Why fine-wire fixators work: an analysis of pressure distribution at the wire-bone interface

Tensioned fine-wire external fixator systems have been used successfully for the treatment of fractures, mal-unions and for limb lengthening for many years. When used in metaphyseal bone, this type of fixator has a lower loosening rate than half-pin fixators. The differing mechanical properties of these fixator systems have been investigated extensively; however, most studies have centered on the mechanical properties of the fixator as a whole. Our knowledge of the interactions occurring at the interface between implant and bone remains sparse. In order to investigate this interaction, we devised a simple experimental model to characterise the distribution of pressure in cancellous bone surrounding a tensioned wire under loading conditions. Comparison was then made to a similar model of a half-pin fixator. Pressure was measured using pressure sensitive film at various distances away from the implant-bone interface. Static, single cycle loading of the model was performed with a Universal Testing Machine. Pressure distribution in the fine-wire model was found to occur in three regions: polar, beam loading and uniform. Polar patterns were seen closest to the wire with pressure concentrated at the entry and exit points of the wire. Beam loading was seen at a distance of 1.5 mm from the wire and pressure reached a uniform distribution at 4.0 mm. Most of the pressure measured was less than 2 MPa, which is less than the yield strength of cancellous bone (2-7 MPa). Higher loads produced higher stresses but the distribution pattern was similar. In contrast, the half-pin model showed far higher pressures (20 MPa), which were present deeper in the bone specimen. These results further our understanding of the biomechanics of fine-wire fixators and may explain the lower loosening rate of this type of fixator when compared to half-pin fixators used in metaphyseal bone.     

The Mu class glutathione transferase is abundant in striated muscle and is an isoform-specific regulator of ryanodine receptor calcium channels

Members of the glutathione transferase (GST) structural family are novel regulators of cardiac ryanodine receptor (RyR) calcium channels. We present the first detailed report of the effect of endogenous muscle GST on skeletal and cardiac RyRs. An Mu class glutathione transferase is specifically expressed in human muscle. An hGSTM2-2-like protein was isolated from rabbit skeletal muscle and sheep heart, at concentrations of approximately 17-93 microM. When added to the cytoplasmic side of RyRs, hGSTM2-2 and GST isolated from skeletal or cardiac muscle, modified channel activity in an RyR isoform-specific manner. High activity skeletal RyR1 channels were inactivated at positive potentials or activated at negative potentials by hGSTM2-2 (8-30 microM). Inactivation became faster as the positive voltage was increased. Channels recovered from inactivation when the voltage was reversed, but recovery times were significantly slowed in the presence of hGSTM2-2 and muscle GSTs. Low activity RyR1 channels were activated at both potentials. In contrast, hGSTM2-2 and GSTs isolated from muscle (1-30 microM) in the cytoplasmic solution, caused a voltage-independent inhibition of cardiac RyR2 channels. The results suggest that the major GST isoform expressed in muscle regulates Ca2+ signalling in skeletal and cardiac muscle and conserves Ca2+ stores in the sarcoplasmic reticulum.     

Does adiposity affect muscle function during walking in children?

The biomechanical mechanisms responsible for the altered gait in obese children are not well understood, particularly as they relate to increases in adipose tissue. The purpose of this study was to test the hypotheses that as body-fat percentage (BF%) increased: (1) knee flexion during stance would decrease while pelvic obliquity would increase; (2) peak muscle forces normalized to lean-weight would increase for gluteus medius, gastrocnemius, and soleus, but decrease for the vasti; and (3) the individual muscle contributions to center of mass (COM) acceleration in the direction of their primary function(s) would not change for gluteus medius, gastrocnemius, and soleus, but decrease for the vasti. We scaled a musculoskeletal model to the anthropometrics of each participant (n=14, 8–12 years old, BF%: 16–41%) and estimated individual muscle forces and their contributions to COM acceleration. BF% was correlated with average knee flexion angle during stance (r=−0.54, p=0.024) and pelvic obliquity range of motion (r=0.78, p<0.001), as well as with relative vasti (r=−0.60, p=0.023), gluteus medius (r=0.65, p=0.012) and soleus (r=0.59, p=0.026) force production. Contributions to COM acceleration from the vasti were negatively correlated to BF% (vertical— r=−0.75, p=0.002, posterior— r=−0.68, p=0.008), but there were no correlation between BF% and COM accelerations produced by the gastrocnemius, soleus and gluteus medius. Therefore, we accept our first, partially accept our second, and accept our third hypotheses. The functional demands and relative force requirements of the hip abductors during walking in pediatric obesity may contribute to altered gait kinematics.     

Reduction of benzoquinones to hydroquinones via spontaneous reaction with glutathione and enzymatic reaction by S-glutathionyl-hydroquinone reductases

S-Glutathionyl-hydroquinone reductases (GS-HQRs) are a new class of glutathione transferases, widely present in bacteria, halobacteria, fungi, and plants. They catalyze glutathione (GSH)-dependent reduction of GS-trichloro-p-hydroquinone to trichloro-p-hydroquinone. Since GS-trichloro-p-hydroquinone is uncommon in nature, the extensive presence of GS-HQRs suggests they use common GS-hydroquinones. Here we demonstrate that several benzoquinones spontaneously reacted with GSH to form GS-hydroquinones via Michael addition, and four GS-HQRs from yeast and bacteria reduced the GS-hydroquinones to the corresponding hydroquinones. The spontaneous and enzymatic reactions led to the reduction of benzoquinones to hydroquinones with the concomitant oxidation of GSH to oxidized glutathione (GS-SG). The enzymes did not use GS-benzoquinones or other thiol-hydroquinones, for example, S-cysteinyl-hydroquinone, as substrates. Apparent kinetic parameters showed the enzymes preferred hydrophobic, bulky substrates, such as GS-menadiol. The broad substrate range and their wide distribution suggest two potential physiological roles: channeling GS-hydroquinones back to hydroquinones and reducing benzoquinones via spontaneous formation of GS-hydroquinones and then enzymatic reduction to hydroquinones. The functions are likely important in metabolic pathways with quinone intermediates.     

