With the vast array of genetically altered (knockout) mice becoming available there is a need for quantitative, repeatable, and efficient methodologies to characterize the phenotypic consequences of knocking out specific genes. Since knockout animals often have the ability to compensate for a single missing gene, it is important to examine the structural, material and morphological properties to obtain a thorough understanding of the changes occurring. For this project, femurs of knockout mice were first scanned using microcomputed tomography (micro-CT) to obtain high-resolution images of the trabecular bone in the distal femur, as well as cortical bone in the mid-diaphysis. After scanning, the femurs were tested to destruction in four-point bending at the mid-diaphysis about the medial lateral axis of the femur. These methodologies allowed quantification of (1) morphologic properties such as bone volume fraction, trabecular properties and 2nd moment of the area (2) structural properties such as stiffness, maximum load at failure, and post yield deformation and (3) material properties such as bone mineral density, elastic modulus and yield strength. As part of two independent studies, two different knockout mice, cyclooxygenase-2 (COX-2 -/-) and Apolipoprotein E (APOE -/-), were examined for structure-function relationships using these methodologies. COX-2 knockout mice were found to have decreased mineral content in their femurs, and increased post yield deformation. APOE knockout mice at 10 weeks of age had decreased bone mass and structural properties. However, by 40 weeks of age APOE deficient mice caught up to and exceeded the structural properties and bone mass of their wild type counterparts.

No abstract available. / Thesis (M.Med.Sc.)-University of KwaZulu-Natal, Durban, 2005.

Ratas adultas foram tratadas com 2.5, 5, 10, 20 e 40x a dose fisiológica de triiodotironina (T3) por 10 semanas. Houve redução, dose-dependente, da massa óssea, volume e espessura trabecular (Tb.Th), e espessura de osso cortical. Até a dose de 10xT3, houve aumento na taxa de formação óssea (BFR). Doses maiores de T3 reduziram a BFR, e a reabsorção óssea. Esses dados mostram que, até um certo grau de tireotoxicose, a osteopenia é resultado de aumento da formação e reabsorção ósseas, com predomínio da última. Em graus mais elevados, a osteopenia é causada por redução no remodelamento ósseo, com predomínio da atividade reabsortiva. Em seguida, comparamos os efeitos do T3 e GC-1, um tireomimético seletivo pelo receptor b de T3 (TRb), no osso de camundongos fêmeas adultas. Na tíbia, o T3 reduziu a resistência, rigidez e resiliência, enquanto o GC-1 aumentou esses parâmetros e a Tb.Th, o que mostra que o GC-1 melhora a qualidade óssea. Considerando-se a seletividade do GC-1 pelo TRb, esses achados sugerem que o TRb medeia predominantemente ações positivas do T3 no osso. / Adult rats were treated with 2.5, 5, 10, 20 and 40x the physiological dose of triiodothyronine (T3) for 10 weeks. There was a dose-dependent reduction in bone mass, trabecular volume and thickness (Tb.Th), and cortical thickness. Up to 10xT3, there was an increase in the bone formation rate (BFR). Higher doses of T3 decreased BFR and bone resorption. These data show that, up to a certain degree of thyrotoxicosis, osteopenia is the result of an increase in bone formation and resorption, with a prevalence of resorption. In more severe thyrotoxicosis, the osteopenia is caused by a reduction in bone remodeling, with a predominance of bone resorption. Then, we compared the effects of T3 and GC-1, a thyromimetic that is selective for T3 receptor b (TRb), on bone of adult female mice. T3 treatment decreased tibial resistance, stiffness and resilience, while GC-1 increase these parameters and Tb.Th, showing that GC-1 improves bone quality. Considering the selectiveness of GC-1 for TRb, these findings suggest that TRb mediates mainly positive actions of T3 in the bone.

Indiana University-Purdue University Indianapolis (IUPUI) / Mechanical loading associated with exercise is known to benefit bone health; however, most studies explore exercise benefits on bone mass independent of bone structure and strength. The purpose of this dissertation is to explore the response of the skeleton to exercise across the translational divide between animal- and human-based studies, with a particular emphasis on exercise-induced changes in bone structure and estimated strength. To explore the skeletal benefits of exercise, models were used wherein loading is introduced unilaterally to one extremity. Unilateral exercise enables the contralateral, non-exercised extremity to be used as an internal control site for the influences of systemic factors, such as genetics and circulating hormones. In study 1, a dose response between load magnitude and tibial midshaft cortical bone adaptation was observed in mice that had their right tibia loaded in axial compression at one of three load magnitudes for 3 d/wk over 4 weeks. In study 2, the ability of peripheral quantitative computed tomography to provide very good prediction of midshaft humerus mechanical properties with good short-term precision in human subjects was demonstrated. In study 3, collegiate-level jumping (long and/or high jump) athletes were shown to have larger side-to-side differences in tibial midshaft structure and estimated strength between their jump and lead legs than observed in non-jumping athletes. In study 4, prepubertal baseball players followed for 12 months were shown to gain more bone mass, structure and estimated strength in their throwing arm relative to their nonthrowing arm over the course of 12 months. These cumulative data using a combination of experimental models ranging from animal to cross-sectional and longitudinal human models demonstrate the ability of the skeleton to adapt its structure and estimated strength to the mechanical loading associated with exercise. Study of these models in future work may aid in optimizing skeletal responses to exercise.

