Preparation of Albumin Microspheres Grafted Galactose Residues Through Polyethylene-Glycol Spacers,Release Behavior of 5-Fluorouracil from Them,and Their Lectin-Mediated Aggregation

Small-sized albumin gel microspheres, MSs, containing 5-fluorouracil (5FU) with targeting moieties on their surfaces (average diameter: 1.5 μm) were prepared by the glutaraldehyde crosslinking method and suspension technique. Since galactose is known to interact specifically with the asialoglycoprotein receptor on hepato-cyte, the galactose residues were introduced on the surface of MSs as the targeting moieties for hepatoma through polyethylene glycol (PEG) spacers. PEG spacers were employed to depress the immu-nogenicity of albumin, to keep the mobility of the galactose residues, and to heighten the distributive stability of the MSs in aqueous solution. It was confirmed by ESC A analysis that the PEG chains were introduced onto the surfaces of MSs. The amount of galactose residues introduced to MS were estimated to be 0.013 wt%. The intra-MSs aggregation was observed by the addition of Ricinus Communis Agglutinin I (RCA120) into the MS suspension, and then the aggregation of MSs was dissociated by addition of free lactose. Moreover, by incubation of the MSs with human hepatoma HLE cells, the phenomena of MS's specific binding onto HLE cell surfaces and phagocytosis of MSs by HLE cells were observed. These results suggested that the galactose residues on the surface of MSs were recognized with the galactose receptors on hepatoma cell surfaces. The release rate of 5FU from MSs was investigated in vitro in physiological saline at 37OC. About 90% of encapsulated 5FU were found to be released from MSs through incubation for 8 h.     

Copolymers for hepatocyte-specific targeting carrying galactose and hydrophobic alkyl groups

Core-forming hydrophobic alkyl groups were incorporated into amphiphilic PVLA to enhance the stability and inclusion ability of the homopolymeric micelles in water. The CAC and hepatocyte targeting in the synthesized P(VLA-co-VBH) were investigated in vitro. The CAC of the copolymers decreased and the stability of the copolymeric micelles increased with the incorporation of hydrophobic groups into the homopolymer. The galactose moieties in the copolymer could be recognized by the ASGP-R of the hepatocytes through a receptor-mediated mechanism. The copolymers efficiently delivered a water-insoluble drug to the hepatocytes. Hence, they be used to deliver hydrophobic drugs to cure various liver diseases.     

Interaction of hepatocytes with polyelectrolyte complex(I): the effect of nonspecific interaction on adhesion of the cell

The adhesion of hepatocytes to polyelectrolyte complex (PEC), prepared by mixing of aqueous solutions of polycation and polyanion, is discussed. Four PECs, poly((dimethyliminio)ethylene(dimethyliminio)methylene‐1,4‐phenylenemethylene dichloride)–poly(acrylic acid)2X–(PAA), 2X‐poly(acrylic acid–co‐2‐ethylhexylacrylate(COA), 2X‐poly(acrylicacid–co‐butylacrylate) (CBA) and 2X‐poly(acrylic acid–co‐laurylacrylate)(CLA) were prepared. Hydrophobic properties of these PECs increased in the order of 2X–PAA < 2X–COA < 2X–CLA ≤ 2X–CBA. About all the hepatocytes adhered rapidly to various PECs even in the absence of serum, while fewer cells adhered to polystrene (tissue culture grade) dishes. At 37 °C (biological condition) about 70–80% of cells adhered to a collagen‐coated dish, but at 4 °C (nonbiological condition), no cell adhered to it. Nonactive cells (prepared by a single cycle of freezing and thawing) did not adhere to collagen, either. On the contrary about 40% of cells adhered to PEC‐coated dishes even at 4 °C, and nonactive cells also adhered to them. Cytocharosine B and colchicine, which are known as inhibitors of the polymerization of intracellular matrix, did not prevent cell adhesion to PECs. From these results it was suggested that hepatocytes adhered to PEC‐coated dishes mainly through a nonbiological interaction. Copyright © 1999 John Wiley & Sons, Ltd.     

