Shape-induced force fields in optical trapping

Advances in optical tweezers, coupled with the proliferation of two-photon polymerization systems, mean that it is now becoming routine to fabricate and trap non-spherical particles. The shaping of both light beams and particles allows fine control over the flow of momentum from the optical to mechanical regimes. However, understanding and predicting the behaviour of such systems is highly complex in comparison with the traditional optically trapped microsphere. In this Article, we present a conceptually new and simple approach based on the nature of the optical force density. We illustrate the method through the design and fabrication of a shaped particle capable of acting as a passive force clamp, and we demonstrate its use as an optically trapped probe for imaging surface topography. Further applications of the design rules highlighted here may lead to new sensors for probing biomolecule mechanics, as well as to the development of optically actuated micromachines

Evaluating the impact of trigeminal neuralgia

Patients with idiopathic trigeminal neuralgia (TN) were categorised into 3 subtypes (n 5 225). Group 1 (n 5 155, 68.9%) had TN without concomitant pain, group 2 (n532, 14.2%) had TN with intermittent concomitant pain, and group 3 (n539, 16.9%) had TN with autonomic symptoms. We tested 2 hypotheses: (1) that different pain profiles would be associated with the different groups; (2) that the severe pain associated with TN would impact negatively on activities of daily living and thereby result in disability as defined by the World Health Organisation. A different pain profile was found across the groups. We obtained unequivocal evidence that TN causes disability with up to 45% of patients being absent from usual daily activities 15 days or more in the past 6 months. On the Hospital Anxiety and Depression Scale, 35.7% patients had mild-to-severe depression and over 50% were anxious. The Pain Catastrophizing Scale showed that 78% of patients had considerable negative thoughts with scores.20 and a mean score of 36.4. Prior to referral, only 54% had been prescribed carbamazepine while opioids had been prescribed in 14.6% of the patients. Prior to referral, over 80% had already been to 1 specialist centre which had not provided appropriate management. Patients with TN report varied characteristics but all result in some degree of psychosocial disability especially before adequate therapy is attained

A comparison of liquid and solid culture for determining relapse and durable cure in phase III TB trials for new regimens

BACKGROUND: Tuberculosis kills more people than any other infectious disease, and new regimens are essential. The primary endpoint for confirmatory phase III trials for new regimens is a composite outcome that includes bacteriological treatment failure and relapse. Culture methodology is critical to the primary trial outcome. Patients in clinical trials can have positive cultures after treatment ends that may not necessarily indicate relapse, which was ascribed previously to laboratory cross-contamination or breakdown of old lesions. Löwenstein-Jensen (LJ) medium was the previous standard in clinical trials, but almost all current and future trials will use the Mycobacteria Growth Indicator Tube (MGIT) system due to its simplicity and consistency of use, which will affect phase III trial results. LJ was used for the definition of the primary endpoint in the REMoxTB trial, but every culture was also inoculated in parallel into the MGIT system. The data from this trial, therefore, provide a unique opportunity to investigate and compare the incidence of false 'isolated positives' in liquid and solid media and their potential impact on the primary efficacy results. METHODS: All post-treatment positive cultures were reviewed in the REMoxTB clinical trial. Logistic regression models were used to model the incidence of isolated positive cultures on MGIT and LJ. RESULTS: A total of 12,209 sputum samples were available from 1652 patients; cultures were more often positive on MGIT than LJ. In 1322 patients with a favourable trial outcome, 126 (9.5%) had cultures that were positive in MGIT compared to 34 (2.6%) patients with positive cultures on LJ. Among patients with a favourable outcome, the incidence of isolated positives on MGIT differed by study laboratory (p < 0.0001) with 21.9% of these coming from one laboratory investigating only 4.9% of patients. No other baseline factors predicted isolated positives on MGIT after adjusting for laboratory. There was evidence of clustering of isolated positive cultures in some patients even after adjusting for laboratory, p < 0.0001. The incidence of isolated positives on MGIT did not differ by treatment arm (p = 0.845, unadjusted). Compared to negative MGIT cultures, positive MGIT cultures were more likely to be associated with higher grade TB symptoms reported within 7 days either side of sputum collection in patients with an unfavourable primary outcome (p < 0.0001) but not in patients with a favourable outcome (p = 0.481). CONCLUSIONS: Laboratory cross-contamination was a likely cause of isolated positive MGIT cultures which were clustered in some laboratories. Certain patients had repeated positive MGIT cultures that did not meet the definition of a relapse. This pattern was too common to be explained by cross-contamination only, suggesting that host factors were also responsible. We conclude that MGIT can replace LJ in phase III TB trials, but there are implications for the definition of the primary outcome and patient management in trials in such settings. Most importantly, the methodologies differ in the incidence of isolated positives and in their capacity for capturing non-tuberculosis mycobacteria. It emphasises the importance of effective medical monitoring after treatment ends and consideration of clinical signs and symptoms for determining treatment failure and relapse

