定量研究碳酸氢钠和氟化钠混合液对牙本质小管的堵塞作用

目的：用激光荧光仪（KaVo DIALNOdent）定量研究碳酸氢钠和氟化钠混合液对牙本质小管的堵塞作用。方法：分别使用1mol/L碳酸氢钠液和0．12mol/L氟化钠液混合液、1mol/L碳酸氢钠液、0．12mol/L氟化钠液浸泡牙本质面，记录三组牙本质面激光荧光峰值，比较用药前后激光荧光峰值均数的变化。结果：三组用药前后激光荧光峰值均数的变化均有显著差异（P＜0．05），其中混合液组与另两溶液组比较，激光荧光峰值均数的变化有显著差异（P＜0．05），而两溶液组之间比较则无显著差异（P＞0．05）。结论：1mol/L碳酸氢钠液和0．12mol/L氟化钠液混合液，对牙本质小管有较好的堵塞作用。     

An immunohistochemical study of the effects of pulsed neodymium:yttrium-aluminium-garnet laser irradiation in root canals on the eruption of rat incisors

The incisors of 21 Wistar rats were transected, pulp tissue was extirpated for 10mm from the level of the gingival margin and each canal was prepared with files. The fibre tip of a pulsed neodymium:yttrium-aluminium-garnet laser was inserted into the root canal for 10mm and laser irradiation delivered at 2 W and 20 pulses/s for 10s. After 6 weeks the mandibles were removed and sectioned. Sections were stained either with haematoxylin and eosin or immunohistochemically using polyclonal antibodies against keratin/cytokeratin, amelogenin and type I collagen. The inner epithelial cells on the labial side differentiated into ameloblasts in animals where eruption had recovered. The pulp cells differentiated into odontoblast-like cells and staining for type I collagen was evident in pulp cells, odontoblast-like cells and inside dentinal tubules. In animals where eruption had ceased, the inner epithelial cells on the labial side did not differentiate into ameloblasts. Staining for type I collagen was observed in the mineralized nodules and tubules of dentine-like hard tissues in the pulp cavity. These results suggest that differentiation of epithelial cells on the labial side into ameloblasts is involved in the re-eruption process.     

Maspin基因甲基化对口腔黏膜鳞癌细胞的调控作用

目的：分析口腔黏膜鳞癌细胞增殖过程中Maspin基因甲基化的调控作用机制,为相关研究及临床治疗提供借鉴。方法：鳞状细胞癌HIOEC-B(a)P细胞系随机分为4组：5-氮-2-脱氧胞苷+曲古抑菌素A(ADC+TSA)阴性组、低(0.1 μmol/L +0.05 μmol/L)、中(1 μmol/L+0.5 μmol/L)和高剂量组(10 μmol/L+5 μmol/L)。以正常口腔黏膜上皮细胞作为对照组。实时定量PCR检测各组细胞Maspin基因的甲基化程度,MTT法检测各组细胞的增殖情况,4',6-二脒基-2-苯基吲哚（DAPI）染色检测细胞凋亡。采用SPSS20.0软件包对数据进行单因素方差分析。结果：与对照组相比,癌细胞的Maspin基因甲基化程度显著增强（P<0.01）,与阴性组相比,中剂量和高剂量组Maspin基因甲基化程度显著减弱（P<0.05,P<0.01）,低、中、高剂量组的细胞增殖抑制率显著高于阴性组（P<0.05,P<0.01）。随着药物剂量的提高,细胞凋亡程度逐渐增加（P<0.05）。结论：Maspin基因甲基化可能参与口腔黏膜鳞癌细胞的增殖过程。     

小分子鹰嘴豆芽素A对人牙髓干细胞成骨分化的影响

目的:探讨不同浓度鹰嘴豆芽素A (biochanin A,BCA)对人牙髓干细胞(human dental pulp stem cells,hDPSCs)成骨分化的影响及相关分子机制.方法:通过组织块法分离培养原代人牙髓干细胞,流式细胞术鉴定其细胞表型.通过CCK-8法检测不同浓度BCA对hDPSCs增殖活性的影响,通...     

