an infective endocarditis (IE) isolate, expresses a dipeptidyl-carboxypeptidase with activity homologous to angiotensin-converting enzyme (ACE). The carboxypeptidase activity was purified to homogeneity as a complex/aggregate from a bacterial surface extract and was also active as a 165 kDa monomer. The specific activity for the carboxypeptidase activity was eightfold higher than that for recombinant human ACE. Selected ACE inhibitors, captopril, lisinopril and enalapril, did not inhibit the ACE activity. The carboxypeptidase also hydrolysed the A?? and B??-chains of human fibrinogen, which resulted in impaired fibrin formation by thrombin. The gene encoding ACE carboxypeptidase activity was sequenced and the inferred polypeptide product showed 99???% amino acid homology to SGO_0566, sgc, 'challisin' of S. gordonii CL1 Challis, and had no significant amino acid sequence homology to human ACE. Homologues of challisin ACE activity were commonly detected among the viridans group streptococci most often associated with IE.
%Z FOR Codes: 1105



%0 Journal Article
%~ Pubmed
%A Zou, W
%A Hunter, N
%A Swain, M V
%T Application of polychromatic ??CT for mineral density determination.
%B Journal of dental research
%D 2011
%V 90
%N 1
%P 18-30
%@ 1544-0591
%X Accurate assessment of mineral density (MD) provides information critical to the understanding of mineralization processes of calcified tissues, including bones and teeth. High-resolution three-dimensional assessment of the MD of teeth has been demonstrated by relatively inaccessible synchrotron radiation microcomputed tomography (SR??CT). While conventional desktop ??CT (C??CT) technology is widely available, polychromatic source and cone-shaped beam geometry confound MD assessment. Recently, considerable attention has been given to optimizing quantitative data from C??CT systems with polychromatic x-ray sources. In this review, we focus on the approaches that minimize inaccuracies arising from beam hardening, in particular, beam filtration during the scan, beam-hardening correction during reconstruction, and mineral density calibration. Filtration along with lowest possible source voltage results in a narrow and near-single-peak spectrum, favoring high contrast and minimal beam-hardening artifacts. More effective beam monochromatization approaches are described. We also examine the significance of beam-hardening correction in determining the accuracy of mineral density estimation. In addition, standards for the calibration of reconstructed grey-scale attenuation values against MD, including K(2)PHO(4) liquid phantom, and polymer-hydroxyapatite (HA) and solid hydroxyapatite (HA) phantoms, are discussed.
%Z FOR Codes: 110501 90303



%0 Journal Article
%~ Pubmed
%A Farahani, Ramin M
%A Simonian, Mary
%A Hunter, Neil
%T Blueprint of an ancestral neurosensory organ revealed in glial networks in human dental pulp.
%B The Journal of Comparative Neurology
%D 2011
%V 519
%N 16
%P 3306-26
%@ 1096-9861
%X Sensory function of human dental pulp has long been known. A composite role has been suggested for odontoblasts as sensory cells in addition to the synthesis of dentinal matrix. However, the neural basis for such a composite sensory activity remains enigmatic. Here, we aimed to probe the question by pursuing an evolutionary logic; if dental pulp is a vestigial sensory organ co-opted to a function of synthesis of mineralized matrix, essential elements of neurosensory organs may persist in dental pulp. Through structural analysis by confocal laser scanning microscopy, three distinct cell populations adjacent to odontoblasts, glial fibrillary acidic protein (GFAP)(+) seracytes, S100(+) telacytes, and HLA-II(+) alacytes were identified in peripheral human dental pulp. Subsequent molecular fingerprinting by quantitative reverse transcriptase-polymerase chain reaction established these cells as analogous to radial glia (GFAP(+) cells), astrocytes (S100(+) cells), and microglia (HLA-II(+) cells) of central nervous system organs. In the cell-rich zone of the pulp, S100(+) cells formed a network, ensheathed unmyelinated axons, and extended end-feet around the capillaries. The microcirculation adjacent to the glial cells in the cell-rich zone possessed ultrastructural features and a gene expression profile typical of the blood-brain barrier system. These novel findings support a new paradigm for understanding sensory functionality of dental pulp by the demonstration of a sophisticated neural structure in the human dental pulp that is analogous to other central sensory organs. Further, the structure that is revealed informs the concept of the evolutionary origin of the dental pulp, suggesting that a neurosensory organ was the precursor structure of teeth.
%Z FOR Codes: 1105



