Unexpected potential protective associations between outdoor air pollution and cataracts

Abstract

Air pollution is one of the biggest public health issues, and the eye is continuously exposed to multiple outdoor air pollution. However, to date, no large-scale study has assessed the relationship between air pollutants and cataracts. We investigated associations between outdoor air pollution and cataracts in the Korean population. A population-based cross-sectional study was performed using data from the Korea National Health and Nutrition Examination Survey, including 18,622 adults more than 40 years of age. The presence of cataracts and their subtypes were evaluated by ophthalmologists. Air pollution data (levels of particulate matter, ozone, nitrogen dioxide, and sulfur dioxide) for the 2 years prior to the ocular examinations were collected from national monitoring stations. The associations of multiple air pollutants with cataracts were assessed by multivariate logistic regression analyses. Sociodemographic factors and previously known risk factors for cataracts were controlled as covariates (model 1 included sociodemographic factors, sun exposure, and behavioral factors, while model 2 further included clinical factors). Higher ozone concentrations were protectively associated with overall cataract which included all subtypes [single pollutant model: 0.003 ppm increase—model 1 (OR 0.89, p = 0.014), model 2 (OR 0.87, p = 0.011); multi-pollutant model: 0.003 ppm increase—model 1 (OR 0.80, p = 0.002), model 2 (OR 0.87, p = 0.002)]. Especially, higher ozone concentrations showed deeply protective association with nuclear cataract subtype [0.003 ppm increase—single pollutant model: model 2 (OR 0.84, p = 0.006), multi-pollutant model: model 2 (OR 0.73, p < 0.001)]. Higher tropospheric ozone concentrations showed protective associations with overall cataract and nuclear cataract subtype in the Korean population.

T-type calcium channels drive migration/invasion in BRAFV600E melanoma cells through Snail1

Abstract

Melanoma is a malignant tumor derived from melanocytes. Once disseminated, it is usually highly resistant to chemotherapy and is associated with poor prognosis. We have recently reported that T-type calcium channels (TTCCs) are overexpressed in melanoma cells and play an important role in melanoma progression. Importantly, TTCC pharmacological blockers reduce proliferation and deregulate autophagy leading to apoptosis. Here, we analyze the role of autophagy during migration/invasion of melanoma cells.

TTCC Cav3.1 and LC3-II protein are highly expressed in BRAFV600E compared to NRAS mutant melanomas, both in cell lines and biopsies. Chloroquine, pharmacological blockade or gene silencing of TTCCs inhibits the autophagic flux and impairs the migration and invasion capabilities, specifically in BRAFV600E melanoma cells.

Snail1 plays an important role in motility and invasion of melanoma cells. We show that Snail1

is strongly expressed in BRAFV600E melanoma cells and patient biopsies, and its expression decreases when autophagy is blocked. These results demonstrate a role of Snail1 during BRAFV600E melanoma progression and strongly suggest that targeting macroautophagy and, particularly TTCCs, might be a good therapeutic strategy to inhibit metastasis of the most common melanoma type (BRAFV600E).

This article is protected by copyright. All rights reserved.

Fate of Lu(III) sorbed on 2-line ferrihydrite at pH 5.7 and aged for 12 years at room temperature. I: insights from ICP-OES, XRD, ESEM, AsFlFFF/ICP-MS, and EXAFS spectroscopy

Abstract

Two-line ferrihydrite (2LFh) was aged for 12 years under ambient conditions and sheltered from light in the presence of Lu(III) used as surrogate for trivalent actinides. 2LFh aging produced hematite rhombohedra with overgrown acicular goethite particles. Analysis of the homogeneous suspension by asymmetrical flow field-flow fractionation (AsFlFFF) coupled to ICP-MS indicated that particles have a mean hydrodynamic diameter of about 140 nm and the strong correlation of the Fe and Lu fractograms hinted at a structural association of the lanthanide with the solid phase(s). Unfortunately, recoveries were low and thus results cannot be considered representative of the whole sample. The suspension was centrifuged and X-ray absorption spectroscopy (XAS) at the Lu L₃ -edge on the settled particles indicated that Lu(III) is sixfold coordinated by oxygen atoms, pointing to a retention by structural incorporation within particles. This result is consistent with AsFlFFF results on the same suspension without centrifugation. The detection of next nearest Fe and O atoms were consistent with the structure of goethite, ruling out incorporation within hematite. After centrifugation of the suspension, only nanoparticulate needle-like particles, very likely goethite, could be detected in the supernatant by ESEM. AsFlFFF data of the supernatant were comparable to that obtained for the homogeneous suspension, whereas XAS indicated that Lu(III) is predominantly present as dissolved species in the supernatant. Results from both techniques can be interpreted as a major fraction of Lu present as aqueous ions and a minor fraction as structurally incorporated. Findings from this study are corroborated by STEM-HAADF data and results from DFT calculations in a companion paper.

Blaschkoid Acute Graft-vs-Host Disease

This case report describes a patient who developed acute graft-vs-host disease along the lines of Blaschko.

Trends in Melanoma Incidence Among Non-Hispanic Whites

This analysis examines data from 2 major databases on melanoma incidence trends among the non-Hispanic white population in the United States by 10-year age groups.

Rapid Repigmentation of Vitiligo-Affected Skin After Tofacitinib Plus UV-B

This case report describes 2 patients who experienced rapid repigmentation of vitiligo-affected skin after tofacitinib plus low-dose, narrowband UV-B therapy.

Follicular and Epidermal Cell Suspension for Vitiligo Treatment

This randomized clinical trial compares the effects of combined epidermal cell suspension and follicular cell suspension with those of epidermal cell suspension alone in treatment of vitiligo.

Prevalence and Types of Genital Lesions in Organ Transplant Recipients

This medical record review assesses the prevalence and types of genital lesions observed in organ transplant recipients.

Attention Shifts Recruit the Monkey Default Mode Network

A unifying function associated with the default mode network (DMN), which is more active during rest than under active task conditions, has been difficult to define. The DMN is activated during monitoring the external world for unexpected events, as a sentinel, and when humans are engaged in high-level internally focused tasks. The existence of DMN correlates in other species, such as mice, challenge the idea that internally focused, high-level cognitive operations, such as introspection, autobiographical memory retrieval, planning the future, and predicting someone else's thoughts, are evolutionarily preserved defining properties of the DMN. A recent human study demonstrated that demanding cognitive shifts could recruit the DMN, yet it is unknown whether this holds for nonhuman species. Therefore, we tested whether large changes in cognitive context would recruit DMN regions in female and male nonhuman primates. Such changes were measured as displacements of spatial attentional weights based on internal rules of relevance (spatial shifts) compared with maintaining attentional weights at the same location (stay events). Using fMRI in macaques, we detected that a cortical network, activated during shifts, largely overlapped with the DMN. Moreover, fMRI time courses sampled from independently defined DMN foci showed significant shift selectivity during the demanding attention task. Finally, functional clustering based on independent resting state data revealed that DMN and shift regions clustered conjointly, whereas regions activated during the stay events clustered apart. We therefore propose that cognitive shifting in primates generally recruits DMN regions. This might explain a breakdown of the DMN in many neurological diseases characterized by declined cognitive flexibility.

SIGNIFICANCE STATEMENT Activation of the human default mode network (DMN) can be measured with fMRI when subjects shift thoughts between high-level internally directed cognitive states, when thinking about the self, the perspective of others, when imagining future and past events, and during mind wandering. Furthermore, the DMN is activated as a sentinel, monitoring the environment for unexpected events. Arguably, these cognitive processes have in common fast and substantial changes in cognitive context. As DMN activity has also been reported in nonhuman species, we tested whether shifts in spatial attention activated the monkey DMN. Core monkey DMN and shift-selective regions shared several functional properties, indicating that cognitive shifting, in general, might constitute one of the evolutionarily preserved functions of the DMN.

This Week in The Journal

In Vivo Analysis of a Gain-of-Function Mutation Confirms Unc18/Munc18's Role in Priming

Selective Suppression of Local Interneuron Circuits in Human Motor Cortex Contributes to Movement Preparation

Changes in neural activity occur in the motor cortex before movement, but the nature and purpose of this preparatory activity is unclear. To investigate this in the human (male and female) brain noninvasively, we used transcranial magnetic stimulation (TMS) to probe the excitability of distinct sets of excitatory inputs to corticospinal neurons during the warning period of various reaction time tasks. Using two separate methods (H-reflex conditioning and directional effects of TMS), we show that a specific set of excitatory inputs to corticospinal neurons are suppressed during motor preparation, while another set of inputs remain unaffected. To probe the behavioral relevance of this suppression, we examined whether the strength of the selective preparatory inhibition in each trial was related to reaction time. Surprisingly, the greater the amount of selective preparatory inhibition, the faster the reaction time was. This suggests that the inhibition of inputs to corticospinal neurons is not involved in preventing the release of movement but may in fact facilitate rapid reactions. Thus, selective suppression of a specific set of motor cortical neurons may be a key aspect of successful movement preparation.

SIGNIFICANCE STATEMENT Movement preparation evokes substantial activity in the motor cortex despite no apparent movement. One explanation for the lack of movement is that motor cortical output in this period is gated by an inhibitory mechanism. This notion was supported by previous noninvasive TMS studies of human motor cortex indicating a reduction of corticospinal excitability. On the contrary, our data support the idea that there is a coordinated balance of activity upstream of the corticospinal output neurons. This includes a suppression of specific local circuits that supports, rather than inhibits, the rapid generation of prepared movements. Thus, the selective suppression of local circuits appears to be an essential part of successful movement preparation instead of an external control mechanism.

Phosphorylation Switch of Orco Shapes the Sense of Smell in Insects

Attention Is Required for Knowledge-Based Sequential Grouping: Insights from the Integration of Syllables into Words

How the brain groups sequential sensory events into chunks is a fundamental question in cognitive neuroscience. This study investigates whether top–down attention or specific tasks are required for the brain to apply lexical knowledge to group syllables into words. Neural responses tracking the syllabic and word rhythms of a rhythmic speech sequence were concurrently monitored using electroencephalography (EEG). The participants performed different tasks, attending to either the rhythmic speech sequence or a distractor, which was another speech stream or a nonlinguistic auditory/visual stimulus. Attention to speech, but not a lexical-meaning-related task, was required for reliable neural tracking of words, even when the distractor was a nonlinguistic stimulus presented cross-modally. Neural tracking of syllables, however, was reliably observed in all tested conditions. These results strongly suggest that neural encoding of individual auditory events (i.e., syllables) is automatic, while knowledge-based construction of temporal chunks (i.e., words) crucially relies on top–down attention.

SIGNIFICANCE STATEMENT Why we cannot understand speech when not paying attention is an old question in psychology and cognitive neuroscience. Speech processing is a complex process that involves multiple stages, e.g., hearing and analyzing the speech sound, recognizing words, and combining words into phrases and sentences. The current study investigates which speech-processing stage is blocked when we do not listen carefully. We show that the brain can reliably encode syllables, basic units of speech sounds, even when we do not pay attention. Nevertheless, when distracted, the brain cannot group syllables into multisyllabic words, which are basic units for speech meaning. Therefore, the process of converting speech sound into meaning crucially relies on attention.