Subunit positioning and stator filament stiffness in regulation and power transmission in the V1 motor of the manduca sexta V-ATpase

The vacuolar H(+)-ATPase (V-ATPase) is an ATP-driven proton pump essential to the function of eukaryotic cells. Its cytoplasmic V1 domain is an ATPase, normally coupled to membrane-bound proton pump Vo via a rotary mechanism. How these asymmetric motors are coupled remains poorly understood. Low energy status can trigger release of V1 from the membrane and curtail ATP hydrolysis. To investigate the molecular basis for these processes, we have carried out cryo-electron microscopy three-dimensional reconstruction of deactivated V1 from Manduca sexta. In the resulting model, three peripheral stalks that are parts of the mechanical stator of the V-ATPase are clearly resolved as unsupported filaments in the same conformations as in the holoenzyme. They are likely therefore to have inherent stiffness consistent with a role as flexible rods in buffering elastic power transmission between the domains of the V-ATPase. Inactivated V1 adopted a homogeneous resting state with one open active site adjacent to the stator filament normally linked to the H subunit. Although present at 1:1 stoichiometry with V1, both recombinant subunit C reconstituted with V1 and its endogenous subunit H were poorly resolved in three-dimensional reconstructions, suggesting structural heterogeneity in the region at the base of V1 that could indicate positional variability. If the position of H can vary, existing mechanistic models of deactivation in which it binds to and locks the axle of the V-ATPase rotary motor would need to be re-evaluated

Cognitive and sensorimotor function in participants being treated for trigeminal neuralgia pain

Background Trigeminal neuralgia (TN) is an orofacial condition defined by reoccurring, spontaneous, short-lived but excruciating stabbing pain. Pharmacological interventions constitute the first-line treatment for TN, with antiepileptic drugs commonly prescribed. People treated for TN pain with antiepileptic drugs describe cognitive and motor difficulties affecting activities of daily living, and report poorer quality of life. We undertook the first comprehensive objective evaluation of sensorimotor and cognitive performance in participants being treated for TN pain with antiepileptic drugs relative to age-matched controls. Methods Participants (43 TN, 41 control) completed a battery of sensorimotor (steering, aiming and tracking) and cognitive (working memory, processing speed, inhibition) tasks. Results The TN group performed significantly worse than controls on the sensorimotor tracking and aiming tasks and across all cognitive measures. Conclusions The data explain why patients treated with antiepileptic drugs report impairment when conducting activities of daily living (given the need for cognitive and motor capability within most of these). The study is an important first step in: (i) ensuring there is adequate information on the impact of pharmacological treatment; (ii) identifying measures to determine optimal medication dosage and track change over time; (iii) creating an evidence base that could allow scientific justification of alternative pain treatment options for TN (e.g. the costs/benefits of surgery)

