Substrate Concentration Gradient Research Articles

Local Self-Assembly of Dissipative Structures Sustained by Substrate Diffusion.

The coupling between energy-consuming molecular processes and the macroscopic dimension plays an important role in nature and in the development of active matter. Here, we study the temporal evolution of a macroscopic system upon the local activation of a dissipative self-assembly process. Injection of surfactant molecules in a substrate-containing hydrogel results in the local substrate-templated formation of assemblies, which are catalysts for the conversion of substrate into waste. We show that the system develops into a macroscopic (pseudo-)non-equilibrium steady state (NESS) characterized by the local presence of energy-dissipating assemblies and persistent substrate and waste concentration gradients. For elevated substrate concentrations, this state can be maintained for more than 4 days. The studies reveal an interdependence between the dissipative assemblies and the concentration gradients: catalytic activity by the assemblies results in sustained concentration gradients and, vice versa, continuous diffusion of substrate to the assemblies stabilizes their size. The possibility to activate dissipative processes with spatial control and create long lasting non-equilibrium steady states enables dissipative structures to be studied in the space-time domain, which is of relevance for understanding biological systems and for the development of active matter.

Local Self‐Assembly of Dissipative Structures Sustained by Substrate Diffusion

The coupling between energy‐consuming molecular processes and the macroscopic dimension plays an important role in nature and in the development of active matter. Here, we study the temporal evolution of a macroscopic system upon the local activation of a dissipative self‐assembly process. Injection of surfactant molecules in a substrate‐containing hydrogel results in the local substrate‐templated formation of assemblies, which are catalysts for the conversion of substrate into waste. We show that the system develops into a macroscopic (pseudo‐)non‐equilibrium steady state (NESS) characterized by the local presence of energy‐dissipating assemblies and persistent substrate and waste concentration gradients. For elevated substrate concentrations, this state can be maintained for more than 4 days. The studies reveal an interdependence between the dissipative assemblies and the concentration gradients: catalytic activity by the assemblies results in sustained concentration gradients and, vice versa, continuous diffusion of substrate to the assemblies stabilizes their size. The possibility to activate dissipative processes with spatial control and create long lasting non‐equilibrium steady states enables dissipative structures to be studied in the space‐time domain, which is of relevance for understanding biological systems and for the development of active matter.

Numerical modeling of senile plaque development under conditions of limited diffusivity of amyloid-β monomers

This paper introduces a new model to simulate the progression of senile plaques, focusing on scenarios where concentrations of amyloid beta (Aβ) monomers and aggregates vary between neurons. Extracellular variations in these concentrations may arise due to limited diffusivity of Aβ monomers and a high rate of Aβ monomer production at lipid membranes, requiring a substantial concentration gradient for diffusion-driven transport of Aβ monomers. The dimensionless formulation of the model is presented, which identifies four key dimensionless parameters governing the solutions for Aβ monomer and aggregate concentrations, as well as the radius of a growing Aβ plaque within the control volume. These parameters include the dimensionless diffusivity of Aβ monomers, the dimensionless rate of Aβ monomer production, and the dimensionless half-lives of Aβ monomers and aggregates. A dimensionless parameter is then introduced to evaluate the validity of the lumped capacitance approximation. An approximate solution is derived for the scenario involving large diffusivity of Aβ monomers and dysfunctional protein degradation machinery, resulting in infinitely long half-lives for Aβ monomers and aggregates. In this scenario, the concentrations of Aβ aggregates and the radius of the Aβ plaque depend solely on a single dimensionless parameter that characterizes the rate of Aβ monomer production. According to the approximate solution, the concentration of Aβ aggregates is linearly dependent on the rate of monomer production, and the radius of an Aβ plaque is directly proportional to the cube root of the rate of monomer production. However, when departing from the conditions of the approximate solution (e.g., finite half-lives), the concentrations of Aβ monomers and aggregates, along with the plaque radius, exhibit complex dependencies on all four dimensionless parameters. For instance, under physiological half-life conditions, the plaque radius reaches a maximum value and stabilizes thereafter.

