bluuebunny/crossref_metadata_2025 huggingface dataset:
Comparison of the pro‐inflammatory cytokine‐stimulating activity of the surface‐associated proteins of periodontopathic bacteria
Authors: Reddi K. , Wilson M. , Nair S. , Poole S. , Henderson B.
Published: July 2006

Saline extraction of the periodontopathic bacterium, Actinobacillus actinomycetemcomitans, releases surface‐associated material (SAM), a complex mixture of proteins and carbohydrates with potent biological actions on isolated bone and on various mammalian cell populations. In this study, the relative ability of the SAM from 5 organisms, implicated in the pathology of periodontal disease, to stimulate human mesenchymal and myelomonocytic cells to synthesize the pro‐inflammatory cytokines ‐ interleukin (IL)‐1β, IL‐6 and tumour necrosis factor (TNF)α has been investigated. The bacteria investigated were Actinobacillus actinomycetemcomitans, Eikenella corrodens, Porphyromonas gingivalis, Prevotella intermedia and Campylobacter rectus. Human cells were exposed to a four log order range of concentrations of the SAM, or of Escherichia coli lipopolysaccharide, to provide full agonist dose responses in order to allow comparison of the potency and efficacy of each SAM, All SAMs demonstrated the capacity to stimulate human gingival fibroblasts (HGFs), human peripheral blood mononuclear cells (PBMCs) or the myelomonocytic cell line ‐ Mono‐Mac‐6 to release one or all of the cytokines assayed. Activity was heat‐ and trypsin‐sensitive suggesting that the active components were proteinaceous. However, there were substantial differences in the potency and efficacy of each SAM when compared on a concentration basis (w/v). The most active SAM was from A. actinomycetemcomitans with those from E. corrodens and P. gingivalis being slightly less active. The least active cytokine‐stimulating SAMs were from C. rectus and Pr. intermedia. One major difference between the SAMs and E. coli LPS was the inability of the former to stimulate HGFs to release IL‐1β or TNFα although they could stimulate PBMCs to release these cytokines. This may have relevance to the pathology of the periodontal diseases.
Fluoride distribution in human dental calculus obtained from different sites on the tooth surface
Authors: Huang S. , Nakagaki H. , Okumura H. , Morita I. , Strong M. , Robinson C. , Pearce E.
Published: July 2006

We examined the site specificity of fluoride (F) distribution in human dental calculus. Teeth with supra‐ and subgingival calculus were obtained from patients who resided in non‐fluoridated areas in Japan and China. Sequential layers of the dental calculus (30 μm thick) were abraded by an abrasive micro‐sampling technique and fluoride and phosphorus in the powdered samples were analyzed. Fluoride concentrations were highest in the outer, lowest in the middle and intermediate in the inner layers of dental calculus in general. In the outermost layers fluoride concentrations were highest in calculus found near the tooth cervix both in supra‐ and subgingival calculus. Fluoride concentrations decreased markedly toward the apical region in subgingival calculus. while it did not change toward the incisal or occlusal region in supragingival calculus. In the inner layers, fluoride concentrations in both supra‐ and subgingival calculus were not affected by position on the teeth. Fluoride concentrations in subgingival calculus near the apex were lower than in supragingival calculus near the incisal or occlusal region. It was concluded that the fluoride concentrations differ in different regions of dental calculus, probably due to their different mechanisms of formation.
Efficacy of quantitative digital subtraction radiography using radiographs exposed in a multicenter trial
Authors: Jeffcoat M. K. , Reddy M. S. , Magnusson I. , Johnson B. , Meredith M. P. , Cavanaugh P. F. , Gerlach R. W.
Published: July 2006

Digital subtraction radiography (DSR) has been shown to be a sensitive and specific method for the detection of small bony changes in periodontitis. The purpose of this study was to perform a multicenter validation of the DSR in human subjects. Seventeen subjects were enrolled at 3 centers. Feather‐edged hydroxyapatite chips (approximately 1,7 and 10 mg) were used to simulate osseous lesions. Bilateral radiographs were taken with and without chips. Geometry was standardized using a cephalostat and the order of radiographs was determined using a randomization plan. Radiographs were subtracted, lesions isolated, and quantified at a single center without knowledge of the randomization code or location of the chips used in each subject. The overall sensitivity and specificity in detecting 1 mg changes was 87.8% and 100%, respectively. Sensitivity and specificity in detecting 7 mg and 10 mg chips was 100%. A strong linear relationship between actual lesion mass and calculated mass was observed (R2=0.94, slope=0.98, p<0.0001). No significant differences were observed by center. These data indicate that the DSR is a valid technique for the assessment of alveolar bone changes in multicenter trials.
Digital image ratio: a new radiographic method for quantifying changes in alveolar bone. Part 1: theory and methodology
Authors: Jean A. , Epelboin Y. , Rimsky A. , Soyer A. , Ouhayoun J. P.
Published: July 2006

A new method, digital image ratio (DIR), has been developed for directly measuring changes in alveolar bone. The image on the computer monitor represents the relative mass change between two radiographs. Fourier filtering is used to reduce noise artefacts. This method is validated through an experiment with a step wedge. DIR needs only a preliminary calibration of the experimental conditions of operation and avoids tedious calibrations for each measurement as in the case of digital image substraction. Low‐voltage X‐ray techniques are suggested for long‐term quantitative studies of patients to minimize irradiation doses.
Protease repertoires of cells adherent to membranes recovered after guided tissue regeneration
Authors: Wakabayashi R. C. , Wong F. , Richards D. W. , Johnson P. W.
Published: July 2006

