Combined Atomic Force Microscope and Volumetric Light Sheet System for Correlative Force and Fluorescence Mechanobiology Studies.

The central goals of mechanobiology are to understand how cells generate force and how they respond to environmental mechanical stimuli. A full picture of these processes requires high-resolution, volumetric imaging with time-correlated force measurements. Here we present an instrument that combines an open-top, single-objective light sheet fluorescence microscope with an atomic force microscope (AFM), providing simultaneous volumetric imaging with high spatiotemporal resolution and high dynamic range force capability (10 pN - 100 nN). With this system we have captured lysosome trafficking, vimentin nuclear caging, and actin dynamics on the order of one second per single-cell volume. To showcase the unique advantages of combining Line Bessel light sheet imaging with AFM, we measured the forces exerted by a macrophage during FcɣR-mediated phagocytosis while performing both sequential two-color, fixed plane and volumetric imaging of F-actin. This unique instrument allows for a myriad of novel studies investigating the coupling of cellular dynamics and mechanical forces.

Search for γ-Ray Line Signals from Dark Matter Annihilations in the Inner Galactic Halo from 10 Years of Observations with H.E.S.S.

Spectral lines are among the most powerful signatures for dark matter (DM) annihilation searches in very-high-energy γ rays. The central region of the Milky Way halo is one of the most promising targets given its large amount of DM and proximity to Earth. We report on a search for a monoenergetic spectral line from self-annihilations of DM particles in the energy range from 300 GeV to 70 TeV using a two-dimensional maximum likelihood method taking advantage of both the spectral and spatial features of the signal versus background. The analysis makes use of Galactic center observations accumulated over ten years (2004-2014) with the H.E.S.S. array of ground-based Cherenkov telescopes. No significant γ-ray excess above the background is found. We derive upper limits on the annihilation cross section ⟨σv⟩ for monoenergetic DM lines at the level of 4×10^{-28} cm^{3} s^{-1} at 1 TeV, assuming an Einasto DM profile for the Milky Way halo. For a DM mass of 1 TeV, they improve over the previous ones by a factor of 6. The present constraints are the strongest obtained so far for DM particles in the mass range 300 GeV-70 TeV. Ground-based γ-ray observations have reached sufficient sensitivity to explore relevant velocity-averaged cross sections for DM annihilation into two γ-ray photons at the level expected from the thermal relic density for TeV DM particles.

H.E.S.S. Limits on Linelike Dark Matter Signatures in the 100 GeV to 2 TeV Energy Range Close to the Galactic Center.

A search for dark matter linelike signals iss performed in the vicinity of the Galactic Center by the H.E.S.S. experiment on observational data taken in 2014. An unbinned likelihood analysis iss developed to improve the sensitivity to linelike signals. The upgraded analysis along with newer data extend the energy coverage of the previous measurement down to 100 GeV. The 18 h of data collected with the H.E.S.S. array allow one to rule out at 95% C.L. the presence of a 130 GeV line (at l=-1.5°, b=0° and for a dark matter profile centered at this location) previously reported in Fermi-LAT data. This new analysis overlaps significantly in energy with previous Fermi-LAT and H.E.S.S. RESULTS: No significant excess associated with dark matter annihilations was found in the energy range of 100 GeV to 2 TeV and upper limits on the gamma-ray flux and the velocity weighted annihilation cross section are derived adopting an Einasto dark matter halo profile. Expected limits for present and future large statistics H.E.S.S. observations are also given.

Search for Dark Matter Annihilations towards the Inner Galactic Halo from 10 Years of Observations with H.E.S.S.

The inner region of the Milky Way halo harbors a large amount of dark matter (DM). Given its proximity, it is one of the most promising targets to look for DM. We report on a search for the annihilations of DM particles using γ-ray observations towards the inner 300 pc of the Milky Way, with the H.E.S.S. array of ground-based Cherenkov telescopes. The analysis is based on a 2D maximum likelihood method using Galactic Center (GC) data accumulated by H.E.S.S. over the last 10 years (2004-2014), and does not show any significant γ-ray signal above background. Assuming Einasto and Navarro-Frenk-White DM density profiles at the GC, we derive upper limits on the annihilation cross section ⟨σv⟩. These constraints are the strongest obtained so far in the TeV DM mass range and improve upon previous limits by a factor 5. For the Einasto profile, the constraints reach ⟨σv⟩ values of 6×10^{-26} cm^{3} s^{-1} in the W^{+}W^{-} channel for a DM particle mass of 1.5 TeV, and 2×10^{-26} cm^{3} s^{-1} in the τ^{+}τ^{-} channel for a 1 TeV mass. For the first time, ground-based γ-ray observations have reached sufficient sensitivity to probe ⟨σv⟩ values expected from the thermal relic density for TeV DM particles.