Glutamatergic Transmission to Hypothalamic Kisspeptin Neurons Is Differentially Regulated by Estradiol through Estrogen Receptor {alpha} in Adult Female Mice

Estradiol feedback regulates gonadotropin-releasing hormone (GnRH) neurons and subsequent luteinizing hormone (LH) release. Estradiol acts via estrogen receptor α (ERα)-expressing afferents of GnRH neurons, including kisspeptin neurons in the anteroventral periventricular (AVPV) and arcuate nuclei, providing homeostatic feedback on episodic GnRH/LH release as well as positive feedback to control ovulation. Ionotropic glutamate receptors are important for estradiol feedback, but it is not known where they fit in the circuitry. Estradiol-negative feedback decreased glutamatergic transmission to AVPV and increased it to arcuate kisspeptin neurons; positive feedback had the opposite effect. Deletion of ERα in kisspeptin cells decreased glutamate transmission to AVPV neurons and markedly increased it to arcuate kisspeptin neurons, which also exhibited increased spontaneous firing rate. KERKO mice had increased LH pulse frequency, indicating loss of negative feedback. These observations indicate that ERα in kisspeptin cells is required for appropriate differential regulation of these neurons and neuroendocrine output by estradiol.

SIGNIFICANCE STATEMENT The brain regulates fertility through gonadotropin-releasing hormone (GnRH) neurons. Ovarian estradiol regulates the pattern of GnRH (negative feedback) and initiates a surge of release that triggers ovulation (positive feedback). GnRH neurons do not express the estrogen receptor needed for feedback (estrogen receptor α [ERα]); kisspeptin neurons in the arcuate and anteroventral periventricular nuclei are postulated to mediate negative and positive feedback, respectively. Here we extend the network through which feedback is mediated by demonstrating that glutamatergic transmission to these kisspeptin populations is differentially regulated during the reproductive cycle and by estradiol. Electrophysiological and in vivo hormone profile experiments on kisspeptin-specific ERα knock-out mice demonstrate that ERα in kisspeptin cells is required for appropriate differential regulation of these neurons and for neuroendocrine output.

Tenuous Inhibitory GABAergic Signaling in the Reticular Thalamus

Maintenance of a low intracellular Cl^– concentration ([Cl^–]_i) is critical for enabling inhibitory neuronal responses to GABA_A receptor-mediated signaling. Cl^– transporters, including KCC2, and extracellular impermeant anions ([A]_o) of the extracellular matrix are both proposed to be important regulators of [Cl^–]_i. Neurons of the reticular thalamic (RT) nucleus express reduced levels of KCC2, indicating that GABAergic signaling may produce excitation in RT neurons. However, by performing perforated patch recordings and calcium imaging experiments in rats (male and female), we find that [Cl^–]_i remains relatively low in RT neurons. Although we identify a small contribution of [A]_o to a low [Cl^–]_i in RT neurons, our results also demonstrate that reduced levels of KCC2 remain sufficient to maintain low levels of Cl^–. Reduced KCC2 levels, however, restrict the capacity of RT neurons to rapidly extrude Cl^– following periods of elevated GABAergic signaling. In a computational model of a local RT network featuring slow Cl^– extrusion kinetics, similar to those we found experimentally, model RT neurons are predisposed to an activity-dependent switch from GABA-mediated inhibition to excitation. By decreasing the activity threshold required to produce excitatory GABAergic signaling, weaker stimuli are able to propagate activity within the model RT nucleus. Our results indicate the importance of even diminished levels of KCC2 in maintaining inhibitory signaling within the RT nucleus and suggest how this important activity choke point may be easily overcome in disorders such as epilepsy.

SIGNIFICANCE STATEMENT Precise regulation of intracellular Cl^– levels ([Cl^–]_i) preserves appropriate, often inhibitory, GABAergic signaling within the brain. However, there is disagreement over the relative contribution of various mechanisms that maintain low [Cl^–]_i. We found that the Cl^– transporter KCC2 is an important Cl^– extruder in the reticular thalamic (RT) nucleus, despite this nucleus having remarkably low KCC2 immunoreactivity relative to other regions of the adult brain. We also identified a smaller contribution of fixed, impermeant anions ([A]_o) to lowering [Cl^–]_i in RT neurons. Inhibitory signaling among RT neurons is important for preventing excessive activation of RT neurons, which can be responsible for generating seizures. Our work suggests that KCC2 critically restricts the spread of activity within the RT nucleus.

HRPU-2, a Homolog of Mammalian hnRNP U, Regulates Synaptic Transmission by Controlling the Expression of SLO-2 Potassium Channel in Caenorhabditis elegans

Slo2 channels are large-conductance potassium channels abundantly expressed in the nervous system. However, it is unclear how their expression level in neurons is regulated. Here we report that HRPU-2, an RNA-binding protein homologous to mammalian heterogeneous nuclear ribonucleoprotein U (hnRNP U), plays an important role in regulating the expression of SLO-2 (a homolog of mammalian Slo2) in Caenorhabditis elegans. Loss-of-function (lf) mutants of hrpu-2 were isolated in a genetic screen for suppressors of a sluggish phenotype caused by a hyperactive SLO-2. In hrpu-2(lf) mutants, SLO-2-mediated delayed outward currents in neurons are greatly decreased, and neuromuscular synaptic transmission is enhanced. These mutant phenotypes can be rescued by expressing wild-type HRPU-2 in neurons. HRPU-2 binds to slo-2 mRNA, and hrpu-2(lf) mutants show decreased SLO-2 protein expression. In contrast, hrpu-2(lf) does not alter the expression of either the BK channel SLO-1 or the Shaker type potassium channel SHK-1. hrpu-2(lf) mutants are indistinguishable from wild type in gross motor neuron morphology and locomotion behavior. Together, these observations suggest that HRPU-2 plays important roles in SLO-2 function by regulating SLO-2 protein expression, and that SLO-2 is likely among a restricted set of proteins regulated by HRPU-2. Mutations of human Slo2 channel and hnRNP U are strongly linked to epileptic disorders and intellectual disability. The findings of this study suggest a potential link between these two molecules in human patients.

SIGNIFICANCE STATEMENT Heterogeneous nuclear ribonucleoprotein U (hnRNP U) belongs to a family of RNA-binding proteins that play important roles in controlling gene expression. Recent studies have established a strong link between mutations of hnRNP U and human epilepsies and intellectual disability. However, it is unclear how mutations of hnRNP U may cause such disorders. This study shows that mutations of HRPU-2, a worm homolog of mammalian hnRNP U, result in dysfunction of a Slo2 potassium channel, which is critical to neuronal function. Because mutations of Slo2 channels are also strongly associated with epileptic encephalopathies and intellectual disability in humans, the findings of this study point to a potential mechanism underlying neurological disorders caused by hnRNP U mutations.

Visually-Driven Maps in Area 3b

Sensory perception relies on the precise neuronal encoding of modality-specific environmental features in primary sensory cortices. Some studies have reported the penetration of signals from other modalities even into early sensory areas. So far, no comprehensive account of maps induced by "foreign sources" exists. We addressed this question using surface-based topographic mapping techniques applied to ultra-high resolution fMRI neuroimaging data, measured in female participants. We show that fine-grained finger maps in human primary somatosensory cortex, area 3b, are somatotopically activated not only during tactile mechanical stimulation, but also when viewing the same fingers being touched. Visually-induced maps were weak in amplitude, but overlapped with the stronger tactile maps tangential to the cortical sheet when finger touches were observed in both first- and third-person perspectives. However, visually-induced maps did not overlap tactile maps when the observed fingers were only approached by an object but not actually touched. Our data provide evidence that "foreign source maps" in early sensory cortices are present in the healthy human brain, that their arrangement is precise, and that their induction is feature-selective. The computations required to generate such specific responses suggest that counterflow (feedback) processing may be much more spatially specific than has been often assumed.

SIGNIFICANCE STATEMENT Using ultra-high field fMRI, we provide empirical evidence that viewing touches activates topographically aligned single finger maps in human primary somatosensory cortical area 3b. This shows that "foreign source maps" in early sensory cortices are topographic, precise, and feature-selective in healthy human participants with intact sensory pathways.

A{beta} mediates F-actin disassembly in dendritic spines leading to cognitive deficits in Alzheimer's disease

Dendritic spine loss is recognized as an early feature of Alzheimer's disease (AD), but the underlying mechanisms are poorly understood. Dendritic spine structure is defined by filamentous actin (F-actin) and we observed depolymerization of synaptosomal F-actin accompanied by increased globular-actin (G-actin) at as early as 1 month of age in a mouse model of AD (APPswe/PS1E9, male mice). This led to recall deficit after contextual fear conditioning (cFC) at 2 months of age in APPswe/PS1E9 male mice, which could be reversed by the actin-polymerizing agent jasplakinolide. Further, the F-actin-depolymerizing agent latrunculin induced recall deficit after cFC in WT mice, indicating the importance of maintaining F-/G-actin equilibrium for optimal behavioral response. Using direct stochastic optical reconstruction microscopy (dSTORM), we show that F-actin depolymerization in spines leads to a breakdown of the nano-organization of outwardly radiating F-actin rods in cortical neurons from APPswe/PS1E9 mice. Our results demonstrate that synaptic dysfunction seen as F-actin disassembly occurs very early, before onset of pathological hallmarks in AD mice, and contributes to behavioral dysfunction, indicating that depolymerization of F-actin is causal and not consequent to decreased spine density. Further, we observed decreased synaptosomal F-actin levels in postmortem brain from mild cognitive impairment and AD patients compared with subjects with normal cognition. F-actin decrease correlated inversely with increasing AD pathology (Braak score, Aβ load, and tangle density) and directly with performance in episodic and working memory tasks, suggesting its role in human disease pathogenesis and progression.

SIGNIFICANCE STATEMENT Synaptic dysfunction underlies cognitive deficits in Alzheimer's disease (AD). The cytoskeletal protein actin plays a critical role in maintaining structure and function of synapses. Using cultured neurons and an AD mouse model, we show for the first time that filamentous actin (F-actin) is lost selectively from synapses early in the disease process, long before the onset of classical AD pathology. We also demonstrate that loss of synaptic F-actin contributes directly to memory deficits. Loss of synaptosomal F-actin in human postmortem tissue correlates directly with decreased performance in memory test and inversely with AD pathology. Our data highlight that synaptic cytoarchitectural changes occur early in AD and they may be targeted for the development of therapeutics.