Guided tissue regeneration is a clinical procedure used to restore mineralized tissue that has been lost to periodontal disease or after tooth extraction. The procedure makes use of Gore‐tex® membranes or Gore‐tex® augmentation membranes (GTAM) to prevent migration of keratinocytes and gingival fibroblasts into healing wounds. To begin to characterize the regenerative cells associated with these membranes, human cells have been rescued from membranes retrieved after bone‐inductive procedures. Cell lines were established from tissue adherent to Gore‐tex® membranes used to regenerate bone around periodontally compromised teeth, and from tissue adherent to GTAM used in edentulous ridge augmentation procedures or in conjunction with implant placement. Cell lines were screened for mineralized nodule formation in vitro prior to their subsequent analysis. All but one of the lines selected for this study formed mineralized nodules in vitro with cells from GTAM tending to form modules more quickly than cells from Gore‐tex®. Zymograms and Western blots were used to compare protease profiles of these cells with those of human gingival fibroblasts, keratinocytes and periodontal ligament (PDL) cells. All cell types except for keratinocytes produced a 72 kD protease. In contrast, keratinocytes were the only cells that produced 92 kD gelatinase. In some cell lines, notably those removed from patients after short periods of regeneration, collagenase was the major protease detected on gelatin substrate gels. Some of these cell lines also produced additional proteases including a low molecular weight protease (30 kD) not seen in gingival fibroblasts, PDL cells or keratinocytes.
Immunocytochemical study of nerve fibers containing substance P in the junctional epithelium of rats
Authors: Tanaka T. , Kido M. A. , Ibuki T. , Yamaza T. , Kondo T. , Nagata E.
Published: July 2006

Nerve fibers with substance P‐like immunoreactivity (SP‐IR) in the junctional epithelium (JE) of 32‐42‐d‐old rats were examined by both light and electron microscopy using the avidin‐biotin‐peroxidase complex method. The density of nerve fibers with SP‐IR was highest in the middle portion of the JE; however, a few fibers were localized in the coronal portion of the JE and close to the enamel surface. Also, rich innervation was found especially in the basal cell layer of the JE. Unmyelinated axons with SP‐IR in the connective tissue underlying the JE were enveloped by Schwann cells but lost their Schwann cell sheath almost completely in the JE. The axons often formed varicosities with SP‐IR as terminals in various areas of the JE. The terminals contained numerous large granular vesicles, small clear vesicles and a few mitochondria, and were surrounded by the cytoplasmic processes of the junctional epithelial cells. These terminals were sometimes located close to neutrophils in the JE; the minimum gap distance between the terminals and the processes of junctional epithelial cells or neutrophils was about 20 nm. A few terminals with SP‐IR came close to the enamel surface, and the minimal distance between the terminals and the enamel surface was about 5 μm. SP‐IR in the nerve terminals in the JE fixed with 0.1% or 0.25% glutaraldehyde was distributed diffusely in the axoplasm or was confined to the granular vesicles. These findings show that substance P is contained in the large granular vesicles in the nerve terminals, and suggest that these terminals may function as modulators of junctional epithelial cells and neutrophils.
Localization of glycosaminoglycans in periodontal ligament during physiological and experimental tooth movement
Authors: Kagayama M. , Sasano Y. , Mizoguchi I. , Kamo N. , Takahashi I. , Mitani H.
Published: July 2006

Localization of chondroitin sulphates in periodontal ligaments (PDL) of rat molar roots during physiological and experimental tooth movement were analysed immunohistochemically with the use of monoclonal antibodies, 3B3 and 2B6, specific to chondroitin 6‐sulphate (CH‐6S) and chondroitin 4‐sulphate/dermatan sulphate(CH‐4S/DS), respectively. The maxillary first molars of experimental animals were forced to move laterally with a 10 g weight by U‐shaped wires for 3 and 7 d. In control animals, 3B3 epitope was seen in the PDL near to the bone surface facing the distal half of roots, which corresponded to the compressive side during physiological tooth movement. Immunoreactivity for 2B6 was weak or negative in the PDL. Both epitopes were present at osteoid, precementum, lacunae and canaliculli of osteocytes and cementocytes. In 3‐d‐treated animals, the early stage of hyalinization characterized with visible cells and fibres was observed in the PDL at the buccal side of the mesial root, which showed intense immunoreactivity for 3B3. Further 3B3 positive area seen in control animals changed its position from the distal to the buccal side of the PDL. Immunoreactivity for 2B6 did not change in the PDL of 3‐d‐treated animals. In 7‐d‐treated animals, the typical hyalinization characterized with no visible cells and fibres was seen in the PDL at the buccal sides of both mesial and disto‐buccal roots, where both epitopes were present at the peripheral part of the tissue. Observation of serial sections suggested that the 3B3‐positive area was present at the peripheral part of the 2B6‐positive area. The present results suggest that the expression of CH‐6S is related to the compressive force in non‐hyalinized and hyalinized PDL, whereas that of CH‐4S/ DS is not influenced by the mechanical stress.
Responses of periodontal nerve terminals to experimentally induced occlusal trauma in rat molars: an immunohistochenmical study using PGP 9.5 antibody
Authors: Sodeyama T. , Maeda T. , Takano Y. , Hara K.
Published: July 2006

