Localization of glycosaminoglycans in periodontal ligament during physiological and experimental tooth movement

Localization of chondroitin sulphates in periodontal ligaments (PDL) of rat molar roots during physiological and experimental tooth movement were analysed immunohistochemically with the use of monoclonal antibodies, 3B3 and 2B6, specific to chondroitin 6-sulphate (CH-6S) and chondroitin 4-sulphate/dermatan sulphate(CH-4S/DS), respectively. The maxillary first molars of experimental animals were forced to move laterally with a 10 g weight by U-shaped wires for 3 and 7 d. In control animals, 3B3 epitope was seen in the PDL near to the bone surface facing the distal half of roots, which corresponded to the compressive side during physiological tooth movement. Immunoreactivity for 2B6 was weak or negative in the PDL. Both epitopes were present at osteoid, precementum, lacunae and canaliculli of osteocytes and cementocytes. In 3-d-treated animals, the early stage of hyalinization characterized with visible cells and fibres was observed in the PDL at the buccal side of the mesial root, which showed intense immunoreactivity for 3B3. Further 3B3 positive area seen in control animals changed its position from the distal to the buccal side of the PDL. Immunoreactivity for 2B6 did not change in the PDL of 3-d-treated animals. In 7-d-treated animals, the typical hyalinization characterized with no visible cells and fibres was seen in the PDL at the buccal sides of both mesial and disto-buccal roots, where both epitopes were present at the peripheral part of the tissue. Observation of serial sections suggested that the 3B3-positive area was present at the peripheral part of the 2B6-positive area. The present results suggest that the expression of CH-6S is related to the compressive force in non-hyalinized and hyalinized PDL, whereas that of CH-4S/ DS is not influenced by the mechanical stress.     

Ultrastructural changes of the periodontal fibers and their attachment in rat molar periodontium incident to orthodontic tooth movement

abstract — Changes of the periodontal fibers and their attachment to cementum and alveolar bone in pressure zones of rat molars subjected to experimental forces were studied in the transmission electron microscope. In 67 rats, one maxillary first molar was moved buccally with a fixed appliance, using forces of 5–25 g. After experimental periods of 30 min to 28 d the animals were sacrificed and the experimental teeth with surrounding periodontal structures were processed for electron microscopy. After 24 h individual fibrils revealed bending and loss of functional orientation, while dense packing of fibrillar elements in irregular arrangement was a characteristic finding between 3 and 7 d. Within localized areas non-striated filaments had formed by longitudinal splitting of collagen fibrils. Even in the hyalinized zones of longest duration (7–14 d) less than 5% of the area seen in the electron microscope had degradèd in this manner. Regenerative processes predominated in the 14–28 d specimens. The apposition of cementum and alveolar bone had come to a halt during hyalinization, as indicated by distinct demarcation lines. The matrix of the new cementum and alveolar bone which had been deposited during the period of repair of the tooth attachment appeared more granular than the old hard tissues, due to a lower content of fibrillar elements.     

Erythrocytic crystallization in rat molar periodontium incident to tooth movement

abstract – From a material of 55 rats in which one maxillary molar had been moved buccally with a fixed orthodontic appliance, four specimens showing crystal-like structures in pressure zones in the periodontal ligament were studied with the electron microscope. Crystals which occurred within the lumen of enlarged blood vessels appeared to have formed by direct transformation of erythrocytes. Crystals were also seen in extravascular locations. These particles seemed to have formed by solidification outside the blood vessels of substances released from degraded red blood cells. Many particles revealed a highly ordered internal structure thought to reflect the unit cell arrangement of the crystalline substance. On the basis of their location, internal structure, and the experimental circumstances, it was concluded that the crystals represented hemoglobin, rather than hematoidin, ferritin or hemosiderin. The study indicated that under the special conditions of pressure and hemostasis that existed in the periodontal ligament as a result of the experimental procedures, local degradation of erythrocytes occurred in a few specimens by mechanisms different from the physiologic breakdown of red blood cells in the spleen, liver and bone marrow.     

