Integration of Functional Imaging, Cytometry, and Unbiased Proteomics Reveals New Features of Endothelial-to-Mesenchymal Transition in Ischemic Mitral Valve Regurgitation in Human Patients

Background: Following myocardial infarction, mitral regurgitation (MR) is a common complication. Previous animal studies demonstrated the association of endothelial-to-mesenchymal transition (EndMT) with mitral valve (MV) remodeling. Nevertheless, little is known about how MV tissue responds to ischemic heart changes in humans. Methods: MVs were obtained by the Cardiothoracic Surgical Trials Network from 17 patients with ischemic mitral regurgitation (IMR). Echo-doppler imaging assessed MV function at time of resection. Cryosections of MVs were analyzed using a multi-faceted histology and immunofluorescence examination of cell populations. MVs were further analyzed using unbiased label-free proteomics. Echo-Doppler imaging, histo-cytometry measures and proteomic analysis were then integrated. Results: MVs from patients with greater MR exhibited proteomic changes associated with proteolysis-, inflammatory- and oxidative stress-related processes compared to MVs with less MR. Cryosections of MVs from patients with IMR displayed activated valvular interstitial cells (aVICs) and double positive CD31+ αSMA+ cells, a hallmark of EndMT. Univariable and multivariable association with echocardiography measures revealed a positive correlation of MR severity with both cellular and geometric changes (e.g., aVICs, EndMT, leaflet thickness, leaflet tenting). Finally, proteomic changes associated with EndMT showed gene-ontology enrichment in vesicle-, inflammatory- and oxidative stress-related processes. This discovery approach indicated new candidate proteins associated with EndMT regulation in IMR. Conclusion: We describe an atypical cellular composition and distinctive proteome of human MVs from patients with IMR, which highlighted new candidate proteins implicated in EndMT-related processes, associated with maladaptive MV fibrotic remodeling. Graphical Abstract

[1]  L. Allen,et al.  2022 ACC Expert Consensus Decision Pathway for Integrating Atherosclerotic Cardiovascular Disease and Multimorbidity Treatment: A Framework for Pragmatic, Patient-Centered Care: A Report of the American College of Cardiology Solution Set Oversight Committee. , 2022, Journal of the American College of Cardiology.

[2]  J. Michel,et al.  Red Blood Cells and Hemoglobin in Human Atherosclerosis and Related Arterial Diseases , 2020, International journal of molecular sciences.

[3]  J. Bavaria,et al.  2020 Focused Update of the 2017 ACC Expert Consensus Decision Pathway on the Management of Mitral Regurgitation. , 2020, Journal of the American College of Cardiology.

[4]  S. Kletukhina,et al.  Role of Mesenchymal Stem Cell-Derived Extracellular Vesicles in Epithelial–Mesenchymal Transition , 2019, International journal of molecular sciences.

[5]  Yan Wu,et al.  Downregulation of IQGAP1 inhibits epithelial‐mesenchymal transition via the HIF1α/VEGF‐A signaling pathway in gastric cancer , 2019, Journal of cellular biochemistry.

[6]  O. J. Semmes,et al.  Endothelial-to-Mesenchymal Transition in Human Adipose Tissue Vasculature Alters the Particulate Secretome and Induces Endothelial Dysfunction. , 2019, Arteriosclerosis, thrombosis, and vascular biology.

[7]  L. Fusini,et al.  Endothelial Dysfunction in Patients with Severe Mitral Regurgitation , 2019, Journal of clinical medicine.

[8]  J. Vilo,et al.  g:Profiler: a web server for functional enrichment analysis and conversions of gene lists (2019 update) , 2019, Nucleic Acids Res..

[9]  J. Michel,et al.  Relationship of Iron Deposition to Calcium Deposition in Human Aortic Valve Leaflets. , 2019, Journal of the American College of Cardiology.

[10]  H. Ebhardt,et al.  Platelet-Derived Microparticles From Obese Individuals: Characterization of Number, Size, Proteomics, and Crosstalk With Cancer and Endothelial Cells , 2019, Front. Pharmacol..

[11]  J. Kovacic,et al.  Endothelial to Mesenchymal Transition in Cardiovascular Disease: JACC State-of-the-Art Review. , 2019, Journal of the American College of Cardiology.

[12]  E. Puré,et al.  CD44-dependent inflammation, fibrogenesis, and collagenolysis regulates extracellular matrix remodeling and tensile strength during cutaneous wound healing. , 2019, Matrix biology : journal of the International Society for Matrix Biology.

