Yi-Xian Qin

Quantity and Quality of Trabecular Bone in the Femur Are Enhanced by a Strongly Anabolic, Noninvasive Mechanical Intervention

The skeletons sensitivity to mechanical stimuli represents a critical determinant of bone mass and morphology. We have proposed that the extremely low level (<10 microstrain), high frequency (20‐50 Hz) mechanical strains, continually present during even subtle activities such as standing are as important to defining the skeleton as the larger strains typically associated with vigorous activity (>2000 microstrain). If these low‐level strains are indeed anabolic, then this sensitivity could serve as the basis for a biomechanically based intervention for osteoporosis. To evaluate this hypothesis, the hindlimbs of adult female sheep were stimulated for 20 minutes/day using a noninvasive 0.3g vertical oscillation sufficient to induce approximately 5 microstrain on the cortex of the tibia. After 1 year of stimulation, the physical properties of 10‐mm cubes of trabecular bone from the distal femoral condyle of experimental animals (n = 8) were compared with controls (n = 9), as evaluated using microcomputed tomography (μCT) scanning and materials testing. Bone mineral content (BMC) was 10.6% greater (p < 0.05), and the trabecular number (Tb.N) was 8.3% higher in the experimental animals (p < 0.01), and trabecular spacing decreased by 11.3% (p < 0.01), indicating that bone quantity was increased both by the creation of new trabeculae and the thickening of existing trabeculae. The trabecular bone pattern factor (TBPf) decreased 24.2% (p < 0.03), indicating trabecular morphology adapting from rod shape to plate shape. Significant increases in stiffness and strength were observed in the longitudinal direction (12.1% and 26.7%, respectively; both, p < 0.05), indicating that the adaptation occurred primarily in the plane of weightbearing. These results show that extremely low level mechanical stimuli improve both the quantity and the quality of trabecular bone. That these deformations are several orders of magnitude below those peak strains which arise during vigorous activity indicates that this biomechanically based signal may serve as an effective intervention for osteoporosis.

Mechanical signals as anabolic agents in bone

Aging and a sedentary lifestyle conspire to reduce bone quantity and quality, decrease muscle mass and strength, and undermine postural stability, culminating in an elevated risk of skeletal fracture. Concurrently, a marked reduction in the available bone-marrow-derived population of mesenchymal stem cells (MSCs) jeopardizes the regenerative potential that is critical to recovery from musculoskeletal injury and disease. A potential way to combat the deterioration involves harnessing the sensitivity of bone to mechanical signals, which is crucial in defining, maintaining and recovering bone mass. To effectively utilize mechanical signals in the clinic as a non-drug-based intervention for osteoporosis, it is essential to identify the components of the mechanical challenge that are critical to the anabolic process. Large, intense challenges to the skeleton are generally presumed to be the most osteogenic, but brief exposure to mechanical signals of high frequency and extremely low intensity, several orders of magnitude below those that arise during strenuous activity, have been shown to provide a significant anabolic stimulus to bone. Along with positively influencing osteoblast and osteocyte activity, these low-magnitude mechanical signals bias MSC differentiation towards osteoblastogenesis and away from adipogenesis. Mechanical targeting of the bone marrow stem-cell pool might, therefore, represent a novel, drug-free means of slowing the age-related decline of the musculoskeletal system.

Fluid pressure gradients, arising from oscillations in intramedullary pressure, is correlated with the formation of bone and inhibition of intracortical porosity

Fluid flow that arises from the functional loading of bone tissue has been proposed to be a critical regulator of skeletal mass and morphology. To test this hypothesis, the bone adaptive response to a physiological fluid stimulus, driven by low magnitude, high frequency oscillations of intramedullary pressure (ImP), were examined, in which fluid pressures were achieved without deforming the bone tissue. The ulnae of adult turkeys were functionally isolated via transverse epiphyseal osteotomies, and the adaptive response to four weeks of disuse (n=5) was compared to disuse plus 10 min per day of a physiological sinusoidal fluid pressure signal (60 mmHg, 20Hz). Disuse alone resulted in significant bone loss (5.7+/-1.9%, p< or =0.05), achieved by thinning the cortex via endosteal resorption and an increase in intracortical porosity. By also subjecting bone to oscillatory fluid flow, a significant increase in bone mass at the mid-diaphysis (18.3+/-7.6%, p<0.05), was achieved by both periosteal and endosteal new bone formation. The spatial distribution of the transcortical fluid pressure gradients (inverted Delta P(r)), a parameter closely related to fluid velocity and fluid shear stress, was quantified in 12 equal sectors across a section at the mid-diaphyses. A strong correlation was found between the inverted Delta P(r) and total new bone formation (r=0.75, p=0.01); and an inverse correlation (r=-0.75, p=0.01) observed between inverted Delta P(r) and the area of increased intracortical porosity, indicating that fluid flow signals were necessary to maintain bone mass and/or inhibit bone loss against the challenge of disuse. By generating this fluid flow in the absence of matrix strain, these data suggest that anabolic fluid movement plays a regulatory role in the modeling and remodeling process. While ImP increases uniformly in the marrow cavity, the distinct parameters of fluid flow vary substantially due to the geometry and ultrastructure of bone, which ultimately defines the spatial non-uniformity of the adaptive process.