Interaction-motif-based classification of self-organizing metabolic cycles

Particles that are catalytically-active and chemotactic can interact through the concentration fields upon which they act, which in turn may lead to wide-scale spatial self-organization. When these active particles interact through several fields, these interactions gain an additional structure, which can result in new forms of collective behavior. Here, we study a mixture of active species which catalyze the conversion of a substrate chemical into a product chemical, and chemotax in concentration gradients of both substrate and product. Such species develop non-reciprocal, specific interactions that we coarse-grain into attractive and repulsive, which can lead to a potentially complex interaction network. We consider the particular case of a metabolic cycle of three species, each of which interacts with itself and both other species in the cycle. We find that the stability of a cycle of species that only chemotax in gradients of their substrate is piloted by a set of two parameter-free conditions, which we use to classify the low number of corresponding interaction networks. In the more general case of substrate- and product-chemotactic species, we can derive a set of two high-dimensional stability conditions, which can be used to classify the stability of all the possible interaction networks based on the self- and pair-interaction motifs they contain. The classification scheme that we introduce can help guide future studies on the dynamics of complex interaction networks and explorations of the corresponding large parameter spaces in such metabolically active complex systems.

Predictive effect of the decline in CD4+ T cell levels in blood on infection in patients with severe hemorrhagic stroke and mechanism

Objective The purpose of this research was to evaluate the influence of immunity on infection in patients with severe hemorrhagic stroke and explore the mechanism underlying this connection. Methods Clinical data obtained from 126 patients with severe hemorrhagic stroke were retrospectively analyzed, and the factors affecting infection were screened by multivariable logistic regression models. Nomograms, calibration curves, the Hosmer–Lemeshow goodness-of-fit test, and decision curve analysis were used to examine the effectiveness of the models in evaluating infection. The mechanism underlying the reduction in CD4 + T-cell levels in blood was explored by analysis of lymphocyte subsets and cytokines in cerebrospinal fluid (CSF) and blood. Results The results showed that CD4 + T-cell levels of <300/μL was an independent risk factor for early infection. The models for multivariable logistic regression involving the CD4 + T-cell levels and other influencing factors had good applicability and effectiveness in evaluating early infection. CD4 + T-cell levels decreased in blood but increased in CSF. Similarly, interleukin (IL)-6 and IL-8 levels in CSF had a significant increase, generating a substantial concentration gradient between the CSF and the blood. Conclusion Reduced blood CD4 + T-cell counts among patients who had severe hemorrhagic stroke increased the risk of early infection. CSF IL-6 and IL-8 may be involved in inducing the migration of CD4 + T cells into the CSF and decreasing blood CD4 + T-cell levels.

Hydrolysis capacity of different sized granules in a full-scale aerobic granular sludge (AGS) reactor

In aerobic granular sludge (AGS) reactors, granules of different sizes coexist in a single reactor. Their differences in settling behaviour cause stratification in the settled granule bed. In combination with substrate concentration gradients over the reactor height during the anaerobic plug-flow feeding regime, this can result in functional differences between granule sizes. In this study, we compared the hydrolytic activity in granules of 4 size ranges (between 0.5 and 4.8 mm diameter) collected from a full-scale AGS installation. Protease and amylase activities were quantified through fluorescent activity assays. To visualise where the hydrolytic active zones were located within the granules, the hydrolysis sites were visualized microscopically after incubating intact and sliced granules with fluorescent casein and starch. The microbial community was studied using fluorescent in situ hybridization (FISH) and sequencing. The results of these assays indicated that hydrolytic capacity was present throughout the granules, but the hydrolysis of bulk substrates was restricted to the outer 100 µm, approximately. Many of the microorganisms studied by FISH, such as polyphosphate and glycogen accumulating organisms (PAO and GAO), were abundant in the vicinity of the hydrolytically active sites. The biomass-specific hydrolysis rate depended mainly on the available granule surface area, suggesting that different sized granules are not differentiated in terms of hydrolytic capacity. Thus, the substrate concentration gradients that are present during the anaerobic feeding in AGS reactors do not seem to affect hydrolytic activity at the granule surfaces. In this paper, we discuss the possible reasons for this and reflect about the implications for AGS technology.

