Molecular engineering of materials for bioreactivity

Current Opinion in Solid State and Materials Science 4 (1999) 381–387

Molecular engineering of materials for bioreactivity Kevin E. Healy a,b , * a

Division of Biological Materials, Northwestern University Medical School, 311 East Chicago Avenue, Ward Building 10 -116, Chicago, IL 60611 -3008, USA b Department of Biomedical Engineering, McCormick School of Engineering and Applied Science, Technological Institute, Evanston, IL 60201, USA Keywords: Molecular engineering; Bioreactivity

1. Introduction New classes of materials are being designed to interact specifically with mammalian cells to control their behavior and subsequently direct the formation of organ specific tissue. When a material comes in contact with biological systems, initial events are dominated by protein adsorption, and platelet, blood and inflammatory cell adhesion. These events constitute what is regarded as the native response to the material and do not represent the optimal behavior between a material and host tissue. A common theme in engineering mammalian cell and tissue behavior is to modify the material to selectively interact with a cell through bimolecular recognition events. Thus, the first step in the process is the rational design of a biomolecular component of the material. This approach to designing materials is commonly referred to as biomimetic engineering of materials. Core hypotheses of biomimetic materials engineering are that monolayer (i.e. one molecular layer) coatings of biologically active peptides affect cell attachment to materials, and that surfaces, linear polymers, or three-dimensional structures modified with these peptides preferentially induce tissue formation consistent with the cell type seeded either on or within the device. In addition to peptide modification, a large variety of biological functions can be built into materials including: solid phase growth factors and cytokines to promote cell differentiation and commitment; enzymes to catalyze reactions; drugs for site-specific delivery; plasmid vectors for cell transfection; and, antibodies for analyte detection or chelation. Finally, synthetic polymers themselves can act as the biomimetic signaling component. Materials modified in a biomimetic manner can be used in sensors, diagnostic assays, drug delivery devices, separation and purification systems, hybrid artificial organs and devices, and either *Tel.: 11-312-503-4735; fax: 11-312-503-2440. E-mail address: [email protected] (K.E. Healy)

medical or dental devices and implants. This review focuses on recent papers (i.e. published during late 1997 through early 1999) that incorporate biomimetic engineering to control the bioreactivity of materials; either solid surfaces, linear polymers, or three-dimensional hydrogels.

2. Biomimetic surface engineering

2.1. Controlling cell behavior with proteins adsorbed to engineered surfaces The adsorption of proteins from either serum-supplemented media or monocomponent solutions to surfaces with well defined chemistry is an attractive approach for studying the influence of surface chemistry on adsorbate function. Webb et al. [1] used organosilanes with post deposition processing to create surfaces that were either terminated by thiol, oxidized thiol, amine, quaternary amine, or methyl groups to assess how these chemistries affected protein adsorption and subsequent fibroblast attachment, spreading, and cytoskeletal organization. Cell attachment on serum (5% v / v) coated surfaces was most pronounced on positively charged amine and quaternary amine surfaces, but the morphology of cells on the modified surfaces was not remarkably different. Of note, were the observations that adsorption of BSA, FN, or LN from monocomponent protein solutions led to differential cell adhesion depending on whether the protein was adsorbed on either thiol or oxidized thiol-terminated surfaces. Although conclusions regarding how these surfaces affect protein deposition were preliminary, the findings indicate that a single protein can have differential effects on cell behavior depending on substrate surface chemistry. Scotchford et al. [2] adsorbed octanethiol and 3-mercaptopropanoic acid (carboxylic acid terminated) to gold surfaces and assessed osteoblast growth to these chemistries in serum-containing (10% v / v) conditions.

