Effects of in vivo mechanical loading on large bone defect regeneration

Fracture healing is highly sensitive to mechanical conditions; however, the effects of mechanical loading on large bone defect regeneration have not been evaluated. In this study, we investigated the effects of functional loading on repair of critically sized segmental bone defects. About 6-mm defects were created in rat femora, and each defect received 5 μg recombinant human bone morphogenetic protein-2 (rhBMP-2), delivered in alginate hydrogel. Limbs were stabilized by either stiff fixation plates for the duration of the study or compliant plates that allowed transfer of compressive ambulatory loads beginning at week 4. Healing was assessed by digital radiography, microcomputed tomography, mechanical testing, histology, and finite element modeling. Loading significantly increased regenerate bone volume and average polar moment of inertia. The response to loading was location-dependent with the polar moment of inertia increased at the proximal end of the defect but not the distal end. As a result, torsional stiffness was 58% higher in the compliant plate group, but failure torque was not altered. In single samples assessed for histology from each group, a qualitatively greater amount of cartilage and a lesser degree of remodeling to lamellar bone occurred in the loaded group compared to the stiff plate group. Finally, principal strain histograms, calculated by FE modeling, revealed that the compliant plate samples had adapted to more efficiently distribute loads in the defects. Together, these data demonstrate that functional transfer of axial loads alters BMP-induced large bone defect repair by increasing the amount and distribution of bone formed within the defect.     

Mixed-mode toughness of human cortical bone containing a longitudinal crack in far-field compression

Bone is generally loaded under multiaxial conditions in vivo; as it invariably contains microcracks, this leads to complex mixed-mode stress-states involving combinations of tension, compression and shear. In previous work on the mixed-mode loading of human cortical bone (using an asymmetric bend test geometry), we found that the bone toughness was lower when loaded in far-field shear than in tension (opposite to the trend in most brittle materials), although only for the transverse orientation. This is a consequence of the competition between preferred mechanical vs. microstructural crack-path directions, the former dictated by the direction of the maximum mechanical “driving force” (which changes with the mode-mixity), and the latter by the “weakest” microstructural path (which in human bone is along the osteonal interfaces or cement lines). As most microcracks are oriented longitudinally, we investigate here the corresponding mixed-mode toughness of human cortical bone in the longitudinal (proximal-distal) orientation using a “double cleavage drilled compression” test geometry, which provides a physiologically-relevant loading condition for bone in that it characterizes the toughness of a longitudinal crack loaded in far-field compression. In contrast to the transverse toughness, results show that the longitudinal toughness, measured using the strain-energy release rate, is significantly higher in shear (mode II) than in tension (mode I). This is consistent, however, with the individual criteria of preferred mechanical vs. microstructural crack paths being commensurate in this orientation.     

Properties of deproteinized bone for reparation of big segmental defect in long bone

Objective： To explore suitable scaffold material for big segmental long bone defect by studying the properties of the prepared deproteinized bone.
Methods： Cancellated bone were made as 30 mm × 3 mm × 3 mm bone blocks from inferior extremity of pig femur along bone trabecula. The deproteinized bone was prepared with an improved method. Their morphological features, components, cell compatibility, mechanical and immunological properties were investigated respectively. Results： Deproteinized bone maintained natural re-ticular pore system. The main organic material is collagen I and inorganic composition is hydroxyapatite. It has good mechanical properties, cell adhesion rate and histocompatibility.
Conlusion： This deproteinized bone can be applicable as scaffold for reparation of big segmental defect in long bone.     

A method to prepare bone-implant specimens for laboratory analysis

Summary A method for the preparation of bone samples containing metallic implants is presented. Technics are detailed for samples to be analyzed by mechanical, microradiographical and microangio-graphical means, and by tetracycline labeled fluorescent and polarized light microscopy.     

