A RUNX2/PEBP2αA/CBFA1 mutation displaying impaired transactivation and Smad interaction in cleidocranial dysplasia

Cleidocranial dysplasia (CCD), an autosomal-dominant human bone disease, is thought to be caused by heterozygous mutations in runt-related gene 2 (RUNX2)/polyomavirus enhancer binding protein 2αA (PEBP2αA)/core-binding factor A1 (CBFA1). To understand the mechanism underlying the pathogenesis of CCD, we studied a novel mutant of RUNX2, CCDαA376, originally identified in a CCD patient. The nonsense mutation, which resulted in a truncated RUNX2 protein, severely impaired RUNX2 transactivation activity. We show that signal transducers of transforming growth factor β superfamily receptors, Smads, interact with RUNX2 in vivo and in vitro and enhance the transactivation ability of this factor. The truncated RUNX2 protein failed to interact with and respond to Smads and was unable to induce the osteoblast-like phenotype in C2C12 myoblasts on stimulation by bone morphogenetic protein. Therefore, the pathogenesis of CCD may be related to the impaired Smad signaling of transforming growth factor β/bone morphogenetic protein pathways that target the activity of RUNX2 during bone formation.

Phosphorylation and microtubule association of the Opitz syndrome protein mid-1 is regulated by protein phosphatase 2A via binding to the regulatory subunit α4

Opitz syndrome (OS) is a human genetic disease characterized by deformities such as cleft palate that are attributable to defects in embryonic development at the midline. Gene mapping has identified OS mutations within a protein called Mid1. Wild-type Mid1 predominantly colocalizes with microtubules, in contrast to mutant versions of Mid1 that appear clustered in the cytosol. Using yeast two-hybrid screening, we found that the α4-subunit of protein phosphatases 2A/4/6 binds Mid1. Epitope-tagged α4 coimmunoprecipitated endogenous or coexpressed Mid1 from COS7 cells, and this required only the conserved C-terminal region of α4. Localization of Mid1 and α4 was influenced by one another in transiently transfected cells. Mid1 could recruit α4 onto microtubules, and high levels of α4 could displace Mid1 into the cytosol. Metabolic 32P labeling of cells showed that Mid1 is a phosphoprotein, and coexpression of full-length α4 decreased Mid1 phosphorylation, indicative of a functional interaction. Association of green fluorescent protein–Mid1 with microtubules in living cells was perturbed by inhibitors of MAP kinase activation. The conclusion is that Mid1 association with microtubules, which seems important for normal midline development, is regulated by dynamic phosphorylation involving MAP kinase and protein phosphatase that is targeted specifically to Mid1 by α4. Human birth defects may result from environmental or genetic disruption of this regulatory cycle.

The product of the ataxia-telangiectasia group D complementing gene, ATDC, interacts with a protein kinase C substrate and inhibitor.

Ataxia-telangiectasia (AT) is an autosomal recessive human genetic disease characterized by immunological, neurological, and developmental defects and an increased risk of cancer. Cells from individuals with AT show sensitivity to ionizing radiation, elevated recombination, cell cycle abnormalities, and aberrant cytoskeletal organization. The molecular basis of the defect is unknown. A candidate AT gene (ATDC) was isolated on the basis of its ability to complement the ionizing radiation sensitivity of AT group D fibroblasts. Whether ATDC is mutated in any AT patients is not known. We have found that the ATDC protein physically interacts with the intermediate-filament protein vimentin, which is a protein kinase C substrate and colocalizing protein, and with an inhibitor of protein kinase C, hPKCI-1. Indirect immunofluorescence analysis of cultured cells transfected with a plasmid encoding an epitope-tagged ATDC protein localizes the protein to vimentin filaments. We suggest that the ATDC and hPKCI-1 proteins may be components of a signal transduction pathway that is induced by ionizing radiation and mediated by protein kinase C.

Phototaxis away from blue light by an Escherichia coli mutant accumulating protoporphyrin IX.

