Protein adsorption and cell adhesion on cationic,neutral, and anionic 2-methacryloyloxyethyl phosphorylcholine copolymer surfaces

Protein adsorption and cell adhesion on cationic, neutral, and anionic water-soluble 2-methacryloyloxyethyl phosphorylcholine (MPC) copolymer surfaces were compared. These model MPC copolymers coated SiO₂ surfaces exhibited comparable surface ζ-potentials of 26.1 mV, near 0 mV, and ?24.2 mV, respectively. X-ray photoelectron spectroscopy analyses indicated the similarities and the differences in the surface composition between the sample surfaces. Atomic force microscopy analyses revealed that the type of the charged moiety did not affect the surface roughness. Static contact angle measurements and dynamic contact angle analyses not only indicated that the surfaces were very hydrophilic in general, but also provided information on the surface mobility and the dominant role of MPC at the surface in aqueous conditions. Comparing with the SiO₂ substrates on which protein seriously adsorbed and cell heavily adhered, three MPC copolymers coated surfaces, despite their different charge properties, exhibited significantly low adsorbed amounts of different proteins having various electrical natures and totally no cell adhesion. This suggested that the incorporation of charged moieties in the MPC copolymers did not significantly inspire both the protein adsorption and cell adhesion. The MPC moieties were predominant at the surface when in contact with aqueous conditions and thereby dominated the bio-adsorptions, while the possible effect from electrostatic interactions would be too small and too limited to influence the overall situation. Therefore, these MPC copolymer surfaces can satisfy those biological applications requiring not only electrical but also non-biofouling properties.     

Nanoscale presentation of cell adhesive molecules via block copolymer self-assembly

Precise control over the nanoscale presentation of adhesion molecules and other biological factors represents a new frontier for biomaterials science. Recently, the control of integrin spacing and cellular shape has been shown to affect fundamental biological processes, such as differentiation and apoptosis. Here, we present the self-assembly of maleimide functionalised polystyrene-block-poly (ethylene oxide) copolymers as a simple, yet highly precise method for controlling the position of cellular adhesion molecules. By manipulating the phase separation of the functional PS-PEO block copolymer used in this study, via a simple blending technique, we alter the nanoscale (on PEO domains of 8–14 nm in size) presentation of the adhesion peptide, GRGDS, decreasing lateral spacing from 62 nm to 44 nm and increasing the number density from ～450 to ～900 islands per μm². The results indicate that the spreading of NIH-3T3 fibroblasts increases as the spacing between domains of RGD binding peptides decreases. Further, the same functional PS-PEO surfaces have been utilised to immobilise, via a zinc chelating peptide sequence, poly-histidine tagged proteins and extracellular matrix (ECM) fragments. This method is seen as an ideal platform for investigations into the role of spatial arrangements of cell adhesion molecules and ECM molecules on cell function and, in particular, control of cell phenotype.     

Adsorption state of fibronectin on poly(dimethylsiloxane) surfaces with varied stiffness can dominate adhesion density of fibroblasts

The state of adsorbed fibronectin and the subsequent cell adhesion behavior on polydimethylsiloxane (PDMS) substrates with varied stiffness were investigated. The bulk elastic modulus as well as the macroscale and nanoscale surface repulsion forces on PDMS substrates with five different cross-linker concentrations (2.5, 5, 10, 20 and 40 wt.%) were evaluated by using tensile and compression tests as well as atomic force microscopy (AFM) indentation. The PDMS substrate with 10 wt.% cross-linker showed the maximum stiffness in the bulk elastic modulus and macroscale compression test. In contrast, PDMS substrates with 2.5 and 5 wt.% cross-linker concentration showed the maximum stiffness in the nanoscale compression test, which indicates that the physical properties of the nanoscale outermost surface are different from the bulk and macroscale surface properties. The fibronectin-treated PDMS substrates showed almost the same amount of fibronectin adsorption. However, the outermost surface density of fibronectin was related to the macroscale surface stiffness, and the exposure of the cell-binding motif was related to the nanoscale surface stiffness. Moreover, the different adsorption state of fibronectin was further confirmed by quartz crystal microbalance-dissipation (QCM-D) monitoring. The adhesion behavior of NIH3T3 mouse fibroblasts was in turn related to the exposure of the cell-binding motif. These results suggest that the well-known differences in cell adhesion behavior on PDMS substrates with varied stiffness are primarily induced by different responses of fibronectin to the PDMS substrates.     

