BHK cell attachment and growth on EDA-plasma-modified poly(L-lactide/ε-caprolactone) biodegradable films

In this study, attachment and growth of Baby Hamster Kidney (BHK) cells on ethylene diamine (EDA)-plasma-treated poly(L-lactide/ε-caprolactone) biodegradable copolymer films were investigated. The co-polymer (M_w: 58 000; M_n: 35 000 and PI 1.60) was synthesised by ring-opening polymerization of the respective dimers with using stannous octoate as the catalyst. The final ratio of L-lactide to ε-caprolactone obtained by ¹H-NMR was 87 :13. The co-polymer films were treated with the EDA-plasma in a glow-discharge apparatus. The BHK-30 cell line was cultured on plain and EDA-plasma-treated films and their pre-wetted forms (with ethanol and/or cell culture medium before use). Cell attachment and growth were followed. Alkaline phosphatase (ALP) activity and glucose uptake in cell culture medium were also investigated. There was no attachment in the first 12 h. Glow-discharge treatment increased significantly the attachment and growth. Pre-wetting with ethanol and cell culture medium was also increase significantly both the attachment and growth.     

Attachment and growth of fibroblasts on poly(L-lactide/epsilon-caprolactone) scaffolds prepared in supercritical CO2 and modified by polyethylene imine grafting with ethylene diamine-plasma in a glow-discharge apparatus

In this study, a copolymer of L-lactide and epsilon-caprolactone (Mn: 73,523, Mw: 127,990 and PI: 1.74) was synthesized by ring-opening polymerization by using stannous octoate as the catalyst. FTIR, 1H-NMR and DSC confirmed the copolymer formation. The copolymer films were prepared and a novel method was developed to produce highly porous sponges for potential use in tissue engineering. Films were subjected to supercritical CO2 at 3300 psi and 70 degrees C to create porous structures for production of possible tissue engineering scaffolds. The pore sizes were in the range of 40-80 microm. The copolymer films were pre-wetted with polyethylene imine (PEI) and then treated with ethylene diamine (EDA)-plasma in glow-discharge apparatus. Gas plasma surface modification of three-dimensional scaffolds fabricated by supercritical carbon dioxide technique was demonstrated to enhance cell adhesion, proliferation, and differentiation over 6 days in culture using L929 fibroblast cell line. Alkaline phosphatase (ALP) activity and glucose uptake in cell culture medium were followed in the cell culture experiments. Fibroblastic cell attachment and growth on the EDA-plasma treated scaffolds were rather low. However, both cell attachment and growth were significantly increased by PEI pre-treatment before EDA-plasma. The changes in ALP activity and glucose uptake also supported the cell growth behavior on these PEI and EDA-plasma treated scaffolds.     

Human articular chondrocyte adhesion and proliferation on synthetic biodegradable polymer films

The effect of polymer chemistry on adhesion, proliferation, and morphology of human articular cartilage (HAC) chondrocytes was evaluated on synthetic degradable polymer films and tissue culture polystyrene (TCPS) as a control. Two-dimensional surfaces of poly(glycolide) (PGA), poly(L-lactide) (L-PLA), poly(D,L-lactide) (D,L-PLA), 85:15 poly(D,L-lactide-co-glycolide) (D,L-PLGA), poly(epsilon-caprolactone) (PCL), 90:10 (D,L-lactide-co-caprolactone) (D,L-PLCL), 9:91 D,L-PLCL, 40:60 L-PLCL, 67:33 poly(glycolide-co-trimethylene carbonate) (PGTMC), and poly(dioxanone) (PDO) were made by spin-casting into uniform thin films. Adhesion kinetics were studied using TCPS and PCL films and revealed that the rate of chondrocyte adhesion began to level off after 6 h. Degree of HAC chondrocyte adhesion was studied on all the substrates after 8 h, and ranged from 47 to 145% of the attachment found on TCPS. The greatest number of chondrocytes attached to PGA and 67:33 PGTMC polymer films, and attachment to PCL and L-PLA films was statistically lower than that found on PGA (p < 0.05). There was no correlation between amount of chondrocyte attachment to the substrates and the substrates' water contact angle. Chondrocytes proliferated equally well on all the substrates resulting in equivalent cell numbers on all the substrates at both day 4 and day 7 of the culture. However, these total cell numbers were reached as a result of a 88- and 42-fold expansion on PDO and PLA, respectively, which was significantly higher than the 11-fold expansion found on TCPS (p < 0.05). The greater fold expansion of the cells on PDO and L-PLA films may be attributed to the availability of space for cells to grow, since their numbers at the start of culture were fewer following the 8 h attachment period. This suggests that regardless of initial seeding density on these degradable polymer substrates (i.e., if some minimum number of cells are able to attach), they will eventually populate the surfaces of all these polymers given sufficient space and time.     