The hemH gene of Escherichia coli encodes ferrochelatase (EC 4.99.1.1), the enzyme that catalyzes the last step in the production of heme, namely the synthesis of heme from protoporphyrin IX plus Fe2+. The behavioral responses to light were studied in E. coli carrying a hemH mutation. It was shown that the hemH mutant displayed a tumbling response upon illumination and a running response upon removal of the light. The most effect light to induce a tumbling response in the hemH mutant was blue light (396-450 nm). The chemotaxis machinery was needed for the light-induced tumbling response in the hemH mutant. The bacterial defect is an analog of the human inherited disease erythropoietic protoporphyria.

Estudo retrospectivo das afecções orais em 754 felinos domésticos (Felis catus) atendidos no Laboratório de Odontologia Comparada da Universidade de São Paulo

Objetivou-se fazer um estudo retrospectivo avaliando quais as afecções da cavidade oral foram mais frequentes nos gatos domésticos atendidos no Laboratório de Odontologia Comparada da Faculdade de Medicina Veterinária e Zootecnia da Universidade de São Paulo, relatando estatisticamente a prevalência das afecções da cavidade oral de gatos, enfatizando se há correlação entre elas e com características como raça, sexo, faixa etária e estado reprodutivo. Os dados analisados dos 754 prontuários foram raça, idade, sexo, estado reprodutivo, diagnóstico, tratamento e, no caso de neoplasia, sua localização e diagnóstico histopatológico. As principais doenças diagnosticadas foram doença periodontal, fratura dentária, gengivoestomatite crônica felina, lesão de reabsorção dentária felina, neoplasia oral e traumatismo do sistema estomatognático (luxação de articulação temporomandibular, fenda palatina, fratura de processo coronoide, fratura de zigomático, disjunção de sínfise, fratura de maxila e mandíbula). A idade dos animais variou de menos de um ano a 20 anos, sendo que, os animais tinham, em média 7,2 anos (desvio padrão = 4,9) e a faixa etária mais frequente foi de um a cinco anos. Os gatos sem raça definida (66,5%), siameses (19,0%) e persas (10,2%) totalizaram 95,7% de todos os felinos atendidos no LOC. A doença periodontal foi a afecção mais frequente e esteve presente em 38,3% da população estudada. A fratura dentária, segunda mais frequente, esteve presente em 27,2% dos animais. Houve associação estatisticamente significativa (p=0,026) entre fratura dentária e faixa etária, já que a proporção de animais entre um e cinco anos de idade com fratura foi maior do que a das outras faixas etárias. A lesão de reabsorção dentária felina (LRDF) esteve presente em 19,6% dos gatos estudados, sendo a terceira afecção mais prevalente dentre as pesquisadas. Esta lesão foi mais frequente em gatos com idade entre 11 e 15 anos e houve associação estatisticamente significativa entre a LRDF e a doença periodontal e entre LRDF e gengivite. A prevalência de gengivoestomatite crônica felina foi de 15,7% entre os felinos pesquisados e a proporção de animais com idades entre seis e dez anos com esta doença foi maior do que em outras faixas etárias. As neoplasias estavam presentes em 9,8% dos gatos, sendo que em 46 dos 72 animais que apresentaram alguma neoplasia tinham mais de dez anos de idade. O carcinoma de células escamosas foi o neoplasma mais comum, correspondendo a 63,2% das neoformações que foram submetidas ao exame histopatológico. As fraturas ósseas do sistema estomatognático corresponderam a 19,3% dos atendimentos, sendo a sínfise mentoniana e o corpo da mandíbula os locais mais comuns de fraturas. Concluiu-se que: existe grande variedade de afecções que acometem a cavidade oral de gatos, sendo a doença periodontal, fratura dentária, lesão de reabsorção dentária, gengivite, gengivoestomatite crônica, neoplasias orais e fraturas dos ossos do sistema estomatognático as mais prevalentes delas; é de extrema importância que as anotações nas fichas de atendimento sejam feitas da maneira mais completa possível, para que informações não sejam perdidas

Modulation of the antibody response by Porphyromonas gingivalis and Fusobacterium nucleatum in a mouse model

