Understanding genetic factors in idiopathic scoliosis, a complex disease of childhood.

Idiopathic scoliosis (AIS) is the most common pediatric spinal deformity, affecting ~3% of children worldwide. AIS significantly impacts national health in the U. S. alone, creating disfigurement and disability for over 10% of patients and costing billions of dollars annually for treatment. Despite many investigations, the underlying etiology of IS is poorly understood. Twin studies and observations of familial aggregation reveal significant genetic contributions to IS. Several features of the disease including potentially strong genetic effects, the early onset of disease, and standardized diagnostic criteria make IS ideal for genomic approaches to finding risk factors. Here we comprehensively review the genetic contributions to IS and compare those findings to other well-described complex diseases such as Crohn's disease, type 1 diabetes, psoriasis, and rheumatoid arthritis. We also summarize candidate gene studies and evaluate them in the context of possible disease aetiology. Finally, we provide study designs that apply emerging genomic technologies to this disease. Existing genetic data provide testable hypotheses regarding IS etiology, and also provide proof of principle for applying high-density genome-wide methods to finding susceptibility genes and disease modifiers.

Calcium modulates the mechanical properties of anionic phospholipid membranes.

Using micropipette aspiration and fluorescence techniques, we have studied the material properties of charged lipid vesicles in calcium solutions. Vesicles were composed of phosphatidylglycerol (PG)/phosphatidylcholine (PC) or phosphatidic acid (PA)/PC mixtures. For the case of PG/PC membranes, we measure no effect of anionic lipid fraction on elasticity but a monotonic decrease up to 20% for tension required to induce membrane failure. Both of these observations are rationalized by a model we have developed to describe membrane electrostatic interactions in a two-component salt solution and the resulting changes in membrane properties. Critical tensions measured for PA/PC membranes, on the other hand, did not depend on anionic lipid fraction and were uniformly approximately 35% lower than PG/PC vesicles. This is likely due to a lateral phase separation in the membrane. By combining mechanical properties with fluorescence observations we propose that the PA-rich phase separates into small unconnected domains.

Material studies of lipid vesicles in the L(alpha) and L(alpha)-gel coexistence regimes.

In this work, we utilize micropipette aspiration and fluorescence imaging to examine the material properties of lipid vesicles made from mixtures of palmitoyloleoylphosphocholine (POPC) and dipalmitoylphosphatidylcholine (DPPC). At elevated temperatures/low DPPC fractions, these lipids are in a miscible liquid crystalline (L(alpha)) state, whereas at lower temperatures/higher DPPC fractions they phase-separate into L(alpha) and gel phases. We show that the elastic modulus, K, and critical tension, tau(c), of L(alpha) vesicles are independent of DPPC fraction. However, as the sample temperature is increased from 15 degrees C to 45 degrees C, we measure decreases in both K and tau(c) of 20% and 50%, respectively. The elasticity change is likely driven by a change in interfacial tension. We describe the reduction in critical tension using a simple model of thermally activated membrane pores. Vesicles with two-phase coexistence exhibit material properties that differ from L(alpha) vesicles including critical tensions that are 20-40% lower. Fluorescence imaging of phase coexistent POPC/DPPC vesicles shows that the DPPC-rich domains exist in an extended network structure that exhibits characteristics of a solid. This gel network explains many of the unusual material properties of two-phase membranes.

The effect of the amount of limb lengthening on skeletal muscle.

The adaptation of tibialis anterior muscles after 20% and 30% gradual limb lengthening was evaluated. Eight skeletally mature neutered male goats had 20% (n = 4) or 30% (n = 4) tibial distraction at a rate of 0.25 mm three times per day. Muscles from lengthened and contralateral control limbs were harvested on completion of distraction. Fiber length and sarcomere length were measured followed by calculation of sarcomere number and muscle fiber-to-bone lengthening ratio. Fiber length and sarcomere number after 20% and 30% limb lengthening were significantly greater in the distracted muscles, whereas no difference in sarcomere length was detected. The difference in muscle fiber length and sarcomere number between distracted and control limbs was greater in the 30% than in the 20% group. The disproportion between the amounts of muscle fiber and bone length increase was similar after 20% and 30% lengthening. The results show that muscular adaptation continues during 20% to 30% limb lengthening by increasing fiber length. It seems that this increase occurs through serial sarcomere addition rather than sarcomere length alteration. The higher rate of musclerelated clinical complications after limb lengthening beyond 20% does not seem to be related to a failure of muscle fiber contractile elements to adapt to increasing limb length.

Intramembrane electrostatic interactions destabilize lipid vesicles.

Membrane stability is of central concern in many biology and biotechnology processes. It has been suggested that intramembrane electrostatic interactions play a key role in membrane stability. However, due primarily to a lack of supporting experimental evidence, they are not commonly considered in mechanical analyses of lipid membranes. In this paper, we use the micropipette aspiration technique to characterize the elastic moduli and critical tensions of lipid vesicles with varying surface charge. Charge was induced by doping neutral phosphatidylcholine vesicles with anionic lipids phosphatidylglycerol and phosphatidic acid. Measurements were taken in potassium chloride (moderate ion-lipid binding) and tetramethylammonium chloride (low ion-lipid binding) solutions. We show that inclusion of anionic lipid does not appreciably alter the areal dilation elasticity of lipid vesicles. However, the tension required for vesicle rupture decreases with increasing anionic lipid fraction and is a function of electrolyte composition. Using vesicles with 30% charged (i.e., unbound) anionic lipid, we measured critical tension reductions of 75%, demonstrating the important role of electrostatic interactions in membrane stability.