Functional characterization of QT interval associated SCN5A enhancer variants identify combined additive effects.

Several empirical and theoretical studies suggest presence of multiple enhancers per gene that collectively regulate gene expression, and that common sequence variation impacting on the activities of these enhancers is a major source of inter-individual variability in gene expression. However, for vast majority of genes, enhancers and the underlying regulatory variation remains unknown. Even for the genes with well-characterized enhancers, the nature of the combined effects from multiple enhancers and their variants, when known, on gene expression regulation remains unexplored. Here, we have evaluated the combined effects from five SCN5A enhancers and their regulatory variants that are known to collectively correlate with SCN5A cardiac expression and underlie QT interval association in the general population. Using small deletions centered at the regulatory variants in episomal reporter assays in a mouse cardiomyocyte cell line we demonstrate that the variants and their flanking sequences play critical role in individual enhancer activities, likely being a transcription factor (TF) binding site. By performing oligonucleotide-based pulldown assays on predicted TFs we identify the TFs likely driving allele-specific enhancer activities. Using all 32 possible allelic synthetic constructs in reporter assays, representing the five biallelic enhancers in tandem in their genomic order, we demonstrate combined additive effects on overall enhancer activities. Using transient enhancer assays in developing zebrafish embryos we demonstrate the four out the five enhancer elements act as enhancers in vivo . Together, these studies extend the previous findings to uncover the TFs driving the enhancer activities of QT interval associated SCN5A regulatory variants, reveal the additive effects from allelic combinations of these regulatory variants, and prove their potential to act as enhancers in vivo .

Identification of candidate causal cis -regulatory variants underlying electrocardiographic QT interval GWAS loci.

Identifying causal variants among tens or hundreds of associated variants at each locus mapped by genome-wide association studies (GWAS) of complex traits is a challenge. As vast majority of GWAS variants are noncoding, sequence variation at cis -regulatory elements affecting transcriptional expression of specific genes is a widely accepted molecular hypothesis. Following this cis -regulatory hypothesis and combining it with the observation that nucleosome-free open chromatin is a universal hallmark of all types of cis -regulatory elements, we aimed to identify candidate causal regulatory variants underlying electrocardiographic QT interval GWAS loci. At a dozen loci, selected for higher effect sizes and a better understanding of the likely causal gene, we identified and included all common variants in high linkage disequilibrium with the GWAS variants as candidate variants. Using ENCODE DNase-seq and ATAC-seq from multiple human adult cardiac left ventricle tissue samples, we generated genome-wide maps of open chromatin regions marking putative regulatory elements. QT interval associated candidate variants were filtered for overlap with cardiac left ventricle open chromatin regions to identify candidate causal cis -regulatory variants, which were further assessed for colocalizing with a known cardiac GTEx expression quantitative trait locus variant as additional evidence for their causal role. Together, these efforts have generated a comprehensive set of candidate causal variants that are expected to be enriched for cis -regulatory potential and thereby, explaining the observed genetic associations.

Advances in two-dimensional transition metal dichalcogenides-based sensors for environmental, food, and biomedical analysis: A review.

Transition metal dichalcogenides (TMDCs) are versatile two-dimensional (2D) nanomaterials used in biosensing applications due to their excellent physical and chemical properties. Due to biomaterial target properties, biosensors' most significant challenge is improving their sensitivity and stability. In environmental analysis, TMDCs have demonstrated exceptional pollutant detection and removal capabilities. Their high surface area, tunable electronic properties, and chemical reactivity make them ideal for sensors and adsorbents targeting various contaminants, including heavy metals, organic pollutants, and emerging contaminants. Furthermore, their unique electronic and optical properties enable sensitive detection techniques, enhancing our ability to monitor and mitigate environmental pollution. In the food analysis, TMDCs-based nanomaterials have shown remarkable potential in ensuring food safety and quality. These nanomaterials exhibit high specificity and sensitivity for detecting contaminants, pathogens, and adulterants in various food matrices. Their integration into sensor platforms enables rapid and on-site analysis, reducing the reliance on centralized laboratories and facilitating timely interventions in the food supply chain. In biomedical studies, TMDCs-based nanomaterials have demonstrated significant strides in diagnostic and therapeutic applications. Their biocompatibility, surface functionalization versatility, and photothermal properties have paved the way for novel disease detection, drug delivery, and targeted therapy approaches. Moreover, TMDCs-based nanomaterials have shown promise in imaging modalities, providing enhanced contrast and resolution for various medical imaging techniques. This article provides a comprehensive overview of 2D TMDCs-based biosensors, emphasizing the growing demand for advanced sensing technologies in environmental, food, and biomedical analysis.

