Work distribution of a colloid in an elongational flow field and under Ornstein-Uhlenbeck noise.

The study of thermodynamic properties of microscopic systems, such as a colloid in a fluid, has been of great interest to researchers since the discovery of the fluctuation theorem and associated laws of stochastic thermodynamics. However, most of these studies confine themselves to systems where effective fluctuations acting on the colloid are in the form of delta-correlated Gaussian white noise (GWN). In this study, instead, we look into the work distribution function when a colloid trapped in a harmonic potential moves from one position to another in a fluid medium with an elongational flow field where the effective fluctuations are given by the Ornstein-Uhlenbeck noise, a type of colored noise. We use path integrals to calculate this distribution function and compare and contrast its properties to the case with GWN. We find that the work distribution function turns out to be non-Gaussian as a result of the elongational flow field but continues to obey the fluctuation theorem in both types of noise. Further, we also look into the effects of the various system parameters on the behavior of work fluctuations and find that although the distribution tends to broaden with increasing noise intensity, increased correlation in fluctuations acts to oppose this effect. Additionally, the system is found to consume heat from the surroundings at early times and dissipate it into the media at later times. This study, therefore, is a step towards gaining a better understanding of the thermodynamic properties of colloidal systems under nonlinear complex flows that also display correlated fluctuations.

Unveiling autophagy and aging through time-resolved imaging of lysosomal polarity with a delayed fluorescent emitter.

Detecting the lysosomal microenvironmental changes like viscosity, pH, and polarity during their dynamic interorganelle interactions remains an intriguing area that facilitates the elucidation of cellular homeostasis. The subtle variation of physiological conditions can be assessed by deciphering the lysosomal microenvironments during lysosome-organelle interactions, closely related to autophagic pathways leading to various cellular disorders. Herein, we shed light on the dynamic lysosomal polarity in live cells and a multicellular model organism, Caenorhabditis elegans (C. elegans), through time-resolved imaging employing a thermally activated delayed fluorescent probe, DC-Lyso. The highly photostable and cytocompatible DC-Lyso rapidly labels the lysosomes (within 1 min of incubation) and exhibits red luminescence and polarity-sensitive long lifetime under the cellular environment. The distinct variation in the fluorescence lifetime of DC-Lyso suggests an increase in local polarity during the lysosomal dynamics and interorganelle interactions, including lipophagy and mitophagy. The lifetime imaging analysis reveals increasing lysosomal polarity as an indicator for probing the successive development of C. elegans during aging. The in vivo microsecond timescale imaging of various cancerous cell lines and C. elegans, as presented here, therefore, expands the scope of delayed fluorescent emitters for unveiling complex biological processes.

Microfluidics based bioimaging with cost-efficient fabrication of multi-level micrometer-sized trenches.

Microfluidic devices, through their vast applicability as tools for miniaturized experimental setups, have become indispensable for cutting edge research and diagnostics. However, the high operational cost and the requirement of sophisticated equipment and clean room facility for the fabrication of these devices make their use unfeasible for many research laboratories in resource limited settings. Therefore, with the aim of increasing accessibility, in this article, we report a novel, cost-effective micro-fabrication technique for fabricating multi-layer microfluidic devices using only common wet-lab facilities, thereby significantly lowering the cost. Our proposed process-flow-design eliminates the need for a mastermold, does not require any sophisticated lithography tools, and can be executed successfully outside a clean room. In this work, we also optimized the critical steps (such as spin coating and wet etching) of our fabrication process and validated the process flow and the device by trapping and imaging Caenorhabditis elegans. The fabricated devices are effective in conducting lifetime assays and flushing out larvae, which are, in general, manually picked from Petri dishes or separated using sieves. Our technique is not only cost effective but also scalable, as it can be used to fabricate devices with multiple layers of confinements ranging from 0.6 to more than 50 µm, thus enabling the study of unicellular and multicellular organisms. This technique, therefore, has the potential to be adopted widely by many research laboratories for a variety of applications.

Lessons Learned from Two Decades of Modeling the Heat-Shock Response.