The response of periodontal nerves to experimentally induced occlusal trauma in rat molars was assessed by immunohistochemistry for protein gene product 9.5 (PGP 9.5) at light and electron microscopic levels, and by computerized image analysis. The occlusal surface on the left upper first molar of 8‐wk‐old male Wistar rats was raised approximately 1 min under ether anaesthesia. The rats were perfusion‐fixed on d 1, 2, 3, 4, and 7 after bite‐raising and then decalcified for 2–3 wk. Frozen sagittal cryostat sections were stained by the avidin‐biotin complex method. By the second day after bite‐raising many Ruffini endings were swollen and their outline unclear at the light microscopic level. Transmission electron microscopy disclosed PGP 9.5 reaction products within Ruffini endings that had unusually long cytoplasmic projections extending through enlarged slits of the Schwann sheaths and also diffuse extracellular PGP 9.5‐immunoreactivity near the Ruffini endings. From d 2 to 4, thin nerve fibres on the pressure side of the periodontal ligament were orientated irregularly and had a prominent beaded appearance. An increase in beaded nerve terminals occurred at d 2–4 post elevation, and decreased later. These results suggest that occlusal trauma induces specific changes in the distribution and shape of nerve terminals in the periodontal ligament.
Aggravation of gingival inflammatory symptoms during pregnancy associated with the concentration of plasminogen activator inhibitor type 2 (PAI‐2) in gingival fluid
Authors: Kinnby B. , Matsson L. , Åstedt B.
Published: July 2006

Gingival inflammatory symptoms are aggravated during pregnancy. In vitro studies suggest a hormonal influence on the plasminogen activator inhibitor type 2 (PAI‐2), and a disturbed balance of the fibrinolytic system could help to explain pregnancy gingivitis. Gingival crevicular fluid (GCF) was sampled in 14 women in pregnant and post‐pregnant states. The gingival condition was assessed by the gingival index of Løe & Silness (GI) and the amount of bacterial plaque by the plaque index of Silness & Løe (PI). The ratio of sites with gingivitis to sites with bacterial plaque was calculated (G/P‐ratio). Antigen levels of tissue plasminogen activator (t‐PA), urokinase‐type plasminogen activator (u‐PA), plasminogen activator inhibitors type 1 (PAI‐1) and PAI‐2 in GCF were determined with ELISAs and 17 β‐oestradiol and progesterone in serum with radioimmunoassays. For each individual the differences (Δ) in hormone levels and PAs and PAIs between pregnancy and post‐pregnancy were calculated. Based on differences in G/P‐ratio between pregnancy and post‐pregnancy, subgrouping was done into a high‐reacting and a low‐reacting group. For the total group, the mean G/P‐ratio was 2.0 during and 1.2 after pregnancy (p=0.064). A statistically significant correlation between Δprogesterone and ΔPAI‐2 was noted: the higher Δprogesterone, the lower ΔPAI‐2. No other significant correlations between hormone levels and components of the fibrinolytic system were found. For the total group of women, the concentrations of PAI‐2, PAI‐1 and t‐PA were significantly higher during than after pregnancy. The individuals in the high‐reacting group, however, showed a lower or unchanged production of PAI‐2 during pregnancy, while those in the low‐reacting group showed a greatly increased production. The lower inhibitory capacity in terms of a low production of PAI‐2 during pregnancy in women with a higher inflammatory reaction indicates that the components of the fibrinolytic system may be involved in the development of pregnancy gingivitis and implies that PAI‐2 serves as an inhibitor of importance for tissue proteolysis. The present finding contributes to the explanation of pregnancy gingivitis.
Novel polysaccharide capsular serotypes in Porphyromonas gingivalis
Authors: Laine M. L. , Appelmelk B. J. , van Winkelhoff A. J.
Published: July 2006

Recently van Winkelhoff et al. (1) described 3 novel serotypes in virulent Porphyromomas gingivalis strains, which were based on different polysaccharide antigens. These antigens probably represent capsular structures and have been designated K1, K2 and K3. In the present study we report on 3 novel capsular serotypes, which are represented by P. gingivalis strains ATCC 49417, HG 1690 and HG 1691. The strains, isolated from patients with periodontitis, showed obvious encapsulation in wet India ink preparations. Thermostable antigens could be detected in the supernatant fractions of autoclaved cells. These antigens appeared to be negatively charged, sensitive to periodate degradation, and resistant to proteinase K treatment. On the basis of these characteristics we conclude that the antigens are probably extra‐cellular polysaccharides representing a bacterial capsular structure. These K‐antigens did not cross‐react with K1, K2 or K3 immunesera of P. gingivalis, with the exception of the K2 antiserum, which partially recognized K5‐ and K6‐antigens. In contrast, K5 and K6 antisera did not react with the K2‐antigen. After absorbtion of the K2 antiserum with cells of strains HG 1690 (K5) and HG 1691 (K6) cross‐reactivity was no longer present. We propose these novel serotypes to be designated: K4 (ATCC 49417), K5 (HG 1960) and K6 (HG 1691).
Isolation of alkaline phosphatase‐positive gingival fibroblasts from patients with chronic inflammatory periodontal disease
Authors: Abe Tatsuya , Hara Yoshitaka , Abe Yukiko , Aida Yoshitomi , Maeda Katsumasa
Published: July 2006