%0 Journal Article
%~ Pubmed
%A Gao, Jin-Long
%A Nguyen, Ky-Anh
%A Hunter, Neil
%T Characterization of a hemophore-like protein from Porphyromonas gingivalis.
%B Journal of Biological Chemistry
%D 2010
%V 285
%N 51
%P 40028-38
%@ 1083-351X
%X The porphyrin auxotrophic pathogen Porphyromonas gingivalis obtains the majority of essential iron and porphyrin from host hemoproteins. To achieve this, the organism expresses outer membrane gingipains containing cysteine proteinase domains linked to hemagglutinin domains. Heme mobilized in this way is taken up by P. gingivalis through a variety of potential portals where HmuY/HmuR of the hmu locus are best described. These receptors have relatively low binding affinities for heme. In this report, we describe a novel P. gingivalis protein, HusA, the product of PG2227, which rapidly bound heme with a high binding constant at equilibrium of 7 ?? 10(-10) M. HusA is both expressed on the outer membrane and released from the organism. Spectral analysis indicated an unusual pattern of binding where heme was ligated preferentially as a dimer. Further, the presence of dimeric heme induced protein dimer formation. Deletional inactivation of husA showed that expression of this moiety was essential for growth of P. gingivalis under conditions of heme limitation. This finding was in accord with the pronounced increase in gene expression levels for husA with progressive reduction of heme supplementation. Antibodies reactive against HusA were detected in patients with chronic periodontitis, suggesting that the protein is expressed during the course of infection by P. gingivalis. It is predicted that HusA efficiently sequesters heme from gingipains and fulfills the function of a high affinity hemophore-like protein to meet the heme requirement for growth of P. gingivalis during establishment of infection.
%Z FOR Codes: 60501 110508



%0 Journal Article
%~ Pubmed
%A Farahani, Ramin M
%A Nguyen, Ky-Anh
%A Simonian, Mary
%A Hunter, Neil
%T Adaptive calcified matrix response of dental pulp to bacterial invasion is associated with establishment of a network of glial fibrillary acidic protein+/glutamine synthetase+ cells.
%B The American Journal of Pathology
%D 2010
%V 177
%N 4
%P 1901-14
%@ 1525-2191
%X We report evidence for anatomical and functional changes of dental pulp in response to bacterial invasion through dentin that parallel responses to noxious stimuli reported in neural crest-derived sensory tissues. Sections of resin-embedded carious adult molar teeth were prepared for immunohistochemistry, in situ hybridization, ultrastructural analysis, and microdissection to extract mRNA for quantitative analyses. In odontoblasts adjacent to the leading edge of bacterial invasion in carious teeth, expression levels of the gene encoding dentin sialo-protein were 16-fold greater than in odontoblasts of healthy teeth, reducing progressively with distance from this site of the carious lesion. In contrast, gene expression for dentin matrix protein-1 by odontoblasts was completely suppressed in carious teeth relative to healthy teeth. These changes in gene expression were related to a gradient of deposited reactionary dentin that displayed a highly modified structure. In carious teeth, interodontoblastic dentin sialo-protein(-) cells expressing glutamine synthetase (GS) showed up-regulation of glial fibrillary acidic protein (GFAP). These cells extended processes that associated with odontoblasts. Furthermore, connexin 43 established a linkage between adjacent GFAP(+)/GS(+) cells in carious teeth only. These findings indicate an adaptive pulpal response to encroaching caries that includes the deposition of modified, calcified, dentin matrix associated with networks of GFAP(+)/GS(+) interodontoblastic cells. A regulatory role for the networks of GFAP(+)/GS(+) cells is proposed, mediated by the secretion of glutamate to modulate odontoblastic response.
%Z FOR Codes: 110599