体外培养的冠髓和根髓细胞中碱性磷酸酶的活性

目的：比较根部和冠部牙髓培养细胞的碱性磷酸酶活性，探讨牙髓干细胞在牙髓中的定位。方法：用组织块法分别培养根髓、冠髓细胞，用Gomori碱性磷酸酶染色法对根、冠髓培养细胞在加入条件孵育液(20％DMEM 10nmol／L地塞米松 10mmol/L β-甘油酸钠 50mg／L L-抗坏血酸)后的第5、10、15天的碱性磷酸酶活性进行测定，从牙髓干细胞的功能角度，探讨其在牙髓中的定位。结果：根髓、冠髓细胞均出现碱性磷酸酶染色阳性，染色强度在第5天无差别，在第10、15天根髓染色强度大于冠髓。结论：碱性磷酸酶的活性在根髓中大于冠髓。牙髓干细胞可能存在于全部牙髓之中．其密度在根髓中大于冠髓。     

碳酸氢钠溶液对早期龋再矿化作用的定量分析

目的　探讨碳酸氢钠溶液对早期光滑面釉质龋再矿化作用的影响。方法　在人牙釉质标本上形成早期龋后将标本分组 ,分别用 0 .12mol/LNaF再矿化液、1mol/LNaHCO3 再矿化液、0 .12mol/LNaF与 1mol/LNaHCO3 混合再矿化液及双蒸水对照组中浸泡。KaVoDIAGNOdent激光荧光仪测定釉质表面荧光峰值 ,比较实验前后荧光峰值均数的变化。结果　 3组实验组再矿化前后荧光峰值均数变化差异均有显著性 (P <0 .0 1) ,对照组处理前后差异无显著性 (P >0 .0 5 ) ;实验组中混合矿化液组与另两组比较荧光峰值变化差异有显著性 (P <0 .0 1) ,而 0 .12mol/LNaF组与 1mol/LNaHCO3 组之间比较差异无显著性。结论　碳酸氢钠溶液可促进早期龋的再矿化 ,且能加强氟化钠对早期龋的再矿化作用     

Bacterial invasion of dentinal tubules beneath apparently intact but hypomineralized enamel in molar teeth with molar incisor hypomineralization

Background. The most common problems for a patient with molar incisor hypomineralization (MIH) are the collapse of enamel and cavitations, loss of fillings, and secondary caries, but most of all, severe hypersensitivity. Objective. The aim of this paper was therefore to histologically study possible bacterial invasion of dentinal tubules beneath apparently intact, but hypomineralized enamel in permanent molars with MIH. Material and methods. Five extracted permanent first molars diagnosed with MIH were fixated, demineralized, and sagittally serially sectioned in a bucco-lingual direction in a microtome with a thickness of 4–5 µm. Sections were stained with a modified Brown and Benn staining for bacteria, unstained sections were analysed in field emission SEM. Results. Stained sections from the cuspal areas, below the hypomineralized enamel, the staining indicated the presence of bacteria in the dentinal tubules. The HTX staining showed that the pulp in sections without any findings was normal and free from bacteria or infiltrates from inflammatory cells. In sections where bacteria were found in the cuspal areas or deeper in the dentin, a zone of reparative dentin was found, and in sections from one tooth, the coronal pulp showed an inflammatory reaction with inflammatory cells. In sections adjacent to those without any bacterial staining, the SEM analyses revealed empty dentinal tubules without any odontoblast processes or signs of bacteria. When odontoblast processes were found, the dentinal tubules were filled with bacteria located on the surface of the odontoblast processes. In some areas, a large number of tubules were found with bacteria. No bacteria were found close to the pulp. The odontoblast processes appeared larger in areas where bacteria were found. Conclusions. The presence of bacteria in the dentinal tubules and inflammatory reactions in the pulp indicate that oral bacteria may penetrate through the hypomineralized enamel into the dentin, thus possibly contribute to hypersensitivity of teeth with MIH.     

