Canada Research Chair Research Articles

Implementation Research on Shared Decision Making in Primary Care: Inventory of Intracluster Correlation Coefficients.

Background. Cluster randomized trials are important sources of information on evidence-based practices in primary care. However, there are few sources of intracluster correlation coefficients (ICCs) for designing such trials. We inventoried ICC estimates for shared decision-making (SDM) measures in primary care. Methods. Data sources were studies led by the Canada Research Chair in Shared Decision Making and Knowledge Transition. Eligible studies were conducted in primary care, included at least 2 hierarchical levels, included SDM measures for individual units nested under any type of cluster (area, clinic, or provider), and were approved by an ethics committee. We classified measures into decision antecedents, decision processes, and decision outcomes. We used Bayesian random-effect models to estimate mode ICCs and the 95% highest probability density interval (HPDI). We summarized estimates by calculating median and interquartile range (IQR). Results. Six of 14 studies were included. There were 97 ICC estimates for 17 measures. ICC estimates ranged from 0 to 0.5 (median, 0.03; IRQ, 0-0.07). They were higher for process measures (median, 0.03; IQR, 0-0.07) than for antecedent measures (0.02; 0-0.07) or outcome measures (0.02; 0-0.06), for which, respectively, "decisional conflict" (mode, 0.48; 95% HPDI, 0.39-0.57), "reluctance to disclose uncertainty to patients" (0.5; 0.11-0.89), and "quality of the decision" (0.45; 0.14-0.84) had the highest ICCs. ICCs for provider-level clustering (median, 0.06; IQR, 0-0.13) were higher than for other levels. Limitations. This convenience sample of studies may not reflect all potential ICC ranges for primary care SDM measures. Conclusions. Our inventory of ICC estimates for SDM measures in primary care will improve the ease and accuracy of power calculations in cluster randomized trials and inspire its further expansion in SDM contexts.

A Global Look at Mercury Exposures: Supporting the Goals of the Minamata Convention.

A Global Look at Mercury Exposures: Supporting the Goals of the Minamata Convention.

Impact of a Free Health Care Policy in the Democratic Republic of the Congo During an Ebola Outbreak: An Interrupted Time-Series Analysis

Background: During past outbreaks of Ebola Virus Disease (EVD) and other infectious diseases, health service utilization declined among the general public, delaying health seeking behavior and affecting population health. In 2018, the Democratic Republic of Congo (DRC) announced a free health care policy during an EVD outbreak in affected and neighboring areas. We evaluated the impact of this policy on health service utilization. Methods: Using monthly data from the national Health Management Information System from January 2017 to January 2019, we examined rates of the use of nine health services at primary health facilities: total visits; first and fourth antenatal care visits; institutional deliveries; postnatal care visits; DTP vaccinations; and visits for uncomplicated malaria, pneumonia, and diarrhea. We used controlled interrupted time series analysis with a mixed effects model to estimate changes in the rates of services use during the policy (June -September 2018) and afterward. Findings: Use of most services increased, including EVD affected zones. Total visits and visits for pneumonia and diarrhea increased more than two-fold relative to the control areas (p<0·001), while institutional deliveries and first antenatal care increased between 20-50% (p<0·01). Visits for DTP, fourth antenatal care visits and postnatal care visits were not significantly affected. Most increases did not persist after the policy ended. Interpretation: The free care policy was effective at rapidly increasing the use of health services, but this effect was not sustained post-FCP. Such policies may mitigate the impact of infectious disease outbreaks on health service utilization. Funding Statement: This project received funding from the Rapid Research Fund for Ebola supported by the International Development Research Centre, Social Sciences and Humanities Research Council, and Canadian Institutes of Health Research, and AXA Research Fund. Dr. Law received salary support through a Canada Research Chair and a Michael Smith Foundation for Health Research Scholar Award. Dr. Grepin also received salary support through a Canada Research Chair. Declaration of Interests: Dr. Law has consulted for Health Canada, the Health Employees’ Union, the Conference Board of Canada, and provided expert witness testimony for the Attorney General of Canada. All other authors report no potential conflicts of interest. Ethics Approval Statement: The research protocol was approved by the Ethics Committees at Wilfrid Laurier University and Kinshasa School of Public Health.

240-LB: Angiotensin-Converting Enzyme and Type 2 Diabetes Risk: A Mendelian Randomization Study

