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Search indexed bioRxiv preprints in genomics, neuroscience, cell biology and bioinformatics. Read source abstracts and check manuscript versions; preprints are not peer reviewed.

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At least 307 records · Page 17Linked to original sources
bioRxiv2017

Denitrifying metabolism of the methylotrophic marine bacterium Methylophaga nitratireducenticrescens strain JAM1

Methylophaga nitratireducenticrescens strain JAM1 is a methylotrophic, marine bacterium that was isolated from a denitrification reactor treating a closed-circuit seawater aquarium. It can sustain growth under anoxic conditions by reducing nitrate (NO3-) to nitrite (NO2-), which accumulates in the medium. These physiological traits are attributed to gene clusters that encode two dissimilatory nitrate reductases (NarGHJI). M. nitratireducenticrescens strain JAM1 also contains gene clusters encoding two putative nitric oxide reductase (NO) reductases and one putative nitrous oxide (N2O) reductase, suggesting that NO and N2O can be reduced by strain JAM1. In this study, we show that strain JAM1 can reduce NO to N2O and N2O to N2 and can sustain growth under anoxic conditions by reducing N2O as the sole electron acceptor. Although strain JAM1 lacks a gene encoding a dissimilatory copper-(NirK) or cytochrome cd1-type (NirS) NO2- reductase, NO3--amended strain JAM1 cultures produce N2O, representing up to 6% of the N-input. NO2- was shown to be the key intermediate of this production process. In NO3-amended cultures, we analyzed denitrification genes in succession of net N2O-production and -consumption phases at the gene expression level. These phases were found to correlate with changes in the expression levels of the NO reductase gene cnorB1 and nnrS, which indicated NO production in the cultures.\n\nImportanceBy showing that all the three denitrification reductases are active, this demonstrates that Methylophaga nitratireducenticrescens JAM1 is one of many bacteria species that maintain genes associated primarily with denitrification, but not necessarily related to the maintenance of the entire pathway. The reason to maintain such incomplete pathway could be related to the specific role of strain JAM1 in the denitrifying biofilm of a denitrification reactor from which it originates. The small production of N2O via NO in strain JAM1 did not involve Nar contrary to what was demonstrated in Escherichia coli. M. nitratireducenticrescens JAM1 is the only reported Methylophaga species that has the capacity to grow under anoxic conditions by using NO3- and N2O as sole electron acceptors for its growth. It is also one of a few marine methylotrophs that is studied at the physiological and genetic levels in relation to its capacity to perform denitrifying activities.

microbiology↗
bioRxiv2017

Human macrophages survive and adopt activated genotypes in living zebrafish

The inflammatory response, modulated both by tissue resident macrophages and recruited monocytes from peripheral blood, plays a critical role in human diseases such as cancer and neurodegenerative disorders. Here, we sought a model to interrogate human immune behavior in vivo. We determined that primary human monocytes and macrophages survive in zebrafish for up to two weeks. Flow cytometry revealed that human monocytes cultured at the physiological temperature of the zebrafish survive and differentiate comparable to cohorts cultured at human physiological temperature. Moreover, key genes that encode for proteins that play a role in tissue remodeling were also expressed. Human cells migrated within multiple tissues at speeds comparable to zebrafish macrophages. Analysis of gene expression of in vivo educated human macrophages confirmed expression of activated macrophage phenotypes. Here, human cells adopted phenotypes relevant to cancer progression, suggesting that we can define the real time immune modulation of human tumor cells during the establishment of a metastatic lesion in zebrafish.

