大鼠正畸牙移动压力侧牙槽骨中cathK、RANKL和OPG的表达

目的检测大鼠正畸牙移动压力侧cathK、RANKL和OPG蛋白表达变化及时间分布特点。方法选用80只6周龄SD雄性大鼠建立正畸牙移动模型,分别在加力后2d、5d、7d、10d和14d各处死16只大鼠。HE染色观察大鼠牙周组织的形态学变化;TRAP染色计数压力侧牙槽骨组织中的破骨细胞数量;免疫组化方法定位及相对定量检测压力侧牙槽骨中cathK、RANKL和OPG蛋白表达变化及时间分布特点。结果压力侧牙槽骨组织中的TRAP染色阳性破骨细胞计数随加力时间的增加而增加,第7d达到高峰,此后逐渐降低;压力侧牙槽骨组织中的cathK、RANKL和OPG蛋白的表达水平均随加力时间的增加而增加,第7d达到高峰,以后均逐渐降低。结论cathK、RANKL和OPG蛋白表达的变化规律与骨改建过程一致,与正畸牙移动骨改建过程中破骨细胞的分化、形成和功能密切相关。     

大鼠正畸压力侧牙槽骨改建中RANKL和OPG mRNA的表达

目的研究破骨细胞核因子kB受体活化因子配基（receptor activator nuclear factor kappa B ligand,RANKL）及其伪受体骨保护因子（osteoprotegerin,OPG）的mRNA在大鼠正畸牙移动压力侧牙槽骨改建中的表达变化及时问分布特点。方法选用80只6周龄SD雄性大鼠建立正畸牙移动模型,分别在加力后2d、5d、7d、10d和14d各处死16只大鼠。HE染色观察大鼠牙周组织的形态学变化;TRAP染色计数压力侧牙槽骨组织中的破骨细胞数量;实时定量PCR方法检测RANKL和OPGmRNA的表达变化及时问分布特点。结果骨改建的最活跃期为正畸加力后的第7d,压力侧牙槽骨组织中的TRAP染色阳性破骨细胞计数随加力时间的增加而增加,第7d达到高峰,而后逐渐降低。压力侧牙槽骨组织中的RANKL和OPGmRNA表达水平均随加力时间的增加而增加,第7d达到高峰,而后均逐渐降低。结论RANKL和OPG mRNA表达的变化规律不仅与骨改建过程一致,而且也与TRAP染色阳性破骨细胞数量的变化规律一致。RANKL和OPG与正畸牙移动骨改建过程中破骨细胞的分化、形成和功能密切相关。     

正畸牙牙根吸收过程中骨膜蛋白与Ⅰ型胶原表达的相关性

目的:探讨大鼠正畸牙牙根吸收过程中破骨细胞及牙周组织中骨桥蛋白(OPN)和Ⅰ型胶原(COL-I)的表达。方法:选取30只健康雄性Wistar大鼠(6～8周龄),建立正畸牙牙根吸收模型，对侧不加力作为对照，分别在加力第1、3、5、7、14和21天后，每组各处死5只大鼠。采用抗酒石酸酸性磷酸酶(TRAP)染色和免疫组化染色分别观察TRAP⁺表达细胞数及OPN和COL-I的表达情况。采用Pearson相关系数检验分析OPN与破骨细胞和COL-I之间的相关性。结果:TRAP染色结果显示，与对照组相比，实验组出现破骨细胞，破骨细胞数量随时间延长呈现先增多后减少的趋势。免疫组化染色结果显示，与对照组相比，实验组OPN表达增多，并且随时间延长OPN表达量逐渐增多，7 d时达到峰值，随后其表达逐渐降低(P<0.05);COL-I的表达呈先降低后增多的趋势(P<0.05)。Pearson相关分析显示，OPN表达与COL-I表达呈负相关(P<0.05)。结论:在正畸牙牙根吸收过程中牙周组织中破骨细胞数呈动态变化，OPN蛋白表达水平与COL-I表达趋势有关。     