The causal relationship between angiotensin-converting enzyme (ACE) inhibition and protection from type 2 diabetes (T2D) remains uncertain. While three large randomized control trials (RCTs) suggested a protective effect of ACE inhibitors compared to placebo, three other RCTs failed to observe an effect, including the only trial dedicated to this question. We hypothesized that ACE concentration-lowering genetic variants could be used to infer the pharmacological effect of ACE inhibitors on T2D risk using a Mendelian Randomization (MR) approach. We first assessed the association between T2D prevalence and ACE serum level in the Outcome Reduction with Initial Glargine Intervention (ORIGIN) trial (N=8,197). We next investigated whether genetically lowered ACE level was causally linked to the risk of T2D using (i) a two-sample MR analysis applied to the DIAbetes Genetics Replication And Meta-analysis consortium (n=26,676 T2D cases; 132,532 controls), and (ii) an ACE concentration-lowering genetic risk score in the UK Biobank cohort (N=341,872). We then compared the genetically determined effect of lower ACE level on T2D risk to the pharmacological inhibition of ACE vs. placebo, which was evaluated through a meta-analysis of six RCTs (N=31,200). Lower ACE serum level was associated with reduced T2D prevalence (OR, 0.89; 95% CI, 0.82-0.96; P=3.50x10-3) in the ORIGIN trial. Through MR analyses, a 1 SD lower genetically determined ACE level predicted a lower risk of T2D (OR, 0.92; 95% CI, 0.89-0.95; P=1.79x10-7). This result was replicated in the UK Biobank (OR, 0.97; 95% CI, 0.96-0.99, P=8.73x10-4). RCT meta-analysis showed a reduction in T2D incidence on ACE inhibitors vs. placebo (OR, 0.76; 95% CI, 0.60-0.97; P=2.59x10-2), which was consistent with the findings from MR analyses (P for comparison=0.95). Our results support a protective effect of long-term ACE inhibition on T2D risk and suggest to consider patient's risk for T2D when prescribing blood-pressure lowering drugs. Disclosure M. Pigeyre: None. J. Sjaarda: None. M. Chong: None. S. Hess: None. J. Bosch: Advisory Panel; Self; Bayer AG. S. Yusuf: None. H. Gerstein: Advisory Panel; Self; Abbott, AstraZeneca, Boehringer Ingelheim International GmbH, Eli Lilly and Company, Janssen Pharmaceuticals, Inc., Merck &amp; Co., Inc., Novo Nordisk A/S, Sanofi. Research Support; Self; AstraZeneca, Eli Lilly and Company, Merck &amp; Co., Inc., Novo Nordisk A/S, Sanofi. Other Relationship; Self; AstraZeneca, Boehringer Ingelheim International GmbH, Eli Lilly and Company, Sanofi. G. Pare: Consultant; Self; Amgen Inc., Bristol-Myers Squibb Company, Lexicomp, Sanofi. Research Support; Self; Canada Research Chair in Genetic and Molecular Epidemiology, CISCO Professorship in Integrated Health Biosystems, Sanofi. Funding Sanofi; Canadian Institutes of Health Research

The Jejunum and Ileum Mediate Increased Calcium Absorption for Bone Mineralization During Postnatal Development via TRPV6 and Cav1.3 (OR26-07-19)

ObjectivesIntestinal Ca2+ absorption early in life is vital to achieving optimal bone mineralization. The molecular details of intestinal Ca2+ absorption have been defined in adults, after peak bone mass has been reached, but are largely unexplored during development. We sought to delineate the molecular details of transcellular Ca2+ absorption across the small intestine which facilitate a positive calcium balance during growth. MethodsWe used wildtype, Cav1.3 knockout and Trpv6 mutant mice. Expression of small intestinal and renal calcium transport genes was assessed using quantitative PCR. Net transcellular 45-calcium flux across intestinal segments was measured in Ussing chambers. Femurs we analyzed using micro-CT and histology. ResultsSignificant TRPV6 mediated Ca2+ flux across the duodenum was absent in pre-weaned (P14) mice but occurred post-weaning. In contrast, we found significant transcellular Ca2+ absorption in the jejunum and ileum at P14 but not 2 months. TRPV6 and Cav1.3 are necessary for this jejunal absorption and Cav1.3 appears to mediate absorption across the ileum although compensation is present in knockout pups. Knockout of Cav1.3 induces a compensatory increase in renal Ca2+ reabsorption in P14 mice although these pups have increased growth plate thickness suggesting delayed bone mineralization. ConclusionsThis work provides molecular details of how the small intestine facilitates increased demand for Ca2+ early in life to meet the requirements of growth and highlights the complexity of the multiple mechanisms involved in achieving a positive Ca2+ balance. Funding SourcesThis work is funded by grants from the Women and Children’s Health Research Institute, supported by the Stollery Children’s Hospital Foundation, and the National Sciences and Engineering Research Council to RTA, who is the Canada Research Chair in Renal Epithelial Transport Physiology. MRB is supported by a Vanier Canada Graduate Scholarship, Alberta Innovates Clinician Fellowship and an NSERC Michael Smith Foreign Study Supplement. H. Dimke is funded by the Danish Medical Research Council. Work at UdS was funded by Deutsche Forschungsgemeinschaft (DFG) by IRTG1830 (to JE, VF), Sonderforschungsbereich (SFB) 894 (to JE, PW) and SFB TRR152 (to VF).

254-OR: Novel Biomarkers Predicting Renal Dysfunction in People with Dysglycemia in the ORIGIN Trial