Loss of Intercalated Cells (ITCs) in the Mouse Amygdala of Tshz1 Mutants Correlates with Fear, Depression, and Social Interaction Phenotypes

The intercalated cells (ITCs) of the amygdala have been shown to be critical regulatory components of amygdalar circuits, which control appropriate fear responses. Despite this, the molecular processes guiding ITC development remain poorly understood. Here we establish the zinc finger transcription factor Tshz1 as a marker of ITCs during their migration from the dorsal lateral ganglionic eminence through maturity. Using germline and conditional knock-out (cKO) mouse models, we show that Tshz1 is required for the proper migration and differentiation of ITCs. In the absence of Tshz1, migrating ITC precursors fail to settle in their stereotypical locations encapsulating the lateral amygdala and BLA. Furthermore, they display reductions in the ITC marker Foxp2 and ectopic persistence of the dorsal lateral ganglionic eminence marker Sp8. Tshz1 mutant ITCs show increased cell death at postnatal time points, leading to a dramatic reduction by 3 weeks of age. In line with this, Foxp2-null mutants also show a loss of ITCs at postnatal time points, suggesting that Foxp2 may function downstream of Tshz1 in the maintenance of ITCs. Behavioral analysis of male Tshz1 cKOs revealed defects in fear extinction as well as an increase in floating during the forced swim test, indicative of a depression-like phenotype. Moreover, Tshz1 cKOs display significantly impaired social interaction (i.e., increased passivity) regardless of partner genetics. Together, these results suggest that Tshz1 plays a critical role in the development of ITCs and that fear, depression-like and social behavioral deficits arise in their absence.

SIGNIFICANCE STATEMENT We show here that the zinc finger transcription factor Tshz1 is expressed during development of the intercalated cells (ITCs) within the mouse amygdala. These neurons have previously been shown to play a crucial role in fear extinction. Tshz1 mouse mutants exhibit severely reduced numbers of ITCs as a result of abnormal migration, differentiation, and survival of these neurons. Furthermore, the loss of ITCs in mouse Tshz1 mutants correlates well with defects in fear extinction as well as the appearance of depression-like and abnormal social interaction behaviors reminiscent of depressive disorders observed in human patients with distal 18q deletions, including the Tshz1 locus.

Disruption of Interneuron Neurogenesis in Premature Newborns and Reversal with Estrogen Treatment

Many Preterm-born children suffer from neurobehavioral disorders. Premature birth terminates the hypoxic in utero environment and supply of maternal hormones. As the production of interneurons continues until the end of pregnancy, we hypothesized that premature birth would disrupt interneuron production and that restoration of the hypoxic milieu or estrogen treatment might reverse interneuron generation. To test these hypotheses, we compared interneuronal progenitors in the medial ganglionic eminences (MGEs), lateral ganglionic eminences (LGEs), and caudal ganglionic eminences (CGEs) between preterm-born [born on embryonic day (E) 29; examined on postnatal day (D) 3 and D7] and term-born (born on E32; examined on D0 and D4) rabbits at equivalent postconceptional ages. We found that both total and cycling Nkx2.1⁺, Dlx2⁺, and Sox2⁺ cells were more abundant in the MGEs of preterm rabbits at D3 compared with term rabbits at D0, but not in D7 preterm relative to D4 term pups. Total Nkx2.1⁺ progenitors were also more numerous in the LGEs of preterm pups at D3 compared with term rabbits at D0. Dlx2⁺ cells in CGEs were comparable between preterm and term pups. Simulation of hypoxia by dimethyloxalylglycine treatment did not affect the number of interneuronal progenitors. However, estrogen treatment reduced the density of total and proliferating Nkx2.1⁺ and Dlx2⁺ cells in the MGEs and enhanced Ascl1 transcription factor. Estrogen treatment also reduced Ki67, c-Myc, and phosphorylation of retinoblastoma protein, suggesting inhibition of the G1-to-S phase transition. Hence, preterm birth disrupts interneuron neurogenesis in the MGE and estrogen treatment reverses interneuron neurogenesis in preterm newborns by cell-cycle inhibition and elevation of Ascl1. We speculate that estrogen replacement might partially restore neurogenesis in human premature infants.

SIGNIFICANCE STATEMENT Prematurity results in developmental delays and neurobehavioral disorders, which might be ascribed to disturbances in the development of cortical interneurons. Here, we show that preterm birth disrupts interneuron neurogenesis in the medial ganglionic eminence (MGE) and, more importantly, that estrogen treatment reverses this perturbation in the population of interneuron progenitors in the MGE. The estrogen seems to restore neurogenesis by inhibiting the cell cycle and elevating Ascl1 expression. As preterm birth causes plasma estrogen level to drop 100-fold, the estrogen replacement in preterm infants is physiological. We speculate that estrogen replacement might ameliorate disruption in production of interneurons in human premature infants.

Prismatic Adaptation Modulates Oscillatory EEG Correlates of Motor Preparation but Not Visual Attention in Healthy Participants

Prismatic adaption (PA) has been proposed as a tool to induce neural plasticity and is used to help neglect rehabilitation. It leads to a recalibration of visuomotor coordination during pointing as well as to aftereffects on a number of sensorimotor and attention tasks, but whether these effects originate at a motor or attentional level remains a matter of debate. Our aim was to further characterize PA aftereffects by using an approach that allows distinguishing between effects on attentional and motor processes. We recorded EEG in healthy human participants (9 females and 7 males) while performing a new double step, anticipatory attention/motor preparation paradigm before and after adaptation to rightward-shifting prisms, with neutral lenses as a control. We then examined PA aftereffects through changes in known oscillatory EEG signatures of spatial attention orienting and motor preparation in the alpha and beta frequency bands. Our results were twofold. First, we found PA to rightward-shifting prisms to selectively affect EEG signatures of motor but not attentional processes. More specifically, PA modulated preparatory motor EEG activity over central electrodes in the right hemisphere, contralateral to the PA-induced, compensatory leftward shift in pointing movements. No effects were found on EEG signatures of spatial attention orienting over occipitoparietal sites. Second, we found the PA effect on preparatory motor EEG activity to dominate in the beta frequency band. We conclude that changes to intentional visuomotor, rather than attentional visuospatial, processes underlie the PA aftereffect of rightward-deviating prisms in healthy participants.

SIGNIFICANCE STATEMENT Prismatic adaptation (PA) has been proposed as a tool to induce neural plasticity in both healthy participants and patients, due to its aftereffect impacting on a number of visuospatial and visuomotor functions. However, the neural mechanisms underlying PA aftereffects are poorly understood as only little neuroimaging evidence is available. Here, we examined, for the first time, the origin of PA aftereffects studying oscillatory brain activity. Our results show a selective modulation of preparatory motor activity following PA in healthy participants but no effect on attention-related activity. This provides novel insight into the PA aftereffect in the healthy brain and may help to inform interventions in neglect patients.

Decoding Trans-Saccadic Memory

We examine whether peripheral information at a planned saccade target affects immediate postsaccadic processing at the fovea on saccade landing. Current neuroimaging research suggests that presaccadic stimulation has a late effect on postsaccadic processing, in contrast to the early effect seen in behavioral studies. Human participants (both male and female) were instructed to saccade toward a face or a house that, on different trials, remained the same, changed, or disappeared during the saccade. We used a multivariate pattern analysis of electroencephalography data to decode face versus house processing directly after the saccade. The classifier was trained on separate trials without a saccade, where a house or face was presented at the fovea. When the saccade target remained the same across the saccade, we could reliably decode the target 123 ms after saccade offset. In contrast, when the target was changed during the saccade, the new target was decoded at a later time-point, 151 ms after saccade offset. The "same" condition advantage suggests that congruent presaccadic information facilitates processing of the postsaccadic stimulus compared with incongruent information. Finally, the saccade target could be decoded above chance even when it had been removed during the saccade, albeit with a slower time course (162 ms) and poorer signal strength. These findings indicate that information about the (peripheral) presaccadic stimulus is transferred across the saccade so that it becomes quickly available and influences processing at its expected new retinal position (the fovea).

SIGNIFICANCE STATEMENT Here we provide neural evidence for early information transfer across saccades. Specifically, we examined the effect of presaccadic sensory information on the initial neuronal processing of a postsaccadic stimuli. Using electroencephalography and multivariate pattern analysis, we found the following: (1) that the identity of the presaccadic stimulus modulated the postsaccadic latency of stimulus relevant information; and (2) that a saccadic neural marker for a saccade target stimulus could be detected even when the stimulus had been removed during saccade. These results demonstrate that information about the peripheral presaccadic stimulus was transferred across the saccade and influenced processing at a new retinal position (the fovea) directly after the saccade landed.

Conditional Deletion of Hippocampal CA2/CA3a Oxytocin Receptors Impairs the Persistence of Long-Term Social Recognition Memory in Mice

Oxytocin (OXT) receptors (OXTRs) are prominently expressed in hippocampal CA2 and CA3 pyramidal neurons, but little is known about its physiological function. As the functional necessity of hippocampal CA2 for social memory processing, we tested whether CA2 OXTRs may contribute to long-term social recognition memory (SRM) formation. Here, we found that conditional deletion of Oxtr from forebrain (Oxtr^–/–) or CA2/CA3a-restricted excitatory neurons in adult male mice impaired the persistence of long-term SRM but had no effect on sociability and preference for social novelty. Conditional deletion of CA2/CA3a Oxtr showed no changes in anxiety-like behavior assessed using the open-field, elevated plus maze and novelty-suppressed feeding tests. Application of a highly selective OXTR agonist [Thr⁴,Gly⁷]-OXT to hippocampal slices resulted in an acute and lasting potentiation of excitatory synaptic responses in CA2 pyramidal neurons that relied on N-methyl-d-aspartate receptor activation and calcium/calmodulin-dependent protein kinase II activity. In addition, Oxtr^–/– mice displayed a defect in the induction of long-term potentiation, but not long-term depression, at the synapses between the entorhinal cortex and CA2 pyramidal neurons. Furthermore, Oxtr deletion led to a reduced complexity of basal dendritic arbors of CA2 pyramidal neurons, but caused no alteration in the density of apical dendritic spines. Considering that the methodologies we have used to delete Oxtr do not rule out targeting the neighboring CA3a region, these findings suggest that OXTR signaling in the CA2/CA3a is crucial for the persistence of long-term SRM.

SIGNIFICANCE STATEMENT Oxytocin receptors (OXTRs) are abundantly expressed in hippocampal CA2 and CA3 regions, but there are little known about their physiological function. Taking advantage of the conditional Oxtr knock-out mice, the present study highlights the importance of OXTR signaling in the induction of long-term potentiation at the synapses between the entorhinal cortex and CA2 pyramidal neurons and the persistence of long-term social recognition memory. Thus, OXTRs in the CA2/CA3a may provide a new target for therapeutic approaches to the treatment of social cognition deficits, which are often observed in patients with neuropsychiatric disorders.

DRG Voltage-Gated Sodium Channel 1.7 Is Upregulated in Paclitaxel-Induced Neuropathy in Rats and in Humans with Neuropathic Pain

Chemotherapy-induced peripheral neuropathy (CIPN) is a common adverse effect experienced by cancer patients receiving treatment with paclitaxel. The voltage-gated sodium channel 1.7 (Na_v1.7) plays an important role in multiple preclinical models of neuropathic pain and in inherited human pain phenotypes, and its gene expression is increased in dorsal root ganglia (DRGs) of paclitaxel-treated rats. Hence, the potential of change in the expression and function of Na_v1.7 protein in DRGs from male rats with paclitaxel-related CIPN and from male and female humans with cancer-related neuropathic pain was tested here. Double immunofluorescence in CIPN rats showed that Na_v1.7 was upregulated in small DRG neuron somata, especially those also expressing calcitonin gene-related peptide (CGRP), and in central processes of these cells in the superficial spinal dorsal horn. Whole-cell patch-clamp recordings in rat DRG neurons revealed that paclitaxel induced an enhancement of ProTx II (a selective Na_v1.7 channel blocker)-sensitive sodium currents. Bath-applied ProTx II suppressed spontaneous action potentials in DRG neurons occurring in rats with CIPN, while intrathecal injection of ProTx II significantly attenuated behavioral signs of CIPN. Complementarily, DRG neurons isolated from segments where patients had a history of neuropathic pain also showed electrophysiological and immunofluorescence results indicating an increased expression of Na_v1.7 associated with spontaneous activity. Na_v1.7 was also colocalized in human cells expressing transient receptor potential vanilloid 1 and CGRP. Furthermore, ProTx II decreased firing frequency in human DRGs with spontaneous action potentials. This study suggests that Na_v1.7 may provide a potential new target for the treatment of neuropathic pain, including chemotherapy (paclitaxel)-induced neuropathic pain.

