Pump‐Less, Recirculating Organ‐on‐Chip (rOoC) Platform to Model the Metabolic Crosstalk between Islets and Liver

Advanced Healthcare Materials, Journal Year: 2024, Volume and Issue: 13(13)

Published: Jan. 15, 2024

Abstract Type 2 diabetes mellitus (T2DM), obesity, and metabolic dysfunction‐associated steatotic liver disease (MASLD) are epidemiologically correlated disorders with a worldwide growing prevalence. While the mechanisms leading to onset development of these conditions not fully understood, predictive tissue representations for studying coordinated interactions between central organs that regulate energy metabolism, particularly pancreatic islets, needed. Here, dual pump‐less recirculating organ‐on‐chip platform combines human pluripotent stem cell (sc)‐derived sc‐liver sc‐islet organoids is presented. The reproduces key aspects cross‐talk both organs, including glucose levels selected hormones, supports viability functionality while preserving reduced release pro‐inflammatory cytokines. In model disruption in response treatment high lipids fructose, exhibit hallmarks steatosis insulin resistance, sc‐islets produce cytokines on‐chip. Finally, known effects anti‐diabetic drugs Taken together, provides basis functional studies T2DM, MASLD on‐chip, as well testing potential therapeutic interventions.

Language: Английский

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Online LC-MS/MS Analysis for Profiling Peptide Hormone Secretion Dynamics from Islets of Langerhans

Analytical Chemistry, Journal Year: 2025, Volume and Issue: unknown

Published: Feb. 11, 2025

Multiple peptide hormones are secreted from islets of Langerhans to maintain blood glucose homeostasis. Defects in the amount and patterns hormone secretion can lead metabolic disorders, such as diabetes. To understand relationships between peptides, analytical methods that quantify multiple short time increments required. this end, an automated online system was developed for sampling perfusate ∼30 human held on a glass microfluidic device with sequential liquid chromatography (LC)-MS/MS runs every 2 min resolve dynamics. Islet mixed isotopically labeled internal standard loaded into μL sample loop which injected 0.1 1.9 onto 2.1 mm × 30 (I.D. length) C18 column at 70 °C. Online detection insulin, C-peptide, glucagon, somatostatin levels performed using triple quadrupole mass spectrometer. Optimization separation conditions linear solvent strength theory enabled rapid four peptides. Calibration curves were 0.5 50 nM RSD all analytes 3-15% <3% retention times. Results showed dynamics first-phase insulin release negatively correlated glucagon insulin. This simple LC-MS method used single 6-port valve is expected be useful examining other biologically relevant molecules could applied biological systems investigate cellular communication.

Language: Английский

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Electromembrane Extraction Provides Unprecedented Selectivity for Drugs in Cell Culture Media Used in Organoid and Organ-on-Chip Systems

Analytical Chemistry, Journal Year: 2025, Volume and Issue: unknown

Published: Feb. 24, 2025

The use of organoids and organ-on-chip technologies as nonanimal methodologies in drug discovery personalized medicine is rapidly expanding. However, the complexity small volumes organoid culture samples present significant analytical challenges, e.g., analysis using liquid chromatography–mass spectrometry (LC–MS). Essentially an electrophoresis across oil membrane, electromembrane extraction (EME) offers a promising approach for measuring drugs, it is, example, compatible with such formats. Given potential technology, there need to assess purity EME extracts ensure EME's compatibility high-throughput, downstream analysis. This study evaluates effectiveness sample cleanup various common cell media used organs-on-chips. were spiked 90 small-molecule drugs. Using gel (sodium dodecyl sulfate polyacrylamide electrophoresis), high-resolution nuclear magnetic resonance spectroscopy, LC–MS, we demonstrate that provides exhaustive removal unwanted medium components (proteins, polar molecules, apolar/neutral molecules) while selectively extracting was demonstrated human stem-cell-derived liver organoids, allowing simple detection monitoring telltale cytochrome P450 metabolism. Taken together, our observations highlight unprecedented ability provide matrixes technology.

