牵张成骨矫治猕猴腭部软硬组织缺损的组织学及荧光标记研究

目的 观察猕猴牵张成骨术整复腭裂过程中,新骨组织形成与改建活动的特点,探讨新骨生成的规律.方法 建立猕猴腭裂动物模型,以牵张成骨术整复其腭部软硬组织缺损,以每天2次、每次0.4 mm的速率牵引,直至骨运送盘移动关闭裂隙后原位固定.分别于固定期第1、2、4、6、8、12及24周取材,每时相点3只实验动物.取材前6 d,实验组动物(21只)均肌肉注射四环素(30 mg/kg)标记.各组标本包埋切片行组织学及荧光标记观察.结果 与实验对照组及空白对照组(动物各2只,同法标记)对比.结果 实验组骨运送盘移动至裂隙对侧关闭缺损,牵开间隙内以膜内成骨方式原位成骨,成功整复腭部软硬组织缺损.牵张成骨术后固定期从1至24周逐渐延长,呈现出成骨-改建-成熟的动态变化过程:牵张区新骨形成的量及其钙化程度不断增加,与此同时,新骨组织亦呈现不断改建成熟的趋势,软组织随着新骨的形成不断地延伸,对照组裂隙无法自行修复.结论 应用牵张成骨术整复腭裂缺损,在良好的固定下,通过膜内成骨的方式,新骨不断的成熟和改建,直至成功修复软硬组织缺损.     

牵张成骨术整复猕猴腭裂后胰岛素样生长因子-1与碱性磷酸酶表达研究

目的 以牵张成骨术整复猕猴腭裂骨缺损,定量分析不同时段新生成骨胰岛素样生长因子-1(insulin.1ike growth factor-I,IGF-1)与碱性磷酸酶(alkaline phosphatase,ALP)表达水平,探讨其成骨调控机制.方法 用猕猴建立腭裂动物模型.实验组动物21只以牵张成骨术整复其腭部软硬组织缺损,关闭裂隙后固定.于牵张成骨术后第1、2,4…6 8 12及24周分别取材,各3只动物.采用实时定量PCR法分析比较IGF-I与ALP的mRNA表达水平,并以酶联免疫吸附试验法(ELISA)定量分析其IGF-I与ALP含量,结果与实验对照及健康对照组(动物各2只)进行比较.结果 固定期第1～2周为成骨早期,第1周时IGF-1和AIJP的mRNA表达明显上调,分别为3.67±0.35和3.30±0.21,第2周时达最高峰,分别为7.55±0.32和5.91±0.21,随后逐渐下降,至第12周与健康对照组无明显差异(P>0.05).ELISA结果显示:固定期第1～2周,IGF-1和ALP表达均增强.IGF-1含量于第2周表达最高,为(2.0±0.06)ng/mg,第4周为(1.46±0.08)ns/mg,第6周为(0.84±0.11)ng/mg,其表达逐渐下降;同期ALP则呈高水平表达,第4周为(25.34±0.44)U/mg,第6周为(26.21±0.82)U/mg.第8～12周,IGF-1和ALP的表达均下降至接近健康对照组.结论 腭骨牵张成骨区域,新骨的原位增量生成明确,增殖过程正常,最终以膜内成骨的方式形成新骨整复了腭裂骨切开牵张间隙区域.     