Sludge granulation in ASBR: Reactor performance, sludge physiochemical properties evolution and the unique clustered structure of the granular sludge

Anaerobic granular sludge was successfully cultivated in the anaerobic sequencing batch reactor (ASBR) after 45 days of operation. The results showed that the mature granular sludge exhibited better physicochemical characteristics: granule percentage of 62.6 ± 8.8 %, 0.8–1.2 mm in diameter, with smooth edges and regular shapes, and sludge volume index of 20.0 mL/g SS. 16S rRNA-targeted fluorescence in situ hybridisation (FISH) and scanning electron microscope (SEM) examination revealed that the granular sludge had a unique clustered microbial community structure, which would to be an optimal microbial arrangement for syntrophic colonies. The large substrate concentration gradient that derived from the feast-famine alternation regime of the substrate in a running cycle of ASBR was the major cause of the clustered microbial distribution structure. Lots of syntrophic consortia formed by acetogens and methanogens randomly distributed throughout the interior of the granules, furthermore, the cavities existing amongst the syntrophic consortia were convenient for substrate transfer and biogas liberation, thus enhancing the digestion performance of ASBR and preventing granule disintegration caused by gas scour. With sludge granulation, the methane yield of ASBR increased from the initial 0.23 L/g COD to 0.28 L/g COD, meanwhile, the maximum specific methanogenic activities (SMAs) of acetoclastic and hydrogenotrophic methanogens respectively increased from 0.51 gCOD/(gVSS·d) to 0.73 gCOD/(gVSS·d) and from 0.35 gCOD/(gVSS·d) to 0.62 gCOD/(gVSS·d). The exploration of granular sludge formation, sludge characteristic evolution and treatment performance in ASBR reactor has theoretical value for its engineering promotion.

Shifts of microbial community structure along substrate concentration gradients in immobilized biomass for nitrogen removal

Immobilized biomass technology has been regarded as an effective strategy to enhance simultaneous nitrification and denitrification (SND) in existing aerobic biological wastewater treatment processes. Nevertheless, the mechanisms of SND in an aerobic immobilized biomass need to be proven. In this study, waste sludge from municipal wastewater treatment plants was immobilized by cellulose triacetate as bioplates, and an immobilized bioplate reactor (IBPR) was successfully established for nitrogen removal tests. The SND efficiency of the IBPR was increased 18% under the intermittent aeration (IA) mode compared with that under the continuous aeration (CA) mode. During IA operation, the IBPR achieved 96% COD removal and 76% NH4+-N removal, with 71% SND. The results of microbial community analysis by high-throughput sequencing showed that nitrogen-related functional bacteria were more abundant in the bioplates than in the attached biofilms. The colocalization of nitrifiers and denitrifiers in the bioplates was observed, and the microbial community of nitrogen-related functional bacteria clearly shifted with the substrate concentration gradients.

Assessing CDK5 as a Nanomotor for Chemotactic Drug Delivery

Enzyme chemotaxis is a thermodynamic phenomenon in which enzymes move along a substrate concentration gradient towards regions with higher substrate concentrations. Mathematical models have been developed to understand and define this phenomenon using Michaelis-Menten kinetics and substrate binding coefficients. One key application of this phenomenon is in the field of drug delivery, where chemotaxis can steer nanovehicles towards targets along natural substrate concentrations. Such gradients exist throughout the body, including around the blood-brain barrier, and thus targeted nanovehicles can bring therapeutics through such a gradient and target themselves at a given toxic species. In patients with Alzheimer’s disease, a gradient of tau protein forms in the bloodstream, and this protein is a substrate of the enzyme CDK5, a tau protein kinase that catalyzes the phosphorylation of tau protein. Through chemotaxis, CDK5 can travel along the tau protein gradient towards increasing concentrations of tau tangles and amyloid-beta proteins that are the cause of neuronal cell death in Alzheimer’s patients. In order to create the momentum necessary to propel a nanovehicle along the substrate concentration gradient, an enzyme’s binding affinity towards its substrate must overcome obstacles such as Brownian motion. We hypothesized that CDK5 would be able to overcome these barriers and developed a quantitative model using Michaelis-Menten kinetics to define the necessary parameters to confirm and characterize CDK5’s chemotactic behavior to establish its utility in drug delivery and other applications. Our results found that CDK5’s movement significantly outpaced random motion and characterized its movement for usage in drug delivery system design.

