功能性电刺激在实验性骨折愈合过程中的作用

应用＾３Ｈ－脯氨酸和＾１５Ｃａ－氯化钙无载体放射性核素体内示踪法，检测了周围神经功能性电刺激对实验性骨折胶愈合过程中胶原蛋白形成与基质钙化的影响。发现：生理性电刺激周围神经，使＾３Ｈ和＾１５Ｃａ掺入早期骨痂的量增加（与对照组比Ｐ〈０．０５），＾１５Ｃａ／＾３Ｈ的比率提高迅速。表明早期骨痂内胶原合成量增加，基质钙化迅速、完全，骨性骨痂发生早，骨转化过程缩短。认为周围神经功能性电刺激可作为促进骨修复的     

低强度超声波促进骨折愈合过程中I、X型胶原蛋白的基因表达

目的 研究低强度超声波刺激下骨折处I、X型胶原的基因表达规律,从分子水平探索低强度超声波促进骨折愈合的作用机制.方法 应用兔双侧桡骨骨折模型,采用左右自身对照.36只新西兰雄性大白兔,分为6组.右侧骨折处每日行超声刺激20rain,左侧不行刺激.分别于术后3、7、10、14、21、28d取骨折处骨痂及纤维组织,进行骨痂厚度的测量和组织学观察.RT-PCR方法半定量分析I、X型胶原mRNA的表达规律.结果 自术后10d开始至28d,实验侧的骨痂厚度明显高于对照侧.术后10、14、21、28d HE染色显示成软骨细胞及成骨细胞提早出现而且量多,细胞外基质合成增加.I型胶原mRNA表达于术后14、21d含量明显高于对照侧,软骨内细胞外基质蛋白合成增加,增加新骨基质的含量.X型胶原mRNA表达于术后14d明显高于对照侧,术后21d达高峰,然后下降.结论 低强度超声波作用可以增加骨折处骨痂含量,其加快骨折愈合通过刺激骨痂增生,使矿化过程提前发生所致.低强度超声波刺激可以增加I、X型胶原的mRNA含量,促使软骨内成骨过程中软骨细胞的增多和细胞外基质的钙化.     

左旋多巴对^35S—硫酸钠掺入骨折骨痂不同时期的影响

作者用~(35)S-Na_2SO_4无载体溶液为示踪核素,观察了口服左旋多巴对~(35)S掺入大鼠胫骨骨折后骨痂形成不同时期的影响.结果证实:第5天、第10天,实验组~(35)S掺入骨痂量(0.11±0.02DPM%和0.19±0.05DPM%)明显大于对照组(0.08±0.02DPM%和0.13±3.05DPM%),统计学上有显著性差异(P<0.01).说明左旋多巴能促进早期骨痂对硫的利用,使外骨痂中软骨合成量增加,加快了软骨内成骨的过程.作者认为:左旋多巴促进骨折愈合作用主要是在纤维和纤维软骨骨痂形成期.     

红景天甙联合碳酸钙对老年雌性大鼠骨量、骨代谢和氧化应激水平影响

目的探索红景天甙联合碳酸钙对老年雌性大鼠骨量、骨代谢和氧化应激水平影响。方法将40只24月龄雌性Sprague-Dawley(SD)大鼠随机分为模型组(OP组),红景天甙组(H组),碳酸钙组(Ca组)和红景天甙联合碳酸钙组(H+Ca组),而将10只3个月大鼠作为对照组(NC组)。观察各组大鼠腰椎和双侧股骨骨密度和骨矿物质水平的变化。测定各组大鼠骨代谢指标、超氧化物歧化酶(SOD)、丙二醛(MDA)和谷胱甘肽过氧化物酶(GSH-Px)水平变化。结果与OP组比较,H、Ca和H+Ca组大鼠腰椎骨密度,双侧股骨骨密度和骨矿物质水平显著升高;治疗12周,H、Ca和H+Ca组血清I型胶原C端肽(CTX-1)、骨钙素、碱性磷酸酶(ALP)、I型前胶原N端肽(PINP)和抗酒石酸酸性磷酸酶5b(TRACP-5b)水平均显著降低,比较差异有统计学意义(P 0.05);治疗12周,H、Ca和H+Ca组血清SOD和GSH-Px活性明显升高,MDA含量显著降低,差异有统计学意义(P 0.05或P0.01);其中H+Ca组上述指标变化最为显著;差异有统计学意义(P 0.05)。结论红景天甙联合碳酸钙可显著提高老年大鼠的抗氧化能力,增加骨密度,降低骨代谢水平,减少骨矿物质流失,从而预防和治疗骨质疏松症。     