Successive immunization of mice with Fusobacterium nucleatum and Porphyromonas gingivalis has been shown to modulate the specific serum IgG responses to these organisms. The aim of this study was to investigate these antibody responses further by examining the IgG subclasses induced as well as the opsonizing properties of the specific antibodies. Serum samples from BALB/c mice immunized with F. nucleatum (gp1-F), P. gingivalis (gp2-P), P. gingivalis followed by F. nucleatum (gp3-PF) F. nucleatum followed by P. gingivalis (gp4-FP) or saline alone (gp5-S) were examined for specific IgG1 (Th2) and IgG2a (Th1) antibody levels using an ELISA and the opsonizing properties measured using a neutrophil chemiluminescence assay. While IgG1 and IgG2a subclasses were induced in all immunized groups, there was a tendency towards an IgG1 response in mice immunized with P. gingivalis alone, while immunization with F. nucleatum followed by P. gingivalis induced significantly higher anti-P. gingivalis IgG2a levels than IgG1. The maximum light output due to neutrophil phagocytosis of P. gingivalis occurred at 10 min using nonopsonized bacteria. Chemiluminescence was reduced using serum-opsonized P. gingivalis and, in particular, sera from P. gingivalis-immunized mice (gp2-P), with maximum responses occurring at 40 min. In contrast, phagocytosis of immune serum-opsonized F. nucleatum demonstrated peak light output at 10 min, while that of F. nucleatum opsonized with sera from saline injected mice (gp5-S) and control nonopsonized bacteria showed peak responses at 40 min. The lowest phagocytic response occurred using gp4-FP serum-opsonized F. nucleatum. In conclusion, the results of the present study have demonstrated a systemic Th1/Th2 response in mice immunized with P. gingivalis and/or F. nucleatum with a trend towards a Th2 response in P. gingivalis-immunized mice and a significantly increased anti-P. gingivalis IgG2a (Th1) response in mice immunized with F. nucleatum prior to P. gingivalis. Further, the inhibition of neutrophil phagocytosis of immune serum-opsonized P. gingivalis was modulated by the presence of anti-F. nucleatum antibodies, while anti-P. gingivalis antibodies induced an inhibitory effect on the phagocytic response to F. nucleatum.

L-arginine-dependent nitric oxide production of a murine macrophage-like RAW 264.7 cell line stimulated with Porphyromonas gingivalis lipopolysaccharide

The aim of this study was to determine nitric oxide (NO) production of a murine macrophage cell line (RAW 264.7 cells) when stimulated with Porphyromonas gingivalis lipopolysaccharides (Pg-LPS). RAW264.7 cells were incubated with i) various concentrations of Pg-LPS or Salmonella typhosa LPS (St-LPS), ii) Pg-LPS with or without L-arginine and/or N-G-monomethyl-L-arginine (NMMA), an arginine analog or iii) Pg-LPS and interferon-gamma (IFN-gamma) with or without anti-IFN-gamma antibodies or interleukin-10 (IL-10). Tissue culture supernatants were assayed for NO levels after 24 h in culture. NO was not observed in tissue culture supernatants of RAW 264.7 cells following stimulation with Pg-LPS, but was observed after stimulation with St-LPS. Exogenous L-arginine restored the ability of Pg-LPS to induce NO production; however, the increase in NO levels of cells stimulated with Pg-LPS with exogenous L-arginine was abolished by NMMA. IFN-gamma induced independent NO production by Pg-LPS-stimulated macrophages and this stimulatory effect of IFN-gamma could be completely suppressed by anti-IFN-gamma antibodies and IL-10. These results suggest that Pg-LPS is able to stimulate NO production in the RAW264.7 macrophage cell model in an L-arginine-dependent mechanism which is itself independent of the action of IFN-gamma.

