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Engineering of a cell-loaded patch as a controlled VEGF-releasing device to treat cardiac ischemia

English title Engineering of a cell-loaded patch as a controlled VEGF-releasing device to treat cardiac ischemia
Applicant Marsano Anna
Number 149274
Funding scheme Project funding (Div. I-III)
Research institution Departement Biomedizin Universität Basel
Institution of higher education University of Basel - BS
Main discipline Cardiovascular Research
Start/End 01.10.2013 - 30.09.2016
Approved amount 275'000.00
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All Disciplines (2)

Discipline
Cardiovascular Research
Molecular Biology

Keywords (7)

tissue engineering; angiogenesis; Controlled VEGF delivery; mesenchymal progenitor cells; perfusion bioreactor; cardiac ischemia; cell-based gene therapy

Lay Summary (German)

Lead
Unkontrollierte Expression von Vascular Endothelial Growth Factor (VEGF) induzieren können abweichende Angiogenese. Wir entwickelten eine Methode, um sicherzustellen, gesteuert VEGF-Freisetzung durch FACS-basierte Reinigung von transduzierten Fettgewebe gewonnene mesenchymale Stromazellen (ASC).Wir wollen zell-basierte Gentherapie mit Tissue Engineering zu kombinieren, um ASC-basierten Patches in der Lage, normale und effiziente Angiogenese in der Umgebung zu fördern generieren.
Lay summary

Titel: Planung einer Zelle geladenen Patch als kontrollierte VEGF-Releasing-Gerät, um kardiale Ischämie zu behandeln

 Hintergrund

Unkontrollierte Expression von VEGF induzieren können abweichende Angiogenese. Wir entwickelten eine Methode, um sicherzustellen, erlitten aber gesteuert VEGF-Freisetzung durch FACS-basierte Reinigung von transduzierten Fettgewebe gewonnene mesenchymale Stromazellen (ASC). In dieser Studie wollen wir zell-basierte Gentherapie mit Tissue Engineering zu kombinieren, um ASC-basierten Patches in der Lage, normale und effiziente Angiogenese in der Umgebung zu fördern generieren.

 Spezifische Ziele

1. Besorgen Sie sich die proof of principle, dass VEGF-Expression ASC zuverlässig induzieren normalen Angiogenese im und um das engineered Patch.
2. Untersuchung der Rolle von frischem Stromazellen vaskulären Fraktion (SVF) abgeleitete Zellen als Quelle von endothelialen Vorläuferzellen und Perizyten Ursprungs-Zellen, in der Patch-Vaskularisierung Dynamik und in der Stabilisierung der Blutgefäßnetze wenn mit ASC co-kultiviert werden, die entweder gesteuert oder heterogenen VEGF.

Methoden

ASC wurden mit retroviralen Vektoren, VEGF164 verbunden mit einem FACS-quantifizierbaren Zelloberflächenmarker und einer Bevölkerung homogen Expression eines VEGF-Ebene definiert (VEGF) transduziert war FACS-gereinigt. Patches wurden durch Kultivieren von Zellen auf Kollagen Biomaterial in einer Perfusions-basierten Bioreaktor erzeugt. Die extrinsische angiogene Potential sowohl subkutane und ischämischen Myokard Nacktratte Modelle beurteilt werden.

 Mögliche Bedeutung der Ergebnisse

Das vorgeschlagene Projekt wird erwartet, um (1) eine grundlegende proof-of-principle einen Zell-basierten angiogenen Faktor Release-System zur Förderung der Durchblutung zu verwenden, (2) Erkenntnisse der Angiogenese in vivo Dynamik und wird die Rolle des SVF in der Aufklärung Vaskularisierung Prozess.

