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BACKGROUND: The high prevalence of low bone mass in young women in Japan has emerged as a serious health issue in recent years. Therefore, the aim of the present study was to reevaluate the relationship between genetic and dietary factors, as well as its influence on bone mass in young Japanese women, with particular emphasis on vitamin D receptor (VDR) gene polymorphisms and calcium intake. METHODS: A total of 499 Japanese women aged 20-24 years were enrolled in the study. The bone mass of the calcaneus was assessed using the quantitative ultrasound method and expressed as the osteo sono-assessment index (OSI). VDR gene polymorphisms (BsmI, TaqI, ApaI, and FokI) were analyzed using DNA extracted from saliva. Calcium intake was assessed using the Food Frequency Questionnaire based on food groups (FFQg) and adjusted with the energy intake. Participants were divided into two groups based on the median calcium intake (250 mg/1000 kcal). RESULTS: Consequently, bone mass was significantly different among the BsmI and TaqI genotypes after adjusting for body mass index (BMI) (p = 0.030 and 0.019, respectively). In addition, the BsmI AA and ApaI GT genotypes showed significant differences in bone mass between the calcium-intake groups, with low OSI in the low-calcium intake group and high OSI in the high-calcium intake group, respectively, even after adjusting for BMI (p = 0.020 and 0.038, respectively). CONCLUSIONS: These findings may prove instrumental in developing a logical approach towards preventing bone loss in young Japanese women.
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Cálcio , Receptores de Calcitriol , Densidade Óssea/genética , Feminino , Predisposição Genética para Doença , Genótipo , Humanos , Japão , Polimorfismo Genético , Receptores de Calcitriol/genéticaRESUMO
Resolution of dysbiosis following treatment for periodontal disease and tobacco dependence has been reported in longitudinal intervention studies. In the present report, we evaluated the biological findings regarding the effect of smoking on the periodontal microbiome. A standardized electronic search was conducted using MEDLINE; overall, 1099 papers were extracted. Studies that addressed the relationship between tobacco and periodontal pathogens were included. Finally, 42 papers were deemed appropriate for the present review. Functional changes in periodontal pathogens exposed to nicotine and cigarette smoke extract support the clinical findings regarding dysbiosis of the subgingival microbiome. Dysbiosis of the periodontal microbiome was presented in smokers regardless of their periodontal condition (healthy, gingivitis, or periodontitis) and remained significant only in smokers even after the resolution of experimentally-induced gingivitis and following reduction of clinical signs of periodontitis with non-surgical periodontal treatment and over 3 months post-therapy. Based on these findings, smoking cessation in periodontitis patients is beneficial for promoting a health-compatible subgingival microbial community. To maximize the benefits of these interventions in dental settings, further studies on periodontal microbiome are needed to elucidate the impact of tobacco intervention on preventing recurrence of periodontal destruction in the susceptible subjects.
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Cellulitis accompanied by gas gangrene is a rapidly-spreading and potentially fatal infection. Here, we present a case of gas gangrene in the deep spaces of the head and neck in an elderly woman, diagnosed by computed tomography (CT). An 86-year-old woman with Alzheimer's disease, hypertension, hyperlipidemia, and osteoporosis was referred to our institute by her local dentist. The patient exhibited trismus caused by severe swelling in the left submandibular area. CT images of the head and neck area showed swelling of the cervical tissue with air in the parapharyngeal and masticator spaces. She was treated with antibiotics, followed by drainage. Although the therapy was continued, the patient died from a cardiac complication on hospital day 42. Our case highlights the usefulness of CT for diagnosing gas gangrene in the deep spaces of the head and neck in a woman with Alzheimer's disease.