1359-0286 / 99 / $ – see front matter  1999 Elsevier Science Ltd. All rights reserved. PII: S1359-0286( 99 )00038-8

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After one day, cell adhesion was an order of magnitude greater on carboxylic acid-terminated surfaces compared to the methyl-terminated alkanethiol, thus supporting the concept of differential cell behavior, in serum containing environments, based on material surface chemistry. A consistent observation appears to be that methyl-terminated surfaces show minimal cell adhesion in serum-containing environments; [1,2] however, a universal theory regarding cell adhesion to a surface remains a challenge, in part, due to the disparate observations with different cell types. For example, Jenny et al. [3] synthesized 14 different organosilane overlayers (e.g. amine-, hydroxyl-, 13F-terminated, etc.) on glass substrates to test the effect of surface chemistry on human peripheral blood monocyte adhesion, motility, and macrophage fusion. In contrast to observations with connective tissue cell types, monocyte / macrophage adhesion and motility was unaffected by the surface chemistry (10% v / v autologous serum), with the exception of low cell adhesion or proliferation on methyl-terminated surfaces. Foreign body giant cell formation correlated to the carbon content in the silane layer, as determined by XPS. The data led to the interpretation that either surface coverage or silane thickness positively influenced the formation of macrophage fusion into foreign body giant cells. Tang et al. [4] took a similar approach using selfassembled thiol-linked molecules on gold coated poly(ethylene terephthalate) films. Ellipsometry was used to confirm that the self-assembled thiol-terminated molecules formed monolayers upon physisorption to gold. Both in vitro and more importantly in vivo experiments supported the hypothesis that hydroxyl terminated surfaces, such as those prepared from mercaptoglycerol or mercaptoethanol, elicited the strongest inflammatory responses. The accumulation of neutrophils and monocytes / macrophages during this inflammatory response was dependent on the activation of the complement system. Surfaces terminating in either amine or carboxylic acid functionalities (e.g. Lcysteine, glutathione) neither initiated complement activation nor an appreciable inflammatory response. The authors correctly point out that their in vivo observations must be interpreted with caution, since the stability of self-assembled monolayers based on gold-thiol chemistry has not been determined in vivo. Collectively, these observations regarding cell behavior on engineered material surfaces with adsorbed protein, suggest one must consider the unique nature of the cell type employed in a specific study, and that a unified ‘theory’ regarding cell behavior on engineered surfaces may not be attainable.

2.2. Peptide modified surfaces to control cell behavior Modification of material surfaces with peptides from ECM proteins is a very active area of research and offers the potential to control the behavior of mammalian cells in contact with the surface of either a device or implant. Tong and Shoichet [*5] modified the surface of poly(tetrafluoro-

ethylene-co-hexafluoropropylene) (FEP) with peptides designed to mimic the ECM found in peripheral nerves to promote neurite extension of central nervous system neurons. Peptides such as GYIGSR, GRGDS, and SIKVAV were covalently grafted to FEP by first introducing nitrogen functionality via vapor phase mercury photosensitization with ammonia, and then coupling the peptides to the amine on the FEP through either an amine or thiol on the peptides using standard procedures with either tresyl chloride or bifunctional crosslinkers. Significant observations included confirmation of amine derivatization and peptide immobilization by surface analytical techniques (e.g. X-ray photoelectron spectroscopy, XPS), and enhanced hippocampal neuron interaction with FEP modified with GYIGSR. This work has significant impact, since FEP is a clinically relevant material and the peptide modification directly affected the number of neurites extending from each neuron. Ultimately, modification of FEP with selected peptides could affect the development of grafts for nerve regeneration. Peptides designed to promote the attachment of osteogenic cells and foster their differentiation have been covalently grafted to various materials with a wide range of coupling strategies. A number of studies have addressed coupling peptides to optically transparent materials (e.g. glass or quartz) to study the mechanisms of interaction between osteogenic cells and the peptides. Rezania et al. [6] covalently modified the surfaces of metal oxides (e.g. quartz, silicon, or titanium) with peptides that mimic the cell binding (–RGD–) and heparin binding domains found within bone sialoprotein (BSP). Peptide domains of BSP were selected based on the hypothesis that the capacity for specific attachment of osteoblasts to the bone extracellular matrix resides in enzymatically generated fragments of BSP, and that peptide ligands containing attachment signals within BSP can modulate osteoblast attachment and differentiation. The immobilization strategy involved coupling of an aminofunctional organosilane (N-(2-aminoethyl)-3-aminopropyl-trimethoxysilane) to either quartz or metal oxide surfaces and derivatizing the terminal amine to a maleimide functional surface by coupling a heterobifunctional crosslinker (4-(N-maleimidomethyl) cyclohexane-1carboxylate). The maleimide terminated surfaces were then used to immobilize a peptide from the cell-binding domain of bone sialoprotein, CGGNGEPRGDTYRAY, via the thiol group on the terminal cysteinyl residue in the peptide [6,7]. This methodology ensured that the molecule could freely interact with the cell-surface receptors and that the orientation of the coupled peptide was known with respect to the substrate. Surface characterization by spectroscopic ellipsometry and XPS confirmed the chemistry and monolayer deposition of the peptides at densities of |4–6 pmol / cm 2 . A similar three step method of coupling RGD-containing peptides to sputter deposited titanium films yielded an order of magnitude increase in peptide surface density, |43 to 61 pmol / cm 2 [**8]. The strength of adhesion, spread-