Bone drilling is a major part of orthopaedic surgery performed during the internal fixation of fractured bones. At present, information related to drilling force, drilling torque, rate of drill bit penetration and drill bit rotational speed is not available to orthopaedic surgeons, clinicians and researchers as bone drilling is performed manually. This research demonstrates that bone drilling force data if recorded in-vivo, during the repair of bone fractures, can provide information about the strength/quality of the bone. Drilling force does not give a direct measure of bone strength; therefore it has been correlated with the shear strength and screw pullout strength to determine the efficacy in estimating the bone strength. Various synthetic bone material densities and animal bones have been tested to demonstrate the use of drilling force data. A novel automated experimental test rig, which enables drilling tests, screw insertion and screw pullout tests to be carried out in a controlled environment, has been developed. Both drilling and screw pullout tests have been carried out in a single setting of the specimen to reduce the experimental errors and increase repeatability of the results. A significantly high value of correlation (r² > 0.99) between drilling force & shear strength and also between drilling force & normalised screw pullout strength in synthetic bone material was found. Furthermore, a high value of correlation (r² = 0.958 for pig bones and r² = 0.901 for lamb bones) between maximum drilling force & normalised screw pullout strength was also found. The result shows that drilling data can be used to predict material strength. Bone screws are extensively used during the internal fixation of fractured bones. The amount of screw been tightened is one of the main factor which affects the bone-screw fixation quality. Over tightening of screw can result into the loss of bone-screw fixation strength, whereas under tightening can result in the screw loosening. Therefore, optimum tightening of the screw is important to achieve the maximum bone-screw fixation strength. At present, optimum tightening of the screw is entirely dependent upon the skill and judgment of the surgeon, which is predominantly based on the feel of the screw tightening torque. Various studies have been reported in the literature to develop an algorithm to set an optimum tightening torque value to be used in surgery. A method which is based on the use of rotation angle of the screw while tightening, rather than using screw insertion/tightening torque, to optimise the bone-screw fixation strength is proposed in this research. The effectiveness of the proposed method has been successfully demonstrated on the synthetic bone material using the designed test rig. The optimum angle for the tested screw was found to be 120° which is equivalent to 33% of the screw pitch.

Indiana University-Purdue University Indianapolis (IUPUI) / Down syndrome (DS), a genetic disorder that affects ~1 in 700 live births, is caused by trisomy of human chromosome 21 (Hsa21). Individuals with DS are affected by a wide spectrum of phenotypes which vary in severity and penetrance. However, cognitive and skeletal impairments can be commonly observed in all individuals with DS. To study these phenotypes, we utilized the Ts65Dn mouse model that carries three copies of approximately half the gene orthologs found on Hsa21 and exhibit similar phenotypes as observed in humans with DS. Individuals with DS and Ts65Dn mice have deficits in bone mineral density (BMD), bone architecture, bone strength, learning and memory. Over-expression of DYRK1A, a serine-threonine kinase encoded on Hsa21, has been linked to deficiencies in DS bone homeostasis and cognition. Epigallocatechin-3-gallate (EGCG), an aromatic polyphenol found in high concentrations in green tea, is a selective inhibitor of DYRK1A activity. Normalization of DYRK1A activity by EGCG therefore may have the potential to ameliorate skeletal and cognitive deficits. We hypothesized that supplements containing EGCG obtained from health food stores/ online vendors will not be as effective as EGCG from a chemical company in correcting bone deficits associated with DS. Our results suggest that EGCG improves the bone mineral density of trisomic femurs significantly better than the supplements while the EGCgNOW supplement from NOW FOODS improves trabecular and cortical bone structure. The results from HPLC analysis of supplements showed the presence of other catechins in EGCgNOW and degradation analysis revealed the rapid degradation of supplements. Therefore we hypothesize that the presence of EGCG degradation products and other green tea catechins in supplements may play a role in the differential skeletal effects we observed. We further hypothesized that a three week treatment of adolescent mice with EGCG will lead to an improvement in the learning and memory deficits that are observed in trisomic animals in comparison to control mice. However, our results indicate that three weeks of low-dose EGCG treatment during adolescence is insufficient to improve hippocampal dependent learning and memory deficits of Ts65Dn mice. The possibility remains that a higher dose of EGCG that begins at three weeks but lasts throughout the behavioral test period may result in improvement in learning and memory deficit of Ts65Dn mice.

Indiana University-Purdue University Indianapolis (IUPUI) / Signal Transducer and Activator of Transcription 3 (STAT3) is known to be related to bone metabolism. Mutation of STAT3 causes a rare disorder in which serum levels of IgE are elevated. This causes various skeletal problems similar to osteoporosis. To examine the effect of STAT3 in the osteoclast, we obtained two osteoclast specific STAT3 knockout mouse models: one using the CTSK promoter to drive Cre recombinase and another using a TRAP promoter. Examination of these mice at 8 weeks of age revealed a decreased trabecular bone volume in CTSK specific STAT3 knockout mice along with a slight decrease in osteoclast number in both CTSK and TRAP specific STAT3 knockout females. We also noticed changes in bone mineral density and bone mechanical strength in females. These data suggest that STAT3 plays a part in the function of the osteoclast.