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1.  Characterization of bioeffects on endothelial cells under acoustic droplet vaporization 
Ultrasound in medicine & biology  2015;41(12):3241-3252.
Gas embolotherapy is achieved by locally vaporizing microdroplets through acoustic droplet vaporization (ADV), which result in bubbles that are large enough to occlude blood flow directed to tumors. Endothelial cells, lining blood vessels, can be affected by these vaporization events leading to cell injury and cell death. An idealized monolayer of endothelial cells was exposed to ADV using a 3.5 MHz transducer and dodecafluoropentane droplets. Treatments included insonation pressures that varied from 2 to 8 MPa (rarefactional), and pulse lengths that varied from 4 to 16 input cycles. The generated bubble cloud was directly dependent on pressure, but not on pulse length. Cellular damage increased with increasing bubble cloud size, but was limited to the bubble cloud area. These results suggest that vaporization near the endothelium may impact the vessel wall, an effect that could be either deleterious or beneficial depending on the intended overall therapeutic application.
doi:10.1016/j.ultrasmedbio.2015.07.019
PMCID: PMC4794981  PMID: 26403698
gas embolotherapy; acoustic droplet vaporization; endothelial cells; bioeffects
2.  Predictive modelling-based design and experiments for synthesis and spinning of bioinspired silk fibres 
Nature communications  2015;6:6892.
Scalable computational modelling tools are required to guide the rational design of complex hierarchical materials with predictable functions. Here, we utilize mesoscopic modelling, integrated with genetic block copolymer synthesis and bioinspired spinning process, to demonstrate de novo materials design that incorporates chemistry, processing and material characterization. We find that intermediate hydrophobic/hydrophilic block ratios observed in natural spider silks and longer chain lengths lead to outstanding silk fibre formation. This design by nature is based on the optimal combination of protein solubility, self-assembled aggregate size and polymer network topology. The original homogeneous network structure becomes heterogeneous after spinning, enhancing the anisotropic network connectivity along the shear flow direction. Extending beyond the classical polymer theory, with insights from the percolation network model, we illustrate the direct proportionality between network conductance and fibre Young's modulus. This integrated approach provides a general path towards de novo functional network materials with enhanced mechanical properties and beyond (optical, electrical or thermal) as we have experimentally verified.
doi:10.1038/ncomms7892
PMCID: PMC4996357  PMID: 26017575
3.  Long-term safety and tolerability of bapineuzumab in patients with Alzheimer’s disease in two phase 3 extension studies 
Background
Immunotherapy with monoclonal antibodies that target amyloid beta has been under investigation as a treatment for patients with Alzheimer’s disease (AD). The 3000 and 3001 phase 3 clinical studies of intravenous bapineuzumab assessed safety and efficacy in patients with mild to moderate AD recruited in over 26 countries. This article describes the long-term safety and tolerability of bapineuzumab in the extension studies for these two protocols.
Methods
The long-term safety and tolerability of intravenous-administered bapineuzumab in patients with AD was evaluated in apolipoprotein E ε4 allele noncarriers (Study 3002, extension of Study 3000) and apolipoprotein E ε4 allele carriers (Study 3003, extension of Study 3001). Those receiving bapineuzumab in the parent study were continued at the same dose; if receiving placebo, patients began bapineuzumab. Bapineuzumab doses were 0.5 mg/kg in both studies and also 1.0 mg/kg in the noncarrier study. Clinical efficacy of bapineuzumab was also assessed in exploratory analyses.
Results
Because of lack of efficacy in two other phase 3 trials, the parent protocols were stopped early. As a result, Studies 3002 and 3003 were also terminated. In total, 492 and 202 patients were enrolled in Studies 3003 and 3002, respectively. In apolipoprotein E ε4 carriers (Study 3003), treatment-emergent adverse events occurred in 70.7 % of the patients who originally received placebo and 66.9 % of those who originally received bapineuzumab. In noncarriers, treatment-emergent adverse events occurred in 82.1 % and 67.6 % of patients who received placebo + bapineuzumab 0.5 mg/kg and placebo + bapineuzumab 1.0 mg/kg, respectively, and in 72.7 % and 64.3 % of those who received bapineuzumab + bapineuzumab 0.5 mg/kg and 1.0 mg/kg, respectively. Amyloid-related imaging abnormalities with edema or effusions were the main bapineuzumab-associated adverse events in both studies, occurring in approximately 11 % of placebo + bapineuzumab and 4 % of bapineuzumab + bapineuzumab groups overall. Exploratory analyses of clinical efficacy were not significantly different between groups in either study.
Conclusions
In these phase 3 extension studies, intravenous bapineuzumab administered for up to approximately 3 years showed no unexpected safety signals and a safety profile consistent with previous bapineuzumab trials.
Trial registration
Noncarriers (Study 3002): ClinicalTrials.gov NCT00996918. Registered 14 October 2009.
Carriers (Study 3003): ClinicalTrials.gov NCT00998764. Registered 16 October 2009.
