1. Cardiovascular Tissue Engineering Devin Nelson July 2010 Department of Bioengineering, University of Pittsburgh McGowan Institute for Regenerative Medicine

2. Overview Tissue Engineering Biomaterials Cells Tissue Engineered Heart Valves Tissue Engineered Blood Vessels Tissue Engineered Myocardium Discussion

3. Tissue Engineering In recent years, the field of tissue engineering (TE) has emerged as an alternative to conventional methods for tissue repair and regeneration Health care costs in the U.S. for patients suffering from tissue loss and/or subsequent organ failure are $100,000,000,000’s of dollars a year TE has grown to encompass many scientific disciplines Bioengineers Clinicians Pathologists Material Scientists Molecular Biologists Mechanical Engineers etc

4. SignalsScaffolds Tissue Engineered Construct Cells Autogeneic Allogeneic Xenogeneic Primary Stem Natural Synthetic Growth Factors Cytokines Mechanical Stimulation Differentiation Factors From An Introduction to Biomaterials. Ch 24. Fig. 1. Ramaswami, P and Wagner, WR. 2005.

5. What do these have in common? All Biomaterials

6. Biomaterials Synthetic biomaterials Engineer can control the properties such as mechanical strength, biological activity, degradation rates etc Natural biomaterials Built-in structure, environment and cues similar to native body (extracellular matrix ECM, collagen, etc) Deliver drugs, cytokines, growth factors, and other signals for cell differentiation, growth, and organization Design criteria: proper mechanical and physical properties adequate degradation rate without the production of toxic degradation products suitable cell adhesion integration into surrounding tissue without extensive inflammatory response or support of infection proper mass transfer

7. Cells There has recently been much excitement surrounding the use of stem cells for tissue repair and regeneration In vitro differentiation of stem cells via humoral factors and direct in vivo utilization of these cells have been proposed as a method for tissue regeneration The use of a biomaterial to guide stem cell commitment provides cells a scaffold on which to grow and permits cell differentiation in vivo while minimizing in vitro manipulation The ideal cell source for various TE applications is still elusive

8. 3-Dimensional Environment The context in which a cell is grown is critical to its development and subsequent function The context in which a cell is grown is critical to its development and subsequent function Cells cultured ex vivo on TCPS are in a 2-D environment which is far-removed from the 3-D tissue from which the cells originated as well as the 3-D tissue into which the cells will be implanted Cells cultured ex vivo on TCPS are in a 2-D environment which is far-removed from the 3-D tissue from which the cells originated as well as the 3-D tissue into which the cells will be implanted Culture of cells in a 3-D vs. 2-D environment AND WITH APPROPRIATE MECHANICAL STIMULATION has been shown to alter cell behavior, gene expression, proliferation, and differentiation Culture of cells in a 3-D vs. 2-D environment AND WITH APPROPRIATE MECHANICAL STIMULATION has been shown to alter cell behavior, gene expression, proliferation, and differentiation Especially for cardiovascular applications Especially for cardiovascular applications

9. Tissue Engineered Heart Valves (TEHV) An estimated 87,000 heart valve replacements were performed in 2000 in the United States alone Approximately 275,000 procedures are performed worldwide each year Heart valve disease occurs when one or more of the four heart valves cease to adequately perform their function, thereby failing to maintain unidirectional blood flow through the heart Surgical procedures or total valve replacement are necessary Adapted from http://z.about.com/d/p/440/e/f/19011.jpg

10. TEHV Replacements Mechanical prostheses Bioprostheses Homografts Each of these valve replacements has limitations for clinical use Can you think of any limitations? Infection Thromboembolism Tissue deterioration Cannot remodel, repair, or grow From http://www.rjmatthewsmd.com/Definitions/img/107figure.jpg

11. Requirements for a TEHV Biocompatible Should not elicit immune or inflammatory response Functional Adequate mechanical and hemodynamic function, mature ECM, durability to open and close > 31 million times a year Living G rowth and remodeling capabilities of the construct should mimic the native heart valve structure

12. What’s being done? Cells Vascular cells Valvular cells Stem cells (MSCs) Scaffolds Synthetic (PLA, PGA) Natural (collagen, HA, fibrin) Decellularized biological matrices Mechanical Stimulation Pulsatile Flow Systems Cyclic flexure bioreactors From An Introduction to Biomaterials. Ch 24. Fig.3 Ramaswami, P and Wagner, WR. 2005.

