An Organ-on-Chip Platform for Simulating Drug Metabolism Along the Gut-Liver Axis.

The human microbiome significantly influences drug metabolism through the gut-liver axis, leading to modified drug responses and potential toxicity. Due to the complex nature of the human gut environment, the understanding of microbiome-driven impacts on these processes is limited. To address this, a multiorgan-on-a-chip (MOoC) platform that combines the human microbial-crosstalk (HuMiX) gut-on-chip (GoC) and the Dynamic42 liver-on-chip (LoC), mimicking the bidirectional interconnection between the gut and liver known as the gut-liver axis, is introduced. This platform supports the viability and functionality of intestinal and liver cells. In a proof-of-concept study, the metabolism of irinotecan, a widely used colorectal cancer drug, is imitated within the MOoC. Utilizing liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS), irinotecan metabolites are tracked, confirming the platform's ability to represent drug metabolism along the gut-liver axis. Further, using the authors' gut-liver platform, it is shown that the colorectal cancer-associated gut bacterium, Escherichia coli, modifies irinotecan metabolism through the transformation of its inactive metabolite SN-38G into its toxic metabolite SN-38. This platform serves as a robust tool for investigating the intricate interplay between gut microbes and pharmaceuticals, offering a representative alternative to animal models and providing novel drug development strategies.

Integration of multiple flexible electrodes for real-time detection of barrier formation with spatial resolution in a gut-on-chip system.

In healthy individuals, the intestinal epithelium forms a tight barrier to prevent gut bacteria from reaching blood circulation. To study the effect of probiotics, dietary compounds and drugs on gut barrier formation and disruption, human gut epithelial and bacterial cells can be cocultured in an in vitro model called the human microbial crosstalk (HuMiX) gut-on-a-chip system. Here, we present the design, fabrication and integration of thin-film electrodes into the HuMiX platform to measure transepithelial electrical resistance (TEER) as a direct readout on barrier tightness in real-time. As various aspects of the HuMiX platform have already been set in their design, such as multiple compressible layers, uneven surfaces and nontransparent materials, a novel fabrication method was developed whereby thin-film metal electrodes were first deposited on flexible substrates and sequentially integrated with the HuMiX system via a transfer-tape approach. Moreover, to measure localized TEER along the cell culture chamber, we integrated multiple electrodes that were connected to an impedance analyzer via a multiplexer. We further developed a dynamic normalization method because the active measurement area depends on the measured TEER levels. The fabrication process and system setup can be applicable to other barrier-on-chip systems. As a proof-of-concept, we measured the barrier formation of a cancerous Caco-2 cell line in real-time, which was mapped at four spatially separated positions along the HuMiX culture area.

Emulating the gut-liver axis: Dissecting the microbiome's effect on drug metabolism using multiorgan-on-chip models.

The homeostatic relationship between the gut, its microbiome, and the liver is crucial for the regulation of drug metabolism processes. Gut microbes are known to influence human health and disease by enhancing food metabolism and providing a first line of defense against pathogens. In addition to this, the gut microbiome also plays a key role in the processing of exogenous pharmaceutical compounds. Modeling the highly variable luminal gut environment and understanding how gut microbes can modulate drug availability or induce liver toxicity remains a challenge. However, microfluidics-based technologies such as organ-on-chips could overcome current challenges in drug toxicity assessment assays because these technologies are able to better recapitulate complex human responses. Efforts are being made to create in vitro multiorgan platforms, tailored for an individual patient's microbial background. These platforms could be used as a tool to predict the effect of the gut microbiome on pharmacokinetics in a personalized way.

Short Communication: An Insertion of Seven Amino Acids in the Envelope Cytoplasmic Tail of HIV-2 Selected During Disease Progression Enhances Viral Replication.

The cytoplasmic tail (CT) of the HIV-2 envelope glycoprotein (Env) includes amino acid (aa) sequences that are similar to lentiviral lytic peptides (LLP) described in other lentiviruses. Within the putative LLP-2 region, we previously observed insertions of 3 or 7 aa in sequences deduced from plasma viral RNA of symptomatic HIV-2-infected individuals. Based on these observations, we reproduced the insertions in a molecular clone to assess their impact on replicative fitness and cell death in vitro. Using a molecular clone of the HIV-2ROD reference strain, site-directed mutagenesis experiments allowed the generation of plasmids with the insertion L791TAI or L791QRALTAI in the Env protein. The clone with 7 aa insertion enhanced viral release 8 to 11 times in infected T cells and cell viability was impaired by more than 20%, compared with the wild-type HIV-2ROD virus. The effect of the 3 aa insertion was milder, with a nonsignificant trend to enhance viral replication and cell death compared with the wild-type virus. Interestingly, the insertions in the Env proteins did not induce a significant increase of viral infectivity, as revealed by the infectivity assay using TZM-bl cells. The insertions in the Env CT observed in vivo from disease progressors may, therefore, be involved in the higher viral load observed in these individuals. This study may open the way to the development of a prognostic marker related to the HIV-2 infection progression.

Modulation of the NF-κB signaling pathway by the HIV-2 envelope glycoprotein and its incomplete BST-2 antagonism.

The HIVs have evolved by selecting means to hijack numerous host cellular factors. HIVs exploit the transcription factor NF-κB to ensure efficient LTR-driven gene transcription. However, NF-κB is primarily known to act as a key regulator of the proinflammatory and antiviral responses. Interestingly, retroviruses activate NF-κB during early stages of infection to initiate proviral genome expression while suppressing it at later stages to restrain expression of antiviral genes. During HIV-1 infection, diverse viral proteins such as Env, Nef and Vpr have been proposed to activate NF-κB activity, whereas Vpu has been shown to inhibit NF-κB activation. It is still unclear how HIV-2 regulates NF-κB signaling pathway during its replication cycle. Here we confirm that human BST-2 and HIV-1 Env proteins can trigger potent activation of NF-κB. Importantly, we demonstrate for the first time that the HIV-2 Env induces NF-κB activation in HEΚ293T cells. Furthermore, the anti-BST-2 activity of the HIV-2 Env is not sufficient to completely inhibit NF-κB activity.