Electronic supplementary material
The online version of this article (doi:10.1186/s13195-016-0193-y) contains supplementary material, which is available to authorized users.
doi:10.1186/s13195-016-0193-y
PMCID: PMC4918115  PMID: 27334799
Alzheimer’s disease; Amyloid beta; Amyloid-related imaging abnormalities with edema or effusions/vasogenic edema; Bapineuzumab; Immunotherapy
4.  A Common Ancestral Mutation in CRYBB3 Identified in Multiple Consanguineous Families with Congenital Cataracts 
PLoS ONE  2016;11(6):e0157005.
Purpose
This study was performed to investigate the genetic determinants of autosomal recessive congenital cataracts in large consanguineous families.
Methods
Affected individuals underwent a detailed ophthalmological examination and slit-lamp photographs of the cataractous lenses were obtained. An aliquot of blood was collected from all participating family members and genomic DNA was extracted from white blood cells. Initially, a genome-wide scan was performed with genomic DNAs of family PKCC025 followed by exclusion analysis of our familial cohort of congenital cataracts. Protein-coding exons of CRYBB1, CRYBB2, CRYBB3, and CRYBA4 were sequenced bidirectionally. A haplotype was constructed with SNPs flanking the causal mutation for affected individuals in all four families, while the probability that the four familial cases have a common founder was estimated using EM and CHM-based algorithms. The expression of Crybb3 in the developing murine lens was investigated using TaqMan assays.
Results
The clinical and ophthalmological examinations suggested that all affected individuals had nuclear cataracts. Genome-wide linkage analysis localized the causal phenotype in family PKCC025 to chromosome 22q with statistically significant two-point logarithm of odds (LOD) scores. Subsequently, we localized three additional families, PKCC063, PKCC131, and PKCC168 to chromosome 22q. Bidirectional Sanger sequencing identified a missense variation: c.493G>C (p.Gly165Arg) in CRYBB3 that segregated with the disease phenotype in all four familial cases. This variation was not found in ethnically matched control chromosomes, the NHLBI exome variant server, or the 1000 Genomes or dbSNP databases. Interestingly, all four families harbor a unique disease haplotype that strongly suggests a common founder of the causal mutation (p<1.64E-10). We observed expression of Crybb3 in the mouse lens as early as embryonic day 15 (E15), and expression remained relatively steady throughout development.
Conclusion
Here, we report a common ancestral mutation in CRYBB3 associated with autosomal recessive congenital cataracts identified in four familial cases of Pakistani origin.
doi:10.1371/journal.pone.0157005
PMCID: PMC4915718  PMID: 27326458
5.  Quantifying visual pathway axonal and myelin loss in multiple sclerosis and neuromyelitis optica 
NeuroImage : Clinical  2016;11:743-750.
Background
The optic nerve is frequently injured in multiple sclerosis and neuromyelitis optica, resulting in visual dysfunction, which may be reflected by measures distant from the site of injury.
Objective
To determine how retinal nerve fiber layer as a measure of axonal health, and macular volume as a measure of neuronal health are related to changes in myelin water fraction in the optic radiations of multiple sclerosis and neuromyelitis optica participants with and without optic neuritis and compared to healthy controls.
Methods
12 healthy controls, 42 multiple sclerosis (16 with optic neuritis), and 10 neuromyelitis optica participants (8 with optic neuritis) were included in this study. Optical coherence tomography assessment involved measurements of the segmented macular layers (total macular, ganglion cell layer, inner plexiform layer, and inner nuclear layer volume) and paripapillary retinal nerve fiber layer thickness. The MRI protocol included a 32-echo T2-relaxation GRASE sequence. Average myelin water fraction values were calculated within the optic radiations as a measure of myelin density.
Results
Multiple sclerosis and neuromyelitis optica eyes with optic neuritis history had lower retinal nerve fiber layer thickness, total macular, ganglion cell and inner plexiform layer volumes compared to eyes without optic neuritis history and controls. Inner nuclear layer volume increased in multiple sclerosis with optic neuritis history (mean = 0.99 mm3, SD = 0.06) compared to those without (mean = 0.97 mm3, SD = 0.06; p = 0.003). Mean myelin water fraction in the optic radiations was significantly lower in demyelinating diseases (neuromyelitis optica: mean = 0.098, SD = 0.01, multiple sclerosis with optic neuritis history: mean = 0.096, SD = 0.01, multiple sclerosis without optic neuritis history: mean = 0.098, SD = 0.02; F3,55 = 3.35, p = 0.03) compared to controls. Positive correlations between MRI and optical coherence tomography measures were also apparent (retinal nerve fiber layer thickness and ganglion cell layer thickness: r = 0.25, p = 0.05, total macular volume and inner plexiform layer volume: r = 0.27, p = 0.04).
Conclusions
The relationship between reductions in OCT measures of neuro-axonal health in the anterior visual pathway and MRI-based measures of myelin health in the posterior visual pathway suggests that these measures may be linked through bidirectional axonal degeneration.
