Clock gene expression in the submandibular glands

Clock genes, which mediate molecular circadian rhythms, are expressed in a circadian fashion in the suprachiasmatic nucleus and in various peripheral tissues. To establish a molecular basis for circadian regulation in the salivary glands, we examined expression profiles of clock-related genes and salivary gland-characteristic genes. Clock-related genes-including Per1, Per2, Cry1, Bmal1, Dec1, Dec2, Dbp, and Reverbalpha-showed robust circadian expression rhythms in the submandibular glands in 12:12-hour light-dark conditions. In addition, a robust circadian rhythm was observed in amylase 1 mRNA levels, whereas the expression of other salivary-gland-characteristic genes examined was not rhythmic. The Clock mutation resulted in increased or decreased mRNA levels of Per2, Bmal1, Dec1, Dec2, and Dbp, and in Cry1-/- background, Cry2 disruption also increased or decreased mRNA levels of these clock-related genes and the amylase 1 gene. These findings indicate that the Clock- and Cry-dependent molecular clock system is active in the salivary glands.     

Gene expression in the developing rat mandible: a gene array study

To analyse the molecular events that occur in the developing mandible, gene arrays containing probes for 1176 known genes were used. Total RNA was extracted from the mandibles of 1- and 6-day-old rats. Radiolabeled probes were then synthesised and used to probe the DNA arrays. Of the 1176 genes examined, 306 were detectable, and the latter were analyzed by bioinformatics algorithms. K-means clustering grouped 72 genes into Up, 56 genes into Down and 178 genes into NO change. A large number of genes related to cell receptors (by ligands) were grouped into the Down cluster. Gene array technology appears to be a useful tool for studying the complex process of mandibular development.     

Cytokine gene expression in chronic periodontitis

Background: Cytokines play an important rôle in controlling inflammatory processes and tissue homeostasis. Periodontitis, as any other chronic inflammatory disease, results from a disarrangement of host factors, mainly cytokines and the initiating agent. Modulation of the cytokines is not only controlled by the host but also by infecting bacteria and their products. Aim: In the present study, we examined the cytokine mRNA expression profiles in six patients, each presenting sites affected with (1) severe progressive periodontitis, (2) chronic, but stable periodontal lesions, and (3) with healthy sites. Analysis using a quantitative RT‐PCR included IFN‐γ, IL‐1β, IL‐2, IL‐4, IL‐5, IL‐6, and TNF‐α. Material and Methods: 6 patients with chronic periodontitis were following treatment observed for a period of six years for local sites staying healthy, local sites with periodontal pathology but without signs of progression of attachment loss and sites with verified progression were biosied. The biopsies were lyzed and analyzed for levels of cytokine mRNAs. Results: Results revealed considerable variation not only between patients, but also between individual sites. Each patient’s site has thus to be looked at as an independent entity. Conclusions: The local action of cytokines, which is heavily dependent on recruitment, interaction and activation of immunocompetent cells can explain the site‐specific nature of cytokine expression. Cytokine data from individual sites together with the local clinical status and data from the literature demonstrate the complexity of periodontal disease pathogenesis. To gain insight to specific mechanisms further studies are needed.     

Sugars and beyond. The role of sugars and the other nutrients and their potential impact on caries

The traditional concept of caries as a multifactorial transmittable and infectious disease has been challenged. Novel conceptual ideas have come to add to the complexity of this highly prevalent disease worldwide. Current etiological understanding of the disease has emphasized the pivotal role of sugars in caries. In fact, current definition points toward an ecological disease caused by the commensal microbiota that under ecological imbalances, mainly due to high and or frequent sugars consumption, creates a state of dysbiosis in the dental biofilm. This modern conceptual idea, however, tends to underrate a key issue. As humans are omnivore and consume a mix diet composed by a multitude of substances, the role of the diet in caries must not be restricted only to the presence of fermentable sugars. This review explores the contribution of other food components, ubiquitous to the diet, mostly as potentially protective factors. Anticaries nutrients might determine an environmental change, affecting the ecology of the oral microbiome and partially mitigating the effect of sugars. Understanding the function of the food usually consumed by the people will contribute new knowledge on the mechanisms associated with the onset of caries, on new caries risk variables and on potential novel strategies for the prevention and treatment of the disease.     

c-Jun调控成釉细胞Amelotin基因表达的研究

目的:通过过表达及knockdown c-Jun基因,观察小鼠成釉细胞中釉成熟蛋白(amelotin)的表达变化,以初步确定c-Jun对amelotin的调控作用。方法:①应用免疫组化和细胞免疫荧光方法观察体内外成釉细胞中的c-Jun及amelotin的表达情况;②RT-PCR获得c-Jun基因,克隆至pcDNA3.1/myc-hisA,瞬时转染成釉细胞,real time RT-PCR观察其对amelotin表达的影响;③化学合成c-Jun siRNA,瞬时转染成釉细胞,re-al time RT-PCR观察其对amelotin表达的影响。结果:①体内外成釉细胞中c-Jun和amelotin均有表达,c-Jun主要在成釉细胞胞核中表达,amelotin在成釉细胞和釉质全层均有表达;②成功构建c-Jun真核重组表达载体pcDNA3.1/myc-hisA-c-Jun,转染成釉细胞后amelotin mRNA表达水平显著上升;③成功沉默c-Jun,amelotin在转染c-Jun siRNA组表达水平显著下降。结论:在成釉细胞中,c-Jun在转录水平调控amelotin基因的表达。     