Preparation and characterization of blends of star-poly(epsilon-caprolactone-co-D,L-lactide) and oligo(epsilon-caprolactone)

Polymer blending provides a relatively facile means of combining the separate desirable properties of different polymers into a single material. In this paper blends of a low-molecular-weight star co-polymer of epsilon-caprolactone and D,L-lactide with a linear oligo(epsilon-caprolactone) are prepared and characterized as a possible biodegradable injectable drug-delivery vehicle. The melting characteristics, melt viscosity and degree of crystallinity of the blends were measured, and an in vitro degradation study was performed over a period of 12 weeks. The blends all had a single glass transition temperature and an onset of melting point near body temperature, with the melting point range decreasing as the star co-polymer content increased. The melt viscosity of the blends increased as the star co-polymer content increased, in a manner consistent with miscible blend behavior. The star co-polymer degraded fastest, with a more than 60% mass decrease over the 12-week period. As the oligo(epsilon-caprolactone) content increased, the degradation rate decreased, with the oligo(epsilon-caprolactone) exhibiting a mass loss of only 12% over the 12-week period.     

Cell growth on the porous sponges prepared from poly(depsipeptide-co-lactide) having various functional groups

In tissue engineering, excellent biodegradable materials are desired as temporary scaffolds to support cell growth and disappear with the progress of tissue regeneration. We previously synthesized biodegradable poly(depsipeptide-co-lactide), poly[(Glc-Asp)-co-LA] and poly[(Glc-Lys)-co-LA], having reactive side-chain groups. Then, the effects of reactive and ionic side-chain groups on cell attachment and growth were investigated using co-polymer films with various amounts of carboxyl or amino groups. In this study, to evaluate the utility of these co-polymers as functional scaffolds for tissue regeneration, 3-dimensional porous sponges were prepared by freeze-drying method and the effects of reactive and ionic side-chain groups on cell growth and degradation behavior were investigated using co-polymer sponges with various amounts of carboxyl or amino groups. Good cell growth was observed on the co-polymer sponges. During cell culture, the co-polymer sponges exhibited various degradation rates related to the depsipeptide unit content. Three-dimensional biodegradable polymer matrices with reactive surface, controllable degradation behavior and good cell growth were successfully prepared using these co-polymers. Such kinds of co-polymer matrices are good candidate for scaffold for tissue engineering.     

Apatite formation in composites of alpha-TCP and degradable polyesters

The objective of this study was to investigate the conversion of alpha-Ca3(PO4)2 (alpha-TCP) in composite bone cements based on a water-degradable polyester matrix as a function of the polymer formulation and the alpha-TCP filler content. Cross-linkable dimethacrylates of epsilon-caprolactone/ D,L-lactide co-polymer or of epsilon-caprolactone/glycolide co-polymer were mixed with hydroxyethylmethacrylate, a photo-initiator and alpha-TCP to obtain composites with a filler content of 80 or 40 wt% alpha-TCP. The disk shaped composite samples were set by visible light irradiation and immersed in HEPES at 37 degrees C. At selected times the samples were removed from the solution and analysed with X-ray diffractometry and infrared spectroscopy. Conversion of alpha-TCP into calcium-deficient hydroxyapatite (CDHAp) was observed for all composites, but the reaction was not completed after 8 weeks immersion. The conversion rate of alpha-TCP and the crystallinity of the formed apatite apparently were not affected by the type of polyester used, but significantly depended on the alpha-TCP content of the composites. An increase of the amount of alpha-TCP in the composite resulted in a slower formation of CDHAp with a higher crystallinity.     

