High resolution scanning electron microscopy of the subplasmalemmal cytoskeleton in human odontoblasts

Abstract – The subplasmalemmal cytoskeleton in human odontoblasts was studied by high resolution scanning electron microscopy. The odontoblast layer was isolated and exposed to formaldehyde, glutaraldehyde, and OsO₄ for some specimens, while the membraneous structures and soluble proteins in the dental tissue were removed by Zenker's solution and 1% OsO₄ for other specimens, without further fixation of the remaining components. The cytoskeletal elements comprised a dense network of interlacing filaments of different diameters in the cell body. Most cytoskeletal elements were parallel to the axis of the cell processes situated inside the dentinal tubules. The appearance and orientation of the investigated subplasmalemmal cytoskeletal elements was unaffected by the choice of method. Both methods confirm the presence of a subplasmalemmal cytoskeleton in human odontoblasts.     

Early Cellular events in an actinomy D-Created detin niche in the rat incisor

Admnistration of actinnomycine D at a does level of 0.375 μg/g resulted in the selective disruption of development odontoblast at a critical stage of morphogenesis. A dentin niche was formed which was later repaired by cellular reparative dentin. The cellular changes which resilted in dentin niche formation were studied histologically and Ultrastructurally in serial longitudinal and transverse section from tissue obtained 10 h to 80 h following injection of the drug. Five stages were identified: initial destruction (10–20h), rapid destruction (30–40 h), debris removal (50–60 h), proliferation (60–80 h) and matrix deposition (post 80h). The cellular changes found in the dental papilla were considerably different from those found in inflammation, resolution and repair of fibrous connective tissue. These early stages were dominated by apoptosis and heterophagy. And after 80 h by disordered dentin matrix formation. The three-dimensional morphology of the defect was reconstructed from serial sections. The shape of the neich was the result of interference by actinomycin D in the patterns of proferation and migration of the cells in the apical region of the rat incisor tooth.     

年轻恒牙组织形态的超微结构观察

  目的 观察年轻恒牙形态的超微结构特点,并分析年轻恒牙牙体硬组织结构特点与龋患关系。方法 取临床因正畸治疗需要而拔除的根尖孔尚未完全闭合的恒前磨牙作为标本,应用透射电镜和扫描电镜观察年轻恒牙釉质表面、髓腔内侧壁牙本质及根尖区牙骨质的超微结构形态。结果 电镜下年轻恒牙的釉质表面为多孔状,牙本质小管密集,横断面呈圆形,成牙本质细胞较多。结论 年轻恒牙的组织形态超微结构特点是龋病易感的主要因素。     

尼古丁抑制成牙本质细胞增殖及作用机制的研究

目的观察尼古丁对成牙本质细胞增殖的抑制作用，并检测其对成牙本质细胞中Ca2+浓度的影响，以探讨尼古丁抑制成牙本质细胞的分子机制。方法体外培养成牙本质细胞系MDPC- 23细胞, 按每毫升2×104个接种，随机分为实验组和对照组，对照组不加任何刺激，实验组施加质量浓度为100 μg/mL尼古丁，并于8 h后加入浓度为10 μmol/L BrdU进行细胞周期标记，刺激24 h后固定细胞，行免疫荧光抗BrdU染色，碘化丙锭（PI）复染胞核，荧光显微镜下计数细胞总数与BrdU阳性细胞数，计算S期阳性细胞率并进行统计学分析。培养成牙本质细胞于特制培养皿中，施加质量浓度为100 μg/mL尼古丁刺激，激光共聚焦显微镜下检测成牙本质细胞中Ca2+浓度的动态变化。结果实验组、对照组S期阳性细胞率分别为36.3%、48.2%，实验组显著低于对照组。尼古丁刺激后，成牙本质细胞中Ca2+浓度迅速升高，在较高水平维持一段时间后缓慢下降。结论尼古丁可抑制成牙本质细胞增殖，这种作用同尼古丁升高成牙本质细胞中Ca2+浓度有关。     

