SYNTHETIC EXTRACELLULAR MATRICES FOR PROMOTING ANGIOGENESIS

Eduardo A. SilvaEduardo A. Silva

Harvard University, Cambridge, MA, USAHarvard University, Cambridge, MA, USAandand

Faculdade de Engenharia da Universidade do Porto, Porto, PortugalFaculdade de Engenharia da Universidade do Porto, Porto, Portugal

December 2004December 2004Porto, PortugalPorto, Portugal

MOTIVATION

Cardiovascular Diseases:

- Principal cause of mortality in western countries

Current Approaches to treat Ischemic Heart Diseases:

Pharmacologic

Invasive Surgical

Therapeutic Angiogenesis

ISCHEMIA

Carmeliet & Jain Nature, 2000

Ischemia - isch- is restriction, hemia or haemia is blood

ANGIOGENESISSprouting of new blood vessels from

pre-existent vessels

VASCULAR ENDOTHELIAL GROWTH FACTOR

VEGF

Is a key regulator of blood vessels formation and function

Endothelial Cells are the primary target of VEGF

45 kDa homodimer

Four isoforms obtained by alternative splicing:

VEGF121, VEGF165, VEGF189, VEGF206

Muller et al., Structure, 5:1325-1338 (1997)Muller et al., Structure, 5:1325-1338 (1997) Fairbrother et al., Structure, 6:637-648 (1998)

AIM I: Investigate VEGF signaling in cardiac cells under hypoxia.

AIM I

VEGF SECRETION UNDER HYPOXIA

TISSUE ENGINEERING STRATEGY

Localized, sustained delivery from bioactive polymeric systems

Constant local concentration

Protection from degradation

Minimal side effects

AIM II

Development of an injectable polymeric system for controlled and defined delivery of angiogenic molecules

VEGF incorporated in an injectable polymer

POLYMERIC SYSTEMAlginate

OOH

CO2-

O

O

OH

-O2C

OH

O

HO

CO2- OH

OHO

CO2-

OHO

O

OOH

OHO

CO2-

O

OH

G G MM G

Ca2+

-L- Guluronic acid

-D- Mannuronic acid

Gelation process

Hydrogel

MODEL SYSTEM Distinct approach used to incorporate VEGF

Simple mixing VEGF and alginate

Pre-incorporation of VEGF in polymeric microspheres and immobilization in the alginate

+ +Ca2+

Angiogenic moleculesAlginate

+ + Microspheres

Polymeric system

Bioactive moleculesBioactive molecules

ALGINATE MODIFICATIONSCombination of binary molecular weight distribution and partial oxidation

Oxidation with NaIOOxidation with NaIO44

Irradiation (5 Mrad)Irradiation (5 Mrad)

CHOCHO

Solution Properties

Gel Properties

VEGF RELEASE

BIOACTIVITY OF VEGF RELEASED

0

5000

10000

15000

20000

25000

0 1 4

Time (days)

Cell number

MediaMedia/VEGFMedia/VEGF released

ANGIOGENESIS

PDGF RELEASE

VEGF/PDGF RELEASE

In vivo evaluation of VEGF releasing materials

Model: Ischemic hind limb of Apo E null (Apo E -/-)

LDPI image

In vivo evaluation of VEGF releasing materials

Blank

VEGF&PDGF

VEGF

Week 2 Week 4 Week 6

In vivo evaluation of VEGF releasing materials

CONCLUSIONS

Sustainable and controlled angiogenic molecules delivery can be achieved with

binary alginate

Blood vessel formation and perfusion can be significantly improved by using binary

alginate as local dual delivery vehicle

FUTURE WORKVEGF 121 AND VEGF 165 PLAY DISTINCT ROLES IN THE STIMULATION OF COLLATERAL VESSEL AND BLOOD FLOW RE-ESTABLISHMENT IN AN ISCHEMIC HEART

VEGF 121

µmµm

VEGF 165

-+

ACKNOWLEDGMENTSProfessor David J. Mooney

Professor Mário Barbosa

Department of RadiologyDivision of Cardiovascular DiseasesDepartment of Internal MedicineUniversity of Michigan

Paul M. Grossman

Sanjay Rajagopalan

Qinghua Sun

Mooney Lab

2nd PGDB

FCT - Fundação para a Ciência e a Tecnologia

IGC/FCG - Fundação Calouste Gulbenkian

NIH - National Institutes of Health

Harvard University/University of Michigan & Universidade do Porto

Thinking about Curia 2005

A - Alginate; B - New tissue; C - Blood Vessel; D - Inflammatory cells