developmental biology↗
bioRxiv2017

Tracking dynamic threat imminence

During real-life situations, multiple factors interact dynamically to determine threat level. In the current functional MRI study involving healthy adult human volunteers, we investigated interactions between proximity, direction (approach vs. retreat), and speed during a dynamic threat-of-shock paradigm. As a measure of threat-evoked physiological arousal, skin conductance responses were recorded during fMRI scanning. Whereas some brain regions tracked individual threat-related factors, others were also sensitive to combinations of these variables. In particular, signals in the anterior insula tracked the interaction between proximity and direction where approach vs. retreat responses were stronger when threat was closer compared to farther. A parallel proximity-by-direction interaction was also observed in physiological skin conductance responses. In the right amygdala, we observed a proximity by direction interaction, but intriguingly in the opposite direction as the anterior insula; retreat vs. approach responses were stronger when threat was closer compared to farther. In the right bed nucleus of the stria terminalis, we observed an effect of threat proximity, whereas in the right periaqueductal gray/midbrain we observed an effect of threat direction and a proximity by direction by speed interaction (the latter was detected in exploratory analyses but not in a voxelwise fashion). Together, our study refines our understanding of the brain mechanisms involved during aversive anticipation in the human brain. Importantly, it emphasizes that threat processing should be understood in a manner that is both context sensitive and dynamic.

neuroscience↗
bioRxiv2017

The lamellipodium is a myosin independent mechanosensor

The ability of adherent cells to sense changes in the mechanical properties of their extracellular environments is critical to numerous aspects of their physiology. It has been well documented that cell attachment and spreading are sensitive to substrate stiffness. Here we demonstrate that this behavior is actually biphasic, with a transition that occurs around a Youngs modulus of [~]7 kPa. Furthermore, we demonstrate that, contrary to established assumptions, this property is independent of myosin II activity. Rather, we find that cell spreading on soft substrates is inhibited due to reduced nascent adhesion formation within the lamellipodium. Cells on soft substrates display normal leading edge protrusion activity, but these protrusions are not stabilized due to impaired adhesion assembly. Enhancing integrin-ECM affinity through addition of Mn2+ recovers nascent adhesion assembly and cell spreading on soft substrates. Using a computational model to simulate nascent adhesion assembly, we find that biophysical properties of the integrin-ECM bond are optimized to stabilize interactions above a threshold matrix stiffness that is consistent with the experimentally observations. Together these results suggest that myosin II-independent forces in the lamellipodium are responsible for mechanosensation by regulating new adhesion assembly, which in turn, directly controls cell spreading. This myosin II-independent mechanism of substrate stiffness sensing could potentially regulate a number of other stiffness sensitive processes.\n\nSignificance StatementCell physiology can be regulated by the mechanics of the extracellular environment. Here, we demonstrate that cell spreading is a mechanosensitive process regulated by weak forces generated at the cell periphery and independent of motor activity. We show that stiffness sensing depends on the kinetics of the initial adhesion bonds that are subjected to forces driven by protein polymerization. This work demonstrates how the binding kinetics of adhesion molecules are sensitively tuned to a range of forces that enable mechanosensation.

biophysics↗
bioRxiv2017

Generation and validation of homozygous fluorescent knock-in cells using genome editing

Gene tagging with fluorescent proteins is essential to investigate the dynamic properties of cellular proteins. CRISPR/Cas9 technology is a powerful tool for inserting fluorescent markers into all alleles of the gene of interest (GOI) and permits functionality and physiological expression of the fusion protein. It is essential to evaluate such genome-edited cell lines carefully in order to preclude off-target effects caused by either (i) incorrect insertion of the fluorescent protein, (ii) perturbation of the fusion protein by the fluorescent proteins or (iii) non-specific genomic DNA damage by CRISPR/Cas9. In this protocol1, we provide a step-by-step description of our systematic pipeline to generate and validate homozygous fluorescent knock-in cell lines.\n\nWe have used the paired Cas9D10A nickase approach to efficiently insert tags into specific genomic loci via homology-directed repair with minimal off-target effects. It is time- and cost-consuming to perform whole genome sequencing of each cell clone. Therefore, we have developed an efficient validation pipeline of the generated cell lines consisting of junction PCR, Southern Blot analysis, Sanger sequencing, microscopy, Western blot analysis and live cell imaging for cell cycle dynamics. This protocol takes between 6-9 weeks. Using this protocol, up to 70% of the targeted genes can be tagged homozygously with fluorescent proteins and result in physiological levels and phenotypically functional expression of the fusion proteins.\n\nEditorial SummaryThis protocol provides a detailed workflow describing how to insert fluorescent markers into all alleles of a gene of interest using CRISPR/Cas 9 technology and how to generate and validate homozygous fluorescent knock-in cell lines.

