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Tissue on a chip or organ-on-chip (OOC) is a technology that's dignified to form a transformation in drug discovery through the use of advanced platforms. These are 3D in-vitro cell culture models that mimic micro-environment of human organs or tissues on artificial microstructures built on a portable microfluidic chip without involving sacrificial humans or animals. This review article aims to offer readers a thorough and insightful understanding of technology. It begins with an in-depth understanding of chip design and instrumentation, underlining its pivotal role and the imperative need for its development in the modern scientific landscape. The review article explores into the myriad applications of OOC technology, showcasing its transformative impact on fields such as radiobiology, drug discovery and screening, and its pioneering use in space research. In addition to highlighting these diverse applications, the article provides a critical analysis of the current challenges that OOC technology faces. It examines both the biological and technical limitations that hinder its progress and efficacy and discusses the potential advancements and innovations that could drive the OOC technology forward. Through this comprehensive review, readers will gain a deep appreciation of the significance, capabilities, and evolving landscape of OOC technology.

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This chapter describes the computational pipeline for the processing and visualization of Protec-Seq data, a method for purification and genome-wide mapping of double-stranded DNA protected by a specific protein at both ends. In the published case, the protein of choice was Saccharomyces cerevisiae Spo11, a conserved topoisomerase-like enzyme that makes meiotic double-strand breaks (DSBs) to initiate homologous recombination, ensuring proper segregation of homologous chromosomes and fertility. The isolated DNA molecules were thus termed double DSB (dDSB) fragments and were found to represent 34 to several hundred base-pair long segments that are generated by Spo11 and are enriched at DSB hotspots, which are sites of topological stress. In order to allow quantitative comparisons between dDSB profiles across experiments, we implemented calibrated chromatin immunoprecipitation sequencing (ChIP-Seq) using the meiosis-competent yeast species Saccharomyces kudriavzevii as calibration strain. Here, we provide a detailed description of the computational methods for processing, analyzing, and visualizing Protec-Seq data, comprising the download of the raw data, the calibrated genome-wide alignments, and the scripted creation of either arc plots or Hi-C-style heatmaps for the illustration of chromosomal regions of interest. The workflow is based on Linux shell scripts (including wrappers for publicly available, open-source software) as well as R scripts and is highly customizable through its modular structure.

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Hi-C is a powerful method for obtaining genome-wide chromosomal structural information. The typical Hi-C analysis utilizes a two-dimensional (2D) contact matrix, which poses challenges for quantitative comparisons, visualizations, and integrations across multiple datasets. Here, we present a protocol for extracting one-dimensional (1D) features from chromosome structure data by HiC1Dmetrics. Leveraging these 1D features enables integrated analysis of Hi-C and epigenomic data.

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In recent years, advances in microfabrication technology and tissue engineering have propelled the development of a novel drug screening and disease modelling platform known as organoid-on-a-chip. This platform integrates organoids and organ-on-a-chip technologies, emerging as a promising approach for in vitro modelling of human organ physiology. Organoid-on-a-chip devices leverage microfluidic systems to simulate the physiological microenvironment of specific organs, offering a more dynamic and flexible setting that can mimic a more comprehensive human biological context. However, the lack of functional vasculature has remained a significant challenge in this technology. Vascularisation is crucial for the long-term culture and in vitro modelling of organoids, holding important implications for drug development and personalised medical approaches. This review provides an overview of research progress in developing vascularised organoid-on-a-chip models, addressing methods for in vitro vascularisation and advancements in vascularised organoids. The aim is to serve as a reference for future endeavors in constructing fully functional vascularised organoid-on-a-chip platforms.

