X. Sherry Liu

Site‐specific changes in bone microarchitecture, mineralization, and stiffness during lactation and after weaning in mice

Despite the dramatic bone loss that occurs during lactation, bone mineral density rapidly recovers after offspring are weaned and milk production stops. The goal of this study is to quantify site‐specific changes in bone quantity and quality during and after lactation in a mouse model. We used micro computed tomography (µCT), individual trabecula segmentation (ITS), digital topological analysis (DTA)‐based tissue mineral density (TMD) analysis, and micro finite element analysis (µFEA) to quantify the effects of lactation and weaning on bone microarchitecture, mineralization, and stiffness at the spine, tibia, and femur. We found a significant decrease in trabecular plate microarchitecture, tissue mineralization of the trabecular surface, trabecular central skeleton, and intervening envelopes, and whole bone stiffness in lactating versus nulliparous mice at all three sites. In recovered mice, all these different aspects of bone quality were comparable to nulliparous mice at the spine. In contrast, trabecular plate microarchitecture and whole bone stiffness at the tibia and femur in recovered mice were lower than nulliparous mice, as were central trabecular tissue mineralization and cortical structure at the femur. These findings are consistent with clinical observations of partial recovery of femoral bone mineral density BMD after lactation in humans. The observed differences in trabecular surface tissue mineralization in nulliparous, lactating, and recovered mice are consistent with prior observations that maternal bone turnover shifts from resorption to formation at the time of pup weaning. The significant differences in trabecular central tissue mineralization during these three states suggest that osteocytes may contribute to the reversible loss of mineral during and after lactation. Future studies are necessary to determine whether differing functions of various bone cells at individual skeletal sites cause site‐specific skeletal changes during and after lactation.

Skeletal structure in postmenopausal women with osteopenia and fractures is characterized by abnormal trabecular plates and cortical thinning.

The majority of fragility fractures occur in women with osteopenia rather than osteoporosis as determined by dual‐energy X‐ray absorptiometry (DXA). However, it is difficult to identify which women with osteopenia are at greatest risk. We performed this study to determine whether osteopenic women with and without fractures had differences in trabecular morphology and biomechanical properties of bone. We hypothesized that women with fractures would have fewer trabecular plates, less trabecular connectivity, and lower stiffness. We enrolled 117 postmenopausal women with osteopenia by DXA (mean age 66 years; 58 with fragility fractures and 59 nonfractured controls). All had areal bone mineral density (aBMD) measured by DXA. Trabecular and cortical volumetric bone mineral density (vBMD), trabecular microarchitecture, and cortical porosity were measured by high‐resolution peripheral computed tomography (HR‐pQCT) of the distal radius and tibia. HR‐pQCT scans were subjected to finite element analysis to estimate whole bone stiffness and individual trabecula segmentation (ITS) to evaluate trabecular type (as plate or rod), orientation, and connectivity. Groups had similar age, race, body mass index (BMI), and mean T‐scores. Fracture subjects had lower cortical and trabecular vBMD, thinner cortices, and thinner, more widely separated trabeculae. By ITS, fracture subjects had fewer trabecular plates, less axially aligned trabeculae, and less trabecular connectivity. Whole bone stiffness was lower in women with fractures. Cortical porosity did not differ. Differences in cortical bone were found at both sites, whereas trabecular differences were more pronounced at the radius. In summary, postmenopausal women with osteopenia and fractures had lower cortical and trabecular vBMD; thinner, more widely separated and rodlike trabecular structure; less trabecular connectivity; and lower whole bone stiffness compared with controls, despite similar aBMD by DXA. Our results suggest that in addition to trabecular and cortical bone loss, changes in plate and rod structure may be important mechanisms of fracture in postmenopausal women with osteopenia.

Kidney Transplantation with Early Corticosteroid Withdrawal: Paradoxical Effects at the Central and Peripheral Skeleton