组蛋白去甲基化酶Jmjd3对牙髓干细胞成牙本质向分化的影响研究

目的研究组蛋白去甲基化酶Jmjd3对牙髓干细胞(dental pulp stem cells,DPSCs)成牙本质向分化的影响。方法原代分离培养DPSCs,用流式细胞术和茜素红染色鉴定DPSCs。体外诱导DPSCs成牙本质向分化不同时间(0、3、5、7、14 d),qRT-PCR检测Jmjd3及成牙本质细胞标志物牙本质涎磷蛋白(dentin sialophosphoprotein,DSPP)、牙本质基质蛋白1(dentin matrix protein1,DMP1)的表达情况。用不同浓度(0、1、10μmol/L)的Jmjd3抑制剂GSK-J4处理DPSCs 14 d,q RT-PCR检测DSPP、DMP1的表达情况。结果原代培养的DPSCs表达间充质干细胞表面标记物CD44、CD29、CD146,体外诱导培养具有成骨分化潜能。DPSCs成牙本质向分化过程中,Jmjd3、DSPP、DMP1表达水平均上调(P<0.05),且可能具有时间依赖性。10μmol/L GSK-J4处理DPSCs可明显抑制DSPP、DMP1的表达(P<0.05)。结论Jmjd3在DPSCs成牙本质向分化过程中发挥正调节作用,且与其去甲基化酶活性相关。     

The effect of tissue molecules on bacterial invasion of dentine

Bacterial invasion of dentinal tubules is a critical step in the pathogenesis of dental caries and pulp and periapical disease. The purpose of this study was to determine the effect some molecules commonly found in saliva and dentinal tubule fluid may have on the bacterial invasion of dentine. The results showed that invasion of Streptococcus gordonii or Enterococcus faecalis cells was inhibited when the bacterial cells were in solution with mucin, immunoglobulin G (IgG) and serum, and this was related to bacterial cell aggregation, as a result of interaction with agglutinins, and/or inhibition of collagen binding. When dentine was soaked in growth media containing fibrinogen, IgG, albumin or serum prior to inoculation, bacterial invasion was inhibited. It is suggested that this may be due to reduced dentine permeability as a consequence of the deposition of the compounds within dentinal tubules.     

Expression of fibronectin and type I collagen by human dental pulp cells and gingiva fibroblasts grown on fibronectin substrate

Specific antibodies and indirect immunoperoxidase labelling were used to study the intracellular production of collagen and fibronectin by cells grown on fibronectin-coated glass; the same cell populations seeded on uncoated glass were used as controls. Strong intracellular staining for type I collagen was seen in all cases, but immunostaining for fibronectin was very faint or negative in both gingival and pulp cells grown on the fibronectin substrate, in contrast to control cells. Thus, fibronectin substrate inhibited fibronectin synthesis by the cultured cells, but did not seem to influence type I collagen synthesis.     

表没食子儿茶素没食子酸酯对牙源性角化囊肿上皮细胞增殖和凋亡的影响

目的: 探讨表没食子儿茶素没食子酸酯（EGCG）对牙源性角化囊肿（OKC）上皮细胞增殖、凋亡的影响,以及与Wnt信号通路相关基因FZD3和JNK3表达的关系。方法: 采用原代培养的人牙源性角化囊肿上皮细胞,分别用不同浓度的EGCG处理24、48、72 h,CCK-8法检测细胞增殖,FITC-Annexin V/PI染色法检测细胞凋亡,流式细胞术检测细胞周期分布,RT-PCR、Western 免疫印迹检测Wnt信号通路相关基因FZD3和JNK3的表达。采用SPSS 22.0软件包对数据进行统计学分析。结果: EGCG以剂量-时间依赖方式抑制OKC上皮细胞增殖。低浓度EGCG（0~20 μmol/L）对细胞增殖无显著影响（P>0.05）,高浓度EGCG（40~320 μmol/L）对细胞增殖有显著抑制作用（P<0.05）,且随着药物浓度增加及作用时间延长,抑制作用逐渐增强（P<0.05）。EGCG (20、160、320μmol/L)诱导OKC上皮细胞凋亡,呈剂量依赖性（P<0.05）,并将细胞周期阻滞于G1期。EGCG可下调Wnt信号通路相关基因FZD3和JNK3的表达（P<0.05）。结论: EGCG抑制OKC上皮细胞增殖,诱导细胞凋亡,其机制可能与阻断Wnt信号通路相关基因FZD3和JNK3的表达有关。     

Accumulation of advanced glycation end-products in human dentine

Cross-linking of collagen by Advanced Glycation End-products (AGEs) occurs by non-enzymatic glycation (Maillard reaction). The purpose of this study was to examine whether AGEs are formed in human dentinal collagen, and to consider any possible influence of AGEs on dentinal physiology. Mechanical characteristics, fluorescence spectra and immunohistochemical analyses of demineralized dentine sections from young subjects were compared with those of aged ones. The same investigations were performed with young dentine artificially glycated by incubation in 0.1 M ribose solution. Indentation measurement indicated that the sections from aged dentine were mechanically harder than those from young dentine. The hardness of young dentine increased after incubation in ribose solution. Fluorescence peak wavelength of the young dentine was shorter than that of the aged one, but shifted towards the peak wavelength of the aged one after incubation in ribose solution. These changes were considered to be due to accumulation of AGEs. Existence of AGEs in dentinal collagen was confirmed by immunohistochemical analysis. The obtained results suggest that AGEs accumulation occurs in dentinal collagen and is affected by both human age and physiological conditions such as glucose level in blood because dentinal collagen receives nourishment via dental pulp and tubules.     