Microhardness changes in dentine after neonatal capsaicin application

AIM: To determine if desensitization of the nociceptive innervation in the dental pulp has an effect on odontoblast function in the rat. METHODOLOGY: Neonatal systemic application of capsaicin was used to selectively eliminate nociceptive innervation. 12 capsaicin-treated rats were intravitally perfused at 150 days of life with 4% formaldehyde and jaws were prepared for Vicker's microhardness (VMH) measurement. As a control, 12 rats were injected with vehicle on the 3rd day of life and intravital perfusion was carried out exactly as those used for the experimental group. Immunohistological labeling of CGRP was carried out in both groups to assure the efficiency of desensitization in the experimental group. The VMH was measured in the incisors of each animal for a quantitative analysis of dentine quality. RESULTS: Vicker's microhardness was significantly higher in the control rats compared with the capsaicin-treated rats (P < 0.001). CONCLUSIONS: Neonatal systemic application of capsaicin produces changes in the quality of dentine in the rat over time and therefore it is suggestive that selective elimination of the nociceptive innervation in pulpal tissue may effect odontoblast function.     

细胞极性相关蛋白CDC42和PAR3在小鼠牙胚发育过程中的表达

目的：研究细胞极性相关蛋白CDC42和PAR3在小鼠牙胚发育过程中的表达,探讨其在牙胚发育中的可能作用。方法：取13.5、14.5、16.5和18.5d（E13.5、E14.5、E16.5和E18.5）的小鼠胚胎以及出生后1和5d（PN1和PN5）的小鼠,分离头部,固定、脱钙、脱水、石蜡包埋、切片,HE染色观察牙胚的组织形态表现,采用免疫组织化学染色方法观察CDC42及PAR3在牙胚发育过程中的表达。结果：HE染色观察,E13.5~E18.5分别为小鼠牙胚发育的蕾状期、帽状期、钟状早期和钟状晚期,PN1小鼠的牙胚中可见分化成熟的成牙本质细胞和成釉细胞,PN5小鼠牙胚可见牙冠部发育完成。免疫组织化学染色,CDC42在E13.5、E14.5和E16.5小鼠牙胚中有广泛表达,在E18.5小鼠牙胚中表达较E13.5、E14.5和E16.5减少,在PN1和PN5小鼠牙胚中主要表达于成牙本质细胞和成釉细胞的分泌端;PAR3弱表达于E13.5和E14.5小鼠牙胚,在E16.5和E18.5小鼠牙胚上皮处表达明显增强,在PN1和PN5小鼠牙胚中表达较E18.5减弱。结论：CDC42和PAR3参与小鼠牙发育的过程,在牙胚发育早期可能参与小鼠牙胚的增殖及迁移,在牙胚发育晚期可能参与了成牙本质细胞和成釉细胞的分化,尤其在成牙本质细胞和成釉细胞极性的形成与维持中可能具有重要作用。     

E-Cadherin在修复性牙本质形成中的免疫组化研究

目的：观察细胞粘附分子Ｅ－ｃａｄｈｅｒｉｎ在修复性牙本质表成中的免疫反应和定位。方法：在大鼠上、下颌第一磨牙近中面制备单面洞,分别观察３、１５和３０ｄ。标本行常规组织学处理。利用鼠抗Ｅ－ｃａｄｈｅｒｉｎ为免疫组化ＳＡＢＣ法,对标本进行标记,ＨＥ复染。结果：Ｅ－ｃａｄｈｅｒｉｎ在术后３ｄ成牙本质细胞梁色较弱,牙髓细胞为阳性;１５ｄ后,成牙本质细胞样细胞和牙髓细胞染色阳性;３０ｄ后,成牙本质细胞样细胞     

永生化成牙本质细胞表达牙本质基质蛋白的实验研究

目的　探讨永生化人成牙本质细胞样细胞系hTERT-hOd-l表达牙本质基质蛋白的情况。方法　矿化液培养hTERT-hOd-l细胞5周,检测骨钙素(OC)分泌量和碱性磷酸酶(ALP)活性。采用免疫组织化学、RT-PCR和原位杂交方法检测Ⅰ型胶原、骨涎蛋白(BSP)、牙本质基质蛋白1 (DMP1)以及成牙本质细胞标志物牙本质涎磷蛋白(DSPP) 和牙本质涎蛋白(DSP)在细胞中的表达。结果　在矿化液诱导下,hTERT-hOd-l细胞ALP活性和OC分泌量升高。 hTERT-hOd-l细胞在mRNA水平上表达BSP、DMP1和DSPP,在蛋白质水平上表达DSP和Ⅰ型胶原。结论　hTERT- hOd-l细胞在体外表达牙本质基质蛋白,具有矿化的潜能。