Applying Spinal Cord Organoids as a quantitative approach to study the mammalian Hedgehog pathway.

The Hedgehog (HH) pathway is crucial for embryonic development, and adult homeostasis. Its dysregulation is implicated in multiple diseases. Existing cellular models used to study HH signal regulation in mammals do not fully recapitulate the complexity of the pathway. Here we show that Spinal Cord Organoids (SCOs) can be applied to quantitively study the activity of the HH pathway. During SCO formation, the specification of different categories of neural progenitors (NPC) depends on the intensity of the HH signal, mirroring the process that occurs during neural tube development. By assessing the number of NPCs within these distinct subgroups, we are able to categorize and quantify the activation level of the HH pathway. We validate this system by measuring the effects of mutating the HH receptor PTCH1 and the impact of HH agonists and antagonists on NPC specification. SCOs represent an accessible and reliable in-vitro tool to quantify HH signaling and investigate the contribution of genetic and chemical cues in the HH pathway regulation.

Impact of heteroaggregation between microplastics and algae on particle vertical transport.

Understanding the impacts of microplastics (MPs) on aqueous environments requires understanding their transport dynamics and how their presence affects other natural processes and cycles. In this context, one aspect to consider is how MPs interact with freshwater snow (FWS), a mixture of algae and natural particles. FWS is one of the primary drivers of the flux of organic matter from the water surface to the bottom sediment, where zooplankton, diurnal migration, fish faecal pellets settling and turbulent mixing can also play prominent roles. Understanding how MPs and FWS heteroaggregation affects their respective settling velocities is important to assess not only MPs fate and transport but also their ecological impacts by altering FWS deposition and thereby nutrient cycling. In this present study, we obtained a mechanistic understanding of the processes controlling MPs settling dynamics and heteroaggregation with FWS and the subsequent impacts on the settling rates of both MPs and ballasted FWS. Here we used a plexiglass column equipped with a stereoscopic camera system to track the settling velocities of (1) MPs of various compositions, densities and morphologies, (2) FWS flocs and (3) MP-FWS agglomerates. For each experimental set, thousands of particles were tracked over a series of image sequences. We found that agglomerates with high-density MPs settled at least twofold faster than FWS alone, implying a much smaller residence time in the water column, except for cases with MP fibres or low-density plastics. These findings will help to refine MP fate models and, while contingent on MPs number, may impact biogeochemical cycles by changing the flux of nutrients contained in FWS to the sediment.

Immiscible Rayleigh-Taylor turbulence: Implications for bacterial degradation in oil spills.

An unstable density stratification between two fluids mixes spontaneously under the effect of gravity, a phenomenon known as Rayleigh-Taylor (RT) turbulence. If the two fluids are immiscible, for example, oil and water, surface tension prevents intermixing at the molecular level. However, turbulence fragments one fluid into the other, generating an emulsion in which the typical droplet size decreases over time as a result of the competition between the rising kinetic energy and the surface energy density. Even though the first phenomenological theory describing this emulsification process was derived many years ago, it has remained elusive to experimental verification, hampering our ability to predict the fate of oil in applications such as deep-water spills. Here, we provide the first experimental and numerical verification of the immiscible RT turbulence theory, unveiling a unique turbulent state that originates at the oil-water interface due to the interaction of multiple capillary waves. We show that a single, non-dimensional, and time-independent parameter controls the range of validity of the theory. Our findings have wide-ranging implications for the understanding of the mixing of immiscible fluids. This includes in particular oil spills, where our work enables the prediction of the oil-water interface dynamics that ultimately determine the rate of oil biodegradation by marine bacteria.

In vitro investigation of the blood flow downstream of a 3D-printed aortic valve.

