血管内皮生长因子对骨折愈合相关因子表达的调控

目的探讨应用和拮抗血管内皮生长因子(VEGF）对骨折愈骨相关因子表达的影响，并观察骨折愈台过程中的病理改变。方法用105只天上I=白兔制作骨折模型．随机分为对照组、应用VECF组和拮抗vEGF组，分别于伤后8、24、72b和1．3、5、8周测定各组动物骨折端相关因子的表达变化、同时取骨折端标本脱钙进行兜镜观察。结果应甩VEGF使骨折端埘BMP的表达时同提前和延良，拈抗VEGF抑制rBMP的表达，应用VEGF组伤后3J嗣时骨折端充填有纤维性骨痴，软骨骨痂和骨性骨痂．5周时新生骨以编织骨为主，8周时骨折正常愈合；括抗VH押导受伤后初期骨细胞坏死增多，1～5周各时相点骨折端均有灶性坏死出现，3同时骨折部位血管生成明显减少。结论骨折愈台过程需要多因子参与、协调，VEGF有可能是作为骨折愈台过程中的一种重要因子在发挥作用     

骨折愈合过程中血流量变化与VEGF的相关性研究

目的：探讨骨折愈合过程中血管内皮生长因子（Vascular endothelial growth factor,VEGF)对骨折端血流量变化的影响。方法：大耳白兔105只，随机分对照组，VEGF组抗VEGF组，各线于伤后8、24、72H,1,3,5,8周利用单光子放射计算机断层显像术（single photon emission computerized tomography,SPECT)测定骨折端血流量变化。结果：应用VEGF使骨折端血流量在一定时间内较对照组增高；抗VEGF可导致骨折端血流量明显减少，结论：VGEF对骨折端血压血流量变化可能影响骨折愈合，有重要意义。     

仙灵骨葆胶囊联合阿仑膦酸钠对骨质疏松骨折大鼠骨痂血管形成的影响

目的探讨仙灵骨葆胶囊联合阿仑膦酸钠对骨质疏松性骨折大鼠骨痂血管形成及VEGF、BMP-2表达的影响。方法50只雌性SD大鼠随机分为假手术组(SHAM组)、模型组(MODEL组)、仙灵骨葆组(XLGB组)、阿仑膦酸钠组(ALLSN组)、联合药物组(LHYW组),10只/组,构建骨质疏松性骨折大鼠模型,放射性X线观察评估骨折愈合,双能X线检测骨密度,Mirco-CT检测骨结构形态学参数,番红O固绿染色观察骨痂组织形态学,免疫组化检测骨痂VEGF和BMP-2蛋白表达。结果所有实验大鼠均进入结果分析。与SHAM组比较,MODEL组大鼠骨折愈合评分、骨密度、骨组织形态学参数、骨痂VEGF和BMP-2表达均显著降低(P0.05),与MODEL组比较,XLGB组、ALLSN组和LHYY组骨折愈合评分、骨密度、骨组织形态学参数、骨痂VEGF和BMP-2表达均显著升高(P0.05),尤以LHYY组最高。SHAM组骨小梁结构正常,几乎均为骨性骨痂。MODEL组骨小梁明显稀疏、断裂,未见明显骨性骨痂。XLGB组和ALLSN组骨小梁增多,排列稍紊乱,大部分为骨性骨痂。LHYW组骨小梁明显增多,排列密集整齐,大量骨性骨痂。结论仙灵骨葆胶囊联合阿仑膦酸钠可能通过介导提高骨质疏松性骨折大鼠骨生长因子VEGF和BMP-2表达,促进骨痂血管形成,加速骨痂形成,增加骨密度,改善骨结构形态,促进骨折愈合。     

The effects of cryopreserved human amniotic membrane on fracture healing: Animal study