Development of a microfluidic cell culture system for the adaptive laboratory evolution

We construct a microfluidics-based long-term cell culture system that can perform multiple adaptive laboratory evolution (ALE) experiments in parallel under different conditions. Single E. coli cells were isolated into water droplets in oil (W/O emulsion) containing agarose gel. Micrometric gel balls (GBs) were then extracted from the oil phase and placed in a microfluidic perfusion culture device, which can generate a substrate concentration gradient. The growth of the clonal colonies was monitored under each condition. We tested this GB-based culture device to expand the application of ALE after the gene knockout based on the flux balance analysis (FBA) performed in our previous studies.

Shear localization in large amplitude oscillatory shear (LAOS) flows of particulate suspensions

Significant shear localization, observed as substantial concentration gradients, is discovered in large amplitude oscillatory shear (LAOS) simulations of a complex fluid in a planar Couette setup. The localization is demonstrated to occur due to the inertial effects imposed by the oscillatory shear in LAOS, which serve as perturbations in the nonlinear equations describing the structural response of the fluid to shear. Additionally, a criterion for the onset of this shear localization is presented.

We Are One: Multispecies Metabolism of a Biofilm Consortium and Their Treatment Strategies.

The ecological and medical significance of bacterial biofilms have been well recognized. Biofilms are harder to control than their planktonic free-living counterparts and quite recently, the focus of the study has shifted to the multispecies consortia, which represent the vast majority of real-case infection scenarios. Studies have begun to explore the complex interspecies interactions within these biofilms. However, only little attention is currently given to the role of cellular metabolites in the cell-to-cell communication. The concentration gradients of metabolic substrates and products affect the spatial growth of bacteria in multispecies biofilm. This, if looked into more deeply, can lead to identification of potential therapies targeting the specific metabolites and hence the coordinated protection in the bacterial community. Herein, we review the interspecies communications, including their metabolic cross-talking, in multispecies biofilm, to signify the importance of such interactions on the initial formation and subsequent growth of these biofilms. Multispecies biofilms with their species heterogeneity are more resilient to antimicrobial agents than their single species biofilm counterparts and this characteristic is of particular interest when dealing with pathogenic bacteria. In this Review, we also discuss the treatment options available, to include current and emerging avenues to combat pathogenic multispecies biofilms in the clinical, environmental, as well as industrial settings.

Lactate Attenuates Synaptic Transmission and Affects Brain Rhythms Featuring High Energy Expenditure.

SummaryLactate shuttled from blood, astrocytes, and/or oligodendrocytes may serve as the major glucose alternative in brain energy metabolism. However, its effectiveness in fueling neuronal information processing underlying complex cortex functions like perception and memory is unclear. We show that sole lactate disturbs electrical gamma and theta-gamma oscillations in hippocampal networks by either attenuation or neural bursts. Bursting is suppressed by elevating the glucose fraction in substrate supply. By contrast, lactate does not affect electrical sharp wave-ripple activity featuring lower energy use. Lactate increases the oxygen consumption during the network states, reflecting enhanced oxidative ATP synthesis in mitochondria. Finally, lactate attenuates synaptic transmission in excitatory pyramidal cells and fast-spiking, inhibitory interneurons by reduced neurotransmitter release from presynaptic terminals, whereas action potential generation in the axon is regular. In conclusion, sole lactate is less effective and potentially harmful during gamma-band rhythms by omitting obligatory ATP delivery through fast glycolysis at the synapse.

Positive and negative chemotaxis of enzyme-coated liposome motors.

The ability of cells or cell components to move in response to chemical signals is critical for the survival of living systems. This motion arises from harnessing free energy from enzymatic catalysis. Artificial model protocells derived from phospholipids and other amphiphiles have been made and their enzymatic-driven motion has been observed. However, control of directionality based on chemical cues (chemotaxis) has been difficult to achieve. Here we show both positive or negative chemotaxis of liposomal protocells. The protocells move autonomously by interacting with concentration gradients of either substrates or products in enzyme catalysis, or Hofmeister salts. We hypothesize that the propulsion mechanism is based on the interplay between enzyme-catalysis-induced positive chemotaxis and solute-phospholipid-based negative chemotaxis. Controlling the extent and direction of chemotaxis holds considerable potential for designing cell mimics and delivery vehicles that can reconfigure their motion in response to environmental conditions.