Direct link to Lay Summary Last update: 30.09.2013

Lay Summary (Italian)

Lead
L’espressione incontrollata di Vascular Endothelial Growth Factor (VEGF) può indurre angiogenesi aberrante. Abbiamo sviluppato un metodo per garantire un sostenuto ma controllato rilascio di VEGF grazie alla purificazione di cellule stromali mesenchimali derivate da tessuto adiposo (ASC) trasdotte.In questo studio ci proponiamo di combinare la terapia genica basata su cellule con l'ingegneria dei tessuti per generare patch cellulari in grado di promuovere la normale ed efficiente angiogenesi.
Lay summary

Titolo: Ingegnerizzazione di una patch a base di cellule come dispositivo per il rilascio controllato di VEGF per trattare l'ischemia cardiaca

 

Background
L’espressione incontrollata di VEGF può indurre l'angiogenesi aberrante. Abbiamo sviluppato un metodo per garantire un sostenuto ma controllato rilascio di VEGF grazie alla purificazione di cellule stromali mesenchimali derivate da tessuto adiposo (ASC) trasdotte. In questo studio ci proponiamo di combinare la terapia genica basata su cellule con l'ingegneria dei tessuti per generare patch cellulari in grado di promuovere la normale ed efficiente angiogenesi nella zona circostante.

Obiettivi

1. Ottenere la prova di principio che ASC esprimenti VEGF riescano ad indurre angiogenesi normale all'interno e intorno la patch ingegnerizzato.

2. Indagare il ruolo delle cellule derivate dalla frazione stromale vascolare del tessuto adiposo (SVF), come fonte di cellule progenitrici endoteliali e cellule di origine pericitica, nelle dinamiche di vascolarizzazione della patch e nella stabilizzazione delle reti vascolari in caso di co-coltura con ASC esprimenti livelli di VEGF o controllati o eterogenei

 .
Metodi
ASC sono trasdotte con vettori retrovirali esprimenti VEGF164 collegato a un marcatore di membrana e una popolazione omogenea che esprime un livello definito VEGF (VEGF) e’ stata purificata tramite FACS. Le patches vengono coltivate su scaffolds di collagene in un bioreattore a perfusione. Il loro potenziale angiogenico estrinseco sarà valutata in modelli sia di ischemia cardiaca sia sottocutanei in ratti nudi.

Importanza potenziale dei risultati

Il progetto proposto si prevede di fornire (1) la prova-di-principio per utilizzare un sistema cellulare di rilascio di un fattore angiogenico per promuovere la vascolarizzazione, (2) approfondimenti delle dinamiche angiogeniche in vivo e sul ruolo delle SVF nel processo di vascolarizzazione.

Direct link to Lay Summary Last update: 30.09.2013

Lay Summary (English)

Lead
Uncontrolled expression of Vascular Endothelial Growth Factor (VEGF) can induce aberrant angiogenesis. We developed a method to ensure sustained but controlled VEGF release by FACS-based purification of transduced adipose tissue-derived mesenchymal stromal cells (ASC).In this study we aim to combine cell-based gene therapy with tissue engineering to generate ASC-based patches capable to promote normal and efficient angiogenesis in the surrounding area..
Lay summary

Title: Engineering of a cell-loaded patch as a controlled VEGF-releasing device to treat cardiac ischemia

Background

Uncontrolled expression of VEGF can induce aberrant angiogenesis. We developed a method to ensure sustained but controlled VEGF release by FACS-based purification of transduced adipose tissue-derived mesenchymal stromal cells (ASC), which could already induce effective vascularization when injected in ischemic myocardium despite the poor cell survival.

In this study we aim to combine cell-based gene therapy with tissue engineering to generate ASC-based patches capable to promote normal and efficient angiogenesis in the surrounding area.

Specific Aims

1. Obtain the proof of principle that VEGF-expressing ASC can reliably induce normal angiogenesis inside and around the engineered patch.        

2. Investigate the role of fresh stromal vascular fraction (SVF) derived cells, as source of endothelial progenitors and pericytic origin cells, in the patch vascularization dynamics and in the stabilization of the vascular networks when co-cultured with ASC expressing either controlled or heterogeneous VEGF levels.