The use of early corticosteroid withdrawal (ECSW) protocols after kidney transplantation has become common, but the effects on fracture risk and bone quality are unclear. We enrolled 47 first-time adult transplant recipients managed with ECSW into a 1-year study to evaluate changes in bone mass, microarchitecture, biomechanical competence, and remodeling with dual energy x-ray absorptiometry (DXA), high-resolution peripheral quantitative computed tomography (HRpQCT), parathyroid hormone (PTH) levels, and bone turnover markers obtained at baseline and 3, 6, and 12 months post-transplantation. Compared with baseline, 12-month areal bone mineral density by DXA did not change significantly at the spine and hip, but it declined significantly at the 1/3 and ultradistal radii (2.2% and 2.9%, respectively; both P<0.001). HRpQCT of the distal radius revealed declines in cortical area, density, and thickness (3.9%, 2.1%, and 3.1%, respectively; all P<0.001), trabecular density (4.4%; P<0.001), and stiffness and failure load (3.1% and 3.5%, respectively; both P<0.05). Findings were similar at the tibia. Increasing severity of hyperparathyroidism was associated with increased cortical losses. However, loss of trabecular bone and bone strength were most severe at the lowest and highest PTH levels. In summary, ECSW was associated with preservation of bone mineral density at the central skeleton; however, it was also associated with progressive declines in cortical and trabecular bone density at the peripheral skeleton. Cortical decreases related directly to PTH levels, whereas the relationship between PTH and trabecular bone decreases was bimodal. Studies are needed to determine whether pharmacologic agents that suppress PTH will prevent cortical and trabecular losses and post-transplant fractures.

Trabecular plates and rods determine elastic modulus and yield strength of human trabecular bone

The microstructure of trabecular bone is usually perceived as a collection of plate-like and rod-like trabeculae, which can be determined from the emerging high-resolution skeletal imaging modalities such as micro-computed tomography (μCT) or clinical high-resolution peripheral quantitative CT (HR-pQCT) using the individual trabecula segmentation (ITS) technique. It has been shown that the ITS-based plate and rod parameters are highly correlated with elastic modulus and yield strength of human trabecular bone. In the current study, plate-rod (PR) finite element (FE) models were constructed completely based on ITS-identified individual trabecular plates and rods. We hypothesized that PR FE can accurately and efficiently predict elastic modulus and yield strength of human trabecular bone. Human trabecular bone cores from proximal tibia (PT), femoral neck (FN) and greater trochanter (GT) were scanned by μCT. Specimen-specific ITS-based PR FE models were generated for each μCT image and corresponding voxel-based FE models were also generated in comparison. Both types of specimen-specific models were subjected to nonlinear FE analysis to predict the apparent elastic modulus and yield strength using the same trabecular bone tissue properties. Then, mechanical tests were performed to experimentally measure the apparent modulus and yield strength. Strong linear correlations for both elastic modulus (r(2) = 0.97) and yield strength (r(2) = 0.96) were found between the PR FE model predictions and experimental measures, suggesting that trabecular plate and rod morphology adequately captures three-dimensional (3D) microarchitecture of human trabecular bone. In addition, the PR FE model predictions in both elastic modulus and yield strength were highly correlated with the voxel-based FE models (r(2) = 0.99, r(2) = 0.98, respectively), resulted from the original 3D images without the PR segmentation. In conclusion, the ITS-based PR models predicted accurately both elastic modulus and yield strength determined experimentally across three distinct anatomic sites. Trabecular plates and rods accurately determine elastic modulus and yield strength of human trabecular bone.

Dependence of mechanical properties of trabecular bone on plate–rod microstructure determined by individual trabecula segmentation (ITS)

Individual trabecula segmentation (ITS) technique can decompose the trabecular bone network into individual trabecular plates and rods and is capable of quantifying the plate/rod-related microstructural characteristics of trabecular bone. This novel technique has been shown to be able to provide in-depth insights into micromechanics and failure mechanisms of human trabecular bone, as well as to distinguish the fracture status independent of area bone mineral density in clinical applications. However, the plate/rod microstructural parameters from ITS have never been correlated to experimentally determined mechanical properties of human trabecular bone. In this study, on-axis cylindrical trabecular bone samples from human proximal tibia (n=22), vertebral body (n=10), and proximal femur (n=21) were harvested, prepared, scanned using micro computed-tomography (µCT), analyzed with ITS and mechanically tested. Regression analyses showed that the plate bone volume fraction (pBV/TV) and axial bone volume fraction (aBV/TV) calculated by ITS analysis correlated the best with elastic modulus (R(2)=0.96-0.97) and yield strength (R(2)=0.95-0.96). Trabecular plate-related microstructural parameters correlated highly with elastic modulus and yield strength, while most rod-related parameters were found inversely and only moderately correlated with the mechanical properties. In addition, ITS analysis also identified that trabecular bone at human femoral neck had the highest trabecular plate-related parameters while the other sites were similar with each other in terms of plate-rod microstructure.