We have reported recently that increased expression of membrane alkaline phosphatase (ALP) activity is a phenotypical characteristic of gingival fibroblasts located in chronic inflammatory periodontal lesions. To understand the cellular properties of these cells, we isolated ALP‐positive gingival fibroblasts from patients with adult periodontitis and evaluated their proliferative potential. Using an enzymatic digestion procedure, we prepared gingival cell suspensions containing ALP‐positive fibroblasts without affecting their ALP activities. These cell suspensions were then subjected to 1 g sedimentation, followed by allowing cells to adhere to substrata. Using this procedure, 71.9% of isolated cells were ALP‐positive. Dissociation of ALP‐positive fibroblasts and contamination by non‐fibroblastic cells were examined by cytochemical and immunocytochemical analyses. The proliferative capacity of ALP‐positive fibroblasts in culture was assessed by monitoring the proportion of ALP‐positive cells after repeated subculture passages and by labelling DNA‐synthesizing cells with bromodeoxyuridine (BrdU). The proportion of ALP‐positive fibroblasts decreased during cell culture passages without an apparent change in the ALP‐positive phenotype. The percentage of BrdU‐positive cells was significantly lower among ALP‐positive than among ALP‐negative fibroblasts. These results indicate that ALP‐positive fibroblasts in chronic inflammatory periodontal lesions have low growth potential. We suggest that their reduced capacity to grow in vitro reflects a more differentiated state induced under inflammatory conditions in vivo.
Comparative effects of plateletderived growth factor‐BB and insulin‐like growth factor‐I, individually and in combination, on periodontal regeneration in Macaca fascicularis
Authors: Giannobile William V. , Hernandez Rafaél A , Finkelman Richard D. , Ryarr Susan , Kiritsy Christopher P. , D'Andrea Mark , Lynch Samuel E.
Published: July 2006

Platelet‐derived growth factor (PDGF) and insulin‐like growth factor I (IGF‐I) in combination have previously been shown to enhance periodontal regeneration. The objective of this study was to further characterize the biological effects of this combination of growth factors in non‐human primates and compare the effects to those of each growth factor individually. Ligature‐induced periodontitis was initiated in 10 cynomolgus monkeys. After periodontal lesions were established, surgery was performed, and either a methylcellulose gel vehicle or vehicle containing 10 μg each of either PDGF‐BB, IGF‐I or both PDGF‐BB and IGF‐I was applied to exposed root surfaces. Biopsies were taken 4 and 12 wk after treatment and the extent of periodontal regeneration was assessed by histomorphometry. At both 4 and 12 wk vehicle‐treated lesions generally revealed minimal osseous defect fill (ODF) (8.5±2.1% and 14.5±5.7%, respectively) and new attachment (NA) (34.1±5.2% and 26.6±10.5%, respectively). IGF‐I treatment did not significantly alter healing compared to vehicle in any parameter at both 4 and 12 wk. PDGFBB‐treated sites exhibited significant (p<0.05) regeneration of NA (69.6±12.0%) at 12 wk; trends for PDGF‐BB treatment effect were also observed in other parameters at 4 and 12 wk. although these increases were not statistically significant. Treatment with PDGF‐BB/IGF‐I resulted in 21.6±5.1 % and 42.5±8.3% ODF at 4 and 12 wk, respectively, and 64.1±7.7% and 74.6±7.4% NA at 4 and 12 wk, respectively (all significantly greater than vehicle, p<0.05). The results from this study demonstrated that: 1) IGF‐1 alone at the dose tested did not significantly alter periodontal wound healing; 2) PDGF‐BB alone significantly stimulated NA, with trends of effect on other parameters; and 3) the PDGF‐BB/IGF‐I combination resulted in significant increases in NA and ODF above vehicle at both 4 and 12 wk.
Effect of methyl mercaptan on synthesis and degradation of collagen
Authors: Johnson P. , Yaegaki K. , Tonzetich J.
Published: July 2006

Measurements of the conversion of [14C]‐proline to [14C]‐hydroxyproline were employed to assess the effect of methyl mercaptan on intra‐ and extracellular metabolism of collagenous proteins in human gingival fibroblast cultures. Following a 30‐min pulse, 10 ng of methyl mercaptan per ml of 95% air/5% CO2, headspace suppressed collagen synthesis by 39% and increased the intracellular degradation of newly synthesized collagen from 26% to 42%. Parallel cultures assayed for proline transport demonstrated a 29% inhibition of [14C]‐proline uptake. A similar analysis of cultures exposed to methyl mercaptan for 12 h revealed an increase in intracellular degradation (20% control vs. 30% test) and a marked increase in extracellular collagenolysis (4% control vs. 55% test). While pulsing, collagen synthesis was decreased by 39%. Slab gel electrophoresis also demonstrated that treatment with methyl mercaptan caused reductions both in mature a, and α2 chains of type I collagen and in type III procollagen. Identities of the procollagen species were confirmed by pepsin digestion. Reverse transcribed polymerase chain reaction was utilized to compare expression of a1 chains of type I procollagen with type III procollagen and indicated suppression of mRNA synthesis for type III procollagen in cultures exposed to methyl mercaptan.
Sources of error for periodontal probing measurements
Authors: Grossi Sara G. , Dunford Robert G. , Ho Alex , Koch Gary , Machtei Eli E. , Genco Robert J.
Published: July 2006

