C HEMICAL AND P HYSICAL R EGULATION OF S TEM C ELLS AND P ROGENITOR C ELLS : P OTENTIAL FOR C ARDIOVASCULAR T ISSUE E NGINEERING (R EVIEW ) N GAN F. H.

CHEMICAL AND PHYSICAL REGULATION OF STEM CELLS AND PROGENITOR CELLS: POTENTIAL FOR CARDIOVASCULAR TISSUE ENGINEERING (REVIEW)

NGAN F. HUANG, RANDALL J. LEE, SONG LI

By Deepika Chitturi

BIOE 506

Spring 2009

WHY CARDIOVASCULAR TISSUE ENGINEERING? Leading Cause of Mortality

(every 34 sec) Expensive ($250 billion) Myocardial Infarction (MI aka

heart-attacks)

Coronary Artery Occlusion Cardiomyocyte Cell Death Non-generation Formation of Scar Tissue Dilation of Chamber Cavities Aneurysmal Thinning of Walls

REDUCED PUMPING CAPACITY

Driving Force: Shortage of Donors

POTENTIAL STEM & PROGENITOR CELLS

MSCs: Mesenchymal Stem Cells

HSCs: Hematopoietic Stem Cells

EPCs: Endothelial Precursor Cells

ESCs: Embryonic Stem Cells

Skeletal Myoblasts

Resident Cardiac Stem Cells

PERFECT TISSUE ENGINEERED CONSTRUCT

CELL SOURCE

SOLUBLE CHEMICAL FACTORS

EXTRACELLULAR MATRIX (ECM)

CARDIOVASCULAR TISSUE ENGINEERING (I)

Cell Source

Embryonic Stem Cells Adult Stem Cells

Soluble Chemical Factors

VEGF (ESCs, HSCs, EPCs) TGF-β (ESCs, MSCs, HSCs,

EPCs) BMP (ESCs) 5-azacytidine (MSCs) FGF (ESCs, HSCs, EPCs) IGF (HSCs, EPCs)

CARDIOVASCULAR TISSUE ENGINEERING (II) Extracellular Matrix

Natural Polymers

Matrigel: In vivo injection for MI, ESC differentiation Collagen: In vivo injection for MI, Vascular grafts Hyalinuric Acid: Vascular grafts Alginate: ESC differentiation Fibrin: In vivo injection for MI, Vascular conduits Decellularized Vessel: Vascular conduits

Synthetic Polymers

Poly-L-lactic Acid (PLLA): ESC differentiation Poly-lactic-co-glycolic acid (PLGA): ESC differentiation Polyglycotic Acid (PGA): Vascular grafts Peptide Nanofibers: In vivo injection for MI Poly-diol-citrates and Poly-glycerol-sebacate: General tissue

engineering

EXTRACELLULAR MATRIX

Dr. Vasif Harsirci- Middle East Technical University (Biomedical Unit)

Matrigel Angiogenesis PLLA Angiogenesis

Effects of Cordyceps militaris extract on angiogenesis and tumor growth1 Hwa-seung YOO, Jang-woo SHIN2, Jung-hyo CHO, Chang-gue SON, Yeon-weol LEE, Sang-yong PARK3, Chong-kwan CHO4 Department of East-West Cancer Center, College of Oriental Medicine, Daejeon University, Daejeon 301-724;

ROLE OF MATRIX MATERIALS FOR STRUCTURAL SUPPORT hESCs cultured in porous PLGA/PLLA scaffolds coated

with Matrigel or Fibronectin vs. Matrigel alone or fibronectin-coated dishes (Levenberg et al)

3-D polymer structure promoted differentiation (neural tissue, cartilage, liver and blood vessels)

Formation of 3-D blood vessels Fibronectin-coated dishes:

Failure to organize into 3-D structure Matrigel:

Organization into 3-D structure No cell differentiation

Conclusion:

Large inter-connected pores: cell colonization Pores smaller than 100 nm: limit diffusion of nutrients and gases 3-D: great surface area, higher expression of integrins

