Conformational behavior of chondroitin and chondroitin sulfate in relation to their physical properties as inferred by molecular modeling

Chondroitin and chondroitin sulfates belong to the family of glycosaminoglycans. They are most widely distributed in animal tissues, where they are involved in structural functions and in cell-cell communication. Their basic structures consist of a disaccharidic repeating unit of beta-D-glucuronic acid (GlcA) and 2-acetamido-2-deoxy-beta-D-galactose (GalNAc), this latter being sulfated at different positions. Molecular mechanics has been applied to calculate the adiabatic energy maps for each of the constituting disaccharides of chondroitin, chondroitin 4-sulfate, and chondroitin 6-sulfate using the MM3 force field. Based on these maps, higher levels of structural organization have been simulated. On one hand, the disordered state is studied through a Metropolis-based algorithm; the resulting chains present a behavior of semirigid polymers, with an order of stiffness: chondroitin 4-sulfate > chondroitin > chondroitin 6-sulfate. On the other hand, the exploration of the stable ordered forms leads to numerous helical conformations of comparable energies. Several of these conformations correspond to the experimentally observed ones. The ability of coordination with cations has also been explored, resulting in a preferential stereospecificity for calcium ions when compared to sodium ions.     

Tachykinin NK-1 receptor probed with constrained analogues of substance P

The action of rotameric probes introduced either in position 7 or 8 in the sequence of substance P (SP) was investigated, i.e. -tetrahydroisoquinoleic acid (Tic), -fluorenylglycine (Flg), -diphenylalanine (Dip), the diastereoisomers of -1-indanylglycine (Ing) and -benz[ƒ]indanylglycine (Bfi), the Z- and E-isomers of dehydrophenylalanine and dehydronaphthylalanine (Δ^ZPhe, Δ^EPhe, Δ^ZNal, Δ^ENal) and (Dmp). The aim of this study was the topographical characterization of the binding subsites of human NK-1 receptor expressed in CHO cells, especially the S₇ and S₈ subsites, corresponding to residues Phe⁷ and Phe⁸ of substance P. According to the binding potencies of these substituted-SP analogues, the S₇ binding subsite is smaller than the S₈ subsite: the S₇ subsite accepts only one aromatic nucleus, while the S₈ can accommodate three coplanar nuclei altogether. These findings are compatible with the idea that the S₈ binding subsite may reside in the extracellular loops of the hNK-1 receptor. NK-1 agonists bind to human NK-1 receptor and activate the production of both inositol phosphates and cyclic AMP. As already quoted for septide, [pGlu⁶, Pro⁹]SP(6–11), discrepancies are observed between affinity (K_i) and activity (EC₅₀) values for IPs production. While a weak correlation between K_i and EC₅₀ values for IPs production could be found (r = 0.70), an excellent correlation could be demonstrated between their affinities (K_i) and their potencies (EC₅₀) for cAMP production (r = 0.97). The high potency (EC₅₀) observed for ‘septide-like’ molecules on PI hydrolysis, compared to their affinity is not an artefact related to the high level of NK-1 receptors expressed on CHO cells since a good correlation was found between EC₅₀ values obtained for PI hydrolysis and those measured for spasmogenic activity in guinea pig ileum bioassay (r = 0.94).

According to the binding potencies of constrained analogues of phenylalanine, the S₇ binding subsite of human NK-1 receptor is small, whereas the S₈, which can accommodate three coplanar nuclei, might probably reside in the extracellular loop. The discrepancies observed between affinity (K_i) and activity (EC₅₀) values for IPs production are not an artefact of CHO cells since a good correlation was found between EC₅₀ for PI hydrolysis and those measured in guinea pig ileum bioassay.     

Estimation of spinal joint centers from external back profile and anatomical landmarks

Defining a subject-specific model of the human body is required for motion analysis in many fields, such as in ergonomics and clinical applications. However, locating internal joint centers from external characteristics of the body still remains a challenging issue, in particular for the spine. Current methods mostly require a set of rarely accessible (3D back or trunk surface) or operator dependent inputs (large number of palpated landmarks and landmarks-based anthropometrics). Therefore, there is a need to provide an alternative way to estimate joint centers only using a limited number of easily palpable landmarks and the external back profile. Two methods were proposed to predict the spinal joint centers: one using only 6 anatomical landmarks (ALs) (2 PSIS, T8, C7, IJ and PX) and one using both 6 ALs and the external back profile. Regressions were established using the X-ray based 3D reconstructions of 80 subjects and evaluated on 13 additional subjects of variable anthropometry. The predicted location of joint centers showed an average error 9.7 mm (±5.0) in the sagittal plane for all joints when using the external back profile. Similar results were obtained without using the external back profile, 9.5 mm (±5.0). Compared to other existing methods, the proposed methods offered a more accurate prediction with a smaller number of palpated points. Additional methods have to be developed for considering postures other than standing, such as a sitting position.     

Continuous hypergravity alters the cytoplasmic elasticity of MC3T3-E1 osteoblasts via actin filaments

Osteoblasts are sensitive to altered gravity conditions, displaying changes in RNA and protein expression, proliferation, and differentiation; however, the effect of hypergravity on the mechanical properties of osteoblasts remains unclear. In this study, atomic force microscopy (AFM) was used to evaluate the effect of hypergravity on the elasticity of osteoblasts. We demonstrate that a continuous hypergravitational environment increased the elasticity of the cytoplasm, but not the nuclei zone, of MC3T3-E1 osteoblasts. Actin filaments, but not microtubules, dominated in the increased elasticity. These findings provide new insights on cellular gravity-sensing mechanisms.     

