氯化镧对体外培养的犬骨髓基质细胞增殖的影响

目的:观察3种浓度的氯化镧(LaCl3,La3+)对体外培养的犬骨髓基质细胞(bone marrow stromal cells,BMSCs)的影响。方法:用5.564×102、5.564、5.564×10-2μg/mL3种浓度的La3+干预第3代BMSCs,设空白对照组和50ng/mLBMP-2干预的阳性对照组。通过MTT法、碱性磷酸酶半定量检测分析La3+对BMSCs的影响。结果:5.564μg/mL La3+干预组BMSCs在第6、7天表现出较高的生长趋势;碱性磷酸酶半定量检测发现5.564μg/mL La3+干预组OD值均数明显高于另两个实验组均数(P<0.05)。结论:5.564μg/mL浓度的La3+可促进BMSCs增殖。     

氯化镧对实验犬组织工程骨成骨的影响

目的观察氯化镧对植入实验犬颌骨自体骨缺损区的组织工程骨成骨情况的影响。方法选成年杂种犬9只。从股骨抽取骨髓、以密度梯度离心法获取骨髓基质细胞（BMSCs）并诱导培养为成骨细胞。以5.564μg／ml的氯化镧（La^3＋）干预第三代BMSCs并将其与猪脱钙冻干骨（fdDBM）复合，体外培养7d后植入实验犬颌骨人造骨缺损处（2cm×1cm×0．8cm），另设BMSCs加fdDBM、fdDBM及空白对照组。3个月及6个月后处死实验犬获取颌骨标本并加工处理作形态学观察及X线骨密度分析。所得数据用SAS 6.12软件包进行方差分析。结果BMSCs与fdDBM复合回植3个月后实验犬骨缺损能完全或大体为新生骨修复。La^3＋干预组回植区新生骨的密度值较高．但与对照组的密度值比较差异无统计学意义（P〉0．05）。回植6个月后BMSCs加fdDBM与单纯fdDBM回植区新生骨的骨密度值比较差异无统计学意义（P〉0.05），但该二组的骨密度值明显高于对照组（P〈0．05）。结论5．564μg／ml浓度的氯化镧对回植入实验犬人造颌骨缺损处的组织工程骨成骨作用无明显负面影响。     

氯化镧对人牙龈成纤维细胞增殖的影响

目的:检测不同浓度氯化镧(La3 )对人牙龈成纤维细胞(HGF)增殖的影响。方法:体外培养HGF,用四唑盐法(MTT)检测6种浓度La3 (1×10-6、1×10-5、5×10-5、1×10-4、1×10-3及1×10-2 mol/L)对HGF的作用。结果: 1×10-2 mol/L的La3 能抑制HGF的生长, 1×10-6、1×10-5、5×10-5、1×10-4及1×10-3 mol/L的La3 对HGF生长无抑制作用。结论: 1×10-2 mol/L浓度的La3 对HGF增殖有抑制作用, 1×10-4 mol/L以下浓度的La3 对HGF无抑制作用。     

不同浓度含镧制剂对人牙根面的抗龋作用

目的:比较含不同浓度镧(La)的漱口水和凝胶对根面的防龋作用,并确定这两种剂型的最低有效浓度。方法:在体外模拟口腔pH变化的环境中,采用含不同浓度La的漱口水(100×10-6~2 000×10-6mmol/L)及凝胶(5 000×10-6~10 000×10-6mmol/L),对离体人牙根面进行抑制脱钙的实验研究,并借助偏光显微镜、电子探针及能谱分析手段,对不同浓度的La3 在根面的抑制脱矿作用进行定性和定量分析。结果:>500×10-6mmol/L的含La3 漱口水及>10 000×10-6mmol/L的含La3 凝胶均具有防龋作用,且其脱矿深度无显著差别。结论:500×10-6mmol/L和10 000×10-6mmol/L的La3 含量分别是这两种制剂防治根面龋的最低有效浓度,而使用含镧(La)漱口水制剂可能更适合老年人根面龋的防治。     

