Microfluidic Bioreactor for Dynamic Regulation of Early Mesodermal Commitment in Human Pluripotent Stem Cells

During development and regeneration, tissues emerge from coordinated sequences of stem cell renewal, specialization and assembly that are orchestrated by cascades of regulatory signals. The complex and dynamic in vivo milieu cannot be replicated using standard in vitro techniques. Microscale technologies now offer potential for conducting highly controllable and sophisticated experiments at biologically relevant scales, with real-time insights into cellular responses. We developed a microbioreactor providing time sequences of space-resolved gradients of multiple molecular factors in three-dimensional (3D) cell culture settings, along with a versatile, high-throughput operation and imaging compatibility. A single microbioreactor yields up to 120 data points, corresponding to 15 replicates of a gradient with 8 concentration levels. Embryoid bodies (EBs) obtained from human embryonic and induced pluripotent stem cells (hESC, hiPSC) were exposed to concentration gradients of Wnt3a, Activin A, BMP4 and their inhibitors, to get new insights into the early-stage fate specification and mesodermal lineage commitment. We were able to evaluate the initiation of mesodermal induction by measuring and correlating the gene expression profiles to the concentration gradients of mesoderm-inducing morphogens. We propose that the microbioreactor systems combining spatial and temporal gradients of molecular and physical factors to hESC and hiPSC cultures can form a basis for predictable in vitro models of development and disease

Association Between Residential Greenness and Cardiovascular Disease Risk

Background Exposure to green vegetation has been linked to positive health, but the pathophysiological processes affected by exposure to vegetation remain unclear. To study the relationship between greenness and cardiovascular disease, we examined the association between residential greenness and biomarkers of cardiovascular injury and disease risk in susceptible individuals. Methods and Results In this cross-sectional study of 408 individuals recruited from a preventive cardiology clinic, we measured biomarkers of cardiovascular injury and risk in participant blood and urine. We estimated greenness from satellite-derived normalized difference vegetation index ( NDVI ) in zones with radii of 250 m and 1 km surrounding the participants' residences. We used generalized estimating equations to examine associations between greenness and cardiovascular disease biomarkers. We adjusted for residential clustering, demographic, clinical, and environmental variables. In fully adjusted models, contemporaneous NDVI within 250 m of participant residence was inversely associated with urinary levels of epinephrine (-6.9%; 95% confidence interval, -11.5, -2.0/0.1 NDVI ) and F2-isoprostane (-9.0%; 95% confidence interval, -15.1, -2.5/0.1 NDVI ). We found stronger associations between NDVI and urinary epinephrine in women, those not on β-blockers, and those who had not previously experienced a myocardial infarction. Of the 15 subtypes of circulating angiogenic cells examined, 11 were inversely associated (8.0-15.6% decrease/0.1 NDVI ), whereas 2 were positively associated (37.6-45.8% increase/0.1 NDVI ) with contemporaneous NDVI . Conclusions Independent of age, sex, race, smoking status, neighborhood deprivation, statin use, and roadway exposure, residential greenness is associated with lower levels of sympathetic activation, reduced oxidative stress, and higher angiogenic capacity

High predictability of terrestrial carbon fluxes from an initialized decadal prediction system

Interannual variations in the flux of carbon dioxide (CO2) between the land surface and the atmosphere are the dominant component of interannual variations in the atmospheric CO2 growth rate. Here, we investigate the potential to predict variations in these terrestrial carbon fluxes 1&ndash;10 years in advance using a novel set of retrospective decadal forecasts of an Earth system model. We demonstrate that globally-integrated net ecosystem production (NEP) exhibits high potential predictability for 2 years following forecast initialization. This predictability exceeds that from a persistence or uninitialized forecast conducted with the same Earth system model. The potential predictability in NEP derives mainly from high predictability in ecosystem respiration, which itself is driven by vegetation carbon and soil moisture initialization. Our findings unlock the potential to forecast the terrestrial ecosystem in a changing environment. </div

Simulation model of the F/A-18 high angle-of-attack research vehicle utilized for the design of advanced control laws

The 'f18harv' six degree-of-freedom nonlinear batch simulation used to support research in advanced control laws and flight dynamics issues as part of NASA's High Alpha Technology Program is described in this report. This simulation models an F/A-18 airplane modified to incorporate a multi-axis thrust-vectoring system for augmented pitch and yaw control power and actuated forebody strakes for enhanced aerodynamic yaw control power. The modified configuration is known as the High Alpha Research Vehicle (HARV). The 'f18harv' simulation was an outgrowth of the 'f18bas' simulation which modeled the basic F/A-18 with a preliminary version of a thrust-vectoring system designed for the HARV. The preliminary version consisted of two thrust-vectoring vanes per engine nozzle compared with the three vanes per engine actually employed on the F/A-18 HARV. The modeled flight envelope is extensive in that the aerodynamic database covers an angle-of-attack range of -10 degrees to +90 degrees, sideslip range of -20 degrees to +20 degrees, a Mach Number range between 0.0 and 2.0, and an altitude range between 0 and 60,000 feet

