ICAM-1 nanoclusters regulate hepatic epithelial cell polarity by leukocyte adhesion-independent control of apical actomyosin.

Epithelial intercellular adhesion molecule (ICAM)-1 is apically polarized, interacts with, and guides leukocytes across epithelial barriers. Polarized hepatic epithelia organize their apical membrane domain into bile canaliculi and ducts, which are not accessible to circulating immune cells but that nevertheless confine most of ICAM-1. Here, by analyzing ICAM-1_KO human hepatic cells, liver organoids from ICAM-1_KO mice and rescue-of-function experiments, we show that ICAM-1 regulates epithelial apicobasal polarity in a leukocyte adhesion-independent manner. ICAM-1 signals to an actomyosin network at the base of canalicular microvilli, thereby controlling the dynamics and size of bile canalicular-like structures. We identified the scaffolding protein EBP50/NHERF1/SLC9A3R1, which connects membrane proteins with the underlying actin cytoskeleton, in the proximity interactome of ICAM-1. EBP50 and ICAM-1 form nano-scale domains that overlap in microvilli, from which ICAM-1 regulates EBP50 nano-organization. Indeed, EBP50 expression is required for ICAM-1-mediated control of BC morphogenesis and actomyosin. Our findings indicate that ICAM-1 regulates the dynamics of epithelial apical membrane domains beyond its role as a heterotypic cell-cell adhesion molecule and reveal potential therapeutic strategies for preserving epithelial architecture during inflammatory stress.

Simultaneous Targeting of IL-1-Signaling and IL-6-Trans-Signaling Preserves Human Pulmonary Endothelial Barrier Function During a Cytokine Storm-Brief Report.

BACKGROUND: Systemic inflammatory diseases, such as sepsis and severe COVID-19, provoke acute respiratory distress syndrome in which the pathological hyperpermeability of the microvasculature, induced by uncontrolled inflammatory stimulation, causes pulmonary edema. Identifying the inflammatory mediators that induce human lung microvascular endothelial cell barrier dysfunction is essential to find the best anti-inflammatory treatments for critically ill acute respiratory distress syndrome patients. METHODS: We have compared the responses of primary human lung microvascular endothelial cells to the main inflammatory mediators involved in cytokine storms induced by sepsis and SARS-CoV2 pulmonary infection and to sera from healthy donors and severely ill patients with sepsis. Endothelial barrier function was measured by electric cell-substrate impedance sensing, quantitative confocal microscopy, and Western blot. RESULTS: The human lung microvascular endothelial cell barrier was completely disrupted by IL (interleukin)-6 conjugated with soluble IL-6R (IL-6 receptor) and by IL-1ß (interleukin-1beta), moderately affected by TNF (tumor necrosis factor)-α and IFN (interferon)-γ and unaffected by other cytokines and chemokines, such as IL-6, IL-8, MCP (monocyte chemoattractant protein)-1 and MCP-3. The inhibition of IL-1 and IL-6R simultaneously, but not separately, significantly reduced endothelial hyperpermeability on exposing human lung microvascular endothelial cells to a cytokine storm consisting of 8 inflammatory mediators or to sera from patients with sepsis. Simultaneous inhibition of IL-1 and JAK (Janus kinase)-STAT (signal transducer and activator of transcription protein), a signaling node downstream IL-6 and IFN-γ, also prevented septic serum-induced endothelial barrier disruption. CONCLUSIONS: These findings strongly suggest a major role for both IL-6 trans-signaling and IL-1ß signaling in the pathological increase in permeability of the human lung microvasculature and reveal combinatorial strategies that enable the gradual control of pulmonary endothelial barrier function in response to a cytokine storm.

Plasmolipin regulates basolateral-to-apical transcytosis of ICAM-1 and leukocyte adhesion in polarized hepatic epithelial cells.

Apical localization of Intercellular Adhesion Receptor (ICAM)-1 regulates the adhesion and guidance of leukocytes across polarized epithelial barriers. Here, we investigate the molecular mechanisms that determine ICAM-1 localization into apical membrane domains of polarized hepatic epithelial cells, and their effect on lymphocyte-hepatic epithelial cell interaction. We had previously shown that segregation of ICAM-1 into apical membrane domains, which form bile canaliculi and bile ducts in hepatic epithelial cells, requires basolateral-to-apical transcytosis. Searching for protein machinery potentially involved in ICAM-1 polarization we found that the SNARE-associated protein plasmolipin (PLLP) is expressed in the subapical compartment of hepatic epithelial cells in vitro and in vivo. BioID analysis of ICAM-1 revealed proximal interaction between this adhesion receptor and PLLP. ICAM-1 colocalized and interacted with PLLP during the transcytosis of the receptor. PLLP gene editing and silencing increased the basolateral localization and reduced the apical confinement of ICAM-1 without affecting apicobasal polarity of hepatic epithelial cells, indicating that ICAM-1 transcytosis is specifically impaired in the absence of PLLP. Importantly, PLLP depletion was sufficient to increase T-cell adhesion to hepatic epithelial cells. Such an increase depended on the epithelial cell polarity and ICAM-1 expression, showing that the epithelial transcytotic machinery regulates the adhesion of lymphocytes to polarized epithelial cells. Our findings strongly suggest that the polarized intracellular transport of adhesion receptors constitutes a new regulatory layer of the epithelial inflammatory response.

GC-MS Study of Mono- and Bishaloethylphosphonates Related to Schedule 2.B.04 of the Chemical Weapons Convention: The Discovery of a New Intramolecular Halogen Transfer.

A new class of compounds, mono- and bis-haloethylphosphonates (HAPs and bisHAPs, respectively), listed in Schedule 2.B.04 of the Chemical Weapons Convention (CWC), has been synthesized and studied by GC-MS with two aims. First, to improve the identification of this type of chemicals by the Organization for the Prohibition of Chemical Weapons, (OPCW). Second, to study the synergistic effect of halogen and silicon atoms in molecules undergoing mass spectrometry. Fragmentation patterns of trimethylsilyl derivatives of HAPs were found to depend on the nature of the halogen atom; this was in agreement with DFT-calculations. The data suggest that a novel intramolecular halogen transfer takes place during the fragmentation process. Graphical Abstract ᅟ.