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ing, and focal contact formation of primary bone-derived cells seeded on the RGD modified surfaces prepared by Rezania et al. [6,7] was significantly greater than control surfaces containing peptides where Asp (D) was replaced by Glu (E). To build upon these results, Rezania et al. [**9] designed surfaces that contained a consensus peptide sequence of the heparin-binding domain, FHRRIKA, found within BSP. The FHRRIKA peptide consisted of a segment that included clusters of basic and hydrophobic amino acids to form the putative heparin-binding consensus sequence, XBBXBX [where X5hydrophobic and B5 basic]. Mimetic peptide surfaces (MPS) were made as homogenous layers and in mixed layers with the RGD: FHRRIKA peptides in ratios of 25:75 (MPS I), 75:25 (MPS II), and 50:50 (MPS III). The degree of rat calvaria osteoblast-like cell spreading, focal contact formation, cytoskeletal organization, proliferation, and mineralization of the extracellular matrix (ECM) on model biomaterial surfaces was examined. Although osteoblasts were more strongly attached to MPS II surfaces, the degree of cell proliferation on the peptide surfaces were not significantly different from each other. However, areas of mineralized ECM formed on MPS II and III surfaces were significantly larger than other surfaces, indicating that peptide sequences incorporating both cell- and heparin-adhesive motifs enhanced the degree of cell surface interactions, and influenced the long term formation of a mineralized ECM in vitro. Dee et al. [10] also designed peptides exploiting the heparin-binding amino acid consensus sequence. They immobilized various peptides based on the KRSR sequence on glass via amino-functionalization of the surface with 3-aminopropyltriethoxysilane and coupling the peptide via carbodiimide chemistry. KRSR peptides, immobilized at a density of |80 pM / cm 2 , slightly enhanced osteoblasts adhesion to the surface compared to either fibroblasts or endothelial cells. Grafting of these peptides [6,7,**9,10] to implant surfaces or into the backbone of novel polymers could preferentially promote osteoblast attachment and differentiation, thereby increasing the rate of integration of the implant with the surrounding tissue. The power of this approach was demonstrated by Fujisawa et al. [11], who investigated the attachment of osteoblasts to hydroxyapatite crystals coated by a synthetic peptide of E 7 PRGDT. The poly-Glu sequence was used for assembly of the peptide on the hydroxyapatite, since phosphorylated amino acids and g-carboxyglutamic acid are known binding sites on proteins for hydroxyapatite. The facile deposition from buffer onto HA was successful due to the good affinity between the peptide and HA, 13.5 mM. Attachment and spreading of the osteoblast cell line (MC3T3-E1) was specifically dependent on the adsorbed peptide. In the studies by Rezania et al. [7,**9], Dee et al. [10], and Fujisawa et al. [11] the specific nature of the interaction of the cells with the peptide modified surface was confirmed by soluble peptide inhibition experiments. Clearly cell behavior at surfaces can be controlled in vitro by selective

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immobilization of peptides; however, one drawback to the aforementioned studies was that the biological performance evaluation was initially conducted in either serumfree or defined media conditions and the results may not translate to complex biological environments without alteration of the surface chemistry of the base material to prevent nonspecific protein binding and materials fouling. As pointed out by Xiao et al. [**8], another limitation of silane-based coupling reactions is the hydrolysis of the siloxane films during exposure to water, which could limit the use of this chemistry either in vivo or in long-term cultures in vitro.