molecular biology↗
bioRxiv2017

Reactive astrocytes - comprehending when neurons play 4’33”

Homeostatic regulation is a powerful tool utilized by virtually all biological systems, brain included. Broadly speaking, each homeostatic process embodies two components: the sensing component, whereby a deviation is detected and quantified, and the effecting component that executes the homeostatic adjustment with the goal of alleviating the deviation. In the central nervous system, homeostatic plasticity has been suggested to play an important role in shaping the dynamics of single neurons and neuronal networks. However, existing \"biophysical\" models of homeostatic plasticity are exceedingly simplistic. These models usually describe the sensor component in terms of simple averaging over neuronal activity and offer no explanation of the relevant biochemical pathways. Here, we attempt to fill this gap in our understanding of homeostatic plasticity by proposing a biophysical framework to explain detection of prolonged synaptic inactivity that may occur in some scenarios of brain injury. We propose that sensing of, and response to, synaptic inactivity involves detection of the extracellular glutamate level and occurs via the activation of metabotropic glutamate receptors (mGluRs), while the inactivity-induced synthesis of one of the homeostatic plasticity effectors, tumor necrosis factor alpha (TNF), serves as an effecting component of the system. This model can help to explain the experimental observations linking prolonged neuronal inactivity to TNF signaling. Importantly, the proposed signaling scheme is not limited to mGluRs and astrocytes, but rather is potentially applicable to any cells expressing receptors that activate the relevant G protein units. The proposed signaling scheme is likely to be useful for developing pharmacological interventions targeting homeostatic plasticity pathways.\n\nAuthor summaryHomeostatic regulation refers to the self-regulation ability of a system aimed at remaining in the same (or nearly the same) state. Homeostatic plasticity, a form of homeostatic regulation that arises in the context of neural dynamics, has been shown to shape neuronal and network dynamics by attempting to maintain physiological levels of neuronal activity in brain networks. Thus, it changes the excitation-inhibition balance in a network when sufficiently large and long-lasting deviations from physiological levels of activity are detected. However, the biophysical mechanisms of homeostatic plasticity and in particular those related to the ability to sense neuronal inactivity, remain elusive. We propose a feasible biophysical model of a homeostatic sensor, in which the sensing of inactivity depends on the presence and activation of G-protein-coupled receptors with different activation thresholds. In this model, the activation of the higher-threshold receptor suppresses the detection of inactivity, while the activation of the lower-threshold receptor promotes the detection of inactivity. The proposed model helps to explain a growing body of experimental data relating synaptic inactivity to production of homeostatic plasticity effectors such as tumor necrosis factor alpha. Although we consider a specific case study of metabotropic glutamate receptors on astrocytes, the model conclusions are likely to be applicable to any cells expressing receptors that activate the relevant G protein units.

neuroscience↗
bioRxiv2017

Symbiodinium functional diversity and clade specificity under global change stressors

Coral bleaching episodes are increasing in frequency, demanding examination of the physiological and molecular responses of corals and their Symbiodinium to climate change. Here we quantify bleaching and Symbiodinium photosynthetic performance of Siderastrea siderea from two reef zones after long-term exposure to thermal and CO2-acidification stress. Molecular response of in hospite Symbiodinium to these stressors was interrogated with RNAseq. Elevated temperatures reduced photosynthetic efficiency, which was highly correlated with bleaching status. However, photosynthetic efficiencies of forereef symbionts were more negatively affected by thermal stress than nearshore symbionts, indicating greater thermal tolerance in nearshore corals. At control temperatures, CO2-acidification had little effect on symbiont physiology, although forereef symbionts exhibited greater photosynthetic efficiencies than nearshore symbionts. Transcriptome profiling revealed that S. siderea were dominated by clade C Symbiodinium, except under thermal stress, which caused shifts to thermotolerant clade D. Comparative transcriptomics of conserved genes across symbiotic partners revealed few differentially expressed Symbiodinium genes when compared to corals. Instead of responding to stress, clade C transcriptomes varied by reef zone, with forereef Symbiodinium exhibiting enrichment of genes associated with photosynthesis. Our findings suggest that functional variation in photosynthetic architecture exists between forereef and nearshore Symbiodinium populations.