%0 Journal Article
%~ Pubmed
%A Nadkarni, Mangala A
%A Simonian, Mary R
%A Harty, Derek W S
%A Zoellner, Hans
%A Jacques, Nicholas A
%A Hunter, Neil
%T Lactobacilli are prominent in the initial stages of polymicrobial infection of dental pulp.
%B Journal of Clinical Microbiology
%D 2010
%V 48
%N 5
%P 1732-40
%@ 1098-660X
%X In earlier studies we used molecular methods to identify the major bacterial consortia associated with advanced dentin caries. These consortia are dominated by bacteria from the families Lactobacillaceae, Streptococcaceae, Veillonellaceae (formerly Acidaminococcaceae), Eubacteriaceae, and Lachnospiraceae from the phylum Firmicutes; Coriobacteriaceae, Bifidobacteriaceae, and Propionibacteriaceae from the phylum Actinobacteria; and Prevotellaceae from the phylum Bacteroidetes, as well as fusobacteria. The phases of infection of vital pulp tissue by dentin microorganisms remain obscure. In the present study, fluorescence in situ hybridization was performed on sections of tissue embedded in resin. Probes for 16S rRNA corresponding to the major taxa of bacteria in carious dentin were used to provide information on the characteristics of pulp infection. Lactobacilli were prominent in 7 of 8 pulps determined to be at a limited stage of infection. Established infection (6 pulps) showed a more complex profile, with lactobacilli persisting in all of the lesions and with invasion of the necrotic regions of tissue by Bacteroidetes, fusobacteria, Lachnospiraceae, and Coriobacteriaceae in particular. Advanced infections (7 pulps) were characterized by mixed anaerobic species, with a strong representation by Coriobacteriaceae and Lachnospiraceae. Lactobacilli were not represented at this stage. Typically, groups of organisms were spatially isolated within the pulp tissue. Analysis indicated that lactobacilli could invade vital pulp tissue to achieve a very high biomass that was not associated with a detectable local inflammatory infiltrate. The findings establish that invasion of the dental pulp can be associated with a pronounced selection from the complex microbial populations within carious dentin, suggesting specific pathogenicity.
%Z FOR Codes: 110502



%0 Journal Article
%~ Pubmed
%A Li, Nan
%A Yun, Peter
%A Nadkarni, Mangala A
%A Ghadikolaee, Nazila Babapoor
%A Nguyen, Ky-Anh
%A Lee, Mihwa
%A Hunter, Neil
%A Collyer, Charles A
%T Structure determination and analysis of a haemolytic gingipain adhesin domain from Porphyromonas gingivalis.
%B Molecular microbiology
%D 2010
%V 76
%N 4
%P 861-73
%@ 1365-2958
%X Porphyromonas gingivalis is an obligately anaerobic bacterium recognized as an aetiological agent of adult periodontitis. P. gingivalis produces cysteine proteinases, the gingipains. The crystal structure of a domain within the haemagglutinin region of the lysine gingipain (Kgp) is reported here. The domain was named K2 as it is the second of three homologous structural modules in Kgp. The K2 domain structure is a 'jelly-roll' fold with two anti-parallel beta-sheets. This fold topology is shared with adhesive domains from functionally diverse receptors such as MAM domains, ephrin receptor ligand binding domains and a number of carbohydrate binding modules. Possible functions of K2 were investigated. K2 induced haemolysis of erythrocytes in a dose-dependent manner that was augmented by the blocking of anion transport. Further, cysteine-activated arginine gingipain RgpB, which degrades glycophorin A, sensitized erythrocytes to the haemolytic effect of K2. Cleaved K2, similar to that found in extracted Kgp, lacks the haemolytic activity indicating that autolysis of Kgp may be a staged process which is artificially enhanced by extraction of the protein. The data indicate a functional role for K2 in the integrated capacity conferred by Kgp to enable the porphyrin auxotroph P. gingivalis to capture essential haem from erythrocytes.
%Z FOR Codes: 90301