ROLE OF MATRIX TOPOGRAPHY AND RIGIDITY Topography: Cell Organization, alignment and

differentiation

Nano-scale and micro-scale matrix topography affects organization and differentiation of stem cells

hMSCs undergo skeletal reorganization and orient themselves in the direction of microgrooves and nano-fibers (Patel et al)

Stiffness/Rigidity: Cells tend to migrate toward more-rigid surfaces and cells on soft matrix have a low rate of DNA synthesis and growth (Engler et al)

Assembly of focal adhesions and contractile cytoskeleton structure depend on rigidity

CARDIOVASCULAR TISSUE ENGINEERING MODELS

In vitro differentiation method: engineering constructs with structural and functional properties as native tissues before transplantation

In situ method: relies on host environment to remodel the chemical and physical environment for cell growth and function

Ex vivo approach: excision of native tissues and remodeling them in culture

CARDIOVASCULAR TISSUE ENGINEERING PROPOSED MODELS

Injectable Stem Cells and Progenitor Cells for in situ cardiac tissue engineering

Vascular Conduits

INJECTABLE STEM CELLS AND PROGENITOR CELLS FOR IN SITU CARDIAC TISSUE ENGINEERING

Delivery modes for myocardial constructs:

Cardiac patching

Cell Injection

Cell-polymer injection

Less invasive than solid scaffolds Adopt shape and form of host environment Delivery vehicles (with cells and GFs) Polymers: Collagen I, Matrigel, Fibrin, Alginate and

Peptide Nanofibers

INJECTABLE DELIVERY OF POLYMERS

Collagen I, Matrigel and Fibrin

Higher capillary density than saline control treatment Migration of vascular cells into infarcted region for

neovascularization

Fibrin + MSCs (Huang et al)

Promotes angiogenesis

ESCs + Matrigel (Kofidis et al)

Greater improvements in contractility after 2 weeks

Rat bone marrow mononuclear cells (MNCs) + Fibrin (Ryu et al)

Enhanced neovascularization Development of larger vessels Extensive tissue regeneration Graft survival: 8 weeks

TREATMENT USING STEM AND PROGENITOR CELLS ALONE TGF-β-treated CD117+ rat MNCs (Li et al)

Differentiation into myogenic lineage Enhanced vascular density

Retrovirally transduced Akt1-overexpressing MSCs (Mangi et al, Laflamme et al)

Reduced intramyocardial inflammation 80% of lost myocardial volume regeneration Normal systolic and diastolic functions restoration

Cardiac enriched hESCs in athymic rats (Laflamme et al)

Cardiomyocyte growth No teratomas 7-fold increase in graft size in 4 weeks Potential regeneration of human myocardium in rat heart

VASCULAR CONDUITS

Goal: To create functional conduit as a bypass graft (small, non-thrombogenic, native mechanical properties)

Limitations to vein grafts: Availability 35% 10-year failure

Synthetic Vascular Grafts: Poly-ethylene-terephthalate Expanded poly-

tetrafluoroethylene Polyurethane Limitation:

Inside diameter larger than 5 mm Frequent thrombosis and

occlusions in smaller grafts

VASCULAR CONDUITS—PROPOSED MODELS ECs + SMCs in a tubular PGA porous scaffold (Niklason et

al)

In vivo implantation: patent for 2 weeks; development of histological features consistent with vascular structures

EPC-seeded grafts (Kaushal et al)

Remained patent for more than 130 days Acellular control grafts occluded in 15 days Vessel-like characteristics: contractility and nitric-oxide mediated

vascular relaxation

EPCs derived from umbilical cord blood using 3D porous polyurethane tubular scaffolds in a biomimetic flow system (Schmidt et al)

In 12 days, EPCs lined lumen of VGs and formed endothelial morphology

VASCULAR CONDUITS—PROPOSED MODELS

MSC seeded nanofibrous vascular grafts (Hashi et al)

Patent for at least 8 weeks Synthesis and organization of collagen and

elastin EC monolayer formed on lumen surfaces SMCs were recruited and formed

CONCLUSION

Understanding the effect of chemical and physical cues for regulation of stem-cell survival, differentiation, organization and morphogenesis into tissue-like structures: most important!!

Cardiovascular repair, Cardiac therapies after MI and engineering of vascular conduits