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Gangrena Gasosa , Tomografia Computadorizada por Raios X , Idoso de 80 Anos ou mais , Celulite (Flegmão)/diagnóstico por imagem , Feminino , Gangrena Gasosa/diagnóstico por imagem , Humanos , Boca/diagnóstico por imagemRESUMO
Kampo medicine is a medical system that has been systematically organized based on the reactions of the human body. At present, in Western, herbal medicines means the whole herbal product. It is being called Kampo medicine as a medicinal drug in Japan. Since 2012, the "National Health Insurance Drug Price Standards Related to Dental Treatment" published by the Japan Dental Association have included seven Kampo formulations. In 2015, the Japan Dental Association sent a "Kampo Education Plan for Dentistry" to all dental universities in Japan. Furthermore, the Japanese Society of Oral Therapeutics and Pharmacology compiled a summary of "Evidence for Kampo Treatment in the Field of Oral Surgery." In addition, the phrase "including wakan-yaku" was included in the draft core model curriculum for dental education in 2016. Thus, Kampo medicine is expected to rapidly spread to the field of dental care and dental medical education. Therefore, the training of dentists with knowledge of both Western and Oriental medicine is required for the treatment of oral pain, periodontal disease, stomatitis, xerostomia, and other complaints concerning oral health. It is our hope that this paper provides a footing for dentists who wish to learn about Kampo medicine and incorporate it into clinical practice.
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OBJECTIVE: Dental plaque is a causative factor for oral disease and a potential reservoir for respiratory infection in the elderly. Therefore, there is a critical need for the development of effective methods to remove oral biofilm. The objective of this study was to investigate the effect of proteases on oral biofilm formation andremoval. DESIGN: The in vivo effect of actinidin, a cysteine protease, on the removal of tongue coating was assessed after orally taking a protease tablet. Effects of the proteases trypsin, papain and actinidin on Actinomyces monospecies biofilm and multispecies biofilm that was reconstructed using a plaque sample from the tongue coating were investigated using the microtiter plate method. Antimicrobial tests and limited proteolysis of fimbrial shaft proteins were also performed to clarify underlying mechanisms of oral biofilm removal. RESULTS: Tablets containing actinidin removed tongue coating in elderly subjects. Oral Actinomyces biofilm was significantly reduced by the proteases papain, actinidin and trypsin. Papain and trypsin effectively digested the major fimbrial proteins, FimP and FimA, from Actinomyces. Actinidin, papain and trypsin reduced multispecies biofilm that was reconstructed in vitro. Papain and trypsin inhibited formation of multispecies biofilm in vitro. CONCLUSIONS: This study shows that proteases reduced oral biofilm in vivo in elderly subjects and in vitro, and suggests that protease digests fimbriae and inhibits biofilm formation.
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Actinomyces/efeitos dos fármacos , Biofilmes/efeitos dos fármacos , Cisteína Endopeptidases/farmacologia , Papaína/farmacologia , Peptídeo Hidrolases/farmacologia , Língua/microbiologia , Tripsina/farmacologia , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Estudos Cross-Over , Método Duplo-Cego , Feminino , Humanos , Masculino , Pessoa de Meia-IdadeRESUMO
Here, we present the complete genome sequence of Actinomyces naeslundii strain ATCC 27039, isolated from an abdominal wound abscess. This strain is genetically transformable and will thus provide valuable information related to its crucial role in oral multispecies biofilm development.
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Here, we present the complete genome sequence of Rothia aeria type strain JCM 11412, isolated from air in the Russian space laboratory Mir. Recently, there has been an increasing number of reports on infections caused by R. aeria The genomic information will enable researchers to identify the pathogenicity of this organism.
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Histatins are salivary proteins with antimicrobial activities. We previously reported that histatin 3 binds to heat shock cognate protein 70 (HSC70), which is constitutively expressed, and induces DNA synthesis stimulation and promotes human gingival fibroblast (HGF) survival. However, the underlying mechanisms of histatin 3 remain largely unknown. Here, we found that the KRHH sequence of histatin 3 at the amino acid positions 5-8 was essential for enhancing p27(Kip1) (a cyclin-dependent kinase inhibitor) binding to HSC70 that occurred in a dose-dependent manner; histatin 3 enhanced the binding between p27(Kip1) and HSC70 during the G1/S transition of HGFs as opposed to histatin 3-M(5-8) (substitution of KRHH for EEDD in histatin 3). Histatin 3, but not histatin 3-M(5-8), stimulated DNA synthesis and promoted HGF survival. Histatin 3 dose-dependently enhanced both p27(Kip1) and HSC70 ubiquitination, whereas histatin 3-M(5-8) did not. These findings provide further evidence that histatin 3 may be involved in the regulation of cell proliferation, particularly during G1/S transition, via the ubiquitin-proteasome system of p27(Kip1) and HSC70.