Diabetes is the most common cause of renal failure accounting for more than 50% of cases. Whereas HbA1c, age and albuminuria are independent risk factors for renal decline, adding novel serum biomarkers to routine clinical risk factors may help identify patients at high risk for renal decline. One ml of stored serum from 7,482 ORIGIN participants with a recorded eGFR at baseline and either the 2 year and/or final visit was assayed for 237 biomarkers using Myriad RBM’s customized human discovery multi-analyte profile 250+ panel, and hs troponin and anti-GAD biomarkers were measured locally. During a median follow-up of 6.2 years, 1,646 (22%) developed the renal outcome comprising renal failure, albuminuria regression or doubling of some creatinine. A linear mixed model using a forward selection approach was used to identify the subset of biomarkers that independently predicted a decline in eGFR after accounting for 9 clinical risk factors for renal decline including age, sex, smoking, body mass index, mean arterial pressure, log (albumin:creatinine), cholesterol, baseline eGFR and HbA1c. A Bonferroni-corrected P value for inclusion in the model of less than 0.00021 (i.e., 0.05/239) identified 13 significant and independent biomarkers that included: alpha 1 microglobulin (beta = -0.157); NT-pro BNP (beta = -0.081); IGF binding protein 4 (beta = -0.110); growth differentiation factor 15 (beta = -0.091); RAGE (beta = -0.052); myoglobin (beta = -0.073); growth regulated alpha protein (beta = 0.058); fibulin-1C (beta = -0.048); apolipoprotein A-4 (beta = -0.057); apolipoprotein A-2 (beta = 0.065); fas ligand (beta = -0.099); eotaxin E (beta = -0.052); and beta amyloid 1-40 (beta = -0.057). The addition of these selected biomarkers to the clinical variables improved the variance of the eGFR from 0.053 to 0.155 (difference in log likelihood = 849, P &amp;lt; 0.001). These novel biomarkers are potential therapeutic targets. They can also help identify dysglycemic people at risk for renal decline. (NCT00069784) Disclosure H. Gerstein: Advisory Panel; Self; Abbott, AstraZeneca, Boehringer Ingelheim International GmbH, Eli Lilly and Company, Janssen Pharmaceuticals, Inc., Merck &amp; Co., Inc., Novo Nordisk A/S, Sanofi. Research Support; Self; AstraZeneca, Eli Lilly and Company, Merck &amp; Co., Inc., Novo Nordisk A/S, Sanofi. Other Relationship; Self; AstraZeneca, Boehringer Ingelheim International GmbH, Eli Lilly and Company, Sanofi. G. Pare: Consultant; Self; Amgen Inc., Bristol-Myers Squibb Company, Lexicomp, Sanofi. Research Support; Self; Canada Research Chair in Genetic and Molecular Epidemiology, CISCO Professorship in Integrated Health Biosystems, Sanofi. M.J. McQueen: None. S. Lee: None. A. Kannt: Employee; Self; Sanofi. S. Hess: None. Funding Sanofi; Innovative Medicines Initiative (115974); European Union; European Federation of Pharmaceutical Industries; JDRF

Biomarkers of endothelial activation/dysfunction distinguish sub-groups of Ugandan patients with sepsis and differing mortality risks.

Sepsis is a complex clinical syndrome with substantial heterogeneity. We sought to identify patterns of serum biomarkers of endothelial activation and dysfunction in individuals with sepsis and evaluate subgroup-specific differences in mortality. Adult patients with sepsis (n=426) were consecutively recruited from two hospitals in Uganda. Clinical information was collected and serum concentrations of eleven biomarkers involved in the endothelial response to infection were measured in samples from 315 patients. Latent variable models were fit to evaluate whether the endothelial response to sepsis consists of one unified biological process or multiple processes and to identify subgroups of patients with distinct host-response profiles. Differences in survival at day 28 were evaluated using Kaplan-Meier survival curves. We identified three patient subgroups characterized by unique host endothelial response profiles. Patients fitting Profile 2 had significantly worse survival (log-rank p<0.001). Four latent factors (Factor 1-4) were identified, each potentially representing distinct biological processes for the endothelial response to sepsis: Factor 1 (CHI3L1, sTREM1, sFLT1); Factor 2 (ANGPT1, PF4, VEGF); Factor 3 (CXCL10, VWF, sICAM1); and Factor 4 (ANGPT2, sTEK). Patient profiles based on patterns of circulating biomarkers of endothelial responses may provide a clinically meaningful way to categorize patients into homogeneous subgroups and may identify patients with a high risk of mortality. Profile 2 may represent dysfunction of the endothelial response to infection. Primary funding: Investigator-Initiated Award provided by Pfizer, Inc (WMS, STJ). Additional support: Canadian Institutes of Health Research (CIHR) Foundation grant (KCK; FDN-148439) and the Canada Research Chair program (KCK).

(Invited) Evolutionary Parameter Extraction for Organic TFT Compact Models Including Contact Effects

The development of accurate and computationally efficient models is critical to reduce the cycle time between design, manufacturing and characterization of electronic devices, circuits and systems. The models must include degrading effects in order to better describe the performance of manufactured device. This is the case of contact effects in the modelling of organic thin film transistors (OTFTs). In fact, much effort was made to include contact effects into compact models. In this work, we consider a generic analytical model for the current-voltage (ID-VD) characteristics of OTFTs, valid for all the operation regions of the transistor, including the subthreshold region [1]. This model was later redefined with the inclusion of a model for the current-voltage (ID-VC) curves of the contact region, and a parameter extraction procedure, in which a sequence of iterative steps is carried out until a good agreement between experimental and simulated current voltage curves is obtained [2]. The parameter extraction procedure was accelerated in [3] with the proposal of an evolutionary procedure to extract the parameters that best fit experimental current-voltage characteristics. Later in [4], improvements over this evolutionary procedure were presented. Due to the explosion of different applications of OTFTs such as phototransistors and a wide variety of physical and chemical sensors, novel materials, contacts, and designs for transistor were proposed and used. For these applications, more accurate models and better extraction parameter procedures are needed. These new structures impose more requirements to the extraction methods and models, and therefore to the extracted parameters. From this point of view, the evolutionary extraction procedures proposed in [3, 4] are a good option to be used in combination with the above mentioned generic analytical model for the current-voltage (ID-VD) characteristics of OTFTs [1, 2]. In order to adapt both the compact model and the extraction procedure to new applications of the OTFTs, rules are added in form of optimization objectives and constrains for the different parameters. In this work, we present such rules applied to different sets of thin film phototransistors and sensors. In the first place, we consider the multi-objective evolutionary algorithm (MOEA), NSGA-II [5]. It was used in [3, 4], and seeks values of the parameter in the compact model that best meet some user defined objectives. In [3], two objectives were optimized, while in [4], four objectives were optimized, along with some defined constraints. It is known that classic MOEAs, such as the NSGA-II, have serious limitations when coping with more than three objectives. These problems are referred as many-objective optimization problems (MaOPs). Among the limitations of MOEAs to treat MaOPs are the selection operators, computational cost, visualization of the Pareto optimal front (POF), and more importantly, the convergence to an optimal solution. Recently, a many-objective implementation of the NSGA-II, the NSGA-III, was released [5]. NSGA-III was designed to specially deal with MaOPs, incorporating different operators to the ones used by its predecessor. Thus, in a second part of this work, we substitute the NSGA-II algorithm with the NSGA-III one for the characterization of the same sets of OTFTs. Finally, the results of using both algorithms are compared and will be discussed. Acknowledgments This work was supported by projects MAT2016-76892-C3-3-R and TIN2015-67020-P funded by the Spanish Government, European Regional Development Funds (ERDF) and the Canada Research Chair Program.