Cardiac muscle-restricted partial loss of Nos1ap expression has limited but significant impact on electrocardiographic features.

Genome-wide association studies have identified sequence polymorphisms in a functional enhancer of the NOS1AP gene as the most common genetic regulator of QT interval and human cardiac NOS1AP gene expression in the general population. Functional studies based on in vitro overexpression in murine cardiomyocytes and ex vivo knockdown in zebrafish embryonic hearts, by us and others, have also demonstrated that NOS1AP expression levels can alter cellular electrophysiology. Here, to explore the role of NOS1AP in cardiac electrophysiology at an organismal level, we generated and characterized constitutive and heart muscle-restricted Nos1ap knockout mice to assess whether NOS1AP disruption alters the QT interval in vivo. Constitutive loss of Nos1ap led to genetic background-dependent variable lethality at or right before birth. Heart muscle-restricted Nos1ap knockout, generated using cardiac-specific alpha-myosin heavy chain promoter-driven tamoxifen-inducible Cre, resulted in tissue-level Nos1ap expression reduced by half. This partial loss of expression had no detectable effect on the QT interval or other electrocardiographic and echocardiographic parameters, except for a small but significant reduction in the QRS interval. Given that challenges associated with defining the end of the T wave on murine electrocardiogram can limit identification of subtle effects on the QT interval and that common noncoding NOS1AP variants are also associated with the QRS interval, our findings support the role of NOS1AP in regulation of the cardiac electrical cycle.

Microfluidic paper-based device coupled with 3D printed imaging box for colorimetric detection in resource-limited settings.

Rapid and effective methods for the detection of analytes such as water contaminants, food adulterants and biomolecules are essential for the protection of public health and environmental protection. Most of the currently established analytical techniques need sophisticated equipment, centralized testing facilities, costly operations, and trained personnel. Such limitations make them inaccessible to the general populace, particularly in regions with limited resources. The emergence of microfluidic devices offers a promising alternative to overcome several such constraints. This work describes a protocol for fabricating a low-cost, open-source paper-based microfluidic device using easily available tools and materials for colorimetric detection of analytes. The ease and simplicity of fabrication allow users to design customized devices. The device is coupled with an imaging box assembled from 3D printed parts to maintain uniform lighting conditions during analytical testing. The platform allows digital imaging using smartphones or cameras to instantaneously capture images of reaction zones on the device for quantitative analysis. The system is demonstrated for detecting hexavalent chromium, a toxic water contaminant. The image analysis is performed using open-source ImageJ for quantification of results. The approach demonstrated in this work can be readily adopted for a wide range of sensing applications.

Tribology of the sock-skin Interface - the influence of different fabric parameters on sock friction.

BACKGROUND: The purpose of this parametric design of experiments was to identify and summarize how the influence of knit structure (single jersey vs. terry), fiber composition (polyester vs. cotton), fiber linear density (30/1 Ne vs. 18/1 Ne & 1/150/34 vs. 2/150/34), and yarn type (filament vs. spun) affected the frictional profile across the sock-skin interface. METHODS: Friction testing trials were completed against both a polypropylene probe and a synthetic skin material (Lorica soft®) to determine if there was a difference in friction based on interface interaction. Friction testing was completed by sliding a probe across the inside bottom surface of the sock (the part that is usually in-contact with the bottom of the foot) while instantaneously measuring the frictional force every tenth of a second. RESULTS: For both trials (plastic probe and synthetic skin), in the dry condition, knit structure was found to be the most prominent fabric parameter affecting the frictional force experienced at the sock-skin interface. It was also determined that fiber linear density, and yarn type are tertiary factors affecting the frictional force measured at the sock-skin interface. Finally, in the dry state, it was determined that fiber composition had seemingly no effect on the frictional force experienced at the sock-skin interface. CONCLUSION: This parametric design of experiments has further enhanced the understanding of the tribology at the sock-skin interface. Through strategic design, four different textile parameters have been investigated, measured, and justified as to how each influence the friction measured between the two interfaces. This knowledge can be used to develop socks that mitigate the risk of friction blisters formation.