Microwave-assisted synthesis of poly(epsilon-caprolactone)-poly(ethylene glycol)-poly(epsilon-caprolactone) tri-block co-polymers and use as matrices for sustained delivery of ibuprofen taken as model drug

Poly(epsilon-caprolactone)-poly(ethylene glycol)-poly(epsilon-caprolactone) triblock co-polymers with number-average molar mass (Mn) over 20000 g/mol were prepared by ring-opening polymerization of epsilon-caprolactone initiated by poly(ethylene glycol) under microwave irradiation. This method was proposed as a means to improve in vivo compatibility as no harmful chemicals were involved in the polymerization except epsilon-caprolactone and poly(ethylene glycol). The resulting tri-block co-polymers were characterized by FT-IR, H-NMR, GPC and WAXD. Their Mn and their composition was controlled by the amount and the chain length of the poly(ethylene glycol) macromers involved in the feed. The ability of poly(epsilon-caprolactone)-poly(ethylene glycol)-poly(epsilon-caprolactone) co-polymers to entrap and deliver drugs was investigated with ibuprofen as a model drug. The release of ibuprofen was significantly influenced by the co-polymer composition and the extent of loading. The in vitro release of ibuprofen was sustained from 3 to 15 days for 10% loading, depending on the ratio of epsilon-caprolactone to ethylene glycol-derived subunits in co-polymer chains. This ratio ranged from 0.97 to 9.78. In the case of the co-polymer whose epsilon-caprolactone molar ratio to ethylene glycol-derived subunits was 2.49, the ibuprofen release was sustained for 2 to 24 days for ibuprofen loads going from 5 to 20 wt%.     

Development of bFGF-Chitosan Matrices and Their Interactions with Human Dermal Fibroblast Cells

Chitosan (CH) is a naturally derived, biodegradable polymer of glucosamine with a variable frequency of N-acetyl-D-glucosamine units, and has been demonstrated to have numerous pharmacological and wound-healing properties. Biodegradable chitosan films were fabricated using a solvent casting technique and investigated for skin tissue-engineering applications. Basic fibroblast growth factor (bFGF) was incorporated into the CH matrices (1 μg/film) by 3 methods: adsorption, entrapment and covalent binding. Release rates and biological activity of the incorporated bFGF were monitored. Human dermal fibroblasts (HDF cells) were used as an in vitro model for cell response to CH and bFGF-CH films. Cell attachment, growth and acid-soluble collagen quantification were employed as an assessment of cell function. The fibroblasts were found to remain viable on the chitosan films and scaffolds. CH films without bFGF were compatible with HDF cells; however, the fibroblasts did not proliferate. The release profile of adsorbed and bound bFGF from CH films were similar (indicating that binding was not efficient) while entrapped bFGF was not released in the time frame studied. The concentration of bFGF released to the cell culture medium was not high enough to stimulate HDF proliferation. However, cell attachment was significantly increased in chitosan films with bFGF adsorbed onto the surface as compared to control surfaces. HDF cells grown on CH films produced significantly more collagen than those on control surfaces.     

