Three-Dimensional Morphology of Microdamage in Peri-Screw Bone: A Scanning Electron Microscopy of Methylmethacrylate Cast Replica

Authors

Lei Wang, Jin Shao, Tingjun Yea, Lianfu Deng, and Shijing Qiu

Abstract

Screw implantation inevitably causes microdamage in surrounding bone. However, little is known about the detailed characteristics of microdamage in peri-screw bone. In this study, we developed a method to construct microdamage cast with methylmethacrylate (MMA) and observed the cast using scanning electron microscopy (SEM). In basic fuchsin stained bone sections observed by bright-field and fluorescence microscopy, diffuse damage, cross-hatched damage, and linear cracks were all presented in peri-screw bone. Using MMA casting/SEM method, we found numerous densely packed microcracks in the areas with diffuse damage. The osteocyte canaliculi and the microcracks consisting of diffuse damage had a similar diameter (or width), usually <0.5 μm, but their morphology was largely different. In the area with cross-hatched damage, the orientation of microcracks was similar to that in diffuse damage, but the number was significantly decreased. Many microcracks were thicker than 1 μm and associated with a rough surface. Large linear cracks (∼10 μm in diameter) occurred in different areas. Plenty of microcracks were present on the surface of some linear cracks. In conclusion, the MMA casting/SEM method can demonstrate the three-dimensional morphology of different types of microdamage, particularly the microcracks in diffuse damage, which are unable to be shown by light microscopy.

Link to Article

http://dx.doi.org/10.1017/S1431927612001286

Angiogenesis is required for stress fracture healing in rats

Authors

Ryan E. Tomlinson, Jennifer A. McKenzie, Anne H. Schmieder, Gregory R. Wohl, Gregory M. Lanza, Matthew J. Silva

Abstract

Although angiogenesis and osteogenesis are critically linked, the importance of angiogenesis for stress fracture healing is unknown. In this study, mechanical loading was used to create a non-displaced stress fracture in the adult rat forelimb. Fumagillin, an anti-angiogenic agent, was used as the water soluble analogue TNP-470 (25 mg/kg) as well as incorporated into lipid-encapsulated αvβ3 integrin targeted nanoparticles (0.25 mg/kg). In the first experiment, TNP-470 was administered daily for 5 days following mechanical loading, and changes in gene expression, vascularity, and woven bone formation were quantified. Although no changes in vascularity were detected 3 days after loading, treatment-related downregulation of angiogenic (Pecam1) and osteogenic (Bsp, Osx) genes was observed at this early time point. On day 7, microCT imaging of loaded limbs revealed diminished woven bone formation in treated limbs compared to vehicle treated limbs. In the second experiment, αvβ3 integrin targeted fumagillin nanoparticles were administered as before, albeit with a 100-fold lower dose, and changes in vascularity and woven bone formation were determined. There were no treatment-related changes in vessel count or volume 3 days after loading, although fewer angiogenic (CD105 positive) blood vessels were present in treated limbs compared to vehicle treated limbs. This result manifested on day 7 as a reduction in total vascularity, as measured by histology (vessel count) and microCT (vessel volume). Similar to the first experiment, treated limbs had diminished woven bone formation on day 7 compared to vehicle treated limbs. These results indicate that angiogenesis is required for stress fracture healing, and may have implications for inducing rapid repair of stress fractures.

Link to Article

http://dx.doi.org/10.1016/j.bone.2012.09.035

The kinase TBK1 controls IgA class switching by negatively regulating noncanonical NF-κB signaling

Authors

Jin Jin, Yichuan Xiao, Jae-Hoon Chang, Jiayi Yu, Hongbo Hu, Robyn Starr, George C Brittain, Mikyoung Chang, Xuhong Cheng, and Shao-Cong Sun

Abstract

Immunoglobulin class switching is crucial for the generation of antibody diversity in humoral immunity and, when deregulated, also has severe pathological consequences. How the magnitude of immunoglobulin isotype switching is controlled is still poorly understood. Here we identify the kinase TBK1 as a pivotal negative regulator of class switching to the immunoglobulin A (IgA) isotope. B cell–specific ablation of TBK1 in mice resulted in uncontrolled production of IgA and the development of nephropathy-like disease signs. TBK1 negatively regulated IgA class switching by attenuating noncanonical signaling via the transcription factor NF-κB, an action that involved TBK1-mediated phosphorylation and subsequent degradation of the NF-κB-inducing kinase NIK. Our findings establish TBK1 as a pivotal negative regulator of the noncanonical NF-κB pathway and identify a unique mechanism that controls IgA production.