Atherosclerotic Carotid Plaque Composition: A 3T and 7T MRI-Histology Correlation Study.

BACKGROUND AND PURPOSE: Carotid artery atherosclerotic plaque composition may influence plaque stability and risk of thromboembolic events, and noninvasive plaque imaging may therefore permit risk stratification for clinical management. Plaque composition was compared using noninvasive in vivo (3T) and ex vivo (7T) MRI and histopathological examination. METHODS: Thirty-three endarterectomy cross-sections, from 13 patients, were studied. The data sets consisted of in vivo 3T MRI, ex vivo 7T MRI, and histopathology. Semiautomated segmentation methods were used to measure areas of different plaque components. Bland-Altman plots and mean difference with 95% confidence interval were carried out. RESULTS: There was general quantitative agreement between areas derived from semiautomated segmentation of MRI data and histology measurements. The mean differences and 95% confidence bounds in the relative to total plaque area between 3T versus Histology were: fibrous tissue 4.99%(-4.56 to 14.56), lipid-rich/necrotic core (LR/NC) with hemorrhage -1.81%(-14.11 to 10.48), LR/NC without hemorrhage -2.43%(-13.04 to 8.17), and calcification -3.18%(-11.55 to 5.18). The mean differences and 95% confidence bounds in the relative to total plaque area between 7T and histology were: fibrous tissue 3.17%(-3.17 to 9.52), LR/NC with hemorrhage -0.55%(-9.06 to 7.95), LR/NC without hemorrhage -12.62%(-19.8 to -5.45), and calcification -2.43%(-9.97 to 4.73). CONCLUSIONS: This study provides evidence that semiautomated segmentation of 3T/7T MRI techniques can help to determine atherosclerotic plaque composition. In particular, the high resolution of ex vivo 7T data was able to highlight greater detail in the atherosclerotic plaque composition. High-field MRI may therefore have advantages for in vivo carotid plaque MRI.

A three-dimensional finite element model of maximal grip loading in the human wrist.

The aim of this work was to create an anatomically accurate three-dimensional finite element model of the wrist, applying subject-specific loading and quantifying the internal load transfer through the joint during maximal grip. For three subjects, representing the anatomical variation at the wrist, loading on each digit was measured during a maximal grip strength test with simultaneous motion capture. The internal metacarpophalangeal joint load was calculated using a biomechanical model. High-resolution magnetic resonance scans were acquired to quantify bone geometry. Finite element analysis was performed, with ligaments and tendons added, to calculate the internal load distribution. It was found that for the maximal grip the thumb carried the highest load, an average of 72.2 +/- 20.1 N in the neutral position. Results from the finite element model suggested that the highest regions of stress were located at the radial aspect of the carpus. Most of the load was transmitted through the radius, 87.5 per cent, as opposed to 12.5 per cent through the ulna with the wrist in a neutral position. A fully three-dimensional finite element analysis of the wrist using subject-specific anatomy and loading conditions was performed. The study emphasizes the importance of modelling a large ensemble of subjects in order to capture the spectrum of the load transfer through the wrist due to anatomical variation.

Thyroid hormone effects on LKB1, MO25, phospho-AMPK, phospho-CREB, and PGC-1alpha in rat muscle.

Expression of all of the isoforms of the subunits of AMP-activated protein kinase (AMPK) and AMPK activity is increased in skeletal muscle of hyperthyroid rats. Activity of AMPK in skeletal muscle is regulated principally by the upstream kinase, LKB1. This experiment was designed to determine whether the increase in AMPK activity is accompanied by increased expression of the LKB1, along with binding partner proteins. LKB1, MO25, and downstream targets were determined in muscle extracts in control rats, in rats given 3 mg of thyroxine and 1 mg of triiodothyronine per kilogram chow for 4 wk, and in rats given 0.01% propylthiouracil (PTU; an inhibitor of thyroid hormone synthesis) in drinking water for 4 wk (hypothyroid group). LKB1 and MO25 increased in the soleus of thyroid hormone-treated rats vs. the controls. In other muscle types, LKB1 responses were variable, but MO25 increased in all. In soleus, MO25 mRNA increased with thyroid hormone treatment, and STRAD mRNA increased with PTU treatment. Phospho-AMPK and phospho-ACC were elevated in soleus and gastrocnemius of hyperthyroid rats. Thyroid hormone treatment also increased the amount of phospho-cAMP response element binding protein (CREB) in the soleus, heart, and red quadriceps. Four proteins having CREB response elements (CRE) in promoter regions of their genes (peroxisome proliferator-activated receptor-gamma coactivator-1alpha, uncoupling protein 3, cytochrome c, and hexokinase II) were all increased in soleus in response to thyroid hormones. These data provide evidence that thyroid hormones increase soleus muscle LKB1 and MO25 content with subsequent activation of AMPK, phosphorylation of CREB, and expression of mitochondrial protein genes having CRE in their promoters.

