Effects of gamma-irradiation on physical and biologic properties of crosslinked hyaluronan tissue engineering scaffolds

Hydrogels containing divinyl sulfone (DVS)-crosslinked hyaluronan (HA) (hylans) are potentially useful implant biomaterials because of their non-cytotoxicity and -antigenicity. However, to successfully fulfill their intended role in vivo, their properties (e.g., mechanics, pore size, surface topography, hydrophilicity, swelling) must be modulated to match the demands of the target application. This study explored whether controlled irradiation with gamma (gamma) can strengthen hylans and modulate their physical and biologic properties, as has previously been shown to be possible with other natural and synthetic polymers. Hydrated hylans containing two different amounts of DVS were irradiated in vacuum to increasing doses of gamma (0-13.5 kGy). The properties of the irradiated gels were compared with those of non-irradiated controls. Changes to bulk structure were evaluated using swelling tests, surface topography and pore structure were evaluated using scanning electron microscopy, mechanics were evaluated using unconfined compression tests, and surface hydrophilicity was evaluated by measuring contact angle changes. Irradiated gels exhibited lower swelling capacity, structural weakening, increase in elasticity, surface texturing, increased pore size, and decreased surface hydrophilicity in direct correlation with received dose. Cells adhered and proliferated readily on the irradiated gel surfaces but not on control gels. The irradiated gels, however, deteriorated during long-term (<60 days) storage. Irradiation of hylans in a lyophilized state instead resulted in gels that were more compact, swelled less, and exhibited smaller pores than their hydrated counterparts. The results show that gamma-irradiation, although useful to modulate hylan gel properties, presents challenges of degradation that may be associated with its generation of free-radicals, HA chain fragmentation, and disruption of DVS crosslinks, particularly when the gels are irradiated in their native hydrated state (>98% water content). Future studies will optimize parameters for gamma-mediated modulation of hylan properties through irradiation under water-free conditions.     

Smooth muscle cell adhesion on crosslinked hyaluronan gels

Hyaluronic acid (HA)-based polymers (hylans) are highly biocompatible and can be structurally modified to obtain desired mechanical properties. This study evaluated divinyl sulfone-crosslinked solid and particulate hylans as cellular scaffolds. These two hylan types differ in surface characteristics, mode of preparation, HA content, and extent of crosslinking. Neonatal rat aortic smooth muscle cells were cultured on hylan gels coated with matrix factors including collagen I, ECM gel, laminin, and fibronectin and on uncoated controls for < or =4 weeks. Cell attachment was sparse on uncoated controls but significantly enhanced on coated gels. Cell morphology was influenced by the identity of the matrix factors coated and the surface topography of the hylan gels. Cells attached to coated particulate gels appeared either highly spread (collagen, fibronectin) or irregularly shaped (ECM gel, laminin). Cells on laminin and fibronectin-coated solid gels were rounded and nonproliferative. Cells proliferated most rapidly on ECM gel-coated gels. The uneven surface of particulate gels induced more protein deposition and the subsequent attachment and active proliferation of cells. This study shows that surface texturizing and subsequent surface treatment with matrix factors enhances cell attachment and proliferation of hylans. These results are useful toward developing bioengineered materials based on cell-hylan composites.     

In vitro hemocompatibility testing of UV-modified hyaluronan hydrogels

Hydrogels (hylans) based on cross-linked hyaluronan (HA) are potentially good biomaterials for vascular tissue engineering applications because they are highly non-antigenic and -immunogenic. To facilitate surface endothelialization, vital to vascular deployment, we irradiated the gel surface with low wavelength UV light. This process micro-textures the smooth gel surface to provide sites for cell anchorage and causes limited scission of native long-chain HA yielding smaller fragments that elicit an enhanced cell response. In the current in vitro study, we assessed the effects of UV irradiation on the short-term (<45 min) interaction between hylan gels and human blood cells (RBCs, platelets) and coagulation proteins at physiologic temperature. Although the lowered hydrophilicity of irradiated (UV) hylans elicited greater vascular cell response relative to unmodified (U) hylans, platelet deposition was unaffected and much lower compared to collagen-coated glass controls. The adhered platelets were rounded or mildly pseudopodic and did not express p-selectin, an activation marker. Both gel types induced identical, and minimal platelet release as measured using an platelet factor 4 ELISA, and identically deferred the intrinsic and extrinsic coagulation pathways. Both gel types induced elevated levels of contact activation of bound, but not plasma-phase factor XII relative to controls. Hemolysis rates were also identical and within accepted standards. We conclude that UV-treatment of hylans, useful to improve surface endothelialization, does not compromise their short-term hemocompatibility, vital to their use as vascular implant materials.     

