The focus of our research is on engineering functional human tissues, by an integrated use of stem cells, biomaterial scaffolds and bioreactors. Our long-term goals are to engineer tissue grafts for application in regenerative medicine, develop enabling technologies for stem cell research, and design models for controlled studies of tissue development, remodeling and disease. The main areas of our work are:
Cardiac Tissue Engineering
The cardiac myocyte is the most physically energetic cell in the body, contracting more than 3 billion times in an average human lifespan, and pumping over 7,000 liters of blood per day along 100,000 miles of blood vessels. In native heart, oxygen is supplied by diffusion from capillaries that are spaced ~20 µm apart, by blood containing hemoglobin. We are trying to pursue a “biomimetic” approach to cardiac tissue engineering driven by the conditions derived from developmental biology.
To engineer a thick and compact cardiac muscle consisting of viable cells, we culture the cells on porous and elastic scaffolds containing an array of channels perfused with culture medium (to mimic blood flow through the capillary network) which in turn can be supplemented by an oxygen carrier (to mimic the role of hemoglobin). The relationships between oxygen supply and consumption were studied with the aid of mathematical modeling that enabled us to derive quantitative criteria for the design of cardiac tissue engineering systems.
Another factor crucial for the development and function of native myocardium is the orderly coupling between electrical pacing signals and the macroscopic contractions. The application of electrical signals designed to mimic those in the heart and applied to induce synchronous contractions of cultured tissue constructs markedly enhanced functional assembly of the engineered tissue via physiologically relevant mechanisms. We are now extending these studies to the use of human stem cells, with the goal to engineer thick, compact, electro-mechanically functional, and vascularized cardiac tissue. For these experiments, we are developing customized scaffolds and bioreactors with electrical stimulation, medium perfusion and real-time imaging.
Osteochondral Tissue Engineering
A major challenge in engineering grafts with hierarchical structure, composition and mechanical properties starting from a single cell preparation, is the design of an appropriate bioreactor system. In addition to the nutrient transport to cells in clinically sized constructs, a bioreactor should provide lineage-specific biological stimuli in various regions of the graft. For stratified cartilage-bone grafts, the development of mechanical competence necessitates the application of biophysical stimuli, which is technically challenging particularly for anatomically shaped constructs.
Our lab has begun studies to generate mechanically competent, biological grafts (such as the temporomandibular joint) using anatomically correct scaffolds seeded with human mesenchymal stem cells (hMSCs). To accommodate these grafts, we are developing a bioreactor capable of coordinating biological, physiological and mechanical stimuli, and applying them in a spatially and temporally controlled manner to provide lineage-specific stimulation within the cartilage and bone regions. This includes the use of chondro-inductive growth factors and dynamic stimulation to the cartilage regions of the graft, and osteo-inductive factors combined with medium perfusion (for nutrient transfer and shear stress) in the bone phase of the graft. The combined stimuli should be applied while the two phases are being grown in apposition to stimulate osteochondral integration.
Human Stem Cell Research
New tools are becoming available to enable controlled studies of stem cells under the conditions that mimic some aspects of the developmental milieu. Specialized biomaterial scaffolds are of particular relevance to studies of human stem cells. We found that human embryonic stem cells (hESCs) have active binding sites and receptors for hyaluronic acid (HA) that are involved in standard cultures of these cells on feeder layers, and that hESCs are able to internalize and process HA. We then designed a completely synthetic HA hydrogel matrix, polymerizable by light, that supports long-term self-renewal and directed differentiation of hESCs.
In addition to their developmentally relevant chemical composition, HA hydrogels have the advantage that they can be tailored with respect to architecture, stiffness and degradation. We showed that the encapsulated hESCs maintain their undifferentiated state, preserve normal karyotype, and can be induced to differentiate by simply altering soluble factors (Gerecht et al. 2007). Hydrogels thus enable a simple “switch” between the self-renewal and differentiation of hESCs.
To further enhance control of the vascular differentiation of hESCs, we developed hydrogels containing combinations of regulatory factors, such as a tethered RGD peptide and microencapsulated VEGF165 (Fereira 2007). By controlling the growth factor delivery, it was possible to regulate the fractions of cells expressing specific receptors (such as the VEGF receptor KDR/Flk-1) and differentiation markers (such as ectodermal marker including nestin or endodermal marker-fetoprotein). In a related study, we showed that hESCs contain a population of CD34+ vascular progenitor cells that can be selectively differentiated into endothelial and smooth muscle cells, and form vascular networks that were integrated with the host vasculature (Ferreira 2007).
