Assessing the Permeability of Engineered Capillary Networks in a 3D Culture

Many pathologies are characterized by poor blood vessel growth and reduced nutrient delivery to the surrounding tissue, introducing a need for tissue engineered blood vessels. Our lab has developed a 3D co-culture method to grow interconnected networks of pericyte-invested capillaries, which can anastamose with host vasculature following implantation to restore blood flow to ischemic tissues. However, if the engineered vessels contain endothelial cells (ECs) that are misaligned or contain wide junctional gaps, they may function improperly and behave more like the pathologic vessels that nourish tumors. The purpose of this study was to test the resistance to permeability of these networks in vitro, grown with different stromal cell types, as a metric of vessel functionality. A fluorescent dextran tracer was used to visualize transport across the endothelium and the pixel intensity was quantified using a customized MATLAB algorithm. In fibroblast-EC co-cultures, the dextran tracer easily penetrated through the vessel wall and permeability was high through the first 5 days of culture, indicative of vessel immaturity. Beyond day 5, dextran accumulated at the periphery of the vessel, with very little transported across the endothelium. Quantitatively, permeability dropped from initial levels of 61% to 39% after 7 days, and to 7% after 2 weeks. When ECs were co-cultured with bone marrow-derived mesenchymal stem cells (MSCs) or adipose-derived stem cells (AdSCs), much tighter control of permeability was achieved. Relative to the EC-fibroblast co-cultures, permeabilities were reduced 41% for the EC-MSC co-cultures and 50% for the EC-AdSC co-cultures after 3 days of culture. By day 14, these permeabilities decreased by 68% and 77% over the EC-fibroblast cultures. Co-cultures containing stem cells exhibit elevated VE-cadherin levels and more prominent EC-EC junctional complexes when compared to cultures containing fibroblasts. These data suggest the stromal cell identity influences the functionality and physiologic relevance of engineered capillary networks

Accounting for Diffusion in Agent Based Models of Reaction-Diffusion Systems with Application to Cytoskeletal Diffusion

Diffusion plays a key role in many biochemical reaction systems seen in nature. Scenarios where diffusion behavior is critical can be seen in the cell and subcellular compartments where molecular crowding limits the interaction between particles. We investigate the application of a computational method for modeling the diffusion of molecules and macromolecules in three-dimensional solutions using agent based modeling. This method allows for realistic modeling of a system of particles with different properties such as size, diffusion coefficients, and affinity as well as the environment properties such as viscosity and geometry. Simulations using these movement probabilities yield behavior that mimics natural diffusion. Using this modeling framework, we simulate the effects of molecular crowding on effective diffusion and have validated the results of our model using Langevin dynamics simulations and note that they are in good agreement with previous experimental data. Furthermore, we investigate an extension of this framework where single discrete cells can contain multiple particles of varying size in an effort to highlight errors that can arise from discretization that lead to the unnatural behavior of particles undergoing diffusion. Subsequently, we explore various algorithms that differ in how they handle the movement of multiple particles per cell and suggest an algorithm that properly accommodates multiple particles of various sizes per cell that can replicate the natural behavior of these particles diffusing. Finally, we use the present modeling framework to investigate the effect of structural geometry on the directionality of diffusion in the cell cytoskeleton with the observation that parallel orientation in the structural geometry of actin filaments of filopodia and the branched structure of lamellipodia can give directionality to diffusion at the filopodia-lamellipodia interface

Falls, Depression and Antidepressants in Later Life: A Large Primary Care Appraisal