%0 Journal Article
%~ Pubmed
%A Ye, Ping
%A Nadkarni, Mangala A
%A Simonian, Mary
%A Hunter, Neil
%T CD24 regulated gene expression and distribution of tight junction proteins is associated with altered barrier function in oral epithelial monolayers.
%B BMC cell biology [electronic resource]
%D 2009
%V 10
%N 
%P 2
%@ 1471-2121
%X Control of intercellular penetration of microbial products is critical for the barrier function of oral epithelia. We demonstrated that CD24 is selectively and strongly expressed in the cells of the epithelial attachment to the tooth and the epithelial lining of the diseased periodontal pocket and studies in vitro showed that CD24 regulated expression of the epithelial intercellular adhesion protein E-cadherin.
%Z FOR Codes: 60108



%0 Journal Article
%~ Pubmed
%A Nadkarni, Mangala A
%A Chhour, Kim-Ly
%A Browne, Gina
%A Jacques, Nicholas A
%A Hunter, Neil
%T Lysine gingipain (kgp) biovars of Porphyromonas gingivalis exhibit differential distribution on oral mucosal sites.
%B Journal of Clinical Microbiology
%D 2009
%V 47
%N 10
%P 3350-2
%@ 1098-660X
%X A predominant kgp biovar colonized subgingival sites and buccal and tongue mucosa in 45 of 56 adults in an isolated community. The presence of biovars 381, W83, and W83v, but not HG66, correlated with the Porphyromonas gingivalis load at diseased sites. Biovars W83 and W83v poorly colonized tongue and buccal mucosa.
%Z FOR Codes: 60504



%0 Journal Article
%~ Pubmed
%A Nadkarni, Mangala A
%A Martin, F Elizabeth
%A Hunter, Neil
%A Jacques, Nicholas A
%T Methods for optimizing DNA extraction before quantifying oral bacterial numbers by real-time PCR.
%B FEMS Microbiology Letters
%D 2009
%V 296
%N 1
%P 45-51
%@ 1574-6968
%X Methods for the optimal extraction of genomic DNA for real-time PCR enumeration of oral bacteria using the muramidase, mutanolysin, were developed using a simple in vitro oral flora model comprised of the facultative anaerobic gram-positive bacteria, Lactobacillus acidophilus and Streptococcus mutans, the gram-positive anaerobe, Parvimonas micra, and the gram-negative anaerobes, Porphyromonas gingivalis, Prevotella melaninogenica and Fusobacterium nucleatum. Traditional, as well as more elaborate, methods of quantifying bacterial numbers, including colony counting and estimation of DNA content using 4',6-diamino-2-phenylindole were compared in order to validate the real-time PCR approach. Evidence was obtained that P. gingivalis nuclease activity adversely affected the extraction of double-stranded DNA from this bacterium either alone or when it formed part of a consortium with the other bacteria. This nuclease activity could be overcome by treatment of the bacteria with either 20 mM diethyl pyrocarbonate or 70% ethanol at 4 degrees C overnight. A final purification of the DNA to remove any potential PCR inhibitors was added to the protocol in order to accurately quantify the amount of DNA by real-time PCR and hence the number of bacteria in a sample.
%Z FOR Codes: 60599



%0 Book Section
%A Nadkarni, Mangala
%A Martin, FE
%A Jaques, NA
%A Hunter, Neil
%T Caries
%B Encyclopedia of Molecular Mechanisms of Disease
%D 2009
%C United States
%I Springer New York LLC
%V 
%N 
%P 279-280
%@ 9783540671367
%E Lang, Florian
%X 
%Z FOR Codes: 60504