Advanced bioreactors that provide multiple molecular and physical regulatory signals, when required in form of spatial and temporal gradients, are of great interest for hESCs, because of the complexity of their regulatory pathways, and uncontrolled variables associated with traditional culture methods. We recently developed a microbioreactor that is the size of a microscope slide and contains twelve independent chambers perfused with culture medium, for cell culture in two-dimensional and three-dimensional settings (Figallo et al. 2007). The microbioreactor array combines the advantages of simple multi-well plates (small volume, high throughput, independent culture wells) with those of perfusion bioreactors (steady-state conditions, enhanced mass transport, application of physical signals).
To take advantage of imaging compatibility of this device, we developed an automated image analysis routine for fast analysis of nuclear and cytoplasmic differentiation markers. In studies of hESCs in this system, we established correlations between the expression of smooth muscle actin with hydrodynamic shear and cell density. The microbioreactor arrays of this kind could be used to study the effects of culture parameters on hESC differentiation, in a systematic manner and with only a minimal consumption of cells and reagents. We are working on prototypes of microbioreactors that are suitable for screening of cells and culture conditions and possibly for studies of disease models and drug screening.
Tissue Engineering Resource Center
The Tissue Engineering Resource Center (TERC) was initiated in August 2004, with the unifying mission to engineer human tissue systems for medical impact. The goal of the Center is to impact medicine in a number of important ways, including: (a) engineering functional grafts of clinically relevant human tissues for application in regenerative medicine, (b) developing in vitro models of human disease to provide new experimental tools for disease understanding and therapeutic discovery, and (c) establishing high-fidelity bioengineering tools for rigorous cell biology studies in the context of tissue development and regeneration. In addition, the Center will continue to foster service, dissemination, training and collaborative research in support of the scientific and technological community. The core themes for TERC evolve along with the scientific and technological progress. The main focus is on functional tissue engineering by integration of the key elements – cells, scaffolds and bioreactors - via a systems approach. The Center also has interest in several areas of clinical relevance – human stem cells, disease models, and research tools applicable to biological inquiry. We maintain focus in two critical areas: (a) skeletal systems and (b) cardiovascular systems, while progressing toward new fundamental and translational projects. The common aspects for both areas include: cell sources, genetic tools, imaging (molecular, cellular, tissue levels), biomechanics (from cells to tissues), modeling (computational biology, transport, electrical and mechanical signal transduction) and the utilization of animal models. Two long-time collaborators: David Kaplan at Tufts University and Gordana Vunjak-Novakovic at Columbia University, lead the Center and direct the Biomaterials Core (Kaplan) and Bioreactors Core (Vunjak-Novakovic).
Peer Reviewed Articles: 2010
- 249. Maidhof R, Marsano A, Lee EJ, Vunjak-Novakovic G. Perfusion seeding of channeled elastomeric scaffolds with myocytes and endothelial cells for cardiac tissue engineering. Biotechnol Prog. 2010 Jan 5. [Epub ahead of print]. PMID: 20052737.
- 248. Chang G, Kim HJ, Vunjak-Novakovic G, Kaplan DL, Kandel R. Enhancing annulus fibrosus tissue formation in porous silk scaffolds. J Biomed Mater Res A. 2010 92(1):43-51. PMID:19165797.
Peer Reviewed Articles: 2009
- 247. Tandon N, Goh B, Marsano A, Chao P-hG, Montouri-Sorrentino C, Gimble J, Vunjak-Novakovic G. Alignment and Elongation of Human Adipose-Derived Stem Cells in Response to Direct-Current Electrical Stimulation. Conf Proc IEEE Eng Med Biol Soc. 2009 1:6517-21. PMCID:PMC2791914.
- 246. Trkov S, Eng G, di Liddo R, Parnigott PP and Vunjak-Novakovic G. Micropatterned 3D hydrogel system to study endothelial-mesenchymal stem cell interactions. J Tissue Eng Regen Med. 2009 Dec 8. [Epub ahead of print]. PMID:19998330.
- 245. Zhang YS, Nuglozeh E, Toure F, Schmidt AM and Vunjak-Novakovic G. Controllable expansion of primary cardiomyocytes by reversible immortalization. Hum Gene Ther. 2009 20(12):1687-96. PMCID: PMC2794932.
- 244. Grayson WG, Frohlich M,, Yeager K, Bhumiratana S, Cannizzaro C, Wan LQ, Chan ME, Liu ME, X. Edward Guo EX and Vunjak-Novakovic GV. Engineering anatomically shaped human bone grafts. Proc Natl Acad Sci USA. 2009 Oct 9. [Epub ahead of print]. PMID:19820164.