Mechanical strength of bone in canine osteoporosis model: Relationship between bone mineral content and bone fragility

The purpose of this study was to clarify the relationship between bone mineral content (BMC) and mechanical strength in beagle dog models, and to find whether the mechanical strength changed with changes in BMC. We used 17 beagle dogs to create an experimental osteoporotic model, dividing them into six groups, based on age and experimental period. Presence and absence of ovariectomy (OVX); and calcium content of the diet. BMC was determined by dual-energy quantitative computed tomography in the lumbar vertebrae, and the mechanical strength of cancellous bone harvested from the lumbar vertebral body and femoral neck was measured. OVX alone did not affect either BMC or mechanical strength. However, when the dogs were ovariectomized, and then given a reduced calcium diet, the mechanical strength of the femoral neck decreased in parallel with the cortical BMC of the lumbar vertebrae. The mechanical strength of the vertebral cancellous bone was not decreased when the BMC was reduced by 20%, but was decreased when BMC was reduced by 30%.     

异种脱蛋白松质骨为载体构建组织工程骨用于脊柱横突间融合的实验观察

目的探讨异种脱蛋白松质骨（deproteinization bone,DPB）作为骨组织工程载体的性能及其用于山羊脊柱横突间融合的作用。方法取成年猪股骨远端松质骨通过理化方法制成DPB载体,对其形态结构、组成成分、生物力学特性以及材料对种子细胞生物学行为的影响进行检测分析。将载体材料复合一定量的自体骨髓间充质干细胞（mesenchymal stem cells,MSCs）重组人骨形成蛋白2（recombinant human bone morphogenetic protein 2,rhBMP-2）构建组织工程骨。取6～8月龄雄性山羊12只,制成L3-4双侧横突间植骨模型,左侧植入组织工程骨为A组,右侧植入等体积自体髂骨作对照为B组。分别于4、8及12周行X线片和组织学观察并对比分析。结果采用脱蛋白处理后的松质骨可见大小不等、相互交通、开放孔隙的网架结构。孔径200～500μm,孔隙率60％左右。其无机成分为羟基磷灰石,有机成分为胶原,力学性能保存良好,有良好的细胞相容性。两组移植后不同时间点X线表现：A组植入横突间第4周与横突桥接处部分区域模糊,以内侧明显;第8周上下桥接部间隙变小,大量连续骨痂生成;12周后完全融合。早期A组密度略低于B组,12周后基本相同。移植区组织学观察：A组植入后第4周以多点方式形成新骨,第8周岛状生长的骨组织贯穿整个移植材料,12周编织骨交错排列,髓腔形成,成骨活性接近自体髂骨移植。B组,4周时有较多新骨形成;8周时出现大量胶原纤维,周边成骨明显;12周时,纤维组织减少,成骨活跃。结论异种DPB是一种良好的组织工程骨载体材料,可为再血管化和成骨细胞的分化提供一个稳定的环境。     

Skeletal muscle contractions uncoupled from gravitational loading directly increase cortical bone blood flow rates in vivo

The direct and indirect effects of muscle contraction on bone microcirculation and fluid flow are neither well documented nor explained. However, skeletal muscle contractions may affect the acquisition and maintenance of bone via stimulation of bone circulatory and interstitial fluid flow parameters. The purposes of this study were to assess the effects of transcutaneous electrical neuromuscular stimulation (TENS)‐induced muscle contractions on cortical bone blood flow and bone mineral content, and to demonstrate that alterations in blood flow could occur independently of mechanical loading and systemic circulatory mechanisms. Bone chamber implants were used in a rabbit model to observe real‐time blood flow rates and TENS‐induced muscle contractions. Video recording of fluorescent microspheres injected into the blood circulation was used to calculate changes in cortical blood flow rates. TENS‐induced repetitive muscle contractions uncoupled from mechanical loading instantaneously increased cortical microcirculatory flow, directly increased bone blood flow rates by 130%, and significantly increased bone mineral content over 7 weeks. Heart rates and blood pressure did not significantly increase due to TENS treatment. Our findings suggest that muscle contraction therapies have potential clinical applications for improving blood flow to cortical bone in the appendicular skeleton. © 2008 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 27: 651–656, 2009     

Mechanotransduction and functional response of the skeleton to physical stress: The mechanisms and mechanics of bone adaptation