The Heat Shock Response (HSR) is a highly conserved genetic system charged with protecting the proteome in a wide range of organisms and species. Experiments since the early 1980s have elucidated key elements in these pathways and revealed a canonical mode of regulation, which relies on a titration feedback. This system has been subject to substantial modeling work, addressing questions about resilience, design and control. The compact core regulatory circuit, as well as its apparent conservation, make this system an ideal 'hydrogen atom' model for the regulation of stress response. Here we take a broad view of the models of the HSR, focusing on the different questions asked and the approaches taken. After 20 years of modeling work, we ask what lessons had been learned that would have been hard to discover without mathematical models. We find that while existing models lay strong foundations, many important questions that can benefit from quantitative modeling are still awaiting investigation.

Persistent Correlation in Cellular Noise Determines Longevity of Viral Infections.

The slowly decaying viral dynamics, even after 2-3 weeks from diagnosis, is one of the characteristics of COVID-19 infection that is still unexplored in theoretical and experimental studies. This long-lived characteristic of viral infections in the framework of inherent variations or noise present at the cellular level is often overlooked. Therefore, in this work, we aim to understand the effect of these variations by proposing a stochastic non-Markovian model that not only captures the coupled dynamics between the immune cells and the virus but also enables the study of the effect of fluctuations. Numerical simulations of our model reveal that the long-range temporal correlations in fluctuations dictate the long-lived dynamics of a viral infection and, in turn, also affect the rates of immune response. Furthermore, predictions of our model system are in agreement with the experimental viral load data of COVID-19 patients from various countries.

A near analytic solution of a stochastic immune response model considering variability in virus and T-cell dynamics.

Biological processes at the cellular level are stochastic in nature, and the immune response system is no different. Therefore, models that attempt to explain this system need to also incorporate noise or fluctuations that can account for the observed variability. In this work, a stochastic model of the immune response system is presented in terms of the dynamics of T cells and virus particles. Making use of the Green's function and the Wilemski-Fixman approximation, this model is then solved to obtain the analytical expression for the joint probability density function of these variables in the early and late stages of infection. This is then also used to calculate the average level of virus particles in the system. Upon comparing the theoretically predicted average virus levels to those of COVID-19 patients, it is hypothesized that the long-lived dynamics that are characteristics of such viral infections are due to the long range correlations in the temporal fluctuations of the virions. This model, therefore, provides an insight into the effects of noise on viral dynamics.

Transcription factors and chaperone proteins play a role in launching a faster response to heat stress and aggregation.

Proteins, under conditions of cellular stress, typically tend to unfold and form lethal aggregates leading to neurological diseases like Parkinson's and Alzheimer's. A clear understanding of the conditions that favor dis-aggregation and restore the cell to its healthy state after they have been stressed is therefore important in dealing with these diseases. The heat shock response (HSR) mechanism is a signaling network that deals with these undue protein aggregates and aids in the maintenance of homeostasis within a cell. This framework, on its own, is a mathematically well studied mechanism. However, not much is known about how the various intermediate mis-folded protein states of the aggregation process interact with some of the key components of the HSR pathway such as the Heat Shock Protein (HSP), the Heat Shock Transcription Factor (HSF) and the HSP-HSF complex. In this article, using kinetic parameters from the literature, we propose and analyze two mathematical models for HSR that also include explicit reactions for the formation of protein aggregates. Deterministic analysis and stochastic simulations of these models show that the folded proteins and the misfolded aggregates exhibit bistability in a certain region of the parameter space. Further, the models also highlight the role of HSF and the HSF-HSP complex in reducing the time lag of response to stress and in re-folding all the mis-folded proteins back to their native state. These models, therefore, call attention to the significance of studying related pathways such as the HSR and the protein aggregation and re-folding process in conjunction with each other.

High prevalence of non-alcoholic fatty liver disease among healthy male blood donors of urban India.