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This study aimed to assess the nephrotoxicity associated with VRP-034 (novel formulation of polymyxin B [PMB]) compared to marketed PMB in a three-dimensional (3D) kidney-on-a-chip model. To model the human kidney proximal tubule for analysis, tubular structures were established using 23 triple-channel chips seeded with RPTEC/hTERT1 cells. These cells were exposed to VRP-034 or PMB at seven concentrations (1-200 µM) over 12, 24, and 48 h. A suite of novel kidney injury biomarkers, cell health, and inflammatory markers were quantitatively assessed in the effluent. Additionally, caspase and cytochrome C levels were measured, and cell viability was evaluated using calcein AM and ethidium homodimer-1 (EthD-1). Exposure to marketed PMB resulted in significantly elevated levels (P < 0.05) of four key biomarkers (KIM-1, cystatin C, clusterin, and OPN) compared to VRP-034, particularly at clinically relevant concentrations of ≥10 µM. At 25 µM, all biomarkers demonstrated a significant increase (P < 0.05) with marketed PMB exposure compared to VRP-034. Inflammatory markers (interleukin-6 and interleukin-8) increased significantly (P < 0.05) with marketed PMB at concentrations of ≥5 µM, relative to VRP-034. VRP-034 displayed superior cell health outcomes, exhibiting lower lactate dehydrogenase release, while ATP levels remained comparable. Morphological analysis revealed that marketed PMB induced more severe damage, disrupting tubular integrity. Both treatments activated cytochrome C, caspase-3, caspase-8, caspase-9, and caspase-12 in a concentration-dependent manner; however, caspase activation was significantly reduced (P < 0.05) with VRP-034. This study demonstrates that VRP-034 significantly reduces nephrotoxicity compared to marketed PMB within a 3D microphysiological system, suggesting its potential to enable the use of full therapeutic doses of PMB with an improved safety profile, addressing the need for less nephrotoxic polymyxin antibiotics.

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The availability of high-frequency, real-time measurements of the concentrations of specific metabolites in cell culture systems will enable a deeper understanding of cellular metabolism and facilitate the application of good laboratory practice standards in cell culture protocols. However, currently available approaches to this end either are constrained to single-time-point and single-parameter measurements or are limited in the range of detectable analytes. Electrochemical aptamer-based (EAB) biosensors have demonstrated utility in real-time monitoring of analytes in vivo in blood and tissues. Here, we characterize a pH-sensing capability of EAB sensors that is independent of the specific target analyte of the aptamer sequence. We applied this dual-purpose EAB to the continuous measurement of pH and phenylalanine in several in vitro cell culture settings. The miniature EAB sensor that we developed exhibits rapid response times, good stability, high repeatability, and biologically relevant sensitivity. We also developed and characterized a leak-free reference electrode that mitigates the potential cytotoxic effects of silver ions released from conventional reference electrodes. Using the resulting dual-purpose sensor, we performed hourly measurements of pH and phenylalanine concentrations in the medium superfusing cultured epithelial tumor cell lines (A549, MDA-MB-23) and a human fibroblast cell line (MRC-5) for periods of up to 72 h. Our scalable technology may be multiplexed for high-throughput monitoring of pH and multiple analytes in support of the broad metabolic qualification of microphysiological systems.

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The lack of representative in vitro models recapitulating human tendon (patho)physiology is among the major factors hindering consistent progress in the knowledge-based development of adequate therapies for tendinopathy.Here, an organotypic 3D tendon-on-chip model is designed that allows studying the spatiotemporal dynamics of its cellular and molecular mechanisms.Combining the synergistic effects of a bioactive hydrogel matrix with the biophysical cues of magnetic microfibers directly aligned on the microfluidic chip, it is possible to recreate the anisotropic architecture, cell patterns, and phenotype of tendon intrinsic (core) compartment. When incorporated with vascular-like vessels emulating the interface between its intrinsic-extrinsic compartments, crosstalk with endothelial cells are found to drive stromal tenocytes toward a reparative profile. This platform is further used to study adaptive immune cell responses at the onset of tissue inflammation, focusing on interactions between tendon compartment tenocytes and circulating T cells.The proinflammatory signature resulting from this intra/inter-cellular communication induces the recruitment of T cells into the inflamed core compartment and confirms the involvement of this cellular crosstalk in positive feedback loops leading to the amplification of tendon inflammation.Overall, the developed 3D tendon-on-chip provides a powerful new tool enabling mechanistic studies on the pathogenesis of tendinopathy as well as for assessing new therapies.