牵张成骨术整复猕猴腭裂骨桥蛋白与骨钙蛋白表达研究

目的 以牵张成骨术整复猕猴腭裂骨缺损,定量分析新骨在不同时期骨桥蛋白(osteopetin,OPN)与骨钙蛋白(osteocalcin,OC)的表达水平,探讨新骨生成与改建的规律.方法 以猕猴为对象建立腭裂动物模型.实验组动物21只行牵张成骨术整复其聘部软硬组织缺损,关闭裂隙后固定.固定期第1、2 4、6、8、12及24周分别取材,各3只动物.采用实时定量PeR法(real-time,RT-PCR)定量比较OPN与OC的mRNA表达水平,并以酶联免疫吸附试验法(ELISA)定量分析其OPN与OC含量,与实验对照组及健康对照组(各2只动物)结果进行比较.结果 固定期第2周OPNmRNA表达上调,第4周达最高(7.59±0.37);而OC mRNA表达则自第4周开始上调(4.98±0.21),第6周时达最大值(7.94±0.31);随后开始下降,至第24周时两者的mRNA表达水平接近健康对照组(P>0.05).ELISA结果显示:固定期第4、6周OPN分别为(4.75±0.15)ng/mg和(4.86±0.09)ng/mg,OC分别为(3.18±0.16)ng/mg和(3.63±0.33)ng/mg,两者均为高水平表达.至第8～12周以后蛋白表达趋势与其对应mRNA表达基本一致.结论 应用牵张成骨术整复腭裂骨缺损,其牵张区域新骨生成,裂隙被骨运送盘移动封闭,腭部裂隙被完全修复.     

应用牵张成骨技术修复腭裂的动物实验研究

目的:将DO技术应用于腭裂硬腭裂隙关闭,以探索一种硬腭部分成骨性修复的新方法,从而达到腭裂功能性整复的目的。方法:以1-1.5岁杂种大7只为研究对象,建立人工腭裂的动物模型。2只为对照组,只在硬腭后部形成8mm×30mm全层洞穿缺损裂隙,5只在形成缺损的同时安置DO装置,并形成骨转移盘,术后12天开始以每次0.3mm,每日2次向裂隙一侧牵张移动骨转移盘,至硬腭部软硬组织裂隙完全关闭,10周后处死动物行大体标本观察照相及X线摄片。结果:对照组10周后处死观察裂隙大小形态与手术时无差别,实验组DO装置固定牢靠、无松脱、顺利完成牵张过程,硬腭部裂隙完全关闭,骨标本及X线片示牵张区完全为新生骨组织取代,骨质厚度约0.5-1.0mm。结论:自行设计的牵张器具有稳定可靠的牵张作用,经牵张成骨能成功地完成硬腭缺损骨性修复,从而为腭裂修复提供了一种新的途径。     

腭骨外侧缝牵张成骨闭合硬腭后部裂(动物实验研究)

目的探索腭裂修复新方法,采用缝牵张技术诱导犬腭骨外侧缝骨再生,补偿腭裂骨组织缺损,形成硬腭后部的骨性修复,避免传统手术存在的语音效果较差和面部发育障碍的不足。方法用8周龄杂种犬9只,实验组经手术形成硬腭8mm 宽的裂隙。用牵张力分别为200g,360g和480g 的镍钛形状记忆合金(NiTi-SMA)缝牵张器牵张腭骨外侧缝,使两侧腭骨向中线移动关闭裂隙。并设实验对照组和空白对照组。结果实验对照组腭裂比手术时略有扩大;实验组则先后完全闭合,组织学发现实验组缝的骨缘有明显新骨形成。结论幼犬腭裂可以通过缝牵张诱导骨组织再生补偿缺损,形成骨性修复;NiTi-SMA 缝牵张器具有稳定可靠的牵张作用;三种不同牵张力值均能诱发缝区大量新骨形成,牵张力值较大者形成的新骨骨髓腔较大,骨小梁细长。     

腭骨外侧缝牵张成骨闭合硬腭后部裂(动物实验研究)