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Proliferação de Células/fisiologia , Inibidor de Quinase Dependente de Ciclina p27/metabolismo , Proteínas de Choque Térmico HSC70/metabolismo , Histatinas/metabolismo , Proteínas e Peptídeos Salivares/metabolismo , Ubiquitina/metabolismo , Sítios de Ligação , Sobrevivência Celular/fisiologia , Células HEK293 , Humanos , Ligação Proteica , Ubiquitinação/fisiologiaRESUMO
BACKGROUND: Salivary histatins are bioactive peptides related to the innate immune system associated with antimicrobial activities. However, very little is known about the physiological and biological functions of histatins against host cells or their role in oral cell inflammation. Histatin 3 binds to heat shock cognate protein 70 (HSC70, a constitutively expressed heat shock protein (HSP)). It is unclear whether HSC70 is involved in the inflammatory response in oral cells. Injured oral cells release some intracellular proteins including HSC70. It is possible that released HSC70 induces toll-like receptor (TLR) activation, just as extracellular HSP70 (a stress inducible HSP) does, and that histatin 3 affects this process. Therefore, we tested the hypothesis that HSC70 activates TLR signaling and histatin 3 inhibits this activation and inflammatory cytokine production. METHODS: A nuclear factor (NF)-κB-dependent luciferase reporter plasmid was transfected into HEK293 cells stably expressing TLR2 with coreceptor CD14 (293-TLR2/CD14 cells) or stably expressing TLR4 with CD14 and the accessory molecule MD2 (293-TLR4/MD2-CD14 cells). The cells were stimulated with HSC70 in the presence or absence of histatin 3, and examined using luciferase assays. We also stimulated human gingival fibroblasts (HGFs) with HSC70 with or without histatin 3. Then, we analyzed the levels of inflammatory cytokines (interleukin (IL)-6 and IL-8) in the culture media. Cell proteins were analyzed using enzyme-linked immunosorbent assay and Western blotting with antibodies of mitogen-activated protein kinases and NF-κB inhibitor IκB-α, respectively. Histatin 3-bound form of HSC70 was analyzed using limited V8 protease proteolysis. RESULTS: HSC70 induced NF-κB activation in a dose-dependent manner in 293-TLR2/CD14 and 293-TLR4/MD2-CD14 cells, and histatin 3 inhibited this process and when histatin 3 binding to HSC70 was precluded by 15-deoxyspergualin, which augmented NF-κB-triggered activation. In HGFs, histatin 3 also inhibited HSC70-induced inflammatory cytokine production, extracellular signal-regulated protein kinase phosphorylation, and degradation of IκB-α. Moreover, HSC70 in the presence of histatin 3 was relatively resistant to digestion by V8 protease compared with HSC70 in the presence of control peptide. CONCLUSIONS: Histatin 3 may be an inhibitor of HSC70-triggered activation of TLR signaling and inflammatory cytokine production and may be involved in inflammation processes noted in oral cells.
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Periodontal disease as a biofilm infectious disease is considered. Periodontal disease-associated bacteria formed biofilm in periodontal pockets or on the surface of cementum. Planktonic bacteria from biofilm invade into periodontal tissues and lead to inflammation and destruction of tissues directly and indirectly by eliciting the host defense mechanism. Supragingival dental plaques (biofilm) are easily removed by professional mechanical tooth cleaning, while subgingival dental plaques and bacteria invading into periodontal tissues are difficult to remove. Therefore, the development of a method for periodontal disease based on the concept that regards periodontal disease as a biofilm infectious disease is needed. Hereby, I report the effect of antibiotics on an in vitro biofilm model of periodontal disease and the systemic administration of azithromycin for early-onset (aggressive) periodontitis like a treatment resistant periodontitis.