Link to Article

http://dx.doi.org/10.1038/ni.2423

Central Depletion of Brain-Derived Neurotrophic Factor in Mice Results in High Bone Mass and Metabolic Phenotype

Authors

Shanil S. Juma, Zahra Ezzat-Zadeh, Dania A. Khalil, Shirin Hooshmand, Mohammed Akhter, Bahram H. Arjmandi

Abstract

Brain-derived neurotrophic factor (BDNF) plays important roles in neuronal differentiation/survival, the regulation of food intake, and the pathobiology of obesity and type 2 diabetes mellitus. BDNF and its receptor are expressed in osteoblasts and chondrocyte. BDNF in vitro has a positive effect on bone; whether central BDNF affects bone mass in vivo is not known. We therefore examined bone mass and energy use in brain-targeted BDNF conditional knockout mice (Bdnf2lox/2lox/93). The deletion of BDNF in the brain led to a metabolic phenotype characterized by hyperphagia, obesity, and increased abdominal white adipose tissue. Central BDNF deletion produces a marked skeletal phenotype characterized by increased femur length, elevated whole bone mineral density, and bone mineral content. The skeletal changes are developmentally regulated and appear concurrently with the metabolic phenotype, suggesting that the metabolic and skeletal actions of BDNF are linked. The increased bone development is evident in both the cortical and trabecular regions. Compared with control, Bdnf2lox/2lox/93 mice show greater trabecular bone volume (+50% for distal femur, P < 0.001; +35% for vertebral body, P < 0.001) and midfemoral cortical thickness (+11 to 17%, P < 0.05), measured at 3 and 6 months of age. The skeletal and metabolic phenotypes were gender dependent, with female being more affected than male mice. However, uncoupling protein-1 expression in brown fat, a marker of sympathetic tone, was not different between genotypes. We show that deletion of central BDNF expression in mice results in increased bone mass and white adipose tissue, with no significant changes in sympathetic signaling or peripheral serotonin, associated with hyperphagia, obesity, and leptin resistance.

Link to Article

http://dx.doi.org/10.1210/en.2012-1378

Soy protein with or without isoflavones failed to preserve bone density in gonadal hormone–deficient male rat model of osteoporosis

Authors

Shanil S. Juma, Zahra Ezzat-Zadeh, Dania A. Khalil, Shirin Hooshmand, Mohammed Akhter, Bahram H. Arjmandi

Abstract

Soy with its isoflavones has been shown to positively influence bone mineral density in female ovariectomized rats; hence, we hypothesized a similar effect in orchidectomized (ORX) male rats. Forty male Sprague-Dawley rats, aged 95 days, were divided into 4 groups and were either sham operated (Sham) or ORX. The ORX groups were fed a soy protein–based diet (SOY), an isoflavone-depleted soy protein diet (SOY−), or a casein based diet for 65 days after surgery. Orchidectomy increased the rate of bone turnover, resulting in reduced bone mineral density and bone mineral content by 3.5% and 14%, respectively, and compromised biomechanical properties. The mean femoral length of ORX animals was also significantly shorter than Sham animals, but ORX rats that were fed SOY diet did not experience this reduction in bone length, implicating a role for soy protein in bone growth (4.02 ± 0.02, 3.93 ± 0.01, 3.99 ± 0.02, 3.91 ± 0.01 for Sham, ORX, SOY, SOY−, respectively). The SOY and SOY− positively influenced the biomechanical properties of bone such as yield and ultimate force, the measures of bone elasticity, and plasticity. In terms of bone histomorphometry, the data indicate that SOY− tends to reduce ORX-induced increase in bone turnover as evidenced by suppressed bone formation rate/mineralized surface by about 9%. Overall, our results indicated that soy protein, regardless of its isoflavone content, was unable to prevent the ORX-induced femoral decrease in bone density and mineral content. However, soy may enhance the quality of bone as indicated by increased yield force.

Link to Article

http://dx.doi.org/10.1016/j.nutres.2012.08.001

Enhanced Periosteal and Endocortical Responses to Axial Tibial Compression Loading in Conditional Connexin43 Deficient Mice

Authors

Susan K. Grimston, Marcus P. Watkins, Michael D. Brodt, Matthew J. Silva, Roberto Civitelli

Abstract

The gap junction protein, connexin43 (Cx43) is involved in mechanotransduction in bone. Recent studies using in vivo models of conditional Cx43 gene (Gja1) deletion in the osteogenic linage have generated inconsistent results, with Gja1 ablation resulting in either attenuated or enhanced response to mechanical load, depending upon the skeletal site examined or the type of load applied. To gain further insights on Cx43 and mechanotransduction, we examined bone formation response at both endocortical and periosteal surfaces in 2-month-old mice with conditional Gja1 ablation driven by the Dermo1 promoter (cKO). Relative to wild type (WT) littermates, it requires a larger amount of compressive force to generate the same periosteal strain in cKO mice. Importantly, cKO mice activate periosteal bone formation at a lower strain level than do WT mice, suggesting an increased sensitivity to mechanical load in Cx43 deficiency. Consistently, trabecular bone mass also increases in mutant mice upon load, while it decreases in WT. On the other hand, bone formation actually decreases on the endocortical surface in WT mice upon application of axial mechanical load, and this response is also accentuated in cKO mice. These changes are associated with increase of Cox-2 in both genotypes and further decrease of Sost mRNA in cKO relative to WT bones. Thus, the response of bone forming cells to mechanical load differs between trabecular and cortical components, and remarkably between endocortical and periosteal envelopes. Cx43 deficiency enhances both the periosteal and endocortical response to mechanical load applied as axial compression in growing mice.

Link to Article

http://dx.doi.org/10.1371/journal.pone.0044222