Virtual screening using molecular simulations

Effective virtual screening relies on our ability to make accurate prediction of protein-ligand binding, which remains a great challenge. In this work, utilizing the molecular-mechanics Poisson-Boltzmann (or Generalized Born) surface area approach, we have evaluated the binding affinity of a set of 156 ligands to seven families of proteins, trypsin β, thrombin α, cyclin-dependent kinase (CDK), cAMP-dependent kinase (PKA), urokinase-type plasminogen activator, β-glucosidase A, and coagulation factor Xa. The effect of protein dielectric constant in the implicit-solvent model on the binding free energy calculation is shown to be important. The statistical correlations between the binding energy calculated from the implicit-solvent approach and experimental free energy are in the range of 0.56-0.79 across all the families. This performance is better than that of typical docking programs especially given that the latter is directly trained using known binding data whereas the molecular mechanics is based on general physical parameters. Estimation of entropic contribution remains the barrier to accurate free energy calculation. We show that the traditional rigid rotor harmonic oscillator approximation is unable to improve the binding free energy prediction. Inclusion of conformational restriction seems to be promising but requires further investigation. On the other hand, our preliminary study suggests that implicit-solvent based alchemical perturbation, which offers explicit sampling of configuration entropy, can be a viable approach to significantly improve the prediction of binding free energy. Overall, the molecular mechanics approach has the potential for medium to high-throughput computational drug discovery.     

Radiation pressure on a biconcave human Red Blood Cell and the resulting deformation in a pair of parallel optical traps

We calculated the three‐dimensional optical stress distribution and the resulting deformation on a biconcave human red blood cell (RBC) in a pair of parallel optical trap. We assumed a Gaussian intensity distribution with a spherical wavefront for each trapping beam and calculated the optical stress from the momentum transfer associated with the reflection and refraction of the incident photons at each interface. The RBC was modelled as a biconcave thin elastic membrane with uniform elasticity and a uniform thickness of 0.25 μm. The resulting cell deformation was determined from the optical stress distribution by finite element software, Comsol Structure Mechanics Module, with Young's modulus (E) as a fitting parameter in order to fit the theoretical results for cell elongation to our experimental data. (© 2014 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)     

Characteristics of dynamic postural reactions in the locust hindleg

Summary The use of the locust (Schistocerca americana) hindleg in postural control was examined in animals that stood on a repeatedly swayed vertical substrate. Myograms were recorded from leg muscles and the angle of the femoro-tibial joint was monitored photographically. Two discrete strategies were observed,; in compensatory reactions the hindleg was held in place, while in flexion reactions, the leg was moved, most often to complete flexion of the femoro-tibial joint. Tightly coupled, rhythmic bursting occurred in the flexor and levator muscles of the leg during compensatory reactions. Bursting was initiated repeatedly when the substrate was being pulled away from the animal. Bursting was correlated with subsequent decreases in the rate of change of the femorotibial joint angle. Compensatory and flexion reactions occurred preferentially in different ranges of joint angles: most often, compensatory reactions occurred in the midrange, while flexion reactions were elicited in the extremes of joint angle. These differences may be due to the mechanical advantages of the tibial muscles and the leg may be moved to full flexion because of a locking mechanism of the flexor muscle tendon. These reactions are compared with known reflexes of hindleg proprioceptors and contrasted with similar responses of vertebrates.     

Biomechanical evaluation of walking and cycling in children

Physical activity in children is important as it leads to healthy growth due to physiological benefits. However, a physiological benefit can be partially negated by excessive or unphysiological loads within the joints. To gain an initial understanding into this, the present study sought to compare joint loading between walking and cycling in children. With institutional ethical approval, 14 pre-pubertal children aged 8–12 walked on an instrumented treadmill and cycled on a stationary ergometer. Two methods were used to match physiological load. Cardiovascular loads between walking and cycling were matched using heart rate. Metabolic load was normalised by matching estimates of oxygen consumption. Joint reaction forces during cycling and walking as well as joint moments were derived using inverse dynamics. Peak compressive forces were greater on the knees and ankles during walking than during cycling. Peak shear peak forces at the knee and ankle were also significantly larger during walking than during cycling, independent of how physiological load was normalised. For both cycling conditions, ankle moments were significantly smaller during cycling than walking. No differences were found for knee moments. At equivalent physiological intensities, cycling results in less joint loading than walking. It can be speculated that for certain populations and under certain conditions cycling might be a more suitable mode of exercise than weight bearing activities to achieve a given metabolic load.     

Role of diastolic vortices in flow and energy dynamics during systolic ejection

MRI-based computational fluid dynamics simulations were performed in the left ventricles of two adult porcine subjects with varying physiological states (before and after an induced infarction). The hypothesis that diastolic vortices store kinetic energy and assist systolic ejection was tested, by performing systolic simulations in the presence and absence of diastolic vortices. The latter was achieved by reinitializing the entire velocity field to be zero at the beginning of systole. A rudimentary prescribed motion model of a mitral valve was included in the simulations to direct the incoming mitral jet towards the apex. Results showed that the presence or absence of diastolic vortex rings had insignificant impact on the energy expended by walls of the left ventricles for systolic ejection for both the porcine subjects, under all physiological conditions. Although substantial kinetic energy was stored in diastolic vortices by end diastole, it provided no appreciable savings during systolic ejection, and most likely continued to complete dissipation during systole. The role of diastolic vortices in apical washout was investigated by studying the cumulative mass fraction of passive dye that was ejected during systole in the presence and absence of vortices. Results indicated that the diastolic vortices play a crucial role in ensuring efficient washout of apical blood during systolic ejection.