High-resolution 3D isotropic MR imaging of mouse flank tumours obtained in vivo with solenoid RF micro-coil.

The investigation of mouse flank tumours by magnetic resonance imaging (MRI) is limited by the achievable spatial resolution, which is generally limited by the critical problem of signal-to-noise ratio. Sensitivity was improved by using an optimized solenoid RF micro-coil, built into the animal cradle. This simple design did not require extensive RF engineering expertise to construct, yet allowed high-resolution 3D isotropic imaging at 60 x 60 x 60 microm(3) for a flank tumour in vivo, revealing the heterogeneous internal structure of the tumour. It also allowed dynamic contrast enhanced (DCE) experiments and angiography (MRA) to be performed at 100 x 100 x 100 microm(3) resolution. The DCE experiments provided an excellent example of the diffusive spreading of contrast agent into less vascularized tumour tissue. This work is the first step in using high-resolution 3D isotropic MR to study transport in mouse flank tumours.

LKB1 and the regulation of malonyl-CoA and fatty acid oxidation in muscle.

5'-AMP-activated protein kinase (AMPK), by way of its inhibition of acetyl-CoA carboxylase (ACC), plays an important role in regulating malonyl-CoA levels and the rate of fatty acid oxidation in skeletal and cardiac muscle. In these tissues, LKB1 is the major AMPK kinase and is therefore critical for AMPK activation. The purpose of this study was to determine how the lack of muscle LKB1 would affect malonyl-CoA levels and/or fatty-acid oxidation. Comparing wild-type (WT) and skeletal/cardiac muscle-specific LKB1 knockout (KO) mice, we found that the 5-aminoimidazole-4-carboxamide-1-beta-d-ribofuranoside (AICAR)-stimulated decrease in malonyl-CoA levels in WT heart and quadriceps muscles was entirely dependent on the presence of LKB1, as was the AICAR-induced increase in fatty-acid oxidation in EDL muscles in vitro, since these responses were not observed in KO mice. Likewise, the decrease in malonyl-CoA levels after muscle contraction was attenuated in KO gastrocnemius muscles, suggesting that LKB1 plays an important role in promoting the inhibition of ACC, likely by activation of AMPK. However, since ACC phosphorylation still increased and malonyl-CoA levels decreased in KO muscles (albeit not to the levels observed in WT mice), whereas AMPK phosphorylation was entirely unresponsive, LKB1/AMPK signaling cannot be considered the sole mechanism for inhibiting ACC during and after muscle activity. Regardless, our results suggest that LKB1 is an important regulator of malonyl-CoA levels and fatty acid oxidation in skeletal muscle.

Proton magnetic resonance spectroscopy of basal ganglia in chronic fatigue syndrome.

Fatigue is a common symptom of neurological diseases that affect basal ganglia function. We used proton magnetic resonance spectroscopy ((1)H MRS) to study the metabolic functions of the basal ganglia in chronic fatigue syndrome (CFS) to test the hypothesis that fatigue in CFS may have a neurogenic component. (1)H MRS of left basal ganglia was carried out in eight non-psychiatric patients with CFS and their results were compared to age- and sex-matched healthy asymptomatic healthy controls. A highly significant increase in the spectra from choline-containing compounds was seen in the CFS patient group (p < 0.001). In the absence of regional structural or inflammatory pathology, increased choline resonance in CFS may be an indicator of higher cell membrane turnover due to gliosis or altered intramembrane signalling.

The ferromagnetic pillow: a potential MR hazard not detectable by a hand-held magnet.

This paper describes an incident in which an apparently normal hospital pillow became a ferromagnetic missile when brought into the proximity of a 1.5 T MR system owing to a fine internal spring system within the pillow. Measurements revealed that the 1 kg pillow reached a maximum velocity of 33.7 km h(-1) after undergoing a maximum acceleration of 9.9g. Non-pathological cervical spines should sustain the measured forces and torques without significant injury. However, the effect could be injurious or even fatal to patients suffering from an existing cervical instability, for example due to rheumatoid arthritis. Of more general concern is the fact that the use of a powerful hand-held magnet did not reveal the presence of ferromagnetic components in this instance. Large objects containing sparsely distributed ferromagnetic materials may not be deflected by such a magnet but could still represent a hazard in the MR environment.

Method of determination of three dimensional index finger moment arms and tendon lines of action using high resolution MRI scans.