目的探索腭裂修复新方法,采用缝牵张技术诱导犬腭骨外侧缝骨再生,补偿腭裂骨组织缺损,形成硬腭后部的骨性修复,避免传统手术存在的语音效果较差和面部发育障碍的不足。方法用８周龄杂种犬９只,实验组经手术形成硬腭８ｍｍ宽的裂隙。用牵张力分别为２００ｇ,３６０ｇ和４８０ｇ的镍钛形状记忆合金（ＮｉＴｉ－ＳＭＡ）缝牵张器牵张腭骨外侧缝,使两侧腭骨向中线移动关闭裂隙。并设实验对照组和空白对照组。结果实验对照组腭裂比手术时略有扩大;实验组则先后完全闭合,组织学发现实验组缝的骨缘有明显新骨形成。结论幼犬腭裂可以通过缝牵张诱导骨组织再生补偿缺损,形成骨性修复;ＮｉＴｉＳＭＡ缝牵张器具有稳定可靠的牵张作用;三种不同牵张力值均能诱发缝区大量新骨形成,牵张力值较大者形成的新骨骨髓腔较大,骨小梁细长。     

牵张成骨关闭牙槽突裂的实验研究及组织学观察

目的:应用牵张成骨技术进行牙槽突裂关闭术,观测牙槽突裂硬软组织修复效果和牵张间隙新骨生成的过程、机制,探索一种牙槽突裂整复治疗的新途径。方法:以10只成年杂种犬为实验对象,建立人工上颌牙槽突裂的动物模型,其中2只为对照组。另8只为实验组,以牙骨复合体作为转运盘,以每次0.4mm,2次/天的速度沿牙弓方向行牵张成骨术,直到关闭硬组织裂隙。于原位固定0,14,28,63天分别处死动物各2只,对标本进行X线摄片、大体和组织学观察。结果:利用牵张成骨术成功地进行牙槽突裂整复术,硬组织裂隙关闭, 同时软组织得到扩张;骨牵张间隙完全被新生骨组织取代并沿牵张方向生长,随固定时间的延长改建、成熟。结论:牵张成骨术提供了牙槽突裂治疗的新途径,新骨的生成是膜内成骨方式。     

腭骨外侧缝牵张成骨闭合硬腭后部裂

目的 探索腭裂修复新方法,采用缝牵张技术诱导犬腭骨外侧缝骨再生,补偿腭裂骨组织缺损,形成硬腭后部的骨性修复,避免传统手术的语音效果较差和面部发育障碍的不足。方法 用８周龄杂种犬９只,实验组经手术形成硬腭８ｍｍ宽的裂隙。用牵张力分别为２００ｇ、３６０ｇ和４８０ｇ的镍钛形状合金缝牵张器牵张腭骨外侧缝,使两侧腭骨向中线移动关闭裂隙。并设实验对照组和空白对照组。结果 实验对照组腭裂比手术时略有扩大;实验组     

腭横缝牵张成骨延长硬腭的实验研究

目的传统腭裂软组织后推延长手术由于瘢痕挛缩和缺乏骨性支持而难以维持软腭的正常长度、位置和发育,手术损伤和瘢痕还导致面部生长障碍。本研究旨在用缝牵张技术延长硬腭,以避免传统手术存在的语音和面部发育问题。方法 8周龄杂种犬6只,腭骨内种置汞齐标记,用 NiTi-SMA 弓形牵张器前后方向牵张一侧腭横缝。维持8周。经临床,X 线,干骨观察缝牵张后变化。结果 4周时腭横缝牵张达到预定限度。X 线片显示,牵张侧腭横缝前后两点汞齐标志的间距增加1.5cm;前颌在牵张早期显著前突,随后逐渐回缩至两侧平齐。9个月头颅干骨发现,牵张侧腭骨后缘明显后移,上颌骨前移,鼻腭孔至硬腭后缘的距离较对侧增加4.6～5.9mm。结论腭横缝牵张可以造成永久性腭骨后退和上颌前移。     

