1. Endoglin Is an endothelial housekeeper against inflammation: insight in ECFC-related permeability through LIMK/cofilin pathway
- Author
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Elisa, Rossi, Alexandre, Kauskot, François, Saller, Elisa, Frezza, Sonia, Poirault-Chassac, Anna, Lokajczyk, Pierre, Bourdoncle, Bruno, Saubaméa, Pascale, Gaussem, Miguel, Pericacho, Regis, Bobe, Christilla, Bachelot-Loza, Samuela, Pasquali, Carmelo, Bernabeu, David M, Smadja, Promex Stiftung Fur Die Forschung, Université de Paris, Institut National de la Santé et de la Recherche Médicale (France), Rossi, Elisa, Kauskot. Alexander, Saller, François, Frezza, Elisa, Saubaméa, Bruno, Gaussem, Pascale, Pericacho, Miguel, Bobe, Regis, Bachelot-Loza, Christilla, Pasquali, Samuela, Bernabéu, Carmelo, Smadja, David M., Innovations thérapeutiques en hémostase (IThEM - U1140), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Paris (UP), Hémostase, Inflammation, Thrombose (HITH - U1176 Inserm - CHU Bicêtre), Institut National de la Santé et de la Recherche Médicale (INSERM)-AP-HP Hôpital Bicêtre (Le Kremlin-Bicêtre)-Université Paris-Saclay, Cibles Thérapeutiques et conception de médicaments (CiTCoM - UMR 8038), Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Université de Paris (UP), Institut Cochin (IC UM3 (UMR 8104 / U1016)), Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Université de Paris (UP), Optimisation thérapeutique en Neuropsychopharmacologie (OPTeN (UMR_S_1144 / U1144)), Universidad de Salamanca, Centro de Investigaciones Biológicas Margarita Salas, Consejo Superior de Investigaciones Científicas [Madrid] (CSIC), Centro de Investigación Biomédica en Red de Enfermedades Raras, CIBER de Enfermedades Raras (CIBERER), Rossi, Elisa [0000-0002-0570-6104], Kauskot. Alexander [0000-0002-4064-8114], Saller, François [0000-0001-5255-2630], Frezza, Elisa [0000-0003-0122-7859], Saubaméa, Bruno [0000-0002-6218-0460], Gaussem, Pascale [0000-0002-9139-2147], Pericacho, Miguel [0000-0003-0226-482X], Bobe, Regis [0000-0002-8225-9117], Bachelot-Loza, Christilla [0000-0002-6264-902X ], Pasquali, Samuela [0000-0003-1487-0894], Bernabéu, Carmelo [0000-0002-1563-6162], and Smadja, David M. [0000-0001-7731-9202]
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Inflammation ,Cell Membrane Permeability ,Neovascularization, Pathologic ,QH301-705.5 ,[SDV]Life Sciences [q-bio] ,Endoglin ,Endothelial Cells ,Lim Kinases ,macromolecular substances ,Article ,Mice ,Chemistry ,Actin Depolymerizing Factors ,Cofilin ,HHT1 ,ECFC ,TNFα ,Animals ,Biology (General) ,QD1-999 - Abstract
Endoglin (Eng) is an endothelial cell (EC) transmembrane glycoprotein involved in adhesion and angiogenesis. Eng mutations result in vessel abnormalities as observed in hereditary hemorrhagic telangiectasia of type 1. The role of Eng was investigated in endothelial functions and permeability under inflammatory conditions, focusing on the actin dynamic signaling pathway. Endothelial Colony-Forming Cells (ECFC) from human cord blood and mouse lung/aortic EC (MLEC, MAEC) from Eng+/+ and Eng+/− mice were used. ECFC silenced for Eng with Eng-siRNA and ctr-siRNA were used to test tubulogenesis and permeability +/− TNFα and +/− LIM kinase inhibitors (LIMKi). In silico modeling of TNFα–Eng interactions was carried out from PDB IDs 5HZW and 5HZV. Calcium ions (Ca2+) flux was studied by Oregon Green 488 in epifluorescence microscopy. Levels of cofilin phosphorylation and tubulin post-translational modifications were evaluated by Western blot. F-actin and actin–tubulin distribution/co-localization were evaluated in cells by confocal microscopy. Eng silencing in ECFCs resulted in a decrease of cell sprouting by 50 ± 15% (p <, 0.05) and an increase in pseudo-tube width (41 ± 4.5%, p <, 0.001) compared to control. Upon TNFα stimulation, ECFC Eng–siRNA displayed a significant higher permeability compared to ctr-siRNA (p <, 0.01), which is associated to a higher Ca2+ mobilization (p <, 0.01). Computational analysis suggested that Eng mitigated TNFα activity. F-actin polymerization was significantly increased in ECFC Eng-siRNA, MAEC+/−, and MLEC+/− compared to controls (p <, 0.001, p <, 0.01, and p <, 0.01, respectively) as well as actin/tubulin distribution (p <, 0.01). Furthermore, the inactive form of cofilin (P-cofilin at Ser3) was significantly decreased by 36.7 ± 4.8% in ECFC Eng-siRNA compared to ctr-siRNA (p <, 0.001). Interestingly, LIMKi reproduced the absence of Eng on TNFα-induced ECFC-increased permeability. Our data suggest that Eng plays a critical role in the homeostasis regulation of endothelial cells under inflammatory conditions (TNFα), and loss of Eng influences ECFC-related permeability through the LIMK/cofilin/actin rearrangement-signaling pathway.
- Published
- 2021
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