Wood Smoke Disrupts Alveolar Epithelial Barrier Function through a p44/42 MAPK Signaling Pathway

BackgroundWood smoke (WS) contains many of the same toxic compounds as those found in cigarette smoke (CS). Although CS is a leading cause of respiratory disease, limited studies investigating the mechanisms involved in WS mediated respiratory disease have been undertaken. Exposure to WS particulate reduces epithelial cell viability and barrier function. Both CS and WS induce MAPK signaling and contribute to airway remodeling. With an increase in the size, duration, and number of forest fires annually, there is a need for a better understanding with regards to the acute and chronic effects of WS on respiratory health. Therefore, we undertook studies to evaluate the role of WS exposure on the function and remodeling of the alveolar epithelium.MethodsBirch wood was combusted and the smoke bubbled into PBS. The pH pf the resulting solution was adjusted to 7.1 and was considered 100% WS solution. Monolayers of A549 alveolar epithelial cells were exposed to increasing concentrations of WS for up to 48 hours. Cell viability was determined using MTT and trypan blue exclusion. Epithelial barrier function was monitored via electrical cell‐substrate impedance sensing (ECIS). Adhesion junctional structure was observed by immunofluorescent staining for E‐cadherin. Western blot was used to assess activation of MAPK signaling pathways.ResultsConcentrations of WS up to 10% had no effect on cell viability. WS promoted a concentration dependent reduction in epithelial barrier function after 24 hours which was maintained out to 48 hours. Cells not exposed to WS demonstrated distinct cobblestone E‐cadherin staining. Exposure to 2.5% WS diminished the cobblestone patterning after 24 hours and was completely lost after exposure to 5.0% WS. The wound healing capacity of cells after WS exposure was assessed using a scratch assay. After 48 hours, cells not exposed to WS had closed the scratched area by &gt;50% whereas those cells exposed to 10% WS had closed the area by &lt;20%. Exposure to WS induced phosphorylation of the p44/42, but not p38, MAPK signaling pathway. Pre‐treatment with the MAPK inhibitor U0126 prevented WS mediated epithelial barrier dysfunction. Additionally, U0126 preserved E‐cadherin cobblestone patterning even after WS exposure and had no effect on cell viability.ConclusionsExposure to WS disrupts alveolar epithelial adhesion that is partially regulated through the p44/42 MAPK signaling pathway. Pharmacological inhibition of p44/42 phosphorylation preserved epithelial barrier function, which may be as a result of maintained E‐cadherin‐mediated cell adhesion.Support or Funding InformationGrant in aide funding was provided by provided CIHR (DJG). MRZ and CTT are recipients of CIHR Fellowships. MRZ also received a MSFHR Trainee Award. CC is a Canada Research Chair and holds the Astra‐Zeneca Chair in Occupational and Environmental Lung Disease at the University of British Columbia.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Bacteria degrade the specialized basal lamina attaches the gingiva to teeth

IntroductionThe junctional epithelium is a specialized portion of the gingiva that seals off the tooth supporting tissues from the oral environment. This relationship is achieved via a specialized basal lamina (sBL). Three unique proteins ‐AMTN, ODAM and SCPPPQ1‐ together with Laminin‐332 structure the supramolecular organization of this sBL and determine its adhesive capacity. Despite its critical and strategic importance and continued exposure to bacteria, little is known on the susceptibility of the sBL to bacterial activity.MethodsThe objective of this study was to evaluate the impact of various oral bacteria on the sBL using molecular biology and complementary imaging approaches.ResultsSDS‐PAGE and western blot analysis of degradation assays with trypsin like proteases as well as incubation with Porphyromonas gingivalis (P. gingivalis) revealed that all sBL constituents, except SCPPPQ1, were rapidly degraded. Mass spectrometry also confirmed that SCPPPQ1 was not degraded. We have previously shown that mixing the recombinant sBL proteins results in the formation a supramolecular network that mimicked the sBL. To observe the impact of oral bacteria on this ‘reconstituted’ sBL network, we exposed it to P. gingivalis and observed the results by atomic force microscopy and scanning electron microscopy (SEM). The bacteria destroyed the supramolecular network in their vicinity creating a peripheral space around them. SEM imaging further showed the presence of fine filamentous residual matrix on the exposed HOPG surface, suggesting the incomplete degradation of some component and a layered destruction of the network. Finally, to determine the effect of P. gingivalis on the native sBL itself, we exposed it ex‐vivo to bacteria. After two hours of exposure, SEM analysis revealed the presence of a shallow depression around the bacteria and after 6 h, the affected peri‐bacterial area became deeper and cavitated , suggesting active degradation of the native sBL as well.ConclusionWe show here for the first time that enzymes and P. gingivalis, one of the major bacteria implicated in chronic periodontitis, can attack the individual components of the sBL as well as alter the supramolecular organization of this critical adhesive extracellular matrix. Because destruction of the sBL would open the door to bacterial infiltration around teeth. These results highlight the importance of the sBL in PD and open the door for new treatment paradigms.Support or Funding InformationThis work was supported by CIHR, Canada Research Chair, RSBO, and Shriners Hospital.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Muscle Sympathetic Activity Kinetics during One‐leg Cycling in Men and Women with and without Heart Failure: Evidence for Preserved Cardiopulmonary Baroreflex Sympathoinhibition