Patterns of strain in the macaque tibia during functional activity

The strain environment of the tibial midshaft of two female macaques was evaluated through in vivo bone strain experiments using three rosette gauges around the circumference of the bones. Strains were collected for a total of 123 walking and galloping steps as well as several climbing cycles. Principal strains and the angle of the maximum (tensile) principal strain with the long axis of the bone were calculated for each gauge site. In addition, the normal strain distribution throughout the cross section was determined from the longitudinal normal strains (strains in the direction of the long axis of the bone) at each of the three gauge sites, and at the corresponding cross-sectional geometry of the bone. This strain distribution was compared with the cross-sectional properties (area moments) of the midshaft. For both animals, the predominant loading regime was found to be bending about an oblique axis running from anterolateral to posteromedial. The anterior and part of the medial cortex are in tension; the posterior and part of the lateral cortex are in compression. The axis of bending does not coincide with the maximum principal axis of the cross section, which runs mediolaterally. The bones are not especially buttressed in the plane of bending, but offer the greatest strength anteroposteriorly. The cross-sectional geometry therefore does not minimize strain or bone tissue. Peak tibial strains are slightly higher than the peak ulnar strains reported earlier for the same animals (Demes et al. [1998] Am J Phys Anthropol 106:87-100). Peak strains for both the tibia and the ulna are moderate in comparison to strains recorded during walking and galloping activities in nonprimate mammals.

Two-Dimensional Nanostructure- Reinforced Biodegradable Polymeric Nanocomposites for Bone Tissue Engineering

This study investigates the efficacy of two-dimensional (2D) carbon and inorganic nanostructures as reinforcing agents for cross-linked composites of the biodegradable and biocompatible polymer polypropylene fumarate (PPF) as a function of nanostructure concentration. PPF composites were reinforced using various 2D nanostructures: single- and multiwalled graphene oxide nanoribbons (SWGONRs, MWGONRs), graphene oxide nanoplatelets (GONPs), and molybdenum disulfide nanoplatelets (MSNPs) at 0.01-0.2 weight% concentrations. Cross-linked PPF was used as the baseline control, and PPF composites reinforced with single- or multiwalled carbon nanotubes (SWCNTs, MWCNTs) were used as positive controls. Compression and flexural testing show a significant enhancement (i.e., compressive modulus = 35-108%, compressive yield strength = 26-93%, flexural modulus = 15-53%, and flexural yield strength = 101-262% greater than the baseline control) in the mechanical properties of the 2D-reinforced PPF nanocomposites. MSNP nanocomposites consistently showed the highest values among the experimental or control groups in all the mechanical measurements. In general, the inorganic nanoparticle MSNP showed a better or equivalent mechanical reinforcement compared to carbon nanomaterials, and 2D nanostructures (GONPs, MSNPs) are better reinforcing agents compared to one-dimensional (1D) nanostructures (e.g., SWCNTs). The results also indicated that the extent of mechanical reinforcement is closely dependent on the nanostructure morphology and follows the trend nanoplatelets > nanoribbons > nanotubes. Transmission electron microscopy of the cross-linked nanocomposites indicated good dispersion of nanomaterials in the polymer matrix without the use of a surfactant. The sol-fraction analysis showed significant changes in the polymer cross-linking in the presence of MSNP (0.01-0.2 wt %) and higher loading concentrations of GONP and MWGONR (0.1-0.2 wt %). The analysis of surface area and aspect ratio of the nanostructures taken together with the above results indicated differences in nanostructure architecture (2D vs 1D nanostructures), and the chemical compositions (inorganic vs carbon nanostructures), number of functional groups, and structural defects for the 2D nanostructures may be key properties that affect the mechanical properties of 2D nanostructure-reinforced PPF nanocomposites and the reason for the enhanced mechanical properties compared to the controls.