Highlights
•First study to assess relationship between segmented retinal layers and MRI in NMO•First study to use optic radiation myelin water imaging in demyelinating diseases•Inner nuclear layer thickening in MS with ON may occur independently of microcystic macular edema.•Myelin density reduction in the optic radiation observed in demyelinating diseases•Myelin loss may be due to subclinical MS disease activity in subjects without ON.•ON may lead to retinal and optic radiation pathology via bidirectional degeneration.
doi:10.1016/j.nicl.2016.05.014
PMCID: PMC4908282  PMID: 27330974
Multiple sclerosis; Neuromyelitis optica; Optical coherence tomography; Magnetic resonance imaging; Myelin water imaging; Optic neuritis
6.  Bapineuzumab for mild to moderate Alzheimer’s disease in two global, randomized, phase 3 trials 
Background
Our objective was to evaluate the efficacy (clinical and biomarker) and safety of intravenous bapineuzumab in patients with mild to moderate Alzheimer’s disease (AD).
Methods
Two of four phase 3, multicenter, randomized, double-blind, placebo-controlled, 18-month trials were conducted globally: one in apolipoprotein E ε4 carriers and another in noncarriers. Patients received bapineuzumab 0.5 mg/kg (both trials) or 1.0 mg/kg (noncarrier trial) or placebo every 13 weeks. Coprimary endpoints were change from baseline to week 78 on the 11-item Alzheimer’s Disease Assessment Scale–Cognitive subscale and the Disability Assessment for Dementia.
Results
A total of 683 and 329 patients completed the current carrier and noncarrier trials, respectively, which were terminated prematurely owing to lack of efficacy in the two other phase 3 trials of bapineuzumab in AD. The current trials showed no significant difference between bapineuzumab and placebo for the coprimary endpoints and no effect of bapineuzumab on amyloid load or cerebrospinal fluid phosphorylated tau. (Both measures were stable over time in the placebo group.) Amyloid-related imaging abnormalities with edema or effusion were confirmed as the most notable adverse event.
Conclusions
These phase 3 global trials confirmed lack of efficacy of bapineuzumab at tested doses on clinical endpoints in patients with mild to moderate AD. Some differences in the biomarker results were seen compared with the other phase 3 bapineuzumab trials. No unexpected adverse events were observed.
Trial registration
Noncarriers (3000) ClinicalTrials.gov identifier NCT00667810; registered 24 Apr 2008.
Carriers (3001) ClinicalTrials.gov identifier NCT00676143; registered 2 May 2008.
Electronic supplementary material
The online version of this article (doi:10.1186/s13195-016-0189-7) contains supplementary material, which is available to authorized users.
doi:10.1186/s13195-016-0189-7
PMCID: PMC4866415  PMID: 27176461
Alzheimer’s disease; Bapineuzumab; Immunotherapy; Amyloid β; Clinical trial; ARIA-E; Vasogenic edema
7.  Integrin Alpha-9 Mediates Lymphatic Valve Formation in Corneal Lymphangiogenesis 
Purpose
We recently reported that corneal lymphatic vessels develop integrin alpha-9 (Itga-9)-positive valves during inflammatory lymphangiogenesis. The purpose of this study was to further investigate the role of Itga-9 in corneal lymphatic valve formation in vivo and lymphatic endothelial cell (LEC) functions in vitro.
Methods
Standard murine suture placement model was used to study the effect of Itga-9 blockade on lymphatic valve formation in vivo using Itga-9 neutralizing antibody. Whole-mount corneas were harvested for immunofluorescent microscopic analysis. Additionally, human LEC culture system was used to examine the effect of Itga-9 gene knockdown on cell functions using small interfering RNAs (siRNAs).
Results
Itga-9 blockade in vivo significantly reduced the number of lymphatic valves formed in the inflamed cornea. Moreover, Itga-9 gene knockdown in human LECs suppresses cell functions of proliferation, adhesion, migration, and tube formation.
Conclusions
Itga-9 is critically involved in corneal lymphatic valve formation. Further investigation of the Itga-9 pathway may provide novel strategies to treat lymphatic-related diseases occurring both inside and outside the eye.
This study presents the first evidence showing that integrin α-9 (Itga-9) mediates inflammatory corneal lymphatic valve formation. Further investigation promises for divulging new therapies for lymphatic diseases.
doi:10.1167/iovs.15-17509
PMCID: PMC4911102  PMID: 26431485
lymphatic valve; lymphangiogenesis; inflammation; integrin alpha-9
8.  Experimental Evaluation and Computational Modeling of Tissue Damage from Low-Flow Push-Pull Perfusion Sampling In Vivo 
Background
Neurochemical monitoring via sampling probes is valuable for deciphering neurotransmission in vivo. Microdialysis is commonly used; however, the spatial resolution is poor.
New Method
Recently push-pull perfusion at low flow rates (50 nL/min) has been proposed as a method for in vivo sampling from the central nervous system. Tissue damage from such probes has not been investigated in detail. In this work, we evaluated acute tissue response to low-flow push-pull perfusion by infusing the nuclear stains Sytox Orange and Hoechst 33342 through probes implanted in the striatum for 200 min, to label damaged and total cells, respectively, in situ.
Results
Using the damaged/total labeled cell ratio as a measure of tissue damage, we found that 33 ± 8% were damaged within the dye region around a microdialysis probe. We found that low-flow push-pull perfusion probes damaged 24 ± 4% of cells in the sampling area. Flow had no effect on the number of damaged cells for low-flow push-pull perfusion. Modeling revealed that shear stress and pressure gradients generated by the flow were lower than thresholds expected to cause damage.