The skeleton's primary mechanical function is to provide rigid levers for muscles to act against as they hold the body upright in defiance of gravity. Many bones are exposed to thousands of repetitive loads each day. During growth and development, the skeleton optimizes its architecture by subtle adaptations to these mechanical loads. The mechanisms for adaptation involve a multistep process of cellular mechanotransduction including: mechanocoupling— conversion of mechanical forces into local mechanical signals, such as fluid shear stresses, that initiate a response by bone cells; biochemical coupling— transduction of a mechanical signal to a biochemical response involving pathways within the cell membrane and cytoskeleton; cell-to-cell signaling from the sensor cells (probably osteocytes and bone lining cells) to effector cells (osteoblasts or osteoclasts) using prostaglandins and nitric oxide as signaling molecules; and effector response— either bone formation or resorption to cause appropriate architectural changes. These architectural changes tend to adjust and improve the bone structure to its prevailing mechanical environment. Structural changes can be predicted, to some extent, by mathematical formulas derived from three fundamental rules: (1) bone adaptation is driven by dynamic, rather than static, loading; (2) extending the loading duration has a diminishing effect on further bone adaptation; (3) bone cells accommodate to a mechanical loading environment, making them less responsive to routine or customary loading signals. Received for publication on Dec. 25, 1997; accepted on Feb. 24, 1998     

骨膜带血管的自体骨改善股骨上段异体骨固定13例

目的：探讨骨膜带血管自体骨对促进异体骨固定的治疗。方法：回顾性分析总结了13例膜带血管自体骨促进异体骨固定的临床资料。结果：13例因人工关节翻修手术造成股骨上段骨缺损经骨膜带血管的自体骨促进异体骨固定的治疗，术后10例随访1－4．5年，效果满意。结论：股骨上段骨缺损的病例较少，但治疗困难。除应用定制的长柄假体外，采用异体骨移植同时应用骨膜带血管的自体骨促进其固定也不失为一种良策。     

Reduced bone loss in a murine model of postmenopausal osteoporosis lacking complement component 3

The growing field of osteoimmunology seeks to unravel the complex interdependence of the skeletal and immune systems. Notably, we and others have demonstrated that complement signaling influences the differentiation of osteoblasts and osteoclasts, the two primary cell types responsible for maintaining bone homeostasis. However, the net effect of complement on bone homeostasis in vivo was unknown. Our published in vitro mechanistic work led us to hypothesize that absence of complement component 3 (C3), a central protein in the complement activation cascade, protects against bone loss in the ovariectomy‐based model of postmenopausal osteoporosis. Indeed, we report here that, when compared to their C57BL/6J (WT) counterparts, ovariectomized C3 deficient mice experienced reduced bone loss at multiple sites and increased stiffness at the femoral neck, the latter potentially improving mechanical function. WT and B6;129S4‐C3^tm1Crr/J (C3^‐/‐) mice were either ovariectomized or sham‐operated at 6 weeks of age and euthanized at 12 weeks. MicroCT on harvested bones revealed that the trabecular bone volume fraction in the metaphyses of both the proximal tibiae and distal femora of ovariectomized C3^‐/‐ mice is significantly greater than that of their WT counterparts. Lumbar vertebrae showed significantly greater osteoid content and mineral apposition rates. Mechanical testing demonstrated significantly greater stiffness in the femoral necks of ovariectomized C3^‐/‐ mice. These results demonstrate that C3 deficiency reduces bone loss at ovariectomy and may improve mechanical properties. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 36:118–128, 2018.     

Indomethacin modulation of load-related stimulation of new bone formationin vivo

Summary The capacity of bone to organize and reorganize its structure in response to changing mechanical demands is well recognized. However, the mechanism by which the changing mechanical environment is detected, and the means by which this information is translated into a stimulus for structural modification, are not understood. A group of substances suggested to be involved in the initial transduction of strain information are the prostaglandins. In this experiment we used a single period of dynamic loading to stimulate an adaptive osteogenic responsein vivo. Loading was performed in the presence and absence of indomethacin. Measurements of the periosteum 5 days after loading showed that the presence of indomethacin at the time of loading reduced the osteogenic response. Though consistent with the hypothesis that prostaglandins are involved in the initial transduction of tissue strain into a biochemical response, this result is not sufficient to demonstrate this conclusively because reduced prostaglandin levels during the 24 hours immediately after the period of loading may affect many other points in the cascade of events between strain transduction and adaptive new bone formation. Furthermore, indomethacin at the relatively high levels we used (40 mg/kg) may have effects other than those on prostaglandin synthesis.     

