Pilot Study: Unique Response of Bone Tissue During an Investigation of Radio-Adaptive Effects in Mice

PURPOSE: We obtained bone tissue to evaluate the collateral effects of experiments designed to investigate molecular mechanisms of radio-adaptation in a mouse model. Radio-adaptation describes a process by which the prior exposure to low dose radiation can protect against the toxic effect of a subsequent high dose exposure. In the radio-adaptation experiments, C57Bl/6 mice were exposed to either a Sham or a priming Low Dose (5 cGy) of Cs-137 gamma rays before being exposed to either a Sham or High Dose (6 Gy) 24 hours later. ANALYSIS: Bone tissue were obtained from two experiments where mice were sacrificed at 3 days (n=3/group, 12 total) and at 14 days (n=6/group, 24 total) following high dose exposure. Tissues were analyzed to 1) evaluate a radio-adaptive response in bone tissue and 2) describe cellular and microstructural effects for two skeletal sites with different rates of bone turnover. One tibia and one lumbar vertebrae (LV2), collected at the 3-day time-point, were analyzed by bone histomorphometry and micro-CT to evaluate the cellular response and any evidence of microarchitectural impact. Likewise, tibia and LV2, collected at the 14-day time-point, were analyzed by micro-CT alone to evaluate resulting changes to bone structure and microarchitecture. The data were analyzed by 2-way ANOVA to evaluate the effects of the priming low dose radiation, of the high dose radiation, and of any interaction between the priming low and high doses of radiation. Bone histomorphometry was performed in the cancellous bone (aka trabecular bone) compartments of the proximal tibial metaphysis and of LV2. RESULTS: Cellular Response @ 3 Days The priming Low Dose radiation decreased osteoblast-covered bone perimeter in the proximal tibia and the total cell density in the bone marrow in the LV2. High Dose radiation, regardless of prior exposure to priming dose, dramatically reduced total cell density in bone marrow of both the long bone and vertebra. However, in the proximal tibia, High Dose radiation increased the osteoclast-covered bone perimeters, the density of adipocytes in bone marrow, and the area of bone marrow occupied by fat cells -- while in the LV2, adipocytes were rare and not stimulated by High Dose radiation. In an unexpected response, High Dose radiation dramatically increased (10-fold) osteoblast-covered bone perimeter in the LV2

Pinwheel patterns and powder diffraction

Pinwheel patterns and their higher dimensional generalisations display continuous circular or spherical symmetries in spite of being perfectly ordered. The same symmetries show up in the corresponding diffraction images. Interestingly, they also arise from amorphous systems, and also from regular crystals when investigated by powder diffraction. We present first steps and results towards a general frame to investigate such systems, with emphasis on statistical properties that are helpful to understand and compare the diffraction images. We concentrate on properties that are accessible via an alternative substitution rule for the pinwheel tiling, based on two different prototiles. Due to striking similarities, we compare our results with the toy model for the powder diffraction of the square lattice.Comment: 7 pages, 4 figure

Doubly connected minimal surfaces and extremal harmonic mappings

The concept of a conformal deformation has two natural extensions: quasiconformal and harmonic mappings. Both classes do not preserve the conformal type of the domain, however they cannot change it in an arbitrary way. Doubly connected domains are where one first observes nontrivial conformal invariants. Herbert Groetzsch and Johannes C. C. Nitsche addressed this issue for quasiconformal and harmonic mappings, respectively. Combining these concepts we obtain sharp estimates for quasiconformal harmonic mappings between doubly connected domains. We then apply our results to the Cauchy problem for minimal surfaces, also known as the Bjorling problem. Specifically, we obtain a sharp estimate of the modulus of a doubly connected minimal surface that evolves from its inner boundary with a given initial slope.Comment: 35 pages, 2 figures. Minor edits, references adde

Erythroid Promoter Confines FGF2 Expression to the Marrow after Hematopoietic Stem Cell Gene Therapy and Leads to Enhanced Endosteal Bone Formation