A surface-tethered model to assess size-specific effects of hyaluronan (HA) on endothelial cells

Crosslinked gels (hylans) containing long-chain (MW>1 x 10(6)Da) hyaluronan (HA), a connective tissue GAG, show exceptional biocompatibility for vascular implantation but poorly interact with vascular endothelial cells (ECs). Previous studies showed in situ fragmentation of HA by UV light to bioactivate hylan gels and elicit enhanced EC responses. Since fragmented HA can be pro-inflammatory, it is important to define an optimal size distribution of HA fragments on the hylan surface that will recruit and support normally functional ECs and limit ulterior responses. Related studies have shown that exogenous models of HA do not necessarily replicate cell responses to HA scaffolds. Since scaffolds cannot be created based on fragmented HA alone, we sought to determine size-specific responses of ECs to HA substrates of defined fragment sizes by creation of HA-tethered culture surfaces. HA (1000, 200, 20 kDa) and an oligomer mixture were tethered onto an aminosilane (APTMS)-treated glass surfaces using a carbodiimide reaction. MALDI-TOF showed the HA digests to contain HA 4-8mers with a 75+/-0.4% w/w of 4mers. Immuno-fluorescence, SEM, AFM and XPS analysis revealed homogeneous amine and HA surfaces. An amine s-SDTB assay and HA fluorophore-assisted carbohydrate electrophoresis (FACE) indicated surface densities of 9+/-3 amine groups/nm(2) and 0.57+/-0.44 microg/cm(2), respectively. HA/HA fragments/oligomers were stable over 21 days of incubation in serum-free culture media. EC proliferation on these surfaces resulted was limited, a possible effect of smooth surface topography, high anionicity, and in case of 4mers, non-interaction with primary HA cell-surface receptors (CD44). This work is significant in that it allows testing of cell responses to substrates composed of single-sized fragments of HA that cannot by themselves be cross-linked into a gel. Future work in our lab will use this model to assess the effects of other HA oligomer sizes on EC behavior.     

Study on Chemical Cross-linking Modification of Hyaluronan and the Biocompatibility of its Derivatives

Objective: Prepare cross-linked HA gels with higher mechanical stability,lower degradation velocity and desirable biocompatibility,so as to extend the usage of HA.Method: 1.Test molecular weight of HA (MrHA) by viscosimetry;2.Prepare cross-linked HA gels by DVS,GTA,DEC;3.Discuss the cross-linking and degradation procedure;4,evaluate the biocompatibility of the best HA gels.Results: The mechanical stability and durability to degradation of HA-DVS gels are superior to those of other gels,and when HA :DVS = 40:1 (g/g),at 35℃ and in 0.2M NaOH solution,the HA-DVS gel shows the best mechanical stability,and its cytotoxicity reaches class I,hemolysis ratio is lower than 5 %.Conclusion:Our HADVS gel can be used to prepare biologic scaffolds.     

Evaluation of the matrix-synthesis potential of crosslinked hyaluronan gels for tissue engineering of aortic heart valves

Our goal is to fabricate continuous sheets of elastin atop non-biodegradable hydrogels (hylans) containing crosslinked hyaluronan, a glycosaminoglycan. Such elastin-hyaluronan composites may be useful to tissue engineer replacements for the glycosaminoglycan- and elastin-rich layers of the native aortic valve cusp. Neonatal rat aortic smooth muscle cells were cultured atop hylan gels with micro-textured surfaces, and on plastic, and the components of the extracellular matrix (collagen, elastin) were periodically analyzed. The hylan substrates induced the cells to proliferate more rapidly and over longer time periods (approximately 4 weeks) relative to those cultured on plastic (2-3 weeks). Consequently, at all assay times, the amounts of elastin was derived from the hylan-based cell cultures was 25% or more than that derived from cells cultured on plastic. However, when elastin content was normalized to the cell DNA content, no significant differences were found in the two substrates beyond the first two weeks of culture. Conversely, at culture times greater than 2 weeks, cells cultured atop hylan gels produced amounts of collagen/nanogram of DNA that were approximately 56% less than that synthesized by cells cultured on plastic. Cells grown on hylan deposited an unusual matrix layer, rich in elastin, at the hylan-cell interface. This elastin was found to be organized into fenestrated sheets and loose elastin fibers, structures that were also isolated from the elastin matrix of the ventricularis layer of porcine aortic valve cusps. We have thus demonstrated that hylan gels are useful as substrates to induce elastin synthesis in culture to obtain structures that resemble the elastin matrix of the native aortic valve.     