正畸牙移动压力侧牙槽骨中cathepsin K、RANKL和OPG mRNA及蛋白的表达

目的检测大鼠正畸牙移动压力侧cathepsinK、RANKL和OPGmRNA及蛋白表达变化及时间分布特点。方法选用80只6周龄SD雄性大鼠建立正畸牙移动模型,分别在加力后2d、5d、7d、10d和14d各处死16只大鼠。HE染色观察牙周组织的形态学变化。TRAP染色计数压力侧牙槽骨中的破骨细胞数量。免疫组化方法定位及相对定量检测cathepsinK、RANKL和OPG蛋白表达变化及时间分布特点。Real—timePCR检测cathepsinK、RANKL和OPGmRNA表达变化及时间分布特点。结果骨改建的最活跃期为正畸加力后的第7d。压力侧牙槽骨中的TRAP染色阳性破骨细胞计数随加力时间的增加而增加,第7d达到高峰,而后逐渐降低。压力侧牙槽骨中的cathepsinK、RANKL和OPGmRNA及蛋白的表达水平均随加力时间的增加而增加,第7d达到高峰,而后均逐渐降低。结论cathepsinK、RANKL和OPGmRNA及蛋白表达的变化规律不仅与骨改建过程一致,而且也与TRAP染色阳性破骨细胞数量的变化规律一致。cathepsinK、RANKL和OPG与正畸牙移动骨改建过程中破骨细胞的分化、形成和功能密切相关。     

正畸牙槽骨改建中RANKL/OPG表达动态变化特征及牙周组织细胞凋亡观察

目的系统研究正畸牙槽骨改建过程中RNAKL/OPG的表达动态变化规律,以及对破骨细胞形成和骨组织改建的影响,同时观察牙周组织中细胞凋亡关键因子的变化特征。方法我们建立小鼠上颌左侧正畸模型,并在正畸开始后第5、7、14和21天进行连续观察,通过Micro CT分析各个时间点第一磨牙移动距离以及远中根压力侧牙槽骨变化。并通过HE染色和TRAP染色检测不同时间点远中根压力侧组织学和破骨方面的变化。进一步通过免疫组化染色和免疫荧光染色检测不同时间点远中根压力侧RANKL、OPG和Caspase3的表达变化。结果在正畸力施加后,小鼠左侧第一磨牙和第二磨牙之间的距离逐渐增加,第一磨牙远中根压力侧BV/TV降低,RNAKL和OPG表达逐渐升高,且在加力第7天RANKL表达出现高峰,随后OPG表达高峰出现在第14天,RANKL先于OPG出现表达高峰,并且伴随出现破骨细胞生成高峰。同时,牙周组织中细胞凋亡数量逐渐增加,牙齿移动明显。结论正畸过程中,RANKL/OPG表达变化呈现时序性特征性变化,同时伴随关键细胞凋亡分子的表达变化,其变化和牙槽骨改建和牙齿移动密切相关。     

核因子-κB受体活化因子配体在大鼠正畸牙压力侧骨改建中的作用

目的：初步探讨在正畸牙移动压力侧核因子-κB受体活化因子配基(receptor activator of nuclearfactor-κB ligand，RANKL)的表达在破骨细胞诱导和骨改建中的调节作用。方法：建立大鼠正畸牙移动模型，利用免疫组化的方法对压力侧RANKL的表达及其变化进行检测；并进一步观察了RANKL对大鼠骨髓破骨细胞形成的影响。结果：正畸牙移动压力侧组化结果显示，RANKL阳性表达位于牙周膜细胞和位于骨陷窝内的破骨细胞中，在正畸牙移动第3、5和7天时呈强阳性表达。体外破骨细胞诱导实验结果显示，在巨噬细胞集落刺激因子(macrophage clone stimulating factor，M-CSF)协同作用下，RANKL呈剂量依赖型诱导TRAP阳性细胞生成。结论：大鼠正畸牙移动中，RANKL在压力侧的表达上调有诱导破骨细胞形成的作用，并导致牙槽骨吸收。     