Development and terminal differentiation of pulp and periodontal nerve elements in subcutaneous transplants of molar tooth germs and incisors of the rat

Ectopic tooth transplants are known to receive rich innervation of local neurons, but the precise location and structural features of neurites in the pulp and periodontal ligament (PDL) of such transplants are unclear. In this experiment, the molar tooth germs of rat embryos and incisors of young rats were subcutaneously transplanted into the dorsal regions of rats and processed, at various time intervals, for immunohistochemical demonstration of neural elements. Teeth with periodontal tissue elements developed in most of the molar transplants in 6 or 8 wk and received rich innervation, including some autonomic fibres, in the pulp. Nerve elements were also confirmed to be present in the PDL of these transplants, including specialized nerve ending-like structures reminiscent of the periodontal Ruffini endings. Mechanoreceptor-like structures were also induced in the regenerated PDL of similarly transplanted incisors, although the success rate was low. We conclude that rich and highly ordered innervation of the pulp, and occasional development of mechanoreceptors in the regenerated PDL of ectopic dental transplants, imply a high probability of successful induction of teeth with both nociceptive and mechanical sensations in the ectopic tooth and/or tooth germ transplant systems, although differentiation of mechanoreceptor-like nerve endings occured in only a few rare cases.     

Cementocyte cell death occurs in rat cellular cementum during orthodontic tooth movement

Objective:To clarify the mechanism of root resorption during orthodontic treatment, we examined cementocyte cell death and root resorption in the cellular cementum on the pressure side during experimental tooth movement.Materials and Methods:Using 8-week-old male Wistar rats, the right first molar was pushed mesiobuccally with a force of 40 g by a Ni-Ti alloy wire while the contralateral first molar was used as a control. Localization and number of cleaved caspase-3-positive and single-stranded DNA (ssDNA) - positive cells were evaluated using dual-label immunohistochemistry with anticleaved caspase-3 and anti-ssDNA antibodies. In addition, tartrate-resistant acid phosphatase (TRAP)-positive cells in the cellular cementum were evaluated using TRAP histochemical staining.Results:Caspase-3- and ssDNA-positive cells appeared at 12 hours, but were restricted to the compressed periodontal ligament (PDL) and not the cellular cementum. Cleaved caspase-3-positive cementocytes were observed in the cellular cementum adjacent to the compressed PDL on day 1. From days 2 to 4, the number of caspase-3- and ssDNA-positive cementocytes increased. TRAP-positive cells appeared on the cellular cementum at the periphery of the hyalinized tissue on day 7, and resorption progressed into the broad surface of the cementum by day 14.Conclusion:Cementocytes adjacent to the hyalinized tissue underwent apoptotic cell death during orthodontic tooth movement, which might have been associated with subsequent root resorption.     

The effects of orthodontic tooth movement on the glycosaminoglycan components of gingival crevicular fluid

Abstract In this study, gingival crevicular fluid (GCF) was collected from around a canine tooth, in children, before and during orthodontic tooth movement. The aim was to identify and quantify the glycosaminoglycan (GAG) components of GCF and relate them to tooth movement, gingival inflammation, plaque accumulation, pocket probing depth and GCF volume recorded at the site of sampling. GAG in GCF samples, collected for a 15-min period into microcapillary tubes, were separated electrophoretically, stained with Alcian blue and quantified using a laser densitometer. 2 GAG components of hyaluronic acid (HA) and chondroitin sulphate (CS) were identified. The increase in GCF volume during orthodontic tooth movement was only partly due to increased gingival inflammation, GAG levels varied with different types of orthodontic tooth movement. In GCF, levels of CS, in particular, may reflect the changes in the deeper periodontal tissues which could be monitored during orthodontic tooth movements.     