Bone Morphometry Strongly Predicts Cortical Bone Stiffness and Strength,but Not Toughness,in Inbred Mouse Models of High and Low Bone Mass

Inbred strains of mice make useful models to study bone properties. Our aim was to compare bone competence and cortical morphometric parameters of two inbred strains to better determine the role of bone structure and geometry in the process of bone failure. Morphometric analysis was performed on 20 murine femora with a low bone mass (C57BL/6J; B6) and 20 murine femora with a high bone mass (C3H/HeJ; C3H) using desktop μCT. The bones were tested under three‐point bending to measure their mechanical properties. Results showed that the C3H strain is a more reproducible model regarding bone morphometric and mechanical phenotypes than the B6 strain. Bone strength, stiffness, yield force, yield displacement, and toughness, as well as morphometric traits, were all significantly different between the two strains, whereas postyield displacement was not. It was found that bone volume, cortical thickness, and cross‐sectional area predicted almost 80% (p < 0.05) of bone stiffness, strength, and yield force. Nevertheless, cortical bone postyield properties such as bone toughness could not be explained by morphometry, but postyield whitening was observed in that phase. In conclusion, we found that morphometric parameters are strong predictors of preyield but not postyield properties. The lack of morphometric influence on bone competence in the postyield phase in combination with the observed postyield whitening confirmed the important contribution of ultrastructure and microdamage in the process of overall bone failure behavior, especially in the postyield phase.     

The Weight-Rearing Capacity of Structural Elements in Femoral Necks

1. Up to the age about 60 the resistance of the femoral neck to compression forces is the same on the left and the right side. After that age the mechanical resistance may decrease considerably on one side. This might be of clinical significance in the explanation of neck fractures.

2. The inferior cortical layer of the femoral neck has a weight-bearing capacity of 40 per cent in adults.

3. The superior cortical layer has a weight-bearing capacity of 20 per cent.

4. The superior and inferior cancellous tissue are of approximately equal value and each takes care of 15 per cent of the weight-bearing function.

5. If both cancellous layers are destroyed at the same time, the weight- bearing capacity is reduced by 30 per cent.

6. If the inferior compact layer and the inferior cancellous layer are eliminated or the superior cortical and trabecular system are removed, the weight-bearing capacity will be 50 per cent of normal.

7. The measurements sem to indicate that the trabecular tissue corresponds to a reinforcing system which helps to increase stability. It might be compared with cables in a bridge construction, the actual foundation of which is represented by the compact tissue.

     

Young's modulus, bending strength, and tissue physical properties of human compact bone

The Young's modulus, bending strength, apparent density, and ash content of 155 human compact bone bending specimens were determined. Both Young's modulus (E) and bending strength (S) were strongly correlated to tissue dry apparent density (rho a). Based upon the correlation coefficient (R) and the percent deviation of the data from the regression curve (% dev.), these correlations were best described by power law relationships: E infinity rho a 1.54 (R2 = 0.79, % dev. = 2.4) and S infinity rho a 2.18 (R2 = 0.80, % dev. = 6.4). Bending strength was related to Young's modulus raised to the 1.26 power, implying a nonlinear relationship for these variables. We found a weak correlation between ash content and the mechanical behavior of the compact bone specimens, particularly Young's modulus, but could not statistically justify formulation of a more complex multivariate power model incorporating both density and ash content. Regional variations in strength and stiffness along the femoral shaft and within the cortex were also noted and were attributed primarily to differences in apparent density. The relationships formulated for the mechanical behavior of human compact bone are discussed in terms of the results of previous investigations of the mechanical behavior of nonhuman compact bone and human cancellous bone.     