- 243. Ifkovits JL, Devlin JJ, Eng G, Martens TP, Vunjak-Novakovic G and Burdick JA. Photocrosslinked biodegradable fibrous scaffolds with tunable properties for tissue engineering applications. ACS Appl. Mater. Interfaces. 2009 1(9):1878-86. (cover)
- 242. Freytes DO, Wan LQ, Vunjak-Novakovic G. Geometry and force control of cell function. J Cell Biochem. 2009 108(5):1047-58. PMID:19795385. (cover)
- 241. Serena E, Figallo E, Tandon N, Cannizzaro C, Gerecht S, Elvassore N, Vunjak-Novakovic G. Electrical stimulation of human embryonic stem cells: Cardiac differentiation and the generation of reactive oxygen species. Exp Cell Res. 2009 315(20):3611-9. PMCID:PMC2787733.
- 240. Vunjak-Novakovic G, Zhang Y, Nuglozeh E, Toure F, Schmidt AM. Controllable Expansion of Primary Cardiomyocytes by Reversible Immortalization. Hum Gene Ther. 2009 20(12):1687-96. PMCID:PMC2794932.
- 239. Vunjak-Novakovic G, Tandon N, Godier A, Maidhof R, Marsano A, Martens T, Radisic M. Challenges in Cardiac Tissue Engineering. Tissue Eng Part B Rev. 2009 Aug 21. [Epub ahead of print] PMID:19698068.
- 238. Frohlich M, Grayson W, Marolt D, Gimble J, Velikonja NK, Vunjak-Novakovic G. Bone Grafts Engineered from Human Adipose-Derived Stem Cells in Perfusion Bioreactor Culture. Tissue Eng Part A. 2009 Aug 13. [Epub ahead of print] PMID:19678762.
- 237. Grayson WL, Martens TP, Eng GM, Radisic M, Vunjak-Novakovic G. Biomimetic approach to tissue engineering. Semin Cell Dev Biol. 2009 20(6):665-73. PMCID:PMC2710409.
- 236. Lee EJ, Vunjak-Novakovic G, Wang Y, Niklason LE. A biocompatible endothelial cell delivery system for in vitro tissue engineering. Cell Transplant. 2009 Apr9. pii:CT-2052. PMID:19500475.
- 235. Radisic M, Fast VG, Sharifov OF, Iyer RK, Park H, Vunjak-Novakovic G. Optical mapping of impulse propagation in engineered cardiac tissue. Tissue Eng Part A. 2009 15(4):851-60. PMCID:PMC2759871.
- 234. Wang X, Wenk E, Zhang X, Meinel L, Vunjak-Novakovic G, Kaplan DL. Growth factor gradients via microsphere delivery in biopolymer scaffolds for osteochondral tissue engineering. J Control Release. 2009 134(2):81-90. PMCID:PMC2698962.
- 233. Cimetta E, Figallo E, Cannizzaro C, Elvassore N, Vunjak-Novakovic G. Micro-bioreactor arrays for controlling cellular environments: design principles for human embryonic stem cell applications. Methods. 2009 47(2):81-9. PMCID:PMC2744042.
- 232. Burdick JA, Vunjak-Novakovic G. Engineered microenvironments for controlled stem cell differentiation. Tissue Eng Part A. 2009 15(2):205-19. PMCID:PMC2716398.
- 231. Ma T, Grayson WL, Frohlich M, Vunjak-Novakovic G. Hypoxia and stem cell-based engineering of mesenchymal tissues. Biotechnol Prog. 2009 25(1):32-42. PMCID:PMC2771546 (review)
- 230. Martens TP, Godier AF, Parks JJ, Wan LQ, Koeckert MS, Eng GM, Hudson BI, Sherman W, Vunjak-Novakovic G. Percutaneous cell delivery into the heart using hydrogels polymerizing in situ. Cell Transplant. 2009 18(3):297-304. PMCID:PMC2771541.
- 229. Tandon N, Cannizzaro C, Chao PH, Maidhof R, Marsano A, Au HT, Radisic M, Vunjak-Novakovic G. Electrical stimulation systems for cardiac tissue engineering. Nat Protoc. 2009 4(2):155-73. PMCID:PMC2775058.