Adsorption behaviors of poly(N-p-vinylbenzyl-4-o-beta-d-galactopyranosyl-[1-->4]-d-gluconamide) by quartz-crystal microbalance

Adsorption behaviors of amphiphilic poly(N-p-vinylbenzyl-4-o-beta-d-galactopyranosyl-[1-->4]-d-gluconamide) (PVLA) on the polystyrene (PS) surface was studied using 27 MHz quartz-crystal microbalance (QCM). The amount of adsorbed PVLA on PS surface was increased with an increase of PVLA concentration as a Langmuir-type in a monolayer. The saturated mass change (DeltaM(max)) and association constant (K(a)) of PVLA on PS surface were 498.6 ng/cm(2) and 1.93 x 10(7)M(-1), respectively. The adsorbed PVLA on PS surface was specifically recognized by Allo A lectin due to specific interaction between galactose moieties in the PVLA and Allo A. The hydrophobic interaction between hydrophobic main chain of PVLA and hydrophobic surface of PS was reduced in the presence of urea and the diameter of PVLA aqueous solution was decreased with an increase of urea concentration.     

Biopolymer/Glycopolypeptide‐Blended Scaffolds: Synthesis,Characterization and Cellular Interactions

Three‐dimensional (3D) scaffolds formed from natural biopolymers gelatin and chitosan that are chemically modified by galactose have shown improved hepatocyte adhesion, spheroid geometry and functions of the hepatocytes. Galactose specifically binds to the hepatocytes via the asialoglycoprotein receptor (ASGPR) and an increase in galactose density further improves the hepatocyte proliferation and functions. In this work, we aimed to increase the galactose density within the biopolymeric scaffold by physically blending the biopolymers chitosan and gelatin with an amphiphlic β‐galactose polypeptide (PPO‐GP). PPO‐GP, is a di‐block copolymer with PPO and β‐galactose polypeptide, exhibits lower critical solution temperature and is entrapped within the scaffold through hydrophobic interactions. The uniform distribution of PPO‐GP within the scaffold was confirmed by fluorescence microscopy. SEM and mechanical testing of the hybrid scaffolds indicated pore size, inter connectivity and compression modulus similar to the scaffolds made from 100 % biopolymer. The presence of the PPO‐GP on the surface of the scaffold was tested monitoring the interaction of an analogous mannose containing PPO‐GP scaffold and the mannose binding lectin Con‐A. In vitro cell culture experiments with HepG2 cells were performed on GLN‐GP and CTS‐GP and their cellular response was compared with GLN and CTS scaffolds for a period of seven days. Within three days of culture the Hep G2 cells formed multicellular spheroids on GLN‐GP and CTS‐GP more efficiently than on the GLN and CTS scaffolds. The multicellular spheroids were also found to infiltrate more in GLN‐GP and CTS‐GP scaffolds and able to maintain their round morphology as observed by live/dead and SEM imaging.     

Electrochemical evidence for asialoglycoprotein receptor – mediated hepatocyte adhesion and proliferation in three dimensional tissue engineering scaffolds

Asialoglycoprotein receptor (ASGPR) is one of the recognition motifs on the surface of hepatocytes, which promote their adhesion to extracellular matrix in liver tissue and appropriate artificial surfaces. ASGPR-mediated adhesion is expected to minimize trans-differentiation of hepatocytes in vitro that is generally observed in integrin-mediated adhesion. The aim of the present study is to verify the role of ASGPR in hepatocyte adhesion and proliferation in scaffolds for hepatic tissue engineering. Scanning Electrochemical Microscopy (SECM) is emerging as a suitable non-invasive analytical tool due to its high sensitivity and capability to correlate the morphology and activity of live cells. HepG2 cells and rat primary hepatocytes cultured in Polyvinyl alcohol (PVA)/Gelatin hydrogel scaffolds with and without galactose (a ligand for ASGPR) modification are studied using SECM. Systematic investigation of live cells cultured for different durations in scaffolds of different compositions (9:1 and 8:2 PVA:Gelatin with and without galactose) reveals significant improvement in cell–cell communication and proliferation on galactose incorporated scaffolds, thereby demonstrating the positive influence of ASGPR-mediated adhesion. In this work, we have also developed a methodology to quantify the respiratory activity and intracellular redox activity of live cells cultured in porous tissue engineering scaffolds. Using this methodology, SECM results are compared with routine cell culture assays viz., MTS ((1-Oxyl-2,2,5,5,-tetramethyl-Δ3-pyrroline-3-methyl) Methanethiosulfonate) and Albumin assays to demonstrate the better sensitivity of SECM. In addition, the present study demonstrates SECM as a reliable and sensitive tool to monitor the activity of live cells cultured in scaffolds for tissue engineering, which could be used on a routine basis.     