Finger-Actuated Microfluidic Concentration Gradient Generator Compatible with a Microplate.

The generation of concentration gradients is an essential part of a wide range of laboratory settings. However, the task usually requires tedious and repetitive steps and it is difficult to generate concentration gradients at once. Here, we present a microfluidic device that easily generates a concentration gradient by means of push-button actuated pumping units. The device is designed to generate six concentrations with a linear gradient between two different sample solutions. The microfluidic concentration gradient generator we report here does not require external pumps because changes in the pressure of the fluidic channel induced by finger actuation generate a constant volume of fluid, and the design of the generator is compatible with the commonly used 96-well microplate. Generation of a concentration gradient by the finger-actuated microfluidic device was consistent with that of the manual pipetting method. In addition, the amount of fluid dispensed from each outlet was constant when the button was pressed, and the volume of fluid increased linearly with respect to the number of pushing times. Coefficient of variation (CV) was between 0.796% and 13.539%, and the error was between 0.111% and 19.147%. The design of the microfluidic network, as well as the amount of fluid dispensed from each outlet at a single finger actuation, can be adjusted to the user’s demand. To prove the applicability of the concentration gradient generator, an enzyme assay was performed using alkaline phosphatase (ALP) and para-nitrophenyl phosphate (pNPP). We generated a linear concentration gradient of the pNPP substrate, and the enzyme kinetics of ALP was studied by examining the initial reaction rate between ALP and pNPP. Then, a Hanes–Woolf plot of the various concentration of ALP was drawn and the Vmax and Km value were calculated.

The effect of antibiotic and nutrient limitation to antibiotic resistant bacteria in single-cell level

Heterogeneity in bacterial populations can manifest in various ways, such as resistant cells, which can be observed in harsh environments after the use of antibiotics. Many studies have looked at the evolution of resistance and the effect of inhibitory and sub-inhibitory concentrations of antibiotics by batch culture measurements without considering the heterogeneity of bacterial populations. But antibiotic susceptibility and fitness costs of resistance mutations or plasmids are affected by the growth rate and physiology of individual cells. Single-cell analysis in microfluidic systems has opened up new possibilities enabling us to investigate the various putative mechanisms behind the persistence phenomenon required direct observation under the microscope. In this study, we use a gradient mixer and a novel micro-chemostat, to create concentration gradients of growth substrates and/or antibiotics to study the effect of nutrient and antibiotic concentration on individual cells growing under constant and defined conditions in cell-sized channels. The single-cell elongation, morphology and growth rate of ribosome-targeting antibiotics resistant E. coli was tracked by combining the microfluidics, microscope phase contrast imaging and fluorescent tag in high throughput mode. A mechanistic cellular model was used to describe the reaction between antibiotics and ribosome and the resulting effects on bacterial growth; then we linked the intracellular chemical-reaction kinetics processes to the population level and predicted the behaviour of population responses. Our approach has enabled the investigation of single-cell individuality and predictions of population dynamics under different environment.

Influence of pulsed and continuous substrate inputs on freshwater bacterial community composition and functioning in bioreactors.

Aquatic environments are typically not homogenous, but characterized by changing substrate concentration gradients and nutrient patches. This heterogeneity in substrate availability creates a multitude of niches allowing bacteria with different substrate utilization strategies to hypothetically coexist even when competing for the same substrate. To study the impact of heterogeneous distribution of organic substrates on bacterioplankton, bioreactors with freshwater bacterial communities were fed artificial freshwater medium with acetate supplied either continuously or in pulses. After a month-long incubation, bacterial biomass and community-level substrate uptake rates were twice as high in the pulsed treatment compared to the continuously fed reactors even if the same total amount of acetate was supplied to both treatments. The composition of the bacterial communities emerging in the two treatments differed significantly with specific taxa overrepresented in the respective treatments. The higher estimated growth yield in cultures that received pulsed substrate inputs, imply that such conditions enable bacteria to use resources more efficiently for biomass production. This finding agrees with established concepts of basal maintenance energy requirements and high energetic costs to assimilate substrates at low concentration. Our results further imply that degradation of organic matter is influenced by temporal and spatial heterogeneity in substrate availability.