Comparative in vitro and in vivo studies using a bioactive poly(epsilon-caprolactone)-organosiloxane nanohybrid containing calcium salt

Comparative in vitro and in vivo studies were conducted using a bioactive poly(epsilon-caprolactone)-organosiloxane nanohybrid containing calcium, which was prepared by sol-gel method. The behavior of human bone marrow stromal cells (hBMSCs) during in vitro osteogenic differentiation were evaluated on poly(epsilon-caprolactone)-organosiloxane nanohybrid and poly(epsilon-caprolactone)-organosiloxane nanohybrid coated with apatite, which mimicked in vivo events. hBMSCs cultured on tissue culture plates (TCPs) were used as a control. For comparative studies, in vivo testing was also conducted using poly(epsilon-caprolactone)-organosiloxane nanohybrid and poly(epsilon-caprolactone)-organosiloxane nanohybrid coated with apatite in the diaphyseal bone defects of rabbit tibiae. Initial attachments and early proliferations of hBMSCs onto poly(epsilon-caprolactone)-organosiloxane with or without the apatite layer were comparable those onto TCPs. However, the late proliferation and the osteogenic differentiation activities on poly(epsilon-caprolactone)-organosiloxane nanohybrid were significantly lower than the hybrid coated with apatite or TCPs. These results were caused by the delayed detachment of hBMSCs induced by the upward growth of spire-shaped apatite granules on the flat apatite layer through mixed nucleation (heterogeneous and homogeneous nucleation) and growth of apatite crystals during cell culture. However, the poly(epsilon-caprolactone)-organosiloxane nanohybrid showed excellent osteoconductivity as same as poly(epsilon-caprolactone)-organosiloxane nanohybrid coated with apatite in vivo even though the cell testing results in vitro were poor. This discrepancy can be explained by the difference in initial degree of supersaturation of apatite in cell culture medium and buffering ability between cell culture medium and body fluid with respect to calcium, which directly affects the nucleation mechanism of apatite crystals and the morphology of grown apatite granules. These findings imply that much attention is required and an optimal method should be used to assess cell responses in vitro. Our results suggest that precoating the apatite layer before in vitro testing is desirable for bioactive materials that release calcium quickly and in large amount because this treatment can more closely mimic in vivo events.     

Improved cell adhesion by plasma-induced grafting of L-lactide onto polyurethane surface

Lactide-grafted polyurethanes were prepared by exposing the polyurethane films to argon plasma discharge, followed by grafting L-lactide onto the plasma-treated surface. The modified surfaces were characterized by measuring the static contact angle and by electron spectroscopy for chemical analysis (ESCA). The water contact angle of polyurethanes was decreased by L-lactide grafting, indicating hydrophilicity of the modified surface. Grafting also increased the O/C atomic ratio and C(C=O)/Ctotal percentage on the surfaces as detected by ESCA. The grafted surfaces showed enhanced attachment and growth in both 3T3 fibroblast and human umbilical vein endothelial cell culture tests. Platelet adhesion to the modified surfaces was also reduced in vitro. L-Lactide monomers grafted onto polyurethane substrates could therefore be useful in facilitating endothelial cell seeding process in small vascular graft applications.     

Poly(N-isopropylacrylamide) co-polymer films as potential vehicles for delivery of an antimitotic agent to vascular smooth muscle cells.