Peer Reviewed Articles: 2008
- 228. Vunjak-Novakovic G. Engineered tissue grafts - a new class of biomaterials for medical use. Chemical Industry & Chemical Engineering Quarterly. 2008 14(4):211-4
- 227. Lovett ML, Cannizzaro CM, Vunjak-Novakovic G, Kaplan DL. Gel spinning of silk tubes for tissue engineering. Biomaterials. 2008 Dec;29(35):4650-7. PMID:18801570.
- 226. Godier AF, Marolt D, Gerecht S, Tajnsek U, Martens TP, Vunjak-Novakovic G. Engineered microenvironments for human stem cells. Birth Defects Res C Embryo Today. 2008 84(4):335-47. PMID:19067427.
- 225. Frohlich M, Grayson WL, Wan LQ, Marolt D, Drobnic M, Vunjak-Novakovic G. Tissue engineered bone grafts: biological requirements, tissue culture and clinical relevance. Curr Stem Cell Res Ther. 2008 3(4):254-64. PMCID:PMC2773298.
- 224. Pei M, He F, Vunjak-Novakovic G. Synovium-derived stem cell-based chondrogenesis. Differentiation. 2008 76(10):1044-56. PMCID: PMC2772098.
- 223. Lima EG, Grace Chao PH, Ateshian GA, Bal BS, Cook JL, Vunjak-Novakovic G, Hung CT. The effect of devitalized trabecular bone on the formation of osteochondral tissue-engineered constructs. Biomaterials. 2008 29(32):4292-9. PMCID:PMC2562244.
- 222. Grayson WL, Bhumiratana S, Cannizzaro C, Chao PH, Lennon DP, Caplan AI, Vunjak-Novakovic G. Effects of initial seeding density and fluid perfusion rate on formation of tissue-engineered bone. Tissue Eng Part A. 2008 14(11):1809-20. PMCID:PMC2773295.
- 221. Vunjak-Novakovic G. Patterning stem cell differentiation. Cell Stem Cell. 2008 3(4):362-3. PMCID:PMC2773296.
- 220. Radisic M, Park H, Martens TP, Salazar-Lazaro JE, Geng W, Wang Y, Langer R, Freed LE, Vunjak-Novakovic G. Pre-treatment of synthetic elastomeric scaffolds by cardiac fibroblasts improves engineered heart tissue. J Biomed Mater Res A. 2008 86(3):713-24. PMCID:PMC2775086.
- 219. Augst A, Marolt D, Freed LE, Vepari C, Meinel L, Farley M, Fajardo R, Patel N, Gray M, Kaplan DL, Vunjak-Novakovic G. Effects of chondrogenic and osteogenic regulatory factors on composite constructs grown using human mesenchymal stem cells, silk scaffolds and bioreactors. J R Soc Interface. 2008 5(25):929-39. PMCID:PMC2607468.
- 218. Pei M, He F, Kish VL, Vunjak-Novakovic G. Engineering of functional cartilage tissue using stem cells from synovial lining: a preliminary study. Clin Orthop Relat Res. 2008 466(8):1880-9. PMCID:PMC2584267.
- 217. Park H, Bhalla R, Saigal R, Radisic M, Watson N, Langer R, Vunjak-Novakovic G. Effects of electrical stimulation in C2C12 muscle constructs. J Tissue Eng Regen Med. 2008 2(5):279-87. PMCID:PMC2782921.
- 216. Grayson WL, Chao PH, Marolt D, Kaplan DL, Vunjak-Novakovic G. Engineering custom-designed osteochondral tissue grafts. Trends Biotechnol. 2008 26(4):181-9. PMCID: PMC2771165.
- 215. Jakab K, Norotte C, Damon B, Marga F, Neagu A, Besch-Williford CL, Kachurin A, Church KH, Park H, Mironov V, Markwald R, Vunjak-Novakovic G, Forgacs G. Tissue engineering by self-assembly of cells printed into topologically defined structures. Tissue Eng Part A. 2008 14(3):413-21. PMID:18333793.
- 214. Kahn CJ, Vaquette C, Rahouadj R, Wang X. A novel bioreactor for ligament tissue engineering. Biomed Mater Eng. 2008 18(4-5):283-7. PMID:19065035.
- 213. Tandon N, Marsano A, Cannizzaro C, Voldman J, Vunjak-Novakovic G. Design of electrical stimulation bioreactors for cardiac tissue engineering. Conf Proc IEEE Eng Med Biol Soc. 2008 2008:3594-7. PMCID: PMC2771167.
- 212. Marsano A, Maidhof R, Tandon N, Gao J, Wang Y, Vunjak-Novakovic G. Engineering of functional contractile cardiac tissues cultured in a perfusion system. Conf Proc IEEE Eng Med Biol Soc. 2008 2008:3590-3. PMID:19163485.