Fibroblast growth factor-2 (FGF2) has been demonstrated to be a promising osteogenic factor for treating osteoporosis. Our earlier study shows that transplantation of mouse Sca-1+ hematopoietic stem/progenitor cells that are engineered to express a modified FGF2 leads to considerable endosteal/trabecular bone formation, but it also induces adverse effects like hypocalemia and osteomalacia. Here we report that the use of an erythroid specific promoter, β-globin, leads to a 5-fold decrease in the ratio of serum FGF2 to the FGF2 expression in the marrow cavity when compared to the use of a ubiquitous promoter spleen focus-forming virus (SFFV). The confined FGF2 expression promotes considerable trabeculae bone formation in endosteum and does not yield anemia and osteomalacia. The avoidance of anemia in the mice that received Sca1+ cells transduced with FGF2 driven by the β-globin promoter is likely due to attenuation of high-level serum FGF2-mediated stem cell mobilization observed in the SFFV-FGF2 animals. The prevention of osteomalacia is associated with substantially reduced serum Fgf23/hypophosphatemia, and less pronounced secondary hyperparathyroidism. Our improved stem cell gene therapy strategy represents one step closer to FGF2-based clinical therapy for systemic skeletal augmentation

Pathogenesis of adolescent idiopathic scoliosis in girls - a double neuro-osseous theory involving disharmony between two nervous systems, somatic and autonomic expressed in the spine and trunk: possible dependency on sympathetic nervous system and hormones with implications for medical therapy

Anthropometric data from three groups of adolescent girls - preoperative adolescent idiopathic scoliosis (AIS), screened for scoliosis and normals were analysed by comparing skeletal data between higher and lower body mass index subsets. Unexpected findings for each of skeletal maturation, asymmetries and overgrowth are not explained by prevailing theories of AIS pathogenesis. A speculative pathogenetic theory for girls is formulated after surveying evidence including: (1) the thoracospinal concept for right thoracic AIS in girls; (2) the new neuroskeletal biology relating the sympathetic nervous system to bone formation/resorption and bone growth; (3) white adipose tissue storing triglycerides and the adiposity hormone leptin which functions as satiety hormone and sentinel of energy balance to the hypothalamus for long-term adiposity; and (4) central leptin resistance in obesity and possibly in healthy females. The new theory states that AIS in girls results from developmental disharmony expressed in spine and trunk between autonomic and somatic nervous systems. The autonomic component of this double neuro-osseous theory for AIS pathogenesis in girls involves selectively increased sensitivity of the hypothalamus to circulating leptin (genetically-determined up-regulation possibly involving inhibitory or sensitizing intracellular molecules, such as SOC3, PTP-1B and SH2B1 respectively), with asymmetry as an adverse response (hormesis); this asymmetry is routed bilaterally via the sympathetic nervous system to the growing axial skeleton where it may initiate the scoliosis deformity (leptin-hypothalamic-sympathetic nervous system concept = LHS concept). In some younger preoperative AIS girls, the hypothalamic up-regulation to circulating leptin also involves the somatotropic (growth hormone/IGF) axis which exaggerates the sympathetically-induced asymmetric skeletal effects and contributes to curve progression, a concept with therapeutic implications. In the somatic nervous system, dysfunction of a postural mechanism involving the CNS body schema fails to control, or may induce, the spinal deformity of AIS in girls (escalator concept). Biomechanical factors affecting ribs and/or vertebrae and spinal cord during growth may localize AIS to the thoracic spine and contribute to sagittal spinal shape alterations. The developmental disharmony in spine and trunk is compounded by any osteopenia, biomechanical spinal growth modulation, disc degeneration and platelet calmodulin dysfunction. Methods for testing the theory are outlined. Implications are discussed for neuroendocrine dysfunctions, osteopontin, sympathoactivation, medical therapy, Rett and Prader-Willi syndromes, infantile idiopathic scoliosis, and human evolution. AIS pathogenesis in girls is predicated on two putative normal mechanisms involved in trunk growth, each acquired in evolution and unique to humans

Effects of alcohol on skeletal response to growth hormone in hypophysectomized rats.

Chronic alcohol abuse is an established risk factor for osteoporosis. However, the precise mechanisms for the bone loss are largely unknown. Alcohol decreases skeletal expression of insulin-like growth factor-I (IGF-I), an important growth hormone (GH)-regulated skeletal growth factor. Therefore, we investigated the effects of alcohol on the skeletal response to GH in male Sprague-Dawley rats made GH-deficient by hypophysectomy (HYPOX). Four groups of sexually mature (3-month-old) rats were studied: pituitary-intact (control), HYPOX, HYPOX + GH, and HYPOX + alcohol + GH. All animals were transferred to a liquid diet 6 days following surgery. The alcohol-fed group was adapted to a graded increase in alcohol beginning 11 days following surgery. GH or vehicle was administered during the final 8 days of study and all animals were sacrificed 25 days following surgery. HYPOX resulted in cessation of body weight gain and tibial growth. Compared to controls, longitudinal bone growth and cancellous bone formation were lower following HYPOX. The latter was associated with lower mineralizing perimeter/bone perimeter. Bone marrow adiposity was higher following HYPOX. Compared to HYPOX, GH treatment increased body weight gain and bone formation rate, and decreased bone marrow adiposity. In contrast to the effects of GH treatment without alcohol, bone marrow adiposity did not differ between HYPOX and alcohol-fed GH-treated HYPOX rats. Alcohol did not alter GH-induced weight gain or increases in serum IGF-I levels, but significantly impaired the effects of GH on tibial growth and cancellous bone formation. We conclude that the detrimental skeletal effects of alcohol abuse observed in this experiment are mediated, at least in part, by skeletal resistance to GH