INTRODUCTION: Local delivery of antimitotic agents is a potential therapeutic strategy for protection of injured coronary vasculature against intimal hyperplasia and restenosis. This study sought to establish the principle that thermoresponsive poly(N-isopropylacrylamide) co-polymer films can be used to deliver, in a controlled manner, an antimitotic agent to vascular smooth muscle cells (VSMC). METHODS: A series of co-polymer films was prepared, using varying ratios (w/w) of N-isopropylacrylamide (NiPAAm) monomer to N-tert-butylacrylamide (NtBAAm) and loaded with the antimitotic agent colchicine (100 nmol/film) at room temperature. RESULTS: The extent of colchicine release at 37 degrees C was inversely proportional to the amount of NtBAAm in co-polymer films: release after 48 h from 85:15, 65:35 and 50:50 (NiPAAm:NtBAAm) films was 26, 17 and 0.5 nmol, respectively. In cytotoxicity studies, when medium incubated with co-polymers for 24 h (in the absence of colchicine) was further incubated with target bovine aortic smooth muscle cells (BASMC), no loss of cell viability occurred. Colchicine released from all three co-polymer films significantly inhibited proliferation and random migration of BASMC: 100 nM colchicine (released from 65:35 NiPAAm:NtBAAm) reduced cell proliferation to 25.7+/-1.7% of levels seen in the absence of colchicine (control) and random cell migration to 37.7+/-5.7% of control (mean+/-S.E.M., n = 3, P < .01 and P < .05, respectively). The magnitudes of these effects were comparable to those seen in separate experiments with native colchicine and were observed in samples of released colchicine which had been stored at -20 degrees C for up to 6 months. CONCLUSIONS: This study has shown that the release of the antimitotic agent colchicine, from NiPAAm/NtBAAm co-polymer films can be manipulated by changes in co-polymer composition. Furthermore, such drug released at 37 degrees C retains comparable bioactivity to that of native colchicine.     

Cytotoxicity of glutaraldehyde crosslinked collagen/poly(vinyl alcohol) films is by the mechanism of apoptosis

Collagen has been investigated as a potential natural biomaterial, because of its occurrence in the extracellular matrix. Collagen requires crosslinking in this context, by reagents that are often cytotoxic. Glutaraldehyde is one such agent that is potentially cytotoxic. The aim of this study was to determine the cause of poor cell attachment and growth on collagen/poly(vinyl alcohol) bioartificial composite films, when crosslinked with glutaraldehyde. Dehydrothermal crosslinking was used as a comparison. Human osteoblasts were observed to undergo apoptosis on glutaraldehyde crosslinked films dependent on concentration of collagen present. Higher collagen content resulted in higher levels of apoptosis with poor cell attachment and spreading of remaining cells. Post-treatment of films with 8% L-glutamic acid prevented the apoptotic response of osteoblasts and allowed attachment and spreading. The addition of 100 nM insulin-like growth factor-1 to the culture medium also prevented apoptosis. Glutaraldehyde toxicity of crosslinked collagen has been demonstrated in this study, the mechanism of which is apoptosis. This study indicates that poor biocompatibility and induction of apoptosis on collagen/poly(vinyl alcohol) films crosslinked by glutaraldehyde are attributed to glutaraldehyde components on the surface of the films (not residual glutaraldehyde), whose effects can be quenched by glutamic acid, and prevented by insulin-like growth factor-1.     

Cytocompatibility of aliphatic polyesters--in vitro study on fibroblasts and macrophages

A resorbable copolymer of glycolide and L-lactide (PGLA), a terpolymer of glycolide, L-lactide, and epsilon-caprolactone (PGLCL), and a copolymer of glycolide and epsilon-caprolactone (PGCL) were synthesized by ring opening polymerization using zirconium acetylacetonate (Zr(acac)(4)) as an initiator. The structure and physicochemical surface properties of the materials were studied by NMR spectroscopy, gel permeation chromatography, differential scanning calorimetry, X-ray photoelectron spectroscopy, atomic force microscopy, and contact angle measurements. On the basis of contact angle measurements and the Owens-Wendt approach, the surface free energy was calculated. The effect of polymeric films produced by solvent casting on morphology and activity of L929 fibroblasts, and two types of macrophages (macrophages from peritoneal exudates and RAW 264.7 monocytes/macrophages), was analyzed. It was found that viability, adhesion, and morphology of fibroblasts on PGLA were very similar to control glass. On PGLCL more adhering cells were round, while on PGCL only single, poorly spread cells were seen, and their viability was significantly reduced. This may suggest that the interaction of fibroblasts with PGCL was due to its hydrophobicity and a very low polarity. Adhesion and viability of RAW 264.7 cells was significantly enhanced on PGLA but reduced on both PGLCL and PGCL. The increased synthesis/release of chemoattractants and metalloproteinases-2 and -9 was observed in the macrophages from peritoneal exudates cultured on PGLA and PGLCL. The viability of cells decreased in the following order: PGLA > PGLCL > PGCL. It is worth noting that glass transition temperature and susceptibility to mechanical deformation of the polymeric materials also decreased in the same order. It may imply that those physical parameters should be also considered as potential factors affecting cell behavior.     

