Innate immune signaling via TLR4 plays critical roles in pathogenesis of metabolic disorders, but the contribution of different lipid species to metabolic disorders and inflammatory diseases is less clear. GM3 ganglioside in human serum is composed of a variety of fatty acids, including long‐chain (LCFA) and very‐long‐chain (VLCFA). Analysis of circulating levels of human serum GM3 species from patients at different stages of insulin resistance and chronic inflammation reveals that levels of VLCFA‐GM3 increase significantly in metabolic disorders, while LCFA‐GM3 serum levels decrease. Specific GM3 species also correlates with disease symptoms. VLCFA‐GM3 levels increase in the adipose tissue of obese mice, and this is blocked in TLR4‐mutant mice. In cultured monocytes, GM3 by itself has no effect on TLR4 activation; however, VLCFA‐GM3 synergistically and selectively enhances TLR4 activation by LPS/HMGB1, while LCFA‐GM3 and unsaturated VLCFA‐GM3 suppresses TLR4 activation. GM3 interacts with the extracellular region of TLR4/MD2 complex to modulate dimerization/oligomerization. Ligand‐molecular docking analysis supports that VLCFA‐GM3 and LCFA‐GM3 act as agonist and antagonist of TLR4 activity, respectively, by differentially binding to the hydrophobic pocket of MD2. Our findings suggest that VLCFA‐GM3 is a risk factor for TLR4‐mediated disease progression.
Examining intestine–liver interactions is important for achieving the desired physiological drug absorption and metabolism response in in vitro drug tests. Multi-organ microphysiological systems (MPSs) constitute promising tools for evaluating inter-organ interactions in vitro. For coculture on MPSs, normal cells are challenging to use because they require complex maintenance and careful handling. Herein, we demonstrated the potential of coculturing normal cells on MPSs in the evaluation of intestine–liver interactions. To this end, we cocultured human-induced pluripotent stem cell-derived intestinal cells and fresh human hepatocytes which were isolated from PXB mice with medium circulation in a pneumatic-pressure-driven MPS with pipette-friendly liquid-handling options. The cytochrome activity, albumin production, and liver-specific gene expressions in human hepatocytes freshly isolated from a PXB mouse were significantly upregulated via coculture with hiPS-intestinal cells. Our normal cell coculture shows the effects of the interactions between the intestine and liver that may occur in vivo. This study is the first to demonstrate the coculturing of hiPS-intestinal cells and fresh human hepatocytes on an MPS for examining pure inter-organ interactions. Normal-cell coculture using the multi-organ MPS could be pursued to explore unknown physiological mechanisms of inter-organ interactions in vitro and investigate the physiological response of new drugs.
In drug discovery, the emergence of unexpected toxicity is often a problem resulting from a poor understanding of the pharmacokinetics of drug–drug interactions (DDI). Organ-on-a-chip (OoC) has been proposed as an in vitro model to evaluate drug efficacy and toxicity in pharmacology, but it has not been applied to DDI studies yet. In this study, we aim to evaluate whether organ-on-a-chip technologies can be applied to DDI studies. To assess the usefulness of OoC for DDI studies, we proposed a multi-organ-on-a-chip (MOoC) with a liver part as the metabolic model and a cancer part as the drug target model, and a pharmacokinetic–pharmacodynamic (PK–PD) model describing the MOoC. An anticancer prodrug, CPT-11, was used to evaluate the drug efficacy of the metabolite in the liver part of the MOoC. To evaluate DDI using the MOoC, the inhibitory effect of simvastatin and ritonavir on the metabolism of CPT-11 was tested. The DDI estimation method was evaluated by comparing the results of the concomitant administration experiment using the MOoC and the results of simulation using the proposed PK–PD model with the estimated parameters. The results were similar, suggesting that the combination of the PK–PD model and the MOoC is a useful way to predict DDI. We conclude that OoC technologies could facilitate a better understanding of pharmacokinetic mechanisms with DDI.
Microphysiological systems (MPSs), including organ-on-a-chip (OoC), have attracted attention as a novel method for estimating the effects and side effects of drugs in drug discovery. To reproduce the dynamic in vivo environment, previous MPSs were connected to pump systems to perfuse culture medium. Therefore, most MPSs are not user-friendly and have poor throughput. We aimed to develop a kinetic pump integrated microfluidic plate (KIM-Plate) by applying the stirrer-based micropump to an open access culture plate to improve the usability of MPSs. The KIM-Plate integrates six multiorgan MPS (MO-MPS) units and meets the ANSI/SBS microplate standards. We evaluated the perfusion function of the kinetic pump and found that the KIM-Plate had sufficient agitation effect. Coculture experiments with PXB cells and hiPS intestinal cells showed that the TEER of hiPS intestinal cells and gene expression levels related to the metabolism of PXB cells were increased. Hence, the KIM-Plate is an innovative tool for the easy coculture of highly conditioned cells that is expected to facilitate cell-based assays in the fields of drug discovery and biology because of its usability and high throughput nature.
The core fucose, a major modification of N‐glycans, is implicated in immune regulation, such as the attenuation of the antibody‐dependent cell‐mediated cytotoxicity of antibody drugs and the inhibition of anti‐tumor responses via the promotion of PD‐1 expression on T cells. Although the core fucose regulates many biological processes, no core fucose recognition molecule has been identified in mammals. Herein, we report that Dectin‐1, a known anti‐β‐glucan lectin, recognizes the core fucose on IgG antibodies. A combination of biophysical experiments further suggested that Dectin‐1 recognizes aromatic amino acids adjacent to the N‐terminal asparagine at the glycosylation site as well as the core fucose. Thus, Dectin‐1 appears to be the first lectin‐like molecule involved in the heterovalent and specific recognition of characteristic N‐glycans on antibodies.
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