Effects of parathyroid hormone (1-34) on tibia in an adult rat model for chronic alcohol abuse.

Chronic alcohol abuse is a risk factor for osteoporosis in men. Human recombinant parathyroid hormone (1-34) (PTH) therapy increases bone mass in patients with osteoporosis. The purpose of the present study was to determine whether PTH is effective in increasing bone formation and bone mass in a rat model for established osteopenia caused by chronic alcohol abuse. Eight-month-old male Sprague Dawley rats were fed the Lieber-DeCarli liquid diet in which 35% of the calories were derived from either maltose-dextran or ethanol. Measurements were performed 16 weeks later to establish the magnitude of bone changes in the rats fed alcohol. High dose PTH (80 microg/kg/day) was administered 5 days/week for 6 weeks to establish the differential efficacy of hormone therapy on bone formation in alcohol consuming and alcohol withdrawn rats. The effects of alcohol and PTH on cancellous and cortical bone mass, architecture and turnover were determined by densitometry and histomorphometry. Rats fed alcohol had reduced bone mineral contents and densities, cancellous and cortical bone areas and cancellous bone formation rates compared to pair-fed controls. Following the withdrawal of alcohol, indices of bone formation increased compared to baseline values. PTH treatment increased bone mineral content and density, bone formation rates, cortical bone area, cancellous bone area and trabecular number and thickness, but several indices of bone formation were reduced in the presence of continued alcohol consumption. These results suggest that alcohol consumption, in addition to inducing bone loss, may reduce the efficacy of PTH therapy to reverse osteoporosis

Effects of low-dose parathyroid hormone on bone mass, turnover, and ectopic osteoinduction in a rat model for chronic alcohol abuse.

Parathyroid hormone (PTH) is used clinically in osteoporotic patients to increase bone mass by enhancing bone formation. PTH therapy is not uniformly effective at all skeletal sites and life-style factors may modulate the skeletal response to PTH. Alcohol may represent one of these factors. Chronic alcohol abuse is associated with osteoporosis and impaired fracture healing. Therefore, the present study investigated the effects of alcohol on the bone anabolic response to a dose of PTH similar to a human therapeutic dose 1) during normal cancellous and cortical bone growth and turnover, and 2) in a model of demineralized allogeneic bone matrix (DABM)-induced osteoinduction. Three-month-old male Sprague Dawley rats were fed a Lieber-DeCarli liquid diet with 35% of the calories derived from ethanol. The controls were pair-fed an alcohol-free isocaloric diet containing maltose-dextran. Following adaptation to the liquid diets, the rats were implanted subcutaneously with DABM cylinders prepared from cortical bone of rats fed normal chow. The rats were subsequently treated daily with PTH (1 microg/kg/d sc, 5 d/week) or vehicle and measurements on bone and DABM implants performed 6 weeks later. Total bone mass was evaluated on the day of necropsy using DXA. Tibiae were processed for histomorphometry. Bone mass and architecture in tibial diaphysis and DABM implants were evaluated by muCT. PTH treatment increased whole body bone mineral content (BMC) and bone mineral density (BMD). The hormone also increased bone formation and bone area/tissue area in the proximal tibial metaphysis. In contrast, PTH treatment had no effect on periosteal bone formation and minimal effects on DABM-induced osteoinduction. Alcohol consumption decreased whole body BMC. Alcohol also decreased cancellous as well as cortical bone formation and bone mass in tibia and impaired DABM-mediated osteoinduction. There was no interaction between PTH treatment and alcohol consumption for any of the endpoints evaluated. Our results indicate that the bone anabolic response to a therapeutic dose of PTH in the rat is largely confined to cancellous bone. In contrast, alcohol consumption inhibits bone formation at all sites. Furthermore, alcohol inhibits osteoinduction and reduces periosteal and cancellous bone formation, irrespective of therapeutic PTH administration. Based on the animal model, our findings suggest that alcohol consumption could impair the beneficial effects of PTH therapy in osteoporosis