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Front. Physiol. | doi: 10.3389/fphys.2018.01393

Epsin15 homology domains: role in the pathogenesis of pulmonary arterial hypertension

 Dan N. Predescu1, Shanshan Qin2,  Monal Patel3, Cristina Bardita4, Rabia Bhalli1 and  SANDA A. PREDESCU1*
  • 1Internal Medicine, Rush University, United States
  • 2Rush University Medical Center, United States
  • 3Feinberg School of Medicine, Northwestern University, United States
  • 4Department of Critical Care Transport, School of Medicine, University of Alabama at Birmingham, United States

Intersectin-1s (ITSN) deficiency and expression of a biologically active ITSN fragment, result of granzyme B cleavage under inflammatory conditions associated with pulmonary arterial hypertension (PAH), are characteristics of lung tissue of human and animal models of PAH. Recently, we have shown that this ITSN fragment comprising two Epsin15 homology domains (EHITSN) triggers endothelial cell (EC) proliferation and the plexiform arteriopathy in PAH. Limited evidence also indicates that the EH domains of endocytic proteins such as ITSN, upregulate compensatory endocytic pathways in cells with impaired vesicular trafficking. Thus, we sought to investigate whether the EHITSN may be involved in this compensatory mechanism for improving the EC endocytic dysfunction induced by ITSN deficiency and possibly contribute to PAH pathogenesis.
We used stably-transfected human pulmonary artery ECs expressing the Myc-EHITSN (ECEH-ITSN) and ITSN knockout heterozygous mice (K0ITSN+/-) transduced with the Myc-EHITSN, in conjunction with functional assays: the biotin assay for caveolae internalization and 8 nm gold (Au)- and dinitrophenylated (DNP)-albumin perfusion of murine lung microvasculature. Pulmonary artery ECs of PAH patients (ECPAH), ITSN knockdown ECs (ECKD-ITSN), the monocrotaline (MCT)-induced mouse and rat models of PAH as well as untreated animals served as controls. ELISA via streptavidin-HRP or anti-DNP antibody (Ab), applied on ECs and lung lysates indicated greater than 30% increase in biotin internalization in ECEH-ITSN compared to ECCtrl. Despite their endocytic deficiency, ECPAH internalized biotin similar to ECCtrl which is 2-fold higher compared to ECKD-ITSN. Moreover, the lung microvascular bed of Myc-EHITSN-transduced mice and MCT-treated animals showed greater than 2-fold increase in DNP-BSA transendothelial transport, all compared to untreated controls. Electron microscopy (EM) revealed increased occurrence of non-conventional endocytic/transcytotic structures (i.e., caveolae clusters, tubulo-vesicular and enlarged endocytic structures, membranous rings), usually underrepresented. Most of these structures were labeled by Au-BSA, consistent with their involvement in the transendothelial transport. Furthermore, ITSN deficiency and EHITSN expression alter the subcellular localization of the EH-binding protein 1 (EHBP1) and cortical actin organization, altogether supporting the increase occurrence/trafficking of the alternative endocytic structures.
Thus, the EHITSN contributes to the endocytic activity of dysfunctional ECPAH, their proliferation and overgrowth, and hence, the development and progression of the disease.

Keywords: intersectin, pulmonary arterial hypertension, Endocytic deficiency, alternative transport pathways, non-conventional endocytic/transcytotic structures, EH-binding protein 1, Cortical actin

Received: 21 Jun 2018; Accepted: 13 Sep 2018.

Edited by:

Qiaobing Huang, Southern Medical University, China

Reviewed by:

Donna L. Cioffi, University of South Alabama, United States
Jean-Francois Quignard, Université de Bordeaux, France  

Copyright: © 2018 Predescu, Qin, Patel, Bardita, Bhalli and PREDESCU. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

* Correspondence: PhD. SANDA A. PREDESCU, Rush University, Internal Medicine, Chicago, 60612, Illinois, United States,