ObjectiveThe aim of this study was to investigate the effects of human amniotic membrane (HAM) on fracture healing in an animal model.MethodsStandard tibial diaphysial fractures were created in twenty-eight Wistar-Albino rats and treated with intramedullary Kirschner wire (K-wire) and HAM (HAM (+) group) or K-wire only (HAM (–) group). Fracture healing was evaluated by histological analysis, radiologic X-ray views and callus diameter measurements at 3rd and 6th weeks postoperatively.ResultsFracture healing was histologically better in the HAM (+) group and the difference was statistically significant at both 3rd and 6th weeks postoperatively (p < 0.05). The highest histologic scores and entire woven bone formation (Huo Stage 8–9) were obtained at 6th weeks postoperatively in the HAM (+) group. Histological examination also revealed predominant fibrous tissue and partial cartilage formation (Huo Stage 2) at the postoperative 3rd week in the HAM (-) group. Equal amounts of woven bone and cartilage formation (Huo Stage 6–7) were observed at 3rd weeks postoperatively in the HAM (+) group and at 6th weeks postoperatively in the HAM (-) group. The callus diameters were greater in the HAM (+) group and the difference was statistically significant (p < 0.05) at 3rd and 6th weeks postoperatively. Although there was only a statistically significant difference (p < 0.05) at the postoperative 3rd week, radiological scores tended to be higher in the HAM (+) group at both the 3rd and 6th weeks postoperatively.ConclusionHAM is a cheap and easily accessible alternative biological material. HAM may be used to support surgical treatment of fractures, particularly where bone healing is expected to last longer.     

一氧化氮合酶在大鼠股骨闭合性骨折愈合中的表达及其意义

目的　探讨一氧化氮合酶 (NOS)在大鼠骨折愈合过程中的表达及其意义。方法　Wistar大鼠 40只建立股骨闭合性骨折模型。骨折后第 3天、1、2、4周分别处死每组 10只大鼠 ,取骨痂和对侧正常骨组织 ,苏木素 伊红 (HE)染色和免疫组织化学链霉抗生物素蛋白 过氧化物酶(SP)法观察骨折愈合及诱生性一氧化氮合酶 (iNOS)、内皮性一氧化氮合酶 (eNOS)和神经性一氧化氮合酶 (bNOS)在骨痂中的表达情况。结果　iNOS主要在成纤维细胞和成软骨细胞中表达 ;eNOS主要在增生血管内膜表达 ,bNOS没有表达。正常的股骨组织内没有NOS的表达。表达iN OS和eNOS的阳性细胞不同 ,并且不同时期其阳性细胞数也不同 ,iNOS在骨折后 1周最多 (P <0 .0 5 ) ,eNOS在骨折后 2周最多 (P <0 .0 1)。结论　iNOS和eNOS在骨折愈合中的表达具有时效性 ,它们在不同细胞内的表达提示一氧化氮可能对骨折愈合起一定的调节作用     

Effect of fluvastatin on vascular endothelial growth factor in rats with osteoporosis in process of fracture healing

OBJECTIVE: To explore the effect of fluvastatin on vascular endothelial growth factor (VEGF) in rats with osteoporosis in the process of fracture healing. METHODS: Fractures at the intermediate piece of the femur were made on 72 Sprague Dawley (SD) rats (weighing initially 290-340 g and aged 6 months) with osteoporosis after ovariectomy for three months, then these rats were divided randomly into the medication administration group (the experimental group) and the control group, 36 rats each. In the experimental group, the rats received fluvastatin lavage (10 mg/kg per day) since the next day of operation lasting for 6 weeks, and the rats in the control group received placebo. Then the expression of VEGF and VEGF mRNA in bony callus of the two groups was measured respectively with immunohistochemistry and in situ hybridization on days of 3rd, 7th, 14th, 21st, 28th, and 42nd, and image analysis was made with real-color image analysis machine. RESULTS: No difference was found in the cellular localization of VEGF and VEGF mRNA gene expression between the experimental group and the control group in process of fracture healing and their expression modes were almost similar. On the 14th day postoperatively, the positive extent of positive cells in the experimental group was higher than that of the control group (P < 0.05). CONCLUSION: Fluvastatin can promote the VEGF level in rats with osteoporosis in process of fracture healing.     