可溶性白细胞介素-1和肿瘤坏死因子受体对大鼠正畸牙移动影响的研究

目的研究局部注射重组人类可溶性受体对大鼠正畸牙移动进程的影响。方法选取64只雄性SD大鼠,从加力第1天起在大鼠牙齿局部分别注射重组人类可溶性白细胞介素- 1受体Ⅱ（sIL- 1- RⅡ）、可溶性肿瘤坏死因子受体Ⅰ（sTNF- RⅠ）、两者混合物,测量磨牙移动距离,用苏木精- 伊红（HE）染色观察被移动磨牙牙周组织形态学变化,并应用抗酒石酸盐酸性磷酸酶（TRAP）组织化学染色对破骨细胞和破牙骨质细胞的数量及分布的时相性改变进行分析。结果各受体组大鼠磨牙牙周组织形态学时相性变化与对照组相似,但各时间点牙槽骨吸收程度均较轻微,多数大鼠牙根表面很少或几乎没有明显牙骨质吸收迹象。与对照组相比,在第14天所有受体注射组牙槽骨及牙根表面TRAP染色阳性细胞降低了约50%（P<0.05）,但受体组组间差异无统计学意义（P>0.05）。结论在大鼠正畸牙局部注射重组人类可溶性白细胞介素- 1和肿瘤坏死因子受体,可以减少牙齿移动距离并减少牙根吸收。     

人牙周膜成纤维细胞来源外泌体对大鼠延期牙再植术后牙根吸收的调控作用

目的: 探讨健康的人牙周膜成纤维细胞（hPDLFs）来源外泌体对大鼠延期牙再植术后牙根吸收的作用及其可能机制。方法: 分离提取hPDLFs来源的外泌体并鉴定。选择30只6周龄雄性SD大鼠,随机分为对照组和外泌体组,建立上颌第一磨牙延期牙再植术模型,牙脱位30 min后植回牙槽窝。对照组牙周膜局部注射40 μL汉克斯平衡盐溶液（HBSS）,实验组牙周膜局部注射40 μL含外泌体的HBSS。术后1、2、4周收样,苏木精-伊红（H-E）染色观察牙根表面吸收陷窝,抗酒石酸酸性磷酸酶（TRAP）染色观察破骨细胞数量,免疫组织化学染色观察牙周膜内骨保护素（OPG）的表达。采用SPSS 17.0软件包对数据进行统计学处理。结果: 鉴定表明提取的细胞外囊泡为外泌体。与对照组相比,hPDLFs来源的外泌体减少了延期牙再植术后牙根吸收陷窝的数量,降低TRAP阳性破骨细胞的表达（P<0.05）,促进牙周膜内OPG的表达（P<0.05）。结论: 延期牙再植术后,hPDLFs来源的外泌体减少了破骨细胞的数量,促进牙周膜内OPG表达,减少了再植术后的牙根吸收。     

牙周炎大鼠上颌骨来源的BM-MSCs中破骨细胞相关因子的表达

目的 :研究来源于脂多糖(LPS)诱导的牙周炎大鼠上颌骨的间充质干细胞(MSC)表达破骨细胞因子的变化。方法:注射脂多糖构建牙周炎大鼠模型,6周后,处死大鼠,利用Micro CT观察上颌骨骨吸收情况,HE染色观察牙周组织的变化,抗酒石酸磷酸酶(TRAP)染色观察上颌骨内破骨细胞的数量;体外分离和培养大鼠上颌骨来源的MSCs,荧光定量PCR(q PCR)检测两组间充质干细胞中M-CSF、OPG和RANKL m RNA表达量,并计算OPG与RANKL的比值。结果:Micro CT显示实验组大鼠上颌骨牙槽骨吸收明显;HE染色显示实验组切片中牙龈明显退缩,结合上皮根向增殖,并有炎性细胞浸润;TRAP染色显示实验组上颌骨内破骨细胞数量明显增多;荧光定量PCR显示实验组BMMSCs表达的M-CSF增加,表达的OPG和RANKL减少,但OPG与RANKL的比值减小,差异具有统计学意义。结论:来源于LPS诱导的牙周炎大鼠,其颌骨的间充质干细胞在炎症状态下,表达的与破骨相关因子上调,可能有增强破骨细胞的功能,能加速骨吸收进程。     