ecology↗
bioRxiv2017

MICCS: A Fully Programmable Multipurpose Integrated Cell Culture System

With the rise of research utilizing microphysiological systems (MPSs), the need for tools that enable the physiological mimicking of the relevant cellular environment is vital. The limited ability to reproduce crucial features of the microenvironment, such as surrounding fluid flow and dynamic changes in biochemical stimuli, severely limits the types of experiments that can be carried out. Current equipment to achieve this, such as syringe and peristaltic pumps, is expensive, large, difficult to program and has limited potential for scalability. Here, we present a new pumping platform that is open-source, low-cost, modular, scalable, fully-programmable and easy to assemble that can be incorporated into cell culture systems to better recapitulate physiological environments. By controlling two commercially available piezoelectric pumps using a Raspberry Pi Zero microcontroller, the system is capable of producing arbitrary dynamic flow profiles with reliable flow rates ranging from 1 to 3,000 {micro}L/min as specified by an easily programmable Python-based script. We validated the accuracy of the flow rates, the use of time-varying profiles, and the practicality of the system by creating repeatable dynamic concentration profiles using a 3D-printed static micromixer.

bioengineering↗
bioRxiv2017

Natural changes in light interact with circadian regulation at promoters to control gene expression in cyanobacteria

The circadian clock interacts with other regulatory pathways to tune physiology to predictable daily changes and unexpected environmental fluctuations. However, the complexity of circadian clocks in higher organisms has prevented a clear understanding of how natural environmental conditions affect circadian clocks and their physiological outputs. Here, we dissect the interaction between circadian regulation and responses to fluctuating light in the cyanobacterium Synechococcus elongatus. We demonstrate that natural changes in light intensity substantially affect the expression of hundreds of circadian-clock-controlled genes, many of which are involved in key steps in metabolism. These changes in expression arise from control of RNA polymerase recruitment to promoters by circadian and light-responsive regulation of a network of transcription factors including RpaA and RpaB. Using phenomenological modeling constrained by our data, we reveal simple principles that underlie the small number of stereotyped responses of dusk circadian genes to changes in light.

bioinformatics↗
bioRxiv2017

Using Temporal ICA to Selectively Remove Global Noise While Preserving Global Signal in Functional MRI Data

Temporal fluctuations in functional Magnetic Resonance Imaging (fMRI) have been profitably used to study brain activity and connectivity for over two decades. Unfortunately, fMRI data also contain structured temporal \"noise\" from a variety of sources, including subject motion, subject physiology, and the MRI equipment. Recently, methods have been developed to automatically and selectively remove spatially specific structured noise from fMRI data using spatial Independent Components Analysis (ICA) and machine learning classifiers. Spatial ICA is particularly effective at removing spatially specific structured noise from high temporal and spatial resolution fMRI data of the type acquired by the Human Connectome Project and similar studies. However, spatial ICA is mathematically, by design, unable to separate spatially widespread \"global\" structured noise from fMRI data (e.g., blood flow modulations from subject respiration). No methods currently exist to selectively and completely remove global structured noise while retaining the global signal from neural activity. This has left the field in a quandary--to do or not to do global signal regression--given that both choices have substantial downsides. Here we show that temporal ICA can selectively segregate and remove global structured noise while retaining global neural signal in both task-based and resting state fMRI data. We compare the results before and after temporal ICA cleanup to those from global signal regression and show that temporal ICA cleanup removes the global positive biases caused by global physiological noise without inducing the network-specific negative biases of global signal regression. We believe that temporal ICA cleanup provides a \"best of both worlds\" solution to the global signal and global noise dilemma and that temporal ICA itself unlocks interesting neurobiological insights from fMRI data.