Engineering of human cardiac muscle electromechanically matured to an adult-like phenotype

Author ManuscriptThe application of tissue-engineering approaches to human induced pluripotent stem (hiPS) cells enables the development of physiologically relevant human tissue models for in vitro studies of development, regeneration, and disease. However, the immature phenotype of hiPS-derived cardiomyocytes (hiPS-CMs) limits their utility. We have developed a protocol to generate engineered cardiac tissues from hiPS cells and electromechanically mature them toward an adult-like phenotype. This protocol also provides optimized methods for analyzing these tissues' functionality, ultrastructure, and cellular properties. The approach relies on biological adaptation of cultured tissues subjected to biomimetic cues, applied at an increasing intensity, to drive accelerated maturation. hiPS cells are differentiated into cardiomyocytes and used immediately after the first contractions are observed, when they still have developmental plasticity. This starting cell population is combined with human dermal fibroblasts, encapsulated in a fibrin hydrogel and allowed to compact under passive tension in a custom-designed bioreactor. After 7 d of tissue formation, the engineered tissues are matured for an additional 21 d by increasingly intense electromechanical stimulation. Tissue properties can be evaluated by measuring contractile function, responsiveness to electrical stimuli, ultrastructure properties (sarcomere length, mitochondrial density, networks of transverse tubules), force-frequency and force-length relationships, calcium handling, and responses to β-adrenergic agonists. Cell properties can be evaluated by monitoring gene/protein expression, oxidative metabolism, and electrophysiology. The protocol takes 4 weeks and requires experience in advanced cell culture and machining methods for bioreactor fabrication. We anticipate that this protocol will improve modeling of cardiac diseases and testing of drugs.NIBIB and NCATS grant EB17103 (G.V.-N.); NIBIB, NCATS, NIAMS, NIDCR, and NIEHS grant EB025765 (G.V.-N.); NHLBI grants HL076485 (G.V.-N.) and HL138486 (M.Y.); NSF grant 16478 (G.V.-N.); the University of Minho MD/PhD program (D.T.); a Japan Society for the Promotion of Science fellowship (K.M.); and the Columbia University Stem Cell Initiative (L.S., M.Y.

Human bone perivascular niche-on-a-chip for studying metastatic colonization

Eight out of 10 breast cancer patients die within 5 years after the primary tumor has spread to the bones. Tumor cells disseminated from the breast roam the vasculature, colonizing perivascular niches around blood capillaries. Slow flows support the niche maintenance by driving the oxygen, nutrients, and signaling factors from the blood into the interstitial tissue, while extracellular matrix, endothelial cells, and mesenchymal stem cells regulate metastatic homing. Here, we show the feasibility of developing a perfused bone perivascular niche-on-a-chip to investigate the progression and drug resistance of breast cancer cells colonizing the bone. The model is a functional human triculture with stable vascular networks within a 3D native bone matrix cultured on a microfluidic chip. Providing the niche-on-a-chip with controlled flow velocities, shear stresses, and oxygen gradients, we established a long-lasting, self-assembled vascular network without supplementation of angiogenic factors. We further show that human bone marrow-derived mesenchymal stem cells, which have undergone phenotypical transition toward perivascular cell lineages, support the formation of capillary-like structures lining the vascular lumen. Finally, breast cancer cells exposed to interstitial flow within the bone perivascular niche-ona- chip persist in a slow-proliferative state associated with increased drug resistance. We propose that the bone perivascular niche-ona- chip with interstitial flow promotes the formation of stable vasculature and mediates cancer cell colonization

Transient study of the oxygen reduction reaction on reduced Pt and Pt alloys microelectrodes: evidence for the reduction of pre-adsorbed oxygen species linked to dissolved oxygen

Using chronoamperometry at preconditioned oxide-free Pt microdisc electrodes in aqueous media, we investigated the oxygen reduction reaction (ORR) on the millisecond timescale and obtained results consistent with the reduction of oxygen species which adsorb on the electrode before the ORR is electrochemically driven. Furthermore these adsorbed species are clearly linked to oxygen in solution. At long times, the amperometric response is solely controlled by the diffusion of dissolved oxygen towards the microelectrode. However, at short times, typically below 50 ms, the reduction of pre-adsorbed oxygen produces a large extra current whose magnitude depends on the oxygen concentration in solution, deliberate electrode poisoning and the rest time before the potential step. Using sampled current voltammetry we show that this extra current affects the entire potential range of the ORR. Using microdisc electrodes made with Pt alloys we find that the amperometric response is sufficiently sensitive to distinguish oxygen coverage differences between Pt, Pt0.9Rh0.1 and Pt0.9Ir0.1 microdiscs. These unexpected and, to our knowledge, never previously reported results provide new insight into the oxygen reduction reaction on Pt. The existence over a wide potential range of irreversibly adsorbed oxygen species arising from dissolved oxygen and different from Pt oxide is particularly relevant to the development of oxygen reduction catalysts for low temperature fuel cells