淫羊藿苷对内毒素抑制牙周膜细胞ALP表达的影响

目的：探讨淫羊藿苷对人牙周膜细胞（periodontal ligament cells,PDLC）增殖及对内毒素（Lipopolysaccha-ride,LPS）干扰下碱性磷酸酶（alkaline phosphatase,ALP）的影响。方法：原代培养PDLC,MTT法检测不同浓度（0、10^-5～10^-9mol／L）淫羊藿苷对PDLC增殖的影响;RT—PCR、对硝基苯酚法测定淫羊藿苷对LPS抑制PDLC的ALPmRNA表达和分泌的影响。结果：淫羊藿苷在一定浓度下（10^-6～10^-7mol／L）可促进PDLC增殖;10υg／mLLPS可抑制PDLC的ALP活性;加入10^-6mol／L淫羊藿苷干扰后,对LPS抑制PDLC的ALP有拮抗作用,可提高其mRNA表达和活性。结论：淫羊藿苷在一定浓度时可促进PDLC增殖;可能通过拮抗LPS抑制其ALP的活性。     

A light and electron microscopic study of the early stages of root surface formation in molar teeth in the rat

The forming root surfaces of first molar teeth of twelve-day old rats were examined by light and transmission electron microscopy. A thin layer of the outermost dentinal matrix, approx. 1.0–1.5 μm wide, did not mineralize initially. The layer was lined externally by epithelial cells containing organelles suggestive of secretory activity. Electron-dense granular material resembling that produced by ameloblasts was observed in the epithelial intercellular spaces and in the unmineralized layer. The so-called initial layer of acellular cementum in the rats is therefore not cementum but a dentinal matrix to which epithelial secretory products are added.     

碳酸氢钠和氟化钠混合液治疗牙本质过敏症的临床研究

目的：观察碳酸氢钠和氟化钠混合液治疗牙本质症过敏的疗效。方法：实验组使用1mol/L浓度的碳酸氢钠和0．12mol/L浓度的氟化钠混合液；对照组用0．48mol/L浓度的氟化钠溶液，记录第3次治疗后的疗效及实验组3个月后的随访结果。结果：实验组，显效54例（90．00％），有效4例（6．67％），无效2例（3．33％），总有效58例（96．67％）。对照组，显效24例（40．00％），有效33例（55．00％），无效3例（5．00％），总有效57例（95．00％）。两组总有效率无显著差异（P＞0．05），显效率有显著差异（P＜0．05）。3个月后实验组疗效；显效47例（88．68％），有效4例（7．55％），无效2例（3．77％），总有效51例（96．23％）。结论：1mol/L浓度的碳酸氢钠和0．12mol/L浓度的氟化钠混合液是一种高效、作用持久、使用安全的牙本质脱敏剂。     

Perfusion fixation of rat molar pulp tissue for light microscopy

abstract – Molar pulp tissue of 42 rats was subjected to cardiac perfusion fixation or to immersion fixation with glutaraldehyde and formaldehyde. The importance of variations in pressure and duration of cardiac perfusion was studied. The results indicated that the perfusion method was superior to immersion fixation. Best preservation of the tissues was obtained when perfusion was performed with 1.7% glutaraldehyde for 10–12 min under a pressure of 130 cmH₂O, and leaving the animals for 4 h, without immersion fixation. Also perfusion with formaldehyde resulted in good preservation, provided the solutions were prepared from paraformaldehyde powder. In contrast, solutions made from commercial stock solutions of formaldehyde gave inferior results. For sections stained with hematoxylin-eosin, perfusion with glutaraldehyde was preferable, while perfusion with formaldehyde showed increased sensitivity in demonstration of dentinal tubules stained with alcian blue at pH 3.6. It is suggested that in future studies on rat pulp tissue, perfusion with glutaraldehyde and formaldehyde should replace immersion fixation.     