Role of platelet-rich plasma in acceleration of bone fracture healing

Platelet-rich plasma (PRP) is a common therapy for acceleration of maxillofacial and spinal fusion bone-graft healing. This study analyzes the therapeutic role of PRP during long-bone fracture healing evaluated Lewis rats. Following creation of unilateral open femur fractures, either 500 microL thrombin-activated PRP (PRP treated group) or 500 microL saline (control group) were applied once to the fracture site. Fracture healing was analyzed after 1 and 4 weeks. Following 4 weeks of fracture healing, radiographic analysis demonstrated higher callus to cortex width ratio (P < 0.05) in the PRP group (PRP: 1.65 +/- 0.06; control: 1.48 +/- 0.05). Three-point load bearing showed increased bone strength following PRP treatment (PRP: 60.85 +/- 6.06 Newton, control: 47.66 +/- 5.49 Newton). Fracture histology showed enhanced bone formation in the PRP group. Immunohistochemistry and Western-blotting demonstrated healing-associated changes in transforming growth factor (TGF)-beta1 and bone morphogenetic protein (BMP)-2. Our results suggest that PRP accelerates bone fracture healing of rat femurs via modulation of TGF-beta1 and BMP-2 growth factor expression.     

转化生长因子-β1在糖尿病骨折延迟愈合大鼠中的表达

目的 观察转化生长因子-β1(TGF-β1)的表达,探讨其对糖尿病骨折延迟愈合的影响.方法 70只大鼠随机分为对照组和链脲佐菌素按60 mg/kg诱导的糖尿病组,血糖>11.2mmol/L为诱导成功.均造成左侧胫骨骨折,于1、2、3、4、6、8周摄X片,取骨痂苏木素-伊红(HE)染色,免疫组织化学和酶联免疫吸附试验(ELISA)检测骨痂及血清TGF-β1.结果 X片和HE染色均示实验组较对照组骨形成滞后;实验组4周TGF-β1表达达高峰,迟于对照组3周,3周灰度值:实验组189.59±2.76,对照组166.74±1.33(P<0.05),3周血清含量:实验组(12.05±1.56) μg/L,对照组(17.48±0.56)μg/L(P<0.05).结论 糖尿病大鼠骨折后血清及骨痂TGF-β1减少是致其延迟愈合的原因之一.     

成骨细胞移植促进老龄鼠骨质疏松性骨折愈合过程中VEGF的动态变化

目的：动态观察在幼龄成骨细胞移植促进骨质疏松性骨折愈合过程中，VEGF在不同时相的表达及其生物学意义。方法：通过建立老龄骨质疏松SD大鼠骨折的动物模型，并将体外培养的SD雄性乳鼠成骨细胞移动到SD雌性鼠老年骨质疏松性骨缺损部位，利用免疫组化及原位杂交检测骨折愈合过程中不同时间相的移植标本VEGF、VEGFmRNA的表达，并作图像分析、绘出其动态变化图。结果：实验组VEGF、VEGFmRNA均在7d左右可见有阳性表达的细胞，14d有分泌,高峰其中以软骨细胞中阳性最强，21 d分泌量开始下降，56d后基本消失。而对照且未见明显分泌高峰。结论；成骨细胞细胞促进老龄鼠骨质疏松性骨折愈合，其机制可能是通过促进VEGF的转录和表达，从而促进骨折部位建立良好的血液循环，加速骨形成。     