Design Specification for a Thrust-Vectoring, Actuated-Nose-Strake Flight Control Law for the High-Alpha Research Vehicle

Specifications for a flight control law are delineated in sufficient detail to support coding the control law in flight software. This control law was designed for implementation and flight test on the High-Alpha Research Vehicle (HARV), which is an F/A-18 aircraft modified to include an experimental multi-axis thrust-vectoring system and actuated nose strakes for enhanced rolling (ANSER). The control law, known as the HARV ANSER Control Law, was designed to utilize a blend of conventional aerodynamic control effectors, thrust vectoring, and actuated nose strakes to provide increased agility and good handling qualities throughout the HARV flight envelope, including angles of attack up to 70 degrees

Skill metrics for confronting global upper ocean ecosystem-biogeochemistry models against field and remote sensing data

Author Posting. © Elsevier B.V., 2009. This is the author's version of the work. It is posted here by permission of Elsevier B.V. for personal use, not for redistribution. The definitive version was published in Journal of Marine Systems 76 (2009): 95-112, doi:10.1016/j.jmarsys.2008.05.015.We present a generalized framework for assessing the skill of global upper ocean ecosystem-biogeochemical models against in-situ field data and satellite observations. We illustrate the approach utilizing a multi-decade (1979-2004) hindcast experiment conducted with the Community Climate System Model (CCSM-3) ocean carbon model. The CCSM-3 ocean carbon model incorporates a multi-nutrient, multi-phytoplankton functional group ecosystem module coupled with a carbon, oxygen, nitrogen, phosphorus, silicon, and iron biogeochemistry module embedded in a global, threedimensional ocean general circulation model. The model is forced with physical climate forcing from atmospheric reanalysis and satellite data products and time-varying atmospheric dust deposition. Data-based skill metrics are used to evaluate the simulated time-mean spatial patterns, seasonal cycle amplitude and phase, and subannual to interannual variability. Evaluation data include: sea surface temperature and mixed layer depth; satellite derived surface ocean chlorophyll, primary productivity, phytoplankton growth rate and carbon biomass; large-scale climatologies of surface nutrients, pCO2, and air-sea CO2 and O2 flux; and time-series data from the Joint Global Ocean Flux Study (JGOFS). Where the data is sufficient, we construct quantitative skill metrics using: model-data residuals, time-space correlation, root mean square error, and Taylor diagrams.This work was supported in part by grants from the NSF/ONR National Ocean Partnership Program (N000140210370), the NASA Ocean Biology and Biogeochemistry Program (NNX07AL80G), and the NSF Center for Microbial Oceanography Research and Education (C-MORE)

Impacts of increasing anthropogenic soluble iron and nitrogen deposition on ocean biogeochemistry

Author Posting. © American Geophysical Union, 2009. This article is posted here by permission of American Geophysical Union for personal use, not for redistribution. The definitive version was published in Global Biogeochemical Cycles 23 (2009): GB3016, doi:10.1029/2008GB003440.We present results from transient sensitivity studies with the Biogeochemical Elemental Cycling (BEC) ocean model to increasing anthropogenic atmospheric inorganic nitrogen (N) and soluble iron (Fe) deposition over the industrial era. Elevated N deposition results from fossil fuel combustion and agriculture, and elevated soluble Fe deposition results from increased atmospheric processing in the presence of anthropogenic pollutants and soluble Fe from combustion sources. Simulations with increasing Fe and increasing Fe and N inputs raised simulated marine nitrogen fixation, with the majority of the increase in the subtropical North and South Pacific, and raised primary production and export in the high-nutrient low-chlorophyll (HNLC) regions. Increasing N inputs alone elevated small phytoplankton and diatom production, resulting in increased phosphorus (P) and Fe limitation for diazotrophs, hence reducing nitrogen fixation (∼6%). Globally, the simulated primary production, sinking particulate organic carbon (POC) export. and atmospheric CO2 uptake were highest under combined increase in Fe and N inputs compared to preindustrial control. Our results suggest that increasing combustion iron sources and aerosol Fe solubility along with atmospheric anthropogenic nitrogen deposition are perturbing marine biogeochemical cycling and could partially explain the observed trend toward increased P limitation at station ALOHA in the subtropical North Pacific. Excess inorganic nitrogen ([NO3 −] + [NH4 +] − 16[PO4 3−]) distributions may offer useful insights for understanding changing ocean circulation and biogeochemistry.This work was supported by funding from NSF grant OCE-0452972 to J. K. Moore and C. S. Zender. Computations were supported by the Earth System Modeling Facility at UCI (NSFATMO321380) and by the Climate Simulation Laboratory at National Center for Atmospheric Research. The National Center for Atmospheric Research is sponsored by the U.S. National Science Foundation. N.M. would like to acknowledge the assistance of NSF– Carbon and Water (ATM-0628472), and N.M., S.D., and C.L. would like to acknowledge the assistance of NASA-IDS (NNX07AL80G)