BACKGROUND: There is limited data on the community prevalence of non-alcoholic fatty liver disease (NAFLD). The present study evaluated the prevalence of NAFLD in a large number of healthy male blood donors of urban north India. METHODOLOGY: In a prospective study performed over 18 months, voluntary blood donors fulfilling the requisite blood donation criteria and consenting to participate in the study were evaluated. The study received the approval of the institute's ethics committee. Diagnosis of NAFLD was made by excluding significant alcohol intake, ultrasound showing hepatic steatosis, and exclusion of transfusion associated infections. Subjects were also evaluated for various metabolic risk factors and the presence of metabolic syndrome. RESULTS: Of 1388 subjects who consented for participation, 386 did not come for evaluation. Three females, nine (0.9%) HBsAg-positive, and four (0.4%) anti-HCV positive subjects were excluded. Of the 986 males evaluated with hepatobiliary ultrasound, 543(55.1%) had fatty liver on ultrasonography [15 (1.5%) alcoholic fatty liver and 528 (53.5%) NAFLD]. Among those with NAFLD, 469 (88.8%), 54 (10.2%), and 5 (0.9%) had mild, moderate, and severe hepatic steatosis, respectively. Subjects with NAFLD, when compared to those without NAFLD, had significantly higher age, BMI, waist circumference, blood pressure, total cholesterol and triglycerides, low-density lipoprotein, and fasting plasma glucose. Multivariate regression analysis demonstrated age, BMI, waist circumference, systolic blood pressure, total cholesterol, and number of metabolic syndrome criteria as independent predictors of NAFLD. CONCLUSIONS: Urban Indian healthy male blood donors have a high prevalence of NAFLD.

Redefining the Normal Values of Serum Aminotransferases in Healthy Indian Males.

BACKGROUND: The normal range for Aspartate and Alanine Aminotransferases (AST and ALT) levels (<40 IU/L) were set in 1950s. Recent data from certain countries suggest lower levels of AST and ALT. Aim of the study was to redefine the normal values of aminotransferases in healthy Indian adults. METHODOLOGY: In a cross sectional prospective study, 1002 blood donors were evaluated to isolate a healthy cohort. Four and 9 subjects positive for HBsAg and anti-Hepatitis C Virus (HCV) respectively and three females were excluded. 986 male subjects were evaluated for levels of serum aminotransferases. RESULTS: Of total 986 subjects (Group I), 543 (55.1%) had fatty liver on ultrasound [15 (1.5%) alcoholic fatty liver and 528 (53.5%) Nonalcoholic Fatty Liver Disease (NAFLD)]. Median AST and ALT in total group (Group I) were 27.69 (Interquartile Range (IQR) 22.33-37.04) and 34.19 IU/L (IQR 23.12-54.87) and in NAFLD (Group II) were 35.67 (IQR -27.49-47.43) and 50.36 (IQR 37.70-76.58) IU/L. Of remaining 443 subjects without fatty liver, 288 had one or more components of metabolic syndrome. Out of 155 patients with no fatty liver and no component of metabolic syndrome (Group III), 103 subjects had normal Body Mass Index (BMI) and normal cholesterol and Low Density Lipoprotein (LDL) (Group IIIB). Median AST and ALT in Group IIIB were 22.56 (IQR 20.23-26.91) and 21.36 (IQR 17.49-27.21) U/L respectively with a 95th percentile of 34.28 and 36.57 U/L for AST and ALT, respectively. CONCLUSION: Levels of AST and ALT in healthy men are lower than the conventional values in India.

Biospark: scalable analysis of large numerical datasets from biological simulations and experiments using Hadoop and Spark.

Data-parallel programming techniques can dramatically decrease the time needed to analyze large datasets. While these methods have provided significant improvements for sequencing-based analyses, other areas of biological informatics have not yet adopted them. Here, we introduce Biospark, a new framework for performing data-parallel analysis on large numerical datasets. Biospark builds upon the open source Hadoop and Spark projects, bringing domain-specific features for biology. AVAILABILITY AND IMPLEMENTATION: Source code is licensed under the Apache 2.0 open source license and is available at the project website: https://www.assembla.com/spaces/roberts-lab-public/wiki/Biospark CONTACT: eroberts@jhu.eduSupplementary information: Supplementary data are available at Bioinformatics online.