2.3. Cell adhesion and morphology on peptide modified non-fouling surfaces In an effort to exploit peptide modification of surfaces in complex biological environments containing proteins and lipids (e.g. in vivo), it is essential to prevent non-specific adsorption of these macromolecules so that the designed modification is not rendered inactive. Thus, since the work of Drumheller, Elbert, and Hubbell [12] materials scientists have been designing and synthesizing surfaces that first resist protein adsorption and fouling, and then build back in the chemistry to promote specific cellular interactions. To passivate the surface, most recent studies exploit hydrophilic PEG in some capacity due to its demonstrated low protein, cell, and bacterial binding characteristics. PEG surfaces do not adsorb proteins because of the unique way that PEG molecules bind water, which forms a protective hydration shell around the PEG molecules. When any type of molecule attempts to adsorb to a PEG rich area, the energetic result is a negative entropic contribution due to the confinement of the PEG chains. Thus, the overall relevance to the system is that the adsorption event is improbable, making subsequent cell and bacteria adhesion unlikely. Hern and Hubbell [**13] incorporated RGD-containing peptides into poly(ethylene glycol)-based hydrogels to control wound healing and tissue formation with these in situ polymerizable materials. Peptide-modified networks were synthesized from PEG diacrylate (8000 MW PEG) and acrylamidoyl peptides that were synthesized from either the N-hydroxysuccinimide ester of acrylic acid, no spacer arm monomer, or from acryloyl-PEG-N-hydroxy succinimide ester to introduce a 3400 MW PEG spacer arm. Mixtures of the PEG diacrylate and acrylamidoyl peptides were polymerized via photoinitiation from an aqueous buffer (PBS). A significant finding was that without the 3400 MW PEG spacer, the effect of peptide on human foreskin fibroblast spreading was nonspecific; however, specificity was introduced by incorporation of the PEG spacer. It is envisioned that these materials will ultimately find use in tissue resurfacing and promoting the re-endothelialization of the blood vessel surface after balloon angioplasty treatment.

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Bearinger, Castner, and Healy [*14] developed a surface interpenetrating polymer network (IPN) based on PEG chemistry designed to prevent non-specific protein adsorption and promote cell attachment specifically dependent on peptide signals conjugated to the IPN. These IPNs were thin films (| 20nm) covalently grafted to either metal oxides, silicon, quartz, or glass, and were synthesized by photoinitiated free-radical polymerization. IPNs synthesized from acrylamide (AAm), PEG, and acrylic acid (AAc) [poly(AAm-co-EG /AAc)] were further modified to promote cell attachment by tethering RGD-containing peptides via a bis-amino PEG (3400 MW) spacer that was covalently linked to the acrylic acid sites within the IPN. Peptides were linked to the free amine on the bis-amino PEG via chemistry addressed previously [6]. Surface characterization by XPS supported the model of an IPN and presentation of the peptide at the near surface region. In culture conditions containing fetal bovine serum, rat calvarial osteoblast adhesion, proliferation, and formation of mineralized tissue was specifically dependent on the presence of the RGD peptide. Similar to the work of Hern and Hubbell [**13], the RGE signal enhanced cell adhesion in the presence of serum, presumably due to protein adsorption to this peptide sequence, but this level of cell adhesion was well below that observed with RGD peptide grafted surfaces regardless of the culture conditions. This peptide-IPN is intended as a coating to improve the initial wound healing and stability of metallic and polymeric implants.

2.4. The use of self-assembly in creating peptide modified non-fouling surfaces Neff et al. [**15] have modified the commercially available PluronicE F108 triblock copolymer of (PEO) 129 –(PPO) 56 –(PEO) 129 with GRGDSY peptides to create a surfactant that easily coats the surfaces of hydrophobic materials. The terminal hydroxyls on PEO / PPO / PEO triblocks were converted to sulfhydryl groups and then physisorbed to tissue culture polystyrene (used for cell culture dishes). Peptides modified with heterobifunctional crosslinkers such as N-Succinimidyl 3-(2-pyridyldithio)propionate (SPDP) were covalently linked to the activated physisorbed PEO / PPO / PEO triblocks. Fibroblasts seeded on TCPS with physisorbed activated PEO / PPO / PEO triblocks resisted cell adhesion, whereas coupling GRGDSY peptides to these surfaces significantly increased the number of cells attached and the degree of spreading of the attached cells. Cell spreading was an indication that bimolecular interaction between the surface bound ligand (GRGDSY) and cell surface receptors occurred. A variation on the theme of using PEG to passivate the surface prior to peptide immobilization has been the use of stable lipid films on solid substrates as the platform for peptide modification. Fields et al. [**16] developed a series of peptide-amphiphiles with triple-helical motifs that