Comparison with existing methods
Push-pull perfusion caused less tissue damage but yielded 1500-fold better spatial resolution.
Conclusions
Push-pull perfusion at low flow rates is a viable method for sampling from the brain with potential for high temporal and spatial resolution. Tissue damage is mostly caused by probe insertion. Smaller probes may yield even lower damage.
doi:10.1016/j.jneumeth.2015.01.019
PMCID: PMC4331210  PMID: 25614385
In vivo sampling; brain tissue damage; push-pull perfusion; microdialysis; cell viability; computational modeling
9.  First-In-Human safety and long-term exposure data for AAB-003 (PF-05236812) and biomarkers after intravenous infusions of escalating doses in patients with mild to moderate Alzheimer’s disease 
Background
In the First-In-Human (FIH), 39-week, randomized, adaptive design study, safety, tolerability, pharmacokinetics and biomarkers were measured in patients with mild-to-moderate Alzheimer’s disease (AD) after infusion of a humanized monoclonal antibody to amyloid β, AAB-003 (NCT01193608; registered 19 August 2010). AAB-003 was developed by modifying bapineuzumab to reduce Fc-receptor-mediated effector function as a strategy to reduce the removal of amyloid from vessel walls associated with amyloid-related imaging abnormalities with edema/effusions (ARIA-E) without diminishing overall amyloid clearance.
Methods
Eighty-eight patients with AD received up to three infusions of AAB-003 (or placebo) 13 weeks apart at doses of 0.5, 1, 2, 4 or 8 mg/kg in the FIH trial. Dose escalation was based on safety data reviews using a Bayesian escalation algorithm. Subjects who completed the FIH study were permitted to enter a 1-year open-label extension trial with four additional intravenous infusions of AAB-003 (NCT01369225; registered 10 May 2011).
Results
Dose-dependent increases in plasma amyloid β and AAB-003 were observed. No significant changes in cerebral spinal fluid biomarkers were observed. Pharmacokinetics elimination half-life (21–28 days) clearance and volume of distribution values were consistent across dose groups indicating linearity. ARIA-E was the most notable safety finding detected by magnetic resonance imaging (MRI) at 8 mg/kg in two patients. Three cases of microhemorrhage were observed. No new safety findings or MRI abnormalities were observed for the 52 subjects who received AAB-003 in the extension trial.
Conclusion
Based on integrated review of laboratory, electrocardiogram, adverse events, and MRI, AAB-003 was safe and well tolerated up to 8 mg/kg for up to 91 weeks (FIH and extension trials) in patients with mild to moderate AD. Asymptomatic and resolvable ARIA-E was observed after the first or second infusion of AAB-003, similar to bapineuzumab. The AAB-003 dose at which ARIA-E was observed was higher compared to bapineuzumab, supporting the hypothesis that reducing Fc-receptor effector function may reduce the ARIA associated with monoclonal antibodies targeting cerebral amyloid.
Electronic supplementary material
The online version of this article (doi:10.1186/s13195-016-0177-y) contains supplementary material, which is available to authorized users.
doi:10.1186/s13195-016-0177-y
PMCID: PMC4772335  PMID: 26925577
Alzheimer's disease; Pharmacokinetics; Monoclonal antibody; Humans; Safety; Immunotherapy; ARIA
10.  Myelin Water Fraction Is Transiently Reduced after a Single Mild Traumatic Brain Injury – A Prospective Cohort Study in Collegiate Hockey Players 
PLoS ONE  2016;11(2):e0150215.
Impact-related mild traumatic brain injuries (mTBI) are a major public health concern, and remain as one of the most poorly understood injuries in the field of neuroscience. Currently, the diagnosis and management of such injuries are based largely on patient-reported symptoms. An improved understanding of the underlying pathophysiology of mTBI is urgently needed in order to develop better diagnostic and management protocols. Specifically, dynamic post-injury changes to the myelin sheath in the human brain have not been examined, despite ‘compromised white matter integrity’ often being described as a consequence of mTBI. In this preliminary cohort study, myelin water imaging was used to prospectively evaluate changes in myelin water fraction, derived from the T2 decay signal, in two varsity hockey teams (45 players) over one season of athletic competition. 11 players sustained a concussion during competition, and were scanned at 72 hours, 2 weeks, and 2 months post-injury. Results demonstrated a reduction in myelin water fraction at 2 weeks post-injury in several brain areas relative to preseason scans, including the splenium of the corpus callosum, right posterior thalamic radiation, left superior corona radiata, left superior longitudinal fasciculus, and left posterior limb of the internal capsule. Myelin water fraction recovered to pre-season values by 2 months post-injury. These results may indicate transient myelin disruption following a single mTBI, with subsequent remyelination of affected neurons. Myelin disruption was not apparent in the athletes who did not experience a concussion, despite exposure to repetitive subconcussive trauma over a season of collegiate hockey. These findings may help to explain many of the metabolic and neurological deficits observed clinically following mTBI.