The hemodynamics in the aorta as well as the durability of aortic valve prostheses vary greatly between different types of devices. Although placement and sizing of surgical aortic valve prostheses are excellent, the valve geometry of common devices cannot be customized to fit the patient's anatomy perfectly. Similarly, transcatheter aortic valve implantation (TAVI) devices are not customizable and may be orientated unfavorably during implantation. Imperfect fit of an aortic valve prosthesis may result in suboptimal performance and in some cases the need for additional surgery. Leveraging the advent of precision, multi-material 3D-printing, a bioinspired silicone aortic valve was developed. The manufacturing technique makes it fully customizable and significantly cheaper to develop and produce than common prostheses. In this study, we assess the hemodynamic performance of such a 3D-printed aortic valve and compare it to two TAVI devices as well as to a severely stenosed valve. We investigate the blood flow distal to the valve in an anatomically accurate, compliant aorta model via three-dimensional particle tracking velocimetry measurements. Our results demonstrate that the 3D-printed aortic valve induces flow patterns and topology compatible with the TAVI valves and showing similarity to healthy aortic blood flow. Compared to the stenosis, the 3D-printed aortic valve reduces turbulent kinetic energy levels and irreversible energy losses by over 75%, reaching values compatible with healthy subjects and conventional TAVIs. Our study substantiates that the 3D-printed heart valve displays a hemodynamic performance similar to established devices and underscores its potential for driving innovation towards patient specific valve prostheses.

An engineered Sox17 induces somatic to neural stem cell fate transitions independently from pluripotency reprogramming.

Advanced strategies to interconvert cell types provide promising avenues to model cellular pathologies and to develop therapies for neurological disorders. Yet, methods to directly transdifferentiate somatic cells into multipotent induced neural stem cells (iNSCs) are slow and inefficient, and it is unclear whether cells pass through a pluripotent state with full epigenetic reset. We report iNSC reprogramming from embryonic and aged mouse fibroblasts as well as from human blood using an engineered Sox17 (eSox17FNV). eSox17FNV efficiently drives iNSC reprogramming while Sox2 or Sox17 fail. eSox17FNV acquires the capacity to bind different protein partners on regulatory DNA to scan the genome more efficiently and has a more potent transactivation domain than Sox2. Lineage tracing and time-resolved transcriptomics show that emerging iNSCs do not transit through a pluripotent state. Our work distinguishes lineage from pluripotency reprogramming with the potential to generate more authentic cell models for aging-associated neurodegenerative diseases.

Universal alignment in turbulent pair dispersion.

Countless processes in nature and industry, from rain droplet nucleation to plankton interaction in the ocean, are intimately related to turbulent fluctuations of local concentrations of advected matter. These fluctuations can be described by considering the change of the separation between particle pairs, known as pair dispersion, which is believed to obey a cubic in time growth according to Richardson's theory. Our work reveals a universal, scale-invariant alignment between the relative velocity and position vectors of dispersing particles at a mean angle that we show to be a universal constant of turbulence. We connect the value of this mean angle to Richardson's traditional theory and find agreement with data from a numerical simulation and a laboratory experiment. While the Richardson's cubic regime has been observed for small initial particle separations only, the constancy of the mean angle manifests throughout the entire inertial range of turbulence. Thus, our work reveals the universal nature of turbulent pair dispersion through a geometrical paradigm whose validity goes beyond the classical theory, and provides a framework for understanding and modeling transport and mixing processes.

Global warming accelerates soil heterotrophic respiration.

Carbon efflux from soils is the largest terrestrial carbon source to the atmosphere, yet it is still one of the most uncertain fluxes in the Earth's carbon budget. A dominant component of this flux is heterotrophic respiration, influenced by several environmental factors, most notably soil temperature and moisture. Here, we develop a mechanistic model from micro to global scale to explore how changes in soil water content and temperature affect soil heterotrophic respiration. Simulations, laboratory measurements, and field observations validate the new approach. Estimates from the model show that heterotrophic respiration has been increasing since the 1980s at a rate of about 2% per decade globally. Using future projections of surface temperature and soil moisture, the model predicts a global increase of about 40% in heterotrophic respiration by the end of the century under the worst-case emission scenario, where the Arctic region is expected to experience a more than two-fold increase, driven primarily by declining soil moisture rather than temperature increase.

Internal Biofilm Heterogeneities Enhance Solute Mixing and Chemical Reactions in Porous Media.