Entropy Production in an Elementary, Light Driven Micro-Machine

This is the final version. Available on open access from Frontiers media via the DOI in this recordData Availability Statement: The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.We consider the basic, thermodynamic properties of an elementary micro-machine operating at colloidal length scales. In particular, we track and analyze the driven stochastic motion of a carefully designed micro-propeller rotating unevenly in an optical tweezers, in water. In this intermediate regime, the second law of macroscopic thermodynamics is satisfied only as an ensemble average, and individual trajectories can be temporarily associated with decreases in entropy. We show that our light driven micro-propeller satisfies an appropriate fluctuation theorem that constrains the probability with which these apparent violations of the second law occur. Implications for the development of more complex micro-machines are discussed.Czech Science FoundationEuropean Regional Development Fund (ERDF)Engineering and Physical Sciences Research Council (EPSRC)Royal Academy of Engineering (RAE

Entropic Tension in Crowded Membranes

Unlike their model membrane counterparts, biological membranes are richly decorated with a heterogeneous assembly of membrane proteins. These proteins are so tightly packed that their excluded area interactions can alter the free energy landscape controlling the conformational transitions suffered by such proteins. For membrane channels, this effect can alter the critical membrane tension at which they undergo a transition from a closed to an open state, and therefore influence protein function \emph{in vivo}. Despite their obvious importance, crowding phenomena in membranes are much less well studied than in the cytoplasm. Using statistical mechanics results for hard disk liquids, we show that crowding induces an entropic tension in the membrane, which influences transitions that alter the projected area and circumference of a membrane protein. As a specific case study in this effect, we consider the impact of crowding on the gating properties of bacterial mechanosensitive membrane channels, which are thought to confer osmoprotection when these cells are subjected to osmotic shock. We find that crowding can alter the gating energies by more than $2\;k_BT$ in physiological conditions, a substantial fraction of the total gating energies in some cases. Given the ubiquity of membrane crowding, the nonspecific nature of excluded volume interactions, and the fact that the function of many membrane proteins involve significant conformational changes, this specific case study highlights a general aspect in the function of membrane proteins.Comment: 20 pages (inclduing supporting information), 4 figures, to appear in PLoS Comp. Bio

Comparison of DC Bead-irinotecan and DC Bead-topotecan drug eluting beads for use in locoregional drug delivery to treat pancreatic cancer

DC Bead is a drug delivery embolisation system that can be loaded with doxorubicin or irinotecan for the treatment of a variety of liver cancers. In this study we demonstrate that the topoisomerase I inhibitor topotecan hydrochloride can be successfully loaded into the DC Bead sulfonate-modified polyvinyl alcohol hydrogel matrix, resulting in a sustained-release drug eluting bead (DEBTOP) useful for therapeutic purposes. The in vitro drug loading capacity, elution characteristics and the effects on mechanical properties of the beads are described with reference to our previous work with irinotecan hydrochloride (DEBIRI). Results showed that drug loading was faster when the solution was agitated compared to static loading and a maximum loading of ca. 40–45 mg topotecan in 1 ml hydrated beads was achievable. Loading the drug into the beads altered the size, compressibility moduli and colour of the bead. Elution was shown to be reliant on the presence of ions to perform the necessary exchange with the electrostatically bound topotecan molecules. Topotecan was shown by MTS assay to have an IC50 for human pancreatic adenocarcinoma cells (PSN-1) of 0.22 and 0.27 lM compared to 28.1 and 19.2 lM for irinotecan at 48 and 72 h, respectively. The cytotoxic efficacy of DEBTOP on PSN-1 was compared to DEBIRI. DEPTOP loaded at 6 & 30 mg ml-1, like its free drug form, was shown to be more potent than DEBIRI of comparable doses at 24, 48 & 72 h using a slightly modified MTS assay. Using a PSN-1 mouse xenograft model, DEBIRI doses of 3.3–6.6 mg were shown to be well tolerated (even with repeat administration) and effective in reducing the tumour size. DEBTOP however, was lethal after 6 days at doses of 0.83–1.2 mg but demonstrated reasonable efficacy and tolerability (again with repeat injection possible) at 0.2–0.4 mg doses. Care must therefore be taken when selecting the dose of topotecan to be loaded into DC Bead given its greater potency and potential toxicity