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Antibacterianos/administração & dosagem , Azitromicina/administração & dosagem , Infecções Bacterianas , Biofilmes , Periodontite Crônica/tratamento farmacológico , Periodontite Crônica/microbiologia , Adulto , Feminino , HumanosRESUMO
In the present study, we investigated the effects of a Kampo medicine Orento (TJ-120) on the production of prostaglandin E(2) (PGE(2)), interleukin (IL)-6 and IL-8 by human gingival fibroblasts (HGFs) treated with lipopolysaccharide from Porphyromonas gingivalis (PgLPS). HGFs proliferation was dose-dependently decreased with Orento at days 3 and 7. However, treatment with PgLPS (10 ng/ml), Orento (up to 1 mg/ml) and their combinations for 24 h did not affect the viability of HGFs. Orento suppressed PgLPS-induced PGE(2) production in a dose-dependent manner but did not alter basal PGE(2) level. In contrast, Orento did not alter PgLPS-induced IL-6 and IL-8 productions. These alterations by Orento were similar to those by a mitogen-activated protein kinase kinase (MAPKK/MEK) inhibitor, PD98059. A Orento showed no effect on cyclooxygenase (COX)-1 and COX-2 activities, and increased cytoplasmic phospholipase A(2) (cPLA(2)) expression and increased PgLPS-induced COX-2 expression. Orento suppressed PgLPS-induced mobility retardation of cPLA(2) band on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) gels, that is cPLA(2) phosphorylation and its activation, while Orento alone did not alter cPLA(2) phosphorylation. Orento suppressed PgLPS-induced extracellular signal-regulated kinase (ERK) phosphorylation, which is known to lead to ERK activation and cPLA(2) phosphorylation. These results suggest that Orento decreased PGE(2) production by inhibition of cPLA(2) phosphorylation and its activation via inhibition of ERK phosphorylation, and also that Orento may be useful to improve gingival inflammation in periodontal disease.
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Anti-Inflamatórios/farmacologia , Dinoprostona/biossíntese , Medicamentos de Ervas Chinesas/farmacologia , Gengiva/efeitos dos fármacos , Interleucinas/biossíntese , Magnoliopsida , Fitoterapia , Acetiltransferases/metabolismo , Anti-Inflamatórios/uso terapêutico , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Medicamentos de Ervas Chinesas/uso terapêutico , Eletroforese em Gel de Poliacrilamida , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Flavonoides , Gengiva/metabolismo , Humanos , Inflamação/induzido quimicamente , Inflamação/tratamento farmacológico , Inflamação/metabolismo , Lipopolissacarídeos , Medicina Kampo , Doenças Periodontais/tratamento farmacológico , Fosforilação , Fatores de Transcrição/metabolismoRESUMO
Chronic periodontitis is a widespread and major dental disease. Recent studies have analyzed a possible relationship between polymorphism of several genes and periodontitis. Histatins are salivary polypeptides with fungicidal activities against Candida albicans and yeast and bactericidal activities against Porphyromonas gingivalis and Streptococcus mutans. Histatins are part of the innate defense of the oral cavity. We examined the frequency of the polymorphism codon 23 of the histatin 3 gene (HIS2 allele) in relation to periodontitis in the Japanese population. The subjects were 143 Japanese individuals, of which 63 were healthy control subjects and 80 were periodontal patients. We isolated genomic DNA from lingual mucosal cells and tested them for single nucleotide polymorphism (SNP) by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). The incidence of polymorphism was analyzed statistically by Fisher's exact test. The results indicated that the gene polymorphism at codon 23 of the histatin 3 gene was not associated with periodontitis in the Japanese population (p = 0.166). Rather, if at all, it appeared to be associated with resistance to periodontitis.