SIGNIFICANCE STATEMENT This work demonstrates that the expression and function of the voltage-gated sodium channel Na_v1.7 are increased in a preclinical model of chemotherapy-induced peripheral neuropathy (CIPN), the most common treatment-limiting side effect of all the most common anticancer therapies. This is key as gain-of-function mutations in human Na_v1.7 recapitulate both the distribution and pain percept as shown by CIPN patients. This work also shows that Na_v1.7 is increased in human DRG neurons only in dermatomes where patients are experiencing acquired neuropathic pain symptoms. This work therefore has major translational impact, indicating an important novel therapeutic avenue for neuropathic pain as a class.

Gonadotropin-Releasing Hormone (GnRH) Neuron Excitability Is Regulated by Estradiol Feedback and Kisspeptin

Gonadotropin-releasing hormone (GnRH) neurons produce the central output controlling fertility and are regulated by steroid feedback. A switch from estradiol negative to positive feedback initiates the GnRH surge, ultimately triggering ovulation. This occurs on a daily basis in ovariectomized, estradiol-treated (OVX+E) mice; GnRH neurons are suppressed in the morning and activated in the afternoon. To test the hypotheses that estradiol and time of day signals alter GnRH neuron responsiveness to stimuli, GFP-identified GnRH neurons in brain slices from OVX+E or OVX female mice were recorded during the morning or afternoon. No differences were observed in baseline membrane potential. Current-clamp revealed GnRH neurons fired more action potentials in response to current injection during positive feedback relative to all other groups, which were not different from each other despite reports of differing ionic conductances. Kisspeptin increased GnRH neuron response in cells from OVX and OVX+E mice in the morning but not afternoon. Paradoxically, excitability in kisspeptin knock-out mice was similar to the maximum observed in control mice but was unchanged by time of day or estradiol. A mathematical model applying a Markov Chain Monte Carlo method to estimate probability distributions for estradiol- and time of day–dependent parameters was used to predict intrinsic properties underlying excitability changes. A single identifiable distribution of solutions accounted for similar GnRH neuron excitability in all groups other than positive feedback despite different underlying conductance properties; this was attributable to interdependence of voltage-gated potassium channel properties. In contrast, redundant solutions may explain positive feedback, perhaps indicative of the importance of this state for species survival.

SIGNIFICANCE STATEMENT Infertility affects 15%–20% of couples; failure to ovulate is a common cause. Understanding how the brain controls ovulation is critical for new developments in both infertility treatment and contraception. Gonadotropin-releasing hormone (GnRH) neurons are the final common pathway for central neural control of ovulation. We studied how estradiol feedback regulates GnRH excitability, a key determinant of neural firing rate using laboratory and computational approaches. GnRH excitability is upregulated during positive feedback, perhaps driving increased neural firing rate at this time. Kisspeptin increased GnRH excitability and was essential for estradiol regulation of excitability. Modeling predicts that multiple combinations of changes to GnRH intrinsic conductances can produce the firing response in positive feedback, suggesting the brain has many ways to induce ovulation.

Cholinergic Overstimulation Attenuates Rule Selectivity in Macaque Prefrontal Cortex

Acetylcholine is released in the prefrontal cortex (PFC) and is a key modulator of cognitive performance in primates. Cholinergic stimulation has been shown to have beneficial effects on performance of cognitive tasks, and cholinergic receptors are being actively explored as promising targets for ameliorating cognitive deficits in Alzheimer's disease. We hypothesized that cholinergic stimulation of PFC during performance of a cognitive task would augment neuronal activity and neuronal coding of task attributes. We iontophoretically applied the general cholinergic receptor agonist carbachol onto neurons in dorsolateral PFC (DLPFC) of male rhesus macaques performing rule-guided prosaccades and antisaccades, a well established oculomotor task for testing cognitive control. Carbachol application had heterogeneous effects on neuronal excitability, with both excitation and suppression observed in significant proportions. Contrary to our prediction, neurons with rule-selective activity exhibited a reduction in selectivity during carbachol application. Cholinergic stimulation disrupted rule selectivity regardless of whether it had suppressive or excitatory effects on these neurons. In addition, cholinergic stimulation excited putative pyramidal neurons, whereas the activity of putative interneurons remained unchanged. Moreover, cholinergic stimulation attenuated saccade direction selectivity in putative pyramidal neurons due to nonspecific increases in activity. Our results suggest excessive cholinergic stimulation has detrimental effects on DLPFC representations of task attributes. These findings delineate the complexity and heterogeneity of neuromodulation of cerebral cortex by cholinergic stimulation, an area of active exploration with respect to the development of cognitive enhancers.

SIGNIFICANCE STATEMENT The neurotransmitter acetylcholine is known to be important for cognitive processes in the prefrontal cortex. Removal of acetylcholine from prefrontal cortex can disrupt short-term memory performance and is reminiscent of Alzheimer's disease, which is characterized by degeneration of acetylcholine-producing neurons. Stimulation of cholinergic receptors is being explored to create cognitive enhancers for the treatment of Alzheimer's disease and other psychiatric diseases. Here, we stimulated cholinergic receptors in prefrontal cortex and examined its effects on neurons that are engaged in cognitive behavior. Surprisingly, cholinergic stimulation decreased neurons' ability to discriminate between rules. This work suggests that overstimulation of acetylcholine receptors could disrupt neuronal processing during cognition and is relevant to the design of cognitive enhancers based on stimulating the cholinergic system.

Meis1 Coordinates Cerebellar Granule Cell Development by Regulating Pax6 Transcription, BMP Signaling and Atoh1 Degradation

Cerebellar granule cell precursors (GCPs) and granule cells (GCs) represent good models to study neuronal development. Here, we report that the transcription factor myeloid ectopic viral integration site 1 homolog (Meis1) plays pivotal roles in the regulation of mouse GC development. We found that Meis1 is expressed in GC lineage cells and astrocytes in the cerebellum during development. Targeted disruption of the Meis1 gene specifically in the GC lineage resulted in smaller cerebella with disorganized lobules. Knock-down/knock-out (KO) experiments for Meis1 and in vitro assays showed that Meis1 binds to an upstream sequence of Pax6 to enhance its transcription in GCPs/GCs and also suggested that the Meis1–Pax6 cascade regulates morphology of GCPs/GCs during development. In the conditional KO (cKO) cerebella, many Atoh1-positive GCPs were observed ectopically in the inner external granule layer (EGL) and a similar phenomenon was observed in cultured cerebellar slices treated with a bone morphogenic protein (BMP) inhibitor. Furthermore, expression of Smad proteins and Smad phosphorylation were severely reduced in the cKO cerebella and Meis1-knock-down GCPs cerebella. Reduction of phosphorylated Smad was also observed in cerebellar slices electroporated with a Pax6 knock-down vector. Because it is known that BMP signaling induces Atoh1 degradation in GCPs, these findings suggest that the Meis1–Pax6 pathway increases the expression of Smad proteins to upregulate BMP signaling, leading to degradation of Atoh1 in the inner EGL, which contributes to differentiation from GCPs to GCs. Therefore, this work reveals crucial functions of Meis1 in GC development and gives insights into the general understanding of the molecular machinery underlying neural differentiation from neural progenitors.

SIGNIFICANCE STATEMENT We report that myeloid ectopic viral integration site 1 homolog (Meis1) plays pivotal roles in the regulation of mouse granule cell (GC) development. Here, we show Meis1 is expressed in GC precursors (GCPs) and GCs during development. Our knock-down and conditional knock-out (cKO) experiments and in vitro assays revealed that Meis1 is required for proper cerebellar structure formation and for Pax6 transcription in GCPs and GCs. The Meis1–Pax6 cascade regulates the morphology of GCs. In the cKO cerebella, Smad proteins and bone morphogenic protein (BMP) signaling are severely reduced and Atoh1-expressing GCPs are ectopically detected in the inner external granule layer. These findings suggest that Meis1 regulates degradation of Atoh1 via BMP signaling, contributing to GC differentiation in the inner EGL, and should provide understanding into GC development.

Itraconazole Induced Hypertension and Hypokalemia: Mechanistic Evaluation

Abstract

We describe a case of apparent mineralocorticoid excess (hypertension, hypokalemia, metabolic alkalosis, and low plasma renin activity) secondary to itraconazole therapy. Inhibition of 11β-hydroxysteroid dehydrogenase 2 was demonstrated, and withholding itraconazole led to resolution of adverse effects that did not recur with voriconazole. This report adds to a growing body of evidence linking apparent mineralocorticoid excess with certain triazoles.

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Exposure to the antifouling chemical medetomidine slows development, reduces body mass, and delays metamorphosis in wood frog ( Lithobates sylvaticus ) tadpoles

Abstract

Antifouling chemicals have a long history of causing toxicity to aquatic organisms. We measured growth and developmental timing in wood frog tadpoles exposed to the antifouling chemical medetomidine (10 nM–10 μM) starting at two different developmental stages in static renewal experiments. For tadpoles hatched from egg masses and exposed for 3 weeks to 100 nM and 1 μM, head width/total body length ratio was significantly shorter compared to control. For field-collected tadpoles at Gosner stage 24–25 and exposed for 2 weeks, 1 and 10 μM medetomidine significantly slowed development as measured by Gosner stage. Medetomidine (1 and 10 μM) significantly increased the time to metamorphosis by over 16 days on average, and at 100 nM and 1 μM, it significantly decreased mass at metamorphosis. We discuss the possible effects of antifouling chemicals containing medetomidine on globally threatened groups such as amphibians.