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Metastasis contributes to the increased mortality rate of cancer, but the intricate mechanisms remain unclear. Cancer cells from a primary tumor invade nearby tissues and access the lymphatic or circulatory system. If these cells manage to survive and extravasate from the vasculature into distant tissues and ultimately adapt to survive, they will proliferate and facilitate malignant tumor formation. Traditional two-dimensional (2D) cell cultures offer a rapid and convenient method for validating the efficacy of anticancer drugs within a reasonable cost range, but their utility is limited because of tumors' high heterogeneity in vivo and spatial complexities. Three-dimensional (3D) cell cultures that mimic the physiological conditions of cancer cells in vivo have gained considerable interest. In these cultures, cells assemble into spheroids through gravity, magnetic forces, or their low-adhesion to the plates. Although these approaches address some of the limitations of 2D cultures, they often require a considerable amount of time and cost. Therefore, this study aims to enhance the effectiveness of 3D culture techniques by using microfluidic systems to provide a high-throughput and sensitive pipeline for drug screening. Using these systems, we studied the effects of lanthanide elements, which have garnered interest in cancer treatment, on spheroid formation and cell spreading. Our findings suggest that these elements alter the compactness of cell spheroids and decrease cell mobility.

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Impaired neutrophil migration in sepsis is associated with a poor prognosis. The potential of utilizing neutrophil chemotaxis to assess immune function, disease severity, and patient prognosis in sepsis remains underexplored. This study employed an innovative approach by integrating a multi-tip pipette with a Six-Unit microfluidic chip (SU6-chip) to establish gradients in six microchannels, thereby analyzing neutrophil chemotaxis in sepsis patients. We compared chemotactic parameters between healthy controls (NH = 20) and sepsis patients (NS1 = 25), observing significant differences in gradient perception time (GP), migration distance (MD), peak velocity (Vmax), chemotactic index (CI), reverse migration rate (RM), and stop migration number (SM). A novel composite indicator, the Sepsis Neutrophil Migration Evaluation (SNME) index, was developed by integrating these six chemotactic migration parameters. The SNME index and individual chemotaxis parameters showed significant correlations with the Sequential Organ Failure Assessment (SOFA) score, Acute Physiology and Chronic Health Evaluation (APACHE II) score, hypersensitivity C-reactive protein (hs-CRP), and heparin-binding protein (HBP). Moreover, the SNME index demonstrated potential for monitoring sepsis progression, with ROC analysis confirming its predictive accuracy (area under the curve [AUC] = 0.895, cutoff value = 31.5, specificity = 86.73 %, sensitivity = 86.71 %), outperforming individual neutrophil chemotactic parameters. In conclusion, the SNME index represents a promising new tool for adjunctive diagnosis and prognosis assessment in patients with sepsis.

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Lung cancer remains a global health concern, demanding the development of noninvasive, prompt, selective, and point-of-care diagnostic tools. Correspondingly, breath analysis using nanobiosensors has emerged as a promising noninvasive nose-on-chip technique for the early detection of lung cancer through monitoring diversified biomarkers such as volatile organic compounds/gases in exhaled breath. This comprehensive review summarizes the state-of-the-art breath-based lung cancer diagnosis employing chemiresistive-module nanobiosensors supported by theoretical findings. It unveils the fundamental mechanisms and biological basis of breath biomarker generation associated with lung cancer, technological advancements, and clinical implementation of nanobiosensor-based breath analysis. It explores the merits, challenges, and potential alternate solutions in implementing these nanobiosensors in clinical settings, including standardization, biocompatibility/toxicity analysis, green and sustainable technologies, life-cycle assessment, and scheming regulatory modalities. It highlights nanobiosensors' role in facilitating precise, real-time, and on-site detection of lung cancer through breath analysis, leading to improved patient outcomes, enhanced clinical management, and remote personalized monitoring. Additionally, integrating these biosensors with artificial intelligence, machine learning, Internet-of-things, bioinformatics, and omics technologies is discussed, providing insights into the prospects of intelligent nose-on-chip lung cancer sniffing nanobiosensors. Overall, this review consolidates knowledge on breathomic biosensor-based lung cancer screening, shedding light on its significance and potential applications in advancing state-of-the-art medical diagnostics to reduce the burden on hospitals and save human lives.