Immunocompetent cells in rat periodontal ligament and their recruitment incident to experimental orthodontic tooth movement

The aims of this study were to evaluate the number and distribution of immunocompetent cells in normal rat periodontal ligament (PDL) and to quantify their recruitment incident to experimental tooth movement. 27 young animals had the 1st right maxillary molar moved mesially by an orthodontic appliance for 1, 3, 7 and 14 days, respectively. 4 animals served as untreated controls. An immunohistochemical procedure was carried out on alternate serial cryostat sections, and monoclonal antibodies against CD1 II (macrophages, dendritic cells), CD43⁺ (lymphocytes, polymorphs), CD4 (helper T-lymphocytes), and class II MHC molecules were used. Mean counts of the immunolabeled cells in the control group showed the highest number of GDI lb⁺ and class II molecule expressing cells, while CD4⁺ and CD43⁺ cells were scarcely found. Significant increase in the number of CD1 lb⁺, CD43⁺ cells and class II molecules was found in the PDL of the experimentally moved 1st molars compared with the contralateral side and the control group, while CD4⁺ cells showed no significant increase. CD11b⁺ and cells expressing class II molecules were found around hyalinized tissue, between dentin and cellular cementum and close to Malassez'epithelial cells. In conclusion, normal rat PDL has high number of macrophage and dendritic-like cells, but few lymphocytes and granulocytes. Furthermore, experimental tooth movement leads to significant recruitment of cells belonging to the mononuclear phagocytic system, but has no significant effect on the number of lymphocytes and granulocytes in the rat PDL.     

正畸移动狗乳磨牙对其恒牙胚的影响

目的:研究正畸力作用下,狗乳磨牙移动对恒牙胚以及恒牙萌出的影响。方法:选用健康15～16周龄雄性中国狗20只,左侧下颌为实验侧,右侧下颌为对照侧,以狗乳尖牙为支抗移动第二乳磨牙。观察正畸加力0d、7d、14d和21d后狗第二乳磨牙的移动以及第三恒前磨牙牙胚的变化。同时对乳牙牙周组织、恒牙牙胚的牙囊、周围牙槽骨进行组织学观察,并对牙槽骨内的破骨细胞进行计数。结果:正畸加力21d后,与对照侧相比,实验侧第二乳磨牙近移31.5%,X线片显示第三恒前磨牙牙胚位置与对照侧相比近移31.9%,二者有明显差异。恒牙萌出后近移30.3%。组织学观察:第二乳磨牙和第三恒前磨牙牙胚近中侧均见骨质吸收,可见破骨细胞;远中侧可见新骨形成。第三恒前磨牙牙胚牙囊近中侧纤维受压、变窄,远中侧纤维受到牵张、变宽。结论:正畸力移动乳牙对其继承恒牙的萌出具有诱导作用,可以使恒牙胚在牙槽骨中有同方向移动趋势,萌出位置改变。     

大鼠磨牙牙根根端组织异体移植实验

目的:探讨大鼠磨牙牙根根端组织发育牙根和牙周组织的能力.方法:选择出生后25 d的SD大鼠,切取下颌第一磨牙牙根端组织,移植于成年大鼠肾被膜下,4周后取材,组织学观察.结果:大鼠磨牙牙根端组织发育出根尖样组织结构.结论:牙根根端组织具有发育牙根和牙周组织的能力.     

大鼠磨牙根尖组织的组织学观察

目的：观察大鼠根尖组织的组织结构特点,了解根尖诱导成形术的组织学基础。方法：制备SD大鼠（出生后20d）下颌的石蜡组织切片,通过HE染色、免疫组化染色（抗增殖细胞核抗原、抗角蛋白）和天狼星红染色观察SD大鼠下颌第一磨牙根尖组织。结果：下颌第一磨牙牙根发育约2／3,根尖区组织细胞密集且有较多增殖细胞,胶原纤维规律排列,根尖端的上皮根鞘仍保持完整。结论：发育牙根的根尖组织具有较强的组织修复再生能力,保留生活的根尖组织是根尖诱导成形术成功的关键。     

Comparative study of gene expression during tooth eruption and orthodontic tooth movement in mice