[13]  C. Cicchini,et al.  Exosome-Mediated Signaling in Epithelial to Mesenchymal Transition and Tumor Progression , 2018, Journal of clinical medicine.

[14]  Martin Eisenacher,et al.  The PRIDE database and related tools and resources in 2019: improving support for quantification data , 2018, Nucleic Acids Res..

[15]  P. Gao,et al.  Suppression of Endothelial-to-Mesenchymal Transition by SIRT (Sirtuin) 3 Alleviated the Development of Hypertensive Renal Injury , 2018, Hypertension.

[16]  L. Ke,et al.  The CXCL5/CXCR2 axis contributes to the epithelial-mesenchymal transition of nasopharyngeal carcinoma cells by activating ERK/GSK-3β/snail signalling , 2018, Journal of Experimental & Clinical Cancer Research.

[17]  Keyuan Zhou,et al.  Increased expression of monoamine oxidase A is associated with epithelial to mesenchymal transition and clinicopathological features in non-small cell lung cancer. , 2017, Oncology letters.

[18]  A. Kobori,et al.  Renin-angiotensin system inhibitors in patients with or without ischaemic mitral regurgitation after acute myocardial infarction , 2017, Open Heart.

[19]  R. Levine,et al.  Mitral Leaflet Changes Following Myocardial Infarction: Clinical Evidence for Maladaptive Valvular Remodeling , 2017, Circulation. Cardiovascular imaging.

[20]  R. Levine,et al.  Effect of Losartan on Mitral Valve Changes After Myocardial Infarction. , 2017, Journal of the American College of Cardiology.

[21]  J. Michel,et al.  Erythrocyte Efferocytosis by the Arterial Wall Promotes Oxidation in Early-Stage Atheroma in Humans , 2017, Front. Cardiovasc. Med..

[22]  Yuchi Han,et al.  Recommendations for Noninvasive Evaluation of Native Valvular Regurgitation: A Report from the American Society of Echocardiography Developed in Collaboration with the Society for Cardiovascular Magnetic Resonance. , 2017, Journal of the American Society of Echocardiography : official publication of the American Society of Echocardiography.

[23]  V. Afshar-Kharghan.  The role of the complement system in cancer , 2017, The Journal of clinical investigation.

[24]  Jason C. Kovacic,et al.  Corrigendum: Endothelial to mesenchymal transition is common in atherosclerotic lesions and is associated with plaque instability , 2017, Nature Communications.

[25]  Liang Chang,et al.  CXCL7 promotes proliferation and invasion of cholangiocarcinoma cells. , 2017, Oncology reports.

[26]  R. Levine,et al.  CD45 Expression in Mitral Valve Endothelial Cells After Myocardial Infarction. , 2016, Circulation research.

[27]  V. Fuster,et al.  Endothelial to mesenchymal transition is common in atherosclerotic lesions and is associated with plaque instability , 2016, Nature Communications.

[28]  M. Frid,et al.  Endothelial-to-Mesenchymal Transition: An Evolving Paradigm and a Promising Therapeutic Target in PAH. , 2016, Circulation.

[29]  R. Levine,et al.  Myocardial Infarction Alters Adaptation of the Tethered Mitral Valve. , 2016, Journal of the American College of Cardiology.

[30]  Francesca N. Delling,et al.  Mitral valve disease—morphology and mechanisms , 2015, Nature Reviews Cardiology.

[31]  Michael S Sacks,et al.  On the presence of affine fibril and fiber kinematics in the mitral valve anterior leaflet. , 2015, Biophysical journal.

[32]  J. Mayer,et al.  Reciprocal interactions between mitral valve endothelial and interstitial cells reduce endothelial-to-mesenchymal transition and myofibroblastic activation , 2015, Journal of molecular and cellular cardiology.

[33]  J. Hung,et al.  Echocardiographic assessment of ischemic mitral regurgitation , 2014, Cardiovascular Ultrasound.

[34]  Z. Fu,et al.  Overexpression of peroxiredoxin 2 inhibits TGF-β1-induced epithelial-mesenchymal transition and cell migration in colorectal cancer. , 2014, Molecular medicine reports.

[35]  M. Hance,et al.  Secreted Hsp90 Is a Novel Regulator of the Epithelial to Mesenchymal Transition (EMT) in Prostate Cancer* , 2012, The Journal of Biological Chemistry.