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Histatinas/genética , Periodontite/genética , Polimorfismo Genético/genética , Adulto , Idoso , Alelos , Códon/genética , DNA/análise , Feminino , Frequência do Gene/genética , Genótipo , Humanos , Imunidade Inata/genética , Japão , Masculino , Pessoa de Meia-Idade , Mutação/genética , Reação em Cadeia da Polimerase , Polimorfismo de Fragmento de Restrição/genética , Polimorfismo de Nucleotídeo Único/genética , Adulto JovemRESUMO
OBJECTIVE: Macrolide antibiotics are reported to modulate the production of cytokines in various type of cells. We examined the effect of macrolide antibiotics on inflammatory cytokines (IL-6 and IL-8) and chemical mediator (PGE(2)) and also matrix metalloproteinases (MMPs) productions by human gingival fibroblasts (HGFs) treated with lipopolysaccharide (LPS). METHODS: The effect of macrolide antibiotics [erythromycin (EM), azithromycin (AZM) and josamycin (JOM)] on HGFs proliferation were examined by MTT assay. HGFs were treated with LPS from Porphyromonas gingivalis (PgLPS) and macrolide antibiotics, and IL-6, IL-8 and PGE(2) levels were evaluated by ELISA. MMPs were detected by gelatin zymography. RESULTS: AZM slightly but significantly decreased HGFs proliferation, while EM and JOM did not affected. AZM increased PgLPS-induced IL-8 production dose-dependently, while AZM did not alter IL-6 and PGE2 productions. EM and JOM did not altered PgLPS-induced IL-6, IL-8 and PGE(2) productions. All macrolide antibiotics did not alter MMPs production. These results indicate that macrolide antibiotics have no direct anti-inflammatory effect. However, the use of the inhibitors of cell signaling pathway failed to reveal the mechanism that AZM enhanced PgLPS-induced IL-8 production. CONCLUSION: These results suggest macrolide antibiotics have an indirect anti-inflammatory effect as a result of their antimicrobial properties. Because AZM increased LPS-induced IL-8 production by HGFs, the possibility is considered that neutrophils may be migrated to periodontal tissue and phagocytize the periodontopathic bacteria more efficiently.
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Azitromicina/farmacologia , Fibroblastos/efeitos dos fármacos , Interleucina-8/biossíntese , Lipopolissacarídeos/farmacologia , Antibacterianos/farmacologia , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Dinoprostona/biossíntese , Fibroblastos/citologia , Fibroblastos/metabolismo , Gengiva/citologia , Humanos , Interleucina-6/biossíntese , Metaloproteinase 2 da Matriz/metabolismo , Metaloproteinase 9 da Matriz/metabolismo , Fatores de TempoRESUMO
BACKGROUND AND OBJECTIVE: A major factor in the pathogenesis of periodontal disease, which is one of the biofilm infectious diseases, is thought to be lipopolysaccharide (LPS), owing to its ability to cause inflammation and promote tissue destruction. Moreover, the elimination of pathogens and their component LPSs is essential for the successful treatment of periodontal disease. Lipopolysaccharide tolerance is a mechanism that prevents excessive and prolonged responses of monocytes and macrophages to LPS. Since persistence of inflammation is necessary for inflammatory cytokine production, cells other than monocytes and macrophages are thought to maintain the production of cytokines in the presence of LPS. In this study, we investigated whether human gingival fibroblasts (HGFs), the most abundant structural cell in periodontal tissue, might be able to maintain inflammatory cytokine production in the presence of LPS bynot displaying LPS tolerance. MATERIAL AND METHODS: Human gingival fibroblasts were pretreated with LPS (from Porphyromonas gingivalis and Escherichia coli) and then treated with LPS, and the amounts of interleukin (IL)-6 and IL-8 in the cell culture supernatants were measured. The expression of negative regulators of LPS signalling (suppressor of cytokine signalling-1, interleukin-1 receptor-associated-kinase M and SH2 domain-containing inositol-5-phosphatase-1) was also examined in LPS-treated HGFs. RESULTS: Human gingival fibroblasts did not display LPS tolerance but maintained production of IL-6 and IL-8 when pretreated with LPS, followed by secondary LPS treatment. Lipopolysaccharide-treated HGFs did not express negative regulators. CONCLUSION: These results demonstrate that HGFs do not show LPS tolerance and suggest that this characteristic of HGFs sustains the inflammatory response in the presence of virulence factors.