26,27-F6-1,25(OH)2D3, 1,25(OH)2D3和17-βEstradiol对培养胎鼠头盖骨的影响

26，26．26，27，27，27-Hexafuoro-la，25-dihydroxy vitamin D3[26，27-F8-1,25(()H)2D2]是一种新合成的具有刺激骨代谢和细胞分裂生物活性的药物。实验研究了26，27-F6-1，1，25(()H)2D2、1a，25-dihvdroxvvitamin D3[1,25(()H)2D4和17β-Estradio! Estradiol]在骨形成和骨吸收等方面的作用，以探讨其对治疗骨质疏松症的基本作用。实验使用妊娠20日龄大鼠胎鼠的成对头盖骨，于强化BGJb培养基中培养7天，测定骨干重、Ca含量及羟脯氨酸含量。26，27-Fa-25(OH)2DD3对胎鼠头盖骨干重、Ca含量和羟晡氨酸含量都有较明显的增加作用。其作用优于Estradiol单纯提高胎鼠头盖骨中Ca的古量．而对骨的干重和羟脯氨酸含量无显著影响；也强于1，25(OH)2D3仅增加胎鼠头盖骨干重和羟脯氨酸含量，却不能提高骨Ca古量的作用，由于26．27-F3-1，25(OH)2D2在骨形成中对胎鼠头盖骨包括骨盐和骨基质中的重要物质Ca和羟晡氨酸都有促进作用。目此它很可能成为一种有潜力的冶疔骨质疏松症的新型药物。     

补钙与骨质疏松症防治研究的进展

骨质疏松症的发生是一个渐进的过程,其临床表现为疼痛、驼背和骨折等。不管是原发性还是继发性骨质疏松症,均有一个共同的防治原则:延缓骨量丢失或增加骨量,对症处理和预防骨折发生。这就需要补充钙制剂。补钙与骨质疏松症防治研究主要进展包括:(1)青少年期摄入钙对其骨矿化的作用。(2)妊娠、哺乳期钙需要量与骨矿含量的关系。(3)老年人的维生素D和钙摄入。(4)缺钙与异位钙化(Cetopiccal-cification)。(5)口服钙补充剂。(6)补钙新问题。现综述如下。一、青少年时期摄入钙对其骨矿化的作用体内99.9%的钙在骨骼中,青少年骨骼生长迅速,是骨量…     

应用双极矩形脉冲电流装置刺激骨痂形成

近年来,不少学者对于电刺激促进骨痂形成这一课题颇感兴趣,并在动物实验和临床上观察其加快骨痂形成的效果。在应用过程中,直流电刺激常不够满意,而应用单极矩形脉冲、中频电流或电动势效应等方法都可以促进骨愈合。除了应用不同的电极外,电流的频率、强度以及作用时间为决定性因素。骨愈合的加速为阴极刺激的结果。但在电刺激的阳极附近可以发生不利的溶骨现象。为了排除这种不利的影响,作者采用能     

矩形髓内钉对实验性骨折愈合过程中骨痂含量的影响

目的　探讨矩形髓内钉 (Rectangle shapedIntramedullaryNail,简称RIN)对骨折愈合过程中骨痂含量的影响。方法　用兔胫骨骨折内固定模型将RIN和传统的Ender s钉 (Ender sNails,简称END)、四孔接骨板 (StainlessSteelPlates ,简称SPL)进行对比研究 ,通过阅片及计算机图像分析系统对不同内固定后不同时期的骨痂X线片进行分析。结果　骨痂直径测量显示 :同组相比 ,8周时骨痂直径最大 ;同期比较 ,RIN组骨痂直径最大 ,END组次之 ,SPL组最少。RIN、END组和SPL组组间均有显著差异 ,RIN和END组相比则无显著差异 ;骨痂灰度密度分析显示 :同组的骨痂灰度密度值随术后时间的增加而增大 ;同期比较 ,8周组的骨痂灰度密度值最高 ,END组次之 ,SPL组最少。RIN、END组和SPL组组间均有显著差异 ,RIN和END组相比 ,RIN组 8周后组间差异显著。结论　RIN组和END组在骨折的不同时期的骨痂量较SPL组多 ,而且骨痂的钙化也较早 ,促进了骨折愈合     