BACKGROUND: Depression and falls are common and co-exist for older people. Safe management of each of these conditions is important to quality of life. METHODS: A cross-sectional survey was used to examine medication use associated with injurious and non-injurious falls in 21,900 community-dwelling adults, aged 60 years or over from 383 Australian general practices recruited for the DEPS-GP Project. Falls and injury from falls, medication use, depressive symptoms (Primary Health Questionnaire (PHQ-9)), clinical morbidity, suicidal ideation and intent, health status (SF-12 Health Survey), demographic and lifestyle information was reported in a standardised survey. FINDINGS: Respondents were 71.8 years (sd 7.7) of age and 58.4% were women. 24% 11% and 8% reported falls, fall related injury, and sought medical attention respectively. Antidepressant use (odds ratio, OR: 1.46; 95% confidence interval, 95%CI: 1.25, 1.70), questionable depression (5-14 on PHQ OR: 1.32, 95%CI: 1.13, 1.53) and clinically significant symptoms of depression (15 or more on PHQ OR: 1.70, 95%CI: 1.14, 1.50) were independently associated with multiple falls. SSRI use was associated with the highest risk of multiple falls (OR: 1.66, 95%CI: 1.36, 2.02) amongst all psychotropic medications. Similar associations were observed for injurious falls. Over 60% of those with four accumulated risk factors had multiple falls in the previous year (OR: 3.40, 95%CI: 1.79, 6.45); adjusted for other demographic and health factors. INTERPRETATION: Antidepressant use (particularly SSRIs) was strongly associated with falls regardless of presence of depressive symptoms. Strategies to prevent falls should become a routine part of the management of older people with depression

A Multiwell Platform for Studying Stiffness-Dependent Cell Biology

Adherent cells are typically cultured on rigid substrates that are orders of magnitude stiffer than their tissue of origin. Here, we describe a method to rapidly fabricate 96 and 384 well platforms for routine screening of cells in tissue-relevant stiffness contexts. Briefly, polyacrylamide (PA) hydrogels are cast in glass-bottom plates, functionalized with collagen, and sterilized for cell culture. The Young's modulus of each substrate can be specified from 0.3 to 55 kPa, with collagen surface density held constant over the stiffness range. Using automated fluorescence microscopy, we captured the morphological variations of 7 cell types cultured across a physiological range of stiffness within a 384 well plate. We performed assays of cell number, proliferation, and apoptosis in 96 wells and resolved distinct profiles of cell growth as a function of stiffness among primary and immortalized cell lines. We found that the stiffness-dependent growth of normal human lung fibroblasts is largely invariant with collagen density, and that differences in their accumulation are amplified by increasing serum concentration. Further, we performed a screen of 18 bioactive small molecules and identified compounds with enhanced or reduced effects on soft versus rigid substrates, including blebbistatin, which abolished the suppression of lung fibroblast growth at 1 kPa. The ability to deploy PA gels in multiwell plates for high throughput analysis of cells in tissue-relevant environments opens new opportunities for the discovery of cellular responses that operate in specific stiffness regimes

Potential Impact of Benzodiazepine Use on the Rate of Hip Fractures in Five Large European Countries and the United States

Benzodiazepine use increases the risk of falls and has been associated with an increased risk of hip fractures. Our aim was to estimate the possible population impact of the use of benzodiazepines on the rate of hip fracture in France, Germany, Italy, Spain, the United Kingdom, and the United States. We conducted a literature review to estimate the pooled relative risk (RR) for hip fractures and use of benzodiazepines. Prevalence rates of benzodiazepine use in 2009 were calculated for each country using the IMS MIDAS database and three public databases in Denmark, the Netherlands, and Norway. Both the RR and prevalence rates were used for calculation of population attributable risks (PARs) of hip fractures associated with benzodiazepine use. The literature review showed an increased risk of hip fractures in benzodiazepine users (RR = 1.4, 95 % CI 1.2–1.6). Rate of benzodiazepine use showed considerable differences between countries, ranging from 4.7 % to 22.3 % of population ever in a 1-year period. These are reflected in results for the PARs; estimated attributions of benzodiazepines to the rate of hip fractures were 1.8 %, 95 % CI 1.1–2.6 (Germany); 2.0 %, 95 % CI 1.2–2.8 (United Kingdom); 5.2 %, 95 % CI 3.2–7.3 (Italy); 7.4 %, 95 % CI 4.5–10.0 (France); 8.0 %, 95 % CI 4.9–11.0 (United States); and 8.2 %, 95 % CI 5.1–12.0 (Spain). PAR estimates suggest that the potential attribution of benzodiazepine use on the population rate of hip fractures in the five specified European countries and the United States varies between 1.8 % and 8.2 %. During the next phase of the IMI-PROTECT study, a comparison with individual patient data will show whether this approach is valid