doi:10.1371/journal.pone.0150215
PMCID: PMC4767387  PMID: 26913900
11.  A Prospective Pilot Investigation of Brain Volume, White Matter Hyperintensities, and Hemorrhagic Lesions after Mild Traumatic Brain Injury 
Traumatic brain injury (TBI) is among the most common neurological disorders. Hemorrhagic lesions and white matter hyperintensities (WMH) are radiological features associated with moderate and severe TBI. Brain volume reductions have also been observed during the months following injury. In concussion, no signs of injury are observed on conventional magnetic resonance imaging (MRI), which may be a true feature of concussion or merely due to the limited sensitivity of imaging techniques used so far. Moreover, it is not known whether volume reductions are due to the resolution of trauma-related edema or a true volume loss. Forty-five collegiate-level ice hockey players (20 females) and 15 controls (9 females), 40 players underwent 3-T MRI for hemorrhages [multi-echo susceptibility-weighted imaging (SWI)], WMH (three-dimensional fluid-attenuated inversion recovery), and brain volume at the beginning and the end of the hockey season. Concussed athletes underwent additional imaging and neuropsychological testing at 3 days, 2 weeks, and 2 months after injury. At the end of the hockey season, brain volume was reduced compared to controls by 0.32% (p < 0.034) in the whole cohort and by 0.26% (p < 0.09) in the concussed athletes. Two weeks and 2 months after concussion, brain volume was reduced by −0.08% (p = 0.027) and −0.23% (p = 0.035), respectively. In athletes, the WMH were significantly closer to the interface between gray matter and white matter compared to controls. No significant changes in the number of WMH over the duration of the study were found in athletes. No microhemorrhages were detected as a result of concussion or playing a season of ice hockey. We conclude that mild TBI does not lead to transient increases in brain volume and no new microbleeds or WMH are detectable after concussion. Brain volume reductions appear by 2 weeks after concussion and persist until at least 2 months after concussion. Brain volume is reduced between the beginning and the end of the ice hockey season.
doi:10.3389/fneur.2016.00011
PMCID: PMC4751255  PMID: 26903944
concussion; mTBI; MRI; neuroimaging; brain volume; susceptibility-weighted imaging; white matter hyperintensities
12.  Orientation Dependent MR Signal Decay Differentiates between People with MS, Their Asymptomatic Siblings and Unrelated Healthy Controls 
PLoS ONE  2015;10(10):e0140956.
R2* relaxometry of the brain is a quantitative magnetic resonance technique which is influenced by iron and myelin content across different brain regions. Multiple sclerosis (MS) is a common inflammatory, demyelinating disease affecting both white and grey matter regions of the CNS. Using R2*, increased iron deposition has been described in deep gray matter of MS patients. Iron accumulation might promote oxidative stress in the brain, which can lead to cell death and neurodegeneration. However, recent histological work indicates that iron may be reduced within the normal appearing white matter (WM) in MS. In the present study we analyzed the R2* signal across the white matter in 39 patients with MS, 31 asymptomatic age matched siblings of patients and 30 age-matched controls. The measurement of R2* in white matter is affected by the signal's dependency on white matter fibre orientation with respect to the main magnetic field which can be accounted using diffusion tensor imaging. We observed a clear separation of the three study groups in R2*. The values in the MS group were significantly lower compared to the siblings and controls, while the siblings group presented with significantly higher R2* values than both unrelated healthy controls and patients. Furthermore, we found significantly decreased normal-appearing white matter R2* values in patients with more severe disease course. Angle resolved analysis of R2* improves the sensitivity for detecting subtle differences in WM R2* compared to standard histogram based analyses. Our findings suggest that the decreased R2* values in MS are due to diffuse tissue damage and decreased myelin in the normal appearing and diffusely abnormal WM. The increased R2* in unaffected siblings may identify a predisposition to increased iron and the potential for oxidative stress as a risk factor for developing MS.
doi:10.1371/journal.pone.0140956
PMCID: PMC4619399  PMID: 26489078
13.  Effect of Sequence Features on Assembly of Spider Silk Block Copolymers 
Journal of structural biology  2014;186(3):412-419.
Bioengineered spider silk block copolymers were studied to understand the effect of protein chain length and sequence chemistry on the formation of secondary structure and materials assembly. Using a combination of in vitro protein design and assembly studies, we demonstrate that silk block copolymers possessing multiple repetitive units self-assemble into lamellar microstructures. Additionally, the study provides insights into the assembly behavior of spider silk block copolymers in concentrated salt solutions.
doi:10.1016/j.jsb.2014.03.004
PMCID: PMC4040161  PMID: 24613991
14.  Canadian Expert Panel Recommendations for MRI Use in MS Diagnosis and Monitoring 
Background: A definitive diagnosis of multiple sclerosis (MS), as distinct from a clinically isolated syndrome, requires one of two conditions: a second clinical attack or particular magnetic resonance imaging (MRI) findings as defined by the McDonald criteria. MRI is also important after a diagnosis is made as a means of monitoring subclinical disease activity. While a standardized protocol for diagnostic and follow-up MRI has been developed by the Consortium of Multiple Sclerosis Centres, acceptance and implementation in Canada have been suboptimal. Methods: To improve diagnosis, monitoring, and management of a clinically isolated syndrome and MS, a Canadian expert panel created consensus recommendations about the appropriate application of the 2010 McDonald criteria in routine practice, strategies to improve adherence to the standardized Consortium of Multiple Sclerosis Centres MRI protocol, and methods for ensuring effective communication among health care practitioners, in particular referring physicians, neurologists, and radiologists. Results: This article presents eight consensus statements developed by the expert panel, along with the rationale underlying the recommendations and commentaries on how to prioritize resource use within the Canadian healthcare system. Conclusions: The expert panel calls on neurologists and radiologists in Canada to incorporate the McDonald criteria, the Consortium of Multiple Sclerosis Centres MRI protocol, and other guidance given in this consensus presentation into their practices. By improving communication and general awareness of best practices for MRI use in MS diagnosis and monitoring, we can improve patient care across Canada by providing timely diagnosis, informed management decisions, and better continuity of care.