neuroscience↗
bioRxiv2017

Peatland Acidobacteria with a dissimilatory sulfur metabolism

Sulfur-cycling microorganisms impact organic matter decomposition in wetlands and consequently greenhouse gas emissions from these globally relevant environments. However,their identities and physiological properties are largely unknown. By applying a functional metagenomics approach to an acidic peatland, we recovered draft genomes of seven novel Acidobacteria species with the potential for dissimilatory sulfite (dsrAB, dsrC, dsrD, dsrN, dsrT, dsrMKJOP) or sulfate respiration (sat, aprBA, qmoABC plus dsr genes). Surprisingly, the genomes also encoded dsrL, a unique gene of the sulfur oxidation pathway. Metatranscriptome analysis demonstrated expression of acidobacterial sulfur-metabolism genes in native peat soil and their upregulation in diverse anoxic microcosms. This indicated an active sulfate respiration pathway, which, however, could also operate in reverse for sulfur oxidation as recently shown for other microorganisms. Acidobacteria that only harbored genes for sulfite reduction additionally encoded enzymes that liberate sulfite from organosulfonates, which suggested organic sulfur compounds as complementary energy sources. Further metabolic potentials included polysaccharide hydrolysis and sugar utilization, aerobic respiration, several fermentative capabilities, and hydrogen oxidation. Our findings extend both, the known physiological and genetic properties of Acidobacteria and the known taxonomic diversity of microorganisms with a DsrAB-based sulfur metabolism, and highlight new fundamental niches for facultative anaerobic Acidobacteria in wetlands based on exploitation of inorganic and organic sulfur molecules for energy conservation.

microbiology↗
bioRxiv2017

Control of Cardiac Mitochondrial Fuel Selection by Calcium

Calcium ion concentration modulates the function of pyruvate dehydrogenase, isocitrate dehydrogenase, and -ketoglutarate dehydrogenase. Previous studies have shown that despite its ability to affect the function of these dehydrogenases, [Ca2+] does not substantially alter mitochondrial ATP synthesis in vitro under physiological sub-strate conditions. We hypothesize that, rather than contributing to respiratory control, [Ca2+] governs fuel selection. Specifically, cardiac mitochondria are able to use different primary carbon substrates to synthesize ATP aerobically. To determine if and how [Ca2+] affects the relative use of carbohydrates versus fatty acids we measured oxygen consumption and tricarboxylic acid cycle intermediate concentrations in suspensions of cardiac mitochondria with different combinations of pyruvate and palmitoyl-L-carnitine in the media at various [Ca2+] and ADP infusion rates. Results reveal that when both fatty acid and carbohydrate substrates are available, fuel selection is sensitive to both calcium and ATP synthesis rate. When no Ca2+ is added under low ATP-demand conditions, {beta}-oxidation provides roughly half of acetyl-CoA for the citrate synthase reaction with the rest coming from the pyruvate dehydrogenase reaction. Under low demand conditions with increasing [Ca2+], the fuel utilization ratio shifts to increased fractional consumption of pyruvate, with 83{+/-}10% of acetyl-CoA derived from pyruvate at the highest [Ca2+] evaluated. With high ATP demand, the majority of acetyl-CoA is derived from pyruvate, regardless of the Ca2+ level. Our results suggest that changes in work rate alone are enough to effect a switch to carbohydrate use while in vivo the rate at which this switch happens may depend on mitochondrial calcium.\n\nKey PointsO_LIDespite its effects on activity of mitochondrial dehydrogenases, Ca2+ does not substantially alter mitochondrial ATP synthesis in vitro under physiological substrate conditions. Nor does is appear to play an important role in respiratory control in vivo in the myocardium.\nC_LIO_LIWe hypothesize that Ca2+ plays a role mediating the switch in fuel selection to increasing carbohydrate oxidation and decreasing fatty acid oxidation with increasing work rate.\nC_LIO_LITo determine if and how Ca2+ affects the relative use of carbohydrates versus fatty acids in vitro we measured oxygen consumption and TCA cycle intermediate concentrations in suspensions of purified rat ventricular mitochondria with carbohydrate, fatty acid, and mixed substrates at various [Ca2+] and ATP demand rates.\nC_LIO_LIOur results suggest that changes in work rate alone are enough to effect a switch to carbohydrate use in vitro while in vivo the rate at which this switch happens may depend on mitochondrial calcium.\nC_LI

