应用Herbst矫治器前伸成年大鼠下颌后髁突软骨RUNX2蛋白表达变化的实验研究

目的 通过研究Herbst矫治器引导成年大鼠下颌前伸后髁突软骨RUNX2蛋白表达水平的变化,探讨Herbst矫治器应用于成年患者的可行性.方法 60只14周龄雄性SD大鼠,根据实验周期(3、7、14、21和30 d)分为5组,每组内随机分为实验组(6只)和对照组(6只).实验组大鼠全天佩戴Herbst矫治器引导下颌前伸,对照组大鼠不佩戴矫治器.实验后3、7、14、21和30 d分别处死各组大鼠,取右侧髁突组织,免疫组化检测髁突软骨中RUNX2蛋白的表达水平.结果 随着观测时间的延长,对照组中大鼠髁突软骨RUNX2表达的IOD值呈现递减趋势,但各时间点差异并无统计学意义(P＞0.05)；除3d组外,其余各时间点实验组大鼠髁突软骨RUNX2表达的IOD值均较对照组高(P＜0.05),且在21d时,其IOD值与对照组相差幅度最大.结论 Herbst矫治器能够诱导成年大鼠髁突发生适应性改建.     

磷脂酶C-γ1tyr783在下颌功能前伸大鼠髁突软骨后部表达变化的研究

目的 研究大鼠下颌功能前伸后髁突软骨中磷脂酶C-γ1 tyr783（PLC-γ1 tyr783）表达及其变化规律,探讨 PLC-γ1tyr783在功能矫治前伸大鼠下颌髁突软骨骨改建中的作用,为临床上骨骼矫形治疗提供实验依据。方法 选取4周龄健康雄性Sprague-Dawley（SD）大鼠60只,适应性饲养1周后随机等量分为实验组和对照组,实验组每天24 h佩戴自制下颌前伸斜面导板矫治器,对照组不戴矫治器。2组大鼠分别于戴用矫治器1、3、7、14、21、28 d时各断颈处死5只,取双侧髁突,常规固定、脱钙、脱水、包埋。石蜡切片采用免疫组织化学方法结合图像分析检测PLC-γ1tyr783在髁突软骨中的表达分布及其变化规律。结果 对照组PLC-γ1tyr783表达水平随着实验周期的推移逐渐减弱（P>0.05）,实验组大鼠髁突软骨处PLC-γ1tyr783的表达在第14天时达到高峰,与对照组相比差异有统计学意义（P<0.01）。结论 PLC-γ1tyr783参与了下颌功能前伸大鼠髁突软骨的骨改建。     

前伸下颌后大鼠髁突软骨内明胶酶表达变化的研究

目的研究前伸下颌后明胶酶（MMP- 2、MMP- 9）在大鼠髁突软骨内表达的变化。方法选取5周龄雄性SD大鼠60只,随机等量分成实验组和对照组,实验组大鼠白天配戴自制的上颌斜面导板式活动矫治器引导下颌前伸,对照组大鼠不戴矫治器。在实验的1、2、4周处死动物,免疫组化检测明胶酶在大鼠髁突软骨内表达的变化。结果在正常髁突软骨内,MMP- 2呈中等强度表达,MMP- 9表达很低。前伸下颌后,髁突软骨内MMP- 2的表达无明显变化,而MMP- 9的表达显著增高（P<0.01）。结论明胶酶参与了功能矫形时下颌髁突软骨的适应性改建。     

功能矫治器引导下颌后退后大鼠髁突软骨组织MMP-3及TIMP-1蛋白的表达及意义

目的:探讨功能矫治器引导下颌后退后,髁突软骨组织MMP-3及TIMP-1蛋白的表达及意义.方法:选用4周龄SD雄性大鼠45只,分为实验组20只及对照组25只.实验组模拟临床功能矫治器引导大鼠下颌后退.分别在0 d、3 d、7 d、14 d及21 d全麻下处死实验组及对照组每时间段各5只动物.采用免疫组化方法观察MMP-3及TIMP-1蛋白表达情况.结果:实验组3 d后,髁突软骨MMP-3及TIMP-1表达量逐渐增加,第7 d达高峰,在中后份表达明显增强,肥大细胞层出现较强阳性反应.实验第14 d后,表达逐渐减弱.结论:后退下颌可导致大鼠髁突软骨组织中MMP-3、TIMP-1表达发生变化,MMP-3和TIMP-1参与了髁突软骨组织改建过程,可能在髁突软骨内成骨及其随后的新骨形成中起着重要的作用.     