Polystyrene-coated micropallets for culture and separation of primary muscle cells

Despite identification of a large number of adult stem cell types, current primary cell isolation and identification techniques yield heterogeneous samples, making detailed biological studies challenging. To identify subsets of isolated cells, technologies capable of simultaneous cell culture and cloning are necessary. Micropallet arrays, a new cloning platform for adherent cell types, hold great potential. However, the microstructures composing these arrays are fabricated from an epoxy photoresist 1002F, a growth surface unsuitable for many cell types. Optimization of the microstructures’ surface properties was conducted for the culture of satellite cells, primary muscle cells for which improved cell isolation techniques are desired. A variety of surface materials were screened for satellite cell adhesion and proliferation and compared to their optimal substrate, gelatin-coated Petri dishes. A 1-μm thick, polystyrene copolymer was applied to the microstructures by contact printing. A negatively charged copolymer of 5% acrylic acid in 95% styrene was found to be equivalent to the control Petri dishes for cell adhesion and proliferation. Cells cultured on control dishes and optimal copolymer-coated surfaces maintained an undifferentiated state and showed similar mRNA expression for two genes indicative of cell differentiation during a standard differentiation protocol. Experiments using additional contact-printed layers of extracellular matrix proteins collagen and gelatin showed no further improvements. This micropallet coating strategy is readily adaptable to optimize the array surface for other types of primary cells.     

Synthetic,Chemically Defined Polymer‐Coated Microcarriers for the Expansion of Human Mesenchymal Stem Cells

Mesenchymal stem cells (MSC), also called marrow stromal cells, are adult cells that have attracted interest for their potential uses in therapeutic applications. There is a pressing need for scalable culture systems due to the large number of cells needed for clinical treatments. Here, a tailorable thin polymer coating—poly(poly(ethylene glycol) methyl ether methacrylate‐ran‐vinyl dimethyl azlactone‐ran‐glycidyl methacrylate) [P(PEGMEMA‐r‐VDM‐r‐GMA); PVG]—to the surface of commercially available polystyrene and glass microcarriers to create chemically defined surfaces for large‐scale cell expansion is applied. These chemically defined microcarriers create a reproducible surface that does not rely on the adsorption of xenogenic serum proteins to mediate cell adhesion. Specifically, this coating method anchors PVG copolymer through ring opening nucleophilic attack by amine residues on poly‐l ‐lysine that is pre‐adsorbed to the surface of microcarriers. Importantly, this anchoring reaction preserves the monomer VDM reactivity for subsequent functionalization with an integrin‐specific Arg‐Gly‐Asp peptide to enable cell adhesion and expansion via a one‐step reaction in aqueous media. MSCs cultured on PVG‐coated microcarriers achieve sixfold expansion—similar to the expansion achieved on PS microcarriers—and retain their ability to differentiate after harvesting.     

Synthesis of polymethacrylamides having a sugar moiety with an aliphatic hydrocarbon spacer and their application to control adhesion of hepatocytes cancer cells on the materials

Polymers having a sugar moiety in the side group have been utilized as artificial matrices for cell adhesion in tissue engineering. In this study, methacrylamide ‐ based polymers having lactose and maltose derivative structures in the side group with various aliphatic hydrocarbon spacers were synthesized, and their cell adhesion properties were examined. Methacrylamide monomers were prepared by two step amidation of a spacer diamine, first with a sugar lactone and then with a methacrylic anhydride. These monomers were radically polymerized in aqueous media using 4,4′‐azobis(4‐cyanovaleric acid) (ACVA) as radical initiator to give the corresponding polymethacrylamide. Specific interaction between these polymers and animal cell was investigated by adhesion of proliferated human liver cancer cell (WRL) to the polymethacrylamides. WRL interacted with polymers having a lactose structure with a hexamethylene or 1,4‐cyclohexylene spacer by a specific manner and was promoted typical spheroid formation, while it did not interacted with polymers having a maltose structure. © 2013 Wiley Periodicals, Inc. J. Polym. Sci., Part A: Polym. Chem. 2013 , 51, 4003–4010     