Methods

ASC were transduced with retroviral vectors expressing VEGF164 linked to a FACS-quantifiable cell surface marker (truncated CD8) and a population homogeneously expressing a defined VEGF level (VEGF) was FACS-purified. Patches were generated by culturing cells for 5 days on collagen scaffolds in a perfusion-based bioreactor. Their extrinsic angiogenic potential will be assessed in both subcutaneous and ischemic myocardial nude rat models.

 

Potential significance of the results

The proposed project is expected to provide (1) a basic proof-of-principle to use a cell-based angiogenic factor release system for promoting vascularization, (2) insights of the angiogenic dynamics in vivo and will elucidate the role of SVF in the vascularization process.

Direct link to Lay Summary Last update: 30.09.2013

Responsible applicant and co-applicants

Employees

Publications

Publication
Engineered mesenchymal cell-based patches as controlled VEGF delivery systems to induce extrinsic angiogenesis
Boccardo Stefano, Gaudiello Emanuele, Melly Ludovic, Cerino Giulia, Ricci Davide, Martin Ivan, Eckstein Friedrich, Banfi Andrea, Marsano Anna, Engineered mesenchymal cell-based patches as controlled VEGF delivery systems to induce extrinsic angiogenesis, in Acta Biomaterialia, 42, 127-135.

Collaboration

Group / person Country
Types of collaboration
Friedrich Eckstein, Professor and Chairman Clinic of Cardiac Surgery, University Hospital Basel Switzerland (Europe)
- Publication
- Research Infrastructure
Ivan Martin, Group Leader of the Tissue Engineering Group, ICFS, Uni Hospital Basel Switzerland (Europe)
- in-depth/constructive exchanges on approaches, methods or results
- Publication
- Research Infrastructure
Andrea Banfi,Group Leader of the Cell and Gene Therapy Group, ICFS, Uni Hospital Basel Switzerland (Europe)
- in-depth/constructive exchanges on approaches, methods or results
- Publication
- Research Infrastructure
Ludovic Melly, Clinic for Cardiac Surgery, University Hospital Basel Switzerland (Europe)
- in-depth/constructive exchanges on approaches, methods or results
- Publication

Scientific events

Active participation

Title Type of contribution Title of article or contribution Date Place Persons involved
TERMIS-EU Talk given at a conference ‘Boosting in vivo angiogenesis by in vitro priming of human stromal vascular fraction cell-based engineered tissues’ 28.06.2016 Uppsala, Sweden Gaudiello Emanuele; Cerino Abdin Giulia; Marsano Anna;
TERMIS-EU Poster Engineering cell-based patches as controlled VEGF-releasing devices for induction of therapeutic angiogenesis 28.06.2016 Uppsala, Sweden Cerino Abdin Giulia; Marsano Anna; Gaudiello Emanuele;
8th Ascona International Workshop on Cardiomyocyte Biology Poster Engineering cell-based patch as a controlled VEGF-releasing device for induction of therapeutic angiogenesis 03.05.2015 Ascona, Switzerland Marsano Anna; Cerino Abdin Giulia; Gaudiello Emanuele;
Cardiovascular and Metabolic Research Meeting Poster Engineering cell-based patch as a controlled VEGF-releasing device for induction of therapeutic angiogenesis 22.01.2015 Fribourg, Switzerland Gaudiello Emanuele; Marsano Anna;
The 4th Lugano Stem Cell Meeting Talk given at a conference Cell-Based Gene Therapy For The Engineering Of A Cardiac Patch With Controlled Angiogenic Potential 24.06.2014 Lugano, Switzerland Marsano Anna; Gaudiello Emanuele;
Termis EU Talk given at a conference Cell-based gene therapy for the engineering of a cardiac patch with controlled angiogenic potential 10.06.2014 Genova, Italy Marsano Anna; Gaudiello Emanuele;


Communication with the public

Communication Title Media Place Year
Talks/events/exhibitions Tag der Biomedizin German-speaking Switzerland 2016

Awards

Title Year
Award for the best poster presentation by the by Swiss Stem Cell Foundation for the study, Cell-Based Gene Therapy For The Engineering Of A Cardiac Patch With Controlled Angiogenic Potential, at the 4th Lugano Stem Cell Meeting, June 2014, University of Lugano, CH. 2014

