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Cardiac Tissue Engineering - Term Paper Example

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The author analyzes the ‘Mending Broken Hearts with Tissue Engineering’ article which describes a novel scaffold for seeding stem cells and heart cells. The article presents results in the new field of cardiac tissue engineering that seeks to revolutionize the treatment of end-stage heart failure. …
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Cardiac Tissue Engineering
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Topic: Transplantation of Selected Article: Mending Broken Hearts with Tissue Engineering Source: Massachusetts Institute of Technology, November 4, 2008. ScienceDaily Introduction The news article, ‘Mending Broken Hearts with Tissue Engineering’ (Massachusetts Institute of Technology, 2008), describes a novel scaffold for seeding stem cells and heart cells that would develop into heart tissues. The accordion-like, honeycomb, polymer scaffold matches the structural and mechanical characteristics of heart tissues by being directionally dependent. This means that the tissues that were cultivated on the scaffold gave similar electrophysiological responses to those of the native heart. The nature of the scaffold also intrinsically guides the alignment of cultured cells even without electrical stimulation (Engelmayr et al., 2008). Compared to previous scaffolds, this one has mechanical properties that are very similar to those of the native heart. The chosen article presents results in the relatively new field of cardiac tissue engineering that seeks to revolutionize treatment of end-stage heart failure and address the scarcity of heart donors.   The term ‘tissue engineering’ was officially coined in 1987 during a meeting of the US National Science Foundation (NSF) in Washington, D.C. It was later defined as the ‘application of principles and methods of engineering and life sciences toward fundamental understanding of structure–function relationship in normal and pathological mammalian tissues and the development of biological substitutes to restore, maintain, or improve functions’ (as quoted in Eschenhagen & Zimmermann, 2005). Tissue engineering employs biological, engineering, and material sciences to replace and improve the function of biological tissues. There are three general strategies used (Langer & Vacanti, 1993): 1. isolated cells or substitutes, which allows for the replacement only of the specific cells that can provide the original function; 2. tissue-inducing substances like growth factors that need to be purified and delivered to target molecules, and 3. implantation of cells on matrices to the body either in closed systems (where the implants are isolated from other tissues to prevent immune system attack), and open systems where the cells are incorporated into the system. Matrices (or scaffolds) may be created from natural materials or from synthetic polymers. The current techniques involve the production of tissues from donor cells that have been seeded on three-dimensional polymeric scaffolds, then inducing growth of new healthy tissues by culture and implantation of these scaffolds to the chosen organ. This approach holds promise for the treatment of heart ailments like myocardial infarction (MI) or heart attack, and congenital heart diseases. During MI, the heart muscle cells (cardiomyocytes) are damaged or die which result in scar tissues, aneurysm, and increased stress on the myocardium, and eventually left ventricular dilatation and heart failure. Apparently, the cardiomyocytes are not normally replaced after injury. The repair of myocardial damage has not been possible despite advances in medicine (Mann, 1999), and the feasible option is heart transplantation, which is limited by the shortage of donor hearts. Therefore, cardiac tissue engineering is a very promising approach because it has the potential to grow new tissue with the potential to multiply and provide self-repair and heart remodelling. Advances in Cardiac Tissue Engineering Even before the name “cardiac tissue engineering” was invented, isolated heart tissues have long been known to beat spontaneously in laboratory dish cultures. The in vitro cultivation of heart tissues, even those of different individuals of the same species, can unite and beat synchronously. This happens when there is cell contact which appeared to be necessary for physiological identity and recognition among the heart cells (Fischer, 1924). Later, the possibility of reproducing heart cells in sufficient