正畸力诱导Th17/Treg平衡与破骨细胞及其相关调节因子的相关性

目的 初步探讨正畸牙齿移动中正畸力诱导Th17/Treg平衡调节牙齿移动的可能机制。方法 20只SD大鼠随机分为空白对照组（C组，4只）与正畸牙齿移动（CM）组(包括CM3-移动3天、CM7-移动7天、CM14-移动14天、CM21-移动21天，4只/组）。使用镍钛拉簧以50g力近中移动双侧上颌第一磨牙，建立正畸加力模型。流式细胞仪检测各时间节点外周血Th17细胞、Treg细胞比例，免疫组化染色检测各时间节点牙周组织中RANK、RANKL、OPG的表达，TRAP染色及破骨细胞计数。使用SPSS 24.0软件行统计分析。结果 Th17/Treg比值先增加再减少，第7天达到峰值；RANKL/OPG比值先增加后减少，峰值在正畸第7天至第14天；破骨细胞数量先增加后减少,第14天达到峰值。Th17细胞、Th17/Treg比值与RANK、RANKL含量、RANKL/OPG比值、破骨细胞数量正相关，与OPG含量负相关。Treg细胞与RANKL含量正相关。结论 Th17/Treg平衡参于调节正畸牙齿移动。正畸力诱导Th17/Treg平衡偏移可能通过调节RANKL/OPG平衡从而调节破骨细胞引起正畸牙...     

正畸力对大鼠正畸牙根吸收后早期修复的影响

目的：研究大鼠正畸牙根吸收后加力方式的改变对牙根早期修复的影响。方法：选用65只SD大鼠,建立正畸牙根吸收模型,随机分为三个实验组：停止加力组（SF组）、间歇加力组（IF组）、持续加力组（CF组）,另选5只作为空白对照组（NTC组）。采用HE和TRAP染色法对牙根组织形态进行观察。结果：随着停止加力时间的延长,破牙细胞数量明显减少,修复性牙骨质明显增多;IF组与SF组组间牙根吸收相对面积及修复相对面积无明显差异（P〉0.05）;在停止加力第17d组,IF组的TRAP阳性细胞数目及根吸收相对面积明显小于CF组（P〈0.05）。结论：正畸牙根吸收发生后,改变加力方式可影响牙根早期修复,停止加力或停止加力后再加力可降低牙根吸收程度。     

增龄因素对鼠正畸牙牙周组织骨保护素及其配体表达的影响

目的:比较幼年鼠和成年鼠在正畸牙移动中牙周组织骨保护素(OPG)及其配体(OPGL)表达的不同比值,探讨增龄因素对正畸骨改建影响的分子机制。方法:制备大鼠正畸牙齿移动模型,于牙齿移动1d、3d、5d、7d、10d、14d及14d后去除正畸力1周后取材,免疫组化检测牙周组织OPG和OPGL的表达。结果:①.牙齿移动3d时,幼年鼠压力侧OPGL的表达明显增强;而成年鼠此时OPGL的表达没有幼年鼠明显。②.牙齿移动5d和7d时,幼年鼠和成年鼠OPGL的表达均成强阳性,破骨细胞多。③.牙齿移动10d、14d时,幼年鼠和成年鼠OPGL的表达逐渐减弱。④.14d时去除正畸力至21d时发现幼年鼠和成年鼠OPGL均已成弱阳性表达,幼年鼠可见部分成骨细胞。结论:在正畸力的作用下,OPGL与OPG的表达比值与年龄关系密切,增龄因素引起的牙周组织内OPGL/OPG表达变化可能是导致成年正畸特点的分子机制之一。     