This study determined the relative contribution to probing measurement reliability of several factors, compared with that of random error. Probing measurements were performed by examiners properly trained and calibrated. A total of 5771 pairs of replicate pocket depth (PD) and relative attachment level (RAL) measurements were performed with the Florida Probe®. A total of 1488 replicate clinical attachment level (CAL) measurements were performed with the North Carolina 15 mm probe. In addition, longitudinal replicate measurements of RAL were performed at 0 and 12 months on 816 sites in 11 patients utilizing the Florida Probe® 20 mm disk probe. Measurement reliability with the Florida Probe® resulted in mean intraexaminer variances of 0.21 and 0.33, for PD and RAL, respectively (s.e.m. 0.46 mm for PD and 0.57 mm for RAL). Measurement reliability with the conventional probe resulted in mean intra‐examiner variances of 0.19 for PD and 0.32 for CAL (s.e.m. 0.44 mm and 0.56 mm). Pocket depth contributed to =5% of the variability of the intra‐examiner variance with both probes with other contributing factors being the individual patient, tooth and site location. Mean intra‐examiner reproducibility for duplicate RAL measurement performed at 0 and 12 months was 0.24 and 0.19, respectively (s.e.m. 0.49 mm and 0.43 mm). In conclusion, a mean intra‐examiner variance of ≤0.24 can be achieved for replicate measurements with both electronic and conventional probes for moderate and severe periodontitis patients. Individual examiner, individual patient and site location contribute up to 10% to the overall variance. Hence, the pattern of variability for intra‐examiner variance of probing measurements performed with either electronic or conventional probes by trained and calibrated examiners is mostly random error.
Influence of conventional forceps extraction and extraction with an extrusion instrument on cementoblast loss and external root resorption of replanted monkey incisors
Authors: Oikarinen Kyösti S. , Stoltze Kaj , Andreasen Jens O.
Published: July 2006

Cementoblast loss and root resorption on the root surface was registered histologically after removal and immediate replantation of maxillary central incisors in monkeys (Cecropithecus aethiops). Incisors were removed with either a conventional extraction technique using forceps or with a special designed extrusion instrument and in both cases teeth were immediately replanted. Altogether 18 monkeys were used for the experiments in which 9 monkeys were sacrificed after 3 d and 9 animals after 8 wk. Histological analysis was made in horizontal sections perpendicular to the long axis of the tooth using either 24 measuring points or 12 sections. Histometric analysis showed extensive cementoblast loss in the periodontal ligament (PDL) and with a maximum of damage occurring on the corner surfaces of the root. Extracted teeth showed significantly more extensive cementoblast loss than extruded teeth. Root resorption was also found in the corner locations and was significantly more frequent in extracted than in extruded teeth. There was a similarity in the distribution of cementoblast loss and root resorption indicating that root resorption develops in the same areas where cementoblast loss takes place, i.e. locations that are more compressed during removal of the tooth and must have caused by mechanical damage to the PDL exerted on the root surface during tooth removal.
Chymotrypsin‐like enzyme secretion is stimulated in cultured epithelial cells during proliferation and in response to Actinobacillus actinomycetemcomitans
Authors: Firth James D. , Sue Evelyn S. , Putnins Edward E. , Oda Dolphine , Uitto Velf‐Jukka
Published: July 2006

A chymotrypsin‐like enzyme was partially purified from culture medium of epithelial cells of human skin, human gingiva and porcine periodontal ligament by aprotinin‐affinity chromatography. The enzyme levels from all three cell types were low in quiescent cultures but increased markedly when the cells were allowed to proliferate. The biphasic elution profile of the enzyme from the affinity column closely matched that of α‐chymotrypsin and the protein comigrated with it on polyacrylamide gels at 27,000 M Synthetic substrate tests of purified fractions showed strong chymotrypsin‐like but no trypsin‐like or elastase‐like activity. Inhibition of protease activity and pH optimum in the range of 7.5–8.0 were consistent with chymotrypsin‐like enzymes. Secreted activity was found to be significantly increased by phorbol myristate acetate treatment in a time‐course that differed from that of elastase‐like activity. Keratinocyte growth factor and epidermal growth factor but not transforming growth factor‐β increased the chymotrypsin‐like activity in a concentration‐dependent manner. The enzyme secretion by epithelial cells was strongly elevated by exposure to 5 of 6 Actinobacillus actinomycetemcomitans strains isolated from plaque samples of juvenile periodontitis patients. These results indicate that chymotrypsin‐like enzymes are secreted by proliferative phenotypes of normal epithelial cells. This enzyme may, therefore, play a role in epithelial physiology and in cell response to certain pathogenic bacteria.
The relationship between gingival crevicular fluid cathepsin B activity and periodontal attachment loss in chronic periodontitis patients: a 2‐year longitudinal study
Authors: Eley B. M. , Cox S. W.
Published: July 2006

This study aims to determine whether gingival crevicular fluid (GCF) cathepsin B levels, total activity (TA) and concentration (EC) predict progressive attachment loss (AL). Seventy‐five previously untreated patients with moderate periodontitis were recruited. GCF was collected from 16 molar and premolar mesiobuccal sites and probing attachment level (PAL) and probing depth (PPD) were measured with an electronic probe. Gingival, gingival bleeding and plaque indices were then scored. Prior to baseline patients were given basic periodontal treatment after which the above procedures were repeated. Carefully localized radiographs were taken of the test teeth and repeated annually. Patients were seen 3‐monthly for 2 yr and the procedures were repeated. One hundred and twenty‐one AL sites, 90 rapid AL (RAL) and 31 gradual AL (GAL), in 49 patients were detected. Cathepsin B levels (TA & EC) at RAL sites were significantly higher (p<0.0001) than paired control sites at the attachment loss time (ALT) and prediction time (PT). Mean levels (TA & EC) over the study period at GAL sites were significantly higher (p<0.0001) than paired control sites. Using a critical value (CV) of 7.5 μU/30 s (TA) and 30 μU /μL (EC) showed a sensitivity of 100% and specificity of 99.83% (TA) and 100% and 99.75%(EC) at both ALT & PT. Mean cathepsin B levels (TA & EC) were significantly higher (p<0.0001) at RAL and GAL sites than nonattachment loss (NAL) sites in AL patients in intrapatient comparisons and mean patient levels were significantly higher (p<0.0001) in AL patients than NAL patients in interpatient comparisons. These results indicate that GCF cathepsin B may serve as a predictor of attachment loss.
Cytokine‐inducing components of periodontopathogenic bacteria
Authors: Wilson Michael , Reddi Krisanavane , Henderson Brian
Published: July 2006