Microneurographic recordings of muscle sympathetic nerve activity (MSNA) are generally analyzed over discrete epochs, usually in minutes. This is sufficient for determining resting sympathetic outflow but may obscure physiologically relevant information regarding temporal changes during an acute stress, such as exercise. To address this limitation, we applied a moving average to examine continuously the effects of one‐leg cycling on MSNA in 22 participants with (n=11) and without (n=11) heart failure with reduced ejection fraction (HFrEF). MSNA burst frequency and incidence were acquired over an 8 minute period: 2 minutes of baseline, zeroload cycling, moderate intensity cycling (50% of VO2 peak), and recovery. MSNA was analyzed in 30 second epochs, advancing in five‐second increments to create a 91 point overlapping moving average plot. Area under the curve analysis demonstrated that both MSNA burst frequency and incidence were higher in HFrEF than controls during zeroload and moderate intensity cycling (both, p&lt;0.05). To estimate the reflex sympathoinhibitory consequences of skeletal muscle‐pump induced increases in cardiopulmonary blood volume at the start of exercise, we calculated the nadir reduction in MSNA burst frequency and incidence during the first 30 seconds of zeroload (HFrEF vs. controls: −6±7 vs. −7±4 bursts/min; −12±11 vs. −15±5 bursts/100 heartbeats) and moderate intensity (HFrEF vs. controls: −7±5 vs. −6±7 bursts/min; −11±7 vs. −8±9 bursts/100 heartbeats) cycling; no differences were observed between HFrEF and controls (all, p&gt;0.05). Such initial reductions in MSNA burst frequency were observed in 9/11 HFrEF and 11/11 controls with zeroload cycling; reductions occurred in 11/11 HFrEF and 7/11 controls with moderate intensity cycling (both, p&gt;0.05). These similar group‐mean magnitude reductions in MSNA at the onset of exercise are consistent with preserved cardiopulmonary‐reflex mediated sympathoinhibition in HFrEF; the subsequent increase in MSNA is consistent with an exaggerated muscle metaboreflex contribution. As peripheral neurogenic vasoconstriction contributes to exercise intolerance in HFrEF, exercise regimens with shorter durations (e.g. high intensity interval training) may be better tolerated and more efficacious elements of conditioning protocols for such patients.Support or Funding InformationThis study was supported by Grants‐in‐Aid from the Heart and Stroke Foundation of Ontario (T4938, NA6298). P.J.M. was supported by a Canadian Institutes of Health Research Postdoctoral Fellowship. J.S.F. was supported by the Canada Research Chair Program.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

The lipid acyltransferase LYCAT controls phosphatidylinositol‐3,4,5‐trisphosphate (PIP3) signaling

Phosphatidylinositol‐3,4,5‐trisphospate (PIP3) is a potent signaling lipid that exerts key control over cell growth, survival, adhesion and migration. PIP3 signaling is often disrupted in various cancers, making understanding the regulation of PIP3 signaling an important priority. Much has been learned from the study of phosphorylation of the inositol headgroup of phosphoinositides, such as the phosphorylation of phosphatidylinositol‐4,5‐bisphospate (PIP2) to produce PIP3, as well as negative regulation of PIP3 by phosphoinositide phosphatases such as PTEN. However, much less is known about another key dimension of control of phosphoinositide action, the regulation of the fatty acyl profile of these lipids. Indeed, phosphatidylinositol and phosphoinositides exhibit remarkable selectivity of acyl chains (&gt;50% harboring 1‐stearoyl, 2‐arachidonoyl), an acyl profile quite distinct from other phospholipids. This suggests that control of fatty acyl profile of phosphoinositides may be an important determinant of the function of these lipids that has to date remained largely unexplored. We recently uncovered that the acyltransferase LYCAT is a key regulator of the acyl profile of specific phosphoinositides such as PIP2, thus exerting control over membrane traffic phenomena dependent on these phosphoinositides (Bone LN et al 2017 Mol Biol Cell. 28:161–172). We now examine how LYCAT controls PIP3 signaling and PIP3‐dependent control of cell physiology, including the activation of Akt and control of actin dynamics. We find that LYCAT perturbation impairs the activation of Akt and the activation of a number of Akt substrates, which in turn impacts cell growth and survival. Furthermore, LYCAT perturbation elicits dramatic changes in actin filament morphology, cell migration and cancer cell invasion. These results indicate that control of phosphoinositide acyl chain profile is an important novel dimension of regulation of PIP3 signaling, impacting proliferative, growth and migration signaling, and as such may be a novel dimension to control of cancer cell growth and progression.Support or Funding InformationThis work was supported by an Ontario Early Researcher Award, a Canada Research Chair Award, and a Natural Sciences and Engineering Research Council Grant to R.J.B, and a Project Grant and New Investigator Salary Award from the Canadian Institutes of Health Research to C.N.A.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Comparative Assessment of Central and Peripheral Chemoreceptor Reflex Regulation of Muscle Sympathetic Nerve Activity and Ventilation