Fabrication using a rapid prototyping system and in vitro characterization of PEG-PCL-PLA scaffolds for tissue engineering

In the field of tissue engineering new polymers are needed to fabricate scaffolds with specific properties depending on the targeted tissue. This work aimed at designing and developing a 3D scaffold with variable mechanical strength, fully interconnected porous network, controllable hydrophilicity and degradability. For this, a desktop-robot-based melt-extrusion rapid prototyping technique was applied to a novel tri-block co-polymer, namely poly(ethylene glycol)-block-poly(epsilon-caprolactone)-block-poly(DL-lactide), PEG-PCL-P(DL)LA. This co-polymer was melted by electrical heating and directly extruded out using computer-controlled rapid prototyping by means of compressed purified air to build porous scaffolds. Various lay-down patterns (0/30/60/90/120/150 degrees, 0/45/90/135 degrees, 0/60/120 degrees and 0/90 degrees) were produced by using appropriate positioning of the robotic control system. Scanning electron microscopy and micro-computed tomography were used to show that 3D scaffold architectures were honeycomb-like with completely interconnected and controlled channel characteristics. Compression tests were performed and the data obtained agreed well with the typical behavior of a porous material undergoing deformation. Preliminary cell response to the as-fabricated scaffolds has been studied with primary human fibroblasts. The results demonstrated the suitability of the process and the cell biocompatibility of the polymer, two important properties among the many required for effective clinical use and efficient tissue-engineering scaffolding.     

Influence of the physical properties of two-dimensional polyester substrates on the growth of normal human urothelial and urinary smooth muscle cells in vitro

Although synthetic biomaterials have a wide range of promising applications in regenerative medicine and tissue engineering, there is limited insight into the basic materials properties that influence cellularisation events. The aim of this study was to investigate the influence of the physical properties of polyester films on the adherence and growth of normal human urothelial and urinary smooth muscle (SM) cells, as part of a programme for the development of potential biomaterials for bladder tissue engineering. Films of different thickness were produced by spin coating from solution. Cell attachment and proliferation were analysed and revealed a reproducible and significant growth advantage over the initial 7 days for both cell types on poly(lactide-co-glycolide) (PLGA) versus poly(epsilon-caprolactone) (PCL), and on thick versus thin films. In order to understand the basis of the variation in cell growth, the surface morphology, degradation behaviour and mechanical properties of the films were investigated. The pattern of cell attachment and growth was found to be unrelated to surface topography and no distinction in film degradation behaviour was found to account for differences in cell growth, except at late time points (14 days), where degradation of thin PLGA films became significant. By contrast, the flexural loss and storage moduli were found to be reduced in films composed of PLGA versus PCL, and also as film thickness increased, indicating that mechanical properties of biomaterials can influence cell growth. We conclude that elastic modulus is relevant to biology at the cellular scale and may also be influential at the tissue/organ level, and is a critical parameter to be considered during development of synthetic biomaterials for tissue engineering.     