amounts using a simple bioreactor. Embryonic chick hearts generated aggregates (as many as 200 cells) under continuous mixing (Moscona & Moscona, 1952). This model was adapted to produce aggregates that were more similar functionally to intact heart tissues (McDonald, Sachs, & DeHaan, 1972), proving that isolated cells have the capacity to re-form the same tissues. Later studies were able to show the preferential aggregation of cardiac myocytes when high densities of cells, cultured in serum media, produced monolayers. Tissue engineering without matrix has refined the detachment of the cellular monolayers after culture at prolonged periods. The use of thermo-sensitive surface coating for culture dishes allows cardiac myocytes to attach at 37°C, but can detach as an intact sheet of beating myocytes (Shimizu et al., 2002). These monolayers form exogenous matrix-free interconnected 3D tissue sheets (Shimizu et al., 2002). The disadvantages of using the monolayer sheets are their fragility, which lead to handling problems, the restriction in terms of moulding the tissues to geometric form and difficulties of imposing mechanical load on the contractile sheets (Eschenhagen & Zimmermann, 2005). In addition, the detachment did not improve cellular differentiation and tissue formation. Since heart cells are continuously mechanically stressed during pumping with fluctuating blood pressure and volume, it was hypothesized that the heart cells will differentiate in response to mechanical load. Cells grown on a membrane were stressed by cyclical distention several times a minute. After 24 h of this, the cells elongated and oriented perpendicularly to the direction of the stretch, indicating that mechanical stimulation directly influenced the heart cellular organization in vitro (Terracio, Miller, & Borg, 1988). Emphasizing the role of mechanical load was the development of the first engineered three dimensional and well-developed muscle strips that were produced from cultured cells subjected to repeated cycles of stretch and relaxation (Vandenburgh, Swasdison, & Karlisch, 1991). Electric stimulation also appears to induce cardiac myocyte differentiation similar to mechanical stimulation (Fink et al., 2000; Zimmermann et al., 2002). The additional use of electrical stimulation in cultures (Radisic et al., 2004) has resulted in new heart muscle constructs that have better tissue forms, and contractile function. Electrical signals were used to improve function and structure of engineered myocardium. After only 8 days in vitro, electric stimulation resulted in a high level of ultrastructural organization concurrent with the development of conductive and contractile properties of cardiac tissue constructs. Another approach used in cardiac tissue engineering is the use of an effective scaffold where tissues can be seeded and transplanted to heart cells in vitro and in vivo. A scaffold is a template, or extracellular matrix (ECM), that supports the formation from cells of large tissues that have distinct three-dimensional form. The scaffold assists in the biosynthesis and proliferation of cells, and if placed in the site of regeneration, the scaffold prevents other cells from disturbing the site of synthesis. Among the early scaffold materials used were polyglycolic acid in bioreactor cultures (Carrier et al., 1999), gelatin scaffolds which were later implanted into infarcted hearts using Gelfoam (Li et al., 1999); alginate scaffolds seeded with foetal cardiac cells that produced tissue grafts (Leor et al., 2000). The tissues formed using these materials survived only for a limited time and did not form beating heart cells. Moreover, upon implantation to heart organs, the scaffolds were not integrated into the host. Spontaneous contractile activity, both in vitro and in vivo, was reported, but the histological microphotographs showed mainly cells of unknown identity embedded in the scaffold material and poor sarcomere development in the few cardiac myocytes present in the construct. Foetal cardiac cells seeded on alginate scaffolds produced vascularized cells when implanted into rat cells but true integration was not shown (Leor et al., 2000). Despite the shortcomings from using preformed scaffolds, these can be designed and shaped into 3D forms that mimic the natural organ. The limitations of this approach are due to inadequacies of the matrix material like limited diffusion and low mechanical compliance. An ideal scaffold would allow for the survival and proliferation of cardiac myocytes that will beat and integrate into the natural healthy heart. The