Discovery of low mucus adhesion surfaces

Mucus secretion from the body is ubiquitous, and finding materials that resist mucus adhesion is a major technological challenge. Here, using a high throughput platform with photo-induced graft polymerization, we first rapidly synthesized, screened and tested a library of 55 different surfaces from six functional monomer classes to discover porcine intestinal low mucus adhesion surfaces using a 1 h static mucus adsorption protocol. From this preliminary screen, two chemistries, a zwitterionic ([2-(acryloyloxy)ethyl] trimethylammonium chloride) and a multiple hydroxyl (N-[tris(hydroxymethyl)methyl]acrylamide) surface, exhibited significantly low mucus adhesion from a Langmuir-type isotherm when exposed to increasing concentrations of mucus for 24 h. Apolar or hydrophobic interactions were likely the dominant attractive forces during mucus binding since many polar or hydrophilic monomers reduced mucus adhesion. Hansen solubility parameters were used to illustrate the importance of monomer polarity and hydrogen bonding in reducing mucus adsorption. For a series of polyethylene glycol (PEG) monomers with changing molecular weight from 144 g mol^?1 to 1100 g mol^?1, we observed an excellent linear correlation (R² = 0.998) between relative amount adsorbed and the distance from a water point in a specialized Hansen solubility parameter plot, emphasizing the role of surface–water interactions for PEG modified surfaces.     

Oriented surfaces of adsorbed cellulose nanowhiskers promote skeletal muscle myogenesis

Cellulose nanowhiskers (CNWs) are high-aspect-ratio rod-like nanoparticles prepared via partial hydrolysis of cellulose. For the first time, CNWs have been extracted from the marine invertebrate Ascidiella aspersa, yielding animal-derived CNWs with particularly small diameters of only a few nanometres. Oriented surfaces of adsorbed CNWs were prepared using a flexible and facile spin-coating method, allowing the modulation of CNW adsorption and relative orientation. Due to the shape and nanoscale dimensions of the CNWs, C2C12 myoblasts adopted increasingly oriented morphologies in response to more densely adsorbed and oriented CNW surfaces. In addition, the degree of myoblast fusion was greatest on the highly oriented CNW surfaces, and even low-orientation CNW surfaces promoted more extensive fusion than flat control surfaces. Highly oriented multinuclear myotubes formed on the oriented CNW surfaces and fibrillar fibronectin deposited on the surfaces was also modelled in a highly oriented arrangement after only 4 days in culture. With a mean feature height of only 5–6 nm, the CNW surfaces present the smallest features ever reported to induce contact guidance in skeletal muscle myoblasts, highlighting the potential for nanoscale materials for engineering oriented tissues such as skeletal muscle.     

Effect of peptide secondary structure on adsorption and adsorbed film properties on end-grafted polyethylene oxide layers

Poly-l-lysine (PLL), in α-helix or β-sheet configuration, was used as a model peptide for investigating the effect of secondary structures on adsorption events to poly(ethylene oxide) (PEO) modified surfaces formed using θ solvents. Circular dichroism results showed that the secondary structure of PLL persisted upon adsorption to Au and PEO modified Au surfaces. Quartz crystal microbalance with dissipation (QCM-D) was used to characterize the chemisorbed PEO layer in different solvents (θ and good solvents), as well as the sequential adsorption of PLL in different secondary structures (α-helix or β-sheet). QCM-D results suggest that chemisorption of PEO 750 and 2000 from θ solutions led to brushes 3.8 ± 0.1 and 4.5 ± 0.1 nm thick with layer viscosities of 9.2 ± 0.8 and 4.8 ± 0.5 cP, respectively. The average number of H₂O per ethylene oxides, while in θ solvent, was determined as ∼0.9 and ∼1.2 for the PEO 750 and 2000 layers, respectively. Upon immersion in good solvent (as used for PLL adsorption experiments), the number of H₂O per ethylene oxides increased to ∼1.5 and ∼2.0 for PEO 750 and 2000 films, respectively. PLL adsorbed masses for α-helix and β-sheet on Au sensors was 231 ± 5 and 1087 ± 14 ng cm⁻², with layer viscosities of 2.3 ± 0.1 and 1.2 ± 0.1 cP, respectively; suggesting that the α-helix layer was more rigid, despite a smaller adsorbed mass, than that of β-sheet layers. The PEO 750 layer reduced PLL adsorbed amounts to ∼10 and 12% of that on Au for α-helices and β-sheets respectively. The PLL adsorbed mass to PEO 2000 layers dropped to ∼12% and 4% of that on Au, for α-helix and β-sheet respectively. No significant differences existed for the viscosities of adsorbed α-helix and β-sheet PLL on PEO surfaces. These results provide new insights into the fundamental understanding of the effects of secondary structures of peptides and proteins on their surface adsorption.     