Bacterial biofilms can form in porous media that are of interest in industrial applications ranging from medical implants to biofilters as well as in environmental applications such as in situ groundwater remediation, where they can be critical locations for biogeochemical reactions. The presence of biofilms modifies porous media topology and hydrodynamics by clogging pores and consequently solutes transport and reactions kinetics. The interplay between highly heterogeneous flow fields found in porous media and microbial behavior, including biofilm growth, results in a spatially heterogeneous biofilm distribution in the porous media as well as internal heterogeneity across the thickness of the biofilm. Our study leverages highly resolved three-dimensional X-ray computed microtomography images of bacterial biofilms in a tubular reactor to numerically compute pore-scale fluid flow and solute transport by considering multiple equivalent stochastically generated internal permeability fields for the biofilm. We show that the internal heterogeneous permeability mainly impacts intermediate velocities when compared with homogeneous biofilm permeability. While the equivalent internal permeability fields of the biofilm do not impact fluid-fluid mixing, they significantly control a fast reaction. For biologically driven reactions such as nutrient or contaminant uptake by the biofilm, its internal permeability field controls the efficiency of the process. This study highlights the importance of considering the internal heterogeneity of biofilms to better predict reactivity in industrial and environmental bioclogged porous systems.

TMED2 binding restricts SMO to the ER and Golgi compartments.

Hedgehog (HH) signaling is important for embryonic pattering and stem cell differentiation. The G protein-coupled receptor (GPCR) Smoothened (SMO) is the key HH signal transducer modulating both transcription-dependent and transcription-independent responses. We show that SMO protects naive mouse embryonic stem cells (ESCs) from dissociation-induced cell death. We exploited this SMO dependency to perform a genetic screen in haploid ESCs where we identify the Golgi proteins TMED2 and TMED10 as factors for SMO regulation. Super-resolution microscopy shows that SMO is normally retained in the endoplasmic reticulum (ER) and Golgi compartments, and we demonstrate that TMED2 binds to SMO, preventing localization to the plasma membrane. Mutation of TMED2 allows SMO accumulation at the plasma membrane, recapitulating early events after HH stimulation. We demonstrate the physiologic relevance of this interaction in neural differentiation, where TMED2 functions to repress HH signal strength. Identification of TMED2 as a binder and upstream regulator of SMO opens the way for unraveling the events in the ER-Golgi leading to HH signaling activation.

Biophysical properties at patch scale shape the metabolism of biofilm landscapes.

Phototrophic biofilms form complex spatial patterns in streams and rivers, yet, how community patchiness, structure and function are coupled and contribute to larger-scale metabolism remains unkown. Here, we combined optical coherence tomography with automated O2 microprofiling and amplicon sequencing in a flume experiment to show how distinct community patches interact with the hydraulic environment and how this affects the internal distribution of oxygen. We used numerical simulations to derive rates of community photosynthetic activity and respiration at the patch scale and use the obtained parameter to upscale from individual patches to the larger biofilm landscape. Our biofilm landscape approach revealed evidence of parallels in the structure-function coupling between phototrophic biofilms and their streambed habitat.

Toward an accurate estimation of wall shear stress from 4D flow magnetic resonance downstream of a severe stenosis.

PURPOSE: First, to investigate the agreement between velocity, velocity gradient, and Reynolds stress obtained from four-dimensional flow magnetic resonance (4D flow MRI) measurements and direct numerical simulation (DNS). Second, to propose and optimize based on DNS, 2 alternative methods for the accurate estimation of wall shear stress (WSS) when the resolution of the flow measurements is limited. Thirdly, to validate the 2 methods based on 4D flow MRI data. METHODS: In vitro 4D MRI has been conducted in a realistic rigid stenosed aorta model under a constant flow rate of 12 L/min. A DNS of transitional stenotic flow has been performed using the same geometry and boundary conditions. RESULTS: Time-averaged velocity and Reynolds stresses are in good agreement between in vitro 4D MRI data and DNS (errors between 2% and 8% of the reference downsampled data). WSS estimation based on the 2 proposed methods applied to MRI data provide good agreement with DNS for slice-averaged values (maximum error is less than 15% of the mean reference WSS for the first method and 25% for the second method). The performance of both models is not strongly sensitive to spatial resolution up to 1.5 mm voxel size. While the performance of model 1 deteriorates appreciably at low signal-to-noise ratios, model 2 remains robust. CONCLUSIONS: The 2 methods for WSS magnitude give an overall better agreement than the standard approach used in the literature based on direct calculation of the velocity gradient close to the wall (relative error of 84%).