Language: Английский

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Preparative agarose gel electrophoresis for reducing matrix interferences of organoid cell medium prior to LC-MS analysis of insulin

Journal of Chromatography A, Journal Year: 2024, Volume and Issue: 1717, P. 464669 - 464669

Published: Jan. 20, 2024

Organoids are 3D cell cultures with microanatomies mimicking aspects of real organs, useful for e.g. animal-free studies development, disease, and drug discovery. The medium organoid models Langerhans islets, regulating blood glucose levels by insulin secretion, can be analyzed liquid chromatography-mass spectrometry (LC-MS). However, complexity is a major challenge, as matrix interferences reduce sensitivity selectivity, even optimized LC-MS conditions. By applying preparative agarose gel electrophoresis-electrodialysis (PGE-ED), we were able to decrease the background signal, allowing reduced affecting analysis human insulin.

Language: Английский

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An FDA-Validated, Self-Cleaning Liquid Chromatography–Mass Spectrometry System for Determining Small-Molecule Drugs and Metabolites in Organoid/Organ-on-Chip Medium

Analytical Chemistry, Journal Year: 2024, Volume and Issue: 96(29), P. 12129 - 12138

Published: July 10, 2024

As organoids and organ-on-chip (OoC) systems move toward preclinical clinical applications, there is an increased need for method validation. Using a liquid chromatography-mass spectrometry (LC-MS)-based approach, we developed measuring small-molecule drugs metabolites in the cell medium directly sampled from liver organoids/OoC systems. The LC-MS setup was coupled to automatic filtration filter flush system with online solid-phase extraction (SPE), allowing robust automated sample cleanup/analysis. For matrix, rich in, e.g., protein, salts, amino acids, no preinjection preparation steps (protein precipitation, SPE, etc.) were necessary. approach demonstrated tolbutamide its metabolite, 4-hydroxytolbutamide (4HT). validated analysis of media human stem cell-derived cultured static conditions on microfluidic platform according Food Drug Administration (FDA) guidelines regards selectivity, matrix effects, accuracy, precision, etc. allows hundreds injections without replacing chromatography hardware. In summary, drug/metabolite organoids/OoCs can be performed robustly minimal preparation.

Language: Английский

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An LC-MS/MS Method for the Simultaneous Quantification of Insulin, Cortisol, Glucagon-like Peptide 1, Ghrelin, and Osteocalcin

Separations, Journal Year: 2024, Volume and Issue: 11(2), P. 41 - 41

Published: Jan. 27, 2024

Hormones are important signaling molecules controlling physiological homeostasis. ELISA kits commonly used to measure hormones; however, few multiplex, not all species-specific commercially available, and typically require a significant volume of biological fluids. Pigs resemble humans in digestive physiology, making them an excellent model preclinical research nutrition metabolism. In this study, we developed validated simple liquid–liquid extraction procedure LC-MS/MS method for the simultaneous quantification insulin, cortisol, glucagon-like peptide-1 (GLP-1) (7-37) (7-36), acyl des-acyl ghrelin, carboxylated osteocalcin pig serum. The proposed is specific, highly sensitive (LOQ ng/mL pg/mL), reasonably accurate (more than 76.2% quality control samples within 20% error from nominal values), precise (intra-day CV ≤ 10% inter-day 23.1%). recoveries analytes corresponding internal standards ranged 83.7 116.0%. also requires low serum 50–100 μL, which invaluable when sample limited. These methods could be easily extended use other mammalian species.

Language: Английский

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Liquid chromatography mass spectrometry-based approaches for determination of pancreatic hormones

Journal of Chromatography Open, Journal Year: 2024, Volume and Issue: 6, P. 100143 - 100143

Published: June 15, 2024

Pancreatic hormones are produced in the islets of Langerhans pancreas and secreted to regulate blood sugar levels. Key pancreatic insulin, glucagon, somatostatin. The measurement these peptides/small proteins is essential study endocrine system, regulating diseases such as diabetes mellitus, but also doping control forensics. In this review, we focus on approaches for measuring using liquid chromatography-mass spectrometry (LC–MS). A rich variety LC–MS used, including variations LC column dimensions chemistry, factors that crucial regarding sensitivity, selectivity, robustness, speed. addition, several sample preparation methods used (protein precipitation, immunopurification, etc.), analysis matrices blood, urine, cell culture medium. There variants mass spectrometric analysis, both targeted non-targeted, lower high-resolution instruments. allows sensitive, multi-hormone measurements, confidently distinguishing between their analogs, metabolites, degradation products. speed can be pushed down a few minutes, with being validated clinically applied. review focuses papers have been published 2019 2024.

Language: Английский

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