Accelerometer-based physical activity in a large observational cohort - study protocol and design of the activity and function of the elderly in Ulm (ActiFE Ulm) study

<p>Abstract</p> <p>Background</p> <p>A large number of studies have demonstrated a positive effect of increased physical activity (PA) on various health outcomes. In all large geriatric studies, however, PA has only been assessed by interview-based instruments which are all subject to substantial bias. This may represent one reason why associations of PA with geriatric syndromes such as falls show controversial results. The general aim of the Active-Ulm study was to determine the association of accelerometer-based physical activity with different health-related parameters, and to study the influence of this standardized objective measure of physical activity on health- and disability-related parameters in a longitudinal setting.</p> <p>Methods</p> <p>We have set up an observational cohort study in 1500 community dwelling older persons (65 to 90 years) stratified by age and sex. Addresses have been obtained from the local residents registration offices. The study is carried out jointly with the IMCA - Respiratory Health Survey in the Elderly implemented in the context of the European project IMCA II. The study has a cross-sectional part (1) which focuses on PA and disability and two longitudinal parts (2) and (3). The primary information for part (2) is a prospective 1 year falls calendar including assessment of medication change. Part (3) will be performed about 36 months following baseline. Primary variables of interest include disability, PA, falls and cognitive function. Baseline recruitment has started in March 2009 and will be finished in April 2010.</p> <p>All participants are visited three times within one week, either at home or in the study center. Assessments included interviews on quality of life, diagnosed diseases, common risk factors as well as novel cognitive tests and established tests of physical functioning. PA is measured using an accelerometer-based sensor device, carried continuously over a one week period and accompanied by a prospective activity diary.</p> <p>Discussion</p> <p>The assessment of PA using a high standard accelerometer-based device is feasible in a large population-based study. The results obtained from cross-sectional and longitudinal analyses will shed light on important associations between PA and various outcomes and may provide information for specific interventions in older people.</p

Evaluation of risk of falls and orthostatic hypotension in older, long-term topical beta-blocker users

Background: Falls are a serious problem in the elderly, and have recently been described as cardiovascular-mediated side effects of beta-blocker eye drops. Therefore, we investigated the possible association between the long-term use of beta-blockers, prostaglandins and their combinations in eye drops, and falls, dizziness and orthostatic hypotension in older patients. Methods: All participants were long-term users of eye drops containing beta-blockers, prostaglandins or their combinations. They underwent a structured falls interview and blood pressure measurement for testing of orthostatic hypotension. The odds ratio for presence of orthostatic hypotension or a positive falls history according to use of beta-blocker eye drops was calculated with a binary logistic regression analysis. The main outcome measures were a positive falls history and the presence of orthostatic hypotension. Results: In total, 148 of 286 subjects participated. After adjustment for age, gender, and use of fall-risk-increasing drugs other than beta-blocker eye drops, we found no significant difference in fall risk [odds ratio (OR): 0.60; 95% confidence interval (CI): 0.268-1.327] between patients using ophthalmic beta-blockers or a combination of ophthalmic beta-blockers and prostaglandins, and patients using ophthalmic prostaglandins only. Although prevalence of orthostatic hypotension was higher in the beta-blocker group (OR: 1.67; 95% CI: 0.731-3.793) compared to the prostaglandin group, this was a non-significant difference. Conclusions: In our study, we did not find a significant association between long-term use of beta-blockers eye drops and falls, dizziness or orthostatic hypotension in older ophthalmic outpatients, compared to long-term use of prostaglandin eye drops