Receptor activator of nuclear factor-kappa ligand,OPG, and IGF-I expression during orthodontically induced inflammatory root resorption in the recombinant human growth hormone–treated rats

Objective:To investigate the effects of growth hormone (GH) on local receptor activator of nuclear factor-kappa ligand (RANKL), OPG, and IGF-I expression during orthodontically induced inflammatory root resorption in rats.Materials and Methods:Forty Wistar rats (gender: male; age: 7 weeks) were randomly divided into control and experimental groups. A force of 50 g was applied to move the right upper first molars mesially. The experimental and control groups received daily subcutaneous injections of recombinant human growth hormone (GH; 2 mg/kg) and equivalent volumes of saline, respectively. The rats were sacrificed on days 1, 3, 7, and 14. Micro–computed tomography–reconstructed images of the upper right first molars were used to survey root resorption and tooth movement. Horizontal sections of the maxillae were prepared for hematoxylin and eosin, tartrate-resistant acid phosphatase, and immunohistochemical staining.Results:Resorption lacunae appeared on the compressed side of the distal buccal root of the right first molar on days 7 and 14. Compared with the control groups, GH-treated groups showed more RANKL-positive cells and osteoclasts on day 3 and more OPG- and IGF-I–positive cells and fewer odontoclasts on days 7 and 14. Indexes of root resorption were lower and tooth movement was faster in the GH-treated groups than in the control groups on days 7 and 14.Conclusions:The inhibitory effect of GH on root resorption by heavy force might be mediated by RANKL/OPG and IGF-I. Short-term GH administration may be a method with which to reduce root resorption and shorten treatment time, especially in patients who are susceptible to root resorption.     

基于RANKL-RANK-OPG轴研究丹参素防治牙槽骨骨质疏松的作用

目的:探讨RANKL-RANK-OPG轴在丹参素防治牙槽骨骨质疏松中的作用。方法:SD大鼠随机分为3组:对照组、去卵巢组和丹参素治疗组。实验90 d后取大鼠牙槽骨,HE染色观察牙槽骨组织形态学改变,组织化学染色方法检测牙槽骨组织中TRAP的活性,免疫组化的方法检测牙槽骨组织中RANKL和OPG的表达情况。结果:与去卵巢组相比:丹参素治疗组牙槽骨骨量明显增多,TRAP阳性的破骨细胞数减少,RANKL/OPG减小。结论:丹参素防治牙槽骨骨质疏松可能与其调控RANKL-RANK-OPG轴有关。     

Primary teeth show less protecting factors against root resorption

International Journal of Paediatric Dentistry 2011; Background. Physiological root resorption differentiates primary from permanent teeth. The understanding of what protects and regulates root resorption might help to develop therapies to its control. Aim. To verify the presence and distribution of ECRM and the expression of CK14, OPG, TRAP and COX‐2 in the periodontal ligament (PDL) of human primary and permanent teeth. Design. Eight primary teeth undergoing physiological or pathological root resorption and 4 permanent teeth were immunohistochemically processed for CK14, TRAP, COX‐2 and OPG expression. Results. PDL from primary and permanent teeth showed similar morphological features; however, fewer ECRM clusters and higher immunoreactivity to CK14 were found in primary PDL. In permanent teeth, ECRM were distributed along the entire PDL tissue. Howship′s lacunae were found only in primary teeth, associated with the presence of TRAP‐positive cells and increase in COX‐2 expression. OPG expression in primary PDL was detected in nonresorptive cervical areas and in lacunae showing reparative tissue. It was observed higher expression of OPG in all permanent teeth when compared to primary specimens. Conclusions. It may be concluded that PDL from primary teeth shows less ECRM clusters and lower expression of OPG. These features may be associated with lower protection against root resorption in primary teeth.     

Expression of osteoprotegerin and receptor activator of nuclear factor ��B ligand in root resorption induced by heavy force in rats

Objective

The aim of this study was to investigate the expression patterns of osteoprotegerin (OPG) and the receptor activator of nuclear factor ??B ligand (RANKL) in root resorption during orthodontic tooth movement.