Impact of delivery mode of hyaluronan oligomers on elastogenic responses of adult vascular smooth muscle cells

Our prior studies demonstrated that exogenous supplements of pure hyaluronan (HA) tetramers (HA4) dramatically upregulate elastin matrix synthesis by adult vascular smooth muscle cells (SMCs). Some studies suggest that exogenous HA likely only transiently contacts and signals cells, and may elicit different cell responses when presented on a substrate (e.g., scaffold surface). To clarify such differences, we used a carbodiimide-based chemistry to tether HA4 onto glass, and compared elastin matrix synthesis by SMCs cultured on these substrates, with those cultured with equivalent amounts of exogenous HA4. Tethered HA4-layers were first characterized for homogeneity, topography, and hydrolytic stability using SEM, XPS, AFM, and FACE. In general, mode of HA4 presentation did not influence its impact on SMC proliferation, or cell synthesis of tropoelastin and matrix elastin, relative to non-HA controls; however, surface-tethered HA4 stimulated SMCs to generate significantly greater amounts of elastin-stabilizing desmosine crosslinks, which partially accounts for the greater resistance to enzymatic breakdown of elastin derived from these cultures. Elastin derived from both sets of cultures contained peptide masses that correspond to the predominant peptides present in rat aortic elastin. SEM and TEM showed that HA4-stimulated fibrillin-mediated elastin matrix deposition, and organization into fibrils. Surface-immobilized HA4 was particularly conducive to organization of elastin into aggregating fibrils, and their networking to form closely woven sheets of elastin fibers, as seen in cardiovascular tissues. The results suggest that incorporation of elastogenic HA4 mers onto cell culture substrates or scaffolds is a better approach than exogenous supplementation for in vitro or in vivo regeneration of architecturally and compositionally faithful-, and more stable mimics of native vascular elastin matrices.     

Attachment of hyaluronic acid to polypropylene, polystyrene, and polytetrafluoroethylene

Surfaces of polypropylene (PP), polystyrene (PS) and polytetrafluoroethylene (PTFE) were activated with radio frequency plasmas Ar and NH3 to aminate the polymer surface and were subsequently reacted with hyaluronic acid (HA) in one of the three different attachment schemes. Results show that ammonia plasma treated polymers were more reactive toward HA attachment. The three chemistry schemes consisted of two distinct approaches: (1) direct attachment of the HA to the aminated surface, and (2) extending the reactive group away from the surface with succinic anhydride and then reacting the newly formed carboxylic acid group with an adipic dihydrazide modified HA (HA-ADH). The latter scheme proved to be more effective, suggesting that steric effects were involved with the reactivity of the HA with surface groups. These HA-coated polymers are a candidate for cell attachment and growth.     

Synthesis and evaluation of collagen-chitosan-hydroxyapatite nanocomposites for bone grafting

Incorporation of hydroxyapatite (HA) into the matrix of collagen (Col) and chitosan (Chi) by in situ synthesis was introduced to prepare nanocomposites. Structural investigations of the pure Col-Chi mixture validated the influence of Chi on Col assembly, but the molecular interactions between Col and Chi was partially depressed during the intervention of in situ HA synthesis, as revealed by FTIR and DSC analyses. A series of Col-Chi-HA (CCHA) nanocomposites with varying HA content were thereby prepared by a sequential method, involving in situ synthesis in the Col-Chi system, then gelling at 25 degrees C and subsequently washing the resultant elastic gel followed by dehydration consolidation. The structural characteristics and biological properties of the dehydrated CCHA nanocomposites were further evaluated by using XRD, FTIR, TG, and SEM analyses and the osteoblast culture experiment. Formation of a well integrated microstructure of organic fibers (ca. 90 nm in size) and dense matrix including inorganic aggregates (less than 30 nm in size) was found in these nanocomposites. Rat Ros 17/2.8 Osteoblasts proliferated and attached well on the surface of both CCHA nanocomposite and Col-Chi mixture. These results indicated that in situ HA synthesis in the Col-Chi system provided a feasible route for bone grafting nanocomposites.     