A Novel Estimation Approach of Pressure Gradient and Haemodynamic Stresses as Indicators of Pathological Aortic Flow Using Subvoxel Modelling.

OBJECTIVE: The flow downstream from aortic stenoses is characterised by the onset of shear-induced turbulence that leads to irreversible pressure losses. These extra losses represent an increased resistance that impacts cardiac efficiency. A novel approach is suggested in this study to accurately evaluate the pressure gradient profile along the aorta centreline using modelling of haemodynamic stress at scales that are smaller than the typical resolution achieved in experiments. METHODS: We use benchmark data obtained from direct numerical simulation (DNS) along with results from in silico and in vitro three-dimensional particle tracking velocimetry (3D-PTV) at three voxel sizes, namely 750  µm, 1 mm and 1.5 mm. A differential equation is derived for the pressure gradient, and the subvoxel-scale (SVS) stresses are closed using the Smagorinsky and a new refined model. Model constants are optimised using DNS and in silico PTV data and validated based on pulsatile in vitro 3D-PTV data and pressure catheter measurements. RESULTS: The Smagorinsky-based model was found to be more accurate for SVS stress estimation but also more sensitive to errors especially at lower resolution, whereas the new model was found to more accurately estimate the projected pressure gradient even for larger voxel size of 1.5 mm albeit at the cost of increased sensitivity at this voxel size. A comparison with other methods in the literature shows that the new approach applied to in vitro PTV measurements estimates the irreversible pressure drop by decreasing the errors by at least 20%. CONCLUSION: Our novel approach based on the modelling of subvoxel stress offers a validated and more accurate way to estimate pressure gradient, irreversible pressure loss and SVS stress. SIGNIFICANCE: We anticipate that the approach may potentially be applied to image-based in vivo, in vitro 4D flow data or in silico data with limited spatial resolution to assess pressure loss and SVS stresses in disturbed aortic blood flow.

Efficient mate finding in planktonic copepods swimming in turbulence.

Zooplankton live in dynamic environments where turbulence may challenge their limited swimming abilities. How this interferes with fundamental behavioral processes remains elusive. We reconstruct simultaneously the trajectories of flow tracers and calanoid copepods and we quantify their ability to find mates when ambient flow imposes physical constrains on their motion and impairs their olfactory orientation. We show that copepods achieve high encounter rates in turbulence due to the contribution of advection and vigorous swimming. Males further convert encounters within the perception radius to contacts and then to mating via directed motion toward nearby organisms within the short time frame of the encounter. Inertial effects do not result in preferential concentration, reducing the geometric collision kernel to the clearance rate, which we model accurately by superposing turbulent velocity and organism motion. This behavioral and physical coupling mechanism may account for the ability of copepods to reproduce in turbulent environments.

Assessment of the Flow Field in the HeartMate 3 Using Three-Dimensional Particle Tracking Velocimetry and Comparison to Computational Fluid Dynamics.

Flow fields in rotary blood pumps (RBPs) have a significant influence on hemocompatibility. Because flow characteristics vary with flow rate, different operating conditions play a role. Furthermore, turbulence is crucial in the evaluation of blood damage potential, but the level of turbulence in implantable RBPs is still unknown. In this study, we addressed both research aspects and for the first time measured turbulent flow fields in the HeartMate 3 (HM3) at different operating flows. The averaged, three-dimensional velocity field including fluctuating velocity components in a HM3 with a transparent lower housing was measured using three-dimensional particle tracking velocimetry (3D-PTV). In vitro results were compared with computational fluid dynamic (CFD) simulations for two flow cases, representing the lower and upper physiologic flow range (2.7 and 5.7 L/min), using two different turbulence models that account for fluctuating velocity fields: the k-ω shear stress transport and the Reynolds stress model (RSM). The measurements revealed higher mean and turbulent kinetic energies (TKEs) for the low-flow condition especially within the gap beneath the impeller. Computed mean fields agree well with 3D-PTV for both models, but the RSM predicts the TKE levels better than the k-ω model. Computational fluid dynamic results further show wall shear stresses higher than 150 Pa, a commonly used damage threshold, in the bottom gap for the lower flow condition. In conclusion, the low-flow condition was found to be more prone to blood damage. Furthermore, CFD predictions for turbulence must be carefully experimentally validated.

