1. Cardiovascular Tissue Engineering Priya Ramaswami July 26, 2006 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 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 estimated to be on the order of hundreds of billions of dollars a year As such, the field of tissue engineering has grown to encompass a number of scientific disciplines with the ever-increasing demand for clinical methods to replace and regenerate tissue

4. Biomaterials Provide cells/tissue with a scaffold on which to grow and/or deliver drugs, cytokines, growth factors, and other signals for cell differentiation, growth, and organization Synthetic biomaterials provide a number of parameters that can be adjusted for optimal mechanical, chemical, and biological properties for a given application 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

5. Embryonic Stem Cells (ESCs) Adapted from Gepstein,L. Circ. Res, 91:866; 2002 Collected at the blastocyst stage (day 6) of embryogenesis Collected at the blastocyst stage (day 6) of embryogenesis Give rise to cells from all three germ layers of the body (ectoderm, endoderm, and mesoderm) Give rise to cells from all three germ layers of the body (ectoderm, endoderm, and mesoderm) Capable of self- renewal and undifferentiated proliferation in culture for extended periods of time Capable of self- renewal and undifferentiated proliferation in culture for extended periods of time

6. Mesenchymal Stem Cells (MSCs) Adapted from www.nih.govwww.nih.gov Have been found in many tissues and organs of the body Have been found in many tissues and organs of the body Are multipotent and possess extensive proliferation potential Are multipotent and possess extensive proliferation potential Bone marrow-derived adult stem cells have been differentiated to a number of cell types including bone, cartilage, and fat Bone marrow-derived adult stem cells have been differentiated to a number of cell types including bone, cartilage, and fat Use of adult stem cells allows for autologous cell transplantation Use of adult stem cells allows for autologous cell transplantation

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 for tissue engineering applications 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 for tissue engineering applications Culture of cells in a 3-D vs. 2-D environment has been shown to alter cell behavior, gene expression, proliferation, and differentiation Culture of cells in a 3-D vs. 2-D environment has been shown to alter cell behavior, gene expression, proliferation, and differentiation

9. 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.

10. 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

11. 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

12. Requirements for a TEHV Biocompatible Should not elicit immune or inflammatory response Functional Adequate mechanical and hemodynamic function, mature ECM, durability Living G rowth and remodeling capabilities of the construct should mimic the native heart valve structure

13. 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.

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 Fail in small-diameter replacements WHY??? Intimal hyperplasia Thrombosis

16. Requirements for a TEBV Biocompatible Biocompatible Should not elicit immune/inflammatory response Functional Functional Adequate mechanical and hemodynamic function, mature ECM, durability 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, PUs) Natural (collagen) Decellularized biological matrices Mechanical Stimulation Pulsatile Flow Systems Cyclic & longitudinal strain From An Introduction to Biomaterials. Ch 24. Fig.5 Ramaswami, P and Wagner, WR. 2005. Signalling Factors Growth Factors (bFGF, PDGF, VEGF) Cytokines

18. 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

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

20. 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 Mechanical Stimulation Pulsatile Flow Systems Rotational seeding Cyclic mechanical strain From An Introduction to Biomaterials. Ch 24. Fig.6 Ramaswami, P and Wagner, WR. 2005. Signalling Factors Growth Factors (Insulin, transferrin, PDGF, 5-azacytidine) Cytokines Conditioned media Co-culture`

21. 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