骨形成蛋白和转化生长因子-β对兔尺骨骨折愈合后生物力学性能的影响

目的 探讨外源性骨形成蛋白(BMP)和转化生长因子—β(TGF—β)对骨折愈合后生物力学性能的影响。方法 36只日本大耳白兔，体重3．5～4．5kg。随机分成4组：空白对照组、TGF—β组、BMP组和TGF—β BMP组，每组9只。采用兔尺骨骨折模型，在骨折局部将聚乳酸(PAL)为载体制备成TGF—β／PAL、BMP／PAL、TGF—β BMP／PLA缓释系统，手术当天将各缓释系统植入骨折区，对照组在相应部位置入PAL，术后50天取材，通过三点弯曲法测骨折愈合后整体骨弯曲结构力学性能的变化。取断端密质骨，用三点弯曲、压缩和拉伸方法测骨试件的材料力学特性的变化，同时测骨痴的几何参数和骨密度的变化来对骨折愈合进行评估。结果 治疗组尺骨的几何参数，整体骨弯曲的破坏载荷、极限强度和弹性棋量，骨试件的弯曲、压缩和拉伸的极限强度，以及弯曲和压缩的弹性棋量明显高于对照组，各治疗组与对照组比较，具有统计学意义(P＜0．01)，TGF—β BMP组高于TGF—β和BMP组；TGF—β组高于BMP组，各治疗组之间两两比较，有统计学意义(P＜0．05)。在骨密度方面，各治疗组与对照组之间无明显差异。结论 兔骨折周围局部应用外源性TGF—β和BMP可促进骨痴形成，增强骨折愈合后骨组织的生物力学强度，TGF—β的作用优于BMP，联合应用优于单用一种生长因子，在促进骨折愈合方面具有协同作用。     

血管内皮生长因子在骨折修复过程中血管生成的促进作用

目的观察血管内皮生长因子(VEGF)在骨折修复过程中对骨折端微血管密度(MVD)的影响，探讨VEGF在骨折端血管生成中的作用。方法用168只大白鼠制作股骨骨折模型，随机分为VEGF组、拮抗VEGF组、对照组，用免疫组织化学分别测定伤后不同时间骨折端MVD的变化。结果应用外源性VEGF后。骨折端MVD明显增加；拮抗VEGF组，骨折端MVD明显下降；对照组MVD低于VEGF组，但高于拮抗VEGF组。结论VEGF在骨折修复过程中，对血管生成具有重要作用，并可能作为一种重要细胞因子参与和调节了骨折修复过程。     

扩髓治疗骨折不愈合中的血管分布研究

目的 研究扩髓治疗骨折不愈合过程中血管生成和血管内皮细胞生长因子(VEGF)表达的时间和空间特点以及相互联系.探讨扩髓并更换粗直径髓内钉治疗骨折不愈合的生物学机理,为临床运用扩髓治疗长骨骨折延迟愈合及不愈合提供理论依据. 方法 3个月龄雄性SD大鼠200只.对大鼠一侧股骨干骨折用不稳定固定方法制备肥大性骨折不愈合模型,随机选取40只作为模型鉴定,剩下160只随机分为试验组扩髓治疗和对照组不扩髓治疗,每组80只进行随机配对研究.术后第1、3、5、7、14、21、28、42天取材.用血管性血友病因子(VWF)标记血管,免疫组化方法检测骨折处VWF和VEGF的表达,来研究骨折处血管生成的时空特点以及与VEGF表达的关系. 结果 实验组的骨折愈合率显著高于对照组.时空分布上,在早期(1～7 d),实验组血管生成主要在骨外膜和软骨痂中.对照组主要在骨内膜;中期(7～14 d)实验组血管随着肉芽组织长入骨折间隙,对照组近骨内膜处血管增加,但是骨折间隙没有血管;后期(14～42 d)实验组骨内膜处出现血管,骨折间隙血管大量增加,对照组血管仍然局限在肉芽组织和骨内膜处,骨折间隙没有血管.实验组血管总数始终高于对照组.VEGF出现的时间和空间顺序与血管生成顺序一致,实验组VEGF表达普遍高于对照组. 结论 在骨折不愈合的治疗中,扩髓可使骨外膜和骨痂血供代偿性增加以及刺激VEGF表达增加,血管总数要高于不扩髓组.VEGF表达时间和分布与血管的生成关系密切,提示治疗骨折不愈合中,VEGF可能起着至关重要的作用,决定着血管形成的时间、部位与数量.     