SOX17 in cellular reprogramming and cancer.

SOX17 is a transcription factor directing the specification and development of the primitive endoderm, primitive germ cells, definitive endoderm and, subsequently, is involved in the cardiovascular system and several endoderm-derived organs. The analysis of cancer genome sequencing data classified SOX17 as mutated cancer driver gene in endometrial cancer. These studies identified hotspot missense mutations within its DNA binding and transactivation domains. In somatic cell reprogramming, structure-based protein re-engineering showed a single missense mutation in SOX17 can change the DNA dependent heterodimer formation with OCT4 and enables the replacement of SOX2 with SOX17 mutants to induce pluripotency. This reveals the profound impact of specific missense mutations on gene function and regulatory activity. Here, we review the roles of SOX17 in cancer and discuss its cross-talk with the WNT/ß-catenin pathway, potentially reconciling its activity as re-engineered reprogramming factor and mutated cancer driver gene.

Pluripotency reprogramming by competent and incompetent POU factors uncovers temporal dependency for Oct4 and Sox2.

Oct4, along with Sox2 and Klf4 (SK), can induce pluripotency but structurally similar factors like Oct6 cannot. To decode why Oct4 has this unique ability, we compare Oct4-binding, accessibility patterns and transcriptional waves with Oct6 and an Oct4 mutant defective in the dimerization with Sox2 (Oct4defSox2). We find that initial silencing of the somatic program proceeds indistinguishably with or without Oct4. Oct6 mitigates the mesenchymal-to-epithelial transition and derails reprogramming. These effects are a consequence of differences in genome-wide binding, as the early binding profile of Oct4defSox2 resembles Oct4, whilst Oct6 does not bind pluripotency enhancers. Nevertheless, in the Oct6-SK condition many otherwise Oct4-bound locations become accessible but chromatin opening is compromised when Oct4defSox2 occupies these sites. We find that Sox2 predominantly facilitates chromatin opening, whilst Oct4 serves an accessory role. Formation of Oct4/Sox2 heterodimers is essential for pluripotency establishment; however, reliance on Oct4/Sox2 heterodimers declines during pluripotency maintenance.

Performance analysis of the transcatheter aortic valve implantation on blood flow hemodynamics: An optical imaging-based in vitro study.

Cardiac implants may have a strong influence on the hemodynamics of the circulatory system. In this study, we aimed at investigating the impact of transcatheter aortic valve implantation (TAVI) devices on blood flow patterns that develop in the ascending aorta under physiological flow conditions in vitro. For this purpose, a noninvasive optical measurement tool, three-dimensional particle tracking velocimetry (3D-PTV), was used in a realistic compliant silicone aortic model. The performance and the influence of two TAVIs and one surgical valve on the aortic flow were investigated. Our results showed that valve design and materials may have a distinct influence on relevant hemodynamic properties, namely kinetic energy, production of turbulence, and shear stresses in the ascending aorta. All properties varied considerably between the different valve models. We found that the total aortic regurgitation composed of the closing volume, transvalvular and paravalvular leakages varied for the three valves investigated. Furthermore, peak mean kinetic energy (MKE) ranged from 61 to 116 J/m3 , whereas peak turbulent kinetic energy (TKE) ranged from 23 to 36 J/m3 . The analysis of shear showed that all the three studied devices had minimal overall risk for thrombus formation. We conclude that the characteristics and material designs of TAVI devices have strong influences on the hemodynamics in the ascending aorta.

Comprehensive In Vitro Study of the Flow Past Two Transcatheter Aortic Valves: Comparison with a Severe Stenotic Case.