We tested the hypothesis that central and peripheral chemoreceptor‐mediated sympathetic activation elicited by increases in CO2 tension (PCO2) mirror concurrent ventilatory responses. Twelve healthy young men (mean±SD; age: 24±4 yr; BMI: 24.5±1.3 kg·m−2) performed a modified rebreathing protocol designed to equilibrate central and peripheral chemoreceptor PCO2 tensions with end‐tidal PCO2 (PETCO2) at two isoxic end‐tidal PO2 (PETO2) tensions such that central chemoreceptor reflex responses (hyperoxia) can be isolated from additive peripheral responses (i.e., hypoxia minus hyperoxia). Subject's ventilation (volume turbine) and muscle sympathetic nerve activity (MSNA; microneurography, right fibular nerve) responses to rebreathing at isoxic PETO2 tensions of 150 mmHg (hyperoxia) and 50 mmHg (hypoxia) were recorded continuously. During rebreathing, the PETCO2 (mmHg) at which ventilation (L·min−1) and total MSNA (au) began to rise were identified as the PETCO2 recruitment thresholds (RT). Slopes of these responses above RT also were determined. In hyperoxic rebreathing (i.e., the central chemoreflex response), the mean RT for ventilation (46±3 mmHg) and MSNA (45±4 mmHg) did not differ (p&gt;0.05). Mean slopes were 2.3±0.9 L·min−1·mmHg−1 and 2.1±1.5 au·mmHg−1, respectively. In hypoxic rebreathing (i.e., the summed central and peripheral chemoreflex response), the RT for both ventilation and MSNA were not different (41±3 mmHg vs 41±3 mmHg, p&gt;0.05) but were lower (p&lt;0.05) compared to hyperoxia. Mean slopes for ventilation (4.0±1.8 L·min−1·mmHg−1) and MSNA (5.1±3.4 au·mmHg−1) in hypoxia were greater than in hyperoxia yielding a mean peripheral chemoreflex sensitivity (hypoxic slope minus hyperoxic slope) of 1.7±0.1 L·min−1·mmHg−1 for ventilation and 3.0±2.6 au·mmHg−1 for MSNA. The 95% limits of agreement of RT between ventilation and MSNA for the central chemoreflex were −7 to 6 mmHg (bias=−1 mmHg, p&gt;0.05) and −5 to 5 mmHg (bias=0 mmHg, p&gt;0.05) for the peripheral chemoreflex. There was no relationship (p&gt;0.05) between the ventilatory and MSNA sensitivity for either the central (r2=0.16) or peripheral (r2=0.01) chemoreceptor reflexes. These data highlight two novel findings: 1) ventilatory and sympathetic arms of the central and peripheral chemoreceptor reflexes are initiated at similar PETCO2 recruitment thresholds and 2) sympathetic responsiveness cannot be predicted from the sensitivities of the ventilatory central and peripheral chemoreceptor reflexes.Support or Funding InformationExperiments were funded by a Project Grant from the Canadian Institutes of Health Research (PJT 159491). Dr. Floras holds the Canada Research Chair in Integrative Cardiovascular Biology.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Non‐Invasive Pulmonary Gas Exchange Measurements Following Deep Breath‐Hold Diving

Breath‐hold diving is a ubiquitous activity, with recreational, fishing, military and competitive divers. During a dive, immersion and the increasing hydrostatic pressure of descent facilitate cardiovascular adjustments that promote a blood volume shift into the heart and chest vasculature, further augmenting autonomic responses (i.e. diving bradycardia, peripheral vasoconstriction, and splenic contraction) to conserve O2. Exceeding the compliant capacity of the lung and associated tissues can result in pulmonary injury called squeeze, which is characterized by post‐dive coughing, wheezing and hemoptysis. To provide insight into the consequences of such lung squeeze, our objective was to evaluate the time‐course of a maximal deep dive on pulmonary gas exchange, pulmonary artery systolic pressures (PASP), and ultrasound lung comets (ULCs). Ten healthy trained breath‐hold divers (33±9 yr; 80±15 kg; 185±9 cm) performed a maximal effort dive (55±12 m; range 40–76 m). Pulmonary gas exchange and PASP were performed at baseline, and repeated at 10 and 60 min post dive. Pulmonary gas exchange was non‐invasively measured by collecting end‐tidal PO2 and PCO2 during steady‐state breathing, and derived arterial PO2 via pulse oximetry. O2 deficit was defined as the difference between end‐tidal and calculated arterial PO2; consistent with the Bohr effect by utilizing end‐tidal PCO2. The ULCs were collected bilaterally, parasternal to mid‐axillary, at baseline and again 2.5 h post dive. Following the dive, although there was an impairment of pulmonary gas exchange, evident by an increase in O2 deficit from 10.9±4.5 mmHg at baseline to 22.9±5.1 mmHg 10‐min post dive (p=0.02), this deficit had normalized by 60‐min post. There was persistent hyperventilation up to 60 min post dive, reflected by reductions in end‐tidal PCO2 (p=0.02). PASP was unaltered post‐dive. Although ULCs trended to increase from baseline to 2.5 h post dive (3.7±3 to 10.4±13.2 comets; p=0.09), the delta change in ULC from baseline to post‐dive was positively correlated with depth (r=0.67, p=0.04) and, to a lesser extent, with both change in SpO2 (r=−0.62, p=0.06) and O2 deficit (r=0.58, p=0.08). The findings of this novel study suggest impairment in pulmonary gas exchange is present following diving to depth, even up to 10 min after surfacing. The mechanism(s) driving the slight hyperventilation are unclear, but potentially could be mediated via mild pulmonary edema and/or stimulation of juxtapulmonary capillary receptors. Together, there is transient impairment in gas exchange and associations with ULC during ‘modest’ dives. The development of pathological pulmonary edema, and severe and sustained impairment in pulmonary gas exchange, would seem likely at the forefront of breath‐hold diving performances (e.g, &gt;100 m). There remains little data regarding safe return‐to‐dive practices following a squeeze, and non‐invasive pulmonary gas exchange monitoring could provide valuable utility.Support or Funding InformationCanada Research Chair, NSERCThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Examining the effects of housing temperature on mitochondrial adaptations to exercise and the browning of white adipose tissue