The effect of dehydration history on PVA/PVP hydrogels for nucleus pulposus replacement

The feasibility of the use of a copolymer gel prepared from blends of polyvinyl alcohol (PVA) and polyvinyl pyrrolidone (PVP) for endoscopic replacement of the nucleus pulposus of a lumbar intervertebral disc was examined in this study. Hydrogels were processed with the use of three freeze/thaw cycles to induce crystallinity. As-prepared samples were dehydrated to various levels: 70.4, 46.3, 25.1, and 10.3% of their as-prepared masses and subsequently rehydrated. The dehydration history controlled the dimensions upon swelling and caused distortion of the material, with major distortion occurring when the hydrogel reached around 25% of the initial hydrated mass. The dehydration history affected the mechanical behavior of the rehydrated gels. Increased dehydration resulted in increased compressive modulus for the reswollen gels. Experiments were performed to investigate the formation of a skin layer that was found on the hydrogels during the dehydration process. The skin was found to dehydrate quickly and form a barrier to further dehydration from the core. Rubber elasticity theory was used to describe the differences in the network characteristics between the skin and the core of a drying hydrogel. The dehydration/rehydration process used in this study and an implantation of a cadaver model demonstrate the feasibility of endoscopic nucleus replacement.     

Macrophage adhesion on gelatin-based interpenetrating networks grafted with PEGylated RGD

Human blood-derived macrophage adhesion on interpenetrating networks (IPNs) composed of PEGylated RGD-modified gelatin and poly(ethylene glycol) diacrylate was studied. The interaction between biomaterial immobilized with biofunctional peptides such as RGD and macrophages is central in the design of tissue-engineering scaffolds. PEGylated RGD-modified gelatin was synthesized via several steps involving PEG derivations and characterized by high-performance liquid chromatography, mass spectroscopy, gel permeation chromatography, and the trinitrobenzenesulfonic acid method. IPNs containing modified or unmodified gelatin were cultured with human macrophages and monitored at 2, 24, 96, and 168 h. At each time point, IPNs containing gelatin modified with PEGylated RGD showed a comparable adherent macrophage density as tissue culture polystyrene and a significantly higher cell density than other IPN formulations containing unmodified gelatin or gelatin modified with PEGylated triglycine. Although surface-immobilized RGD can serve to mediate the adhesion of different cell types on the biomaterial surface, the interaction of RGD with immune/inflammatory cells such as macrophages should also be considered when assessing the potential host response of tissue-engineering scaffolds.     

Quantitative analysis of osteoblast behavior on microgrooved hydroxyapatite and titanium substrata

The effects of implant surface topography and chemistry on osteoblast behavior have been a research focus because of their potential importance in orthopedic and dental applications. This work focused on the topographic effects of hydroxyapatite (HA) and titanium (Ti) surface that had identical micropatterns to determine whether there was synergistic interaction between surface chemistry and surface topography. Surface microgrooves with six different groove widths (4, 8, 16, 24, 30, and 38 microm) and three different groove depths (2, 4, and 10 microm) were made on single crystalline silicon wafers using microfabrication techniques. Ti and HA thin films were coated on the microgrooves by radio-frequency magnetron sputtering. After that, human osteoblast-like cells were seeded and cultured on the microgrooved surfaces for up to 7 days. The cells' behavior was examined using scanning electron microscopy after cells were fixed and dehydrated. Statistical analysis was based on quantitative data of orientation angle, evaluating the contact guidance, and form index, describing cell shape or cell morphology changes. The contact guidance and cell shape changes were observed on the HA and Ti microgrooves. No difference in orientation angle between HA and Ti microgrooves was found. This might suggest that surface chemistry was not a significant influence on cell guidance. However, the form index analysis indicated an interaction between topographic effects and surface chemistry. Thus, conclusions about surface topographic effects on cell behavior drawn from one type of material cannot simply be applied to another type of material.     