Gradient sensing by a bistable regulatory motif enhances signal amplification but decreases accuracy in individual cells.

Many vital eukaryotic cellular functions require the cell to respond to a directional gradient of a signaling molecule. The first two steps in any eukaryotic chemotactic/chemotropic pathway are gradient detection and cell polarization. Like many processes, such chemotactic and chemotropic decisions are made using a relatively small number of molecules and are thus susceptible to internal and external fluctuations during signal transduction. Large cell-to-cell variations in the magnitude and direction of a response are therefore possible and do, in fact, occur in natural systems. In this work we use three-dimensional probabilistic modeling of a simple gradient sensing pathway to study the capacity for individual cells to accurately determine the direction of a gradient, despite fluctuations. We include a stochastic external gradient in our simulations using a novel gradient boundary condition modeling a point emitter a short distance away. We compare and contrast three different variants of the pathway, one monostable and two bistable. The simulation data show that an architecture combining bistability with spatial positive feedback permits the cell to both accurately detect and internally amplify an external gradient. We observe strong polarization in all individual cells, but in a distribution of directions centered on the gradient. Polarization accuracy in our study was strongly dependent upon a spatial positive feedback term that allows the pathway to trade accuracy for polarization strength. Finally, we show that additional feedback links providing information about the gradient to multiple levels in the pathway can help the cell to refine initial inaccuracy in the polarization direction.

Dynamics of simple gene-network motifs subject to extrinsic fluctuations.

Cellular processes do not follow deterministic rules; even in identical environments genetically identical cells can make random choices leading to different phenotypes. This randomness originates from fluctuations present in the biomolecular interaction networks. Most previous work has been focused on the intrinsic noise (IN) of these networks. Yet, especially for high-copy-number biomolecules, extrinsic or environmental noise (EN) has been experimentally shown to dominate the variation. Here, we develop an analytical formalism that allows for calculation of the effect of EN on gene-expression motifs. We introduce a method for modeling bounded EN as an auxiliary species in the master equation. The method is fully generic and is not limited to systems with small EN magnitudes. We focus our study on motifs that can be viewed as the building blocks of genetic switches: a nonregulated gene, a self-inhibiting gene, and a self-promoting gene. The role of the EN properties (magnitude, correlation time, and distribution) on the statistics of interest are systematically investigated, and the effect of fluctuations in different reaction rates is compared. Due to its analytical nature, our formalism can be used to quantify the effect of EN on the dynamics of biochemical networks and can also be used to improve the interpretation of data from single-cell gene-expression experiments.

Efficacy and safety of autologous bone marrow-derived stem cell transplantation in patients with type 2 diabetes mellitus: a randomized placebo-controlled study.

There is a growing interest in cell-based therapies in T2DM as ß-cell failure is progressive and inexorable with the advancing duration of disease. This prospective, randomized, single-blinded placebo-controlled study evaluates the efficacy and safety of autologous bone marrow-derived stem cell transplantation (ABMSCT) in T2DM. Twenty-one patients with triple oral antidiabetic drug failure and requiring insulin ≥0.4 IU per kg per day with HbA1c <7.5% were randomly assigned to an intervention (n = 11) and control group (n = 10) and followed for 12 months. Patients in the intervention group received ABMSCT through a targeted approach, and after 12 weeks, a second dose of stem cells was administered through the antecubital vein after mobilization with G-CSF, while the control group underwent a sham procedure. The primary end point was a reduction in insulin requirement by ≥50% from baseline while maintaining HbA1c <7%. Nine out of the 11 (82%) patients in the intervention group achieved the primary end point, whereas none of the patients in the control group did over the study period (p = 0.002). The insulin requirement decreased by 66.7% in the intervention group from 42.0 (31.0­64.0) IU per day to 14.0 (0.0­30.0) IU per day (p = 0.011), while in controls it decreased by 32.1% from 40.5 (31.8­44.3) IU per day to 27.5 (23.5­33.3) IU per day (p = 0.008) at 12 months. The reduction in insulin requirement was significantly more in the intervention group compared to controls at both 6 (p = 0.001) and 12 months (p = 0.004). There was a modest but nonsignificant increase in HbA1c (%) in cases from 6.9% (6.4­7.2%) to 7.1% (6.6­7.5%) as well as in controls from 6.9% (6.2­7.0%) to 7.0% (6.9­7.5%). Ten out of 11 (91%) patients could maintain HbA1c <7% in the intervention group, whereas 6 out of 10 did (60%) in the control group (p = 0.167). The glucagon-stimulated C-peptide significantly increased in treated cases compared to controls (p = 0.036). The decrease in insulin requirement positively correlated with stimulated C-peptide (r = 0.8, p = 0.001). In conclusion, ABMSCT results in a significant decrease in the insulin dose requirement along with an improvement in the stimulated C-peptide levels in T2DM. However, a greater number of patients with a longer duration of follow-up are required to substantiate these observations.