13. R.T. Tranquillo Biomaterials 30 (2009) 4078–4084.

14. Tissue Engineered Blood Vessels (TEBV) From An Introduction to Biomaterials. Ch 24. Fig.4 Ramaswami, P and Wagner, WR. 2005. Atherosclerosis, in the form of coronary artery disease results in over 515,000 coronary artery bypass graft procedures a year in the United States alone Many patients do not have suitable vessels due to age, disease, or previous use Synthetic coronary bypass vessels have not performed adequately to be employed to any significant degree

15. TEBV Replacements Synthetic Grafts Work well in large- diameter replacements (6-10 mm) Fail in small-diameter replacements (3-5 mm) WHY??? Intimal hyperplasia Thrombosis

16. Requirements for a TEBV Biocompatible Biocompatible Should not elicit immune/inflammatory response Functional Functional Adequate mechanical (burst pressure) and hemodynamic function, mature ECM, durability, nervous system response Living Living Growth and remodeling capabilities of the construct should mimic the native blood vessel structure LOOK FAMILIAR???

17. What’s being done? Cells Endothelial cells Smooth muscle cells Fibroblasts & myofibroblasts Genetically modified cells Stem cells (MSCs & ESCs) Scaffolds Synthetic (PET, ePTFE, PGA, PLA, PU) Natural (collagen) Decellularized biological matrices Mechanical Stimulation Pulsatile Flow Systems Cyclic & longitudinal strain Signalling Factors Growth Factors (bFGF, PDGF, VEGF) Cytokines

18. Cell-Seeded Collagen Cells can remodel and reorganize in collagen Cells can remodel and reorganize in collagen Collagen may be weak but is strengthened through various techniques (magnetic pre- alignment, glycation, mechanical training) Collagen may be weak but is strengthened through various techniques (magnetic pre- alignment, glycation, mechanical training)

19. Mechanical Training Seliktar et al. Ann Biomed Eng 2000

20. Self-Assembled Sheets Good 3D architecture Good 3D architecture Good mechanical strength Good mechanical strength Disadvantages: need cell source, requires > 2 months in vitro to make Disadvantages: need cell source, requires > 2 months in vitro to make

21. Seeding and Culture

22. Electrospinning Stankus et al. Biomaterials 2007

23. Tissue Engineered Myocardium From www.aic.cuhk.edu.hk/web8/Hi%20res/Heart.jpg Ischemic heart disease is one of the leading causes of morbidity and mortality in Western societies with 7,100,000 cases of myocardial infarction (MI) reported in 2002 in the United States alone Within 6 years of MI, 22% of men and 46% of women develop CHF MI and CHF will account for $29 billion of medical care costs this year in the US alone Cardiac transplantation remains the best solution, but there is an inadequate supply of donor organs coupled with the need for life-long immunosuppression following transplantation

24. Requirements for a Myocardial Patch Biological, Functional, and Living (same as TEHV and TEBV) High metabolic demands High cell density Complex cell architecture High vascularity Mechanical and Electrical anisotropy VERY DIFFICULT!!!

25. What’s being done? Cells Cardiocytes Cardiac progenitor cells Skeletal muscle cells Smooth muscle cells Stem cells (MSCs & ESCs) Scaffolds Synthetic (PET, ePTFE, PEUU) Natural (collagen, ECM proteins, alginate) Cell sheets Injectables Mechanical Stimulation Pulsatile Flow Systems Rotational seeding Cyclic mechanical strain Signalling Factors Growth Factors (Insulin-like, bFGF, PDGF, hGH) Cytokines Conditioned media Co-culture

26. Cardiac Patch

27. Cell Sheet Engineering

28. Artificial Muscle – Be Creative NATIVE TISSUE ENGINEERED

29. Extracellular Matrix

30. Injectable Material

32. In Conclusion… We have a lot of work to do We have a lot of work to do Taking these tissue engineered constructs from benchtop to bedside Taking these tissue engineered constructs from benchtop to bedside Better understanding the human body and how to manipulate cells Better understanding the human body and how to manipulate cells