Biomaterials 21 (2000) 2529}2543 Sca!olds in tissue engineering bone and cartilage Dietmar W. Hutmacher Laboratory for Biomedical Engineering, Institute of Engineering Science, Department of Orthopedic Surgery, National University of Singapore, 10 Kent Ridge Crescent, Singapore 119260, Singapore Abstract Musculoskeletal tissue, bone and cartilage are under extensive investigation in tissue engineering research. A number of biodegradable and bioresorbable materials, as well as sca!old designs, have been experimentally and/or clinically studied. Ideally, a sca!old should have the following characteristics: (i) three-dimensional and highly porous with an interconnected pore network for cell growth and #ow transport of nutrients and metabolic waste (ii) biocompatible and bioresorbable with a controllable degradation and resorption rate to match cell/tissue growth in vitro and/or in vivo (iii) suitable surface chemistry for cell attachment, proliferation, and di!erentation and (iv) mechanical properties to match those of the tissues at the site of implantation. This paper reviews research on the tissue engineering of bone and cartilage from the polymeric sca!old point of view. ( 2000 Elsevier Science Ltd. All rights reserved. Keywords: Tissue engineering of bone and cartilage Design and fabrication of 3-D sca!old Biodegradable and bioresorbable polymers 1. Introduction Bone and cartilage generation by autogenous cell/tis- sue transplantation is one of the most promising tech- niques in orthopedic surgery and biomedical engineering [1]. Treatment concepts based on those techniques would eliminate problems of donor site scarcity, immune rejection and pathogen transfer [2]. Osteoblasts, chon- drocytes and mesenchymal stem cells obtained from the patient's hard and soft tissues can be expanded in culture and seeded onto a sca!old that will slowly degrade and resorb as the tissue structures grow in vitro and/or in vivo [3]. The sca!old or three-dimensional (3-D) con- struct provides the necessary support for cells to prolifer- ate and maintain their di!erentiated function, and its architecture de"nes the ultimate shape of the new bone and cartilage. Several sca!old materials have been inves- tigated for tissue engineering bone and cartilage includ- ing hydroxyapatite (HA), poly(a-hydroxyesters), and natural polymers such as collagen and chitin. Several reviews have been published on the general properties and design features of biodegradable and bioresorbable polymers and sca!olds [4}12]. The aim of this paper is to complete the information collected so far, with special emphasis on the evaluation of the material and design characteristics which are of speci"c interest in tissue engineering the mesenchymal tissues bone and cartilage. The currently applied sca!old fabrication technologies, with special emphasis on the so-called solid-free form fabrication technologies, will also be bench marked. Fi- nally, the paper discusses the author's research on the design and fabrication of 3-D sca!olds for tissue engi- neering an osteochondral transplant. 2. Polymer-based sca4old materials The meaning and de"nition of the words biodegrad- able, bioerodable, bioresorbable and bioabsorbable (Table 1)*which are often used misleadingly in the tissue engineering literature*are of importance to discuss the rationale, function as well as chemical and physical proper- ties of polymer-based sca!olds. In this paper, the polymer properties are based on the de"nitions given by Vert [13]. The tissue engineering program for bone and cartilage in the author's multidisciplinary research curriculum has been classi"ed into six phases (Table 2). Each tissue engineering phase must be understood in an integrated manner across the research program*from the polymer material properties, to the sca!old micro- and macro- architecture, to the cell, to the tissue-engineered trans- plant, to the host tissue. Hence, the research objectives in each phase are cross-disciplinary and the sub-projects are linked horizontally as well as vertically. 0142-9612/00/$- see front matter ( 2000 Elsevier Science Ltd. All rights reserved. PII: S 0 1 4 2 - 9 6 1 2 ( 0 0 ) 0 0 1 2 1 - 6