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Fibroblastos/patologia , Gengiva/patologia , Interleucina-6/análise , Interleucina-8/análise , Periodontite/patologia , Actinas/análise , Linhagem Celular , Células Cultivadas , Escherichia coli/imunologia , Fibroblastos/imunologia , Gengiva/imunologia , Humanos , Tolerância Imunológica/imunologia , Inositol Polifosfato 5-Fosfatases , Quinases Associadas a Receptores de Interleucina-1/análise , Interleucina-10/farmacologia , Lipopolissacarídeos/imunologia , Periodontite/imunologia , Monoéster Fosfórico Hidrolases/análise , Porphyromonas gingivalis/imunologia , Pele/imunologia , Pele/patologia , Proteína 1 Supressora da Sinalização de Citocina , Proteínas Supressoras da Sinalização de Citocina/análise , Fator de Crescimento Transformador beta1/farmacologia , Domínios de Homologia de src/imunologiaRESUMO
Histatins, a family of salivary proteins, have antimicrobial activity. Candida albicans, which is killed by histatins, induces oral candidiasis in individuals with compromised immune systems. Although the functional significance of histatins has been documented, their biological and physiological functions against host cells have not been clarified. In this study, we found that histatin 3, a member of the histatin family, binds to heat shock cognate protein 70 (HSC70). These proteins were co-localized in the cytoplasm and nucleus in human gingival fibroblasts following non-heat and heat shock. Histatin 3 induced stimulation of DNA synthesis and cell survival in human gingival fibroblasts in a dose-dependent manner. This DNA synthesis was found to be dependent on HSC70 by knockdown experiments. The effect of heat shock on DNA synthesis induced by histatin 3 was approximately 2-fold higher than that of non-heat shock. When the histatin 3 uptake into cells was inhibited by monodansylcadaverine or when histatin 3 binding to HSC70 was precluded by 15-deoxyspergualin, DNA synthesis by histatin 3 was approximately 2-fold less than that without monodansylcadaverine or 15-deoxyspergualin. Although HSC70 directly bound to p27(Kip1) (a cyclin-dependent kinase inhibitor), histatin 3 increased the binding between those proteins but not with a peptide capable of binding to HSC70. Moreover histatin 3 prevented ATP-dependent dissociation of HSC70-p27(Kip1). ATP was unable to form a histatin 3-HSC70(D10N)-p27(Kip1) complex (HSC70(D10N) is a mutant attenuating ATPase activity). These findings suggest that histatin 3 may be involved in cell proliferation through the regulation of HSC70 and p27(Kip1) in oral cells.
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Fibroblastos/metabolismo , Fase G1 , Gengiva/citologia , Proteínas de Choque Térmico HSC70/metabolismo , Histatinas/metabolismo , Fase S , Proteínas e Peptídeos Salivares/metabolismo , Trifosfato de Adenosina/farmacologia , Sobrevivência Celular/efeitos dos fármacos , Inibidor de Quinase Dependente de Ciclina p27/metabolismo , DNA/biossíntese , Fibroblastos/citologia , Fibroblastos/efeitos dos fármacos , Fase G1/efeitos dos fármacos , Técnicas de Silenciamento de Genes , Guanidinas/farmacologia , Proteínas de Choque Térmico HSC70/química , Resposta ao Choque Térmico/efeitos dos fármacos , Humanos , Ligação Proteica/efeitos dos fármacos , Estrutura Terciária de Proteína , Transporte Proteico/efeitos dos fármacos , Fase S/efeitos dos fármacosRESUMO
Histatins are salivary proteins found and expressed in human salivary glands. They play a role in the non-immune system of antimicrobial defense, for instance, against Candida albicans. The transcriptional regulatory sequences of the histatin gene, HIS1, have remained obscure for a long time. Here, we cloned the putative promoter from human genomic DNA and tested it in a luciferase reporter system. This promoter is much more active in salivary gland cells than in other cell types. Analysis of deletion mutants revealed that the region encompassing -2254 to -1748 is a strong positive transcriptional element, and its functional core sequence (termed HTN27 box) works in correct and reverse orientations in synergy with downstream sequences, the region spanning -680 to +28 and a proximal promoter. The plus single-stranded HTN27 box is specifically bound by a 100 kDa protein that is present in HSG cells, but not in HeLa cells. These findings indicate that the regulation of the histatin gene expression may be intricate, and it seems to have a cell-type preference in the salivary gland cells.