Finding the brake: dissecting the RhoA pathway regulating glioma cell migration

Abstract

Cell migration is one of the hallmarks of cancer. Cancer cells can adopt two main migratory strategies displaying either a mesenchymal or amoeboid phenotype. Targeting cell migration presents an opportunity in improving treatment of invasive and migratory tumours, however the cellular mechanisms that control the cell migration phenotypes in high-grade gliomas, such as Glioblastoma multiforme, are still not fully understood. In previous work we used 2D and 3D migration assays to screen a panel of inhibitors targeting the actin polymerisation pathway, a key regulator of cell migration in glioma. Amongst RhoA pathway inhibitors, which are key negative regulators of migration, we have identified one compound, CCG-1423, that uniquely failed to inhibit migration. CCG-1423 induces a reduced expression of the adhesion protein CCN1 by blocking nuclear import of MKL-1, a transcriptional co-activator. We suggest that CCG-1423 induces a mesenchymal transition, which permits continued migration. To confirm the effect of CCG-1423 on CCN1 levels we used ELISA to detect secreted CCN1 levels in culture supernatants in treated and non-treated U251 and U87 cells. CCG-1423 significantly reduced CCN1 levels in supernatant of both cell lines compared to untreated controls. Further biochemical analysis was performed to establish levels of MKL-1 in treated versus untreated cells and to elucidate its association with other proteins. Preliminary data indicate an increase in MKL-1 concentration in treated cells, consistent with a build-up of MKL-1 in the cytosol due to lack of nuclear import, as previously reported. Further detailed characterisation of the RhoA signalling pathway in glioma will provide an understanding of the role of MKL-1 in mesenchymal migration and the effect of CCG-1423 on mesenchymal switching. We will next characterise the interactions between CCG-1423 and MKL-1, and use pharmacological and/or genetic manipulation to refine combination treatments to target both mesenchymal and amoeboid cell migration.

IDH1 mutations drive an oxygen-sparing metabolic phenotype to permit tumour growth

Abstract

Since the description of mutations in isocitrate dehydrogenase 1 (IDH1) in adult glial tumours in 2008, many studies have focused on the role of the oncometabolite 2-hydroxyglutarate in determining the pheontype of affected gliomas. However, another aspect of IDH1 activity - its redox-dependency - may also represent an important determinant of IDH1-mutant glioma phenotype. To examine the effect of IDH1 mutations on redox homeostasis, we traced their use of redox-active metabolic pathways using nutrients enriched in the stable isotope, 13C. We found that cells expressing the IDH1 R132H mutant demonstrated enhanced proline synthesis, but that unexpectedly the proline produced was excreted rather than used for biosynthetic processes. As proline synthesis is a highly redox-dependent pathway, we hypothesised that this represented a metabolic response to altered isocitrate metabolism by the IDH1 mutant enzyme. We furthermore showed that metabolic response was dependent on pyrroline 5-carboxylate reductase 1 (PYCR1), and was important for the oxidation of mitochondrial NADH. Consistent with this, knockdown of PYCR1 resulted in a compensatory increase in cell respiration. We therefore show that cells with IDH1 mutations require proline synthesis in order to retain normal TCA cycle activity while using less oxygen. This may mean that IDH1 mutant tumours are in general less hypoxic, and therefore more responsive to treatment.

High-resolution RNA-seq profiling of glioblastoma residual invasive cells isolated by a novel 5-ALA fluorescence based method

Abstract

INTRODUCTION

Malignant brain cancer Glioblastoma (GBM) kills 3500 people per year in the UK with an average survival of 6 months. The intra-tumour heterogeneity of GBM has meant tailored therapies based on the genetic mutations within the cancer cells have so far failed to replicate in vitro promise in clinical trials. Previous research has focused on the core of the tumour; however this is removed by surgery and doesn't reflect the residual cancer cells left in the patient's brain. The aim of this study is to generate new potential targets from looking at the expression of those residual cancer cells left behind after surgery that ultimately lead to tumour recurrence.

METHOD

5-aminolevulinic acid (5-ALA) is a clinically used drug that makes cancer cells glow (fluoresce) pink, allowing more complete removal of brain tumours. 10 Tumour samples were dissociated and fluorescent activated cell sorting (FACS) was used to separate the residual fluorescent cancer cells (1%) from normal brain at the tumour's edge. Samples from 3 distinct tumour regions, core, rim and invasive margin without FACS sorting from the same patients were also included for comparison to the FACS samples. Gene expression was analysed by RNA-seq and validated by qPCR.

RESULTS

High-resolution RNA-seq profiles were generated for distinct tumour regions and the fluorescently isolated invasive cell population, allowing potential targetable abnormalities to be identified and validated.

CONCLUSION

This study shows that pure tumour cells can be separated from normal cells using fluorescence generated from tumour metabolism of 5-ALA. Further optimisation of FACS is required to increase RNA yield from FACS sorted samples. Our approach gives hope that we can interrogate the true residual disease, and for the first time gain insight into the source of tumour recurrence.

The identification and validation of novel aptamers for the diagnosis of oligodendroglioma

Abstract

Oligodendroglioma account for nearly 12% of brain tumours (Jain et al., 2017). In the United Kingdom, 0.4 people per 100 000 per year are newly diagnosed with oligodendroglioma (Crocetti et al., 2012). Early diagnosis and treatment of oligodendroglioma is essential as patients given the appropriate chemotherapy survive on average nearly 8 years longer than patients given radiotherapy alone (Cairncross et al., 2014). Aptamers are short, single-stranded nucleotide sequences (DNA or RNA) that uniquely fold to form specific recognition molecules that selectively bind to a molecular target. Aptamers against targets are generated using a process called systematic evolution of ligands by exponential enrichment (SELEX). Since 2002, a number of aptamers have been identified that can selectively identify cancerous cells. These aptamers have been used both diagnostically and therapeutically against a range of different cancers, however, there are currently no aptamers specific to glioma tissue. This first part of this project was to find aptamers specific to the grade IV glioblastoma cell line U87MG. Previous studies have found aptamers that were able to distinguish U87MG from non-malignant cell lines but showed good binding to most highly tumorigenic cell lines (Cerchia et al., 2009; Aptekar et al., 2015). To ensure that highly specific aptamers were identified, a large number of negative controls were utilised, and two variants of the SELEX process were undertaken to ascertain whether the negative controls had to be selected against individually or if they could be pooled. Patient brain tumour samples will be obtained from the Brain Tumour North West (BTNW) tissue bank and aptamers specific to oligodendroglioma will be selected. These aptamers will be used to screen patient samples to determine if they can identify oligodendroglioma tissue and be of use diagnostically, therapeutically and as a drug delivery tool.

Local Delivery Of Irinotecan To Recurrent Glioblastoma At Reoperation Offers A Safe Therapeutic Advantage Over Systemic Delivery

Abstract

Direct drug delivery to a brain tumours offers the surety of access, together with the potential to realise at least therapeutic concentrations. With previously used systemic drugs toxicity and removal mechanisms are avoided. With new formulations drug activation and duration can be explored for low local toxicity, and tumour impact with sequential imaging. In this early study Irinotecan a drug with known efficacy against glioma but with limiting systemic toxicity has been incorporated into biodegradeable hydrogel microspheres and injected into the post-surgical cavity wall in patients with recurrent glioblastoma. 10 patients with focally recurrent GBM were recruited with 9 undergoing injection with up to 3mls of microbeads in alginate suspension in up to 60 ~ 8mm injection tracks after surgical resection (100mg). Patients were assessed for immediate (72hrs) toxicity with clinical assessment and imaging. Pharmacokinetic determination of Irinotecan and SN 38 active metabolite allowed confirmation of activation and comparator with known plasma toxicity profiles. Follow-up was planned for a minimum of 6/12 for SAE but continued to allow overall survival estimation. No overt SAE were determined for this group of patients with imaging and steroid use indicating less local swelling and wound healing issues than have been demonstrated for Carmustine wafers despite early offloading. Phamacokinetic measures (SN38 plasma curves) indicate a much higher activation of Irinotecan (>90%) than expected but shorter period of exposure. Survival curves suggest clinical benefit with 4/9 patient living longer than 8 months warranting further exploration of this safe approach

Regulation of DNA double strand break repair by EGF and VEGF signalling reveals Akt to be a critical therapeutic target in glioblastoma

Abstract

Glioblastoma (GBM) is currently incurable. Its radioresistance has been attributed to a subpopulation of cells termed 'GBM stem-like cells' characterised by multipotentiality and tumorigenicity. The discrepancy between pre-clinical and clinical effects of molecular targeted agents on radiosensitivity indicates that 2D in vitro models of GBM do not recapitulate the clinical scenario. In a 3D model developed in our laboratory, EGFR inhibitors failed to enhance radiosensitivity recapitulating their lack of efficacy in the clinic, contrasting with their radiosensitising activity in 2D cultures. Conversely, inhibition of VEGF signalling caused significant radiosensitisation of 3D cultures but had no effect in 2D conditions. The critical role of the DNA damage response in mediating these effects is illustrated by the consistent correlation between radiosensitivity, unrepaired double-strand breaks (γH2AX foci), mitotic catastrophe and micronuclei in both 2D and 3D models. Further investigation revealed unrepaired DSB to be associated with delayed resolution of phosphorylated DNA-PKcs nuclear foci and reduced formation of Rad51 foci. Hence in 2D conditions, EGFR signalling appeared to promote efficient non-homologous end-joining (NHEJ) repair, while in 3D conditions this process was dependent on VEGF signalling. Motivated by previous reports that radiation induced EGFR signalling promotes NHEJ via Akt mediated phosphorylation of DNA-PKcs, we investigated the role of Akt signalling in 2D and 3D systems. Specific inhibition of Akt using the small molecule inhibitor MK-2206 or Akti knockdown increased radiation sensitivity in both 2D and 3D models to a similar extent as EGFR or VEGF inhibition respectively. In keeping with this, phosphorylation of Akt was EGFR dependent in 2D GSC cultures but VEGF dependent in the 3D model. MK-2206 induced radiosensitivity was correlated with increased unrepaired DSBs and extended mouse survival in an U87MGLuc2 orthotopic model compared to radiation only. Our data identify Akt as a promising therapeutic target in combination with radiation for GBM.

Risk factors for developing post-operative seizures following meningioma resection

Abstract

BACKGROUND

Meningiomas are benign primary tumours treated by surgical resection. Following surgery patients may develop postoperative seizures. The aim of this study was to investigate the risk factors associated with developing postoperative seizures and determine whether prophylactic antiepileptic drugs (AED) should be prescribed.

METHODS

Retrospective analysis of patients undergoing surgical resection of meningioma between 2010–2015. The following clinical factors were assessed: tumour location, grade, Simpson resection, post-operative haemorrhage/infection and use of AED prophylaxis. MRI parameters were assessed including: tumour volume, adjacent parenchymal T2 signal change, midline shift. Logistical regression was performed.

RESULTS

285 patients were identified with a median age of 60 years (range: 16–83 years). 68 (24%) had pre-operative seizures of which 62 were on AEDs and post-operatively 48 (17%) had seizures. 217 (76%) were seizure naïve, 19 were given prophylactic AEDs and post-operatively 5 had seizures. Of the 198 patients who did not receive prophylactic AED, 43 had post-operative seizures (22%). Prophylactic AED did not reduce the risk of seizures compared to no use (OR = 1.85, p= 0.260). Clinical risk factors for developing post-operative seizures were non-skull base location (n= 207, OR = 0.35, p= 0.010) and presence of post-operative haemorrhage/infection (n= 25, OR = 1.69, p= 0.002). MRI factors associated with developing post-operative seizures were presence of T2 signal intensity change greater than 66% of tumour volume (n= 81, OR = 4.71, p= 0.000).

CONCLUSION

Risk factors associated with postoperative seizure development were the non-skull base tumour location and post-operative haemorrhage/infection. The role of prophylactic AED could not be determined in this study due to insufficient sample size. Randomized control trials are required.