%0 Journal Article
%~ Pubmed
%A Smith-Vaughan, Heidi
%A Byun, Roy
%A Halpin, Stephen
%A Nadkarni, Mangala A
%A Jacques, Nicholas A
%A Hunter, Neil
%A Morris, Peter S
%A Leach, Amanda J
%T Interventions for prevention of otitis media may be most effective if implemented in the first weeks of life.
%B International journal of pediatric otorhinolaryngology
%D 2008
%V 72
%N 1
%P 57-61
%@ 0165-5876
%X For Indigenous Australian children living in remote communities, onset of otitis media commences within weeks of birth and is associated with early nasopharyngeal colonisation with multiple respiratory bacterial pathogens: Streptococcus pneumoniae, Haemophilus influenzae, and Moraxella catarrhalis. The high prevalence of eardrum perforation and the failure of standard therapies to cure or prevent OM in this population require urgent attention. The objective of this study was to measure the changes in nasopharyngeal bacterial flora between birth and first episode of otitis media.
%Z FOR Codes: 60504 111403



%0 Journal Article
%~ Pubmed
%A Fong, Michele M
%A Darendeliler, M Ali
%A Hunter, N
%A Shen, Gang
%T Epithelial cells in PDL are critical in resuming the integral relation between tooth root and supporting bone after trauma--a transplantation experiment.
%B Archives of oral biology
%D 2007
%V 52
%N 2
%P 182-8
%@ 0003-9969
%X The aim of this study was to determine the response of the residual epithelial network following traumatic breakdown of the periodontal ligament (PDL).
%Z FOR Codes: 110508



%0 Journal Article
%~ Pubmed
%A Byun, Roy
%A Carlier, Jean-Philippe
%A Jacques, Nicholas A
%A Marchandin, Helene
%A Hunter, Neil
%T Veillonella denticariosi sp. nov., isolated from human carious dentine.
%B International journal of systematic and evolutionary microbiology
%D 2007
%V 57
%N Pt 12
%P 2844-8
%@ 1466-5026
%X Selective culture of human carious dentine for Veillonella strains resulted in the isolation of two strains of a Gram-negative, coccus-shaped bacterium that has not been described previously. Comparative 16S rRNA and dnaK gene sequence analysis indicated that the two strains were homogeneous and comprised a distinct lineage within the genus Veillonella, phylogenetically most closely related to Veillonella rodentium. This was supported by DNA-DNA hybridization, which showed clearly that the two strains were similar and distinct from other Veillonella species, and the production of major cellular fatty acids (C(13 : 0) and C(17 : 1)omega8), which is consistent with other members of the genus Veillonella. Based on these observations, strains RBV81 and RBV106(T) represent a novel species, for which the name Veillonella denticariosi sp. nov. is proposed, with the type strain RBV106(T) (=CIP 109448(T) =CCUG 54362(T) =DSM 19009(T)).
%Z FOR Codes: 110508



%0 Journal Article
%~ Pubmed
%A Yun, L W P
%A Decarlo, A A
%A Hunter, N
%T Blockade of protease-activated receptors on T cells correlates with altered proteolysis of CD27 by gingipains of Porphyromonas gingivalis.
%B Clinical and Experimental Immunology
%D 2007
%V 150
%N 2
%P 217-29
%@ 1365-2249
%X Cysteine proteinases, termed gingipains, of Porphyromonas gingivalis are able to inactivate a broad range of host proteins involved in cellular responses and have been implicated as key virulence factors in the onset and progression of adult periodontitis. In the present study, the high molecular weight Arg-gingipain, RgpA, produced a time- and concentration-dependent hydrolysis of the tumour necrosis factor (TNF)-alpha receptor family member CD27 on resting T cells. As a consequence of CD27 degradation, a reduction in CD27-ligation dependent co-stimulatory CD40L expression was observed. Concomitantly, RgpA activated the protease-activated receptors (PAR)-1, PAR-2 and PAR-4 and induced CD69 and CD25 expression on T cells, thereby demonstrating T cell activation. The Lys-gingipain Kgp demonstrated a low capacity to degrade CD27 but the ability to affect CD27 expression and biological activity was increased when T cells were pretreated with blocking peptide against PAR-2. CD70, the ligand for CD27 induced on activated B cells, was significantly reduced by RgpA treatment and weakly affected by Kgp. These findings suggest that while RgpA can activate T cells through PARs, the parallel action of direct hydrolysis of membrane CD27 as well as CD70 indicates a potential down-regulatory effect through inhibition of CD27/CD70-mediated cell activation in periodontitis.
%Z FOR Codes: 110508