含镧制剂对根面抗龋作用的最低有效浓度的研究

目的 :比较不同浓度的含镧 (La)漱口水和凝胶对根面的防龋作用 ,并确定这两种剂型的最低有效浓度。方法 :在体外模拟口腔 pH循环变化的环境中 ,采用不同浓度的含La漱口水 ( 10 0× 10 -6-2 0 0 0× 10 -6mmol/L)及凝胶 ( 5 0 0 0× 10 -6 10 0 0 0× 10 -6mmol/L) ,对离体人牙根面进行抑制脱钙的实验研究 ,并借助偏光显微镜、电子探针及能谱分析手段 ,对不同浓度的La3 在根面的抑制脱矿作用进行定性和定量分析。结果 :5 0 0× 10 -6mmol/L以上含La3 漱口水及 10 0 0 0× 10 -6mmol/L以上含La3 凝胶均具有防龋作用 ,且其脱矿深度无显著差别 .结论 :5 0 0× 10 -6mmol/L和 10 0 0 0× 10 -6mmol/L的La3 含量分别是这两种制剂防治根龋的最低有效浓度 ,而使用含镧漱口水制剂可能更适合老年人根龋的防治。     

周期性流体静压力对BMSCs细胞增殖活性及超微结构的影响

目的:观察周期性流体静压力作用下骨髓间充质干细胞(Bone Marrow Stem cells,BMSCs)细胞增殖活性及超微结构的改变。方法:贴壁法分离培养大鼠BMSCs。采用自主研发的流体静压力加载装置对大鼠BMSCs进行力学加载,加载压力为0~45 kPa、0~90 kPa,加载频率为0.1 Hz,时间1 h,连续加载2 d。对照组细胞不加压。采用CCK8法检测细胞的增殖活性,采用透射电镜观察大鼠BMSCs超微结构的变化。结果:0~45 kPa 、0~90 kPa周期性流体静压力作用下BMSCs增殖活性显著高于对照组(P<0.05),透射电镜下观察发现,0~45 kPa作用下BMSCs细胞内质网轻度扩张,胶原合成量增加。而0~90 kPa作用下细胞出现了大量胶原及凋亡小体。结论:适宜的周期性流体静压力作用下可提高细胞的增殖活性,但过高的压力刺激可诱导细胞产生凋亡。     

含镧漱口水对人牙釉质及根面抗酸作用的研究

目的比较不同浓度的含镧(La)漱口水的防龋作用,并对其在牙釉质、牙骨质的抗酸作用进行比较.方法在体外模拟口腔pH循环变化的环境中,用含(100×10-6mmol/L)～(2000 ×10-6mmol/L)La3+的漱口水处理离体牙根面,并对其脱矿情况进行定性和定量分析,筛选出最低有效防龋浓度的漱口水.然后用此漱口水每日处理离体牙牙釉质和牙根面1次,共计5h.结果 500 ×10-6mmol/L以上的含La3+漱口水均具有防龋作用.经500 ×10-6mmol/L漱口水处理的牙根面与牙釉质的脱矿深度无显著差别,而处理前后脱矿量的变化显示,釉质的脱矿量大于根面.结论 500×10-6mmol/L的La3+含量是最低有效防龋浓度,且含镧防龋制剂可能更适合于老年人根龋的防治.     

RA和bFGF体外诱导人牙髓干细胞向神经细胞分化的研究

目的:探讨全反式维甲酸(all-trans-retinoic acid,RA)、碱性成纤维生长因子(basic fibroblast growth factor,bFGF)体外诱导人牙髓干细胞(dental pulp stem cells,DPSCs)分化为神经细胞的可行性。方法:体外分离培养DPSCs并进行克隆化,检测STRO-1的表达。将DPSCs分别接种于含有不同浓度RA、bFGF或二者结合的诱导液,MTT法检测细胞增殖能力,免疫荧光法检测微管相关蛋白(microtubule-associated protein-2,MAP-2)、神经元烯醇化酶(neuron-specific enolase,NSE)、胶质原纤维酸性蛋白(glial fibrillary acid protein,GFAP)的表达。透射电镜观察诱导前后细胞超微结构。结果:克隆来源细胞的STRO-1表达阳性。0.5μg/ml RA或20 ng/ml bFGF单独应用促增殖作用最强(P<0.05),bFGF单独作用各组及对照组均未检测出神经元样细胞。RA作用各组检测到阳性细胞。而0.5μg/ml RA与20 ng/ml bFGF联合应用的增殖和分化效应均优于其它组。透射电镜观察到幼稚神经元样细胞。结论:RA和bFGF联合应用可在体外有效诱导人DPSCs转化为神经细胞。     