Pro‐inflammatory cytokines such as interleukin (IL)‐1, IL‐6, TL‐8 and tumour necrosis factor (TNF) are believed to be the major pathological mediators of inflammatory diseases ranging from arthritis to the periodontal diseases. The stimuli inducing proinflammatory cytokine induction in the former disease is unclear but in the periodontal diseases it is obvious that the stimulus is the accumulation of bacteria in the subgingival region. As these bacteria do not invade the lesional tissues in large numbers, it is believed that their soluble components or products interact with host tissues to induce cytokine gene transcription. The paradigm is that lipopolysaccharide is the key bacterial component inducing pro‐inflammatory cytokine gene expression. However, over the past decade a growing number of reports on non‐oral bacteria have established that many other bacterial components, as well as secretory products, have the capacity to induce cytokine synthesis. Some of these, such as the protein pneumolysin from Streptococcus pneumoniae, are incredibly potent (in this case inducing cytokine synthesis at femtomolar concentrations). This review surveys the range of bacterial components and products which have been shown to stimulate cytokine synthesis with particular emphasis on the hypothesis that these components play a role in the pathology of the periodontal diseases.
Effects of T cell adoptive transfer into nude mice on alveolar bone resorption induced by endotoxin
Authors: Ukai Takashi , Hara Yoshitaka , Kato Lhachi
Published: July 2006

Using the method of reconstitution of nude mice with T cells, we examined the effects of T cell on alveolar bone resorption induced by repeated injections of Escherichia coli endotoxin into periodontal tissue. Three mice groups (normal, nude and T cell reconstituted nude mice) were used. Endotoxin derived from E. coli was repeatedly injected into the gingiva of the mice left mandibles every 48 h and the mice were killed on the day after the 1st, 4th, 7th, 10th, 13th and 20th injections of endotoxin. Alveolar bone resorption was examined histopathologically and histomorphometrically. Bone surfaces in contact with the osteoclast were defined as the site of active resorption and the ratios of active resorption were compared among the 3 nice groups. Consequently, no active resorption was found after the first injection of endotoxin in any group. After the 4th injection, active resorption was found in normal nice and T cell reconstituted nude mice and gradually rose with the increase in the injection frequency. In contrast, few osteoclasts were found even after the 10th injection in the nude mice. In addition, there were statistically significant differences between the normal price and nude mice after the 4th and 10th injections (p<0.05). These findings suggested that T cell influences periodontal bone destruction induced by local administration of endotoxin during the early phases.
Clonality of Porphy givens intermedia and Preotella nigrescens isolated from periodontally diseased and healthy sites
Authors: Teanpa San R. , Douglas C. W. I. , Eley A. R. , Walsh T. F.
Published: July 2006

Black‐pigmented anaerobes have been implicated as major pathogens in the aetiology of adult periodontitis but these organisms are also found in healthy sites. This study aimed to examine the relationship between genotypes of black‐pigmented anaerobes and disease status of periodontal sites using restriction endonuclease analysis (REA) and ribotyping. The main black‐pigmented species recovered from sites were Pot phyromonas gingivalis, Prevotella intermedia and Prevotella nigrescens. Each of the 58 subjects investigated harboured distinct genotypes of these three species. Most subjects appeared to be colonized by a single genotype of P. gingivalis and Pr. intermedia. whereas multiple types of Pr. nigrescens colonized many individuals. Plasmids were only found in a few Pr. nigrescens strains. No association was found between the disease status of sites and any specific or group of genotypes of either species or presence of a plasmid. Since the same genotypes of P. gingivalis, Pr. intermedia and Pr. nigrescens were found at both diseased and non‐diseased sites in a subject, adult periodontitis is not explained by the presence of specially virulent clones of these organisms. Their role in periodontitis, therefore, is likely to be opportunistic.
The localization of epithelial root sheath cells during cementum formation in rat molars
Authors: Alatli I. , Lundmark C. , Hammarström L.
Published: July 2006

The purpose of this study was to investigate the distribution of epithelial cells and the fate of the basement membrane along the root surface of rat molars during cementogenesis, and to test the hypothesis that the Hertwig's epithelial root sheath (HERS) cells remain on the root surface if mineralization is inhibited. To demonstrate the HERS cells and basement membrane, immunohistochemistry with antibodies against keratin and laminin were used. The dentin matrix mineralization was inhibited by a single injection of 1‐hydroxyethylidene‐1,1‐bisphosphonate (HEBP). A modified Gomori staining method was used to monitor the inhibition of mineral formation in dentin and cementum. Paraffin sections were stained with haematoxylin‐eosin. and freeze‐dried sections were used for Gomori and immunohistochemical stainings. We found that the formation of acellular cementum was suppressed above the dentin with inhibited mineralization. Instead, a hyperplastic matrix, different from acellular cementum, covered the dentin. This hyperplastic cementum had keratin‐ and laminin‐positive cells incorporated; such cells were never incorporated in normal acellular cementum. The later formation of cellular cementum correlated, in controls, with the disappearance of HERS cells from the root surface. Treatment with HEBP resulted in a persistent presence of epithelial cells, interpreted as an inhibition of their disappearance. In conclusion, there is evidence that the cells of HERS are involved in the development of both acellular and cellular cementum. The developmental processes of these tissues appear in some way to be influenced by or associated with the initial mineralization of the dentin.
Extracts of periodontopathic microorganisms lack functional superantigenic activity for murine T cells
Authors: Petit M. D. A. , Stashenko P.
Published: July 2006