WHO grade III gliomas: a 3-year institutional retrospective case series analysis of the correlation between IDH-1 mutation, Ki67 and progression free survival (2012-1015)

Abstract

OBJECTIVES

To assess the correlation between Ki67 proliferation index, IDH-1 mutation status and perform a subset analysis of progression free survival in World Health Organisation (WHO) Grade III gliomas

DESIGN

A 3 year retrospective case series analysis.

SUBJECTS

Patients newly diagnosed with pathological WHO grade III glioma 2012–2015.

METHODS

Electronic interrogation of case notes in NHS Greater Glasgow & Clyde and Scottish National PACS.

RESULTS

44 patients (male:female ratio 1.2:1). Mean age was 52yrs (range: 20-74yrs). 63.6% were anaplastic astrocytomas, 18.2% anaplastoc oligodendrogliomas, 6.8% oligodendrogliomas, 9.1% were oligoastrocytoma and 2.3% were anaplastic pleomorphic xanthoastrocytoma. IDH-1 mutation occurred in 65.9% of patients. Ki67 ranged from 4–40 (IDH-1 wild type range: 4–40; IDH-1 mutated range: 6–40). Student's t-test showed no statistical significance between IDH-1 mutation status and Ki67 index (p=0.51). Progression free survival was assessed using the Kaplan-Meier method and log-rank test was used for comparison between groups defined by IDH-1 mutation status and high versus low Ki67 proliferation index (high Ki67 ≥ 20). Ki67 status was not significantly linked to progression free survival in patients with IDH-1 mutated disease (z=1.24, p=0.22); however patients with wild type IDH-1and high Ki67 had significantly shorter progression free survival than those with low Ki67 (z=2.15, p<0.05).

CONCLUSIONS

Contrary to the published literature we find no correlation between IDH-1 mutation status and Ki67 in grade III glioma. Progression free survival is not significantly correlated with Ki67 status in IDH-1 mutated disease. Patients with wild type IDH-1 and high Ki67 have significantly shorter progression free survival than those with low Ki67.

Psychometric evaluation of the Caregiver Needs Screen in neuro-oncology family caregivers

Abstract

INTRODUCTION

The informal care demands of brain tumour patients include various issues associated with neurological and cognitive symptoms. Caregiver needs questionnaires developed for cancer populations do not include these disease-specific symptoms. Therefore, we have developed the neuro-oncology Caregiver Needs Screen (CNS) and evaluated its psychometric properties.

METHODS

In-depth qualitative interviews with caregivers were used to generate items, which were reviewed by experts to add or collapse items as necessary. The resulting 32-item instrument was tested in a sample of 122 neuro-oncology caregivers, along measures of depression (Center for Epidemiological Studies – Depression scale), anxiety (Profile of Mood States – Anxiety), caregiver burden (Caregiver Reaction Assessment) and mastery (Mastery Scale). Exploratory factor analysis was used to determine internal structure. Internal consistency reliability and construct validity were assessed.

RESULTS

Six subscales were identified (neurologic symptoms; oncologic symptoms; personal communication; communicating with health care providers; resources; caregiver health). Internal consistency for the subscales ranged between α = .653 and α = .886. A floor effect was found for the personal communication subscale only. Convergent validity was verified by moderate to strong correlations between measures of caregiver wellbeing and CNS scale scores.

DISCUSSION

Findings provide preliminary evidence of reliability and validity for the CNS as a measure of neuro-oncology caregiver distress as a result of common care issues. This instrument can prove useful in research setting as well as in clinical practice when assessing caregivers' needs for supportive care.

Cognitive Rehabilitation & Support Following Cancer Treatment

Abstract

INTRODUCTION

35% of patients treated for cancer will experience cancer-related cognitive changes (CRCC). These cognitive difficulties are potentially long term affecting home life, relationships and employment; additionally, impacting significantly on quality of life, self-confidence and independence. The aim was to provide cognitive rehabilitation, enabling self-management for all those experiencing CRCC.

METHOD

To achieve our aim we:-provided training to health, social care, 3rd sector colleagues and OH departments to give guidance on the management of CRCC.-developed information resources for use in primary care and community settings, outlining cognitive rehabilitation strategies relevant to CRCC.- delivered a group-based intervention for patients presenting with more complex CRCC and their caregivers to provide psycho-education around the nature of CRCC, and supported them in the implementation of evidence-based cognitive compensatory strategies. This provided the opportunity for attendees to receive peer and professional support.The group was delivered in hospital and community-based settings ensuring optimal access.

AIMS/ OBJECTIVES

We aimed to improve awareness of CRCC in professionals throughout the care process, providing them with information resources and training to equip them to support patients experiencing CRCC. Through group intervention we aimed to improved their quality of life using validation, understanding and development of cognitive strategies.

RESULTS/ OUTCOMES

Fifty-nine individuals have attended the programme. Initial analysis indicates a reduction in distress levels associated with cognitive changes. Participants have described benefitting from the programme in helping them understand and self-manage cognitive changes post treatment. They reported that developing cognitive compensation strategies has contributed to increasing their confidence in everyday functioning.

5-ALA fluorescence based isolation of minority population GBM cells identifies putative invasion associated molecular changes

Abstract

INTRODUCTION

Glioblastoma (GBM) is a highly pleomorphic polyclonal tumour with molecular abnormalities varying temporo-spatially (intra-tumour heterogeneity), one mechanism of therapy resistance. Fluorescence guided resection (FGR) is performed with prior administration of 5-aminolevulinic acid (5-ALA) leading to individually fluorescent tumour cells mixed within a background population of non-neoplastic neural cells in the invasive region beyond the "pure" tumour. We have isolated this invasive tumour population by fluorescence activated cell sorting (FACS) to allow the study of invasive tumour cells without an overwhelming background "normal" signal. We conducted genome wide gene expression of phenotypically distinct areas of the tumour and from fluorescent/non-fluorescent cancer/non-cancer cells purified from the invasive zone respectively.

METHODS

We performed genome-wide gene expression analysis on 14 glioma samples from three different GBM patients including samples from tumour core, rim, invasive margin and GBM cells from the invasive margin that were isolated by 5-ALA assisted FACS.

RESULTS

Statistical analysis by linear models for microarray data identified 325 differentially expressed genes between FACS positive cells and other tumour regions (adjusted P-value < 0.05), of these, 50 genes were upregulated in all comparisons. These transcriptomic changes orchestrate MAPK (DUSP1, DUSP2, DUSP10 and FOSB), chemokine signalling pathways (CXCL2, CXCL3, CCL20 and NFKB1) and negative regulation of cell proliferation (EREG and KLF6). In contrast, 29 downregulated genes in FACS positive cells were enriched with signal transduction (DDR2 and MTSS1L) and ECM-receptor interaction (COL4A1, COL4A2, and HSPG2).

DISCUSSION

Residual cells responsible for GBM recurrence in the invasive zone are shown to be not only phenotypically different but also exhibiting activation of distinct molecular pathways and biological processes. These unique molecular features offer hope for developing more efficacious targeted therapies focusing on this population rather than the bulk tumour that has been the subject of most historical analyses.

High content screening of patient-derived cell lines highlights the potential of non-standard chemotherapeutic agents for the treatment of recurrent glioblastoma

Abstract

Glioblastoma Multiforme (GBM) is the most common primary brain tumour in adults, yet survival outcomes remain poor. First line treatment is well established but with disease recurrence almost inevitable, improving prognosis is challenging. With the aim of personalising therapy at recurrence we have established a high content screening (HCS) platform to analyse the sensitivity profile of seven patient-derived cancer stem cell lines to 82 FDA-approved chemotherapy drugs, with and without irradiation. Seven cancer stem cell lines were derived from patients with GBM, or GBM variants. Along with the established cell line U87-MG, each patient-derived line was cultured in tandem in serum-free conditions as adherent monolayers and three-dimensional neurospheres. Chemotherapeutics were screened at multiple concentrations and cells double-stained to observe their effect on both cell death and proliferation. Sensitivity was classified using high-throughput algorithmic image analysis. Cell line specific drug responses were observed across the seven patient-derived cell lines. Few agents were seen to have radio-sensitising effects, yet some drug classes showed a marked difference in efficacy between monolayers and neurospheres. In vivo validation of six drugs indicated that cell death readout in a three-dimensional culture scenario is a more physiologically relevant screening model, and can be used effectively to assess the chemosensitivity of patient-derived GBM lines. The study puts forward a number of non-standard chemotherapeutics that could be useful in the treatment of recurrent GBM, whilst demonstrating the potential of HCS to be used for personalised treatment based on the chemosensitivity profile of patient tumour cells.

An epigenetically controlled PML/Slit axis at the root of cell migration in both normal and neoplastic cells in the CNS

Abstract

In the central nervous system (CNS), regulation of nuclear function has been implicated in the control of cell cycle and migratory processes during neurogenesis. Alterations of these processes can lead to neoplastic transformation of neural stem cells (NSCs) and glioblastoma multiforme (GBM). The ability of GBM cells to migrate through the brain parenchyma represents a key factor underlying GBM aggressiveness and resistance to treatment. Notably, brain cancer cells use the same routes utilized by neuroblasts/immature neurons and NSCs, suggesting a neurobiological root of brain cancer migration. However, our understanding of potentially common mechanisms regulating cell migration/invasion during neurogenesis and brain tumourigenesis remains limited. Our previous work has implicated the Promyelocytic Leukaemia protein (PML), the essential component of the PML nuclear body (PML-NB), in regulation of embryonic neurogenesis via its ability to control proliferation in NSCs. We set out to investigate the role of PML in adult neurogenesis and GBM. Loss of PML leads to impaired NSC and neuroblast migration and to a smaller olfactory bulb in the adult mouse brain. A similar migration defect is observed in primary GBM cells where PML expression has been knocked down. Mechanistically, PML controls cell migration in both mouse NSCs and primary GBM cells via down-regulation of Slit genes, which are key regulators of axon guidance during development. Changes in Slits transcription upon PML kd are caused by global reduction of the repressive H3K27me3 epigenetic mark. This is associated with its redistribution to nuclear lamina-associated domains (LADs). Finally, PML controls tumor invasion and survival in an orthotopic animal model and inversely correlates with patient prognosis in GBM. Taken together, these findings support a model whereby PML-mediated modifications of chromatin structure and function regulate cell migration during normal neurogenesis and brain tumorigenesis, suggesting a neurobiological root of brain cancer invasion.

Impact and effectiveness of ioMRI and awake surgery in glioma surgery

Abstract

Despite all ours "technological bubbles", surgical removal of infiltrating brain tumours remains limited to 65%-76% in the recent large series published in the Literature. Fuzzy tumour/brain margins are the main cause of failure in reaching "complete" removals. Aggressive attempts at "supramarginal" resection often carry a risk of functional cortical and subcortical structures' damage with an unacceptable decrease in quality of life and post-operative functional status and an increase in post-operative psychological, human and financial costs for the patient, his/her carers and the NHS.Awake surgery and ioMRI (intra-operative MRI) proved to be the most effective individual aids in preventing damage to functional brain while maximizing the extent of resection.We present our series of 22 patients operated on at Southmead Hospital (North Brstol NHS Trust) in between July 2014 and Dec 2016 using both ioMRI and awake surgery (MAC, monitored anaesthesia care). Setting, patients' features, indications, type and size of tumours, and surgical times will be analysed and discussed.Using ioMRI together with awake surgery is demanding for the anaesthetic team, stuff nurse and for the patient. Theatre times are about 2 hours longer than the standard awakes, with an average stay of 10 hours. Nevertheless, post-operative patient's satisfaction together with very low morbidity and higher total resections rates (well above the average of the current published data) wipe the aforementioned concerns about ioMRI+awake feasibility in challenging neuro-oncology cases.