Inhibition of osteopenia by low magnitude, high-frequency mechanical stimuli.

The identification of anabolic agents for the treatment of metabolic bone disease is a highly prized, and elusive, goal. In searching for the osteogenic (bone-producing) constituents within mechanical stimuli, it was determined that high frequency (10-100 Hz) and low magnitude (<10 microstrain) stimuli were capable of augmenting bone mass and morphology, thereby benefiting both bone quantity and quality. Using animal models, it is shown that these mechanical signals can double bone-formation rates, inhibit disuse osteoporosis and increase the strength of trabecular bone by 25%. Considering that the magnitude of these mechanical signals are several orders of magnitude below those which cause damage to the bone tissue, it is proposed that this modality could be useful in the treatment of metabolic bone diseases.

Differentiation of the Bone-Tissue Remodeling Response to Axial and Torsional Loading in the Turkey Ulna*†

The ability of bone tissue to differentiate between axial and torsional loading was determined with use of a functionally isolated turkey-ulna model of bone adaptation. Surface modeling and intracortical remodeling were quantified after four weeks of 5000 cycles per day of axial loading sufficient to cause 1000 microstrain normal to the long axis of the bone (five ulnae), 5000 cycles per day of torsional loading sufficient to cause 1000 microstrain of shear strain (five ulnae), or disuse (six ulnae). Of these three distinct regimens, only disuse caused a significant change in gross areal properties (12 per cent loss of bone; p < 0.05) as compared with those in the contralateral, intact control ulnae (sixteen ulnae). This finding suggested that both axial and torsional loading conditions were suitable substitutes for functional signals normally responsible for bone homeostasis. However, the intracortical response was strongly dependent on the manner in which the bone was loaded. Axial loading increased the number of intracortical pores by a factor of seven as compared with that in the controls (246 ± 40.5 compared with 36 ± 8.5 pores); it also increased the area lost because of porosis as compared with that in the controls (1.39 ± 0.252 compared with 0.202 ± 0.062 square millimeter); however, the mean size of the individual pores was similar to that in the controls (0.00565 ± 0.0019 compared with 0.00561 ± 0.0029 square millimeter). Conversely, torsional loading failed to increase substantially the number of pores (67 ± 22.6 pores), the area of bone lost because of porosis (0.352 ± 0.114 square millimeter), or the size of the pores (0.00525 ± 0.0035 square millimeter) as compared with those in the controls. Although disuse failed to increase substantially the number of intracortical pores (59 ± 22.4 pores), significant area (1.05 ± 0.35 square millimeters; p < 0.05) was lost within the cortex because of a threefold increase in the mean size of each pore (0.0178 ± 0.0126 square millimeter). It appears that bone tissue can readily differentiate between distinct components of the strain environment, with strain per se necessary to retain coupled formation and resorption, shear strain achieving this goal by maintaining the status quo, and axial strain increasing intracortical turnover but retaining coupling. While it is clear that load influences bone mass and morphology, it is also clear that specific parameters within the strain environment have distinct strategic roles in defining this architecture. CLINICAL RELEVANCE: These data demonstrate that the processes that control the modeling and remodeling of bone tissue are capable of distinguishing between specific parameters of the functional strain and stress milieu. Shear strain minimizes bone turnover, suggesting that exercise or rehabilitation programs, or both, for the inhibition of osteoporosis should maximize diverse activities. If a high degree of bone turnover is warranted, as in the promotion of fracture-healing or bone ingrowth, axial conditions should prevail. With an improved understanding of the mechanisms that permit the cell to differentiate between changes in its volume and shape, the ability to treat many musculoskeletal disorders as well as to regulate the osseous response to procedures such as joint reconstruction, distraction osteogenesis, and fracture-healing will improve.