骨折愈合机理的研究进展

骨折的愈合是一个复杂的组织学 ,生物学 ,内分泌学及生物力学的动态过程。1　骨折的自然愈合过程按组织学变化分为三个阶段 :(1)血肿机化期 :约 3周 (包括损伤性炎症及血肿形成 ,机化 )骨断裂后 ,髓腔内 ,骨膜下和周围软组织内出血 ,形成血肿。前 2周 ,新生的毛细血管和吞噬细胞 ,成纤维细胞侵入 ,清除机化血肿 ,形成肉芽组织 ,纤维组织。第 3周 ,骨外膜 ,骨内膜的成骨细胞活跃增生 ,由远端骨折处逐渐向骨折处延伸 ,形成骨样组织。 (2 )原始骨痂形成期 :骨内、外膜的成骨细胞形成骨样组织逐渐钙化 ,膜内化骨形成外骨痂和内骨痂。断端间和髓…     

低频微动后转化生长因子β_1与胰岛素样生长因子在新骨组织中的表达研究

目的研究转化生长因子β1(transforminggrowthfactorβ1,TGF-β1)与胰岛素样生长因子(insulin-likegrowthfactor,IGF-)在低频微动引起骨折间接愈合过程中的表达与意义。方法山东雌性高腿绵羊15只,行双侧胫骨中段横形截骨,形成2mm骨缺损,用带有微动装置的单边外固定支架固定。术后10d,随机选择一侧肢体为微动组,以频率1Hz,幅度0.25mm(30min/d)微动,4周结束;另一侧后肢不微动,为对照组。术后3、4和6周分别处死5只动物,应用免疫组织化学与RT-PCR检测骨痂组织中TGF-β1与IGF-的表达。结果免疫组织化学:术后3周,微动组TGF-β1在骨痂边缘的新生软骨细胞各区均有阳性表达,以增殖区最为显著;IGF-主要表达于软骨内骨化带边缘的成骨细胞,钙化并转化为新骨的软骨细胞与骨细胞中。对照组的相应区域,TGF-β1有少量表达,IGF-几乎无表达。4周,微动组新骨组织逐渐成熟,TGF-β1表达强度减弱,阳性信号主要位于细胞外基质与骨化带周围成骨细胞中;IGF-的表达趋于高峰,主要集中在新骨表面的成骨细胞、趋于成熟的骨细胞及趋于钙化的类骨质。对照组TGF-β1与IGF-仅有少量表达。6周,微动组TGF-β1的表达显著衰退,IGF-仍有适量表达,主要在新生骨小梁骨细胞中。对照组TGF-β1与IGF-仅有微量表达。微动组术后3、4周TGF-β1,3、4、6周IGF-吸光度(A)值与对照组相比,差异有统计学意义(P<0.05)。RT-PCR电泳示:术后3、4周,微动组骨痂中TGF-β1与IGF-的mRNA有较高的表达量;对照组骨痂中TGF-β1、IGF-的mRNA有轻度表达。术后6周,微动组TGF-β1、IGF-的mRNA表达量显著降低,但仍略高于对照组。微动组术后3、4周TGF-β1,3、4、6周IGF-A值与对照组相比,差异有统计学意义(P<0.05)。结论低频微动在骨折愈合早期,可以促进TGF-β1与IGF-的表达。它们协同调节软骨内骨化的进程;后期主要由IGF-调节骨细胞的分化与类骨质的矿化,其可能更多的参与调节微动各个时期的细胞生物行为。     