Intervention timing of strontium treatment on estrogen depletion‐induced osteoporosis in rats: Bone microstructure and mechanics

Purpose

To evaluate the effect of intervention timing of Sr treatment on trabecular bone microstructure and mechanics.

Methods

Ninety female rats were randomly divided into three batches with three groups in each batch. Each group was divided according to the initiation timing of vehicle or strontium compound (SrC), which was at week 0 (early), 4 (mid‐term) and 8 (late) after the ovariectomy, respectively. The treatment lasted for 12 weeks. The trabecular bone biomechanical properties, trabecular bone tissue mechanical properties, trabecular bone microstructure, and bone remodeling were analyzed with mechanical testing, nanoindentation, microCT, and histomorphometry, respectively. The osteoblast and osteoclast phenotypic genes were analyzed with real‐time polymerase chain reaction (PCR).

Results

Early and mid‐term Sr treatment significantly increased biomechanical properties of trabecular bone, which was associated with increased microarchitecture parameters, increased bone formation parameters and up‐regulation of osteoblast‐related gene expression. Late Sr treatment failed to exert a beneficial effect on any of those parameters.

Conclusions

The beneficial effect of Sr was dependent on the intervention timing in ovariectomized rats. © 2013 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 32:477–484, 2014.     

Recipient‐derived angiogenesis with short term immunosuppression increases bone remodeling in bone vascularized composite allotransplantation: A pilot study in a swine tibial defect model

Current vascularized composite allotransplantation (VCA) transplantation protocols rely upon life‐long immune modulation to maintain tissue perfusion. Alternatively, bone‐only VCA viability may be maintained in small animal models using surgical angiogenesis from implanted autogenous vessels to develop a neoangiogenic bone circulation that will not be rejected. This study tests the method's efficacy in a large animal model as a bridge to clinical practice, quantifying the remodeling and mechanical properties of porcine tibial VCAs. A segmental tibial defect was reconstructed in Yucatan miniature swine by transplantation of a matched tibia segment from an immunologically mismatched donor. Microsurgical repair of nutrient vessels was performed in all pigs, with simultaneous intramedullary placement of an autogenous arteriovenous (AV) bundle in Group 2. Group 1 served as a no‐angiogenesis control. All received 2 weeks of immunosuppression. After 16 weeks, micro‐CT and histomorphometric analyses were used to evaluate healing and remodeling. Axial compression and nanoindentation studies evaluated bone mechanical properties. Micro‐CT analysis demonstrated significantly more new bone formation and bone remodeling at the distal allotransplant/recipient junction and on the endosteal surfaces of Group 2 tibias (p = 0.03). Elastic modulus and hardness were not adversely affected by angiogenesis. The combination of 2 weeks of immunosuppression and autogenous AV‐bundle implantation within a microsurgically transplanted tibial allotransplant permitted long‐term allotransplant survival over the study period of 16 weeks in this large animal model. Angiogenesis increased bone formation and remodeling without adverse mechanical effects. The method may allow future composite‐tissue allotransplantation of bone without the risks associated with long‐term immunosuppression. © 2016 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 35:1242–1249, 2017.

     

Microencapsulation of rifampicin: A technique to preserve the mechanical properties of bone cement