Table 1 De"nitions given by Vert Biodegradable are solid polymeric materials and devices which break down due to macromolecular degradation with dispersion in vivo but no proof for the elimination from the body (this de"nition excludes environmental, fungi or bacterial degradation). Biodegradable poly- meric systems or devices can be attacked by biological elements so that the integrity of the system, and in some cases but not necessarily, of the macromolecules themselves, is a!ected and gives fragments or other degradation by-products. Such fragments can move away from their site of action but not necessarily from the body. Bioresorbable are solid polymeric materials and devices which show bulk degradation and further resorb in vivo i.e. polymers which are eliminated through natural pathways either because of simple "ltration of degradation by-products or after their metabolization. Bioresorption is thus a concept which re#ects total elimination of the initial foreign material and of bulk degradation by-products (low molecular weight compounds) with no residual side e!ects. The use of the word &bio- resorbable' assumes that elimination is shown conclusively. Bioerodible are solid polymeric materials or devices, which show sur- face degradation and further, resorb in vivo. Bioerosion is thus a con- cept, too, which re#ects total elimination of the initial foreign material and of surface degradation by-products (low molecular weight com- pounds) with no residual side e!ects. Bioabsorbable are solid polymeric materials or devices, which can dissolve in body #uids without any polymer chain cleavage or molecu- lar mass decrease. For example, it is the case of slow dissolution of water-soluble implants in body #uids. A bioabsorbable polymer can be bioresorbable if the dispersed macromolecules are excreted. Table 2 The research program for tissue engineering bone and cartilage classi- "ed into six phases I*Fabrication of bioresorbable sca!old II*Seeding of the osteoblasts/chondrocytes populations into the polymeric sca!old in a static culture (petri dish) III*Growth of premature tissue in a dynamic environment (spinner #ask) IV*Growth of mature tissue in a physiologic environment (bioreactor) V*Surgical transplantation VI*Tissue-engineered transplant assimilation/remodeling The "rst stage of tissue engineering bone or cartilage begins with the design and fabrication of a porous 3-D sca!old, the main topic of this review paper. In general, the sca!old should be fabricated from a highly biocom- patible material which does not have the potential to elicit an immunological or clinically detectable primary or secondary foreign body reaction [9]. Furthermore, a polymer sca!old material has to be chosen that will degrade and resorb at a controlled rate at the same time as the speci"c tissue cells seeded into the 3-D construct attach, spread and increase in quantity (number of cells/per void volume) as well as in quality. Currently, the design and fabrication of sca!olds in tissue engineering research is driven by three material categories: I. Regula- tory approved biodegradable and bioresorbable poly- mers (Table 3), such as collagen, polyglycolide (PGA), polylactides (PLLA, PDLA), polycaprolactone (PCL), etc. II. A number of non-approved polymers, such as polyorthoester (POE), polyanhydrides, etc. which are also under investigation. III. The synthesis of entrepre- neurial polymeric biomaterials, such as poly (lactic acid- co-lysine), etc., which can selectively shepherd speci"c cell phenotypes and guide the di!erentiation and prolifer- ation into the targeted functional premature and/or ma- ture tissue. In general, polymers of the poly(a-hydroxy acids) group undergo bulk degradation. The molecular weight of the polymer commences to decrease on day one (PGA, PDLA) or after a few weeks (PLLA) upon placement in an aqueous media [12]. However, the mass loss does not start until the molecular chains are reduced to a size which allows them to freely di!use out of the polymer matrix [14]. This phenomenon described and analyzed in detail by a number of research groups [15}18], results in accelerated degradation and resorption kinetics until the physical integrity of polymer matrix is compromised. The