氯化镧漱口水防治老年根面龋三年临床观察

目的:观察和评价氯化镧(LaCl3)漱口水防治老年根面龋的临床效果.方法:选择63～84岁患有牙龈萎缩根面暴露的老年人222名,随机分为实验组和对照组,每组均按常规方法记录牙龈退缩数和根面龋补数,实验组每日使用一次500×10-6mmol/L的含La3 漱口水漱口,对照组使用NaCl漱口水漱口,两组均使用1个月、1年及2年后重复使用漱口水1个月,3年后复查每个老年人根面龋情况.结果:实验组根龋指数(RCI)的新增值明显低于对照组.结论:500×10-6 mmol/L的含La3 漱口水能有效地控制老年根面龋.     

低浓度氯化镧对乳牙人工龋再矿化的研究

目的研究低浓度氯化镧(LaCl3)溶液对乳牙人工釉质龋再矿化的影响,比较3种不同浓度氯化镧的再矿化效果。方法选取30名5~7岁乳牙列无龋儿童因滞留拔除的下颌乳中切牙30颗,制备为人工龋后,随机分为3组,每组10颗。将所有标本沿牙体长轴从唇舌方向正中对半切开,分别作为实验组与对照组。将3组实验组牙片分别每日3次悬吊浸泡于LaCl3溶液(La3 质量浓度分别为0.1、0.5、2.0g/L),每次浸泡0.5h,其余时间浸泡在人工唾液中。对照组牙片始终用人工唾液浸泡。共处理10d。使用显微硬度测量仪测量各组釉质的显微硬度。统计采用配对t检验和方差分析。结果3个对照组的釉质显微硬度之间差异无统计学意义(P>0.05)。3个实验组均较相应对照组的釉质显微硬度增加,差别有高度显著性(P<0.01)。3组釉质显微硬度增加值之间差异无统计学意义(P>0.05)。结论低浓度氯化镧对乳牙人工釉质龋有再矿化作用。0.1g/L的镧溶液具有与0.5、2.0g/L镧溶液相同的再矿化效果。     

TEM下自酸蚀粘结剂的两种模式牙本质纳米渗漏观察

目的：观察并评价3种自酸蚀粘结剂(Clearfil^TM SE Bond、Adper^TM Prompt^TM、PANAVIA^TM F)牙本质粘结界面的纳米渗漏。方法：选取12颗无龋坏人磨牙,用600目碳化硅砂纸在流水冲洗下预备出统一的牙本质粘结面玷污层。分别用3种自酸蚀粘结剂按使用说明进行粘结处理。每颗牙齿垂直于粘结面切割出5个0．9mm厚粘结试件,分别避光贮存于硝酸银、氨化硝酸银溶液中24h,在TEM下观察牙本质粘结界面纳米渗漏。结果：在TEM下观察到3种自酸蚀粘结剂牙本质粘结界面混合层中呈网状、点状两种模式分布的纳米渗漏。结论：自酸蚀粘结剂由于其自身的组份特性,牙本质粘结界面均存在不同程度的纳米渗漏。     

侵袭性牙周炎龈上皮细胞凋亡的研究

目的 观察侵袭性牙周炎龈上皮凋亡细胞形态特征,了解牙周炎发病机理.方法 以24例病变龈上皮组织为研究对象,利用透射电镜观察凋亡细胞的超微形态结构改变,流式细胞术测定分析细胞周期,及免疫细胞与细胞凋亡的关系.结果 凋亡细胞形态学改变分为四型,流式细胞术测试在G1期前出现亚G1期峰,凋亡细胞为19.9%;免疫细胞发生活化.结论 龈上皮细胞确有凋亡细胞存在,凋亡是在免疫细胞活化的基础上诱导进行的,由于细胞凋亡而导致牙周袋加深,形成侵袭性牙周炎.     