promoted the adhesion of a model cell type (human melanoma) to standard TCPS. The peptide-amphiphiles were simply physisorbed to the TCPS from a static buffer solution overnight. Collagen-like peptide-amphiphiles were created by combining lipids of various chain length with a peptide sequence, [IV-H1], known to promote adhesion and spreading of melanoma cells. Peptide-amphiphiles with either a C14 or C16 monoalkyl tail and a (Gly-ProHyp) 4 -[IV-H1]-(Gly-Pro-Hyp) 4 head were used in the cell adhesion studies. Most interesting was that ligand-receptor engagement promoted intracellular signal transduction as indicated by the phosphorylation of focal adhesion kinase (p125 FAK ), and that this event was dependent, and increased in duration and level based on the triple-helical conformation of the peptide amphiphile. Both the selfassembling peptide systems developed by Neff et al. [**15] and Fields et al. [**16] have significant promise as biomimetic coatings for hydrophobic materials.

3. Modification of polymers for biospecific molecular recognition

3.1. Random polymers capable of molecular recognition A very exciting twist on the theme of grafting biological ligands to polymers to create sites for molecular recognition is the concept that polymers randomly substituted with specific functional groups can also have biospecific interactions. This concept was proposed some time ago by M. Jozefowicz and J. Jozefonvicz and an elegant review of their work was recently published [17]. The impact of this concept is most notable by recent contributions addressing the interaction of chemically functionalized dextran (FDx) with human endothelial cells [**18]. Cansell et al. [**18] synthesized a series of dextrans modified with carboxymethyl, benzylamide, and sulfonate groups that were then chemically grafted to cholesterol and used to synthesize FDx-functionalized liposomes. Only FDx-functionalized liposomes were preferentially incorporated into the endothelial cells, thus supporting the concept that FDx polymers engage in molecular recognition with receptors on human endothelial cells. These materials may find use in applications that require targeting of endothelial cells for site specific drug delivery.

3.2. Polymers that promote cell aggregation Belcheva et al. [19] synthesized a series of polymerpeptide systems to test the hypothesis that cell aggregation can be controlled by means of multi-peptide-polymer conjugates. Four different classes of PEG-peptide conjugates were synthesized: di-peptide-PEGs; tetra-peptidePEGs; statistical distribution of peptides on a PAAc backbone; and, dendrimer-PEG-(multi)-peptide polymers. In general, the synthesis was performed in organic media

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and exploited condensation reactions with the aid of carbodiimide chemistry in the presence of N-hydroxysuccinimidyl active esters. Intermediate and final products were purified and characterized by spectral and chromatographic methods. As judged by the chromatographic and spectral characterization (IR, GPC) the synthesis generated a family of peptide-polymer conjugates that promote the aggregation of cells in vitro or potentially in vivo.

3.3. Protein conjugates Reversible soluble–insoluble polymer-enzyme conjugates have been synthesized to improve process recovery of both product and enzymes for enzymatically-catalyzed reactions [20]. The thermo-reversible polymer, poly(Nisopropylacrylamide), poly(NIPAAm), was conjugated to trypsin via modification of a mono carboxyl-terminated poly(NIPAAm) synthesized via chain transfer polymerization. The carboxyl end group was activated through carbodiimide chemistry and reacted with the enzyme, trypsin, to create the conjugates. Conjugation of the enzyme to the polymer increased its stability, but decreased the enzyme’s longevity. The thermal recycling process led to greater than 95% recovery of the enzyme after over 10 cycles through the lower critical solution temperature phase separation (e.g. precipitation).