%0 Journal Article
%~ Pubmed
%A Ye, P
%A Nadkarni, M A
%A Hunter, N
%T Regulation of E-cadherin and TGF-beta3 expression by CD24 in cultured oral epithelial cells.
%B Biochemical and Biophysical Research Communications
%D 2006
%V 349
%N 1
%P 229-35
%@ 0006-291X
%X We previously reported evidence that patients with periodontitis have serum antibodies to oral Gram positive bacteria that are cross-reactive with epithelial antigens, including CD24. High level expression of CD24 was confined to the reactive periodontal epithelium and inflamed gingival attachment. As a model for the reactive epithelium of chronic periodontitis, H413 epithelial cells derived from a human oral squamous cell carcinoma were cloned and lines expressing high levels of CD24 were selected. RNA interference protocols were designed to determine if CD24 could modulate intercellular interactions and regulate the biology of these epithelial cells. Knock-down of CD24 protein was demonstrated by Western blot and flow cytometry. The level of mRNA for CD24 was reduced 90% by RNAi treatment as assayed by real-time, reverse transcriptase (RT)-PCR. Gene products known to be important in epithelial biology, including E-cadherin and TGF-beta3 that were demonstrated to undergo altered expression patterns in the periodontal lesion, were investigated. Down-regulation of CD24 mRNA was associated with reduced e-cadherin expression and up-regulated expression of snail, twist, and tgf-beta3. The cells were treated with monoclonal antibodies to CD24 to mimic the action of auto-reactive antibodies to CD24 detected in affected patients. Relative to isotype control antibody, stimulation by anti-CD24 antibodies induced up-regulated expression of e-cadherin and down-regulation of tgf-beta3 as assessed by real-time RT-PCR. No consistent changes for expression of beta-catenin, connexins, integrins, icam-1, tgf-beta1 or tgf-beta2 were observed. CD24 could play an important role in modulating expression of genes that regulate epithelial differentiation in periodontal disease.
%Z FOR Codes: 60108



%0 Journal Article
%~ Pubmed
%A Smith-Vaughan, Heidi
%A Byun, Roy
%A Nadkarni, Mangala
%A Jacques, Nicholas A
%A Hunter, Neil
%A Halpin, Stephen
%A Morris, Peter S
%A Leach, Amanda J
%T Measuring nasal bacterial load and its association with otitis media.
%B BMC ear, nose, and throat disorders [electronic resource]
%D 2006
%V 6
%N 
%P 10
%@ 1472-6815
%X Nasal colonisation with otitis media (OM) pathogens, particularly Streptococcus pneumoniae, Haemophilus influenzae and Moraxella catarrhalis, is a precursor to the onset of OM. Many children experience asymptomatic nasal carriage of these pathogens whereas others will progress to otitis media with effusion (OME) or suppurative OM. We observed a disparity in the prevalence of suppurative OM between Aboriginal children living in remote communities and non-Aboriginal children attending child-care centres; up to 60% and <1%, respectively. This could not be explained by the less dramatic difference in rates of carriage of respiratory bacterial pathogens (80% vs 50%, respectively). In this study, we measured nasal bacterial load to help explain the different propensity for suppurative OM in these two populations.
%Z FOR Codes: 60504