Electrostimulation of rat callus cells and human lymphocytes in vitro

Asymmetrical pulsing low voltage current was supplied via electrodes to cultured rat fracture callus cells and human peripheral blood lymphocytes. The [3H]thymidine incorporation of the callus cells and 5-[125I]iodo-2'-deoxyuridine incorporation of the lymphocytes were determined. The growth pattern of callus cells (estimated by cellular density) did not respond to electrical stimulation. However, the uptake of [3H]thymidine was increased at the early phase of cell proliferation and inhibited at later phases of proliferation. The [3H]thymidine uptake of confluent callus cell cultures did not respond to electrical stimulation. Lymphocytes reacted in a similar way; stimulated cells took up more DNA precursor than control cells at the early phase of stimulation. During cell division, induced by the mitogens phytohemagglutinin and Concanavalin-A, the uptake of DNA precursor by stimulated cells was constantly inhibited. The results suggest that electrical stimuli affect the uptake mechanisms of cell membranes. The duality of the effect seems to be dependent on the cell cycle.     

不同神经损伤对骨折前期愈合影响的实验研究

目的：研究脊髓和周围神经损伤对骨折前期修复的影响。方法：将实验大鼠分成3组，一组为T10脊髓横断伴股骨骨折，另两组分别为周围神经切断伴股骨骨折组和单纯股骨骨折组。骨折后2周和4周摄股骨X线片，4周处死大鼠行骨痂细胞和组织学检查。结果：骨折后2周3组均未出现明显骨痂，术后4周测量x线片骨痂大小显示周围神经切断组骨痂最多，其次分别为对照组和脊髓损伤组。组织学显示周围神经切断组骨痂量多，并有明显的皮质骨桥接断端，但其中类骨质较多，骨钙的沉积相对较少，而且骨痂缺乏血管和正常的骨小梁结构，骨化重建不够理想；而脊髓损伤组骨痂则表现为欠成熟，细胞种类多，分布紊乱，少部分仍处于纤维修复阶段。对照组的组织学改变介于两者之间，但其骨小梁结构良好而且骨钙沉着明显。结论：不同类型神经损伤对骨折愈合前期有着不同的影响。失周围神经支配对骨折修复有一定的加速作用，而脊髓损伤则延缓了骨折前期愈合，其发生机理仍需进一步研究。     

Early mechanical stimulation only permits timely bone healing in sheep

Bone fracture healing is sensitive to the fixation stability. However, it is unclear which phases of healing are mechano‐sensitive and if mechanical stimulation is required throughout repair. In this study, a novel bone defect model, which isolates an experimental fracture from functional loading, was applied in sheep to investigate if stimulation limited to the early proliferative phase is sufficient for bone healing. An active fixator controlled motion in the fracture. Animals of the control group were unstimulated. In the physiological‐like group, 1 mm axial compressive movements were applied between day 5 and 21, thereafter the movements were decreased in weekly increments and stopped after 6 weeks. In the early stimulatory group, the movements were stopped after 3 weeks. The experimental fractures were evaluated with mechanical and micro‐computed tomography methods after 9 weeks healing. The callus strength of the stimulated fractures (physiological‐like and early stimulatory) was greater than the unstimulated control group. The control group was characterized by minimal external callus formation and a lack of bone bridging at 9 weeks. In contrast, the stimulated groups exhibited advanced healing with solid bone formation across the defect. This was confirmed quantitatively by a lower bone volume in the control group compared to the stimulated groups.The novel experimental model permits the application of a well‐defined load history to an experimental bone fracture. The poor healing observed in the control group is consistent with under‐stimulation. This study has shown early mechanical stimulation only is sufficient for a timely healing outcome. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 36:1790–1796, 2018.

     

失神经对大鼠胫骨骨折愈合影响作用的实验研究

目的：通过失神经后大鼠腔骨骨折愈合情况的观察，研究周围神经系统影响骨折愈合的作用。方法：造成大鼠双侧腔骨闭合横行骨折，一侧切断股神经坐骨神经，为骨折失神经组，另一侧保留神经为骨折正常神经支配组。术后4周处死大鼠，取对应双侧腔骨标本称湿重、拍X线片、测定生物力学强度及骨计量学检查。结果：湿重及X线片骨痂评分显示，失神经组骨痂形成量明显大于正常神经支配组，但力学强度降低。骨计量学显示失神经组骨痂类骨质宽度、破骨细胞指数、破骨细胞吸收表面增大，双标记线间距离较窄、矿化沉积速率较慢、类骨质成熟时间相对延长。结论：失神经可使骨折形成有明显缺陷的骨痂，完整的神经支配是骨折正常愈合的必要条件。     