腭横缝牵张成骨延长硬腭的实验研究

目的传统腭裂软组织后推延长手术由于瘢痕挛缩和缺乏骨性支持而难以维持软腭的正常长度、位置和发育,手术损伤和瘢痕还导致面部生长障碍。本研究旨在用缝牵张技术延长硬腭,以避免传统手术存在的语音和面部发育问题。方法８周龄杂种犬６只,腭骨内种置汞齐标记,用ＮｉＴｉＳＭＡ弓形牵张器前后方向牵张一侧腭横缝,维持８周。经临床,Ｘ线,干骨观察缝牵张后变化。结果４周时腭横缝牵张达到预定限度。Ｘ线片显示,牵张侧腭横缝前后两点汞齐标志的间距增加１５ｃｍ;前颌在牵张早期显著前突,随后逐渐回缩至两侧平齐。９个月头颅干骨发现,牵张侧腭骨后缘明显后移,上颌骨前移,鼻腭孔至硬腭后缘的距离较对侧增加４６～５９ｍｍ。结论腭横缝牵张可以造成永久性腭骨后退和上颌前移。     

Distraction osteogenesis for cleft palate closure in a canine model

OBJECTIVE: To assess the utility of distraction osteogenesis (DO) when applied to closure of a hard palate cleft in dogs. METHODS: A midline hard palate cleft was created in 10 mature dogs. Two were controls and had no distraction; the other 8 dogs underwent osteotomies with installation of customized DO devices to the hard palate. After a 10-day latency, distraction commenced at 1 mm/d. After a 14-day consolidation period, the device was removed and the mucosa closed. Each dog was injected with fluorochrome labels and serially killed at 2-week intervals. Bone healing was analyzed further with traditional histologic analysis and fluorochrome labeling. RESULTS: No serious complications occurred. Bone resorption and cleft widening occurred in both control dogs. Complete bone closure of the hard palate cleft was achieved with DO in 5 of 8 experimental dogs. Three experimental dogs had bone resorption and incomplete palatal closure. CONCLUSIONS: The application of DO techniques in closure of a hard palate cleft in a canine model is safe and well tolerated. Furthermore, in some cases, it proved effective in achieving bony closure of the cleft. Further investigation is warranted into innovative use of DO in treating children born with cleft palate.     

Bone formation and remodeling after symmetric and asymmetric physeal distraction

Bone formation after symmetric and asymmetric physeal distraction was studied in growing sheep. After gradual distraction by external fixation, separation of the growth plate occurred. Fluorescence microscopy, microradiography, and histomorphometric methods were used to study neometaphysis formation. The bone was formed from the inner layer of the periosteum to encircle the distraction area, where the osteogenesis occurred according to the collagen bundles. At consolidation, the distraction area was composed of lamellar trabecular and partly woven bone. The bone trabeculae in the distraction area were thicker than in the control radius. After asymmetric physeal distraction, spontaneous correction of the formed angulation occurred through remodeling and asymmetric physeal growth. Although local physeal changes were noted histologically, enchondral growth usually continued after distraction.     

Midface Membranous Bone Lengthening: A One-Year Histological and Morphological Follow-Up of Distraction Osteogenesis

Midface bone lengthening was performed on three young, adult sheep using distraction osteogenesis following osteotomy of the maxilla and mounting of an extraoral fixation device. The midface was gradually distracted, 2 mm/day, for 21 days, up to approximately 40 mm. A marked midface advancement was noted. Following a further 6 weeks of retention, the device was removed and the animals were followed for 1 year. Biopsies specimens were taken from the distracted area at the end of the distraction period, after the additional 6 weeks of retention, and finally 1 year later. A nondistracted area of the maxillary bone served as control. The specimens were analyzed histologically, histochemically, and by scanning electron microscopy for the ultrastructural pattern, mineralization, mineral content, and approximate Ca²⁺ concentration. Clinically and radiographically, all sheep fully bridged the experimental gap. Histologically, at the completion of distraction, collagen bundles and slender bone trabeculae oriented in the direction of the distraction could be seen. At the end of the retention period, the trabeculae thickened noticeably and were partially replaced by mature lamellar bone. At the end of 1 year and after completion of the process of remodeling, the pattern of the distracted area resembled the control area. The mineralization, as reflected by quantitative calcium analysis, compared with the nondistracted area, demonstrated a low rate of mineralization after 3 weeks of lengthening, increased 6 weeks later, and after 1 year became nearly the same as in the nondistracted area. In conclusion, distraction osteogenesis provides satisfactory quantitative and structural new bone. Received: 23 September 1996 / Accepted: 17 June 1997     