植入型复方丹参缓释剂对骨折愈合影响的实验研究

目的:观察丹参缓释剂植入骨折局部对骨折愈合的影响.方法:42只新西兰健康白兔随机分为两组,均于右前肢桡骨中段制成3mm骨缺损的骨折模型,同时实验组放置丹参缓释剂60mg,对照组放置等量赋形剂.分别于术后第2、4、5、6周取右桡骨行X线片及组织学检查;第2、4、6周行电镜及骨密度检查;第5、6周进行生物力学测试.结果:X线片显示实验组较对照组骨折愈合提前.组织学观察实验组纤维性骨痂、软骨性骨痂、骨小梁之间过渡加快,毛细血管增生明显.电镜检查实验组示骨折修复细胞形态学变化均有利于修复骨折.术后第2、4、6周时骨密度值实验组均较对照组高,其中第4周时两者差异具有显著性(P＜0.05).术后第5、6周实验组抗折力均较对照组高(P＜0.05).结论:在骨折局部使用丹参缓释剂对骨折愈合具有一定的促进作用.     

甲状旁腺激素联合降钙素对大鼠骨质疏松性骨折骨生长因子的影响

目的探讨甲状旁腺素联合降钙素对大鼠骨质疏松性骨折骨相关生长因子及骨折愈合的影响。方法75只雌性SD大鼠随机分为:假手术组、去势组、甲状旁腺素组、降钙素组、联合用药组,15只/组,首先构建去卵巢大鼠骨质疏松性骨折动物模型。观察并评估骨折愈合,测定骨痂BMD和BMC,检测血清BMP-2、VEGF、TGF-β、IGF-1水平和骨痂蛋白表达,观察骨痂形态结构变化。结果去势组较假手术组骨折愈合评分、骨痂BMD和BMC、血清BMP-2、VEGF、TGF-β、IGF-1水平和骨痂蛋白表达均显著降低(P均<0.05),而联合用药组较去势组上述指标均显著升高(P均<0.05)。去势组骨痂骨小梁明显减少,生长稀疏,排列紊乱,大量间充质干细胞及纤维软骨细胞,成骨细胞及微血管少见;联合用药组骨痂大量骨小梁,生长旺盛,互相交错网状,排列致密有序,可见较多成熟的骨细胞及成骨细胞。结论甲状旁腺素联合降钙素通过介导提高相关骨生长因子表达,提高骨质疏松性骨折骨密度和矿物含量,改善骨组织形态学,加快骨折愈合。     

高能震波对骨痂中骨形成蛋白表达的影响

目的 观察高能震波（ＨＥＳＷ）对骨痂中骨形成蛋白（ＢＭＰ）表达的影响，探讨其促进骨折愈合的机理。方法 手术造成家兔双侧胫骨中段标准骨折模型，对一侧胫骨骨折部施以ＨＥＳＷ，另一侧为对照组，术后每周进行Ｘ线及组织学检查，并取骨痂组织切片用ＢＭＰ抗体进行免疫组化染色（ＳＡＢＣ法（）。结果 实验组骨痂中ＢＭＰ表达强度在第２－４周时明在于对照组（Ｐ〈０．０１），且Ｘ线摄片及组织学观察均显示实验组骨折愈合时间     

Experimental histopathological studies of electrical callus formation and mechanism of bone healing by direct micro-electrical current

In order to get better understanding of the effects of electrical stimulation on bone healing processes, the author compared the healing processes of the femur in dogs between two groups: a stimulation group and a control group (non-stimulation group) which were experimentally prepared. These bone specimens were periodically extirpated and used for pathological examinations and X-ray micro-analysis. In the stimulation group, strong proliferation of osteoblasts and new trabecular formation in the bone marrow were observed at the 3rd day, and transition from fibrous to bony callus were noted at the 9th day; after the 3rd week bone remodeling was sparsely seen and bone healing period was shortened. In electromicroscopic observation, calcification of bone matrix and bone remodeling also seemed to be facilitated in this group. However, no marked differences in histological process of bone healing were observed between the stimulation group and the control group.     