升高咬合对青春期大鼠髁突软骨中增殖细胞核抗原表达变化的影响

目的：探讨升高咬合后髁突软骨中增殖细胞核抗原（PCNA）的变化情况及意义。方法：40只5周龄雄性SD大鼠,随机分为对照组和实验组（双侧后牙板升高咬合）。分别于术后7、14、21及28d,取其右侧髁突,应用免疫组织化学SABC法检测大鼠髁突软骨中PCNA的表达变化,并作图像分析和统计学处理。结果：与对照组相比,实验组髁突软骨PCNA阳性细胞表达在实验第7、14d明显减少,第28d明显增多（P〈0.05）。结论：咬合升高可以引起大鼠髁突软骨的适应性改建。     

功能矫形前伸大鼠下颌后髁突软骨β转化生长因子-1(TGF-β_1)表达变化的研究

目的：检测快速生长期大鼠髁突软骨在功能矫形前伸下颌后β转化生长因子－１（ＴＧＦ－β１）分布的变化，探讨髁突软骨生长代谢的生长因子控制机理。方法：实验组戴模拟临床功能矫治器，引导大鼠下颌前伸，并在实验后不同时期处死大鼠，取下髁突，应用免疫组织化学方法探测ＴＧＦ－β１在髁突中的含量及分布。结果：髁突各层软骨细胞均有ＴＧＦ－β１的分布，以成熟层阳性强度最高；功能矫形前伸下颌后，髁突软骨细胞各层ＴＧＦ－β１的表达均增强。结论：ＴＧＦ－β１介导了功能矫形的机械刺激作用，使软骨细胞增生，分化功能增强，髁突软骨增生     

导下颌向前对生长期大鼠髁突软骨结缔组织生长因子表达的影响

目的:通过建立导下颌向前的Sprague-Dawley (SD)大鼠的动物模型进行免疫组化实验,观察结缔组织生长因子(CTGF)在髁突软骨中分布的变化,探讨CTGF对前伸下颌后的髁突软骨的影响。方法:建立SD大鼠动物模型,选取45只5周龄SD 雌性大鼠,体重约100 g,随机分为对照组(20只)和实验组(25)只,组内再随机平均分为5组(3、7、14、21 和30 d)。实验组动物24 h佩戴改良可摘式上颌斜面导板矫治器,对照组动物不做处理。免疫组化染色标记CTGF,观察髁突软骨各层CTGF的表达变化。对CTGF的阳性表达量进行半定量分析。结果:对照组和实验组中髁突软骨组织中均有CTGF的表达,主要分布于肥大层和增殖层。实验组不同时间点髁突肥大层和增殖层中CTGF的平均光密度值分别高于对照组的平均光密度值(P<0.05 ) 。与对照组相比,实验组7 d后髁突肥大层和增殖层中CTGF的表达开始上升,到第14天达到最高,之后CTGF表达回落,但仍高于对照组(P<0.05 )。实验3 d、30 d,肥大层和增殖层中CTGF实验组与对照组相比无统计学差异。结论:CTGF可能参与了功能负荷改变所导致的髁突软骨适应性改建活动。     

功能矫形前伸下颌后大鼠髁突软骨雌激素受体的荧光组织化学分析

选用５０只４周龄雌性ＳＤ大鼠，随机等量地分为实验组和对照组，实验组大鼠配戴自制上颌斜面导板式功能矫治器引导下颌前伸，应用荧光组织化学法对大鼠髁突软骨中雌激素受体（ＥＲ）进行定位及半定量分析。结果表明，大鼠髁突软骨中存在ＥＲ，且主要位于胞浆中；ＥＲ在软骨的生发层细胞中分布最多，且髁突软骨增生旺盛时分布较多；功能矫形前伸下颌后，髁突软骨各层细胞中ＥＲ的分布均较对照组明显增加，以生发层细胞的增加最明显。表明ＥＲ与髁突软骨的增生、分化及适应性生长改建关系密切。     