Associated projects

Number Title Start Funding scheme
172989 Engineered patches for cardiac repair 01.06.2017 Project funding (Div. I-III)

Abstract

Background. Revascularization strategies are the current standard clinical procedure for chronic myocardial ischemia which otherwise, if not treated, leads to progressive deterioration of cardiac function and eventually to end-stage heart failure. Nevertheless, some patients might benefit from an adjuvant angiogenic therapy supporting the growth of new blood vessels in the ischemic tissue by delivery of pro-angiogenic factors (e.g. vascular endothelial growth factor, VEGF). However, uncontrolled microenvironmental VEGF expression has been shown to cause the growth of aberrant angioma-like vascular tumors. Therefore, we developed a FACS-based technique to rapidly purify transduced adipose tissue-derived stem cells (ASC) that homogeneously express specific and safe VEGF levels from a heterogeneous primary population. Rationale. Controlled VEGF delivery by direct intra-myocardial injection of retrovirally transduced and FACS-purified ASC induced robust growth of normal and stable vessels both in normal and ischemic myocardium, and reliably avoided the growth of aberrant angioma-like structures. In the rat ischemic model only controlled VEGF expression caused a moderate functional improvement. However, survival of the injected progenitors was very limited, in agreement with reported results in the literature. The limited cell engraftment might explain the moderate success of cell injection-based clinical studies. To overcome this limitation we engineered a cardiac 3D patch, generated by co-culture of neonatal cardiomyocytes and this time skeletal myoblasts expressing a controlled VEGF level. We observed a robust and normal angiogenesis in the engineered graft, which supported the implanted cell survival and differentiation. Surprisingly, angiogenesis was also greatly improved in the underlying myocardium, where VEGF-expressing cells were not present. Cardiac contractility was also significantly improved in a mouse infarction model. These results show that a patch loaded with genetically engineered progenitors not only lead to a high cell survival upon implantation but also has the potential to deliver controlled VEGF levels to the underlying ischemic myocardium and induce therapeutic vascular growth.We propose in this grant application to develop a cell-based controlled VEGF release system in order to induce a normal and an efficient angiogenesis within the patch itself and in the surrounding cardiac tissue.Specific aims. This proposal covers three specific aims: 1. Obtain the proof of principle that VEGF-expressing ASC can reliably induce normal angiogenesis inside and around the engineered patch (Aim 1a) and then determine the minimum amount of engineered cells necessary to ensure the intrinsic angiogenic potential of the patch (Aim 1b).2. Investigate the role of fresh stromal vascular fraction (SVF) derived cells, as source of endothelial progenitors and pericytic origin cells, in the patch vascularization dynamics and in the stabilization of the vascular networks when co-cultured with ASC expressing either specific (Aim 2a) or heterogeneous (Aim 2b) VEGF levels. 3. Test the efficacy and safety of the engineered cell-loaded patch as a controlled VEGF-releasing device to treat cardiac ischemia in rats (Aim 3).Experimental design. FACS purified populations of retrovirally-transduced ASC expressing either heterogeneous or specific rat VEGF164 levels will be loaded on a collagen based scaffold by using bioreactor-based culture system. Co-culture with SVF will be used to accelerate the in vivo patch vascularization. First, we will use an ectopic rat model to screen out the most promising patch composition to induce normal angiogenesis within the patch and in the surrounding tissue. Then, the intrinsic angiogenic potential of the selected engineered patch will be tested in a cardiac ischemic rat model assessing cardiac functionality.Expected value of the proposed project. The proposed project is expected to provide a basic proof-of-principle to use a cell-based angiogenic factor release system for promoting vascularization in the surrounding tissue. The optimization process will be pursued in order to minimize the quantity of transduced ASC needed and to open up a co-culture with SVF. Furthermore, the proposed research will also provide insights of the angiogenic dynamics in vivo and will elucidate the role of SVF in the vascularization process.
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