Spot sputum samples are at least as good as early morning samples for identifying Mycobacterium tuberculosis

Background The use of early morning sputum samples (EMS) to diagnose tuberculosis (TB) can result in treatment delay given the need for the patient to return to the clinic with the EMS, increasing the chance of patients being lost during their diagnostic workup. However, there is little evidence to support the superiority of EMS over spot sputum samples. In this new analysis of the REMoxTB study, we compare the diagnostic accuracy of EMS with spot samples for identifying Mycobacterium tuberculosis pre- and post-treatment. Methods Patients who were smear positive at screening were enrolled into the study. Paired sputum samples (one EMS and one spot) were collected at each trial visit pre- and post-treatment. Microscopy and culture on solid LJ and liquid MGIT media were performed on all samples; those missing corresponding paired results were excluded from the analyses. Results Data from 1115 pre- and 2995 post-treatment paired samples from 1931 patients enrolled in the REMoxTB study were analysed. Patients were recruited from South Africa (47%), East Africa (21%), India (20%), Asia (11%), and North America (1%); 70% were male, median age 31 years (IQR 24–41), 139 (7%) co-infected with HIV with a median CD4 cell count of 399 cells/μL (IQR 318–535). Pre-treatment spot samples had a higher yield of positive Ziehl–Neelsen smears (98% vs. 97%, P = 0.02) and LJ cultures (87% vs. 82%, P = 0.006) than EMS, but there was no difference for positivity by MGIT (93% vs. 95%, P = 0.18). Contaminated and false-positive MGIT were found more often with EMS rather than spot samples. Surprisingly, pre-treatment EMS had a higher smear grading and shorter time-to-positivity, by 1 day, than spot samples in MGIT culture (4.5 vs. 5.5 days, P < 0.001). There were no differences in time to positivity in pre-treatment LJ culture, or in post-treatment MGIT or LJ cultures. Comparing EMS and spot samples in those with unfavourable outcomes, there were no differences in smear or culture results, and positive results were not detected earlier in Kaplan–Meier analyses in either EMS or spot samples. Conclusions Our data do not support the hypothesis that EMS samples are superior to spot sputum samples in a clinical trial of patients with smear positive pulmonary TB. Observed small differences in mycobacterial burden are of uncertain significance and EMS samples do not detect post-treatment positives any sooner than spot samples

Nodal quasiparticle meltdown in ultra-high resolution pump-probe angle-resolved photoemission

High-$T_c$ cuprate superconductors are characterized by a strong momentum-dependent anisotropy between the low energy excitations along the Brillouin zone diagonal (nodal direction) and those along the Brillouin zone face (antinodal direction). Most obvious is the d-wave superconducting gap, with the largest magnitude found in the antinodal direction and no gap in the nodal direction. Additionally, while antinodal quasiparticle excitations appear only below $T_c$, superconductivity is thought to be indifferent to nodal excitations as they are regarded robust and insensitive to $T_c$. Here we reveal an unexpected tie between nodal quasiparticles and superconductivity using high resolution time- and angle-resolved photoemission on optimally doped Bi$_2$Sr$_2$CaCu$_2$O$_{8+\delta}$. We observe a suppression of the nodal quasiparticle spectral weight following pump laser excitation and measure its recovery dynamics. This suppression is dramatically enhanced in the superconducting state. These results reduce the nodal-antinodal dichotomy and challenge the conventional view of nodal excitation neutrality in superconductivity.Comment: 7 pages, 3 figure. To be published in Nature Physic