Interfacial growth as a model of tube-width heterogeneities in concentrated solutions of stiff polymers.

Recent experimental measurements of the distribution P(w) of transverse chain fluctuations w in concentrated solutions of F-actin filaments [B. Wang, J Guan, S. M. Anthony, S. C. Bae, K. S. Schweizer, and S. Granick, Phys. Rev. Lett. 104, 118301 (2010); J. Glaser, D. Chakraborty, K. Kroy, I. Lauter, M. Degawa, N. Kirchgessner, B. Hoffmann, R. Merkel, and M. Giesen, Phys. Rev. Lett. 105, 037801 (2010)] are shown to be well-fit to an expression derived from a model of the conformations of a single harmonically confined weakly bendable rod. The calculation of P(w) is carried out essentially exactly within a path integral approach that was originally applied to the study of one-dimensional randomly growing interfaces. Our results are generally as successful in reproducing experimental trends as earlier approximate results obtained from more elaborate many-chain treatments of the confining tube potential.

Subdiffusion in hair bundle dynamics: the role of protein conformational fluctuations.

The detection of sound signals in vertebrates involves a complex network of different mechano-sensory elements in the inner ear. An especially important element in this network is the hair bundle, an antenna-like array of stereocilia containing gated ion channels that operate under the control of one or more adaptation motors. Deflections of the hair bundle by sound vibrations or thermal fluctuations transiently open the ion channels, allowing the flow of ions through them, and producing an electrical signal in the process, eventually causing the sensation of hearing. Recent high frequency (0.1-10 kHz) measurements by Kozlov et al. [Proc. Natl. Acad. Sci. U.S.A. 109, 2896 (2012)] of the power spectrum and the mean square displacement of the thermal fluctuations of the hair bundle suggest that in this regime the dynamics of the hair bundle are subdiffusive. This finding has been explained in terms of the simple Brownian motion of a filament connecting neighboring stereocilia (the tip link), which is modeled as a viscoelastic spring. In the present paper, the diffusive anomalies of the hair bundle are ascribed to tip link fluctuations that evolve by fractional Brownian motion, which originates in fractional Gaussian noise and is characterized by a power law memory. The predictions of this model for the power spectrum of the hair bundle and its mean square displacement are consistent with the experimental data and the known properties of the tip link.

Confinement and viscoelastic effects on chain closure dynamics.

Chemical reactions inside cells are typically subject to the effects both of the cell's confining surfaces and of the viscoelastic behavior of its contents. In this paper, we show how the outcome of one particular reaction of relevance to cellular biochemistry--the diffusion-limited cyclization of long chain polymers--is influenced by such confinement and crowding effects. More specifically, starting from the Rouse model of polymer dynamics, and invoking the Wilemski-Fixman approximation, we determine the scaling relationship between the mean closure time t(c) of a flexible chain (no excluded volume or hydrodynamic interactions) and the length N of its contour under the following separate conditions: (a) confinement of the chain to a sphere of radius d and (b) modulation of its dynamics by colored Gaussian noise. Among other results, we find that in case (a) when d is much smaller than the size of the chain, t(c) ~ Nd(2), and that in case (b), t(c) ~ N(2/(2-2H)), H being a number between 1/2 and 1 that characterizes the decay of the noise correlations. H is not known a priori, but values of about 0.7 have been used in the successful characterization of protein conformational dynamics. At this value of H (selected for purposes of illustration), t(c) ~ N(3.4), the high scaling exponent reflecting the slow relaxation of the chain in a viscoelastic medium.