Extraction techniques in food industry: Insights into process parameters and their optimization.

This review presents critical evaluation of the key parameters that affect the extraction of targeted components, giving due consideration to safety and environmental aspects. The crucial aspects of the extraction technologies along with protocols and process parameters for designing unit operations have been emphasized. The parameters like solvent usage, substrate type, concentration, particle size, temperature, quality and storage of extract as well as stability of extraction have been elaborately discussed. The process optimization using mathematical and computational modeling highlighting information and communication technologies have been given importance aiming for a green and sustainable industry level scaleup. The findings indicate that the extraction processes vary significantly depending on the category of food and its structure. There is no single extraction method or universal set of process conditions identified for extracting all value-added products from respective sources. A comprehensive understanding of process parameters and their optimization as well as synergistic combination of multiple extraction processes can aid in enhancement of the overall extraction efficiency. Future efforts must be directed toward the design of integrated unit operations that cause minimal harm to the environment along with investigations on economic feasibility to ensure sustainable extraction systems.

Paper-based microfluidic colorimetric sensor on a 3D printed support for quantitative detection of nitrite in aquatic environments.

To ensure safe drinking water, it is necessary to have a simple method by which the probable pollutants are detected at the point of distribution. Nitrite contamination in water near agricultural locations could be an environmental concern due to its deleterious effects on the human population. The development of a frugal paper-based microfluidic sensor could be desirable to achieve the societal objective of providing safe drinking water. This work describes the development of a facile and cost-effective microfluidic paper-based sensor for quantitative estimation of nitrite in aquatic environments. A simple punching machine was used for fabrication and rapid prototyping of paper-based sensors without the need of any specialized equipment or patterning techniques. A reusable 3D printed platform served as the support for simultaneous testing of multiple samples. The nitrite estimation was carried out with smartphone-assisted digital image acquisition and colorimetric analysis. Under optimized experimental conditions, the variation in average grayscale intensity with concentration of nitrite was linear in the range from 0.1 to 10 ppm. The limits of detection and quantitation were 0.12 ppm and 0.35 ppm respectively. The reproducibility, expressed as relative standard deviation was 1.31%. The selectivity of nitrite detection method was determined by performing interference studies with commonly existing co-ions in water, such as bicarbonates, chloride and sulphate. The paper-based sensor was successfully applied for estimation of nitrite in actual water samples and showed high recoveries in the range of 83.5-109%. The results were in good agreement with those obtained using spectrophotometry. The developed paper-based sensor method, by virtue of its simplicity, ease of fabrication and use, could be readily extended for detection of multiple analytes in resource-limited settings.

Evaluation of the relationship between dermatoglyphics and mandibular third molar impaction: A cross-sectional study.

Background: Dermatoglyphics can be utilised in clinical settings to identify those who are more likely to have impacted teeth. Additionally, dermatoglyphics looks to have potential as a non-invasive diagnostic method for predicting the presence or absence of an impacted tooth. The goal of this study was to look at the most common dermatoglyphic pattern in people who had or didn't have an impacted mandibular third molar teeth and see if there was a dermatoglyphic signature. Methods: A cross-sectional study with 180 participants was conducted (90 cases and 90 controls). The rolling impression technique was used to apply blue duplicating ink to participants' fingertips, which was then recorded. There was an increase in the frequency of the whorl-plain pattern in the right-hand ring finger (60%; p=0.028) and left-hand little finger (33.3%; p=0.009), as well as the loop-ulnar pattern in the right-hand middle finger (74.4%; p=0.024) in individuals with a predisposition to the presence of impacted teeth. Results: The left-hand little finger was found to be the most predictive for impaction in a forward stepwise binary logistic regression analysis. Conclusions: Dermatoglyphics could be used as a non-invasive sign to predict whether or not a tooth is affected. Its value comes in early detection, which helps to avoid the surgical problems that come with symptomatic extraction of an impacted tooth.

Current status and future prospects of membrane separation processes for value recovery from wastewater.