Epigenetic inactivation of arginine biosynthesis genes in paediatric intracranial ependymoma

Abstract

Aberrant cellular metabolism is recognized as a major event in the growth and development of many cancers and the targetting of metabolic defects in tumour cells represents a new therapeutic opportunity. For example, cells that do not express sufficient levels of argininosuccinate synthetase-1 (ASS1) or argininosuccinate lyase (ASL) become auxotrophic for arginine and require exogenous supply. Arginine deprivation using arginine deiminase (ADI-PEG20) is currently under evaluation in clinical trials for adult GBM. In this study, we investigated the arginine biosynthesis pathway in paediatric intracranial ependymoma, comprising 24 fresh frozen biopsies and 17 short-term cell cultures of low passage (<10). The methylation status of ASS1 and ASL was assessed by methylation-specific PCR and gene expression levels were measured using real-time Q-PCR analysis. The response of ependymoma cell cultures in vitro to ADI-PEG20 was determined at various time points using the sulphorodamine B (SRB) assay. Promoter hypermethylation of ASS1 was present in 41.5% ependymoma (17/41 samples) and methylation correlated with down-regulation of ASS1 expression (p<0.0001, Fisher's exact test). Importantly, methylation and expression status was maintained in 6 patient-derived cell cultures for which paired biopsies were available. Conversely, methylation of ASL was not detected in any samples. Treatment with ADI-PEG20 inhibited proliferation of ependymoma cells only in those cultures with methylation-dependent silencing of ASS1. Our findings suggest that argininine depletion therapy may benefit a significant proportion of paediatric patients with intracranial ependymoma.

3D bioprinted models of Glioblastoma

Abstract

In-vitro brain tumour models are valuable tools both to study cancer biology and for successful pre-clinical testing of anticancer drugs. However, substantial challenges remain before we can create pathologically-relevant models which closely mimic the microenvironment of human brain tumours, their growth characteristics and their response to therapies. We have developed a 3D bioprinting strategy using modified alginate matrices incorporating hyaluronan and collagen, in order to create tumour-like structures containing both glioma stem cell lines and glioma derived stromal cells. Our results to date show that the printing process can be performed with minimal effects on cell viability, even when cells are printed at high density; that cells proliferate within the gel and that rapid cell-cell adhesion can be promoted with appropriate matrices. We are able to print glioma stem cells with and without stromal cells in separate layers within tumour like 3D constructs with a smallest feature size (resolution) of approximately 300 micrometers. This capability to spatially position cell populations and other components within a 3D tumour construct allows new experimental approaches by controlling individual parameters of the tumour microenvironment and should provide novel highly-reproducible models for preclinical drug testing.

The potential of CRL4DCAF1 and KSR1 as therapeutic targets in merlin-deficient meningioma

Abstract

BACKGROUND

Merlin-deficient meningiomas are caused by mutations in the Neurofibromin 2 gene and occur in approximately 60% of sporadic meningiomas. Merlin loss is commonly associated with the genetic condition Neurofibromatosis type 2, leading to the development of multiple low grade tumours including schwannoma and meningioma. Currently, the only treatment for low grade meningioma is (radio)surgery therefore identification of novel drug targets is vital. Previous studies have shown that Kinase suppressor of Ras 1 (KSR1) is a potential therapeutic target in schwannoma and that the E3 ubiquitin ligase, CRL4DCAF1, binds to KSR1. The aim of this project is to investigate the interaction between CRL4DCAF1 and KSR1 to determine if targeting this protein complex in meningioma holds therapeutic value.

METHODS

HEK293T cells were transfected with KSR1 constructs and interactions with CRL4DCAF1 were investigated by immunoprecipitation. Immunohistochemistry, cell fractionation and Western blot were used to analyse DCAF1 and KSR1 expression and localization in primary human meningioma. A shRNA construct was used to knock down DCAF1 and APS_2_79 was used to inhibit KSR1.

RESULTS

KSR1 interacts via the N-term with DCAF1 and is ubiquitinated at the C-term in the NF+/+ model which may be DCAF1 dependent. Immunohistochemistry showed increased DCAF1 and KSR1 expression in Merlin-deficient meningiomas compared with normal meninges, whereas Western blot analysis showed variable protein expression. DCAF1 knockdown led to a reduction of nuclear pERK1/2 and a significant decrease in proliferation of meningioma cells but pERK1/2 and Cyclin D1 levels were unchanged. Combination of shDCAF1 and APS_2_79, a specific KSR1 inhibitor, reduced pERK and proliferation in both BenMen-1, a benign meningioma cell line and primary meningioma. Therefore, targeting both DCAF1 and KSR1 represents an attractive novel therapeutic strategy in meningioma.

Identifying tumour associated macrophages and microglia in an experimental glioblastoma model

Abstract

Glioblastoma is the most aggressive primary brain cancer, with 14.6 months median survival. Tumour associated macrophages and microglia (TAMM) populate upto 40% of the tumour bulk and represent potential immunotherapy targets. Characterising functional roles of TAMM subpopulations remains elusive due to limitations of existing mouse models and cross-reactive cell-surface markers. We identify a new marker-set using flow cytometry that distinguishes between peripheral macrophages and brain microglia in glioblastoma. Using non-myeloablative busulfan conditioning, we transplant mice expressing the pan leukocyte epitope CD45.1 with haematopoietic donor cells expressing congenic CD45.2, without damaging the recipient blood-brain barrier. Once 90% bone marrow chimerism is achieved at 12 weeks with no brain macrophage repopulation, we stereotactically implant GL261 murine glioblastoma cells into brain of chimeric mice, recapitulating normal tumour development. Brain flow cytometry using a 15-marker panel was then used to identify different immune cell populations and characterise TAMM subpopulations. We identify multiple immune populations migrating to tumours, and demonstrate that marker sets such as CD11b/CD45 and Siglec-H/CX3CR1, used to distinguish between microglia and macrophages, have significant co-staining from other leukocyte populations (both donor and recipient). Importantly we identify a novel 4-marker set that separates TAMM into four subpopulations with ≥ 90% donor/recipient purity, including monocytes, undifferentiated and differentiated macrophages, and microglia. In conclusion, we have developed a novel chimeric mouse model that achieves superior levels of peripheral chimeras, yet retains a homeostatic brain. Our results also highlighted a new 4-marker approach to subcategorising TAMM. RNA sequencing of TAMM subpopulations and downstream transcriptional analysis will provide the first insight into the functional role of TAMM. Further work will necessitate applying this marker set to human glioblastoma specimens, and validate our findings from an experimental glioblastoma model.

Delivery of low intensity/low frequency electric fields via clinical Deep Brain Stimulation electrodes has anti-proliferative effects on Glioblastoma multiforme cell lines

Abstract

INTRODUCTION

Phase III trials of Tumour Treating Fields (TTFields) have shown potentially positive results in both primary and recurrent adult Glioblastoma multiforme (GBM), giving hope that electric fields present a new treatment paradigm for brain tumours. Here we present investigations into the repurposing of deep brain stimulation (DBS) electrodes as a novel delivery method of therapeutic electric fields to high grade brain tumours as a localised, and internalised system.

METHODS

Medtronic DBS electrodes were inserted into cell culture flasks and delivered electric fields over a range of voltages and frequencies to a panel of primary, adult and paediatric GBM cell lines. Cell viability, cell count, cell cycle analyses, as well as combinational studies with an array of mitotic inhibitors were undertaken.

RESULTS

DBS electric fields negatively affect cell counts and viabilities of GBM cells. Cell viability was reduced by up to 70% in U87, 74% in SF188, 71% in KNS42 and 45% in GIN-5 cell lines in a voltage-dependent manner The influence of frequency on cell viability is also demonstrated. Cell counts were undertaken of treated flasks and compared across all intensities. Cell cycle analysis performed via flow cytometry reveals that electrotreatment significantly increases the proportion of cells in G0 phase relative to control flasks in GBM cells. The effects of electrotreatment in combination with mitotic inhibitors are also explored.

CONCLUSIONS

Implantable DBS electrodes have potential as a localised electric field based therapeutic strategy for the treatment of brain tumours. Electric fields delivered from DBS electrodes at significantly lower frequencies and intensities than those utilised by OptuneTM have profound anti-proliferative effects on GBM cell lines, causing measurable alterations in cell cycling. Impact: Implantable multiple DBS electrodes may address the compliance issues associated with Optune and provide a new modality for loco-regional brain tumour treatment.

Epidermal Growth Factor Receptor activity is elevated in glioma cancer stem cells and is required to maintain chemotherapy and radiation resistance

Abstract

Glioblastoma remains among the most aggressive of all human and canine malignancies, displaying high mortality rates and limited treatment options. We propose that given the similarities between canine and human gliomas, such as incidence of occurrence, histopathology, molecular characteristics, and response to therapy, that canine gliomas are a natural model of the human disease. A range of human and canine tumours have been shown to harbor specific subpopulations of cells with stem cell-like properties that initiate and maintain neoplasticity while resisting conventional therapies. Here, we show that both canine and human glioma cell lines contain a small population of cancer stem cells (CSCs), and by molecular profiling highlight the important role of the epidermal growth factor receptor (EGFR) pathway in canine CSCs. EGFR signaling is crucial in the regulation of cancer cell proliferation, migration and survival. To date EGFR-targeted interventions alone have been largely ineffective. Our findings confirm that specifically inhibiting EGFR signaling alone has no significant effect on the viability of CSCs. However inhibition of EGFR did enhance the chemo- and radio-sensitivity of both canine and human glioma CSCs, enabling this resistant, tumourigenic population of cells to be effectively targeted by conventional therapies.

Marine fungal DHICA as a UVB protectant: Assessment under in vitro and in vivo conditions

Publication date: Available online 31 January 2018
Source:Journal of Photochemistry and Photobiology B: Biology
Author(s): S.K. Shanuja, S. Iswarya, A. Gnanamani
The present study explores UVB protective role of a melanin precursor namely DHICA (5,6- Dihydroxyindole-2-carboxylic acid) expressed by the marine imperfect fungus Aspergillus nidulans. In brief, A. nidulans grown in a modified growth medium for the period of 5 days at 25 C under static conditions and the extracellular medium free from fungal biomass used for the extraction of DHICA. The extracted DHICA further exposed to partial purification and subjected to UVB protection studies using HaCaT cells and Balb/c mice independently. DHICA obtained in the present study found soluble in water. Experiments on HaCaT cell compatibility revealed nil cell death up to 500 μM concentration of DHICA. UVB protection studies under in vitro conditions emphasizes DHICA significantly protect HaCaT cells from UVB exposure by quenching the generated ROS, reducing cell apoptosis, maintain the cellular integrity and sequentially down regulating the LPO (Lipid peroxidation) and up-regulating the antioxidant enzyme (SOD (Superoxide Dismutase), Catalase, GPx (Glutathione peroxidase)) respectively. Further, experiments on cell cycle arrest analysis, gelatin zymography, and western blot analysis on COX-2 and TNF-alpha, IHC (Immunohistochemistry) on apoptotic markers (Bax, Bcl2) substantiate the protective role of DHICA. Furthermore, in vivo studies on BALB/c mice carried out and compared with the sunscreen cream with sun protective factor (SPF) of 20. Analysis of skin sections of experimental samples revealed that an appreciable reduction in the epidermal thickness of the skin samples of mice pre-exposed to DHICA followed by UVB exposure compared to UVB exposure alone. RT-PCR results on various inflammatory apoptotic markers also suggested that DHICA has UVB protective potential. The observations made in the present study explore the possible application of DHICA alone as a sun-protective agent for skin care.