The thermoneutral zone for small rodents is ~29–31°C and with decreases in ambient temperature below this there are consequent increases in metabolism to promote thermogenesis. Despite this, many studies report housing mice at room temperature for the convenience of the researchers studying them. As such, the conflicting reports between human and rodent models of exercise‐induced mitochondrial biogenesis in adipose tissue and the browning of white adipose tissue may be explained by the often‐overlooked variable, housing temperature. Male 8‐week‐old C57BL/6 mice were housed in cages with (VWR) or without (SED) running wheels (n = 12/group/condition) for 5–6 weeks at room temperature (RT; 22°C) or at thermoneutrality (TN; ~29°C). Mice housed at TN ran a greater distance. VWR protected against weight gain independent of housing temperature however food intake was greater in both VWR and SED mice housed at RT compared to TN. Inguinal white adipose tissue (iWAT) and epididymal white adipose tissue (eWAT) mass were reduced following VWR independent of housing temperature and there were main effects of both exercise and temperature to reduce brown adipose tissue (BAT) mass. There was no effect of housing temperature on circulating glucose or glycerol concentrations however NEFA, insulin, and the HOMA‐IR were increased at TN. In eWAT there was a main effect of exercise to increase cytochrome c oxidoreductase core I subunit (CORE1), cytochrome c oxidase, and peroxisome proliferator‐activated receptor‐γ coactivator‐1α (PGC‐1α) protein content with a main effect of housing temperature for PGC‐1α as well. There was a strong trend for an exercise‐induced increase in pyruvate dehydrogenase (PDH). In iWAT, there was a main effect of exercise and temperature on mitochondrial proteins with VWR leading to the expected increases in PDH, CORE1, COX IV, and PGC‐1α, but reduced markers of mitochondrial content at TN. Most notably protein content of the thermogenic uncoupling protein‐1 (UCP‐1) was undetectable in iWAT at TN. Qualitative histological analysis revealed patches of multilocular adipocytes in iWAT with VWR at RT but not at TN. Together our findings provide novel evidence that while housing mice at TN reduces markers of mitochondrial content in adipose tissue under sedentary conditions, the ability of exercise to increase mitochondrial protein content remains largely intact.Support or Funding InformationOntario Graduate Scholarship Canada Research Chairs ProgramThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Importance of TFEB and TFE3 in Mediating Lysosomal and Mitochondrial Adaptations

Autophagy is the process through which damaged cellular components are degraded by the lysosome, and it is essential for the overall maintenance of the cell. Mitochondria are recycled through a selective form of autophagy, known as mitophagy, in which dysfunctional mitochondria are poly‐ubiquitinated, engulfed in an autophagosomal membrane and transported to the lysosome for degradation. Chronic contractile activity (CCA) alters mitophagy flux in muscle, and this coincides with the activation of lysosomal biogenesis, which precedes changes in mitochondrial content. Thus, it is proposed that the clearance of dysfunctional mitochondria through the lysosomes may play an important role in mediating mitochondrial adaptations to CCA. We sought to understand the transcriptional regulation of lysosomal biogenesis brought about by CCA, by investigating the transcriptional regulators TFEB and TFE3, and their relationship to mitochondrial content. TFEB and TFE3 were silenced using siRNA in C2C12 myotubes, and subjected to 3 days of stimulation (3hr/day) to induce CA. Following a single bout, TFEB and TFE3 were elevated in both the nuclear and cytosolic fractions, which coincided with increases in Cathepsin D, preceding the increases in VDAC seen following 3 days of stimulation. The loss of TFEB or TFE3 individually had no impact on lysosomal adaptations, since LAMP1 and Cathepsin D remained responsive to CCA, demonstrating redundancy in role of TFEB and TFE3 in mediating lysosomal biogenesis. However, in the absence of both transcription factors, CCA‐induced lysosomal adaptations were reduced, illustrating their collective importance in mediating lysosomal biogenesis. Interestingly, the absence of TFEB had no effect on mitochondrial adaptations, measured by COX I and IV protein content, however this adaptation to CCA was blunted in the absence of TFE3. These results suggest that while TFEB and TFE3 may exhibit compensatory roles in mediating lysosomal biogenesis, TFE3 alone appears to be important for the mitochondrial response to CCA. Since, lysosomal adaptations proceed in the absence of TFE3, these data suggest a more direct role of TFE3 in the regulation of mitochondria in response to chronic contractile activity.Support or Funding InformationANO is funded by an Ontario Graduate Scholarship (OGS). JMM is funded by Natural Science and Engineering Research (NSERC) scholarship. DAH is the holder of a Canada Research Chair in Cell Physiology with funding from NSERC and CIHR.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Using Convolutional Neural Networks for the Detection of Spinal Transverse Processes

