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Endothelial to Mesenchymal Transition (EndoMT) leads to a loss of normal endothelial cell function and may contribute to the development of pulmonary arterial hypertension

Good, R, Gilbane, A, Trinder, S, Denton, C, Coghlan, G, Abraham, D and Holmes, A (2014) Endothelial to Mesenchymal Transition (EndoMT) leads to a loss of normal endothelial cell function and may contribute to the development of pulmonary arterial hypertension In: ASCB/IFCB Meeting, 2014-12-06 - 2014-12-10, Philadelphia, PA.

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Abstract

Background/Purpose Vascular complications in Scleroderma (SSc) patients are associated with high mortality, particularly in patients who develop pulmonary arterial hypertension (SSc-PAH). Vascular complications, thought to arise from initial activation and dysfunction of the endothelium can lead to: elevated vascular leak, inflammation, mesenchymal hypertrophy by activation of resident smooth muscle cells and fibroblasts, and neointima formation. Recent studies suggest that as well as resident mesenchymal cells, endothelial cells can undergo endothelial-mesenchymal transition (EndoMT), and acquire a mesenchymal phenotype which may contribute to the expansion of the mesenchymal cell population. Here we sought to determine the prevalence of EndoMT in SSc-PAH patients and pre-clinical models of PAH, and assess the cellular effects on pulmonary artery endothelial cells (PAECs) functions. Methods Using lung tissue from SSc-PAH patients (n=3), healthy control (HC) donors (n=3), and from the hypoxia/SU5416 pre-clinical murine model of PAH (n=5), EndoMT was determined by immunofluorescence based on co-expression of vWF and αSMA. EndoMT was induced in human PAECs (n=3) in vitro by TNFα [5ng/ml], IL-1β [0.1ng/m;] and TGFβ [5ng/ml] in combination. Morphological changes were assessed by light microscopy and phalloidin staining. Western blotting and immunofluorescence was used to quantify: CD31, vWF, occludin, VE-cadherin, αSMA, calponin and collagen type 1 expression. Conditioned media was collected from PAECs, PAECs following treatment to initiate EndoMT and SSc-PAH and HC fibroblasts; levels of inflammatory secretion was quantified by MSD arrays. The capacity of homogenous EndoMT monolayers (n=6) and mixed cultures of 1:10 EndoMT:PAECs (n=6) cells to form exclusion barriers was assessed using trans-well permeability FITC-albumin assays. Results Co-localisation of vWF and αSMA was observed in ≤5% of pulmonary arteries from SSc-PAH patients and hypoxia/SU5416 mice. PAECs treated with TNFα, IL-1β and TGFβ exhibited significant changes in morphology, loss of endothelial markers and elevated expression of mesenchymal markers by day 6. There was a significant (P<0.05) increase in secretion of pro-inflammatory chemokines by EndoMT cells compared to PAECs including IL-6 [474±95 vs.12±6.6 pg/ml] and IL-8 [620±71 vs.28±6.5 pg/ml]. EndoMT cells alone or in mixed 1:10 ratio cultures with PAECs, exhibited a significant (P>0.01) 5-fold increase in permeability compared to PAECs alone. Consistent with this, EndoMT cells co-cultured with PAECs in a ratio of 1:10 led to 2.5-fold significant (P>0.05) increase in permeability. Conclusion The co-localisation of vWF and αSMA present in the pulmonary arteries of SSc-PAH patients and pre-clinical models of PAH, is indicative of EndoMT. We demonstrate EndoMT leads to a loss of normal PAEC morphology and an enhanced secretion of pro-inflammatory chemokines. Furthermore EndoMT cells failed to form integral biological barriers and contributed to enhanced permeability of PAEC barriers. Collectively our data suggests that EndoMT may contribute to the loss of normal endothelium function and the development of SSc-PAH.

Item Type: Conference or Workshop Item (Conference Abstract)
Authors :
NameEmailORCID
Good, RUNSPECIFIEDUNSPECIFIED
Gilbane, AUNSPECIFIEDUNSPECIFIED
Trinder, Ss.trinder@surrey.ac.ukUNSPECIFIED
Denton, CUNSPECIFIEDUNSPECIFIED
Coghlan, GUNSPECIFIEDUNSPECIFIED
Abraham, DUNSPECIFIEDUNSPECIFIED
Holmes, AUNSPECIFIEDUNSPECIFIED
Date : December 2014
Contributors :
ContributionNameEmailORCID
publisherAmerican Society of Cell Biology, UNSPECIFIEDUNSPECIFIED
Related URLs :
Depositing User : Symplectic Elements
Date Deposited : 17 May 2017 10:45
Last Modified : 17 May 2017 14:52
URI: http://epubs.surrey.ac.uk/id/eprint/829057

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