Development and functional evaluation of biomimetic silicone surfaces with hierarchical micro/nano-topographical features demonstrates favourable in vitro foreign body response of breast-derived fibroblasts

Reproducing extracellular matrix topographical cues, such as those present within acellular dermal matrix (ADM), in synthetic implant surfaces, may augment cellular responses, independent of surface chemistry. This could lead to enhanced implant integration and performance while reducing complications. In this work, the hierarchical micro and nanoscale features of ADM were accurately and reproducibly replicated in polydimethylsiloxane (PDMS), using an innovative maskless 3D grayscale fabrication process not previously reported. Human breast derived fibroblasts (n = 5) were cultured on PDMS surfaces and compared to commercially available smooth and textured silicone implant surfaces, for up to one week. Cell attachment, proliferation and cytotoxicity, in addition to immunofluorescence staining, SEM imaging, qRT-PCR and cytokine array were performed. ADM PDMS surfaces promoted cell adhesion, proliferation and survival (p= < 0.05), in addition to increased focal contact formation and spread fibroblast morphology when compared to commercially available implant surfaces. PCNA, vinculin and collagen 1 were up-regulated in fibroblasts on biomimetic surfaces while IL8, TNFα, TGFβ1 and HSP60 were down-regulated (p= < 0.05). A reduced inflammatory cytokine response was also observed (p= < 0.05). This study represents a novel approach to the development of functionalised biomimetic prosthetic implant surfaces which were demonstrated to significantly attenuate the acute in vitro foreign body reaction to silicone.     

Electrically polarized HAp-coated Ti: In vitro bone cell–material interactions

Electrically polarized bulk sintered hydroxyapatite (HAp) compacts have been shown to accelerate mineralization and bone tissue ingrowth in vivo. In this work, a comprehensive study has been carried out to investigate the influence of surface charge and polarity on in vitro bone cell adhesion, proliferation and differentiation on electrically polarized HAp-coated Ti. Uniform and crack free sol–gel derived HAp coatings of 20 ± 1.38 μm thickness were polarized by application of an external d.c. field of 2.0 kV cm^?1 at 400 °C for 1h. In vitro bioactivity of polarized HAp coatings was evaluated by soaking in simulated body fluid, and bone cell–material interactions were studied by culturing with human fetal osteoblast cells (hFOB) for a maximum period of 11 days. Scanning electron microscopic observation showed that accelerated mineralization on negatively charged surfaces favored rapid cell attachment and faster tissue ingrowth over non-polarized HAp coating surfaces, while positive charge on HAp coating surfaces restricted apatite nucleation with limited cellular response. Immunochemistry and confocal microscopy confirmed that the cell adhesion and early stage differentiation were more pronounced on negatively charged coating surfaces as hFOB cells expressed higher vinculin and alkaline phosphatase proteins on negatively charged surface compared to cells grown on all other surfaces. Our results in this study are process independent and potentially applicable to any other commercially available coating techniques.     

Impact of nanopore morphology on cell viability on mesoporous polymer and carbon surfaces

Topography at the nanoscale can lead to dramatic changes in the adhesion of cells to surfaces and their subsequent viability. For biological applications, including tissue engineering and cell-based sensing, the large internal surface area of ordered mesoporous carbons provides an opportunity for enhanced sensitivity and performance, but the mesostructure also affects the topography of the material. In this work, we probe the viability and adhesion of osteoblasts on ordered mesoporous materials with different morphologies and matrix chemistries. FDU-15 (hexagonal) and FDU-16 (cubic) films were processed at either 350 °C (polymeric) or 800 °C (carbon) to provide these different materials. For the films processed at 350 °C, the cell adhesion was markedly improved on the mesoporous films in comparison to a dense film analog, consistent with many reports in the literature that nanostructuring of surfaces improves the viability and adhesion of osteoblasts. Conversely, osteoblast adhesion was reduced on the carbonized surfaces processed at 800 °C when ordered mesopores were introduced, particularly for the cubic mesostructure (FDU-16). We attribute the decrease in cell adhesion to the propensity of the ordered mesoporous carbon films to sorb organics from aqueous solution, which could lead to removal of adhesion-promoting compounds at the film surface. These results suggest that cell viability on mesoporous polymer and carbon films can be controlled through simple changes in the pyrolysis temperature.     

Studies on bound water restrained by poly(2-methacryloyloxyethyl phosphorylcholine): Comparison with polysaccharide–water systems

The structural change of water restrained by poly(2-methacryloyloxyethyl phosphorylcholine) (PMPC) was investigated by differential scanning calorimetry (DSC), since the biocompatibility of PMPC and related biopolymers is affected by the structure of water on the polymer surface. The phase transition behaviour of PMPC–water systems with a water content (W_c = mass of water/mass of dry sample, g g^?1) in the range 0–2.0 was measured in the temperature range ?150 to 50 °C. Glass transition, cold crystallization and melting were observed. Cold crystallization, which has been suggested as an index of biocompatibility, was detected for PMPC with a W_c in the range 0.5–0.9. The amounts of two types of bound water, non-freezing water and freezing bound water, were calculated from the melting enthalpy. The amount of non-freezing water of PMPC was ～0.48. It was found that the phase transition behaviour and amount of bound water of PMPC were quite similar to those of water-soluble polysaccharide electrolytes. The results indicate that the bound water, not the free water, is restrained by PMPC.     

