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Disturbed flow increases endothelial inflammation and permeability via a Frizzled-4-β-catenin-dependent pathway.

ABSTRACT: Multidirectional or disturbed flow promotes endothelial dysfunction and is associated with early atherogenesis. Here we investigated the role of Wnt signalling in flow-mediated endothelial dysfunction. The expression of Frizzled-4 was higher in cultured human aortic endothelial cells (ECs) exposed to disturbed flow compared to that seen for undisturbed flow, obtained using an orbital shaker. Increased expression was also detected in regions of the porcine aortic arch exposed to disturbed flow. The increased Frizzled-4 expression in cultured ECs was abrogated following knockdown of R-spondin-3. Disturbed flow also increased the nuclear localisation and activation of β-catenin, an effect that was dependent on Frizzled-4 and R-spondin-3. Inhibition of β-catenin using the small-molecule inhibitor iCRT5 or knockdown of Frizzled-4 or R-spondin-3 resulted in reduced expression of pro-inflammatory genes in ECs exposed to disturbed flow, as did inhibition of WNT5A signalling. Inhibition of the canonical Wnt pathway had no effect. Inhibition of β-catenin also reduced endothelial paracellular permeability; this was associated with altered junctional and focal adhesion organisation and cytoskeletal remodelling. These data suggest the presence of an atypical Frizzled-4-β-catenin pathway that promotes endothelial dysfunction in response to disturbed flow.

SUBMITTER: Rickman M

PROVIDER: S-EPMC10112981 | biostudies-literature | 2023 Mar

REPOSITORIES: biostudies-literature

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Disturbed flow increases endothelial inflammation and permeability via a Frizzled-4-β-catenin-dependent pathway.

Rickman Matthew M Ghim Mean M Pang Kuin K von Huelsen Rocha Ana Cristina AC Drudi Elena M EM Sureda-Vives Macià M Ayoub Nicolas N Tajadura-Ortega Virginia V George Sarah J SJ Weinberg Peter D PD Warboys Christina M CM

Journal of cell science 20230324 6

Multidirectional or disturbed flow promotes endothelial dysfunction and is associated with early atherogenesis. Here we investigated the role of Wnt signalling in flow-mediated endothelial dysfunction. The expression of Frizzled-4 was higher in cultured human aortic endothelial cells (ECs) exposed to disturbed flow compared to that seen for undisturbed flow, obtained using an orbital shaker. Increased expression was also detected in regions of the porcine aortic arch exposed to disturbed flow. T ...[more]

PMID: 36846872

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Project description:Pathologically increased vascular permeability is an important dysfunction in the pathomechanism of life-threatening conditions, such as sepsis, ischemia/reperfusion, or hereditary angioedema (HAE), diseases accompanied by uncontrolled activation of the complement system. HAE for example is caused by the deficiency of C1-inhibitor (the main regulator of early complement activation), which leads to edematous attacks threatening with circulatory collapse. We have previously reported that endothelial cells become activated during HAE attacks. A natural target of C1-inhibitor is mannan-binding lectin-associated serine protease-1 (MASP-1), a multifunctional serine protease, which plays a key role in the activation of complement lectin pathway. We have previously shown that MASP-1 induces the pro-inflammatory activation of endothelial cells and in this study we investigated whether MASP-1 can directly affect endothelial permeability. All experiments were performed on human umbilical vein endothelial cells (HUVECs). Real-time micro electric sensing revealed that MASP-1 decreases the impedance of HUVEC monolayers and in a recently developed permeability test (XperT), MASP-1 dose-dependently increased endothelial paracellular transport. We show that protease activated receptor-1 mediated intracellular Ca2+-mobilization, Rho-kinase activation dependent myosin light chain (MLC) phosphorylation, cytoskeletal actin rearrangement, and disruption of interendothelial junctions are underlying this phenomenon. Furthermore, in a whole-transcriptome microarray analysis MASP-1 significantly changed the expression of 25 permeability-related genes in HUVECs-for example it up-regulated bradykinin B2 receptor expression. According to our results, MASP-1 has potent permeability increasing effects. During infections or injuries MASP-1 may help eliminate the microbes and/or tissue debris by enhancing the extravasation of soluble and cellular components of the immune system, however, it may also play a role in the pathomechanism of diseases, where edema formation and complement lectin pathway activation are simultaneously present. Our findings also raise the possibility that MASP-1 may be a promising target of anti-edema drug development.

Project description:Background: Increasing evidence suggests that hemodynamic disturbed flow induces endothelial dysfunction via a complex biological process so-called endothelial to mesenchymal transition (EndoMT). Recently, DNA methyltransferases (DNMTs) was reported as a key molecular mediator to promote EndoMT. Our understanding of how DNMTs, particularly the maintenance DNMTs, DNMT1, coordinate EndoMT is still lacking. Methods: A parallel-plate flow apparatus and perfusion devices were used to apply fluid with endothelial protective pulsatile shear (PS, to mimic the laminar flow) or harmful oscillatory shear (OS, to mimic the disturbed flow) to cultured endothelial cells (ECs). Endothelial lineage tracing mice and conditional EC Dnmt1 knockout mice were subjected to a surgery of carotid partial ligation to generate the flow-accelerated atherogenesis models. Western blotting, quantitative RT-PCR, immunofluorescent staining, methylation-specific PCR, chromatin immunoprecipitation, endothelial functional assays, and assessments for neointimal formation and atherosclerosis were performed. Results: Inhibition of DNMTs with 5-aza-2'-deoxycytidine (5-Aza) suppressed the disturbed flow/OS-induced EndoMT, both in cultured cells and the endothelial lineage tracing mice. 5-Aza also ameliorated the downregulation of aldehyde dehydrogenases (ALDHs) and β-alanine biosynthesis caused by disturbed flow/OS. Knockdown of the ALDH family proteins, ALDH2, ALDH3A1, and ALDH6A1, showed an EndoMT-induction effect as OS. Supplementation of cells with the functional metabolites of β-alanine, carnosine and acetyl-CoA (acetate), reversed EndoMT, likely via inhibiting the phosphorylation of Smad2/3. Endothelial-specific knockout of Dnmt1 protected the vasculature from disturbed flow-induced remodeling and atherosclerosis. Conclusions: Endothelial DNMT1 acts as one of the key epigenetic factors to mediate the hemodynamically regulated EndoMT at least through repressing the expression of ALDH2, ALDH3A1, and ALDH6A1. Supplementation with carnosine and acetate may have a great potential in the prevention and treatment of atherosclerosis.

Dataset Information

Disturbed flow increases endothelial inflammation and permeability via a Frizzled-4-β-catenin-dependent pathway.

Publications

Disturbed flow increases endothelial inflammation and permeability via a Frizzled-4-β-catenin-dependent pathway.

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