Implications for Fracture Healing of Current and New Osteoporosis Treatments: An ESCEO Consensus Paper

Osteoporotic fracture healing is critical to clinical outcome in terms of functional recovery, morbidity, and quality of life. Osteoporosis treatments may affect bone repair, so insights into their impact on fracture healing are important. We reviewed the current evidence for an impact of osteoporosis treatments on bone repair. Treatment with bisphosphonate in experimental models is associated with increased callus size and mineralization, reduced callus remodeling, and improved mechanical strength. Local and systemic bisphosphonate treatment may improve implant fixation. No negative impact on fracture healing has been observed, even after major surgery or when administered immediately after fracture. Experimental data for denosumab and raloxifene suggest no negative implications for bone repair. The extensive experimental results for teriparatide indicate increased callus formation, improved biomechanical strength, and greater external callus volume and total bone mineral content and density. Case reports and a randomized trial have produced mixed results but are consistent with a positive impact of teriparatide on clinical fracture healing. Studies with strontium ranelate in models of fracture healing indicate that it is associated with improved bone microstructure, callus volume, and biomechanical properties. Finally, there is experimental evidence for a beneficial effect of some of the agents currently being developed for osteoporosis, notably sclerostin antibody and DKK1 antibody. There is currently no evidence that osteoporosis treatments are detrimental for bone repair and some promising experimental evidence for positive effects on healing, notably for agents with a bone-forming mode of action, which may translate into therapeutic applications.     

The electrical induction of callus formation and external skeletal fixation using methyl methacrylate for delayed union of open tibial fracture with segmental loss.

A case of delayed union of an open tibial fracture with segmental loss is presented in which electricity was applied to the bone defect through two screws that were connected to a methy methacrylate skeletal fixation device. The fracture developed electrically induced callus formation around the cathode, callus extended to the anode, and solid bony union ensued. External skeletal fixation using methyl methacrylate is convenient and is a suitable adjunct to electrical stimulation of fracture healing.     

Muscle preservation using an implantable electrical system after nerve injury and repair.

The value of continuous electrical stimulation of denervated muscles after nerve injury and repair has been clearly shown in a series of laboratory experiments in three animal models. This experimental background, which showed improved muscle preservation and better functional results, evolved into a clinical study that included 15 patients with peripheral nerve injuries in the upper extremities, 3 patients with brachial plexus injuries, and three patients with facial nerve paralysis. Improved functional results were obtained using this implantable system, which were similar to those achieved with the animal experiments. All patients had muscle stimulation for extended periods ranging from 127 to 346 days. Analysis of the results showed satisfactory nerve regeneration on clinical examination and with electromyographic studies. Functional muscle analysis varied somewhat from patient to patient, but every patient had a satisfactory to excellent recovery. The results from this study have clearly shown the benefits of continuous muscle stimulation using an implantable electrical system after nerve injury and repair expansion of the project to a larger patient cohort is indicated.     

Effect of mechanical stability on fracture healing--an update

The effects of mechanical stability and mechanical stimulation have been studied extensively in vivo using a variety of animal models and stimulators. Early results indicated that stimulation does not significantly contribute to fracture healing. Lately, however, more rigid external stimulators that withstand increased callus formation have identified a contribution of mechanical stimulation in the initial period of fracture healing. However, these studies also show that the same amount of movement inhibits union during the last phase of fracture healing. On the cellular level, most investigations have used 2-dimensional cell culture systems to study the response of different cell phenotypes to mechanical stimulation, shear stress, and hydrostatic pressure. Cell proliferation and differentiation are clearly altered by these stimuli, however, the response depends on the cell type, the magnitude of the strain, and the cofactors applied. Lately, 3-dimensional cell cultures in mechano-bioreactors have been used to investigate the response of bone marrow stromal cells. These results indicate that the predominant stimulus for proliferation is perfusion. Mechanical stimulation affects cell differentiation and depends on the strain magnitude and the cell phenotype. As a consequence, today's implants should be applied in a fashion that supports maximum perfusion at the fracture site. In the early period, the osteosynthesis should facilitate micromotion of the fragments if secondary fracture healing is desired. At the same time, joint congruency, and axial and rotational positions have to be maintained. In the final period of healing, motion within the calcifying callus should be limited, which is naturally achieved by the increasing stiffness of the callus ossification.     