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Regulação da Expressão Gênica , Histatinas/genética , Regiões Promotoras Genéticas/genética , Saliva/metabolismo , Transcrição Gênica , Animais , Sequência de Bases , Células COS , Chlorocebus aethiops , Clonagem Molecular , Perfilação da Expressão Gênica , Células HeLa , Humanos , Camundongos , Dados de Sequência Molecular , Peso Molecular , Células NIH 3T3 , Proteínas Nucleares/metabolismo , Nucleotídeos/genética , Especificidade de Órgãos , Ligação Proteica , Deleção de SequênciaRESUMO
Periodontitis, which is a widespread and major dental disease, is a multifactorial, lifestyle-related disease and has been analyzed for gene polymorphism. We examined the frequency of the polymorphisms of the pro-inflammatory cytokine genes IL-1 A (-889) and IL-1 B (+3953) in relation to periodontitis in the Japanese population. We also examined whether polymorphism of TLR2 (Arg677Trp) and TLR4 (Asp299Gly), which are receptors recognized by periodontopathic bacteria, may also be associated with periodontitis. The subjects were 92 Japanese individuals, among whom 43 had periodontitis and 49 were healthy controls. We isolated genomic DNA from lingual mucosal cells and tested them for single nucleotide polymorphisms by polymerase chain reaction-restriction fragment length polymorphism. The incidence of polymorphisms was analyzed statistically by Fisher's exact test, and the sensitivity and specificity of the gene polymorphisms were calculated. The purpose was to determine whether such polymorphisms might be effectively used in the diagnosis of periodontitis. However, we found no evidence that the gene polymorphism of IL-1A (p = 0.082), IL-1 B (p = 0.180), TLR2 (p = 1.000) or TLR4 (p = 1.000) and overall gene polymorphism in any of the genes (p = 0.752) correlate with periodontitis. The sensitivity (14.0%) and specificity (83.7%) of the mutations found in all of the genes were low. Therefore, we advise against using the analyses of polymorphism of these genes to detect periodontitis in the Japanese population.
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Interleucina-1alfa/genética , Interleucina-1beta/genética , Periodontite/imunologia , Polimorfismo Genético/genética , Receptor 2 Toll-Like/genética , Receptor 4 Toll-Like/genética , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Arginina/genética , Ácido Aspártico/genética , Glicina/genética , Humanos , Japão , Pessoa de Meia-Idade , Mutação/genética , Periodontite/genética , Periodontite/microbiologia , Reação em Cadeia da Polimerase , Polimorfismo de Fragmento de Restrição , Polimorfismo de Nucleotídeo Único/genética , Sensibilidade e Especificidade , Triptofano/genéticaRESUMO
In the present study, we investigated the anti-inflammatory effects of a Kampo medicine Shosaikoto (TJ-9) using in vitro periodontal disease model, in which human gingival fibroblasts (HGFs) treated with lipopolysaccharide (LPS) from Porphyromonas gingivalis (PgLPS) produce IL-6, IL-8 and prostaglandin E2 (PGE2). Treatment with PgLPS (10 ng/ml), TJ-9 (up to 1 mg/ml) and their combinations for 24 h did not affect the viability of HGFs. Moreover, TJ-9 did not alter LPS-induced IL-6 and IL-8 productions. However, TJ-9 significantly suppressed LPS-induced PGE2 production in a dose-dependent manner but TJ-9 alone did not affect basal PGE2 level. Western blotting demonstrated that TJ-9 decreased cyclooxygenase-2 (COX-2) expression in a dose-dependent manner but not phospholipase A2. Moreover, TJ-9 selectively and dose-dependently inhibited COX-2 activity. These results suggest that TJ-9 decreased PGE2 production by inhibition of both COX-2 expression and activity and that TJ-9 may be useful to improve gingival inflammation in periodontal disease.