High-resolution MRI scans, in conjunction with CAD software, were used to determine the three-dimensional moment arms and force vector direction cosines for 11 structures passing the interphalangeal and metacarpophalangeal joints of the index finger. The results are presented for five different angles of joint flexion for a single subject. The moment arm data obtained differ from previous studies, where results have been derived from tendon excursion techniques or geometrical models. These dissimilarities have been accounted for by the differences in experimental techniques.

Thermal injuries associated with MRI.

Most physicians are aware of the absolute contraindications to magnetic resonance imaging (MRI). However, less familiar is the potential for an MRI-induced thermal or electrical burn associated with electrical monitoring devices. Although detailed studies concerning the burn hazard in MRI have not been reported, it is widely believed that direct electromagnetic induction in looped cables associated with the patient is responsible for the excessive heating and it is on this theory that present guidelines are based. Recent reports have however indicated that other mechanisms may cause the heating of metal, either in or on the patient. This document reviews numerous reported burn injuries sustained during MRI and addresses the underlying heating mechanisms possibly causing these events.

Investigation of the factors responsible for burns during MRI.

Numerous reported burn injuries have been sustained during clinical MRI procedures. The aim of this study was to investigate the possible factors that may be responsible for such burns. Experiments were performed to investigate three possible mechanisms for causing heating in copper wire during MRI: direct electromagnetic induction in a conductive loop, induction in a resonant conducting loop, and electric field resonant coupling with a wire (the antenna effect). Maximum recorded temperature rises were 0.6 degrees C for the loop, 61.1 degrees C for the resonant loop, and 63.5 degrees C for the resonant antenna. These experimental findings suggest that, contrary to common belief, it is unlikely that direct induction in a conductive loop will result in thermal injury. Burn incidents are more likely to occur due to the formation of resonant conducting loops and from extended wires forming resonant antenna. The characteristics of resonance should be considered when formulating safety guidelines.

Estimation of torque on mechanical heart valves due to magnetic resonance imaging including an estimation of the significance of the Lenz effect using a computational model.

Mitral and aortic valve replacement is a procedure which is common in cardiac surgery. Some of these replacement valves are mechanical and contain moving metal parts. Should the patient in whom such a valve has been implanted be involved in magnetic resonance imaging, there is a possible dangerous interaction between the moving metal parts and the static magnetic field due to the Lenz effect. Mathematical models of two relatively common forms of single-leaflet valves have been derived and the magnitude of the torque which opposes the motion of the valve leaflet has been calculated for a valve disc of solid metal. In addition, a differential model of a ring-strengthener valve type has been considered to determine the likely significance of the Lenz effect in the context of the human heart. For common magnetic field strengths at present, i.e. 1 to 2 T, the effect is not particularly significant. However, there is a marked increase in back pressure as static magnetic field strength increases. There are concerns that, since field strengths in the range 3 to 4 T are increasingly being used, the Lenz effect could become significant. At 5 to 10 T the malfunction of the mechanical heart valve could cause the heart to behave as though it is diseased. For unhealthy or old patients this could possibly prove fatal.

Potential MR hazard to patients with metallic heart valves: the Lenz effect.

The Lenz effect on the motion of metal containing moving parts of artificial heart valves has not been considered to date. In this paper the basic theory is outlined and a simple model applied to estimate worst-case forces on such valves. The results indicate a potential for forces over 100 times greater than for the more widely recognized ferromagnetic interation. Resistive pressure effects comparable to cardiac pressure differences may occur for such valves in the mitral position even at fields as low as 1.5 T. The effect will increase linearly with field strength. Additional monitoring and perhaps even exclusion of such patients may be prudent, even at 1.5 T.

Functional magnetic resonance imaging in the visual system.

Functional magnetic resonance imaging (fMRI) is a relatively new technique for measuring brain function during resting and activated conditions with good spatial and temporal resolution. Because of a robust and reproducible activation response to visual stimuli in the occipital cortex, many studies have been directed at visual function. The methodology has been refined progressively to allow more accurate detection of the small activation signal, and using computational mapping foci of cerebral activity have been displayed in a two-dimensional format. Several factors modifying the activation signal have been identified. fMRI has been used to define the retinotopic representation of areal boundaries and the localization of higher visual functions in the occipital cortex. Motion perception in area middle temporal (MT) is well-recognized, but eye movement studies are limited. The activated signal may have significant implications for our understanding of brain metabolism, but cerebral blood flow and oxygenation sensitive recordings after prolonged visual stimulation have given conflicting results. Clinically, fMRI can follow changes in cerebral activity during a progressive neurologic illness and measure responses to treatment. Neurosurgical planning in disorders such as epilepsy may be facilitated.