4. Biomimetic engineering of synthetic extracellular matrices

4.1. Synthesis of peptide or growth factor conjugated hydrogels The ECM plays an important role in providing signals for cell adhesion, migration, proliferation, and differentiation. Signals for adhesion are provided as peptide domains, as previously discussed, and signals for differentiation are presented as matrix bound growth or morphogenetic factors. Synthesis of artificial ECMs for tissue regeneration may incorporate either of these signals, or both. Borkenhagen et al. [*21] covalently linked laminin-derived oligopeptides to agarose gels to create an artificial ECM that would promote nerve regeneration. The oligopeptides CDPGYIGSR, CDPGYIGSK, and CDPGRGSYI were coupled to agarose either by activation with 1,1-carbonyldiimidazole or via photoimmobilization with benzophenone. The combination of photoimmobilization with a spatially programmable laser enabled unique pattern formation of the peptide signal. The extension of nerve cell processes, neurite outgrowth, from dorsal root ganglia was specifically dependent on the active peptide, CDPGYIGSR, within the gel, and was confirmed by peptide inhibition experiments. This observation was independent of the synthetic route. Of great interest was the ability of the CDPGYIGSR-derivatized gel to enhance

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nerve regeneration in vivo in a transected rat dorsal root animal model compared to unmodified agarose and agarose modified with the scrambled control sequence, CDPGRGSYI. This study is of critical importance since it clearly demonstrated that a base material that resists significant protein adsorption (agarose), but also contains ligands for specific cellular receptors, can be effective in vivo. Grzesiak et al. [22] covalently modified collagen / glycosaminoglycan (GAG) matrices with a RGD peptide for applications in dermal regeneration. Endothelial cell and keratinocyte attachment and spreading on the RGDcollagen / GAG matrices in vitro were dependent on the peptide signal present, and antibody and peptide inhibition studies confirmed this observation. Although the results of this work are exciting, a potential limitation of the study was the use of collagen, which presents the cells with myriad signals and theoretically makes it difficult to control cell behavior in vivo. In addition to peptide signals, one can incorporate growth factors into the solid phase of the artificial ECM. Tabata et al. [23] created biodegradable hydrogels composed of gelatin (collagen) processed to have an isoelectric point of 4.9 which was used to ionically complex basic fibroblast growth factor (bFGF). Due to short half-life of growth factors in vivo, it is desirous to bind growth factors to a solid matrix to resist protease attack and increase the active half-life. Tabata et al. [23] employed bovine collagen from bone as the basis of their hydrogel and converted the amide groups on the pendant chain of the amino acids to carboxyl groups to control the isoelectric point. Gelatin hydrogels were synthesized by cross-linking the modified collagen with glutaraldehyde, which controlled the water content of the gels. bFGF was complexed to freeze dried, sterilized, hydrogels in doses of 2 to 200 mg per gel used to repair a cranial bone defect. Enhanced bone regeneration within calvarial defects in rabbit skulls was associated with bFGF complexation and low water content of the hydrogel. The results were interpreted as the lower water content gels slowed collagen degradation, thereby limiting protease access to the bFGF and improving bone regeneration. The use of collagen here is, again, a potential limitation of this work, where the matrix material can bind various proteins making control of specific events challenging in vivo. In addition, the transport of oxygen to the cells in the gel would require significant water content, perhaps as high as 80%; [24] thus, eventually an optimization study of the effect of water content of the gel on both growth factor stability and cell viability must be addressed. Recently, West and Hubbell [**25] developed telechelic water-soluble triblock copolymers of oligopeptide-PEGoligopeptide (ABA) capped at each end with reactive acrylate groups. The oligopeptides introduced were sites for protease (e.g. collagenase, plasmin) attack, so when acrylated macromers were synthesized into hydrogels the degradation was enzyme specific. Although data regarding

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exposure of cells to the gels were not presented, theoretically their degradation can be controlled by the oligopeptide sequence’s (e.g. Ala-Pro-Gly-Leu, or Val-Arg-Asn) effect on enzyme kinetic properties. Clearly, a combination of the approaches used by West and Hubbell [**25] and Borkenhagen et al. [*21] would lead to a synthetic hydrogel capable of bimolecular interaction between cell surface receptors and peptides, and controlled degradability by the protease susceptible crosslinks. The clinical impact of this type of material would be important in attempts to regenerate numerous tissues both in vitro and in vivo.