PTH1–34 alleviates radiotherapy-induced local bone loss by improving osteoblast and osteocyte survival

Cancer radiotherapy is often complicated by a spectrum of changes in the neighboring bone from mild osteopenia to osteoradionecrosis. We previously reported that parathyroid hormone (PTH, 1-34), an anabolic agent for osteoporosis, reversed bone structural deterioration caused by multiple microcomputed tomography (microCT) scans in adolescent rats. To simulate clinical radiotherapy for cancer patients and to search for remedies, we focally irradiated the tibial metaphyseal region of adult rats with a newly available small animal radiation research platform (SARRP) and treated these rats with intermittent injections of PTH1-34. Using a unique 3D image registration method that we recently developed, we traced the local changes of the same trabecular bone before and after treatments, and observed that, while radiation caused a loss of small trabecular elements leading to significant decreases in bone mass and strength, PTH1-34 preserved all trabecular elements in irradiated bone with remarkable increases in bone mass and strength. Histomorphometry demonstrated that SARRP radiation severely reduced osteoblast number and activity, which were impressively reversed by PTH treatment. In contrast, suppressing bone resorption by alendronate failed to rescue radiation-induced bone loss and to block the rescue effect of PTH1-34. Furthermore, histological analyses revealed that PTH1-34 protected osteoblasts and osteocytes from radiation-induced apoptosis and attenuated radiation-induced bone marrow adiposity. Taken together, our data strongly support a robust radioprotective effect of PTH on trabecular bone integrity through preserving bone formation and shed light on further investigations of an anabolic therapy for radiation-induced bone damage.

PTH prevents the adverse effects of focal radiation on bone architecture in young rats.

Radiation therapy is a common treatment regimen for cancer patients. However, its adverse effects on the neighboring bone could lead to fractures with a great impact on quality of life. The underlying mechanism is still elusive and there is no preventive or curative solution for this bone loss. Parathyroid hormone (PTH) is a current therapy for osteoporosis that has potent anabolic effects on bone. In this study, we found that focal radiation from frequent scans of the right tibiae in 1-month-old rats by micro-computed tomography severely decreased trabecular bone mass and deteriorated bone structure. Interestingly, PTH daily injections remarkably improved trabecular bone in the radiated tibiae with increases in trabecular number, thickness, connectivity, structure model index and stiffness, and a decrease in trabecular separation. Histomorphometric analysis revealed that radiation mainly decreased the number of osteoblasts and impaired their mineralization activity but had little effects on osteoclasts. PTH reversed these adverse effects and greatly increased bone formation to a similar level in both radiated and non-radiated bones. Furthermore, PTH protects bone marrow mesenchymal stem cells from radiation-induced damage, including a decrease in number and an increase in adipogenic differentiation. While radiation generated the same amount of free radicals in the bone marrow of vehicle-treated and PTH-treated animals, the percentage of apoptotic bone marrow cells was significantly attenuated in the PTH group. Taken together, our data demonstrate a radioprotective effect of PTH on bone structure and bone marrow and shed new light on a possible clinical application of anabolic treatment in radiotherapy.

Premenopausal and postmenopausal differences in bone microstructure and mechanical competence in Chinese-American and white women

Compared to white women, premenopausal Chinese‐American women have more plate‐like trabecular (Tb) bone. It is unclear whether these findings are relevant to postmenopausal women and if there are racial differences in the deterioration of bone microarchitecture with aging. We applied individual trabecula segmentation and finite element analysis to high‐resolution peripheral quantitative computed tomography images in premenopausal and postmenopausal Chinese‐American and white women to quantify within‐race age‐related differences in Tb plate‐versus‐rod microarchitecture and bone stiffness. Race–menopause status interactions were assessed. Comparisons between races within menopause status were adjusted for age, height and weight. Comparisons between premenopausal and postmenopausal women were adjusted for height and weight. Adjusted analyses at the radius indicated that premenopausal Chinese‐Americans had a higher plate bone volume fraction (pBV/TV), Tb plate‐to‐rod ratio (P‐R ratio), and greater plate‐plate junction densities (P‐P Junc.D) versus white women (all p < 0.01), resulting in 27% higher Tb stiffness (p < 0.05). Greater cortical thickness and density (Ct.Th and Dcort) and more Tb plates led to 19% greater whole bone stiffness (p < 0.05). Postmenopausal Chinese‐Americans had similar pBV/TV and P‐P Junc.D, yet a higher P‐R ratio versus white women. Postmenopausal Chinese‐American versus white women had greater Ct.Th, Dcort, and relatively intact Tb plates, resulting in similar Tb stiffness but 12% greater whole bone stiffness (p < 0.05). In both races, Ct.Th and Dcort were lower in postmenopausal versus premenopausal women and there were no differences between races. Tb plate parameters were also lower in postmenopausal versus premenopausal women, but age‐related differences in pBV/TV, P‐R ratio, and P‐P Junc D were greater (p < 0.05) in Chinese‐Americans versus white women. There are advantages in cortical and Tb bone in premenopausal Chinese‐American women. Within‐race cross‐sectional differences between premenopausal and postmenopausal women suggest greater loss of plate‐like Tb bone with aging in Chinese‐Americans, though thicker cortices and more plate‐like Tb bone persists.