Two‐stage exchange with antibiotic‐loaded bone cement spacers remains the gold standard for chronic periprosthetic joint infection (PJI). Rifampicin is highly efficient on stationary‐phase staphylococci in biofilm; however, its addition to PMMA to manufacture spacers prevents polymerization and reduces mechanical properties. Isolation of rifampicin during polymerization by microencapsulation could allow manufacturing rifampicin‐loaded bone cement maintaining elution and mechanical properties. Microcapsules of rifampicin with alginate, polyhydroxybutyratehydroxyvalerate (PHBV), ethylcellulose and stearic acid (SA) were synthesized. Alginate and PHBV microcapsules were added to bone cement and elution, compression, bending, hardness, setting time and microbiological tests were performed. Repeated measures ANOVA and Bonferroni post‐hoc test were performed, considering a p < 0.05 as statistical significance. Bone cement specimens containing alginate microcapsules eluted more rifampicin than PHBV microcapsules or non‐encapsulated rifampicin over time (p < 0.012). Microencapsulation of rifampicin allowed PMMA to preserve mechanical properties in compression and bending tests. Cement with alginate microcapsules showed similar behavior in hardness tests to control cement over the study period (73 ± 1.68H_D). PMMA with alginate microcapsules exhibited the largest zones of inhibition in microbiological tests. Statistically significant differences in mean diameters of zones of inhibition between PMMA loaded with alginate‐rifampicin (p = 0.0001) and alginate‐PHBV microcapsules (p = 0.0001) were detected. Rifampicin microencapsulation with alginate is the best choice to introduce rifampicin in PMMA preserving mechanical properties, setting time, elution, and antimicrobial properties. The main applicability of this study is the opportunity for obtaining rifampicin‐loaded PMMA by microencapsulation of rifampicin in alginate microparticles, achieving high doses of rifampicin in infected tissues, increasing the successful of PJI treatment. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 36:459–466, 2018.

     

Mechanical properties of collagen from decalcified rat femur in relation to age andin vitro maturation

Summary Cortical bone collagen was obtained by decalcifying femoral bones from 2-,5-,15-, and 25-month-old male rats. Collagen specimens were cut longitudinal to the long axis of the femur and tested mechanically. The maximum load (ultimate strength) and maximum slope of the load-strain curve (maximum stiffness) were found to decrease with age. The age-related reduction in the mechanical parameters resulted from a change in the mechanical strength of the constituent collagen and a change in the morphology of the bone tissue.In vitro aging, produced by incubating for 0–5 months, did not change the mechanical strength of bone collagen specimens obtained from 2-month-old rats. The changes in the mechanical characteristics of bone collagen, accompanyingin vivo andin vitro aging are the opposite of those observed in soft tissue collagen. In the latter there is an increase in the mechanical strength duringin vivo andin vitro aging.     

Effects of preexisting microdamage,collagen cross‐links,degree of mineralization,age, and architecture on compressive mechanical properties of elderly human vertebral trabecular bone

Previous studies have shown that the mechanical properties of trabecular bone are determined by bone volume fraction (BV/TV) and microarchitecture. The purpose of this study was to explore other possible determinants of the mechanical properties of vertebral trabecular bone, namely collagen cross‐link content, microdamage, and mineralization. Trabecular bone cores were collected from human L2 vertebrae (n = 49) from recently deceased donors 54–95 years of age (21 men and 27 women). Two trabecular cores were obtained from each vertebra, one for preexisting microdamage and mineralization measurements, and one for BV/TV and quasi‐static compression tests. Collagen cross‐link content (PYD, DPD, and PEN) was measured on surrounding trabecular bone. Advancing age was associated with impaired mechanical properties, and with increased microdamage, even after adjustment by BV/TV. BV/TV was the strongest determinant of elastic modulus and ultimate strength (r² = 0.44 and 0.55, respectively). Microdamage, mineralization parameters, and collagen cross‐link content were not associated with mechanical properties. These data indicate that the compressive strength of human vertebral trabecular bone is primarily determined by the amount of trabecular bone, and notably unaffected by normal variation in other factors, such as cross‐link profile, microdamage and mineralization. © 2010 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 29:481–488, 2011     

机械敏感离子通道蛋白Piezo1响应机械力刺激调节骨代谢的研究进展

随着人们生活方式的改变和人口老龄化,骨质疏松症的发生率逐年增加,已经渐渐成为重大的公共卫生问题。骨骼是一种相对动态的器官,可以为身体结构提供刚度、形状、支撑和运动。骨骼强度直接与死亡率和受伤风险相关,身体活动通常被认为是影响一生骨密度的环境因素,可是骨对于机械负荷的应答过程及其分子机制在很大程度上仍然未知。机械敏感阳离子通道蛋白Piezo1作为机械力刺激间充质干细胞、成骨细胞、破骨细胞、软骨细胞等产生变化的响应靶点,为机械负荷刺激（运动）改善骨质疏松等骨代谢疾病提供了新的思路和方向。