Synthesis and Cytotoxic Activity of Oxidized Galactomannan/ADR Conjugate

Abstract

Many polysaccharides are expected to apply as biomaterials because they generally show good biocompatibilities and biodegradabilities. It has recently been reported that the saccharides play important roles in biological recognition and the transmission of biological information on a cellar surface. Galactomannan (GalM) is a polysaccharide whose main chain is composed of β-1,4-linked mannose units only. It has some branching α-galactose residues at the C-6 position of mannose units. Therefore, it was of interest of us to use GalM as a drug carrier which was targeted to hepatocyte having a galactose receptor on its cellar surface. Dicarboxy-galactomannan (DC-GalM), which has reactive functional groups and is a carboxylic acid derivative of galactomannan, was prepared by IO^4-/CIO^2- oxidation of GalM. The obtained DC-GalM showed specific binding with maclura pomifera (MPA) [1] which has a specificity to α-galactose. Moreover, DC-GalM showed selective incorporation into hepatocyte. Adriamycine (ADR), which is one of the most prominent anticancer agents, was immobilized to DC-GalM. The DC-GalM/ADR conjugate showed specific cytotoxic activity against HepG2 human hepatoma cells which have a galactose receptor on the cell surface, compared with Hela utrocervical carcinoma cells which have no galactose receptor.     

Cell attachment and detachment on micropattern-immobilized poly(N-isopropylacrylamide) with gelatin

To investigate by microscopic observation the effects on cell behaviour of immobilized polymers, a micropattern-immobilization technique using a photo-mask was employed. Poly(acrylic acid) or poly(N-isopropylacrylamide-co-acrylic acid) was coupled with azidophenyl groups to form a photo-reactive polymer. The photo-reactive polymer was coated, with or without gelatin, on a cell-culture polystyrene plate and photo-irradiated through a micropatterned photo-mask. Mouse fibroblast STO cells were cultured on the micropattern-immobilized plate. The surface wettability of the immobilized plate was examined by measurement of the contact angle in the cell culture medium. The attachment of cells on the plate was significantly affected by the surface properties. Although the poly(acrylic acid) has the same effect on cell adhesion as a bare polystyrene surface, co-immobilization with gelatin significantly enhanced cell adhesion, while poly(N-isopropylacrylamide) reduced it. However, co-immobilization with gelatin enhanced cell adhesion, and, on the co-immobilized surface, cell detachment was observed by lowering the temperature. Micropattern immobilization was useful for comparing the effects of materials on cell behavior and for constructing biochips.     

Effect of end groups of poly(n-butyl methacrylate) on its biocompatibility

Telechelic poly(n-butyl methacrylate)s (PBMAs) with various end groups were prepared using nonionic, anionic, cationic or zwitterionic azo-type radical initiators and cell adhesion onto the surfaces of the polymers was investigated. The tendency for cell adhesion to the polymers differed with and without pretreatment with phosphate-buffered saline (PBS, pH 7.4). The cell adhesion to polymer surfaces without pretreatment was lower than that with pretreatment. The effect of pretreatment with PBS was significant for PBMA with ionic end groups. Furthermore, cell adhesion to the surface of PBMA with zwitterionic end groups was suppressed compared with that to the surfaces of other polymers. It was presumed that positive and negative charges of zwitterionic groups in the same molecule negated each other at pH 7.4 and that the polymers with zwitterionic end groups had no effective charges. The results clearly indicated that biocompatibility of polymers can be changed by the introduction of functional groups at the ends of the polymer chains. Fabrication of functional material surfaces will be anticipated by the similar method in the future.     

Synthesis and Cytotoxic Activity of Macromolecular Prodrug of Cisplatin Using Poly(ethylene glycol) with Galactose Residues or Antennary Galactose Units

To provide a macromolecular prodrug with recognition ability for hepatoma cells, we synthesized new conjugates of cisplatin (CDDP) and poly(ethylene glycol) (PEG) with galactose residues or antennary galactose units (Gal4A, four branched galactose residues) at the chain terminus, Gal‐PEG‐DA/CDDP or Gal4A‐PEG‐DA/CDDP conjugates. An antennary (branched) structure of Gal4A was designed based on the fact that saccharide clusters with branched structures show highly effective binding with saccharide receptors, a phenomenon known as the ‘cluster effect’. The cytotoxic activity of the conjugates was investigated against HepG2 human hepatoma cells in vitro and compared with a control conjugate without galactose, MeO‐PEG‐DA/CDDP. Gal‐PEG‐DA/CDDP and Gal4A‐PEG‐DA/CDDP conjugates showed lower IC₅₀ values (3.1×10^–4 and 2.3×10^–4 M , respectively) than the MeO‐PEG‐DA/CDDP conjugate (10.5×10^–4 M ). The cytotoxic activities of these conjugates with galactose residues or antennary galactose units were inhibited as a result of the addition of galactose and strongly inhibited by the addition of Gal4A, however the inclusion of a methoxy group (the MeO‐PEG‐DA/CDDP conjugate) did not affect the activity. These results suggest that the Gal4A unit introduced to the conjugate has effective recognition ability against HepG2 human hepatoma cells.     