Persistent colonization with Tannerella forsythensis and loss of attachment in adolescents

Colonization with Tannerella forsythensis may characterize the conversion of periodontally healthy sites into diseased sites. This three-year study describes the prevalence of T forsythensis and its relationship to clinical loss of attachment (LOA) in a group of adolescents considered at risk of developing early chronic periodontitis. Adolescents with (LOA+) and without (LOA-) loss of attachment were examined at baseline and 1.5 and 3 yrs subsequently. On each occasion, attachment loss was measured on selected teeth, and the presence of T. forsythensis in their subgingival plaque samples was determined by PCR. T. forsythensis prevalence in LOA+ subjects at baseline (64%) increased to 82% and 86% on subsequent examinations. In contrast, prevalence of T. forsythensis in LOA- subjects was always significantly lower (25%, 36%, and 32%, respectively). The odds of loss of attachment were 8.16 times greater in subjects infected with T. forsythensis at each examination. These results suggest that T. forsythensis is strongly associated with loss of attachment in this adolescent population.

Immunohistological analysis of Tannerella forsythia-induced lesions in a murine model

Tannerella forsythia has been implicated as a defined periodontal pathogen. In the present study a mouse model was used to determine the phenotype of leukocytes in the lesions induced by subcutaneous injections of either live (group A) or nonviable (group B) T. forsythia. Control mice (group C) received the vehicle only. Lesions were excised at days 1, 2, 4, and 7. An avidin-biotin immunoperoxidase method was used to stain infiltrating CD4(+) and CD8(+) T cells, CD14(+) macrophages, CD19(+) B cells, and neutrophils. Hematoxylin and eosin sections demonstrated lesions with central necrotic cores surrounded by neutrophils, macrophages and lymphocytes in both group A and group B mice. Lesions from control mice exhibited no or only occasional solitary leukocytes. In both groups A and B, neutrophils were the dominant leukocyte in the lesion 1 day after injection, the numbers decreasing over the 7-day experimental period. There was a relatively low mean percent of CD4(+) and CD8(+) T cells in the lesions and, whereas the percent of CD8(+) T cells remained constant, there was a significant increase in the percent of CD4(+) T cells at day 7. This increase was more evident in group A mice. The mean percent of CD14(+) macrophages and CD19(+) B cells remained low over the experimental period, although there was a significantly higher mean percent of CD19(+) B cells at day 1. In conclusion, the results showed that immunization of mice with live T. forsythia induced a stronger immune response than nonviable organisms. The inflammatory response presented as a nonspecific immune response with evidence of an adaptive (T-cell) response by day 7. Unlike Porphyromonas gingivalis, there was no inhibition of neutrophil migration.

The effects of intra-amniotic injection of periodontopathic lipopolysaccharides in sheep

Objective: Periodontal disease may cause several complications of pregnancy, including fetal death. The purpose of this study was to investigate in sheep the effects of the intra-amniotic injection of lipopolysaccharide from 3 periodontopathic organisms and to compare these effects with those resulting from similar injection of Escherichia coli lipopolysaccharide. The outcomes that were studied included the rates of fetal death and the features of inflammation and lung maturation in survivors. Study design: At 118 days of pregnancy, ewes that were bearing single fetuses were allocated at random to receive intra-amniotic injections of saline solution (n = 13 fetuses), or lipopolysaccharide from Porphyromonas gingivalis (in doses from 0.1 to 10 tug [n = 22 fetuses]), Actinobacillus actinomycetemcomitans (10 mg [n = 6 fetuses]; 1 mg [n = 6 fetuses]), Fusobacterium nucleation (10 mg [n = 6 fetuses]) or Escherichia coli (10 mg [n = 14 fetuses]; 1 mg [n = 7 fetuses]). Surviving fetuses were delivered abdominally at 125 days of gestation (term, 150 days). Results: When compared with Escherichia coli lipopolysaccharide at similar dosages, periodontopathic lipopolysaccharides had high rates of fetal lethality. Only 6 of 22 fetuses that were exposed to intra-amniotic Porphyromonas gingivalis lipopolysaccharide survived doses of 0.1 to 10 mg, and only 3 of 6 fetuses survived 10-mg Actinobacillus actinomycetemcomitans lipopolysaccharide. Escherichia coli lipopolysaccharide did not cause fetal loss when given at doses of 10 mg (n = 14 fetuses) or l mg (n = 7 fetuses). Fetuses that survived exposure to these lipopolysaccharides showed features of inflammation in amniotic fluid and cord blood at birth and enhanced lung maturation. Conclusion: Lipopolysaccharides from these 3 periodontopathic organisms have much higher rates of fetal lethality than Escherichia coli lipopolysaccharide but can cause similar intrauterine inflammatory responses and improvements in lung volumes in survivors. Sources of inflammation that are distant from the uterus may underlie a proportion of unexplained stillbirth and other complications of pregnancy. (c) 2005 Mosby, Inc. All rights reserved.