Co-expression of BMPs and BMP-inhibitors in human fractures and non-unions

Bone morphogenetic proteins (BMPs) are increasingly being used clinically to enhance fracture repair and healing of non-unions. However, the potential efficacy of supraphysiological dosing for clinical results warrants further clarification of the BMP signaling pathway in human fracture healing. As BMP signaling can be fine-tuned at numerous levels, the role of BMP-inhibitors has become a major focus. The aim of the present study was to document co-expression of BMPs, pSmad 1/5/8, and BMP-inhibitors in human fracture callus and human non-unions. Using human tissue of fracture callus (n=14) and non-unions (n=4) we documented expression of BMPs (BMP2, BMP3 and BMP7), pSmad 1/5/8 and the BMP-inhibitors noggin, gremlin, chordin, Smad-6, Smad-7 and BAMBI. Co-expression of pSmad 1/5/8, BMPs and BMP-inhibitors was noted in the osteoblasts of fracture callus as well as of non-unions. Expression of BMP-inhibitors was generally stronger in non-unions than in fracture callus. The most pertinent differences were noted in the cartilaginous tissue components. Expression of BMP2 in chondrocytes was markedly decreased in non-unions compared to fracture callus and that of BMP7 was almost completely absent. Expression of BMP-inhibitors was almost the same in osteoblasts, chondrocytes and fibroblasts of fracture callus and well as in non-unions. Interestingly, although BMP ligands were present in the chondrocytes and fibroblasts of non-unions, they did not co-express pSmad 1/5/8 suggesting that BMP signaling may have been inhibited at some point before Smad 1/5/8 phosphorylation. These results suggest co-expression of BMP, pSmad 1/5/8 and BMP-inhibitors occurs in human fracture callus as well as non-unions but the relative expression of BMPs vs. BMP-inhibitors was different between these two tissue types. In contrast to our expectations, the expression of BMP inhibitors was comparable between fracture callus and non-unions, whereas the expression of BMPs was notably lower in the cartilaginous component of the non-unions in comparison to fracture callus. Based on these results, we believe that aberrations in the BMP-signaling pathway in the cartilaginous component of fracture healing could influence clinical fracture healing. An imbalance between the local presence of BMP and BMP-inhibitors may switch the direction towards healing or non-healing of a fracture.     

ERK5在小鼠骨质疏松性骨折愈合过程中作用的实验研究

目的通过构建小鼠骨质疏松骨折模型,研究ERK5在骨质疏松性骨折愈合过程中的作用。方法将108只6周龄雌性昆明小鼠随机分成4组,通过手术切除小鼠双侧卵巢及小鼠股骨离断分别构建骨质疏松(OVX)和骨折模型(Fracture),然后给实验组小鼠每日腹腔注射ERK5特异性阻断剂XMD8-92,于1 w、2 w、4 w后分别处死一定数量的小鼠,取术侧股骨标本,行X线片检查、股骨骨痂Micro-CT、HE染色及免疫组织化学染色检查,观察骨折端骨小梁生长情况,骨痂内成骨相关蛋白及ERK5表达情况。结果给予实验小鼠注射XMD8-92后第2周及第4周,Fracture组小鼠骨痂生长较快,骨小梁数目较多,厚度较大,成骨相关蛋白ALP、Runx2的表达相对较多(P0.05),而Fracture+XMD8-92组小鼠骨痂生长则相对缓慢,骨小梁稀少且绯薄,结构相对较乱,碱性磷酸酶(alkaline phosphatase,ALP)、Runx2表达较少(P0.05);且OVX+Fracture+XMD8-92组小鼠与OVX+Fracture组小鼠相比,骨小梁生成更少且紊乱,骨痂生长明显延迟,ALP、Runx2表达量显著减少(P0.05)。结论 ERK5影响骨折端骨痂形成的速度和质量,在促进骨质疏松性骨折愈合过程中起着十分重要的生理作用。     

微动对骨折端微循环及血管内皮生长因子(VEGF)表达的影响

目的研究外固定架微动对骨折端微循环及血管内皮生长因子(VEGF)表达的影响。方法60只新西兰大白兔随机分为微动组与固定组,利用微循环检测仪、免疫组化等方法检测固定组和微动组动物骨折端血流量、VEGF蛋白表达。结果微动组骨折端血流量在骨折后2、3、4、5周较固定组有显著增加。骨折后3、4、5周微动组骨折端软骨细胞、成骨细胞中VEGF蛋白表达均较固定组增强。结论微动可以增加VEGF表达,使毛细血管再生增加,促进骨痂生长。     

Effects of recombinant human growth hormone (r-hGH) on experimental osteoporotic fracture healing

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