Objective:  The aim of this study was to understand tooth eruption by comparing the gene expression during tooth eruption and orthodontic tooth movement (OTM).
Materials and methods:  Orthodontic force was applied on maxillary molars for 2, 4, 7 and 14 days to study tooth movement. Mice at PN 0, 7, 10, 15 and 21 were fixed to observe tooth eruption. Comparative study of two procedures was assessed by haematoxylin and eosin, tartrate-resistant acid phosphatase staining and in situ hybridization for matrix metalloproteinase ( Mmp ) 2 , 13 , bone sialoprotein ( Bsp ) and osteocalcin ( Ocn ).
Results:  Tartrate-resistant acid phosphatase activity and expression of Mmp2 , 13 were obviously detectable in the compression region during OTM. They were also identified in the occlusal and apical region of alveolar bone during tooth eruption. Strong expression of Bsp and Ocn was detectable at the tension side during OTM. These genes were also expressed in the inner lateral region of alveolar bone adjacent to the tooth, but absent in the inner surface of the occlusal and root apical regions during tooth eruption.
Conclusion:  The process of alveolar bone metabolism during developmental eruption and OTM shares the same mechanism. Internal force, as the orthodontic force for OTM, may be initiating factor for tooth eruption.     

Crystallite arrangement of hydroxyapatite microcrystals in human tooth cementum as revealed by electron paramagnetic resonance (EPR)

Human dental cementum was analyzed by electron paramagnetic resonance (EPR). The measured EPR powder spectra of γ-irradiated cementum resembled those of γ-irradiated enamel. Both spectra were characterized by the same line shapes and g values. The position of the extreme first derivate peaks can be described by g₁ =2.0023 and g₂= 1.9971 ± 0.0002, and are assignable to the CO₃^3? center. The angular dependence of the cementum EPR spectra indicates a different arrangement of the hydroxyapatite microcrystals compared to that of enamel. A corresponding model of cementum microcrystal alignment has been proposed. The methodology presented can be utilized for studying the mineralization process of root cementum and other mineralized tissues.     

Expression of osteocalcin in cementoblasts forming acellular cementum

To determine the phenotypic expression of cementoblasts responsible for acellular cementum, an immunohistochemical study was performed using a polyclonal antibody raised against the aminoterminal peptide of rat osteocalcin (OC). Maxillary first molars of Wistar male rats aged 2 and 3 wk were used for observations. Serial sections of decalcified paraffin embedded specimens were stained either with hematoxylin and eosin or with the anti-OC antibody. In 2-wk-old rats, apical roots were lined with the epithelial root sheath. A thin layer of acellular cementum was seen at most of the root surface, but was not seen near to root apex. In 3-wk-old rats, cellular cementum began to be formed at root apex, and acellular cementum became more thick than in 2-wk-old rats. Acellular and cellular cementum were lined with the fibroblast-like cells. Osteocalcin staining was detected in cells lining root surface in both 2- and 3-wk-old rats. Almost all cells lining cellular cementum were positive for OC. In contrast OC positive cells lining acellular cementum and root surface devoid of cementum appeared at a specific site of the root. The cells at the interradicular area of root surface were positive but the cells at the outer area (the opposite side of the interradicular area) were negative for OC. Osteoblasts and odontoblasts were positive with the antibody. The present results suggest that the OC expression of cementoblasts forming acellular cementum is similar to that of cells forming cellular cementum as well as osteoblasts and odontoblasts, and has a role for calcification of acellular cementum.     

肌动蛋白在大鼠磨牙及其发育中的分布研究

目的：探讨肌动蛋白在大鼠磨牙及其发育中的时空分布及意义。方法：建立大鼠牙胚发育模型。用免疫组化方法检测肌动蛋白在成年和发育期磨牙中的表达。结果：肌动蛋白在钟状期外釉上皮细胞，分泌前期成釉细胞和成牙本质细胞及中间层细胞之间，硬组织形成期成釉细胞Tomes'突和成牙本质细胞胞浆呈阳性表达。结论：肌动蛋白在大鼠磨牙发育过程中的分布具有时空特异性。与细胞的分化程度和功能状态密切相关。     