Products of periodontopathic bacteria exert immunomodulatory effects on various lymphoid cell populations, some of which have been implicated in the pathogenesis of periodonthis. It has recently been suggested that some of these bacterial products may possess superantigenic (SAg) activity. SAg bind simultaneously to the Vβ chain of T cell receptors and to class II major histocompatibility complex molecules, thereby activating as many as 35% of T cells to proliferate and produce cytokines. In order to examine this question, the proliferation of splenic and thymic T cells from immunologically naive, 3‐6‐wk‐old Balb/c (H‐2d), C57BL/6 (H‐2b) and C3H/HeJ (H‐2k) mice was assessed in response to sonic extracts of periodontopathogens. Laboratory and/or reference strains of a.o. Actinobacillus actinomycetemcomitans, Porphyromonas gingivalis, Prevotella intermedia and Prevotella nigrescens were used as stimulants. Staphylococcal enterotoxin B (SEB), a known superantigen, was utilized as a positive control. Unfractionated spleen cells responded to several of the tested preparations of the different bacteria, as well as to SEB, Con A and Escherichia coli LPS. Thymocytes responded to Con A and SEB, but not to LPS or to any sonic extract. Spleen cells depleted of B cells by panning responded to SEB and Con A, but not to LPS and showed a reduced response to sonicates. The residual response of B cell‐depleted spleen cells was reduced essentially to background by treatment with anti‐Thy 1.2 + C'. Similar results were obtained in the presence of 5% added mitomycin‐treated antigen presenting cells, indicating that these cells were not limiting. These results demonstrate that extracts of periodontopathic bacteria do not stimulate murine T cells in a manner consistent with superantigenic activation.
Use of the polymeric matrix as internal standard for quantitation of in vivo delivery of tetracycline HCI from Actisite® tetracycline fiber during periodontal treatment
Authors: Litch Joyce M. , Encarnacion Myriam , Chen Susan , Leonard Joe , Burkoth Terry L.
Published: July 2006

Actisite® (tetracycline hydrochloride) periodontal fiber is a 23 cm monofilament containing 12.7 mg tetracycline HCI homogeneously dispersed in a polymer. This product sis indicated as an adjunct to scaling and root planing to reduce pocket depth and bleeding on probing in patients with adult periodontitis. The sustained‐release system, placed in the periodontal pocket for 10 d, releases the antibiotic through mechanisms of diffusion and osmosis. A study was conducted in 13 patients with moderate to severe adult periodontitis to evaluate the amount of tetracycline HCI released during therapy (based on residual drug content). Fibers placed in the pocket remained in place for an average of 9 d. Each patient had 1–4 teeth treated with fiber therapy. At the termination of therapy samples from 29 teeth were retrieved and analyzed. The amount of matrix polymer was used as an internal standard for the quantitation of tetracycline, eliminating any uncertainties with respect to recovery or contamination. An average of 31 % (SD 9%) of the tetracycline HCI content was released from the fiber during the treatment period. No single fiber had less than 50% of the original drug remaining. The study demonstrated that a substantial amount of the tetracycline remains in the Actisite® fiber at removal (about 70%), which indicates that substantial drug concentrations are maintained in the pocket for the duration of treatment.
Immunohistochemical localization of transforming growth factor β, basic fibroblast growth factor and heparan sulphate glycosaminoglycan in gingival hyperplasia induced by nifedipine and phenytoin
Authors: Saito K. , Mori S. , Iwakura M. , Sakamoto S.
Published: July 2006

Although drug‐induced gingival hyperplasia has been extensively studied, the pathogenesis of this disorder has not been clarified to date. Transforming growth factorβ (TGFβ) and basic fibroblast growth factor (bFGF) have been shown to be implicated in diverse fibrotic and hyperplastic diseases. Heparan sulphate proteoglycan (HSPG), which is composed of heparan sulphate glycosaminoglycan (HSGAG), has also been shown to play an important role in the pathogenesis of tissue overgrowth by enhancing the functions of bFGF. However, the possible implication of these growth factors in gingival hyperplasia has not been studied. Immunohistochemical localization of TGFβ, bFGF, their receptors and HSGAG was studied in 4 nifedipine‐induced and 5 phenytoin‐induced hyperplastic gingival tissues, and 5 non‐hyperplastic control gingival tissues to elucidate the pathogenesis of this disease. Significant immunostaining against TGFβ, bFGF, the receptors of these two growth factors and HSGAG was observed in the lamina propria of hyperplastic gingival tissues while less immunostaining was observed in the controls. The mean numbers of immunostained cells against TGFβ, bFGF, their receptors in a square unit (0.1 ×0.1 mm) of the lamina propria, which were counted to 10 units of each hyperplastic gingival tissue, were significantly higher than those of the controls. The results suggest that the increased synthesis of TGFβ, bFGF, their receptors and HSGAG may be related to the pathogenesis of drug‐induced gingival hyperplasia.
Lymph and interstitial fluid dynamics in labial gingival tissues of sheep
Authors: Au Binh , Boulton Melfort R. , Narini Phillip P. , McCulloch Christopher A. G. , Hay John B.
Published: July 2006