%0 Journal Article
%~ Pubmed
%A Yun, Peter L W
%A Decarlo, Arthur A
%A Hunter, Neil
%T Gingipains of Porphyromonas gingivalis modulate leukocyte adhesion molecule expression induced in human endothelial cells by ligation of CD99.
%B Infection and Immunity
%D 2006
%V 74
%N 3
%P 1661-72
%@ 0019-9567
%X Porphyromonas gingivalis has been implicated as a key etiologic agent in the pathogenesis of destructive chronic periodontitis. Among virulence factors of this organism are cysteine proteinases, or gingipains, that have the capacity to modulate host inflammatory defenses. Intercellular adhesion molecule expression by vascular endothelium represents a crucial process for leukocyte transendothelial migration into inflamed tissue. Ligation of CD99 on endothelial cells was shown to induce expression of endothelial leukocyte adhesion molecule 1, vascular cell adhesion molecule 1, intercellular adhesion molecule 1, and major histocompatibility complex class II molecules and to increase adhesion of leukocytes. CD99 ligation was also found to induce nuclear translocation of NF-kappaB. These results indicate that endothelial cell activation by CD99 ligation may lead to the up-regulation of adhesion molecule expression via NF-kappaB activation. However, pretreatment of endothelial cells with gingipains caused a dose-dependent reduction of adhesion molecule expression and leukocyte adhesion induced by ligation of CD99 on endothelial cells. The data provide evidence that the gingipains can reduce the functional expression of CD99 on endothelial cells, leading indirectly to the disruption of adhesion molecule expression and of leukocyte recruitment to inflammatory foci.
%Z FOR Codes: 110502



%0 Journal Article
%~ Pubmed
%A Ye, P
%A Simonian, M
%A Nadkarni, M A
%A Decarlo, A A
%A Chapple, C C
%A Hunter, N
%T Identification of epithelial auto-antigens associated with periodontal disease.
%B Clinical and Experimental Immunology
%D 2005
%V 139
%N 2
%P 328-37
%@ 0009-9104
%X We previously reported evidence that patients with periodontitis have serum antibodies to oral Gram positive bacteria that are cross-reactive with epithelial antigens. In the present report cross-reactive epithelial antigens including CD24, lactate dehydrogenase A [LDM-A], antioxidant protein 2 [AOP 2] and nuclear factor of activated T cells 5 [NFAT 5], were identified by screening a cDNA expression library with pooled patient sera. Titres of antibodies to CD24 peptide correlated negatively with indices of periodontal disease severity. Strong expression of CD24 in the reactive periodontal epithelium and inflamed gingival attachment contrasted with low to undetectable expression in the external gingival epithelium. In periodontitis, a local action of these auto-reactive antibodies could modulate the regulatory potential associated with expression of CD24 in this epithelium.
%Z FOR Codes: 110508



%0 Journal Article
%~ Pubmed
%A Chhour, Kim-Ly
%A Nadkarni, Mangala A
%A Byun, Roy
%A Martin, F Elizabeth
%A Jacques, Nicholas A
%A Hunter, Neil
%T Molecular analysis of microbial diversity in advanced caries.
%B Journal of Clinical Microbiology
%D 2005
%V 43
%N 2
%P 843-9
%@ 0095-1137
%X Real-time PCR analysis of the total bacterial load in advanced carious lesions has shown that the total load exceeds the number of cultivable bacteria. This suggests that an unresolved complexity exists in bacteria associated with advanced caries. In this report, the profile of the microflora of carious dentine was explored by using DNA extracted from 10 lesions selected on the basis of comparable total microbial load and on the relative abundance of Prevotella spp. Using universal primers for the 16S rRNA gene, PCR amplicons were cloned, and approximately 100 transformants were processed for each lesion. Phylogenetic analysis of 942 edited sequences demonstrated the presence of 75 species or phylotypes in the 10 carious lesions. Up to 31 taxa were represented in each sample. A diverse array of lactobacilli were found to comprise 50% of the species, with prevotellae also abundant, comprising 15% of the species. Other taxa present in a number of lesions or occurring with high abundance included Selenomonas spp., Dialister spp., Fusobacterium nucleatum, Eubacterium spp., members of the Lachnospiraceae family, Olsenella spp., Bifidobacterium spp., Propionibacterium sp., and Pseudoramibacter alactolyticus. The mechanisms by which such diverse patterns of bacteria extend carious lesions, including the aspect of infection of the vital dental pulp, remain unclear.
%Z FOR Codes: 60504