Monitoring of fracture healing by electrical conduction: A new diagnostic procedure

Background:

Electrical stimulation of fractures has been reported to enhance fracture healing. X-rays are normally used to assess union of fractures. Electrical conduction is not tried as a tool to study fracture healing. The current study focuses on electrical conduction as a diagnostic tool to assess fracture healing and new bone formation. The aim was to find if electrical resistance across the fracture can be used as a tool to study fracture healing which can be verified with simultaneous radiographs.

Materials and Methods:

A prospective study was conducted where 12 open fractures of tibia, including two with bone defects were evaluated. They were debrided and four-carbon ring Ilizarov external fixators were applied. Their healing was followed with clinical assessment and periodical X-rays till the endpoint of fracture union and then the rings were removed. In addition, all these cases also had application of electrical voltage in the range of 0.1–1.0 V DC in 0.1 V increments, across the two wires on either side of fracture. The output current was recorded by an ammeter connected in series. Resistance calculated for various voltages was plotted as a graph for the period of fracture treatment and the characteristics were studied. This graph was compared with the appearance of new bone in the X-rays.

Results:

In all cases, when the above graph stabilized, in the consecutive recordings, the X-rays showed healing (bridging callus) matching the curve and the patient was able to load the limb. The time of stabilization of this graph for a specific voltage was different in individual cases. However, for a given case, the resistance characteristics were the same for the entire voltage range of 0.1–1.0 V.

Conclusion:

If the resistance versus day curve stabilizes on the consecutive recordings, we can predict that the fracture is in the process of healing. This stabilization period also matched the patients’ ability to comfortably load the limb and also the radiographs which showed bridging callus (healing). If this is used as a positive criterion for fracture healing in future, the radiation exposure by X-rays shall be less.     

Mechanisms for the enhancement of fracture healing in rats treated with intermittent low-dose human parathyroid hormone (1-34).

Recent reports have demonstrated that intermittent treatment with parathyroid hormone (1-34) [PTH(1-34)] increases callus formation and mechanical strength in experimental fracture healing. However, little is known about the optimal dose required for enhancement of fracture repair or the molecular mechanisms by which PTH regulates the healing process. In this study, we analyzed the underlying molecular mechanisms by which PTH affects fracture healing and tested the hypothesis that intermittent low-dose treatment with human PTH(1-34) can increase callus formation and mechanical strength. Unilateral femoral fractures were produced and a daily subcutaneous injection of 10 microg/kg of PTH(1-34) was administered during the entire healing period. Control animals were injected with vehicle solution alone. The results showed that on day 28 and day 42 after fracture, bone mineral content (BMC), bone mineral density (BMD), and ultimate load to failure of the calluses were significantly increased in the PTH-treated group compared with controls (day 28, 61, 46, and 32%; day 42, 119, 74, and 55%, respectively). The number of proliferating cell nuclear antigen (PCNA)-positive subperiosteal osteoprogenitor cells was significantly increased in the calluses of the PTH-treated group on day 2, and TRAP+ multinucleated cells were significantly increased in areas of callus cancellous bone on day 7. The levels of expression of type I collagen (COLlA1), osteonectin (ON), ALP, and osteocalcin (OC) mRNA were increased markedly in the PTH-treated group and accompanied by enhanced expression of insulin-like growth factor (IGF)-I mRNA during the early stages of healing (days 4-7). The increased expression of COL1A1, ON, ALP, and OC mRNA continued during the later stages of healing (days 14-21) despite a lack of up-regulation of IGF-I mRNA. These results suggest that treatment of fractures with intermittent low dose PTH(1-34) enhances callus formation by the early stimulation of proliferation and differentiation of osteoprogenitor cells, increases production of bone matrix proteins, and enhances osteoclastogenesis during the phase of callus remodeling. The resultant effect to increase callus mechanical strength supports the concept that clinical investigations on the ability of injectable low-dose PTH(1-34) to enhance fracture healing are indicated.