Material and methods

Forty 12-week-old male SD rats were used with the right maxillary side as the experimental group and the left maxillary side as the control group. After 1?N (100g) force was loaded on the right maxillary first molar, the rats were sacrificed on days 0, 1, 4, 8, and 12. Mesial root resorption of the first molar, the number of odontoclasts and osteoclasts, and OPG and RANKL mRNA expression were determined by hematoxylin-eosin and scanning electron microscopy, tartrate-resistant acid phosphate staining, and in situ hybridization, respectively.

Results

Serious root resorption was apparent on the pressure side of the mesial root of the right maxillary first molar on days 8 and 12. The number of odontoclasts in the cementum lacuna was elevated on days 8 and 12. OPG expression rose significantly on the tensile side, while RANKL expression increased on the pressure side. The mRNA level of RANKL was significantly elevated on days 4, 8, and 12. Moreover, the RANKL/OPG mRNA ratio was increased on the pressure side, but decreased on the tensile side.

Conclusion

Changes in the expression of RANKL mRNA and the RANKL/OPG mRNA ratio are accompanied by a parallel alteration in the number of odontoclasts and tooth resorption, suggesting crucial involvement of RANKL and OPG in tooth resorption.     

Histological and immunohistochemical analyses of the chronology of healing process after immediate tooth replantation in incisor rat teeth

Abstract – Dental tissues have special characteristics, and its regenerative capacity is noteworthy. However, understanding the circumstances that lead to regeneration is challenging. In this study, the chronology of the healing process after immediate replantation of rat incisor teeth was examined by histological and immunohistochemical analyses within a 60‐day period. Thirty‐six male Wistar rats had their maxillary right incisors extracted and replanted after 15 min in saline storage. The rats were sacrificed immediately 3, 7, 15, 28, and 60 days after replantation. The histological analysis showed rupture of the periodontal ligament and formation of a blood clot, which started being replaced by a connective tissue after 3 days. At 7 days, the gingival mucosa epithelium was reinserted and areas of root resorption could be seen. At 15 days, the periodontal ligament was repaired. At 3 days, the pulp presented an absence of the odontoblast layer, which started being replaced by a connective tissue. This tissue suffered gradual calcification, filling the root canal at 28 and 60 days. The root ends were closed. The immunohistochemical analysis revealed greater expression of OP, OPG, and RANK proteins in the initial periods (0 and 3 days), while TRAP expression predominated at 28 and 60 days (P < 0.05). In conclusion, in delayed tooth replantation, there is great new bone formation activity in the earlier periods of the repair process, while a predominance of bone resorption and remodeling is observed in the more advanced periods.     

Effects of calcitonin on orthodontic tooth movement and associated root resorption in rats

Objectives: Our main aim was to evaluate the effects of calcitonin (CT) on orthodontic tooth movement (OTM) and orthodontic root resorption in a rat model.

Material and methods: Eighty male Wistar rats were randomly divided into five groups. Rats in the negative control group were not given any appliances or injections. All the remaining rats were used to establish a model of OTM. The positive control group were then injected with normal saline, while rats in the three experimental groups were injected with 0.2?IU, 1?IU or 5?IU/kg/day CT. Nickel-titanium closed-coil springs were used to deliver an initial 50?g mesial force to the left maxillary first molar for 14 days in rats in the positive control group and the experimental groups. Each group was randomly subdivided into two groups, one for analysis of tooth movement, tissue changes and tartrate-resistant acid phosphatase (TRAP)-positive cells in alveolar bone, the other to examine root resorption by scanning electron microscopy.

Results: The OTM distance, the number of force-induced osteoclasts and root resorption areas were significantly decreased in CT-injected rats in a dose-dependent manner.

Conclusions: Administration of CT reduces the root resorption area and may therefore be effective as a novel adjunctive orthodontic approach to diminish undesired tooth movement via enhancing anchorage or preventing relapse after OTM.