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Somatic activating mutations in PIK3CA are common drivers of vascular and lymphatic malformations. Despite common biophysical signatures of tissues susceptible to lesion formation, including compliant extracellular matrix and low rates of perfusion, lesions vary in clinical presentation from localized cystic dilatation to diffuse and infiltrative vascular dysplasia. The mechanisms driving the differences in disease severity and variability in clinical presentation and the role of the biophysical microenvironment in potentiating progression are poorly understood. Here, we investigate the role of hemodynamic forces and the biophysical microenvironment in the pathophysiology of vascular malformations, and we identify hemodynamic shear stress and defective endothelial cell mechanotransduction as key regulators of lesion progression. We found that constitutive PI3K activation impaired flow-mediated endothelial cell alignment and barrier function. We show that defective shear stress sensing in PIK3CA E542K endothelial cells is associated with reduced myosin light chain phosphorylation, junctional instability, and defective recruitment of vinculin to cell-cell junctions. Using 3D microfluidic models of the vasculature, we demonstrate that PIK3CA E542K microvessels apply reduced traction forces and are unaffected by flow interruption. We further found that draining transmural flow resulted in increased sprouting and invasion responses in PIK3CA E542K microvessels. Mechanistically, constitutive PI3K activation decreased cellular and nuclear elasticity resulting in defective cellular tensional homeostasis in endothelial cells which may underlie vascular dilation, tissue hyperplasia, and hypersprouting in PIK3CA-driven venous and lymphatic malformations. Together, these results suggest that defective nuclear mechanics, impaired cellular mechanotransduction, and maladaptive hemodynamic responses contribute to the development and progression of PIK3CA-driven vascular malformations.

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BACKGROUND: Sepsis, which is characterized by acute systemic inflammation and is associated with high rates of morbidity and mortality, presents a significant challenge in health care. Some scholars have found that the sequential organ failure assessment (SOFA) and quick SOFA scores are not ideal for predicting severe sepsis and mortality. Microbial culture takes a long time (2-3 d) and provides no information for early diagnosis and treatment. Therefore, new diagnostic methods for sepsis need to be explored. AIM: To assess cytokine levels in the plasma of sepsis patients and identify potential biomarkers for diagnosing sepsis. METHODS: Ten sepsis patients admitted to the emergency department within 24 h of onset were enrolled as the observation group, whereas ten noninfected patients served as the control group. Of the 10 noninfected patients, 9 hypertension combined with cerebral infarction, 1 patients with vertiginous syndrome. Plasma Cytokines were measured using the Bio-Plex Pro™ Human Chemokine Panel 40-plex. Differentially expressed cytokines in plasma of sepsis and nonsepsis patients were analyzed using Gene Ontology (GO) functional enrichment and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses. RESULTS: Interleukin (IL)-16, granulocyte-macrophage granulocyte-macrophage colony-stimulating factor (GM-CSF), CX3CL1, CXCL9, CXCL16, CCL25, and CCL23 plasma levels were significantly increased in sepsis patients. GO analysis revealed that these cytokines were mainly associated with cellular structures such as intermediates, nuclear plaques, adhesion plaques, lateral plasma membranes, and cell matrix junctions. These genes were involved in various molecular functions, such as cytokine activity, receptor ligand activity, and signal receptor activator activity, contributing to various biological functions, such as leukocyte chemotaxis, migration, and chemotaxis. KEGG analysis indicated involvement in cytokine cytokine receptor interactions, chemokine signaling pathways, virus-protein interactions with cytokines and cytokine receptors, and the tumor necrosis factor signaling pathway. CONCLUSION: Elevated serum levels of IL-16, GM-CSF, CX3CL1, CXCL9, CXCL16, CCL25, and CCL23 in sepsis patients suggest their potential as diagnostic biomarkers for sepsis.