Resource constraints and deteriorating environment have made it necessary to look for intensification of the industrial processes, to recover value from spent streams for reuse. The development of reverse osmosis has already established that water can be recovered from aqueous streams in a cost-effective and beneficial manner to the industries. With the development of several membrane processes and membrane materials, the possibility of recovering value from the effluents looks like a workable proposition. In this context, the potentialities of the different membrane processes in value recovery are presented. Among the pressure-driven processes, reverse osmosis can be used for the recovery of water as value. Nanofiltration has been used for the recovery of several dyes including crystal violet, congo red, methyl blue, etc., while ultrafiltration has been used in the fractionation of different solute species using membranes of different pore-size characteristics. Diffusion dialysis is found useful in the separation of acids from its salt solutions. Bipolar membrane electrodialysis has the potential to regenerate acid and base from salt solutions. Thermally driven membrane distillation can provide desalinated water, besides reducing the temperature of hot discharge streams. Passive membrane processes such as supported liquid membranes and membrane-assisted solvent extraction have been found useful in separating minor components from the wastewater streams. The details are discussed to drive home that membrane processes can be useful to achieve the objectives of value recovery, in a cost-effective manner through process intensification, as they are more compact and individual streams can be treated and value used seamlessly.

Lab-on-a-chip technologies for food safety, processing, and packaging applications: a review.

The advent of microfluidic systems has led to significant developments in lab-on-a-chip devices integrating several functions onto a single platform. Over the years, these miniature devices have become a promising tool for faster analytical testing, displaying high precision and efficiency. Nonetheless, most microfluidic systems are not commercially available. Research is actually undergoing on the application of these devices in environmental, food, biomedical, and healthcare industries. The lab-on-a-chip industry is predicted to grow annually by 20%. Here, we review the use of lab-on-a-chip devices in the food sector. We present fabrication technologies and materials to developing lab-on-a-chip devices. We compare electrochemical, optical, colorimetric, chemiluminescence and biological methods for the detection of pathogens and microorganisms. We emphasize emulsion processing, food formulation, nutraceutical development due to their promising characteristics. Last, smart packaging technologies like radio frequency identification and indicators are highlighted because they allow better product identification and traceability.

Valorization of food waste as adsorbents for toxic dye removal from contaminated waters: A review.

Industrial contaminants such as dyes and intermediates are released into water bodies, making the water unfit for human use. At the same time large amounts of food wastes accumulate near the work places, residential complexes etc. polluting the air due to putrefaction. The need of the hour lies in finding innovative solutions for dye removal from wastewater streams. In this context, the article emphasizes adoption or conversion of food waste materials, an ecological nuisance, as adsorbents for the removal of dyes from wastewaters. Adsorption, being a well-established technique, the review critically examines the specific potential of food waste constituents as dye adsorbents. The efficacy of food waste-based adsorbents is examined, besides addressing the possible adsorption mechanisms and the factors affecting phenomenon such as pH, temperature, contact time, adsorbent dosage, particle size, and ionic strength. Integration of information and communication technology approaches with adsorption isotherms and kinetic models are emphasized to bring out their role in improving overall modeling performance. Additionally, the reusability of adsorbents has been highlighted for effective substrate utilization. The review makes an attempt to stress the valorization of food waste materials to remove dyes from contaminated waters thereby ensuring long-term sustainability.

Agricultural waste materials for adsorptive removal of phenols, chromium (VI) and cadmium (II) from wastewater: A review.

Management of basic natural resources and the spent industrial and domestic streams to provide a sustainable safe environment for healthy living is a magnum challenge to scientists and environmentalists. The present remedial approach to the wastewater focuses on recovering pure water for reuse and converting the contaminants into a solid matrix for permanent land disposal. However, the ground water aquifers, over a long period slowly leach the contaminants consequently polluting the ground water. Synthetic adsorbents, mainly consisting of polymeric resins, chelating agents, etc. are efficient and have high specificity, but ultimate disposal is a challenge as most of these materials are non-biodegradable. In this context, it is felt appropriate to review the utility of adsorbents based on natural green materials such as agricultural waste and restricted to few model contaminants: phenols, and heavy metals chromium(VI), and cadmium(II) in view of the vast amount of literature available. The article discusses the features of the agricultural waste material-based adsorbents including the mechanism. It is inferred that agricultural waste materials are some of the common renewable sources available across the globe and can be used as sustainable adsorbents. A discussion on challenges for industrial scale implementation and integration with advanced technologies like magnetic-based approaches and nanotechnology to improve the removal efficiency is included for future prospects.