自酸蚀黏结剂的牙本质黏结界面纳米渗漏观察

目的观察并评价3种自酸蚀黏结剂(ClearfilTMSEBond、AdperTMPromptTM、PANAVIATMF)黏结处理后的牙本质纳米渗漏。方法选取9颗无龋坏人磨牙,用600目碳化硅砂纸在流水冲洗下预备出统一的牙本质黏结面玷污层。分别用3种自酸蚀黏结剂按使用说明进行黏结处理。每颗牙齿垂直于黏结面切割出5个0.9mm厚黏结试件,避光贮存于氨化硝酸银溶液中24h,在透射电镜(TEM)下观察牙本质黏结界面纳米渗漏。结果3种自酸蚀黏结剂试件牙本质黏结界面混合层的底部在TEM下均观察到排列规则的银粒子渗漏。结论自酸蚀牙本质黏结界面下可能存在未完全封闭的牙本质脱矿层,存在纳米渗漏。     

细菌胞外膜泡的电镜观察

本实验对分离的５株细菌的ＥＣＶ作了电镜观察，证实４株菌都能形成ＥＣＶ，但在量上有差异。以ＰｇＷ５０最为典型，菌体及其ＥＣＶ的结构层次清晰可见；ＰｅＥＣＶ的荚膜层较薄且直径相对小而均一；Ｐｍ和Ｐｉ的ＥＣＶ大小不均一，标本中破裂现象和不规碎片出现较早。     

Elemental sulphur and alkali elutable melanin detected in oral melanosis and malignant melanoma by energy-filtering transmission electron microscopy

BACKGROUND: The morphology and contents of melanosomes are important features for differentiating melanocyte-derived melanotic lesions such as melanosis and malignant melanoma. METHODS: In this study, we attempted to elucidate the structure of melanin and sulphur content in oral melanosis and malignant melanomas by ultrastructural analysis. RESULTS: In oral melanosis, the essential pathological findings were overproduction of eumelanin and discharge of melanin into keratinocytes. In malignant melanoma in situ, pleomorphic and ellipsoid abnormal melanosomes with an increase in sulphur content and alkali elution rate were detected. In invasive malignant melanoma, the irregular ellipsoid and spheroid melanosomes existing either as discrete bodies or compound melanosomes with furtherly increased sulphur content and alkali elution were detected. CONCLUSIONS: Our findings suggest that abnormal melanosome morphology and high sulphur content are predictive markers for assessment of early or precancerous melanotic lesions and malignant melanoma.     

小鼠磨牙上皮根鞘与牙骨质发育的关系

目的：研究上皮根鞘细胞（Hertwig’s epithelial root sheath,HERS）的发生,非胶原基质蛋白骨桥素（osteopontin,OPN）、骨涎蛋白bone sialoprotein,BSP）及牙骨质发育之间的关系。方法：取出生后5d、9d、15d、19d及4个月的BALB／c小鼠共18只,光镜观察上皮根鞘的发生、牙骨质发育过程,透射电镜观察该阶段上皮根鞘细胞的超微结构。PV免疫组化两步法检测增殖细胞核抗原（proliferating cell nuclear antigen。PCNA）、OPN、BSP在牙骨质形成过程中的表达。结果：超微结构观察发现,内、外釉上皮在融合过程中,2层细胞间包含个别具有丰富细胞器的一类细胞。其胞质中可见大量的高尔基复合体和粗面内质网,通过桥粒与内、外釉上皮连接。免疫组化显示,新形成上皮根鞘细胞呈OPN阳性表达,但未见BSP阳性表达。在牙根的发育过程中,牙本质一牙骨质交界处OPN有较高表达,伴随牙骨质基质在根面不断沉积,OPN在牙骨质中表达增强,成熟牙根表面可见厚度不同的牙骨质覆盖,OPN阳性表达。结论：上皮根鞘发生时,内、外釉上皮间可包容个别源于网状层或中间层、细胞器丰富的细胞,从而形成类似成釉器的结构。其可能在牙根发育起始阶段分泌某些蛋白和信号分子,启动根部牙本质和牙骨质的形成。     

生长期犬上颌缝牵引的组织学研究

目的：了解上颌缝牵张成骨对骨形成蛋白2、4、7及缝组织超微结构的影响。方法：以杂种犬21只为实验动物,实验组在腭横缝及额上颌缝安置内置式镍钛合金牵引丝。对照组不作处理。术后5d、10d、15d、20d、30d、40d切取腭横缝组织,进行免疫组化研究及透射电镜观察。结果：实验组腭横缝被成功延长,BMP-2、BMP-4呈强阳性表达,BMP-7弱阳性表达,阳性染色主要定位于成骨细胞中。结论：缝牵引可以顺利扩张生长期上颌骨缝;BMP-2、BMP-4可以激发和增殖前体间充质细胞,并可诱导间充质细胞转化为成骨细胞;1500g力既可明显增强成骨细胞生物学活性又不会造成器质性损伤。     