The distribution of Tannerella forsythia in an adolescent and adult population

Background: The fact that Tannerella forsythia, an important periopathogen, is difficult to cultivate from mixed infections has impeded precise estimates of its distribution within a given population. In order to discern T. forsythia alone from the mixed infection of plaque, the use of sensitive 16S ribosomal RNA based polymerase chain reaction (PCR) detection is necessary. Objectives: The aim of the present study was to determine the distribution of T. forsythia in an adult and in an adolescent population. Materials and methods: Subgingival plaque samples were obtained from 498 Australian adults and from 228 adolescent subjects from Manchester, UK. Tannerella forsythia was detected using PCR and confirmed by restriction analysis. Semi-quantitation of the organisms was carried out using two specific primers of differing sensitivities. Results: In the adolescent population, 25% were found to carry T. forsythia, albeit in relatively low numbers. In the adult population, a total of 37.8% and 11% were found to carry the organism with primer 2 and primer 1, respectively, suggesting that around 27% had between 10(3) and 10(7) organisms. Although there was an apparent increased proportion of T. forsythia positive subjects in those aged >= 50 years, this was not statistical significant. However, T. forsythia positive male smokers showed increased disease severity compared with T. forsythia negative subjects. Conclusion: This study has shown that at least 25% of the adolescent population carry low numbers of T. forsythia, whereas at least 37% of adults carry the organism, with some 11% having relatively high numbers. The relationship between T. forsythia and disease progression in these populations, however, remains to be determined.

Eating disorders and oral health: A review of the literature

This article is a review of the recent literature pertaining to the oral sequelae of eating disorders (EDs). Dentists are recognized as being some of the first health care professionals to whom a previously undiagnosed eating disorder patient (EDP) may present. However, despite the prevalence (up to 4 per cent) of such conditions in teenage girls and young adult females, there is relatively little published in the recent literature regarding the oral sequelae of EDs. This compares unfavourably with the attention given recently in the dental literature to conditions such as diabetes mellitus, which have a similar prevalence in the adult population. The incidence of EDs is increasing and it would be expected that dentists who treat patients in the affected age groups would encounter more individuals exhibiting EDs. Most of the reports in the literature concentrate on the obvious clinical features of dental destruction (perimolysis), parotid swelling and biochemical abnormalities particularly related to salivary and pancreatic amylase. However, there is no consistency in explanation of the oral phenomena and epiphenomena seen in EDs. Many EDPs are nutritionally challenged; there is a relative lack of information pertaining to non-dental, oral lesions associated with nutritional deficiencies.

Periodontitis and rheumatoid arthritis: A review

Periodontitis and rheumatoid arthritis (RA) appear to share many pathologic features. In this review, the common pathologic mechanisms of these two common chronic conditions are explored. Emerging evidence now suggests a strong relationship between the extent and severity of periodontal disease and RA. While this relationship is unlikely to be causal, it is clear that individuals with advanced RA are more likely to experience more significant periodontal problems compared to their non-RA counterparts, and vice versa. A case is made that these two diseases could be very closely related through common underlying dysfunction of fundamental inflammatory mechanisms. The nature of such dysfunction is still unknown. Nonetheless, there is accruing evidence to support the notion that both conditions manifest as a result of an imbalance between proinflammatory and anti-inflammatory cytokines. As a result, new treatment strategies are expected to emerge for both diseases that may target the inhibition of proinflammatory cytokines and destructive proteases. The clinical implications of the current data dictate that patients with RA should be carefully screened for their periodontal status.