Extraction and detection methods of microplastics in food and marine systems: A critical review.

The ubiquitous presence of microplastics as contaminants in the ecosystem has become a matter of environmental concern gaining considerable attention in the research community as well as public arena. Lack of efficient collection and improper management of plastic have resulted in the enormous amounts of plastic wastes landing into the marine systems with oceans being the ultimate sink. Due to non-biodegradability, these plastics break down into smaller fragments over a period of time leading to consumption by aquatic species, threatening marine life. In the recent years, a wide range of food products has also been contaminated with microplastics directly affecting human health. This review focuses on the separation and identification technologies for extraction and detection of microplastics in food and marine ecosystems. Efficient technologies like floatation, membrane separation, chemical treatment, enzymatic treatment, and other miscellaneous techniques have been discussed considering their merits and demerits. Additionally, identification technologies like optical detection, scanning electron microscopy, Fourier transform infrared spectroscopy, Raman spectroscopy, thermo-analytical methods, and hyperspectral imaging have been emphasized for the detection of microplastic particles. The emerging techniques like enzymatic digestion combined with hyperspectral imaging could be a possible way for obtaining higher separation efficiency and characterization with minimal harm to food sample. This article narrows the gap for choosing a standard separation technology for microplastic detection in food matrices keeping in mind the composition, particle size, shape, data visualization techniques and cost.

A multi-enhancer RET regulatory code is disrupted in Hirschsprung disease.

The major genetic risk factors for Hirschsprung disease (HSCR) are three common polymorphisms within cis-regulatory elements (CREs) of the receptor tyrosine kinase gene RET, which reduce its expression during enteric nervous system (ENS) development. These risk variants attenuate binding of the transcription factors RARB, GATA2, and SOX10 to their cognate CREs, reduce RET gene expression, and dysregulate other ENS and HSCR genes in the RET-EDNRB gene regulatory network (GRN). Here, we use siRNA, ChIP, and CRISPR-Cas9 deletion analyses in the SK-N-SH cell line to ask how many additional HSCR-associated risk variants reside in RET CREs that affect its gene expression. We identify 22 HSCR-associated variants in candidate RET CREs, of which seven have differential allele-specific in vitro enhancer activity, and four of these seven affect RET gene expression; of these, two enhancers are bound by the transcription factor PAX3. We also show that deleting multiple variant-containing enhancers leads to synergistic effects on RET gene expression. These, coupled with our prior results, show that common sequence variants in at least 10 RET enhancers affect HSCR risk, seven with experimental evidence of affecting RET gene expression, extending the known RET-EDNRB GRN to reveal an extensive regulatory code modulating disease risk at a single gene.

Eco-friendly pH detecting paper-based analytical device: Towards process intensification.

This work describes the development of a miniaturized paper-based pH detection platform using natural dye extracted from red cabbage (Brassica oleracea). The easily available paper was used as a substrate and the requisite patterned zone was created with the aid of a punching machine. Experimental parameters were optimized to obtain the best signal readout. The performance of the device at different pH values was quantitatively assessed using digital image analysis with various color space models. Regression analysis suggested that a∗ parameter in CIEL∗a∗b∗ color space model, which captures the variations on the red-green scale, exhibited the best fit with experimental data (R2 = 0.9754). This parameter was used for the quantitative estimation of pH variations in a wide range of pH (1-12). A series of real test samples were examined using the paper-based device and results validated with a standard pH meter. The use of paper and natural dye makes the device eco-friendly. The simplicity of fabrication, ease of usage and low reagent and sample volume requirements render the methodology suitable for in situ measurements of pH. The approach demonstrated here would pave the way for the development of clean, sustainable and intensified chemical sensor technologies.

Sequence-based correction of barcode bias in massively parallel reporter assays.