功能性矫治器引导下颌后退对大鼠髁突软骨新骨形成影响的实验研究

目的探讨功能性矫治器引导下颌后退所致的髁突软骨新骨形成的影响。方法选用4周龄SD雄性大鼠45只,分为实验组及对照组,每组5只。实验组模拟临床功能性矫治器引导大鼠下颌后退。分别在0、3、7、14及21 d全麻下处死动物各5只。苏木精-伊红染色观察细胞形态反应,PAS染色观察新骨形成量。结果大鼠在下颌持续后退情况下,实验组髁突后份软骨组织增殖层和成熟层明显变薄,移行区可见破骨细胞增加,此区附近的骨小梁骨沉积减少;实验第3天及第7天,实验组和对照组髁突软骨新骨形成的量有明显区别,第14天和第21天,二者间的差别趋于减小。结论后退大鼠下颌后,髁突软骨后份新骨形成量减少。     

功能矫形前伸下颌对大鼠咬肌TrkB表达影响的研究

目的：探讨功能矫形前伸大鼠下颌对青春期大鼠咬肌酪氨酸激酶受体-B（Tyrosine kinase receptor-B,TrkB）表达的影响。方法：将40只4周龄雄性SD大鼠随机分为实验组和对照组各20只,实验组大鼠佩戴自制的上颌功能矫治器,对照组不戴,分别在实验第3、7、14、21 d处死大鼠,通过免疫组织化学染色和RT-PCR检测两组大鼠咬肌中TrkB及其mRNA的表达。结果：实验3 d组、7 d组免疫组化阳性染色和mRNA的表达虽有增强,但与对照组相比,均无明显变化;实验14 d组、21 d组TrkB免疫组化阳性染色明显强于对照组,其mRNA表达结果亦同样高于对照组。结论：随着功能矫形前伸下颌时间的延长,TrkB表达含量逐渐增加,TrkB参与了咬肌在下颌前伸中的适应性改建活动。     

Neovascularization and bone formation in the condyle during stepwise mandibular advancement

The aims of this investigation were to identify the temporal expression of vascular endothelial growth factor (VEGF) in the mandibular condyle and to correlate it with the pattern of new bone formation during stepwise mandibular advancement. Two hundred and fifty female, 35-day-old Sprague-Dawley rats were randomly divided into 10 groups, with 10 rats allocated to the single-step bite-jumping subgroup, 10 rats to the stepwise advancement subgroup and five rats to the control subgroup. In the experimental groups, the mandibles were kept in a continuous forward position. The initial stepwise advancement commenced on day 35, whereas the second advancement started on day 65. The rats were sacrificed on experimental days 3, 7, 14, 21, 30, 33, 37, 44, 51 and 60. Sections (7 microm) were cut through the condyle in the parasagittal plane and stained with anti-VEGF antibody. Each section was counter-stained with haematoxylin for observation of the cellular response. The sections were digitized and quantitatively analysed with a computer-assisted image analysing system. The results showed that the initial advancement in the stepwise group led to significantly less expression of VEGF when compared with single advancement. However, the second advancement on day 30 resulted in a significant increase in VEGF expression when compared with the one-step group and the natural growth control group. Thus, it was concluded that changes in the amplitude of mechanical loading, produced by stepwise advancement, have a significant effect on the production of VEGF by the chondrocytes. During the later stages of advancement, more VEGF and more condylar bone was produced.     

The correlation of replicating cells and osteogenesis in the condyle during stepwise advancement

The aim of this study was to quantify the number of replicating mesenchymal cells and to correlate it to the amount of bone formation in the condyle during stepwise advancement of the mandible. Two hundred and fifty female Spraque-Dawley rats, 35 days old, were randomly divided into 10 control groups (n = 5) and 20 experimental groups (n = 10). Fifty rats from the stepwise experimental group relieved a two-mm advancement initially and veneers were added on day 30 with another 1.5 mm advancement. The rats were sacrificed after 3, 7, 14, 21, 30, 33, 37, 44, 51, and 60 days. One hour before death, all rats were injected with bromodeoxyuridine (BrdU) intravenously. Tissue sections of seven microm were cut through the condyle in the sagittal plane and stained with anti-BrdU antibody to evaluate the number of replicating mesenchymal cells. Haematoxylin stain was applied to observe cellular response. The results indicated that during the first advancement, replicating mesenchymal cells in the posterior region of the condyle showed the highest increase on days 7 and 14 when compared with the control. Such an increase preceded the highest level of bone formation between days 30 and 37 of advancement. In response to the second advancement, another increase of replicating cells was evident on day 44, along with a significant increase in bone formation observed on day 60. We concluded that forward positioning of mandible in a stepwise manner delivers a mechanical strain that solicits an increase in the number of replicating mesenchymal cells in the condyle. The increase in the population size of the osteoprogenitor cells subsequently leads to more bone formation.     