Topical review. Dental pain and odontoblasts: facts and hypotheses

Dental pain arises from exposed dentin following bacterial, chemical, or mechanical erosion of enamel and/or recession of gingiva. Thus, dentin tissue and more specifically patent dentinal tubules represent the first structure involved in dentin sensitivity. Interestingly, the architecture of dentin could allow for the transfer of information to the underlying dental pulp via odontoblasts (dentin-forming cells), via their apical extension bathed in the dentinal fluid running in the tubules, or via a dense network of trigeminal sensory axons intimately related to odontoblasts. Therefore, external stimuli causing dentinal fluid movements and odontoblasts and/or nerve complex responses may represent a unique mechanosensory system bringing a new role for odontoblasts as sensor cells. How cells sense signals and how the latter are transmitted to axons represent the main questions to be resolved. However, several lines of evidence have demonstrated that odontoblasts express mechano- and/or thermosensitive transient receptor potential ion channels (TRPV1, TRPV2, TRPV3, TRPV4, TRPM3, KCa, TREK-1) that are likely to sense heat and/or cold or movements of dentinal fluid within tubules. Added to this, voltage-gated sodium channels confer excitable properties of odontoblasts in vitro in response to injection of depolarizing currents. In vivo, sodium channels co-localize with nerve terminals at the apical pole of odontoblasts and correlate with the spatial distribution of stretch-activated KCa channels. This highlights the terminal web as the pivotal zone of the pulp/dentin complex for sensing external stimuli. Crosstalk between odontoblasts and axons may take place by the release of mediators in the gap space between odontoblasts and axons in view of evidence for nociception-transducing receptors on trigeminal afferent fibers and expression of putative effectors by odontoblasts. Finally, how axons are guided to the target cells and which kind of signaling molecules are involved is extensively discussed in this review.     

GSK3beta inhibitor-induced dental mesenchymal stem cells regulate ameloblast differentiation

ObjectivesEpithelial-mesenchymal interactions are extremely important in tooth development and essential for ameloblast differentiation, especially during tooth formation. We aimed to identify the type of mesenchymal cells important in ameloblast differentiation.MethodsWe used two types of cell culture systems with chambers and found that a subset of debtal mesenchimal cells is important for the differentiatiuon of dental spithelial cells into ameloblasts. Further, we induced dental pulp stem cell-like cells from dental pulp stem cells using the small molecule compound BIO ( a GSK-3 inhibitor IX) to clarify the mechanism involved in ameloblast differentiation induced by dental pulp stem cells.ResultsThe BIO-induced dental pulp cells promoted the expression of mesenchymal stem cell markers Oct3/4 and Bcrp1. Furthermore, we used artificial dental pulp stem cells induced by BIO to identify the molecules expressed in dental pulp stem cells required for ameloblast differentiation. Panx3 expression was induced in the dental pulp stem cell through interaction with the dental epithelial cells. In addition, ATP release from cells increased in Panx3-expressing cells. We also confirmed that ATP stimulation is accepted in dental epithelial cells.ConclusionsThese results showed that the Panx3 expressed in dental pulp stem cells is important for ameloblast differentiation and that ATP release by Panx3 may play a role in epithelial–mesenchymal interaction.     

Formation of odontoblast-like cells from cultured human dental pulp cells on dentin in vitro

Recent characterization of human dental pulp stem cells has shed new light on the understanding of the odontoblastic lineage. The purpose of the study was to characterize human adult dental pulp cells isolated and cultured in vitro and to examine the cell differentiation potential grown on dentin. We observed that some pulp cells isolated with an enzyme-digestion approach proliferated at a similar rate as the immortal cell line NIH 3T3. Population doubling time (PDt) for pulp cells at passage 3 was 22.6 +/- 0.5 hours and for NIH 3T3 was 23.1 +/- 2.3 hours. The pulp cells formed mineral nodules stimulated with dexamethasone or dexamethasone plus 1,25-dihydroxyvitamin D3. Pulp cells, after being seeded onto mechanically and chemically treated dentin surface, appeared to establish an odontoblast-like morphology with a cytoplasmic process extending into a dentinal tubule revealed by scanning electron microscopy analysis. Our data demonstrated the formation of cells with odontoblastic morphologies on existing dentin, suggesting that isolated human pulp stem cells may differentiate into odontoblasts on dentin in vitro.