biochemistry↗
bioRxiv2017

The Generation and Propagation of the Human Alpha Rhythm

IntroductionThe alpha rhythm (7-13 Hz) is the longest studied brain oscillation and has been theorized to play a key role in cognition. Still, substantial uncertainty remains over its physiology. In this study, we used micro and macro electrodes in patients undergoing surgery for epilepsy to measure the intracortical and thalamic generators of the human alpha rhythm. We first found that alpha propagates from higher-order anterosuperior cortex towards the lower-order occipital poles, consistent with alpha effecting top-down processing. This cortical alpha drives thalamic alpha, reversing prevailing theories of a thalamic alpha pacemaker. Finally, alpha is dominated by currents and firing in supragranular cortex, contravening the popular conception of alpha as an infragranular rhythm. Together, these results demonstrate that the alpha rhythm reflects short-range supragranular feedback which propagates from higher-order to lower order cortex and cortex to thalamus. These physiological insights explain how alpha could mediate feedback throughout the thalamocortical system.

neuroscience↗
bioRxiv2017

HDAC11 regulates type I interferon signaling through defatty-acylation of SHMT2

The smallest histone deacetylase (HDAC) and the only class IV HDAC member, HDAC11, is reported to regulate immune activation and tumorigenesis, yet its physiological function is largely unknown. Here we identify HDAC11 as an efficient lysine defatty-acylase that is >10,000-fold more efficient than its deacetylase activity. Through proteomics studies, we identified SHMT2 as a defatty-acylation substrate of HDAC11. HDAC11-catalyzed defatty-acylation did not affect the enzymatic activity of SHMT2. Instead, it affects the ability of SHMT2 to regulate type I interferon receptor ubiquitination and internalization. Correspondingly, HDAC11 depletion increased type I interferon signaling in both cell culture and mice. This study is the first time a zinc-dependent HDAC is found to have an activity that is much more efficient than the corresponding deacetylase activity. The finding expands the physiological functions of HDAC11 and protein lysine fatty acylation, and opens up opportunities to develop HDAC11-specific inhibitors as therapeutics to modulate immune responses.

biochemistry↗
bioRxiv2017

Circuit-based models of shared variability in cortical networks

Trial-to-trial variability is a reflection of the circuitry and cellular physiology that makeup a neuronal network. A pervasive yet puzzling feature of cortical circuits is that despite their complex wiring, population-wide shared spiking variability is low dimensional with all neurons fluctuating en masse. Previous model cortical networks are at loss to explain this global variability, and rather assume it is from external sources. We show that if the spatial and temporal scales of inhibitory coupling match known physiology, model spiking neurons internally generate low dimensional shared variability that captures the properties of in vivo population recordings along the visual pathway. Shifting spatial attention into the receptive field of visual neurons has been shown to reduce low dimensional shared variability within a brain area, yet increase the variability shared between areas. A top-down modulation of inhibitory neurons in our network provides a parsimonious mechanism for this attentional modulation, providing support for our theory of cortical variability. Our work provides a critical and previously missing mechanistic link between observed cortical circuit structure and realistic population-wide shared neuronal variability and its modulation.

neuroscience↗
bioRxiv2017

CHRAC/ACF Contribute to the Repressive Ground State of Chromatin

The chromatin remodeling complexes CHRAC and ACF combine the ATPase ISWI with the signature subunit ACF1. These enzymes catalyze well-studied nucleosome sliding reactions in vitro, but how their actions affect physiological gene expression is unclear. Here we explored the influence of Drosophila CHRAC/ACF on transcription by complementary gain- and loss-of-function approaches.\n\nTargeting ACF1 to multiple reporter genes inserted at many different genomic locations revealed a context-dependent inactivation of poorly transcribed reporters in repressive chromatin. Accordingly, single-embryo transcriptome analysis of a Acf knock-out allele showed that only lowly expressed genes are de-repressed in the absence of ACF1. Finally, the nucleosome arrays in Acf-deficient chromatin show loss of physiological regularity, particularly in transcriptionally inactive domains.\n\nTaken together our results highlight that ACF1-containing remodeling factors contribute to the establishment of an inactive ground state of the genome through chromatin organization.

molecular biology↗
bioRxiv2017

Extracting biological age from biomedical data via deep learning: too much of a good thing?