Replicating mesenchymal cells in the condyle and the glenoid fossa during mandibular forward positioning.

The purpose of this study was to identify and quantify the temporal sequence of replicating mesenchymal cells during natural growth and mandibular advancement in the condyle and the glenoid fossa. One hundred fifty 35-day-old female Sprague-Dawley rats were randomly divided into 10 experimental groups (10 rats each) and 10 control groups (5 rats each). The experimental groups were fitted with appliances that positioned the mandible forward. One hour before the rats were killed, bromodeoxyuridine (BrdU) was intravenously injected into them. Sections were cut and stained with anti-BrdU antibody to evaluate the number of replicating mesenchymal cells. Cellular uptake of BrdU was quantified with the Leica Qwin (Leica Microsystem Imaging Solutions, Cambridge, United Kingdom) system. The results showed that the numbers of replicating mesenchymal cells during natural growth were highest in the posterior region of the condyle and the anterior region of the glenoid fossa. In the experimental groups, the posterior region had the highest number of replicating cells for both the condyle and the glenoid fossa, with the condyle having 2 to 3 times more replicating cells than the glenoid fossa. The number of replicating mesenchymal cells, which is genetically controlled, influences the growth potential of the condyle and the glenoid fossa. Mandibular protrusion leads to an increase in the number of replicating cells in the temporomandibular joint. Individual variations in the response to growth modification therapy could be a result of the close correlation between mesenchymal cell numbers and growth.     

Osteogenesis in the glenoid fossa in response to mandibular advancement.

The purpose of this study was to identify the temporal sequence of cellular changes in the glenoid fossa and to quantify the amount of bone formation in response to mandibular advancement. One hundred 35-day-old female Sprague-Dawley rats were randomly divided into 5 experimental groups (15 rats each) and 5 control groups (5 rats each). In the experimental groups, functional appliances were used to create continuous forward mandibular advancement. The rats were killed after 3, 7, 14, 21, and 30 days. Sections were cut through the glenoid fossa in the parasagittal plane and stained with periodic acid and Schiff's reagent for evaluation of bone formation and with hematoxylin and eosin for observation of cellular response. The results showed that, in the control rats, bone formation was initially higher in the posterior and middle regions than in the anterior region then decreased over time in all regions. In the experimental group, bone formation significantly increased from day 7 to day 30 compared with control rats. Day 21 marked the highest levels of bone formation in the middle (+184%) and posterior regions (+300%). Mandibular protrusion resulted in the osteoprogenitor cells being oriented in the direction of the pull of the posterior fibers of the disc and also resulted in a considerable increase in bone formation in the glenoid fossa.     

Vascular endothelial growth factor expression and bone formation in posterior glenoid fossa during stepwise mandibular advancement.

This study assessed the amount of vascular endothelial growth factor (VEGF) expression and related the findings to new bone formation in the posterior glenoid fossa during stepwise mandibular advancement. A total of 250 female Sprague-Dawley rats, 35 days old, were randomly divided into 10 groups, each including 5 control and 20 experimental rats. Within each group, 10 experimental rats were fitted with functional appliances with a 1-step advancement of 3.5 mm. Another 10 were fitted with stepwise appliances with an initial advancement of 2 mm and a subsequent increase to 3.5 mm on day 30. The rats in the experimental groups were killed on days 3, 7, 14, 21, 30, 33, 37, 44, 51, and 60, respectively. The matched controls were killed on the same time points. Sections (7 microm) were cut through the glenoid fossa sagittally and stained with anti-VEGF antibody. VEGF expression in the posterior glenoid fossa was evaluated with a computer-assisted image-analyzing system. Both VEGF expression and new bone formation were greater in the experimental rats than in the controls. During stepwise advancement, initial VEGF expression was less than that of 1-step advancement, but the second advancement elicited another peak on day 44. New bone formation was also less than that of 1-step advancement during early stages of stepwise advancement but then began to increase from day 37 onward. The maximum increase was observed on day 60. Stepwise advancement of the mandible delivers mechanical stimuli that produce a series of tissue responses that lead to increased vascularization and bone formation.     