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The Fengjing pig is one of the local pig breed resources in China and has many excellent germplasm characteristics. However, research on its genome is lacking. To explore the degree of genetic diversity of the Fengjing pig and to deeply explore its excellent traits, this study took Fengjing pigs as the research object and used the Beadchip Array Infinium iSelect-96|XT KPS_PorcineBreedingChipV2 for genotyping. We analyzed the genetic diversity, relatedness, inbreeding coefficient, and population structure within the Fengjing pig population. Our findings revealed that the proportion of polymorphic markers (PN) was 0.469, and the effective population size was 6.8. The observed and expected heterozygosity were 0.301 and 0.287, respectively. The G-matrix results indicated moderate relatedness within the population, with certain individuals exhibiting closer genetic relationships. The NJ evolutionary tree classified Fengjing boars into five family lines. The average inbreeding coefficient based on ROH was 0.318, indicating a high level of inbreeding. GWAS identified twenty SNPs significantly associated with growth traits (WW, 2W, and 4W) and reproductive traits (TNB and AWB). Notably, WNT8B, RAD21, and HAO1 emerged as candidate genes influencing 2W, 4W, and TNB, respectively. Genes such as WNT8B were verified by querying the PigBiobank database. In conclusion, this study provides a foundational reference for the conservation and utilization of Fengjing pig germplasm resources and offers insights for future molecular breeding efforts in Fengjing pigs.

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Ischemic stroke has garnered global medical attention as one of the most serious cerebrovascular diseases. The mechanisms involved in both the development and recovery phases of ischemic stroke are complex, involving intricate interactions among different types of cells, each with its own unique functions. To better understand the possible pathogenesis, neurovascular unit (NVU), a concept comprising neurons, endothelial cells, mural cells, glial cells, and extracellular matrix components, has been used in analysing various brain diseases, particularly in ischemic stroke, aiming to depict the interactions between cerebral vasculature and neural cells. While in vivo models often face limitations in terms of reproducibility and the ability to precisely mimic human pathophysiology, it is now important to establish in vitro NVU models for ischemic stroke research. In order to accurately portray the pathological processes occurring within the brain, a diverse array of NVU 2D and 3D in vitro models, each possessing unique characteristics and advantages, have been meticulously developed. This review presents a comprehensive overview of recent advancements in in vitro models specifically tailored for investigating ischemic stroke. Through a systematic categorization of these developments, we elucidate the intricate links between NVU components and the pathogenesis of ischemic stroke. Furthermore, we explore the distinct advantages offered by innovative NVU models, notably 3D models, which closely emulate in vivo conditions. Additionally, an examination of current therapeutic modalities for ischemic stroke developed utilizing in vitro NVU models is provided. Serving as a valuable reference, this review aids in the design and implementation of effective in vitro models for ischemic stroke research.

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The quality and authenticity of milk are of paramount importance. Cow milk is more allergenic and less nutritious than ewe, goat, or donkey milk, which are often adulterated with cow milk due to their seasonal availability and higher prices. In this work, a silicon photonic dipstick sensor accommodating two U-shaped Mach-Zehnder Interferometers (MZIs) was employed for the label-free detection of the adulteration of ewe, goat, and donkey milk with cow milk. One of the two MZIs of the chip was modified with bovine κ-casein, while the other was modified with bovine serum albumin to serve as a blank. All assay steps were performed by immersion of the chip side where the MZIs are positioned into the reagent solutions, leading to a photonic dipstick immunosensor. Thus, the chip was first immersed in a mixture of milk with anti-bovine κ-casein antibody and then in a secondary antibody solution for signal enhancement. A limit of detection of 0.05% v/v cow milk in ewe, goat, or donkey milk was achieved in 12 min using a 50-times diluted sample. This fast, sensitive, and simple assay, without the need for sample pre-processing, microfluidics, or pumps, makes the developed sensor ideal for the detection of milk adulteration at the point of need.

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As semiconductor chip manufacturing technology advances, chip structures are becoming more complex, leading to an increased likelihood of void defects in the solder layer during packaging. However, identifying void defects in packaged chips remains a significant challenge due to the complex chip background, varying defect sizes and shapes, and blurred boundaries between voids and their surroundings. To address these challenges, we present a deep-learning-based framework for void defect segmentation in chip packaging. The framework consists of two main components: a solder region extraction method and a void defect segmentation network. The solder region extraction method includes a lightweight segmentation network and a rotation correction algorithm that eliminates background noise and accurately captures the solder region of the chip. The void defect segmentation network is designed for efficient and accurate defect segmentation. To cope with the variability of void defect shapes and sizes, we propose a Mamba model-based encoder that uses a visual state space module for multi-scale information extraction. In addition, we propose an interactive dual-stream decoder that uses a feature correlation cross gate module to fuse the streams' features to improve their correlation and produce more accurate void defect segmentation maps. The effectiveness of the framework is evaluated through quantitative and qualitative experiments on our custom X-ray chip dataset. Furthermore, the proposed void defect segmentation framework for chip packaging has been applied to a real factory inspection line, achieving an accuracy of 93.3% in chip qualification.