Anti-inflammatory drug delivery from hyaluronic acid hydrogels

Two different types of hyaluronic acid (HA) hydrogels were synthesized by crosslinking HA with divinyl sulfone (DVS) and poly(ethylene glycol)-divinyl sulfone (VS-PEG-VS). Vitamin E succinate (VES), an anti-inflammatory drug, and bovine serum albumin (BSA), a model of anti-inflammatory protein drugs, were loaded into the gels and their release kinetics were measured in vitro. VES and BSA released with a burst from both HA hydrogels during the first few hours, and release continued gradually for several days. The rate of release from HA-VS-PEG-VS-HA hydrogels was faster than that from HA-DVS-HA hydrogels, presumably due to the lower crosslink density in the former. The anti-inflammatory action of released VES was tested by incubating peripheral blood mononuclear cells (PBMC) on HA hydrogels with and without VES in the gel. The number of cells adhering on HA hydrogels was very low compared to that on tissue culture polystyrene (TCPS), which might be one of the important advantages of using HA hydrogels for implant coatings or tissue engineering applications. ELISA test results showed that the tumor necrosis factor-alpha (TNF-alpha) concentration was very low in the supernatant of the wells containing the HA hydrogel with VES in contact with the activated macrophages compared to that without VES. This is probably the effect of the released VES reducing the production of anti-inflammatory cytokine, TNF-alpha. HA hydrogels containing anti-inflammatory drugs may have potential for use in tissue engineering and also as biocompatible coatings of implants.     

Hyaluronic acid modified biodegradable scaffolds for cartilage tissue engineering

Hyaluronic acid (hyaluronan, HA) was immobilized onto the surface of macroporous biodegradable poly(D,L-lactic acid-co-glycolic acid) [PLGA] scaffolds to enhance the attachment, proliferation, and differentiation of chondrocytes for cartilage tissue engineering. The PLGA scaffolds were prepared by blending PLGA with varying amounts of amine-terminated PLGA-PEG di-block copolymer. They were fabricated by a gas foaming/salt leaching method. HA was chemically conjugated to the surface-exposed amine groups on the pre-fabricated scaffolds. The amount of surface exposed free amine groups was quantitatively determined by conjugating an amine-reactive fluorescent dye to the PLGA blend films. The extent of HA immobilization was also confirmed by measuring water contact angles. When chondrocytes were seeded within HA modified PLGA scaffolds, enhanced cellular attachment was observed compared to unmodified PLGA scaffolds. Furthermore, glycosaminoglycan and total collagen synthesis increased substantially for HA modified PLGA scaffolds. RT-PCR result and histological examination of the resultant cartilage tissue revealed that HA modified scaffolds excelled in inducing cartilage tissue formation in terms of collagen type II expression and tissue morphological characteristics.     

Release of macromolecules and daunomycin from hydrophilic gels containing enzymatically degradable bonds

The synthesis of hydrophilic gels based on N (2-hydroxypropyl)methacrylamide copolymers crosslinked via degradable oligopeptide sequences is described. The influence of the conditions of preparation and of the gel structure on the equilibrium degree of swelling (network density) was determined. To evaluate the potential of such gels for controlled delivery of macrornolecules and drugs, the release of FITC-dextrans of different molecular weights was studied and the rate of release was found to depend mainly on the equilibrium degree of swelling and not on the structure of the crosslinks. However, the degradation of the gels by a mixture of lysosomal enzymes isolated from rat liver (Tritosomes) or chymotrypsin was dependent on both the equilibrium degree of swelling and the structure of the crosslinks (length of the oligopeptide sequence and structure of the amino acid residues). The release of the anticancer drug daunomycin imbibed in gels was pH-dependent, the rate of release being higher at lower pH. In addition, a gel was synthesized which contained a pentapeptide in the crosslinks and daunomycin bound via a tetrapeptide side-chain, and in this case, incubation with Tritosomes led to degradation with simultaneous release of the drug.     

The effect of bioactive hydrogels on the secretion of extracellular matrix molecules by valvular interstitial cells

Valvular interstitial cells (VICs) were encapsulated in enzymatically degradable, crosslinked hydrogels formed from hyaluronic acid (HA) and poly(ethylene glycol) (PEG) macromolecular monomers. Titration of PEG with HA allowed for the synthesis of gels with a broad compositional spectrum, leading to a range of degradation behavior upon exposure to bovine testes hyaluronidase. The rate of mass loss and release of HA fragments from the copolymer gels depended on the PEG content of the network. These hydrogels were shown to have the dual function of permitting the diffusion of ECM elaborated by 3D cultured VICs and promoting the development of a specific matrix composition. Initial cleavage of hydrogel crosslinks, prior to network mass loss, permit the diffusion of collagen, while later stages of degradation promote elastin elaboration and suppress collagen production due to HA fragment release. Exogenous HA delivery through the cell culture media further demonstrated the utility of delivered HA on manipulating the secretory properties of encapsulated VICs.     