Influence of sialic acid and bacterial sialidase on differential adhesion of Pseudomonas aeruginosa to epithelial cells

Epithelial cell lines from several tissues show a differential sensitivity to Pseudomonas aeruginosa adherence. A549 (lung), HepG2 (liver) and Caco-2 (colon) cells presented an adhesion index of about 3, 1.5 and 5 CFU/cell, respectively, whereas Mz-Ch cell lines (gallbladder cholangiocytes) presented adhesion indexes up to 35. These variations could be associated with the variable amount of sialic acid in cell surface glycoconjugates. Moreover, the presence of free sialic acid in culture media induces the secretion by P. aeruginosa of a sialidase which is able to hydrolyze glycoconjugate-linked sialic acid. As shown with A549 cells, this specific hydrolysis increases bacterial adhesion, probably by unmasking new binding sites onto the cell surface.     

Micropatterned designs of thermoresponsive surfaces for modulating cell behaviors

The chemistry and topography of the material surfaces have an important effect on cell behaviors. In this study, we reported the preparation of thermoresponsive micropatterned surfaces (TS) and galactosylated TS for modulating the adhesion/detachment of cells. A thickness of 1 µm of poly(N‐isopropylacrylamide) grafted layer was fabricated on the polystyrene surface with microgrooves using ultraviolet‐induced copolymerization. The thick grafted layer was in favor of the interactions between cells and materials. The following immobilization of galactose ligand with specific affinity to hepatocyte onto TS promoted the adhesion of human hepatocyte line (HL‐7702 cells). The microgrooves structure could facilitate cell adhesion and regulate the oriented growth of cells. Moreover, narrow grooves accelerated the spontaneous detachment of cells only by reducing temperature. Thus, micropatterned biofunctional designs with controlled geometrical features presented in this study have sufficient biofunctional activities in facilitating cell sheet engineering and regenerative medicine. Copyright © 2013 John Wiley & Sons, Ltd.     

Evaluation of boronate-containing polymer brushes and gels as substrates for carbohydrate-mediated adhesion and cultivation of animal cells

Boronate-containing thin polyacrylamide gels (B-Gel), polymer brushes (B-Brush) and chemisorbed organosilane layers (B-COSL) were prepared on the surface of glass slides and studied as substrates for carbohydrate-mediated cell adhesion. B-COSL- and B-Brush-modified glass samples exhibited multiple submicron structures densely and irregularly distributed on the glass surface, as found by scanning electron microscopy and atomic force microscopy. B-Gel was ca. 0.1 mm thick and contained pores with effective size of 1–2 μm in the middle and of 5–20 μm on the edges of the gel sample as found by confocal laser scanning microscopy. Evidence for the presence of phenylboronic acid in the samples was given by time-of-flight secondary ion mass-spectrometry (ToF SIMS), contact angle measurements performed in the presence of fructose, and staining with Alizarin Red S dye capable of formation specific, fluorescent complexes with boronic acids. A comparative study of adhesion and cultivation of animal cells on the above substrates was carried out using murine hybridoma M2139 cell line as a model. M2139 cells adhered to the substrates in the culture medium without glucose or sodium pyruvate at pH 8.0, and then were cultivated in the same medium at pH 7.2 for 4 days. It was found that the substrates of B-Brush type were superior both regarding cell adhesion and viability of the adhered cells, among the substrates studied. MTT assay confirmed proliferation of M2139 cells on B-Brush substrates. Some cell adhesion was also registered in the macropores of B-Gel substrate. The effects of surface microstructure of the boronate-containing polymers on cell adhesion are discussed. Transparent glass substrates grafted with boronate-containing copolymers offer good prospects for cell adhesion studies and development of cell-based assays.     

Effects of Serum on the Kinetics of CHO Attachment to Ultraviolet-Ozone Modified Polystyrene Surfaces

Attachment kinetics of Chinese hamster ovarian (CHO) cells were investigated on ultraviolet-ozone oxidized polystyrene (UVO-PS) dishes in the presence and absence of serum. The surface chemistry of UVO-PS has been extensively characterized. Although cells attached rapidly to the oxidized dishes with serum present it was found that serum actually inhibits the rate of attachment. Spreading of attached cells was favored by the presence of serum. It is suggested that the increased quantity of hydrophilic carboxyl groups on longer exposed UVO-PS leads to a change in the protein layer adsorbed from serum and also a higher affinity of the surface for extracellular proteins secreted by the attached cells. The UVO-PS surfaces present a new way of producing tissue culture grade polystyrene (TCPS) in a highly controllable method, which would ensure greater consistency in TCPS surfaces. Copyright 2001 Academic Press.     