Massively parallel reporter assays (MPRAs) are a high-throughput method for evaluating in vitro activities of thousands of candidate cis-regulatory elements (CREs). In these assays, candidate sequences are cloned upstream or downstream from a reporter gene tagged by unique DNA sequences. However, tag sequences may themselves affect reporter gene expression and lead to major potential biases in the measured cis-regulatory activity. Here, we present a sequence-based method for correcting tag-sequence-specific effects and show that our method can significantly reduce this source of variation and improve the identification of functional regulatory variants by MPRAs. We also show that our model captures sequence features associated with post-transcriptional regulation of mRNA. Thus, this new method helps not only to improve detection of regulatory signals in MPRA experiments but also to design better MPRA protocols.

Multiple, independent, common variants at RET, SEMA3 and NRG1 gut enhancers specify Hirschsprung disease risk in European ancestry subjects.

PURPOSE: Hirschsprung disease (HSCR) is a developmental disorder of the enteric nervous system (ENS) characterized by congenital aganglionosis arising from coding variants in ENS genes causing partial or total loss-of-function. Low-penetrance, common, noncoding variants at RET, SEMA3 and NRG1 loci are also associated with HSCR, with small-to-moderate loss of gene expression mediated through sequence variants in cis-regulatory elements (CRE) as another causal mechanism. Since these latter variants are common, many individuals carry multiple risk variants. However, the extent and combinatorial effects of all putative CRE variants within and across these loci on HSCR is unknown. METHODS: Using 583 HSCR subjects, one of the largest samples of European ancestry studied, and genotyping 56 tag variants, we evaluated association of all common variants overlapping putative gut CREs and fine-mapped causal variants at RET, SEMA3 and NRG1. RESULTS: We demonstrate that 28 and 8 tag variants, several of which are genetically independent, overlap putative-enhancers at the RET and SEMA3 loci, respectively, as well as two fine-mapped tag variants at the NRG1 locus, are significantly associated with HSCR. Importantly, disease risk increases with increasing numbers of risk alleles from multiple variants within and across these loci, varying >25-fold across individuals. CONCLUSION: This increasing allele number-dependent risk, we hypothesize, arises from HSCR-relevant ENS cells sensing the reduced gene expression at multiple ENS genes since their developmental effects are integrated through gene regulatory networks.

Textile-based Pressure Sensors for Monitoring Prosthetic-Socket Interfaces.

Amputees are prone to experiencing discomfort when wearing their prosthetic devices. As the amputee population grows this becomes a more prevalent and pressing concern. There is a need for new prosthetic technologies to construct more comfortable and well-fitted liners and sockets. One of the well-recognized impediments to the development of new prosthetic technology is the lack of practical inner socket sensors to monitor the inner socket environment (ISE), or the region between the residual limb and the socket. Here we present a capacitive pressure sensor fabricated through a simple, and scalable sewing process using commercially available conductive yarns and textile materials. This fully-textile sensor provides a soft, flexible, and comfortable sensing system for monitoring the ISE. We provide details of our low-power sensor system capable of high-speed data collection from up to four sensor arrays. Additionally, we demonstrate two custom set-ups to test and validate the textile-based sensors in a simulated prosthetic environment. Finally, we utilize the textile-based sensors to study the ISE of a bilateral transtibial amputee. Results indicate that the textile-based sensors provide a promising potential for seamlessly monitoring the ISE.

Techniques and modeling of polyphenol extraction from food: a review.

There is a growing demand for vegetal food having health benefits such as improving the immune system. This is due in particular to the presence of polyphenols present in small amounts in many fruits, vegetables and functional foods. Extracting polyphenols is challenging because extraction techniques should not alter food quality. Here, we review technologies for extracting polyphenolic compounds from foods. Conventional techniques include percolation, decoction, heat reflux extraction, Soxhlet extraction and maceration, whereas advanced techniques are ultrasound-assisted extraction, microwave-assisted extraction, supercritical fluid extraction, high-voltage electric discharge, pulse electric field extraction and enzyme-assisted extraction. Advanced techniques are 32-36% more efficient with approximately 15 times less energy consumption and producing higher-quality extracts. Membrane separation and encapsulation appear promising to improve the sustainability of separating polyphenolic compounds. We present kinetic models and their influence on process parameters such as solvent type, solid and solvent ratio, temperature and particle size.