Adaptations of Trabecular Bone to Low Magnitude Vibrations Result in More Uniform Stress and Strain Under Load

Extremely low magnitude mechanical stimuli (<10 microstrain) induced at high frequencies are anabolic to trabecular bone. Here, we used finite element (FE) modeling to investigate the mechanical implications of a one year mechanical intervention. Adult female sheep stood with their hindlimbs either on a vibrating plate (30 Hz, 0.3 g) for 20 min/d, 5 d/wk or on an inactive plate. Microcomputed tomography data of 1 cm bone cubes extracted from the medial femoral condyles were transformed into FE meshes. Simulated compressive loads applied to the trabecular meshes in the three orthogonal directions indicated that the low level mechanical intervention significantly increased the apparent trabecular tissue stiffness of the femoral condyle in the longitudinal (+17%, p < 0.02), anterior–posterior (+29%, p < 0.01), and medial-lateral (+37%, p < 0.01) direction, thus reducing apparent strain magnitudes for a given applied load. For a given apparent input strain (or stress), the resultant stresses and strains within trabeculae were more uniformly distributed in the off-axis loading directions in cubes of mechanically loaded sheep. These data suggest that trabecular bone responds to low level mechanical loads with intricate adaptations beyond a simple reduction in apparent strain magnitude, producing a structure that is stiffer and less prone to fracture for a given load.

The pathway of bone fluid flow as defined by in vivo intramedullary pressure and streaming potential measurements.

AbstractThe pathway for intracortical fluid flow response to a step-load was identified in vivo using intramedullary pressure (ImP) and streaming potential (SP) measurements, and allowed the development of a load-induced flow mechanism which considers mechanotransduction and mechanoelectrotransduction phenomena. An avian model was used for monitoring, simultaneously, ImP and SP under axial loading which generated peak strains of approximately 600 microstrain (με). ImP response to step-load decayed more quickly than SP relaxation, in which multiple time constants were observed during the relaxations. While the initial relaxation of SP showed a decay on the order of 200 ms, ImP decayed on the order of approximately 100 ms. After the initial decay (∼200 ms after loading), ImP quickly relaxed to base line, while SP continued to dominate relaxation. It appears that the decay of ImP is indicative of resistive fluid flow occurring primarily in the vasculature and other intraosseous channels such as lacunar-canalicular pores, and that SP represents the fluid flow in the smaller porosities, i.e., lacunar-canalicular system or even microspores. These results suggest that SP and ImP decays are determined by a hierarchical interdependent system of multiple porosities, and that the temporal dynamics of load-bearing define the manner in which the fluid patterns and pressures are distributed.

Tungsten disulfide nanotubes reinforced biodegradable polymers for bone tissue engineering

In this study, we have investigated the efficacy of inorganic nanotubes as reinforcing agents to improve the mechanical properties of poly(propylene fumarate) (PPF) composites as a function of nanomaterial loading concentration (0.01-0.2 wt.%). Tungsten disulfide nanotubes (WSNTs) were used as reinforcing agents in the experimental group. Single- and multi-walled carbon nanotubes (SWCNTs and MWCNTs) were used as positive controls, and crosslinked PPF composites were used as the baseline control. Mechanical testing (compression and three-point bending) shows a significant enhancement (up to 28-190%) in the mechanical properties (compressive modulus, compressive yield strength, flexural modulus and flexural yield strength) of WSNT-reinforced PPF nanocomposites compared to the baseline control. In comparison to the positive controls, significant improvements in the mechanical properties of WSNT nanocomposites were also observed at various concentrations. In general, the inorganic nanotubes (WSNTs) showed mechanical reinforcement better than (up to 127%) or equivalent to that of carbon nanotubes (SWCNTs and MWCNTs). Sol fraction analysis showed significant increases in the crosslinking density of PPF in the presence of WSNTs (0.01-0.2 wt.%). Transmission electron microscopy (TEM) analysis on thin sections of crosslinked nanocomposites showed the presence of WSNTs as individual nanotubes in the PPF matrix, whereas SWCNTs and MWCNTs existed as micron-sized aggregates. The trend in the surface area of nanostructures obtained by Brunauer-Emmett-Teller (BET) surface area analysis was SWCNTs>MWCNTs>WSNTs. The BET surface area analysis, TEM analysis and sol fraction analysis results taken together suggest that chemical composition (inorganic vs. carbon nanomaterials), the presence of functional groups (such as sulfide and oxysulfide) and individual dispersion of the nanomaterials in the polymer matrix (absence of aggregation of the reinforcing agent) are the key parameters affecting the mechanical properties of nanostructure-reinforced PPF composites and the reason for the observed increases in the mechanical properties compared to the baseline and positive controls.