Adhesion and proliferation of cells on new polymers modified biomaterials

Up to today, several techniques have been used to maintain cells in culture for studying many aspects of cell biology and physiology. More often, cell culture is dependent on proper anchorage of cells to the growth surface. Poly-l-lysine is commonly used as adhesive molecule. In this study, we present, as an alternative to poly-l-lysine, new polymer film substrates, realized by electropolymerization of different monomers on fluorine-doped tin oxide (FTO) surfaces since electropolymerization is a good method to coat selectively metallic or semiconducting electrodes with polymer films. So, the adhesion, proliferation and morphology of rat neuronal cell lines were investigated on polymer treated surfaces. Several amine-based biocompatible polymers were tested: polyethyleneimine (PEI), polypropyleneimine (PPI), polypyrrole (PPy) and poly(p-phenylenediamine) (PPPD). These polymer films were coated on FTO surfaces by electrochemical oxidation. After 8 h in a culture medium, a high percentage of cells was found to be attached to PEI and PPI compared to the other polymers and to the reference surfaces (glass and FTO uncovered). After 24 and 72 h in the culture medium, cells were found to proliferate faster on PEI and PPI than on other polymers and reference surfaces. Consequently, cells have a greater fold expansion on PEI and PPI than on PPPD, PPy or glass and FTO uncoated. From these results, we deduce that PEI and PPI can be useful as coating surface to cultivate neuronal cells.     

Binding of Ricinus communis agglutinin to a galactose-carrying polymer brush on a colloidal gold monolayer

A polymer with many pendent galactose residues was prepared by atom-transfer radical polymerization (ATRP) of galactose-carrying vinyl monomer, 2-lactobionamidoethyl methacrylate (LAMA), with a disulfide-carrying ATRP initiator, 2-(2'-bromoisobutyroyl)ethyl disulfide (DT-Br). The galactose-carrying polymer obtained (DT-PLAMA) was accumulated as a polymer brush via Au-S bond on a colloidal gold monolayer deposited on a cover glass. For comparison, a disulfide which carried one galactose residue at both ends (2-lactobionamidoethyl disulfide, Cys-Lac) was accumulated as a self-assembled monolayer (SAM) on the colloidal gold monolayer, too. The association and dissociation processes of galactose residues on the colloidal gold with a lectin, Ricinus communis agglutinin (RCA(120)), were observed by the increase and decrease in absorbance at 550nm corresponding to localized surface plasmon resonance (LSPR) phenomena. The Cys-Lac SAM-carrying glass chip showed a strong non-specific adsorption of the lectin, whereas the DT-PLAMA brush-carrying one reversibly associated with the lectin, indicating reusability of the latter device. The apparent association constant of the lectin with the galactose residues in the DT-PLAMA brush was much larger than the association constant for free galactose, and the detection limit of RCA(120) by the glycopolymer brush-modified device was satisfactorily low. Furthermore, a microscopic observation clearly indicated that the DT-PLAMA brush could reversibly associate with a HepG2 cell having galactose receptors, though these processes could not be observed spectrophotometrically due to a gigantic size of the cell.     

Protein Adsorption on Derivatives of Hyaluronan

Summary: Serum protein adsorption and fibroblast cell adhesion on photo reactive hyaluronic acid (Hyal-N₃) and its sulfated derivative (HyalS-N₃₎ was analysed using a combination of quartz crystal microbalance (QCM) and cell adhesion assays. There was no significant differences in the amount of protein adsorbed onto the two polymers, however proteins were found to be more loosely bound to HyalS-N₃ compared with Hyal-N₃. Approximately 17% and 31% of the fibronectin interacting with Hyal-N₃ and HyalS-N₃ respectively was found to be irreversibly bound after rinsing with MilliQ water, SDS and urea. Proteins were exposed to the polymers before cell adhesion was monitored for a period of 2 hours in serum free conditions. Minimal cell adhesion was observed on albumin-coated materials as well as serum precoated Hyal-N₃. Precoating the materials with fibronectin enhanced cell adhesion, although HyalS-N₃ experienced higher levels of cell adhesion than Hyal-N₃ and similar results were found for the serum precoated materials.