5. Summary and future directions Design strategies for creating biomimetic materials that direct the interaction with biological systems, such as the formation of tissue surrounding implants or regeneration within artificial matrices, has been a prolific area of research. Results from the studies addressed in this review suggest that biomolecular modification of materials is a potentially fruitful approach to take. However, many problems remain. For example, it is unlikely that materials modified only with the ubiquitous linear RGD signal will lead to controlled responses of a specific cell type in complex environments (e.g. in vivo). In addition, it is clear that any biomimetic modification can be significantly enhanced by first providing a non-protein binding surface for grafting. However, grafting a non-fouling surface to many commonly used materials remains a significant challenge. In addition, it is also unlikely that thiolate or silane chemistry will be robust enough to withstand conditions in vitro or in vivo for long term applications. In the short term, direct conjugation of proteins (e.g. enzymes, growth factors, etc.) to polymers seems to be the most promising approach for biotechnological applications ex vivo. Thus, although biomimetic modification of materials is quite promising as an approach to control the behavior of cells with materials, the aforementioned challenges and potentially new ones must be addressed prior to widespread clinical or industrial use.

Abbreviations AAc bFGF BSA BSP CDPGYIGSR / K CDPGRGSYI E 7 PRGDT

acrylic acid basic fibroblast growth factor bovine serum albumin bone sialoprotein amino acid sequence Cys-Asp-ProGly-Tyr-Ile-Gly-Ser-Arg / Lys amino acid sequence Cys-Asp-ProGly-Arg-Gly-Ser-Tyr-Ile amino acid sequence Glu 7 -Pro-ArgGly-Asp-Thr

FDx FEP

functionalized dextran Poly(tetrafluoroethylene-co-hexafluoropropylene) FHRRIKA amino acid sequence Phe-His-ArgArg-Ile-Lys-Ala FN fibronectin GRGDS / Y amino acid sequence Gly-Arg-GlyAsp-Asp-Ser / Try, also RGD GYIGSR amino acid sequence Gly-Tyr-IleGly-Ser-Arg IPN interpenetrating polymer network [IV-H1] amino acid sequence Gly-Val-LysGly-Asp-Lys-Gly-Asn-Pro-Gly-TrpPro-Gly-Trp-Pro-Gly-Ala-Pro KRSR amino acid sequence Lys-Arg-SerArg LN laminin NGEPRGDTYRAY amino acid sequence Asn-Gly-GluPro-Arg-Gly-Asp-Thr-Tyr-Arg-AlaTyr poly(NIPAAm) poly(N-isopropylacrylamide) PEG Poly(ethylene glycol), same chemistry as PEO PEO Poly(ethylene oxide), same chemistry as PEG PPO Poly(propylene oxide) REDRV amino acid sequence Arg-Glu-AspArg-Val SIKVAV amino acid sequence Ser-Ile-LysVal-Ala-Val TCPS tissue culture polystyrene XPS X-ray photoelectron spectroscopy

References Papers of particular interest, published within the annual period of review, have been highlighted as: * of special interest; ** of outstanding interest. [1] Webb K, Hlady V, Tresco PA. Relative importance of surface wettability and charged functional groups on NIH 3T3 fibroblast attachment, spreading, and cytoskeletal organization. J Biomed Mater Res 1998;41:422–30. [2] Scotchford CA, Cooper E, Leggett GJ, Downes S. Growth of human osteoblast-like cells on alkanethiol on gold self-assembled monolayers: the effect of surface chemistry. J Biomed Mater Res 1998;41:431–42. [3] Jenny CR, DeFife KM, Colton E, Anderson JM. Human monocyte / macrophage adhesion, macrophage motility, and IL-4 induced foreign body giant cell formation on silane-modified surfaces in vitro. J Biomed Mater Res 1998;41:171–82. [4] Tang L, Liu L, Elwing HB. Complement activation and inflammation triggered by model biomaterial surfaces. J Biomed Mater Res 1998;41:333–40. [*5] Tong YW, Shoichet MS. Enhancing the interaction of central nervous system neurons with poly(tetrafluoroethylene-co-hexa-

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