A novel selenium-containing glutathione transferase zeta1-1, the activity of which surpasses the level of some native glutathione peroxidases

Glutathione peroxidase (GPX) is a critical antioxidant selenoenzyme in organisms that protects cells against oxidative damage by catalyzing the reduction of hydroperoxides by glutathione (GSH). Thus, some GPX mimics have been generated because of their potential therapeutic value. The generation of a semisynthetic selenoenzyme with peroxidase activity, which matches the catalytic efficiencies of naturally evolved GPX, has been a great challenge. Previously, we semisynthesized a GPX mimetic with high catalytic efficiency using a rat theta class glutathione transferase (rGST T2-2) as a scaffold, in which the highly specific GSH-binding site is adjacent to an active site serine residue that can be chemically modified to selenocysteine (Sec). In this study, we have taken advantage of a new scaffold, hGSTZ1-1, in which there are two serine residues in the active site, to achieve both high thiol selectivity and highly catalytic efficiency. The GPX activity of Se-hGSTZ1-1 is about 1.5 times that of rabbit liver GPX, indicating that the selenium content at the active site plays an important role in enhancement of catalytic performance. Kinetic studies revealed that the catalytic mechanism of Se-hGSTZ1-1 belong in a ping-pong mechanism similar to that of the natural GPX.     

B cell overexpression of FCRL5 and PD-1 is associated with low antibody titers in HCV infection

Antibodies targeting the hepatitis C virus (HCV) envelope glycoprotein E2 are associated with delayed disease progression, and these antibodies can also facilitate spontaneous clearance of infection in some individuals. However, many infected people demonstrate low titer and delayed anti-E2 antibody responses. Since a goal of HCV vaccine development is induction of high titers of anti-E2 antibodies, it is important to define the mechanisms underlying these suboptimal antibody responses. By staining lymphocytes with a cocktail of soluble E2 (sE2) glycoproteins, we detected HCV E2-specific (sE2+) B cells directly ex vivo at multiple acute infection timepoints in 29 HCV-infected subjects with a wide range of anti-E2 IgG titers, including 17 persistently infected subjects and 12 subjects with spontaneous clearance of infection. We performed multi-dimensional flow cytometric analysis of sE2+ and E2-nonspecific (sE2-) class-switched B cells (csBC). In sE2+ csBC from both persistence and clearance subjects, frequencies of resting memory B cells (rMBC) were reduced, frequencies of activated MBC (actMBC) and tissue-like MBC (tlMBC) were increased, and expression of FCRL5, an IgG receptor, was significantly upregulated. Across all subjects, plasma anti-E2 IgG levels were positively correlated with frequencies of sE2+ rMBC and sE2+ actMBC, while anti-E2 IgG levels were negatively correlated with levels of FCRL5 expression on sE2+ rMBC and PD-1 expression on sE2+ actMBC. Upregulation of FCRL5 on sE2+ rMBC and upregulation of PD-1 on sE2+ actMBC may limit anti-E2 antibody production in vivo. Strategies that limit upregulation of these molecules could potentially generate higher titers of protective antibodies against HCV or other pathogens.     

Interactions between dihydropyridine receptors and ryanodine receptors in striated muscle

Excitation-contraction coupling in both skeletal and cardiac muscle depends on structural and functional interactions between the voltage-sensing dihydropyridine receptor L-type Ca²⁺ channels in the surface/transverse tubular membrane and ryanodine receptor Ca²⁺ release channels in the sarcoplasmic reticulum membrane. The channels are targeted to either side of a narrow junctional gap that separates the external and internal membrane systems and are arranged so that bi-directional structural and functional coupling can occur between the proteins. There is strong evidence for a physical interaction between the two types of channel protein in skeletal muscle. This evidence is derived from studies of excitation–contraction coupling in intact myocytes and from experiments in isolated systems where fragments of the dihydropyridine receptor can bind to the ryanodine receptors in sarcoplasmic reticulum vesicles or in lipid bilayers and alter channel activity. Although micro-regions that participate in the functional interactions have been identified in each protein, the role of these regions and the molecular nature of the protein–protein interaction remain unknown. The trigger for Ca²⁺ release through ryanodine receptors in cardiac muscle is a Ca²⁺ influx through the L-type Ca²⁺ channel. The Ca²⁺ entering through the surface membrane Ca²⁺ channels flows directly onto underlying ryanodine receptors and activates the channels. This was thought to be a relatively simple system compared with that in skeletal muscle. However, complexities are emerging and evidence has now been obtained for a bi-directional physical coupling between the proteins in cardiac as well as skeletal muscle. The molecular nature of this coupling remains to be elucidated.