In vivo transmission electron microscopy of the alternating structure of the protrusions between adjacent macrophage-like cells on micropatterns

ObjectivesWe investigated the behavior of macrophages in the defined microtopography of materials.MethodsPatterned cyclo-olefin polymer films were implanted into the femurs of seven-week-old rats. After 1 and 4 weeks, the rats were fixed with glutaraldehyde and OsO₄, and their bones were observed with transmission electron microscopy (TEM).ResultsTEM and segmentation revealed an alternating structure in which multiple protrusions of adjacent macrophage-like cells overlapped. They were approximately 2 μm long and almost uniform in width, and were induced by the limited topography.ConclusionNew structures appeared between the macrophage-like cells as a result of microtopography.     

Dentin conditioned with a metal salt-based conditioner

ObjectivesUniversal adhesives (UAs) can optionally be applied in either an etch-and-rinse (E&R) or self-etch (SE) bonding mode. As the preferred bonding mode differs for enamel versus dentin, a universal conditioner for both enamel and dentin in replacement of the relatively aggressive phosphoric-acid etchant remains desirable. This study aimed to test if a metal salt-based etchant (ZrO(NO₃)₂) provides as durable bonding to dentin as a classic E&R or SE bonding modeMethodsBefore applying the UA Adhese Universal (‘AdU’; Ivoclar Vivadent) to bur-cut dentin of 24 teeth (n = 8), dentin was conditioned with either (1) an experimental metal salt-based conditioner (‘ZON’; Ivoclar Vivadent) or (2) 37% phosphoric acid (Total Etch gel, Ivoclar Vivadent), representing a classic ‘E&R′ mode; (3) a third experimental group involved AdU applied in SE mode. Bonding effectiveness was determined in terms of immediate (‘1w’) and aged (‘50k’ TC) micro-tensile bond strength (μTBS) to bur-cut dentin. Adhesive-conditioned dentin interfacial interactions were characterized by S/TEM.ResultsLinear mixed-effects modeling revealed significantly higher immediate μTBS to dentin of ZON_AdU than E&R_AdU, while ZON_AdU performed not significantly different from SE_AdU. No significant differences were found between the three experimental groups after 50k TC (aged μTBS). S/TEM disclosed less exposure of dentinal collagen fibrils when AdU was bonded upon ZON etching than when applied in E&R mode. Moreover, ZON resulted in more hydroxyapatite (HAp) crystals remaining at the bottom of the hybrid layer, while dentinal tubule orifices remained nearly fully closed, by which hardly any resin tags were formed.SignificanceThe alternative metal salt-based conditioner revealed at dentin a more HAp-protected hybrid layer with less exposure of collagen fibrils, while a comparable bond strength was obtained to that with a phosphoric-acid E&R as well as with an SE (no conditioner) bonding mode. These findings confirm that the metal salt-based conditioner can be considered as a suitable alternative (enamel/)dentin conditioner to classic phosphoric acid employed in an E&R bonding mode.     

Dentin bonding: effect of tubule orientation on hybrid-layer formation

Schüpbach P. Krejci I, Felix L: Dentin bonding: effect of tubule orientation on hybrid-layer formation.
In an attempt to compare the morphology of the resin-dentin interface in areas where the dentinal tubules run perpendicularly or at an angle to the cavity surface with that of areas where they run parallel to it, we studied a dentin adhesive system using transmission electron microscopy and fluorescence confocal laser scanning microscopy. The design of the study included the simulation of the normal hydrostatic pressure within the pulp and the dentinal tubules. Following acid etching of the dentinal surface with maleic acid/HEMA, the smear layer was removed, and a superficial zone was demineralized in such a way that the exposed collagenous dentin matrix retained its integrity. Confocal laser scanning microscopal investigations using primer labeled with rhodamine B showed that the penetration of the primer occurred not only vertically via surface porosities, but mainly laterally, via the dentinal tubules. The adhesive resin labeled with fluorescein completely infiltrated the demineralized layer, thereby forming a hybrid layer. The orientation of the dentinal tubules had a profound effect on the formation of the hybrid layer. In areas with perpendicular tubule orientation, the layer was 3.2 ± 0.8 μm thick, showing solid 27.2 ± 0.8 μm long resin tags in the dentinal tubules, and a network of tiny tags in their side-branches. In areas with parallel tubule orientation, the layer was significantly thinner (1.3 ± 0.6 μm) and resin tags were absent.