observed limitations have led to the development of new materials (mostly polymers and biodegradable) that are more refined and promote spreading of cells (reviewed by Leor & Cohen, 2004). How the heart works is due to the complexity of the muscle fibers that form a structure with spaces, interconnections and layers in different orientations. Aside from mechanical stability, the structure of heart muscles is contractile and compliant. Gelatin, polyglycolate and other scaffold materials used do not have all these characteristics. The scaffold with honeycomb structure was announced to have three distinct advantages over earlier scaffolding materials used (Engelmayr et al., 2008). Basing their design on the natural structure and mechanical properties of the heart, Englemayr et al. (2008) used laser technology to create a directionally dependent scaffold. The new scaffold has three main advantages over previously designed scaffolds. First, the mechanical properties are highly similar to tissues of the natural heart; being stiffer when stretched circumferentially than longitudinally. The polymer scaffold’s mechanical characteristics can be fine-tuned by varying the time it was required to set. The second advantage was the formation of tissues that were able to contract more readily in one direction when an electrophysiological field was applied. The third advantage was the capability of the scaffold to direct the orientation of cultured heart cells. Traditional scaffolds can align cultured cells too, but only when electrically stimulated. However, despite the advances some limitations still exist, since the material was not thick enough to reconstruct a myocardium of the thickness that native hearts have (Massachusetts Institute of Technology, 2008). Nevertheless, the aspect of mechanical compliance has been addressed and only minor adjustments in the design are necessary. The researchers also noted that the scaffold used in the experiments described above has some limitations due to lack of its ability to reconstruct myocardium of sufficient thickness. However, new honeycomb scaffolds are being created that, among other things, allow much thicker, multi-layered tissue structures. Growth of cells on scaffolds are affected also by the type of cell sources that are used as seed for engineering cardiac tissue. Other heart cells have been used including fibroblasts, smooth muscle cells, and cardiac myocytes. However, these cells produce limited amounts of new myocytes, thus new types of cells are being used in studies to determine their capacity to build new heart muscle. In 2003, studies were published that showed the presence of myocardium progenitor cells in residing in the heart that can differentiate to functional cardiac myocytes that have the capacity to repair damaged cardiac tissues (Oh et al., 2003; Beltrami et al., 2003). Almost all stem cells (except those derived from skeletal myoblast) can transdifferentiate to cardiac tissue (Van Laake et al., 2006). An advantage of stem cells is their high level of reproducibility, in their undifferentiated state, under culture conditions. The cells can be induced to differentiate into cardiomyocytes under special conditions; already human stem cells have been reported to differentiate in polymeric scaffolds (Levenberg et al., 2003). Currently, preferred cell sources for engineered cardiac tissues are human embryonic stem cells (Habib, Caspi, & Gepstein, 2008). Vascularized muscle tissue has already been engineered from human embryonic stem cells (Caspi, et al., 2007). Challenges in Cardiac Tissue Engineering To summarize, the following strategies have been employed in engineering effective myocardium (Figure 1): (I) seeding cardiac myocytes and, recently, stem cells, on synthetic polymeric or biologic matrices (II) supporting the propensity of cardiac myocytes to form contracting aggregates by entrapment in collagen (III) stacking cardiac myocyte monolayers to form multi-layered heart muscle constructs Figure 1. Summary of the different strategies in cardiac muscle engineering using (I) polymeric matrices as scaffold material, (II) soluble collagen and extracellular matrix (ECM) components to entrap cells (II), and (III) monolayer sheets. Figure was adapted from Zimmermann et al., (2006). Although rapid progress in tissue engineering has been achieved, the feasible transplantation of engineered tissues to patients with heart disease have not been realized. The heart, seemingly a mechanical pump, is an extremely coordinated, flexible, and complex organ. For each of the strategy