Epigenetic gene regulation by histone demethylases: emerging role in oncogenesis and inflammation

Histone N‐terminal tails of nucleosomes are the sites of complex regulation of gene expression through post‐translational modifications. Among these modifications, histone methylation had long been associated with permanent gene inactivation until the discovery of Lys‐specific demethylase (LSD1), which is responsible for dynamic gene regulation. There are more than 30 members of the Lys demethylase (KDM) family, and with exception of LSD1 and LSD2, all other KDMs possess the Jumonji C (JmjC) domain exhibiting demethylase activity and require unique cofactors, for example, Fe(II) and α‐ketoglutarate. These cofactors have been targeted when devising KDM inhibitors, which may yield therapeutic benefit. KDMs and their counterpart Lys methyltransferases (KMTs) regulate multiple biological processes, including oncogenesis and inflammation. KDMs’ functional interactions with retinoblastoma (Rb) and E2 factor (E2F) target promoters illustrate their regulatory role in cell cycle progression and oncogenesis. Recent findings also demonstrate the control of inflammation and immune functions by KDMs, such as KDM6B that regulates the pro‐inflammatory gene expression and CD4⁺ T helper (Th) cell lineage determination. This review will highlight the mechanisms by which KDMs and KMTs regulate the target gene expression and how epigenetic mechanisms may be applied to our understanding of oral inflammation.     

体外培养人牙周膜细胞成骨基因表达谱的检测

目的:探讨体外培养人牙周膜细胞(human periodontal ligament cells,hPDL)成骨相关基因的表达谱,明确体外培养人牙周膜细胞的成骨潜能。方法:消化离心法收集体外培养人牙周膜细胞,提取总RNA,使用Super-array公司的点样数96点的人骨再生基因表达谱芯片检测人牙周膜细胞成骨相关基因的表达。结果:体外培养人牙周膜细胞有15种成骨相关基因无表达,81种基因有表达,其中表达较高的基因22种。结论:体外培养人牙周膜细胞具有部分成骨细胞基因表型特征,但成熟成骨细胞的特征基因表达量较低,提示人牙周膜细胞是具有成骨细胞分化潜能的成纤维细胞。     

TIP30蛋白在涎腺腺样囊性癌高转移细胞中的表达鉴定及作用

目的筛选含有Tip30的涎腺腺样囊性癌高转移细胞，检测TIP30蛋白在涎腺腺样囊性癌高转移细胞中的表达，检测携带外源性TIP30细胞的细胞周期分布变化。方法应用脂质体转染方法将TIP30导入ACC-M细胞中，G418筛选阳性克隆，用免疫印记杂交法(Western blot)检测转染细胞中TIP30蛋白的表达；流式细胞仪测定细胞周期分布。结果筛选14天后，含有TIP30的ACC-M单克隆形成，TIP30蛋白在ACC-M中表达稳定；携带外源性TIP30的细胞G0-G1期比例增高，G2-M、S期细胞比例降低。结论ACC-M细胞能稳定表达外源基因TIP30，携带外源性TIP30的细胞G0-G1期细胞比例增高，G2-M、S期细胞比例降低，从而影响ACC-M细胞的生长，是TIP30抑制涎腺腺样囊性癌细胞生长的可能机制之一。     

牙龈卟啉单胞菌菌毛fimA基因在大肠杆菌的融合表达

目的：克隆牙龈卟啉单胞菌菌毛蛋白fimA基因,并使其在大肠杆菌中融合表达。方法：利用PCR方法克隆fimA,并用基因重组融合表达技术获得在大肠杆菌中的表达。结果：克隆基因测序结果与GenBank数据库中的序列呈现99．9％同源性;经诱导表达后可观察到Mr38kDa的融合蛋白。结论：成功克隆了fimA基因,并在大肠杆菌中表达了FimA蛋白,为后续研究奠定了基础。     

牙龈卟啉菌牙龈蛋白酶K基因真核表达载体的构建和体外表达

目的：构建牙龈卟啉菌牙龈蛋白酶K基因的真核表达载体，并检测其在体外真核细胞中的表达情况。方法：利用基因重组技术构建牙龈卟啉菌牙龈蛋白酶K基因重组质粒pcDNA3.1( )-KGPcd，然后通过脂质体将pcDNA3.1( )-KGPcd瞬时转染COS7细胞，以RT-PcR和间接免疫荧光法检测重组质粒的转录水平和蛋白表达情况。结果：成功构建了牙龈蛋白酶K真核表达载体；转染的COS7细胞中可检测到目的蛋白的转录和表达。结论：成功构建了pcDNA3.1( )－KGPcd真核表达载体并在哺乳动物细胞中能够正确转录和表达，为下一步作为基因疫苗免疫动物提供了依据。     

Influence of testosterone on gene expression in the ovariectomized mouse submandibular gland

Androgens exert significant effects on the murine submandibular gland. Our objective in this study was to determine the nature and extent of testosterone regulation of gene expression in the female submandibular gland, and to explore the degree to which this control is the same as in male glands. Ovariectomized female BALB/c mice were treated with placebo- or testosterone-containing hormone pellets for 14 d. Glands were collected and total RNA was isolated. Samples were analyzed for differential expression of mRNA using CodeLink microarrays, and the data were evaluated using genesifter. Testosterone significantly influenced the expression of over 500 genes, and while many (n = 214) of the genes were similarly differentially expressed in androgen-treated males, there were also many that were unique. These findings support our hypotheses that testosterone extensively influences gene expression in the female submandibular gland, and that the nature of this influence is variable between sexes.