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Anti-Inflamatórios não Esteroides , Medicamentos de Ervas Chinesas/farmacologia , Fibroblastos/efeitos dos fármacos , Gengiva/citologia , Gengiva/efeitos dos fármacos , Lipopolissacarídeos/antagonistas & inibidores , Lipopolissacarídeos/toxicidade , Medicina Kampo , Western Blotting , Sobrevivência Celular/efeitos dos fármacos , Ciclo-Oxigenase 1/metabolismo , Ciclo-Oxigenase 2/metabolismo , Dinoprostona/biossíntese , Relação Dose-Resposta a Droga , Ensaio de Imunoadsorção Enzimática , Gengivite/tratamento farmacológico , Gengivite/patologia , Humanos , Indicadores e Reagentes , Interleucina-6/biossíntese , Interleucina-8/biossíntese , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Monócitos/efeitos dos fármacos , Monócitos/metabolismo , Porphyromonas gingivalis/química , Sais de Tetrazólio , TiazóisRESUMO
Calcium (Ca2+) antagonists induce gingival overgrowth as a side effect but the pathogenic mechanism is still unknown. The Ca2+-channel activator Bay K 8644 elevates intracellular Ca2+ concentration ([Ca2+]i) and enhances the cell proliferation of gingival fibroblasts in a dose-dependent manner. Verapamil, an L-type Ca2+-channel blocker, also elevates [Ca2+]i in gingival fibroblasts, but it has no effect on other fibroblasts such as those of the lung, skin, and muscle. Moreover, verapamil enhances the proliferation of fibroblasts of the gingiva but has no effect on the proliferation of those of other tissues. These findings confirm that [Ca2+]i elevation induces the proliferation of gingival fibroblasts.
Assuntos
Sinalização do Cálcio/fisiologia , Cálcio/antagonistas & inibidores , Cálcio/metabolismo , Citosol/metabolismo , Gengiva/citologia , Gengiva/crescimento & desenvolvimento , Éster Metílico do Ácido 3-Piridinacarboxílico, 1,4-Di-Hidro-2,6-Dimetil-5-Nitro-4-(2-(Trifluormetil)fenil)/farmacologia , Agonistas dos Canais de Cálcio/farmacologia , Bloqueadores dos Canais de Cálcio/farmacologia , Linhagem Celular , Proliferação de Células , Fibroblastos/fisiologia , Corantes Fluorescentes , Fura-2 , Humanos , Verapamil/farmacologiaRESUMO
PURPOSE: To investigate whether genomic DNA can be purified in sufficient quantity and quality from the oral cavity. METHODS: One milliliter of peripheral blood and saliva were collected. The buccal and lingual mucosal cells were also obtained using 10 strokes with a swab or a toothbrush, respectively. All materials were centrifuged and the cells were lysed by adding sodium dodecyl-sulfate and proteinase K. The DNAs were extracted with phenol and precipitated with ethanol followed by electrophoresing on 0.8% agarose gel. The purified DNAs were digested with restriction enzyme Dpn I and Mbo I, respectively. Amplification of the IL-1A gene by PCR was carried out using the purified DNAs and electrophoresing on polyacrylamide gel. RESULTS: DNA was obtained from lingual mucosal cells collected with a toothbrush. Only about one-thirtieth of the recovered DNA was of non-human origin (bacterial contaminants from the oral cavity). Judging from the PCR amplifications of the IL-1A gene, the DNA extracted from lingual cells was of sufficient quality, in all respects indistinguishable from the DNAs extracted from the other specimen, such as peripheral blood, saliva and buccal mucosal cells collected with a swab, and in sufficient quantity. Our results indicate that it is possible to purify DNAs from lingual mucosal cells collected with a toothbrush in a simple and safe manner. Compared to DNA samples from patients by blood extraction, the described method also had the advantage of being painless and not inducing mental distress.