rhBMPs对山羊下颌骨较大速率DO成骨的影响

目的研究在下颌骨较大速率牵引成骨术(DO)中应用重组人骨形成蛋白(rhBMPs)对成骨的影响,探讨下颌骨较大速率DO的可行性。方法12只山羊双侧下颌骨行DO,山羊及左右下颌骨随机化分为实验侧、对照侧,一只山羊作为正常标本。术中预置牵开间隙3.5mm,实验侧应用rhBMPs,延迟期为3d,牵引速率为1.5mm/d,分3次牵引,延长幅度为20mm。3、5、7、12周行大体、X线、骨密度、生物力学和组织学观察。结果延长幅度达到20mm,自固定期3周开始观察,牵引间隙新骨的再生形成及成熟改建仍是连续的骨愈合过程。固定期5周时,大体及X线观察表明实验侧已形成良好的骨皮质连续性,密度近于正常骨,而对照侧牵引间隙中间区却存在骨不连或间隙存在。组织学显示DO过程仍然是膜内成骨,对照侧牵引间隙中央可见中央区纤维连接、软骨改变,周边区可见新骨组织形成。生物力学三点弯曲实验显示实验侧在5周时接近正常骨,与对照侧有显著差异。骨密度随时间递增,实验侧在各时间点均高于对照侧。本实验条件下在5周后拆除牵引器较为可靠。结论较大速率DO的骨再生形成也是一个连续的膜内成骨过程。rhBMPs在较大速率和较大幅度DO中的应用能够加速骨的再生和愈合速...     

电穿孔介导的基因治疗对兔下颌骨牵引区新生骨骨密度与强度的影响

目的 探索电穿孔介导的基因治疗对下颌骨DO过程中牵引间隙新生骨密度与强度的影响,从而为促进下颌骨DO新骨生成,缩短牵引周期,减少并发症提供新思路.方法 以新西兰大白兔为实验动物模型,于术后3 d开始下颌骨牵引,每天0.8 mm,连续牵引7 d后,将实验动物分为5组.A组:在牵引区注射2 μg(O.1μg/μl)重组质粒pIRES-hVEGF165-hBMP2;B组:在牵引区注射2 μg(0.1μg/μl)重组质粒pIRES-hBMP2;C组:在牵引区注射2 μg(0.1μg/μl)重组质粒pIRES-hVEGF165;D组:在牵引区注射2 μg(0.1μg/μl)空质粒pIRES;E组:在牵引区注射相同剂量的生理盐水.5组实验动物均施加电穿孔刺激.各组分别于固定期第1、2、4、8周行X线及QCT检查.选整个牵张间隙新生骨痂部分为兴趣区,测定骨密度.然后处死动物.取材测量牵引区新生骨的三点抗压强度.结果 A、B、C组新生骨痂密度各时相点新生骨痂密度明显高于D、E组(P<0.01).固定2周,A组明显高于各组,但B、c组间比较差异无统计学意义.固定4周,A、B组明显高于C、D、E组(P<0.01).固定8周A组明显高于B、c、D、E组(P<0.01).B、C组间比较差异无统计学意义,但高于D、E组(P<0.01).固定4周,A组新生骨的三点抗压强度明显高于B、C、D、E组(P<0.01).固定8周,A组仍明显高于各组(P<0.01),且B组也明显高于c、D、E组(P<0.05).结论 电脉冲介导的pIRES-hVEGF165-hBMP2重组质粒体内转染可使牵引区获得较满意的骨再生和骨化成熟进程,其新骨骨化、改建过程均超过对照组.提示联合应用BMP与VEGF,可能会实现成骨与血供的联合重建,并且使单一生长因子的效应放大,使骨愈合的速度加快.