- 211. Radisic M, Marsano A, Maidhof R, Wang Y, Vunjak-Novakovic G. Cardiac tissue engineering using perfusion bioreactor systems. Nat Protoc. 2008 3(4):719-38. PMCID:PMC2763607.
Peer Reviewed Articles: 2007
- 195. Hoffman S, Hagenmueller H, Koch A, Mueller R, Vunjak-Novakovic G, Kaplan D, Merkle HP and Meinel LW. Control of in vitro tissue-engineered bone-like structures using human mesenchymal stem cells and porous silk scaffolds. Biomaterials 28(6): 1152-1562 (2007)
- 196. Khademhosseini A, Eng G, Yeh J, Kucharczyk S, Langer R, Vunjak-Novakovic G and Radisic M. Microfluidic patterning for fabrication of contractile cardiac organoids. Biomedical Microdevices 9(2): 149-157 (2007)
- 197. Cannizzaro C, Tandon N, Figallo E, Park H, Gerecht S, Radisic M, Elvassore N and Vunjak-Novakovic G. Practical aspects of cardiac tissue engineering with electrical stimulation. Methods in Molecular Medicine, 291-307, 2007.
- 198. Park H, Cannizzaro C, Langer R, Vunjak-Novakovic G, Vacanti CA, Farokhzad OC. Micro- and Nanofabrication of functional materials for tissue engineering (Review article) Tissue Engineering 13(8):1867-1877 (2007)
- 199. Iyer R, Radisic M, Cannizzaro C and Vunjak-Novakovic G. Oxygen carriers in cardiac tissue engineering. Artificial Cells Blood Substitutes and Biotechnology 35 (1): 135-148 (2007)
- 200. Ferreira L, Gerecht-Nir S, Shieh H, Vunjak-Novakovic G and Langer R. Bioactive hydrogel scaffolds for controllable vascular differentiation of human embryonic stem cells. Biomaterials 28(17): 2706-2717 (2007) Highlighted in Materials Today: “Hydrogels make stem cells differentiate” 10(5): 10 (May 2007)
- 201. Ferreira L, Gerecht-Nir S, Shieh H, Vunjak-Novakovic G and Langer R. Vascular progenitor cells isolated from human embryonic stem cells Circulation Research 101(3): 286-294, 2007.
- 202. Radisic M, Park H, Gerecht-Nir S, Cannizzaro C, Langer R., Vunjak-Novakovic G. Biomimetic approach to cardiac tissue engineering. Philosophical Transactions of the Royal Society of London – B Biological Sciences 362(1484):1357-1368 (2007)
- 203. Figallo E., Cannizzaro C, Gerecht-Nir S, Burdick J, Langer R, Elvassore N and Vunjak-Novakovic G. Micro-bioreactor array for controlling cellular environments. Lab on a Chip 7 (6): 710 – 719, 2007, Cover article
- 204. Chang G, Jin HJ, Kaplan DL, Vunjak-Novakovic G and Kandel R. Porous silk scaffolds can be used for tissue engineering annulus fibrosus. European Spine Journal 16(11):1848-1857 (2007)
- 205. Gerecht S, Burdick JA, Ferreira LS, Townsend SA, Langer R. and Vunjak-Novakovic G. Propagation of undifferentiated human embryonic stem cells in hyaluronic acid hydrogels. Proceedings of the National Academy of Sciences USA 104:11298-303, 2007
- 206. Hagenmüller H, Hofmann S, Kohler T, Merkle HP, Kaplan DL, Vunjak-Novakovic G, Müller R and Meinel L. Noninvasive time-lapsed monitoring and quantification of engineered bone-like tissue. Annals of Biomedical Engineering 35(10):1657-1667 (2007)
- 207. Gerecht S, Bettinger CJ, Zhang Z, Borenstein J, Vunjak-Novakovic G, Langer R. The effect of actin disrupting agents on contact guidance of human embryonic stem cells. Biomaterials 28(28):4068-4077 (2007)
- 208. Chao PhG, Grayson W and Vunjak-Novakovic G. Engineering cartilage and bone using human mesenchymal stem cells. Journal of Orthopaedic Science 12(4):398-404 (2007)
- 209. Lovett M, Cannizzaro C, Daheron L, Messmer B, Vunjak-Novakovic G and Kaplan DL. Silk fibroin microtubes for blood vessel engineering. Biomaterials 28(35):5271-5279 (2007)