Aging-related physiological changes are systemic and, at least in humans, are linearly associated with age. Therefore, linear combinations of physiological measures trained to estimate chronological age have recently emerged as a practical way to quantify aging in the form of biological age. Aging acceleration, defined as the difference between the predicted and chronological age was found to be elevated in patients with major diseases and is predictive of mortality. In this work, we compare three increasingly accurate biological age models: metrics derived from unsupervised Principal Components Analysis (PCA), alongside two supervised biological age models; a multivariate linear regression and a state-of-the-art deep convolution neural network (CNN). All predictions were made using one-week long locomotor activity records from a 2003-2006 National Health and Nutrition Examination Survey (NHANES) dataset. We found that application of the supervised approaches improves the accuracy of the chronological age estimation at the expense of a loss of the association between the aging acceleration predicted by the model and all-cause mortality. Instead, we turned to the NHANES death register and introduced a novel way to train parametric proportional hazards models in a form suitable for out-of-the-box implementation with any modern machine learning software. Finally, we characterized a proof-of-concept example, a separate deep CNN trained to predict mortality risks that outperformed any of the biological age or simple linear proportional hazards models. Our findings demonstrate the emerging potential of combined wearable sensors and deep learning technologies for applications involving continuous health risk monitoring and real-time feedback to patients and care providers.

bioinformatics↗
bioRxiv2017

Engineering Human CNS Morphogenesis: Controlled Induction of Singular Neural Rosette Emergence

Human pluripotent stem cell (hPSC)-derived neural organoids have revolutionized in vitro modelling of human neurological disorders. Cell-intrinsic morphogenesis processes displayed within these tissues could serve as the basis for ex vivo manufacture of brain and spinal cord tissues with biomimetic anatomy and physiology. However, we must first understand how to control their emergent properties starting at the genesis of neural organoid formation, i.e. emergence of polarized neuroepithelium. In vivo, all CNS tissues develop from a singular neuroepithelial tube. Yet, current protocols yield organoids with multiple neuroepithelial rings, a.k.a. neural rosettes, each acting as independent centers of morphogenesis and thereby impeding coordinate tissue development. We discovered that the morphology of hPSC-derived neural tissues is a critical biophysical parameter for inducing singular neural rosette emergence. Tissue morphology screens conducted using micropatterned array substrates and an automated image analysis determined that circular morphologies of 200-250 and 150m diameter are optimal for inducing singular neural rosette emergence within 80-85% forebrain and 73.5% spinal tissues, respectively. The discrepancy in optimal circular morphology for Pax6+/N-cadherin+ neuroepithelial forebrain versus spinal tissues was due to previously unknown differences in ROCK-mediated cell contractility. The singular neuroepithelium induced within geometrically confined tissues persisted as the tissues morphed from a 2-D monolayer to multilayered 3-D hemispherical aggregate. Upon confinement release using clickable micropatterned substrates, the tissue displayed radial outgrowth with maintenance of a singular neuroepithelium and peripheral neuronal differentiation. Thus, we have quantitatively defined a pertinent biophysical parameter for effectively inducing a singular neuroepithelium emergence within morphing hPSC-derived neural tissues.\n\nSignificance StatementHuman pluripotent stem cell (hPSC)-derived neural organoids display emergent properties that, if harnessed, could serve as the basis for ex vivo manufacture of brain and spinal cord tissues with biomimetic macroscale anatomy and physiology. Their chaotic terminal structure arises from uncontrolled morphogenesis at their genesis, resulting in spontaneous induction of multiple neuroepithelial morphogenesis centers,a.k.a. neural rosettes. Here, we determined that neural tissue morphology is a pertinent biophysical parameter for controlling subsequent morphogenesis, and identified discrete circular tissue morphologies as optimal and effective at inducing singular neural rosette emergence within forebrain and spinal neural tissues. Thus, we developed an approach to reproducibly control the initial stage of hPSC-derived neural tissue morphogenesis enabling their manufacture with a biomimetic nascent CNS anatomy.

bioengineering↗