Janus Reactors with Highly Efficient Enzymatic CO2 Nanocascade at Air-Liquid Interface.

Though enzymatic cascade reactors have been the subject of intense research over the past few years, their application is still limited by the complicated fabrication protocols, unsatisfactory stability and lack of effective reactor designs. In addition, the spatial positioning of the cascade reactor has so far not been investigated, which is of significant importance for biphase catalytic reaction systems. Inspired by the Janus properties of the lipid cellular membrane, here we show a highly efficient Janus gas-liquid reactor for CO2 hydration and conversion. Within the Janus reactor, nanocascades containing the nanoscale compartmentalized carbonic anhydrase and formic dehydrogenase were positioned at a well-defined gas-liquid interface, with a high substrate concentration gradient. The Janus reactor exhibited 2.5 times higher CO2 hydration efficiency compared with the conventional gas-liquid contactor with pristine membranes, and the formic acid conversion rate can reach approximately 90%. Through this work, we provide evidence that the spatial arrangement of the nanocascade is also crucial to efficient reactions, and the Janus reactor can be a promising candidate for the biphase catalytic reactions in environmental, biological and energy aspects.

Thermodynamic Paradigm for Solution Demixing Inspired by Nuclear Transport in Living Cells.

Living cells display a remarkable capacity to compartmentalize their functional biochemistry. A particularly fascinating example is the cell nucleus. Exchange of macromolecules between the nucleus and the surrounding cytoplasm does not involve traversing a lipid bilayer membrane. Instead, large protein channels known as nuclear pores cross the nuclear envelope and regulate the passage of other proteins and RNA molecules. Beyond simply gating diffusion, the system of nuclear pores and associated transport receptors is able to generate substantial concentration gradients, at the energetic expense of guanosine triphosphate hydrolysis. In contrast to conventional approaches to demixing such as reverse osmosis and dialysis, the biological system operates continuously, without application of cyclic changes in pressure or solvent exchange. Abstracting the biological paradigm, we examine this transport system as a thermodynamic machine of solution demixing. Building on the construct of free energy transduction and biochemical kinetics, we find conditions for the stable operation and optimization of the concentration gradients as a function of dissipation in the form of entropy production.

Depth‐dependent concentrations of hematopoietic stem cells in the adult skeleton: Implications for active marrow dosimetry

The hematopoietically active (or red) bone marrow is the target tissue assigned in skeletal dosimetry models for assessment of stochastic effects (leukemia induction) as well as tissue reactions (marrow toxicity). Active marrow, however, is in reality a surrogate tissue region for specific cell populations, namely the hematopoietic stem and progenitor cells. Present models of active marrow dosimetry implicitly assume that these cells are uniformly localized throughout the marrow spaces of trabecular spongiosa. Data from Watchman etal. and Bourke etal., however, clearly indicate that there is a substantial spatial concentration gradient of these cells with the highest concentrations localized near the bone trabeculae surfaces. The purpose of the present study was thus to explore the dosimetric implications of these spatial gradients on active marrow dosimetry. Images of several bone sites from a 45-yr female were retagged to group active marrow voxels into 50μm increments of marrow depth, after which electron and alpha-particle depth-dependent specific absorbed fractions were computed for four source tissues - active marrow, inactive marrow, bone trabeculae volumes, and bone trabeculae surfaces. Corresponding depth-dependent S values (dose to a target tissue per decay in a source tissue) were computed and further weighted by the relative target cell concentration. These depth-weighted radionuclide S values were systematically compared to the more traditional volume-averaged radionuclide S values of the MIRD schema for both individual bones of the skeleton and their skeletal-averaged quantities. For both beta-emitters and alpha-emitters localized in the active and inactive marrow, depth-weighted S values were shown to differ from volume-averaged S values by only a few percent, as dose gradients across the marrow tissues are nonexistent. For bone volume and bone surface sources of alpha-emitters and lower energy beta-emitters, when marrow dose gradients are expected, explicit consideration of target cell spatial concentration gradients are shown to significantly impact marrow dosimetry. For medical isotopes currently utilized for treatment of skeletal metastases, namely 153 Sm and 223 Ra, accounting for hematopoietic stem and progenitor cell concentration gradients resulted in maximum percent differences to reference skeletal-averaged S values of ~21% and 55%, respectively.