mass loss is accompanied by a release gradient of acidic by-products. In vivo, massive release of acidic degradation and resorption by-products results in in#ammatory reac- tions, as reported in the bioresorbable device literature [19}22]. If the capacity of the surrounding tissue to eliminate the by-products is low, due to the poor vas- cularization or low metabolic activity, the chemical com- position of the by-products may lead to local temporary disturbances. One example of this is the increase of osmo- tic pressure or pH manifested through local #uid accumulation or transient sinus formation from "ber reinforced polyglycolide pins applied in orthopedic sur- gery [21]. Potential problems of biocompatibility in tissue engineering bone and cartilage, by applying degra- dable, erodable, and resorbable polymer sca!olds, may also be related to biodegradability and bioresorbability. Therefore, it is important that the 3-D sca!old/cell con- struct is exposed at all times to su$cient quantities of neutral culture media, especially during the period where the mass loss of the polymer matrix occurs. The incorporation of a tricalciumphosphate (TCP) [23], hydroxyapatite (HA) [24] and basic salts [15] into a polymer matrix produces a hybrid/composite material. These inorganic "llers allow to tailor the desired degra- dation and resorption kinetics of the polymer matrix. A composite material would also improve biocompatibil- ity and hard tissue integration in a way that ceramic particles, which are embedded into the polymer matrix, allow for increased initial #ash spread of serum proteins compared to the more hydrophobic polymer surface [9]. In addition, the basic resorption products of HA or TCP would bu!er the acidic resorption by-products of the aliphatic polyester and may thereby help to avoid the formation of an unfavorable environment for the cells due to a decreased pH [15,23,24]. 2530 D.W. Hutmacher / Biomaterials 21 (2000) 2529}2543

Table 3 Properties of bioresorbable and bioerodable polymers Polymer Comparison of mechanical properties of bioerodable and bioresorbable polymers Degradation and resorption process via hydrolysis Molecular weight loss/loss of mechanical properties (in month) ! Mass loss (in month) ! References (sca ! olds) References (medical device) Area of application Products with regulatory approval Poly( L -lactide) ### Bulk erosion 9 } 15 36 } 48 46, 48, 49, 60 } 64, 70 } 72 19, 20, 24 Orthopedic Surgery, Oral and Maxillofacial Surgery FixSorb System (screws, nails, pins) Neo " x (screws, nails, pins) Poly( L -lactide-co- D , L -lactide) 70/30 ## Bulk erosion 5 } 6 12 } 18 22, 23 Oral and Maxillofacial Surgery, Orthopedic Surgery ResorPin, Lead " x MacroSorb System (screws and plates, mesh, nails, pins) PolyPin Poly( L -lactide-co- glycolide) 10/90 ## Bulk erosion 1 } 2 3 } 4 28, 63 Suture Periodontal Surgery, Surgery, Vicryl Suture, Vicryl Mesh Polyglycolide ### Bulk erosion 0.5 } 1 3 } 4 6, 30 } 35, 41, 65 Orthopedic Surgery Bio " x Poly( D , L -lactide) # Bulk erosion 1 } 2 5 } 6 6, 31, 34, 49, 60 Poly( D , L -lactide- co-glycolide) 85/15 # Bulk erosion 1 } 2 4 } 5 53 } 56, 60, 70 Poly( D , L -lactide- co-glycolide) 75/25 # Bulk erosion 1 } 2 4 } 5 49, 61 Poly( D , L -lactide- co-glycolide) 50/50 ## Bulk erosion 1 } 2 3 } 4 28 Polycaprolactone # Bulk and surface erosion 9 } 12 24 } 36 29 Drug delivery Capranor Polyorthoester ## Surface erosion 4 } 6 12 } 18 Polyanhydrides ## Surface erosion 4 } 6 12 } 18 59 Animal experiments ! Molecular weight and mass loss vary depending on factors such as chemical structure and composition presence of ionic groups and of side group defect s con " guration of the structure molecular weight and molecular weight distribution (polydispersity) presence of low molecular weight components (monomers, oligomers, solvents, softene rs, drugs, growth factors, etc.) production and manufacturing procedures and their process parameters, implant design, sterilization method, morphology (amorphous versus semi-crystalline, p resence of microstructures and stress within the components), tempering, storage, implant site. ### , good ## , average # , poor. D.W. Hutmacher / Biomaterials 21 (2000) 2529}2543 2531