大鼠牙移动中牙周膜功能状态与牙槽骨改建关系的研究

目的:通过检测不同牙周膜功能状态大鼠磨牙移动过程中压力侧牙槽骨表面破骨细胞计数及牙移动距离的差异,探讨牙周膜功能状态与牙槽骨改建的关系.方法:选择48只6周龄体重(250±20)gSD大鼠,随机分为正常对照组(16只)和实验组(32只).通过拔除实验组大鼠右下颌所有磨牙使其上颌左、右第一磨牙咬合力改变,3周后分别形成牙周膜代偿性机能亢进模型和牙周膜废用性萎缩模型,据此实验组又分为萎缩组与亢进组(各16只).在各组大鼠上颌切牙和第一磨牙间放置5mm镍钛拉簧,初始力值60g,近中移动磨牙.分别于加力0d、3d、7d,每组各处死动物2只,14d处死其余动物,制备组织学标本,通过TRAP染色鉴别第一磨牙近中根压力侧牙槽骨表面破骨细胞并记数;对加力14d后的标本拍摄X线片测定牙齿移动距离.结果:牙周膜机能代偿性增强组牙齿移动距离(0.265±0.107mm)明显小于其它两组(0.631±0.142mm,0.679±0.090mm),P<0.01.牙周膜废用性萎缩组与正常对照组移动距离无显著性差异(P>0.05).萎缩组大鼠近中根压力侧牙槽骨表面破骨细胞计数在实验全过程中(除0d外)均低于亢进组和对照组,有显著性差异(P<0.05).结论:处于退化状态的废用性萎缩牙周膜对矫治力抵抗性差,细胞分化增殖不活跃,骨组织改建率低;机能代偿性增强牙周膜对矫治力抵抗性好,细胞分化增殖活跃,成骨活动大于破骨活动.     

The development and structure of principal fibers and cellular cementum in rat molars

The principal fibers and cellular cementum were examined in various developmental stages in rat molars by light and electron microscopy, and their development and structure were studied. The principal fibers increased in thickness with root development. At any given stage, the principal fibers consisted of small fiber bundles on the cementum surface and showed a branching structure. Sharpey's fibers also showed a similar branching structure and increased in thickness, as traced superficially in the cementum. It was suggested that the principal fibers increased in thickness by means of the aggregation of fiber bundles. The development of cellular cementum was divided into two (early and late) phases on the basis of the fiber arrangement. The early cementum was believed to contain a few Sharpey's fibers and many randomly arranged intrinsic fibers. The late cementum contained thicker Sharpey's fibers with branches and the intrinsic fibers which appeared to encircle Sharpey's fibers and/or meander among them.     

正畸牙齿移动过程中大鼠三叉神经节CGRP mRNA表达变化

目的：观察正畸牙齿移动不同时期大鼠三叉神经节CGRP mRNA表达变化。方法：采用反转录。聚合酶链反应（RT-PCR）技术检测正畸加力和撤力后不同时期大鼠三叉神经节CGRP mRNA水平的变化。结果：加力后2h CGRP mRNA的表达量没有明显增加,至加力后8h开始增加,加力1d-1W达到最高峰,至撤力后2～4WCGRP mRNA的表达量开始下降,但CGRP mRNA水平仍然明显高于对照组（P〈0．05）。结论：CGRP可能不仅参与早期正畸牙周组织的炎症损伤过程,而且可能参与了后期的组织修复重建过程。     

骨皮质切开术对大鼠牙移动影响的实验研究

目的建立骨皮质切开术正畸牙移动实验动物模型,观察移动速率和对牙周组织改建的影响。方法 75只SD大鼠,实验组35只行上颌双侧中切牙骨皮质切开术,1周后与对照组35只同时进行正畸加力,空白对照组5只不进行加力。按不同加力时间处死,测量牙齿移动的距离,观察组织学改变。结果实验组2周时牙齿移动距离(3.471±0.359)mm,对照组为(1.247±0.198)mm,具有显著性差异;HE染色结果表明,实验组牙周膜内玻璃样变组织的形成相对于对照组来说,范围缩小,时间缩短。结论骨皮质切开术能够加速正畸牙移动。     

大鼠正畸牙齿移动过程中Smad1的表达和意义

目的：探讨BMP超家族的细胞内信号转导分子Smad1蛋白在大鼠正畸牙齿移动过程中牙周组织的分布和表达。方法：用ABC免疫组织化学法，检测实验性大鼠正畸牙齿移动过程中牙周组织Smad1的表达。结果：Smad1在正畸牙齿加力各个时期存在的特异的分布模式，与BMP有相似之处。结论：Smad1参与了大鼠正畸牙齿移动过程，可能介导了BMP在正畸牙齿加力区的信号。