Biodegradable polymer/hydroxyapatite composites: surface analysis and initial attachment of human osteoblasts

Biodegradable polymer/hydroxyapatite (HA) composites have potential application as bone graft substitutes. Thin films of polymer/HA composites were produced, and the initial attachment of primary human osteoblasts (HOBs) was assessed to investigate the biocompatibility of the materials. Poly(epsilon-caprolactone) (PCL) and poly(L-lactic acid) (PLA) were used as matrix materials for two types of HA particles, 50-microm sintered and submicron nonsintered. Using ESEM, cell morphology on the surfaces of samples was investigated after 90 min, 4 h, and 24 h of cell culture. Cell activity and viability were assessed after 24 h of cell culture using Alamar blue and DNA assays. Surface morphology of the polymer/HA composites and HA exposure were investigated using ESEM and EDXA, respectively. ESEM enabled investigation of both cell and material surface morphology in the hydrated condition. Combined with EDXA it permitted chemical and visual examination of the composite. Differences in HA exposure were observed on the different composite surfaces that affected the morphology of attached cells. In the first 4 h of cell culture, the cells were spread to a higher degree on exposed HA regions of the composites and on PLA than they were on PCL. After 24 h the cells were spread equally on all the samples. The cell activity after 24 h was significantly higher on the polymer/HA composites than on the polymer films. There was no significant difference in the activity of the cells on the various composite materials. However, cells on PCL showed higher activity compared to those on PLA. A polymer surface exhibiting "point exposure" of HA appeared to provide a novel and favorable substrate for primary cell attachment. The cell morphology and activity results indicate a favorable cell/material interaction and suggest that PLA and PCL and their composites with HA may be candidate materials for the reconstruction of bony tissue. Further investigations regarding long-term biomaterial/cell interactions and the effects of acidic degradation products from the biodegradable polymers are required to confirm their utility.     

Evaluation of cell affinity on poly(L-lactide) and poly(epsilon-caprolactone) blends and on PLLA-b-PCL diblock copolymer surfaces

An evaluation of cell proliferation and adhesion on biocompatible film supports was performed. A series of films were compression molded from commercially available poly (L-lactide), PLLA, and poly(epsilon-caprolactone), PCL, and from their melt mixed blends (PLLA/PCL blends). These were compared with compression molded films of PLLA-b-PCL model diblock copolymers. The samples were analyzed by differential scanning calorimetry (DSC), contact angle measurements, and scanning force microscopy (SFM). Cell adhesion and proliferation were performed with monkey derived fibroblasts (VERO) and with osteoblastic cells obtained either enzymatically or from explants cultures of Sprague-Dawley rat calvaria. Migration studies were performed with bone explants of the same origin. The results obtained indicate that although all materials tested were suitable for the support of cellular growth, a PLLA-b-PCL diblock copolymer sample with 93% PLLA was significantly more efficient. This sample exhibited a unique surface morphology with long range ordered domains (of the order of 2-3 mum) of edge-on PLLA lamellae that can promote "cell contact guidance." The influence of other factors such as chemical composition, degree of crystallinity, and surface roughness did not play a major role in determining cell preference toward a specific surface for the materials employed in this work.     

Synthesis and characterization of poly(ethylene glycol)-poly(D,L-lactide-co-glycolide) poly(ethylene glycol) tri-block co-polymers modified with collagen: a model surface suitable for cell interaction

This study focused on the synthesis and characterization of poly(ethylene glycol)-poly(D,L-lactide-co-glycolide)-poly(ethylene glycol) tri-block co-polymer (PEG-PDLLG-PEG), and its modification with type-I collagen. To this aim, a PEG-PDLLG-PEG tri-block co-polymer was synthesized in two steps by reacting poly(ethylene glycol)bis(carboxymethyl)ether with thionyl chloride to obtain an acyl-halide-terminated poly(ethylene glycol) and subsequently coupling this compound to hydroxyl-terminated poly(D,L-lactide-co-glycolide) (PDLLG). The new carboxyl endgroups of PEG-PDLLG-PEG were subsequently reacted with N-hydroxysuccinimide (NHS) in the presence of the hetero-bifunctional cross-linking agent dicyclohexylcarbodiimide (DCC) in order to activate the co-polymer for coupling with collagen. PEG-PDLLG-PEG and its activated form PEG-PDLLG-NHS were characterized by Fourier transform infrared (FT-IR) and 1H-NMR spectroscopy. Molecular weights of the polymeric products were determined by SEC. Type-I collagen in phosphate buffer was reacted with PEG-PDLLG-NHS. The resultant product, PEG-PDLLG-Col, was characterized by FT-IR. This biopolymer was used for preparation of a suitable surface for cell growth experiment. To measure the degree of cell proliferation, the films prepared with PDLLG, PEG-PDLLG-NHS and PEG-PDLLG-Col were seeded with L929 mouse fibroblasts. Cell growth was followed by SEM photography and quantitated by the neutral red uptake assay. It was shown that the attachment of collagen significantly increased the number of cells on the co-polymers.     