Anti-inflammatory drug delivery from hyaluronic acid hydrogels

Two different types of hyaluronic acid (HA) hydrogels were synthesized by crosslinking HA with divinyl sulfone (DVS) and poly(ethylene glycol)-divinyl sulfone (VS-PEG-VS). Vitamin E succinate (VES), an anti-inflammatory drug, and bovine serum albumin (BSA), a model of anti-inflammatory protein drugs, were loaded into the gels and their release kinetics were measured in vitro. VES and BSA released with a burst from both HA hydrogels during the first few hours, and release continued gradually for several days. The rate of release from HA-VS-PEG-VS-HA hydrogels was faster than that from HA-DVS-HA hydrogels, presumably due to the lower crosslink density in the former. The anti-inflammatory action of released VES was tested by incubating peripheral blood mononuclear cells (PBMC) on HA hydrogels with and without VES in the gel. The number of cells adhering on HA hydrogels was very low compared to that on tissue culture polystyrene (TCPS), which might be one of the important advantages of using HA hydrogels for implant coatings or tissue engineering applications. ELISA test results showed that the tumor necrosis factor-α (TNF-α) concentration was very low in the supernatant of the wells containing the HA hydrogel with VES in contact with the activated macrophages compared to that without VES. This is probably the effect of the released VES reducing the production of anti-inflammatory cytokine, TNF-α. HA hydrogels containing anti-inflammatory drugs may have potential for use in tissue engineering and also as biocompatible coatings of implants.     

Release of macromolecules and daunomycin from hydrophilic gels containing enzymatically degradable bonds.

The synthesis of hydrophilic gels based on N-(2-hydroxypropyl)methacrylamide copolymers crosslinked via degradable oligopeptide sequences is described. The influence of the conditions of preparation and of the gel structure on the equilibrium degree of swelling (network density) was determined. To evaluate the potential of such gels for controlled delivery of macromolecules and drugs, the release of FITC-dextrans of different molecular weights was studied and the rate of release was found to depend mainly on the equilibrium degree of swelling and not on the structure of the crosslinks. However, the degradation of the gels by a mixture of lysosomal enzymes isolated from rat liver (Tritosomes) or chymotrypsin was dependent on both the equilibrium degree of swelling and the structure of the crosslinks (length of the oligopeptide sequence and structure of the amino acid residues). The release of the anticancer drug daunomycin imbibed in gels was pH-dependent, the rate of release being higher at lower pH. In addition, a gel was synthesized which contained a pentapeptide in the crosslinks and daunomycin bound via a tetrapeptide side-chain, and in this case, incubation with Tritosomes led to degradation with simultaneous release of the drug.     

Plasma modification of PMMA films: surface free energy and cell-attachment studies

The surface of a material is the most important part determining the acceptance by and compatibility with the environment. In many cases, although the bulk properties are excellent for a specific application, the surface may require to be modified and engineered in the desired direction. This is especially important for materials used in biological media, since the surface charge, hydophilicity and wettability are important for thrombosis formation, cell attachment or cell proliferation. In this study, poly(methyl methacrylate) films were prepared by solvent casting and their surfaces were modified by oxygen plasma treatment by applying powers of 20, 100 and 300 W. The effects of surface chemistry alterations on hydophilicity, work of adhesion, surface free energy and cell adhesion were examined. Cell attachment and proliferation are especially important for the materials used for tissue-engineering purposes. The results demonstrated that there is an optimum value for hydrophilicity and surface free energy which enhance cell attachment.     

Plasma modification of PMMA films: surface free energy and cell-attachment studies

The surface of a material is the most important part determining the acceptance by and compatibility with the environment. In many cases, although the bulk properties are excellent for a specific application, the surface may require to be modified and engineered in the desired direction. This is especially important for materials used in biological media, since the surface charge, hydophilicity and wettability are important for thrombosis formation, cell attachment or cell proliferation. In this study, poly(methyl methacrylate) films were prepared by solvent casting and their surfaces were modified by oxygen plasma treatment by applying powers of 20, 100 and 300 W. The effects of surface chemistry alterations on hydophilicity, work of adhesion, surface free energy and cell adhesion were examined. Cell attachment and proliferation are especially important for the materials used for tissue-engineering purposes. The results demonstrated that there is an optimum value for hydrophilicity and surface free energy which enhance cell attachment.