%0 Journal Article
%~ Pubmed
%A Yun, Peter L W
%A Decarlo, Arthur A
%A Chapple, Cheryl C
%A Hunter, Neil
%T Functional implication of the hydrolysis of platelet endothelial cell adhesion molecule 1 (CD31) by gingipains of Porphyromonas gingivalis for the pathology of periodontal disease.
%B Infection and Immunity
%D 2005
%V 73
%N 3
%P 1386-98
%@ 0019-9567
%X Periodontitis is a response of highly vascularized tissues to the adjacent microflora of dental plaque. Progressive disease has been related to consortia of anaerobic bacteria, with the gram-negative organism Porphyromonas gingivalis particularly implicated. The gingipains, comprising a group of cysteine proteinases and associated hemagglutinin domains, are major virulence determinants of this organism. As vascular expression of leukocyte adhesion molecules is a critical determinant of tissue response to microbial challenge, the objective of this study was to determine the capacity of gingipains to modulate the expression and function of these receptors. Given the potential multifunctional role of platelet endothelial cell adhesion molecule 1 (PECAM-1) in the vasculature, the effect of gingipains on PECAM-1 expression by endothelial cells was examined. Activated gingipains preferentially down-regulated PECAM-1 expression on endothelial cells compared with vascular cell adhesion molecule 1 and endothelial-leukocyte adhesion molecule 1, but the reduction in PECAM-1 expression was completely inhibited in the presence of the cysteine proteinase inhibitor TLCK (Nalpha-p-tosyl-l-lysine chloromethyl ketone). Endothelial monolayers treated with activated gingipains demonstrated progressive intercellular gap formation that correlated with reduced intercellular junctional PECAM-1 expression as determined by Western blotting and immunofluorescence microscopy. This was accompanied by enhanced transfer of both albumin and neutrophils across the monolayer. The results suggest that degradation of PECAM-1 by gingipains contributes to increased vascular permeability and neutrophil flux at disease sites.
%Z FOR Codes: 110502



%0 Journal Article
%~ Pubmed
%A Nguyen, Ky-Anh
%A DeCarlo, Arthur A
%A Paramaesvaran, Mayuri
%A Collyer, Charles A
%A Langley, David B
%A Hunter, Neil
%T Humoral responses to Porphyromonas gingivalis gingipain adhesin domains in subjects with chronic periodontitis.
%B Infection and Immunity
%D 2004
%V 72
%N 3
%P 1374-82
%@ 0019-9567
%X The gingipains have been implicated in the pathogenicity of Porphyromonas gingivalis, a major etiologic agent of chronic periodontitis. Mature gingipains often present as a membrane-bound glycosylated proteinase-adhesin complex comprising multiple adhesin domains (HA1 to -4) and a catalytic domain. Using recombinant adhesin domains, we were able to show that patients with chronic periodontitis produce significantly more immunoglobulin G reactive with gingipain domains than a corresponding group with healthy periodontium. Titers were predominantly directed toward the carbohydrate epitopes shared between the gingipains and the lipopolysaccharide of P. gingivalis with little recognition of the peptide backbone of the catalytic domains. Distribution of titers to peptide epitopes of the adhesin domains was as follows: HA4 approximately HA1 > HA3 >> HA2. No correlation was observed between markers of disease severity and titers to individual adhesins within the disease group. Posttreatment titers showed no change or a decrease in titers for the majority of patients except for titers to the HA2 domain which showed marked increases in a few responding patients. Since the HA2 domain is important in hemoglobin binding and acquisition of essential porphyrin, boosting titers of antibodies to this domain may have the potential to control the growth of this organism.
%Z FOR Codes: 110508 60501