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Extracellular vesicles (EVs) are considered as promising candidates for predicting patients who respond to immunotherapy. Nevertheless, simultaneous detection of multiple EVs markers still presents significant technical challenges. In this work, we developed a high-throughput microdroplet-enhanced chip (MEC) platform, which utilizes thousands of individual microchambers (∼pL) as reactors, accelerating the detection efficiency of the CRISPR/Cas systems and increasing the sensitivity by up to 100-fold (aM level). Ten biomarkers (including 5 RNAs and 5 proteins) from patients' EVs are successfully detected on one chip, and the comprehensive markers show increased accuracy (AUC 0.911) than the individual marker for the efficacy prediction of immunotherapy. This platform provides a high-throughput yet sensitive strategy for screening immunotherapy markers in clinical.

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Clonal hematopoiesis of indeterminate potential (CHIP) is a well-studied phenomenon in hematologic malignancies. With advancements in gene sampling and analysis and the use of large cohort studies, CHIP has recently been linked to cardiovascular disease (CVD). The relationship between CHIP and CVD appears to be bidirectional, with traditional risk factors for cardiovascular disease increasing the mutation burden in CHIP, and CHIP itself effecting the incidence or prognosis of a variety of CVD. The purpose of this review is to understand the epidemiology, risk factors, and pathogenesis of CHIP in the context of various CVD conditions.

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Breast cancer, the most common cancer among women worldwide, lacks specific tumor markers for accurate diagnosis. Recent advances have highlighted tumor-derived exosomes as a promising non-invasive biomarker for cancer detection. Continuous monitoring of surface protein markers on exosomes in the blood could offer valuable insights for breast cancer diagnosis. However, integrating the isolation and detection of exosomes from whole blood is bulky, time-consuming, and requires professional operations. To address this difficulty, we developed a method of integrated centrifugal disk chip (CD chip) exosome enrichment directly from whole blood followed by a colorimetric visualization strategy for multiplex analysis. The disc consists of multi-chambers and multi-microchannels with immediate smartphone-enabled processing of colorimetric results. The combination of CEA + CA125 + EGFR on-chip detection could significantly differentiate the different stages of cancer in tumor-bearing mice and successfully distinguish between breast cancer patients and healthy individuals. Crucially, small volumes (100 µL) of blood samples were adequate. In addition, the chip was simple and fast, with results within 10 min, which provides immediate exosomal information through consecutive blood sampling, which could potentially result in a more timely and well-informed clinical breast cancer diagnosis.

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Excessive fluoride ingestion during tooth development can cause dental fluorosis. Previously, we reported that fluoride activates histone acetyltransferase (HAT) to acetylate p53, promoting fluoride toxicity in mouse ameloblast-like LS8 cells. However, the roles of HAT and histone acetylation status in fluoride-mediated gene expression remain unidentified. Here, we demonstrate that fluoride-mediated histone modification causes gene expression alterations in LS8 cells. LS8 cells were treated with or without fluoride followed by ChIP-Seq analysis of H3K27ac. Genes were identified by differential H3K27ac peaks within ±1 kb from transcription start sites. The levels of mRNA of identified genes were assessed using rea-time PCR (qPCR). Fluoride increased H3K27ac peaks associated with Bax, p21, and Mdm2 genes and upregulated their mRNA levels. Fluoride decreased H3K27ac peaks and p53, Bad, and Bcl2 had suppressed transcription. HAT inhibitors (Anacardic acid or MG149) suppressed fluoride-induced mRNA of p21 and Mdm2, while fluoride and the histone deacetylase (HDAC) inhibitor sodium butyrate increased Bad and Bcl2 expression above that of fluoride treatment alone. To our knowledge, this is the first study that demonstrates epigenetic regulation via fluoride treatment via H3 acetylation. Further investigation is required to elucidate epigenetic mechanisms of fluoride toxicity in enamel development.

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