doi:10.1017/cjn.2015.24
PMCID: PMC4416356  PMID: 25896163
Magnetic resonance imaging; multiple sclerosis; multiple sclerosis MRI; neuroimaging
15.  Role of Embolization in the Treatment of Renal Masses 
Renal arterial embolization (RAE) performed for the treatment of renal masses has been proven to be a safe and effective technique, with several decades of experience. RAE is well tolerated with few complications, particularly if the time interval from embolization to surgery is reduced to less than 48 hours. Review of the literature suggests that RAE is also extremely effective for palliation of symptoms in the setting of nonoperative advanced stage renal cell carcinoma. In addition, this technique plays a large role in the management of angiomyolipomas that are symptomatic or at risk of spontaneous rupture. To date, RAE has not been evaluated in a randomized controlled setting, which has contributed to its underutilization. All of these potential benefits warrant the need for prospective studies for further validation.
doi:10.1055/s-0033-1363845
PMCID: PMC3930649  PMID: 24596442
embolization; renal cell carcinoma; angiomyolipoma; preoperative; interventional radiology
16.  Microfabrication of High-Resolution Porous Membranes for Cell Culture 
Journal of membrane science  2013;452:460-469.
Microporous membranes are widely utilized in cell biology to study cell-cell signaling and cell migration. However, the thickness and low porosity of commercial track-etched membranes limit the quality of cell imaging and the degree of cell-cell contact that can be achieved on such devices. We employ photolithography-based microfabrication to achieve porous membranes with pore diameter as small as 0.9 μm, up to 40% porosity, and less than 5% variation in pore size. Through the use of a soap release layer, membranes as thin as 1 μm can be achieved. The thin membranes minimally disrupt contrast enhancement optics, thus allowing good quality imaging of unlabeled cells under white light, unlike commercial membranes. In addition, the polymer membrane materials display low autofluorescence even after patterning, facilitating high quality fluorescence microscopy. Finally, confocal imaging suggests that substantial cell-cell contact is possible through the pores of these thin membranes. This membrane technology can enhance existing uses of porous membranes in cell biology as well as enable new types of experiments.
doi:10.1016/j.memsci.2013.11.034
PMCID: PMC3931465  PMID: 24567663
microporous; cell biology
17.  Cross-modal induction of thalamocortical potentiation leads to enhanced information processing in the auditory cortex 
Neuron  2014;81(3):664-673.
Summary
Sensory systems do not work in isolation; instead they show interactions that are specifically uncovered during sensory loss. To identify and characterize these interactions, we investigated whether visual deprivation leads to functional enhancement in primary auditory cortex (A1). We compared sound-evoked responses of A1 neurons in visually-deprived animals to those from normally-reared animals. Here we show that visual deprivation leads to improved frequency selectivity as well as increased frequency and intensity discrimination performance of A1 neurons. Furthermore, we demonstrate in vitro that in adults, visual deprivation strengthens thalamocortical (TC-) synapses in A1, but not in primary visual cortex (V1). Because deafening potentiated TC-synapses in V1 but not A1, cross-modal TC-potentiation seems to be a general property of adult cortex. Our results suggest that adults retain the capability for cross-modal changes while such capability is absent within a sensory modality. Thus multimodal training paradigms might be beneficial in sensory processing disorders.
doi:10.1016/j.neuron.2013.11.023
PMCID: PMC4023256  PMID: 24507197
18.  A microRNA-1280/JAG2 network comprises a novel biological target in high-risk medulloblastoma 
Oncotarget  2014;6(5):2709-2724.
Over-expression of PDGF receptors (PDGFRs) has been previously implicated in high-risk medulloblastoma (MB) pathogenesis. However, the exact biological functions of PDGFRα and PDGFRβ signaling in MB biology remain poorly understood. Here, we report the subgroup specific expression of PDGFRα and PDGFRβ and their associated biological pathways in MB tumors. c-MYC, a downstream target of PDGFRβ but not PDGFRα, is involved in PDGFRβ signaling associated with cell proliferation, cell death, and invasion. Concurrent inhibition of PDGFRβ and c-MYC blocks MB cell proliferation and migration synergistically. Integrated analysis of miRNA and miRNA targets regulated by both PDGFRβ and c-MYC reveals that increased expression of JAG2, a target of miR-1280, is associated with high metastatic dissemination at diagnosis and a poor outcome in MB patients. Our study may resolve the controversy on the role of PDGFRs in MB and unveils JAG2 as a key downstream effector of a PDGFRβ-driven signaling cascade and a potential therapeutic target.