mentioned, corresponding challenges are also present. The use of stem cells has great promise, but currently, its reproduction in polymeric scaffolds has been an issue. The evidence for ability of mesenchymal stem cells (MSC) and endothelial precursor cells to transdifferentiate to cardiomyocytes is conflicting. Although MSC can differentiate to heart lineage, beating cells are not produced (Siepe et al., 2008). MSC cells also transdifferentiate in vivo but no proof yet for improving their capacity in vitro. The use of human embryonic cells is the current focus because they have been shown to produce cardiomyocytes in vitro. However, the allogeneic origin of embryonic stem cells can result in immune responses and tumour formation (Habib, Caspi, & Gepstein, 2008; Kofidis, et al., 2005). Nevertheless, embryonic stem cells in rats have been found to reversibly inhibit T-cell proliferation to various stimuli, allowing for engraftment across allogeneic barriers (Koch, Geraldes, & Platt, 2008). However, the same response in human embryonic stem cells has not been demonstrated. As research continues for the ideal cells for heart tissue engineering, so should efforts to design better and more sophisticated scaffolds. Specific scaffold and tissue interactions have to be optimized to supply cell their needs for growth and proper differentiation. Stem cells, despite their utility as source of cardiomyocytes, would still be dependent on scaffolds. Although stem cells have been shown to differentiate to other tissue lineages, differentiation to cardiomyocytes in vitro has not been shown (Levenberg et al., 2003). Thus, this area needs to be addressed by new research. In connection with improving transdifferentiation of stem cells to cardiomyocytes, more research is needed on the incorporation of cytokines (growth factors) and other peptides to improve cell growth. The development of in vitro culture media is also necessary specially the characterization of other growth factors. Vision for the Future of Cardiac Tissue Engineering Advancement in the field of heart tissue engineering has been very rapid in the past ten years. New techniques and discoveries in stem cell research, biomaterials, and bioreactors, coupled with the number of researchers working on this area, will be instrumental in coming up soon with viable engineered heart tissue. In integrative approach, where all aspects of cardiac tissue engineering, is necessary because of the many factors involved in engineering the tissue and the response of the host organ (Efimov, 2008). The importance of collaboration of all stakeholders: biomaterials scientists, tissue culture specialists, immunologists, biochemists, engineers and especially medical doctors, should be emphasized. Surgeons who perform the transplantation and evaluation of the scaffolds play a significant role in the success of the technology. Without the close collaboration of scientists among the different fields, it will be unlikely for tissue engineering to respond to the needs of the patients (Ikada, 2006). Nevertheless, with the speed that advances have been going on, it can be safely predicted that in less than ten years, actual transplantation for heart tissue regeneration and repair will be beyond the clinical trial stage. References Beltrami, A, Barlucchi, L, Torella, D, Baker, M, Limana, F, Chimenti, S, Kasahara, H, Rota, M, Musso, E, Urbanek, K, Leri, A, Kajstura, J, Nadal-Ginard, B & Anversa, P 2003, ‘Adult cardiac stem cells are multipotent and support myocardial regeneration’, Cell, Vol. 114, pp. 763-776. Carrier, R, Papadaki, M, Rupnick, M, Schoen, F, Bursac, N, Langer, R, Freed, LE &Vunjak-Novakovic, G 1999, ‘Cardiac tissue engineering: cell seeding, cultivation parameters, and tissue construct characterization’, Biotechnology and Bioengineering, Vol. 64, pp. 580-589. Caspi, O, Lesman, A, Basevitch, Y, Gepstein, A, Arbel, G, Huber, I, Habib, M, Gepstein, L & Levenberg, S 2007, ‘Tissue engineering of vascularized cardiac muscle from human embryonic stem cells’, Circulation Research, Vol. 100, pp. 263-272. Efimov, I 2008, ‘Nature versus nurture in cardiac conduction: toward integrative paradigm of cardiac tissue engineering’, Circulation Research, Vol. 103, pp. 119-121. Engelmayr, G, Cheng, M, Bettinger, C, Borenstein, J, Langer, R, & Freed, L 2008, ‘Accordion-like honeycombs for tissue engineering of cardiac anisotropy’. Nature Materials, Vol. 7, pp. 1003-1010. Eschenhagen, T & Zimmermann, W 2005, ‘Engineering myocardial tissue’, Circulation Research, Vol. 97, pp.1220-1231. Fink, C, Ergun, S, Kralisch, D, Remmers, U, Weil, J & Eschenhagen, T 2000, ‘Chronic stretch of engineered heart tissue induces hypertrophy and functional improvement’, Faseb Journal, Vol. 14, pp. 669-679. Fischer, A1924, ‘The interaction of two fragments of pulsating heart tissue’, Journal of Experimental Medicine, Vol. 39, pp. 577-583. Habib, M, Caspi, O & Gepstein, L 2008, ‘Human embryonic stem cells for cardiomyogenesis’, Journal of Molecular and Cellular Cardiology, Vol. 45, No. 4, pp. 462-474. Ikada, Y 2006, ‘Challenges in tissue engineering’, Journal of the Royal Society Interface, Vol. 3, pp. 583-589. Koch, C, Geraldes, P & Platt, J 2008, ‘Immunosuppression by embryonic stem cells’, Stem Cells, Vol. 26, pp. 89-98. Kofidis, T, Debruin, J, Tanaka, M, Zwierzchoniewska, M, Weissman, I, Fedoseyeva, E, Haverich, A & Robbins, RC 2005, ‘They are not stealthy in the heart: embryonic stem cells trigger cell infiltration, humoral and T-lymphocyte-based host immune response’, Vol. 28, pp. 461-466. Langer, R & Vacanti, J 1993, ‘Tissue engineering’, Science, Vol. 260, pp. 923-926. Leor, J & Cohen, S 2004, ‘Myocardial tissue engineering: creating a muscle patch for a wounded heart’, Annals of the New York Academy of Science, Vol. 1015, pp.312-331. Leor, J, Aboulafia-Etzion, S, Dar, A, Shapiro, L, Barbash, I, Battler, A, Granot, Y & Cohen, S 2000, ‘Bioengineered cardiac grafts: A new approach to repair the infarcted myocardium?’, Circulation, Vol. 102, pp. III56-III61. Levenberg, S, Huang, N, Lavik, E, Rogers, A, Itskovitz-Eldor, J & Langer, R 2003, ‘Differentiation of human embryonic stem cells on three dimensional polymer scaffolds’, Proceedings of the National Academy of Sciences USA, Vol. 100, No.21, pp. 12741–12746. Li, R, Jia, Z, Weisel, R, Mickle, D, Choi, A & Yau, T 1999, ‘Survival and function of bioengineered cardiac grafts’, Circulation, Vol. 100, pp. II63-II69. Mann, D 1999, ‘Mechanisms and models in heart failure’, Circulation, Vol. 100, pp. 999-1008. Massachusetts Institute of Technology, 2008, November 4, ‘Mending broken hearts with tissue engineering’ ScienceDaily, viewed January 20, 2009, McDonald, T, Sachs, H, & DeHaan, R 1972, ‘Development of sensitivity to tetrodotoxin in beating chick embryo hearts, single cells, and aggregates’, Science, Vol. 176, pp.1248-1250. Moscona, A & Moscona, H 1952, ‘The dissociation and aggregation of cells from organ rudiments of the early chick embryo’, Journal of Anatomy, Vol. 86, pp. 287-301. Oh, H, Bradfute, S, Gallardo, T, Nakamura, T, Gaussin, V, Mishina, Y, Pocius, J, Michael, LH, Behringer, RR, Garry, DJ, Entman, ML & Schneider, MD 2003, ‘Cardiac progenitor cells from adult myocardium: homing, differentiation, and fusion after infarction’, Proceedings of the National Academy of Sciences USA, Vol. 100, No. 21, pp. 12313-12318. Radisic, M, Park, H, Shing, H, Consi, T, Schoen, F, Langer, R, Freed, LE &Vunjak-Novakovic, G 2004, ‘Functional assembly of engineered myocardium by electrical stimulation of cardiac myocytes cultured on scaffolds’, Proceeding of the National Academy of Sciences USA, Vol. 100, pp. 18129-18134. Shimizu, T, Yamato, M, Isoi, Y, Akutsu, T, Setomaru, T, Abe, K, Kikuchi, A, Umezu, M & Okano, T 2002, ‘Fabrication of pulsatile cardiac tissue grafts using a novel 3-dimensional cell sheet manipulation technique and temperature-responsive cell culture surfaces’, Circulation Research, Vol. 90, pp. e40-e48. Siepe, M, Akhyar, P, Lichtenberg, A, Schlensak, C & Beyersdorf, F 2008, ‘Stem cells used for cardiovascular tissue engineering’, European Journal of Cardio-thoracic Surgery, Vol. 34, pp. 242-247. Terracio, L, Miller, B & Borg, T 1988, ‘Effects of cyclic mechanical stimulation of the cellular components of the heart: in vitro’, In Vitro Cellular and Developmental Biology, Vol. 24, pp. 53-58. Van Laake, L, Hassink, R, Doevendans, P & Mummery, C 2006, ‘Heart repair and stem cells’, Journal of Physiology, Vol. 577.2, pp. 467-478. Vandenburgh, H, Swasdison, S & Karlisch, P 1991, ‘Computer-aided mechanogenesis of skeletal muscle organs from single cells in vitro’, Faseb Journal, Vol. 5, pp. 2860-2867. Zimmermann, WH, Didie, M, Doker, S, Melnychenko, I, Naito, H, Rogge, C, Tiburcy, M & Eschenhagen, T 2006, ‘Heart muscle engineering: An update on cardiac muscle replacement therapy’, Cardiovascular Research, Vol. 71, pp. 419-429. Zimmermann, WH, Schneiderbanger, K, Schubert, P, Didie, M, Munzel, F, Heubach, JF, Kostin, S, Neuhuber, WL & Eschenhagen, T 2002, ‘Tissue engineering of a differentiated cardiac muscle construct’, Circulation Research, Vol. 90, pp. 223-230. Read More
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