We investigate the flow past two transcatheter aortic valves (TAVs) and one severely calcified valve in an anatomically realistic aorta geometry to evaluate the ability of the TAVs to establish a healthier aortic flow compared to a diseased case. Velocity measurements of pulsatile flow are carried out using the 3D-particle tracking velocimetry technique. We present a novel approach based on the Smagorinsky model to assess the important subvoxel-scale (here smaller than 750 [Formula: see text]m) shear stress contribution that is usually unavailable in experiments. Both TAV models feature a small retrograde flow of about 5% of the stroke volume and a lower number of coherent vortical structures. Turbulence past the TAVs is strongly suppressed as evidenced by the lower levels of turbulent kinetic energy even though the newer generation TAV performs better than the old one. Also lysis indices are substantially reduced in both models. The new generation TAV displays a slightly higher risk for thrombogenicity due to longer exposure times. We anticipate that our new approach to include turbulence and shear stress related quantities may help to validate the design of cardiovascular devices.

A Novel Approach for 3D-Structural Identification through Video Recording: Magnified Tracking.

Advancements in optical imaging devices and computer vision algorithms allow the exploration of novel diagnostic techniques for use within engineering systems. A recent field of application lies in the adoption of such devices for non-contact vibrational response recordings of structures, allowing high spatial density measurements without the burden of heavy cabling associated with conventional technologies. This, however, is not a straightforward task due to the typically low-amplitude displacement response of structures under ambient operational conditions. A novel framework, namely Magnified Tracking (MT), is proposed herein to overcome this limitation through the synergistic use of two computer vision techniques. The recently proposed phase-based motion magnification (PBMM) framework, for amplifying motion in a video within a defined frequency band, is coupled with motion tracking by means of particle tracking velocimetry (PTV). An experimental campaign was conducted to validate a proof-of-concept, where the dynamic response of a shear frame was measured both by conventional sensors as well as a video camera setup, and cross-compared to prove the feasibility of the proposed non-contact approach. The methodology was explored both in 2D and 3D configurations, with PTV revealing a powerful tool for the measurement of perceptible motion. When MT is utilized for tracking "imperceptible" structural responses (i.e., below PTV sensitivity), via the use of PBMM around the resonant frequencies of the structure, the amplified motion reveals the operational deflection shapes, which are otherwise intractable. The modal results extracted from the magnified videos, using PTV, demonstrate MT to be a viable non-contact alternative for 3D modal identification with the benefit of a spatially dense measurement grid.

Hemodynamic Changes in the Right Ventricle Induced by Variations of Cardiac Output: A Possible Mechanism for Arrhythmia Occurrence in the Outflow Tract.

The rationale of this paper is to investigate right ventricular (RV) hemodynamics in relation to changes in cardiac output, and in particular to study exercise-induced stresses at the RV outflow tract (RVOT), which is a common site of ventricular arrhythmias in the athlete's heart. We hypothesize that the thin-walled RVOT is exposed to high wall shear stresses (WSS) during physiological states associated with high cardiac output such as exercise, and therefore, may be particularly prone to substrate formation leading to ventricular tachyarrhythmias. 3D Particle Tracking Velocimetry (3D-PTV), an optical imaging method, has been performed in a novel anatomically accurate compliant silicone right heart model derived from a high resolution MRI heart scan of a healthy male proband. RV and RVOT flow patterns at resting conditions were obtained from two healthy athletic male proband's hearts and two patients with arrhythmogenic right ventricular cardiomyopathy/dysplasia (ARVC/D) via phase contrast magnetic resonance imaging (PC-MRI). The healthy case was used as a reference for validating the in vitro flow patterns of the silicone model, while the diseased cases were used to generalize our findings and investigate possible changes in hemodynamic stresses with RV morphological remodelling. Our results showed that both healthy and diseased geometries consistently displayed an increased WSS in the RVOT relative to the rest of the RV. We found that increases in cardiac output may lead to increases of mean kinetic energy (MKE), laminar viscous dissipation and WSS at the RVOT. Furthermore, higher peak WSS magnitudes were found for the diseased cases. The identified high WSS regions may correlate with the common site of RVOT ventricular tachycardia in athletes and patients with ARVC/D. Our results imply that exercise, as well as anatomical and functional remodeling might alter RV wall shear stress both in magnitude and spatial distribution, leading to increased hemodynamic stresses in the RVOT.