PMCID: PMC4413612  PMID: 25576913
PDGFR; c-MYC; JAG2; miR-1280; medulloblastoma
19.  A high speed multifocal multiphoton fluorescence lifetime imaging microscope for live-cell FRET imaging 
Biomedical Optics Express  2015;6(2):277-296.
We demonstrate diffraction limited multiphoton imaging in a massively parallel, fully addressable time-resolved multi-beam multiphoton microscope capable of producing fluorescence lifetime images with sub-50ps temporal resolution. This imaging platform offers a significant improvement in acquisition speed over single-beam laser scanning FLIM by a factor of 64 without compromising in either the temporal or spatial resolutions of the system. We demonstrate FLIM acquisition at 500 ms with live cells expressing green fluorescent protein. The applicability of the technique to imaging protein-protein interactions in live cells is exemplified by observation of time-dependent FRET between the epidermal growth factor receptor (EGFR) and the adapter protein Grb2 following stimulation with the receptor ligand. Furthermore, ligand-dependent association of HER2-HER3 receptor tyrosine kinases was observed on a similar timescale and involved the internalisation and accumulation or receptor heterodimers within endosomes. These data demonstrate the broad applicability of this novel FLIM technique to the spatio-temporal dynamics of protein-protein interaction.
doi:10.1364/BOE.6.000277
PMCID: PMC4354599  PMID: 25780724
(180.0180) Microscopy; (170.2520) Fluorescence microscopy; (180.4315) Nonlinear microscopy; (040.1240) Arrays; (090.1970) Diffractive optics
20.  YAP Regulates Cell Proliferation, Migration, and Steroidogenesis in Adult Granulosa Cell Tumors 
Endocrine-related cancer  2014;21(2):297-310.
The Hippo signaling pathway has been implicated as a conserved regulator of organ size in both Drosophila and mammals. Yes associated protein (YAP), the central component of the Hippo signaling cascade, functions as an oncogene in several malignancies. Ovarian granulosa cell tumors (GCT) are characterized by enlargement of ovary, excess production of estrogen, high frequency of recurrence and potential of malignancy and metastasis. Whether the Hippo pathway plays a role in the pathogenesis of GCT is unknown. This study was conducted to examine the expression of YAP in human adult GCTs and to determine the role of YAP in the proliferation and steroidogenesis of GCT cells. Compared with age-matched normal human ovaries, GCT tissues exhibited higher levels of YAP expression. YAP protein was predominantly expressed in the nucleus of tumor cells, whereas the non-tumor ovarian stromal cells expressed very low levels of YAP. YAP was also expressed in cultured primary human granulosa cells and in KGN and COV434 GCT cell lines. siRNA-mediated knockdown of YAP in KGN cells resulted in a significant reduction in cell proliferation (P<0.001). Conversely, overexpression of wild-type YAP or a constitutively active YAP mutant resulted in a significant increase in KGN cell proliferation and migration. Moreover, YAP knockdown reduced FSH-induced aromatase (CYP19A1) protein expression and estrogen production in KGN cells. These results demonstrate that YAP plays an important role in regulating GCT cell proliferation, migration and steroidogenesis. Targeting the Hippo/YAP pathway may provide a novel therapeutic approach for GCT.
doi:10.1530/ERC-13-0339
PMCID: PMC4222524  PMID: 24389730
Hippo/YAP pathway; granulosa cell tumor; cell proliferation; steroidogenesis; migration
21.  Virtually No Thoracic Lesion Inaccessible: A Pictorial Case Review 
Access route considerations in percutaneous intrathoracic biopsy or ablation offers its own unique set of challenges, with special consideration toward reducing the rate of pneumothorax. This review highlights several novel and atypical methods to improve access to intrathoracic lesions through a series of representative cases. These methods include patient positioning, curved needles, hydrodissection, induced/artificial pneumothorax, and use of specialized equipment functions. No intrathoracic lesion should be considered “inaccessible” either for biopsy or treatment by percutaneous approaches without consideration of performing these adjunctive techniques.
doi:10.1055/s-0033-1342963
PMCID: PMC3709986  PMID: 24436538
biopsy; ablation; lung cancer; thoracic; technique; interventional radiology
22.  Effects of Base Polymer Hydrophobicity and End Group Modification on Polymeric Gene Delivery 
Biomacromolecules  2011;12(10):3592-3600.
A new 320 member polymer library of end-modified poly(beta amino) esters was synthesized. This library was chosen such that small differences to the structures of component backbone, side-chain, and end-group monomers could be systematically and simultaneously evaluated. The in vitro transfection efficacy and cytotoxicity of DNA nanoparticles formed from this library was assessed. This library approach enabled us not only to synthesize and test a large variety of structures rapidly, but provided us with a robust dataset to analyze for the effect of small structural permutations to polymer chain structure. Small changes to the side chains, backbones, and end groups within this polymer library produced dramatic results, with transfection efficacy of CMV-Luc varying over 4-orders in a 96-well plate format. Increasing hydrophobicity of the base polymer backbone and side chain tended to increase transfection efficacy, but the most hydrophobic side chains and backbones showed the least requirement for a hydrophobic pair. Optimal PBAE formulations were superior to commercially available non-viral alternatives FuGENE® HD and Lipofectamine™ 2000, enabling ~3-fold increased luminescence (2.2×106 RLU/well vs 8.1×105 RLU/well) and 2-fold increased transfection percentage (76.7% vs 42.9%) as measured by flow cytometry with comparable or reduced toxicity.
doi:10.1021/bm200807s
PMCID: PMC3959121  PMID: 21888340
Non-viral; Gene delivery; Biodegradable polymer; Combinatorial library; Hydrophobicity
23.  Magnetic resonance frequency shifts during acute MS lesion formation 
Neurology  2013;81(3):211-218.