Fast Trabecular Bone Strength Predictions of HR‐pQCT and Individual Trabeculae Segmentation–Based Plate and Rod Finite Element Model Discriminate Postmenopausal Vertebral Fractures

Although high‐resolution peripheral quantitative computed tomography (HR‐pQCT) has advanced clinical assessment of trabecular bone microstructure, nonlinear microstructural finite element (µFE) prediction of yield strength using a HR‐pQCT voxel model is impractical for clinical use due to its prohibitively high computational costs. The goal of this study was to develop an efficient HR‐pQCT‐based plate and rod (PR) modeling technique to fill the unmet clinical need for fast bone strength estimation. By using an individual trabecula segmentation (ITS) technique to segment the trabecular structure into individual plates and rods, a patient‐specific PR model was implemented by modeling each trabecular plate with multiple shell elements and each rod with a beam element. To validate this modeling technique, predictions by HR‐pQCT PR model were compared with those of the registered high‐resolution micro–computed tomography (HR‐µCT) voxel model of 19 trabecular subvolumes from human cadaveric tibia samples. Both the Youngs modulus and yield strength of HR‐pQCT PR models strongly correlated with those of µCT voxel models (r2 = 0.91 and 0.86). Notably, the HR‐pQCT PR models achieved major reductions in element number (>40‐fold) and computer central processing unit (CPU) time (>1200‐fold). Then, we applied PR model µFE analysis to HR‐pQCT images of 60 postmenopausal women with (n = 30) and without (n = 30) a history of vertebral fracture. HR‐pQCT PR model revealed significantly lower Youngs modulus and yield strength at the radius and tibia in fracture subjects compared to controls. Moreover, these mechanical measurements remained significantly lower in fracture subjects at both sites after adjustment for areal bone mineral density (aBMD) T‐score at the ultradistal radius or total hip. In conclusion, we validated a novel HR‐pQCT PR model of human trabecular bone against µCT voxel models and demonstrated its ability to discriminate vertebral fracture status in postmenopausal women. This accurate nonlinear µFE prediction of the HR‐pQCT PR model, which requires only seconds of desktop computer time, has tremendous promise for clinical assessment of bone strength.

3D image registration is critical to ensure accurate detection of longitudinal changes in trabecular bone density, microstructure, and stiffness measurements in rat tibiae by in vivo microcomputed tomography (μCT).

In the recent decade, in vivo μCT scanners have become available to monitor temporal changes in rodent bone in response to diseases and treatments. We investigated short-term and long-term precision of in vivo μCT measurements of trabecular bone density, microstructure and stiffness of rat tibiae and tested whether they can be improved by 3D image registration. Rats in the short-term precision group underwent baseline and follow-up scans within the same day (n = 15) and those in the long-term precision group were scanned at day 0 and day 14 (n = 16) at 10.5 μm voxel size. A 3D image-registration scheme was applied to register the trabecular bone compartments of baseline and follow-up scans. Prior to image registration, short-term precision ranged between 0.85% and 2.65% in bone volume fraction (BV/TV), trabecular number, thickness, and spacing (Tb.N*, Tb.Th*, Tb.Sp*), trabecular bone mineral density and tissue mineral density (Tb.BMD, and Tb.TMD), and was particularly high in structure model index (SMI), connectivity density (Conn.D), and stiffness (4.29%-8.83%). Image registration tended to improve the short-term precision, but the only statistically significant improvement was in Tb.N*, Tb.TMD, and stiffness. On the other hand, unregistered comparisons between day-0 and day-14 scans suggested significant increases in BV/TV, Tb.N*, Tb.Th*, Conn.D, and Tb.BMD and decrease in Tb.Sp* and SMI. However, the percent change in each parameter from registered comparisons was significantly different from unregistered comparisons. Registered results suggested a significant increase in BV/TV, Tb.BMD, and stiffness over 14 days, primarily caused by increased Tb.Th* and Tb.TMD. Due to the continuous growth of rodents, the direct comparisons between the unregistered baseline and follow-up scans were driven by changes due to global bone modeling instead of local remodeling. Our results suggested that 3D image registration is critical for detecting changes due to bone remodeling activities in rodent trabecular bone by in vivo μCT imaging.