[36]  Y. Takayama,et al.  LRP1 Regulates Architecture of the Vascular Wall by Controlling PDGFRβ-Dependent Phosphatidylinositol 3-Kinase Activation , 2009, PloS one.

[37]  Suzanne Sullivan,et al.  Active Adaptation of the Tethered Mitral Valve: Insights Into a Compensatory Mechanism for Functional Mitral Regurgitation , 2009, Circulation.

[38]  F. Torti,et al.  Ferritin for the clinician. , 2009, Blood reviews.

[39]  Robert A Levine,et al.  Mitral Leaflet Adaptation to Ventricular Remodeling: Occurrence and Adequacy in Patients With Functional Mitral Regurgitation , 2008, Circulation.

[40]  Kurt R Stenmark,et al.  Perspectives on endothelial-to-mesenchymal transition: potential contribution to vascular remodeling in chronic pulmonary hypertension. , 2007, American journal of physiology. Lung cellular and molecular physiology.

[41]  Steven P Gygi,et al.  Target-decoy search strategy for increased confidence in large-scale protein identifications by mass spectrometry , 2007, Nature Methods.

[42]  C. Kielty,et al.  Fibrillin-1 regulates the bioavailability of TGFβ1 , 2007, The Journal of cell biology.

[43]  R. Gomer,et al.  Bone marrow-derived fibroblast precursors mediate ischemic cardiomyopathy in mice , 2006, Proceedings of the National Academy of Sciences.

[44]  K. Wilson,et al.  Nuclear Titin interacts with A- and B-type lamins in vitro and in vivo , 2006, Journal of Cell Science.

[45]  K. Kaibuchi,et al.  IQGAP1: a key regulator of adhesion and migration , 2005, Journal of Cell Science.

[46]  M. Enriquez-Sarano,et al.  Heart Failure and Death After Myocardial Infarction in the Community: The Emerging Role of Mitral Regurgitation , 2005, Circulation.

[47]  D. Judge,et al.  TGF-β–dependent pathogenesis of mitral valve prolapse in a mouse model of Marfan syndrome , 2004 .

[48]  S. Dimmeler,et al.  Endothelial Progenitor Cells: Characterization and Role in Vascular Biology , 2004, Circulation research.

[49]  Wen-Fang Tseng,et al.  LRP‐1/TβR‐V mediates TGF‐β1‐induced growth inhibition in CHO cells , 2004 .

[50]  Renu Virmani,et al.  Intraplaque hemorrhage and progression of coronary atheroma. , 2003, The New England journal of medicine.

[51]  F. Tang,et al.  Cellular growth inhibition by IGFBP‐3 and TGF‐β1 requires LRP‐1 , 2003 .

[52]  R. Discipio,et al.  Complement C3a and C5a Induce Different Signal Transduction Cascades in Endothelial Cells1 , 2002, The Journal of Immunology.

[53]  P. Libby,et al.  Activated Interstitial Myofibroblasts Express Catabolic Enzymes and Mediate Matrix Remodeling in Myxomatous Heart Valves , 2001, Circulation.

[54]  K. Bailey,et al.  Ischemic Mitral Regurgitation: Long-Term Outcome and Prognostic Implications With Quantitative Doppler Assessment , 2001, Circulation.

[55]  E. Schwammenthal,et al.  Mechanism of ischemic mitral regurgitation with segmental left ventricular dysfunction: three-dimensional echocardiographic studies in models of acute and chronic progressive regurgitation. , 2001, Journal of the American College of Cardiology.

[56]  F. Yin,et al.  Biaxial mechanical behavior of excised porcine mitral valve leaflets. , 1995, The American journal of physiology.

[57]  K. Eilertsen,et al.  Cellular titin localization in stress fibers and interaction with myosin II filaments in vitro , 1994, The Journal of cell biology.

[58]  R. P. Cochran,et al.  Stress/Strain Characteristics of Porcine Mitral Valve Tissue: Parallel Versus Perpendicular Collagen Orientation , 1992, Journal of cardiac surgery.

[59]  William Stafford Noble,et al.  Assigning significance to peptides identified by tandem mass spectrometry using decoy databases. , 2008, Journal of proteome research.

[60]  Wen-Fang Tseng,et al.  LRP-1/TbetaR-V mediates TGF-beta1-induced growth inhibition in CHO cells. , 2004, FEBS letters.