The correlation between neovascularization and bone formation in the condyle during forward mandibular positioning

The aim of the present study was to investigate the temporal pattern of expression of VEGF (Vascular Endothelial Growth Factor) and new bone formation in the condyle during forward mandibular positioning. The importance of vascularization during endochondral ossification was investigated during natural growth of the condyle and compared to that after forward mandibular positioning. The goal was to further our understanding of the cellular responses during functional appliance therapy with a view to extending the experiment into maturity. One hundred and fifty 35 days old Sprague-Dawley rats, 100 fitted with a bite-jumping appliance and 50 untreated, were divided into 10 groups. One group was sacrificed on each of experimental days 3, 7, 14, 21, 30, 33, 37, 44, 51 and 60 respectively. Sagittal sections were cut and stained with VEGF antibodies and Periodic acid and Schiff's reagent (PAS). Each section was quantitatively analyzed with a computer assisted analyzing program and the temporal sequence of expression of VEGF and new bone formation during natural growth and after mandibular forward positioning was compared. There was significant increase in both vascularization and mandibular bone growth upon forward mandibular positioning and the highest amount of both were expressed in the posterior region of the condyle. The highest acceleration of vascularization preceded that of new bone formation. Thus, forward mandibular positioning was found to solicit a sequence of cellular events leading to increased vascularization and subsequently new bone formation resulting in enhanced condylar growth.     

Forward mandibular positioning enhances condylar adaptation in adult rats

The aim of this investigation was to assess quantitatively the adaptive changes in the condyles of adult rats to forward mandibular positioning. The level of types II and X collagen expressed in the condyles of adult rats was compared with that formed in response to forward mandibular positioning and the levels of expression were correlated to the amount of bone formed in response to mandibular advancement. Seventy-eight 120-day-old female Sprague-Dawley rats were included in this study. The rats were randomly allocated to six groups. Each group consisted of nine rats with bite-jumping devices and four untreated controls. The animals in each group were sacrificed on days 3, 7, 14, 21, 30, and 60. Immunostaining was used for the detection of types II and X collagen, while Alcian blue-PAS was used to observe the extracellular matrix and new bone formation. The results showed that new cartilage was formed in the posterior condyle. The highest level of expression of types II and X collagen were present on day 21, the amount of increase was 247.99 and 540.08 per cent, respectively. The highest level of new bone formation was measured at day 30 of advancement when the amount of increase in new bone formation was 318.91 per cent. These findings indicate that forward mandibular positioning causes changes in the biophysical environment of the temporomandibular joint (TMJ) of adult rats that leads to condylar adaptation.     

不同年龄段小鼠下颌前导后髁突骨骼改建差异的研究

目的：：观测小鼠间歇性下颌前导后髁状突的形态学变化,比较不同年龄段的组织改建差异,为下颌前导的矫治时机和策略提供理论依据。方法：在4周龄和8周龄BABL/c雌鼠上建立间歇性下颌前导模型,于下颌前导后的14 d和28 d取颞下颌关节标本行Trap染色、Godener三色染色、微型计算机断层扫描( Micro-CT)技术来研究髁突破骨细胞及成骨细胞变化和骨骼形态学变化。结果：28 d时4周龄组相对于28 d时8周龄组,骨体积分数( BV/TV)和骨小梁厚度( TB. Th)减小,骨小梁间距(TB. Sp)增大(P<0.05)。骨小梁数量(TB. N)无明显差异(P>0.05)。14 d时4周龄组相对于14 d时8周龄组以及14 d时4周龄组相对于28 d时4周龄组,每视野中破骨细胞和成骨细胞数升高(P<0.05)。结论：小鼠导下颌向前与髁突骨骼改建有密切联系,不同年龄段差异显著。