Lymphatic drainage and circulation in periodontal tissues have been cited as important components of host defence and pathogenic mechanisms, but quantitative data are sparse because of the technical difficulties associated with small animal lymphatic studies. However, the lymphatic vessels draining the periodontal tissues and surrounding region are sufficiently large in sheep to permit surgical placement of lymphatic catheters. Consequently, lymph and recirculating lymphocytes can be continuously collected and this permits the quantitative assessment of local immune responses in these tissues. We have studied the lymphatic drainage pathways from the labial gingival tissues in sheep by two methods. First, in a series of anatomical studies (n=6), a complex of Evan's blue dye and albumin was injected into the labial gingival tissues. One hour after injection the animals were sacrificed and the submandibular and cervical regions were dissected to expose the stained lymphatics. This anatomical study demonstrated 2 major drainage pathways: 1) cervical lymph ducts and; 2) efferent prescapular lymphatics. Secondly, to compare the relative importance of these two drainage pathways, radiolabeled protein (125I‐albumin) was injected directly into the gingival tissues and its appearance in the cervical and prescapular lymph was measured (n=7). Despite the technical difficulties encountered in the experiments, data collected showed that over 7.5 h, 64.7% of the injected protein was recovered in the prescapular and cervical lymph vessels (31.8±6.5% and 32.9±8.5%, respectively). In addition, 11.9±2.1% of the injected protein was transported to the blood by routes not involving the cannulated cervical and prescapular lymph vessels. With most of the remaining radiolabeled protein (17.9±4.9%) recovered from the injection site, we were able to account for approximately 95% of the injected protein. This study suggests that the lymph drainage from this region in the sheep model could provide one of the best described closed and contained systems and thus, could be a useful system for future continous monitoring of inflammatory responses during experimental periodontal diseases.
Role of αβ T cells and γδ T cells in protective immunity to Actinobacillus actinomycetemcomitans in mutant mice by targeted gene disruption
Authors: Takahashi I. , Nakagawa I. , Xu L. , Hamada S.
Published: July 2006

The relative roles of αβ or γδ T cells in protection against Actinobacillus actinomycetemcomitans were analysed by using mutant mice deficient in αβ or γδ T cell receptor (TCR). The dose of A. actinomycetemcomitans producing lethality for the TCR‐α deficient mice was 25% of that for control C57BL/6 mice, while that of TCR‐δ deficient mice was similar to that of C57BL/6 mice. Reverse transcription (RT)‐PCR revealed that γδ T cells from the A. actinomycetemcomitans‐infected αβ TCR deficient mice expressed significant levels of interferon (IFN)‐γ, interleukin (IL)‐5 and IL‐6 mRNA, whereas αβ T cells from A. actinomycetemcomitans‐infected γδ TCR deficient mice exhibited significant levels of IFN‐γ and IL‐2 mRNA. Delayed‐type hypersensitivity (DTH) reactions against heat shock protein (HSP) 60 were prominently observed in A. actinomycetemcomitans‐infected TCR‐δ mutant and C57BL/6 mice, but were absent in TCR‐α mutant mice, suggesting that the DTH response is exclusively dependent on HSP60‐specific αβ T cells producing IFN‐γ and IL‐2 mRNA. It was found that all the mice used exhibited similar levels of serum IgG and IgM responses against A. actinomycetemcomitans‐specific antigens (whole cells and HSP60), suggesting that αβ as well as γδ T cells participate in the serum immune responses. In addition, the humoral antibody responses displayed in the TCR‐α deficient mice implies resistance to A. actinomycetemcomitans infection. Thus, the resistance to A. actinomycetemcomitans infection may be ascribed mainly to αβ T cells, while γδ T cells can partially compensate for the αβ T cell defect.
Biochemical alterations in inflammatory periodontal diseases I. Poly (ADP‐ribose) synthetase activity in gingiva and gingival fibroblasts from humans with periodontitis
Authors: Ghani Q. Perveen , Armitage Gary C. , Hussain M. Zamirul
Published: July 2006

Periodontal diseases are characterized in part by generation of oxygen free radicals, which can cause breaks in cellular DNA strands. Repair of damaged DNA is dependent upon the synthesis of poly(ADP‐ribose)(PADPR) catalyzed by PADPR synthetase, an enzyme known to be activated by the broken ends of DNA strands. We measured the activities of PADPR synthetase and of PADPR glycohydrolase. which degrades PADPRS, in gingival biopsy specimens from 16 sites with adult periodontitis and 12 clinically healthy control sites. The results indicated that sites with periodontitis displayed markedly reduced PADPR synthetase activity compared with healthy control sites, whereas PADPR glycohydrolase activity was not changed. The mean specific activity of PADPR synthetase for the diseased specimens was one‐sixth of that of the healthy specimens (p < 0.001). The PADPR synthetase activity was negatively correlated with the Gingival Index (rs=‐0.60), pocket depth (rs=‐0.70) and bleeding upon probing (rs=‐0.72). Cultured fibroblasts derived from clinically characterized healthy and diseased gingival sites reflected similar patterns of enzyme activity. The mean specific activity of PADPR synthetase for the diseased‐site cultures (n=9) was 56±7% (p < 0.001) of the cultures from healthy control sites (n=6). These results suggest that a reduced level of PADPR synthetase activity is associated with increased inflammation and periodontal destruction, and that the ability to synthesize PADPR is compromised in adult periodontitis.