Introduction: Scoliosis, an excessive curvature of the spine, affects approximately 1 in 1,000 individuals. As a result, there have formerly been implementations of mandatory scoliosis screening procedures. Screening programs are no longer widely used as the harms often outweigh the benefits; it causes many adolescents to undergo frequent diagnosis X-ray procedure This makes spinal ultrasounds an ideal substitute for scoliosis screening in patients, as it does not expose them to those levels of radiation.&#x0D; Spinal curvatures can be accurately computed from the location of spinal transverse processes, by measuring the vertebral angle from a reference line [1]. However, ultrasound images are less clear than x-ray images, making it difficult to identify the spinal processes. To overcome this, we employ deep learning using a convolutional neural network, which is a powerful tool for computer vision and image classification [2].&#x0D; Method: A total of 2,752 ultrasound images were recorded from a spine phantom to train a convolutional neural network. Subsequently, we took another recording of 747 images to be used for testing. All the ultrasound images from the scans were then segmented manually, using the 3D Slicer (www.slicer.org) software. Next, the dataset was fed through a convolutional neural network. The network used was a modified version of GoogLeNet (Inception v1), with 2 linearly stacked inception models. This network was chosen because it provided a balance between accurate performance, and time efficient computations.&#x0D; Results: Deep learning classification using the Inception model achieved an accuracy of 84% for the phantom scan.&#x0D; Conclusion: The classification model performs with considerable accuracy. Better accuracy needs to be achieved, possibly with more available data and improvements in the classification model.&#x0D; Acknowledgements: G. Fichtinger is supported as a Canada Research Chair in Computer-Integrated Surgery. This work was funded, in part, by NIH/NIBIB and NIH/NIGMS (via grant 1R01EB021396-01A1 - Slicer+PLUS: Point-of-Care Ultrasound) and by CANARIE’s Research Software Program. &#x0D; &#x0D; Figure 1: Ultrasound scan containing a transverse process (left), and ultrasound scan containing no transverse process (right). Figure 2: Accuracy of classification for training (red) and validation (blue).&#x0D;

Gabapentin increases expression of δ subunit-containing GABAA receptors.

BackgroundGabapentin is a structural analog of the inhibitory neurotransmitter γ-aminobutyric acid (GABA). Its anticonvulsant, analgesic and anxiolytic properties suggest that it increases GABAergic inhibition; however, the molecular basis for these effects is unknown as gabapentin does not directly modify GABA type A (GABAA) receptor function, nor does it modify synaptic inhibition. Here, we postulated that gabapentin increases expression of δ subunit-containing GABAA (δGABAA) receptors that generate a tonic inhibitory conductance in multiple brain regions including the cerebellum and hippocampus.MethodsCell-surface biotinylation, Western blotting, electrophysiologic recordings, behavioral assays, high-performance liquid chromatography and gas chromatography-mass spectrometry studies were performed using mouse models.FindingsGabapentin enhanced expression of δGABAA receptors and increased a tonic inhibitory conductance in neurons. This increased expression likely contributes to GABAergic effects as gabapentin caused ataxia and anxiolysis in wild-type mice but not δ subunit null-mutant mice. In contrast, the antinociceptive properties of gabapentin were observed in both genotypes. Levels of GABAA receptor agonists and neurosteroids in the brain were not altered by gabapentin.InterpretationThese results provide compelling evidence to account for the GABAergic properties of gabapentin. Since reduced expression of δGABAA receptor occurs in several disorders, gabapentin may have much broader therapeutic applications than is currently recognized.FundSupported by a Foundation Grant (FDN-154312) from the Canadian Institutes of Health Research (to B.A.O.); a NSERC Discovery Grant (RGPIN-2016-05538), a Canada Research Chair in Sensory Plasticity and Reconsolidation, and funding from the University of Toronto Centre for the Study of Pain (to R.P.B.).

Editorial introductions

Editorial introductions

Factors affecting sex-related reporting in medical research: a cross-disciplinary bibliometric analysis

Clinical and preclinical studies have shown that there are sex-based differences at the genetic, cellular, biochemical, and physiological levels. Despite this, numerous studies have shown poor levels of inclusion of female populations into medical research. These disparities in sex inclusion in research are further complicated by the absence of sufficient reporting and analysis by sex of study populations. Disparities in the inclusion of the sexes in medical research substantially reduce the utility of the results of such research for the entire population. The absence of sex-related reporting are problematical for the translation of research from the preclinical to clinical and applied health settings. Large-scale studies are needed to identify the extent of sex-related reporting and where disparities are more prevalent. In addition, while several studies have shown the dearth of female researchers in science, few have evaluated whether a scarcity of women in science might be related to disparities in sex inclusion and reporting. We aimed to do a cross-disciplinary analysis of the degree of sex-related reporting across the health sciences-from biomedical, to clinical, and public health research-and the role of author gender in sex-related reporting. This bibliometric analysis analysed sex-related reporting in medical research examining more than 11·5 million papers indexed in Web of Science and PubMed between 1980 and 2016 and using sex-related Medical Subject Headings as a proxy for sex reporting. For papers that were published between 2008 and 2016 and could be matched with PubMed, we assigned a gender to first and last authors on the basis of their names, according to our gender assignment algorithm. We removed papers for which we could not determine the gender of either the first or last author. We grouped papers into three disciplinary categories (biomedical research, clinical medicine, and public health). We used descriptive statistics and regression analyses (controlling for the number of authors and representation of women in specific diseases, countries, continents, year, and specialty areas) to study associations between the gender of the authors and sex-related reporting. Between Jan 1, 1980, and Dec 31, 2016, sex-related reporting increased from 59% to 67% in clinical medicine and from 36% to 69% in public health research. But for biomedical research, sex remains largely under-reported (31% in 2016). Papers with female first and last authors had an increased probability of reporting sex, with an odds ratio of 1·26 (95% CI 1·24 to 1·27), and sex-related reporting was associated with publications in journals with low journal impact factors. For publications in 2016, sex-related reporting of both male and female is associated with a reduction of -0·51 (95% CI -0·54 to -0·47) in journal impact factors. Gender disparities in the scientific workforce and scarcity of policies on sex-related reporting at the journal and institutional level could inhibit effective research translation from bench to clinical studies. Diversification in the scientific workforce and in the research populations-from cell lines, to rodents, to humans-is essential to produce the most rigorous and effective medical research. Canada Research Chairs.