Objective:
We investigated the evolution of new multiple sclerosis (MS) lesions over time using frequency shifts of the magnetic resonance (MR) signal.
Methods:
Twenty patients with relapsing-remitting MS were serially scanned for 6 months at 1-month intervals. Maps of MR frequency shifts were acquired using susceptibility-weighted imaging. New lesions were identified by enhancement with gadolinium (Gd).
Results:
Forty new lesions were identified as areas of signal increase on Gd-enhanced scans. Up to 3 months before lesion appearance, the frequency in areas of future Gd enhancement was not detectably different from the frequency in normal-appearing white matter. Rapid increase in MR frequency was observed between 1 month before and 1 month after Gd enhancement. Two months postenhancement and later, the frequency stabilized and remained at a constantly increased level.
Conclusions:
These findings suggest that an increase in MR frequency does not simply reflect blood-brain barrier disruption or edema; rather, it reflects a change of tissue architecture as a consequence of new lesion formation. The data demonstrate that the MR frequency of focal MS lesions is increased before the lesions appear on conventional MRI. Unlike many other advanced imaging techniques, the images for frequency mapping can be rapidly acquired at high spatial resolution and standardized on most clinical scanners.
doi:10.1212/WNL.0b013e31829bfd63
PMCID: PMC3770162  PMID: 23761621
24.  AIPL1 implicated in the pathogenesis of two cases of autosomal recessive retinal degeneration 
Molecular Vision  2014;20:1-14.
Purpose
To localize and identify the gene and mutations causing autosomal recessive retinal dystrophy in two consanguineous Pakistani families.
Methods
Consanguineous families from Pakistan were ascertained to be affected with autosomal recessive retinal degeneration. All affected individuals underwent thorough ophthalmologic examinations. Blood samples were collected, and genomic DNA was extracted using a salting out procedure. Genotyping was performed using microsatellite markers spaced at approximately 10 cM intervals. Two-point linkage analysis was performed with the lod score method. Direct DNA sequencing of amplified genomic DNA was performed for mutation screening of candidate genes.
Results
Genome-wide linkage scans yielded a lod score of 3.05 at θ=0 for D17S1832 and 3.82 at θ=0 for D17S938, localizing the disease gene to a 12.22 cM (6.64 Mb) region flanked by D17S1828 and D17S1852 for family 61032 and family 61227, which contains aryl hydrocarbon receptor interacting protein-like 1 (AIPL1), a gene previously implicated in recessive Leber congenital amaurosis and autosomal dominant cone-rod dystrophy. Sequencing of AIPL1 showed a homozygous c.773G>C (p.Arg258Pro) sequence change in all affected individuals of family 61032 and a homozygous c.465G>T (p.(H93_Q155del)) change in all affected members of family 61227.
Conclusions
The results strongly suggest that the c.773G>C (p.R258P) and c.465G>T (p.(H93_Q155del)) mutations in AIPL1 cause autosomal recessive retinal degeneration in these consanguineous Pakistani families.
PMCID: PMC3888496  PMID: 24426771
25.  Stromal modulation of bladder cancer-initiating cells in a subcutaneous tumor model 
The development of new cancer therapeutics would benefit from incorporating efficient tumor models that mimic human disease. We have developed a subcutaneous bladder tumor regeneration system that recapitulates primary human bladder tumor architecture by recombining benign human fetal bladder stromal cells with SW780 bladder carcinoma cells. As a first step, SW780 cells were seeded in ultra low attachment cultures in order to select for sphere-forming cells, the putative cancer stem cell (CSC) phenotype. Spheroids were combined with primary human fetal stromal cells or vehicle control and injected subcutaneously with Matrigel into NSG mice. SW780 bladder tumors that formed in the presence of stroma showed accelerated growth, muscle invasion, epithelial to mesenchymal transition (EMT), decreased differentiation, and greater activation of growth pathways compared to tumors formed in the absence of fetal stroma. Tumors grown with stroma also demonstrated a greater similarity to typical malignant bladder architecture, including the formation of papillary structures. In an effort to determine if cancer cells from primary tumors could form similar structures in vivo using this recombinatorial approach, putative CSCs, sorted based on the CD44+CD49f+ antigenic profile, were collected and recombined with fetal bladder stromal cells and Matrigel prior to subcutaneous implantation. Retrieved grafts contained tumors that exhibited the same structure as the original primary human tumor. Primary bladder tumor regeneration using human fetal bladder stroma may help elucidate the influences of stroma on tumor growth and development, as well as provide an efficient and accessible system for therapeutic testing.
PMCID: PMC3512189  PMID: 23226620
Bladder cancer; cancer stem cell (CSC); subcutaneous tumor model; stroma; sphere

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