Biodegradable poly(L-lactide)-poly(ethylene glycol) multiblock copolymer: synthesis and evaluation of cell affinity

A series of poly(L-lactide)-poly(ethylene glycol) multiblock copolymers (Multi-PLE) with high molecular weight were synthesized and successfully used to fabricate three-dimensional scaffolds. Using mouse NIH 3T3 fibroblasts as model cells, the cell affinity of various Multi-PLE copolymers was evaluated and compared with that of poly(L-lactide) (PLLA) by means of cell attachment efficiency measurement, scanning electron microscopy observation and MTT assay. On one hand, the results showed that the cell attachment efficiency on Multi-PLE 4/1(4/1 refers to the molar ratio of lactidyl units to ethylene oxide units) films was close to that on PLLA film, however, the other Multi-PLE films exhibited much lower cell attachment efficiency than PLLA film, such as Multi-PLE 2/1 and Multi-PLE 1/1, which had higher PEG content. On the other hand, it was interesting to find that cell proliferation on Multi-PLE4/1 and Multi-PLE2/1 scaffolds was better than that on PLLA scaffold, which was closely related to the improved hydrophilicity of Multi-PLE copolymers due to the incorporation of PEG in comparison with pure PLLA. The Multi-PLE copolymer scaffolds with appropriate hydrophilicity were in favor of mass transportation, and then of cell proliferation and cell affinity. It meant that the cell proliferation would be much improved by increasing the hydrophilicity of the three-dimensional scaffolds, which even outweighed the disadvantages of the cell attachment efficiency reduction with the incorporation of PEG.     

Surface-induced changes in protein adsorption and implications for cellular phenotypic responses to surface interaction

Understanding external factors that determine cellular phenotypic responses is of key interest in the field of biomaterials. Currently, material surface characteristics, protein adsorption and cellular phenotypic responses are all considered to be interrelated and ultimately determine the biocompatibility of materials. The exact nature of the relationship between these distinct, yet related, phenomena still remains to be elucidated. Through the use of a series of thermoresponsive N-isopropylacrylamide-based co-polymer films, we aimed to shed light on the relationship between surface hydrophobicity, protein adsorption and subsequent cellular response. Despite changes in co-polymer hydrophobicity mediated by altered ratios of constituent monomers, differential cellular response was only apparent in the presence of serum. Co-polymer films displayed alterations with respect to the amount of protein adsorbed on the surface, with individual serum proteins (albumin and fibronectin) displaying contrasting adsorption characteristics. Changes in protein adsorption corresponded to changes in cell adhesion, cytoskeletal organisation and cell morphology, as well as to changes in cell movement and intracellular signalling events. Examination of focal adhesion kinase (FAK), and extracellular signal-regulated kinase (ERK 1/2), important mediators of adhesion and growth factor-related signalling events, revealed a comparative reduction in phosphorylation of these signalling proteins in cells grown on co-polymers in comparison to those cultured on tissue culture polystyrene (TCP; used as a control surface). We also associated surface-mediated phenotypic alterations of cells grown on TCP and co-polymer films with particular changes in gene expression. These results indicate that cellular response to interaction with our series of co-polymer films is determined by the surface-adsorbed protein layer, which in turn is determined by the changing surface chemistry as the ratio of the co-monomers is altered.