Distribution of transverse chain fluctuations in harmonically confined semiflexible polymers.

Two different experimental studies of polymer dynamics based on single-molecule fluorescence imaging have recently found evidence of heterogeneities in the widths of the putative tubes that surround filaments of F-actin during their motion in concentrated solution. In one [J. Glaser, D. Chakraborty, K. Kroy, I. Lauter, M. Degawa, N. Kirchesner, B. Hoffmann, R. Merkel, and M. Giesen, Phys. Rev. Lett. 105, 037801 (2010)], the observations were explained in terms of the statistics of a worm-like chain confined to a potential determined self-consistently by a binary collision approximation, and in the other [B. Wang, J. Guan, S. M. Anthony, S. C. Bae, K. S. Schweizer, and S. Granick, Phys. Rev. Lett. 104, 118301 (2010)], they were explained in terms of the scaling properties of a random fluid of thin rods. In this paper, we show, using an exact path integral calculation, that the distribution of the length-averaged transverse fluctuations of a harmonically confined weakly bendable rod (one possible realization of a semiflexible chain in a tube), is in good qualitative agreement with the experimental data, although it is qualitatively different in analytic structure from the earlier theoretical predictions. We also show that similar path integral techniques can be used to obtain an exact expression for the time correlation function of fluctuations in the tube cross section.

Microsomal Epoxide Hydrolase Polymorphisms and Haplotypes as Determinants of Hepatitis B Virusand Hepatitis C Virus-related Liver Disease in Indian Population.

BACKGROUND: Hepatocellular carcinoma (HCC) is the fifth most common cancer and third leading cause of death worldwide. Main causes of HCC are hepatitis B virus (HBV) and hepatitis C virus (HCV) infections. mEPHX, a xenobiotic metabolizing enzyme, exhibits a dual role of procarcinogen detoxification and activation, hence considered as a cancer risk factor as well as a protective factor. Two known polymorphic forms of mEPHX, exon in exon 3 and 4, are associated with the development of HCC. OBJECTIVE: To determine the association of genotypes and haplotypes of mEPHX with risk of HCC developments separately in HBV- and HCV-infected carriers and patients with hepatitis. METHODS: Polymerase chain reactions (PCR) were carried out using primers to amplify exon 3 (113 Tyr→His variant) and exon 4 (139 His→Arg) polymorphic sites. To distinguish the wild and variant forms, PCR amplification products were digested with restriction endonucleases EcoRV and Rsa1 for exons 3 and 4, respectively. RESULT: Exon 3 genotypes, Y113H and H113H, shared a protective association with HBV-chronic hepatitis infection (P < 0.001 and P< 0.01, respectively) as well as HBV-HCC development (P < 0.001) among HBV-carrier group, while Y113H acts as a risk factor for HCV-chronic hepatitis development (P < 0.001) as well as for HCC development (P < 0.01) with HCV-carrier group as reference. Both H139R and R139R, exon 4 genotypes, acted as a risk factor for HBV/HCV-chronic hepatitis infection and for HBV/HCV-HCC development (P ranges from < 0.05 to < 0.001) with HBV/HCV carriers as reference. 113His-139His and 113His-139Arg haplotypes shared a significant negative and positive association, respectively, with HBV hepatitis and HBV-HCC risk. 113Tyr-139Arg haplotype acted as a risk for HCV-HCC development. CONCLUSION: Polymorphic and haplotypic variant forms of mEPHX exon 3 and 4 variably determine the susceptibility to develop HCC in HBV- and HCV-carrier subjects.