New cationic and neutral copper(II) complexes containing 7-hydroxy-4-oxo-4[H]-chromene derived ONO pincer ligands: Synthesis, characterization and in vitro biological evaluations

Publication date: Available online 31 January 2018
Source:Journal of Photochemistry and Photobiology B: Biology
Author(s): G. Kalaiarasi, S. Rex Jeya Rajkumar, S. Dharani, Nigam P. Rath, R. Prabhakaran
New Schiff base ligands are prepared by the condensation of 7-hydroxy-3-formylchromone with semicarbazone and phenyl semicarbazone. The complexation of these ligands with Cu(II) ion is proposed in the light of spectral studies (IR, UV–Vis, ¹H NMR, ¹³C NMR, Mass and ESR). In the complexes 1 and 2, the ligands coordinate to the Cu(II) ion in a neutral fashion via ONO donor atoms. The single crystal XRD studies reveal a slightly distorted square-pyramidal geometry for cationic complex (1) and an octahedral geometry for neutral complex (2). Preliminary biological studies such as DNA and Protein binding are carried out by using absorption and emission titration methods. Observation of intercalative mode of binding with Calf Thymus DNA (CT-DNA) is confirmed by means of viscosity measurements. The micro-environmental changes occurring in Bovine Serum Albumin (BSA) and Human Serum Albumin (HSA) are monitored via three dimensional (3D) fluorescence studies. The compounds ability in inhibiting microbial growth is tested against different pathogens. MCF-7 (human breast cancer) and A549 (human lung carcinoma) cell lines are utilized to check the anticancer potential of the synthesized compounds by using MTT, LDH and NO assays. The results show that complexes 1 and 2 exhibited potent cytotoxic activity over standard drug cisplatin.

Graphical abstract

New Hydrogels Enriched with Antioxidants from Saffron Crocus Can Find Applications in Wound Treatment and/or Beautification

Saffron extracts have a long history of application as skin protectant, possibly due to their ability to scavenge free radicals. In this work, the performance of a hydrogel enriched with antioxidant compounds isolated from saffron crocus (Crocus sativus L.) petals was tested. These hydrogels could be considered as new drug delivery system. Hydrogels are crosslinked polymer networks that absorb large quantities of water but retain the properties of a solid, thus making ideal dressings for sensitive skin. We tested antioxidant-enriched hydrogels on primary mouse fibroblasts. Hydrogels enriched with kaempferol and crocin extracted from saffron petals showed good biocompatibility with in vitro cultured fibroblasts. These new types of hydrogels may find applications in wound treatment and/or beautification.
Skin Pharmacol Physiol 2018;31:95-98

Natural history of Kindler syndrome and propensity for skin cancer – case report and literature review

The status of dermoscopy in Germany – results of the cross-sectional Pan-Euro-Dermoscopy Study

Summary

Background

Survey on the current status of dermoscopy in Germany.

Methods

In the context of a pan-European internet-based study (n = 7,480) conducted by the International Dermoscopy Society, 880 German dermatologists were asked to answer questions with respect to their level of training as well as their use and perceived benefit of dermoscopy.

Results

Seven hundred and sixty-two (86.6 %) participants practiced dermatology in a publicly funded health care setting; 98.4 % used a dermoscope in routine clinical practice. About 93 % (n = 814) stated to have had more than five years of experience in the use of dermoscopy. Dermoscopy was considered useful in the diagnosis of melanoma by 93.6 % (n = 824); for pigmented skin tumors, by 92.4 % (n = 813); in the follow-up of melanocytic lesions, by 88.6 % (n = 780); for non-pigmented lesions, by 71.4 % (n = 628), in the follow-up of non-melanocytic lesions, by 52.7 % (n = 464); and for inflammatory skin lesions, by 28.5 % (n = 251). Overall, 86.5 % (n = 761) of participants felt that – compared to naked-eye examination – dermoscopy increased the number of melanomas diagnosed; 77,7 % (n = 684) considered the number of unnecessary excisions of benign lesions to be decreased. Participants who personally felt that dermoscopy improved their ability to diagnose melanoma were significantly i) younger, ii) had been practicing dermatology for a shorter period of time, iii) were less commonly employed by an university-affiliated dermatology department, iv) were more frequently working in an office-based public health care setting, and v) had more frequently been trained in dermoscopy during their dermatology residency.

Conclusions

The findings presented herein ought to be integrated into future residency and continuing medical education programs with the challenge to improve dermato-oncological care and to expand the diagnostic spectrum of dermoscopy to include inflammatory skin diseases.

Histoid leprosy: clinical and histopathological analysis of patients in follow-up in University Clinical Hospital of endemic country

Abstract

Background

Histoid leprosy (HL) is a rare form of lepromatous leprosy, characterized by hyperchromic indurated nodules above normal skin. Its main histopathological aspect is spindle cells. Because it may simulate other aspects, such as dermatofibroma and neurofibroma, histoid leprosy poses itself as a diagnostic challenge.

Methods

This is a retrospective study with all patients having been selected from the leprosy clinic of the Hospital das Clínicas da Universidade de São Paulo from 2006 to 2016.

Results

There were 12 patients in this study, eight in the histoid group and four in the lepromatous leprosy group. The prevalence of HL was 1.12% in all leprosy subjects. All individuals from HL group were "de novo" cases, and the histopathological analysis of skin lesions presented spindle cells generating a storiform pattern. Immunohistochemistry for CD68, vimentin, and anti-BCG were positive in all 12 cases. Factor XIIIa was visualized only in the papillary dermis, and S100 protein was negative in all biopsies. Smooth-muscle actin was present in 62.5% of the HL samples.

Conclusion

The prevalence of HL was similar to previous reports. However, all histoid patients were "de novo" cases, differing from published studies. Fusocellular macrophage transformation could be explained by the differences in cytoskeleton proteins expressed in histoid lesions in comparison to other leprosy variants, with emphasis on vimentin and smooth muscle actin.

Miescher nevus as devil's mark, but also as beauty mark

Retrospective review of 18 British South Asian women with frontal fibrosing alopecia

Boat builders' occupational contact dermatitis – 11-year data from the Finnish register of occupational diseases

Ophthalmic Manifestations of Allergic Fungal Sinusitis

Purposes: To study the ophthalmic manifestations of patients with allergic fungal sinusitis (AFS) and evaluate the importance of early diagnosis and management in preventing the possible future complications of AFS. Methods: Retrospective chart review of 100 patients with the diagnosis of AFS from a single institution was performed. Age, gender, clinical presentation including ophthalmic and radiological findings, immune status, patterns of sinus involvement, medical and surgical intervention needed, laboratory results, and the course of the disease were evaluated. Results: The mean age at presentation was 19.8 years (range, 10–42 years) with no clear gender predominance (52% of patients were female).The most common sinuses involved were ethmoid and maxillary sinuses. All patients underwent functional endoscopic sinus surgery and received systemic and topical steroids. There were no intervention-related complications. Thirty-four of 100 patients had ophthalmic consequences of AFS. The most common ophthalmic presentation was proptosis (n = 21, 61.7%), followed by epiphora (n = 5, 14.7%), visual loss (n = 4, 11.7%), diplopia (n = 3, 8.8%), and dystopia (n = 1, 2.9%) in addition to 1 patient having ptosis beside proptosis. CT scans of these 34 patients showed that 82.3% had nonhomogenous opacification of sinuses, 52% had erosion of lamina papyracea, 17.6% had intraorbital extension, and 8.8% had intracranial extension. Conclusions: Ophthalmologists may be the first who encounter these patients which necessitates familiarity with AFS presentations and complications. In cases of sudden visual loss, early intervention may prevent permanent vision loss. Functional endoscopic sinus surgery and postoperative systemic and topical steroids resulted in dramatic improvement of ophthalmic symptoms and signs. Accepted for publication November 18, 2017. The authors have no financial or conflicts of interest to disclose. Address correspondence and reprint requests to Adel H. Alsuhaibani, M.D., F.R.C.S., Department of Ophthalmology, King Abdulaziz University Hospital, King Saud University, P.O. Box 245, Riyadh 11411, Kingdom of Saudi Arabia. E-mail: adelsu@yahoo.com © 2018 by The American Society of Ophthalmic Plastic and Reconstructive Surgery, Inc., All rights reserved.

Coccidioidomycosis: Imported and Possible Domestic Cases in China A Case Report and Review, 1958-2017

Abstract

We report a case of imported pulmonary coccidioidomycosis caused by Coccidioides posadasii in a patient who was misdiagnosed as tuberculosis and mistreated with antituberculosis medications for 18 months. The symptoms were not relieved until antifungal treatment was started. An extensive review of the coccidioidomycosis cases occurring in China reveals 38 cases, 16 of which had no associated history of travel to any traditional endemic areas. We speculate that some factors may drive Coccidioides spp. transference to China, which then causes those domestic infections. Moreover, we indicate the first, to our knowledge, possible endemic areas in China.

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Clinical experience of ingenol mebutate gel for the treatment of Bowen's disease

Abstract

There are only a few anecdotal case reports about Bowen's disease (BD) treated with ingenol mebutate (IM) gel but no clinical study has been published yet. The aim of this study was to evaluate the effectiveness of IM gel in the treatment of BD and to observe the therapeutic efficacy of IM alone or IM with ablative fractional laser pretreatment. Nineteen patients with BD or squamous cell carcinoma in situ confirmed by skin biopsy were enrolled. IM was applied with 0.015% gel on facial lesions for 3 days consecutively and 0.05% gel on other sites for 2 days consecutively, with a 5-mm application margin around the visible lesion. Nine patients applied IM gel immediately following fractional CO₂ laser treatment. Two patients were lost to follow up and a total of 17 patients were enrolled. Nine patients (9/17, 52.9%) had a clinically complete response at 2 months after treatment. Among the patients treated with the fractional CO₂ laser before applying IM gel, eight (8/9, 88.9%) showed a complete response and one (1/9, 11.1%) showed partial response. Among the patients treated with IM gel alone, only one patient (1/8, 12.5%) showed a complete response, four (4/8, 50%) showed a partial response and three (3/8, 37.5%) did not respond to therapy. IM gel alone seems to have limited value for treatment of BD; however, a combination therapy with the ablative fractional laser can increase its therapeutic effectiveness.

Mild case of Hailey–Hailey disease caused by a novel ATP2C1 mutation

Detection of human papillomavirus in actinic keratosis