Human insulin adsorption kinetics,conformational changes and amyloidal aggregate formation on hydrophobic surfaces

The formation of insulin amyloidal aggregates on material surfaces is a well-known phenomenon with important pharmaceutical and medical implications. Using surface plasmon resonance imaging, we monitor insulin adsorption on model hydrophobic surfaces in real time. Insulin adsorbs in two phases: first, a very fast phase (less than 1 min), where a protein monolayer forms, followed by a slower one that can last for at least 1 h, where multilayered protein aggregates are present. The dissociation kinetics reveals the presence of two insulin populations that slowly interconvert: a rapidly dissociating pool and a pool of strongly bound insulin aggregates. After 1 h of contact between the protein solution and the surface, the adsorbed insulin has practically stopped dissociating from the surface. The conformation of adsorbed insulin is probed by attenuated total reflection–Fourier transform infrared spectroscopy. Characteristic shifts in the amide A and amide II′ bands are associated with insulin adsorption. The amide I band is also distinct from that of soluble or aggregated insulin, and it slowly evolves in time. A 1708 cm^?1 peak is observed, which characterizes insulin adsorbed for times longer than 30 min. Finally, Thioflavin T, a marker of extended β-sheet structures present in amyloid fibers, binds to adsorbed insulin after 30–40 min. Altogether, these results reveal that the conformational change induced in insulin upon binding to hydrophobic surfaces allows further insulin binding from the solution. Adsorbed insulin is thus an intermediate along the α-to-β structural transition that results in the formation of amyloidal fibers on these material surfaces.     

Cyclodextrin-functionalized biomaterials loaded with miconazole prevent Candida albicans biofilm formation in vitro

Polyethylene (PE) and polypropylene (PP) were functionalized at their surfaces with cyclodextrins (CDs) in order to prevent the adhesion and proliferation of Candida albicans on medical devices made from these polymers. The surface functionalization involved the grafting of glycidyl methacrylate (GMA) after oxidative γ-ray pre-irradiation, followed by the attachment of β-CD and HP-β-CD to PE-g-GMA and PP-g-GMA surfaces. The yield of CD functionalization directly depended on the amount of GMA grafted. The presence of CDs on the surface of the polymers did not compromise their cell compatibility, but remarkably changed their protein adsorption profile. In contrast to unmodified PE and PP that adsorb significant amounts of fibrinogen (～0.047 mg cm^?2) but not albumin, the CD-modified polyethers promoted the adsorption of albumin (between 0.015 and 0.155 mg cm^?2) and reduced the adsorption of fibrinogen. Furthermore, functionalization with CDs provided PE and PP with the capability to incorporate the anti-fungal drug miconazole (up to 0.27 mg cm^?2), leading to reduced biofilm formation by C. albicans in an in vitro biofilm model system. Overall, the results of the work indicate that the novel approach for functionalization of PE and PP is potentially useful to reduce the likelihood of foreign body-related infections.     

Two amino acid-based superlow fouling polymers: Poly(lysine methacrylamide) and poly(ornithine methacrylamide)

We developed and investigated two new antifouling zwitterionic polymers, poly(lysine methacrylamide) (pLysAA) and poly(ornithine methacrylamide) (pOrnAA), both derived from natural amino acids – lysine and ornithine, respectively. The pLysAA and pOrnAA brushes were grafted on gold via the surface-initiated photoiniferter-mediated polymerization, with the polymer film thickness controlled by the UV-irradiation time. Nonspecific adsorption from human blood serum and plasma was investigated by surface plasmon resonance. Results show that the adsorption level decreased with the increasing film thickness. With the thin films of ∼14.5 nm, the minimal adsorption on pLysAA was 3.9 ng cm⁻² from serum and 5.4 ng cm⁻² from plasma, whereas the lowest adsorption on pOrnAA was 1.8 and 3.2 ng cm⁻², from serum and plasma, respectively. Such protein resistance is comparable to other widely reported antifouling surfaces such as poly(sulfobetaine methacrylate) and polyacrylamide, with a much thinner polymer film thickness. Both pLysAA and pOrnAA showed better protein resistance than the previously reported serine-based poly(serine methacrylate), whereas the pOrnAA is the best among three. The pLysAA- and pOrnAA-grafted surfaces also highly resisted the endothelial cell attachment and Escherichia coli K12 bacterial adhesion. Nanogels made of pLysAA and pOrnAA were found to be ultrastable in undiluted serum, with no aggregation observed after culturing for 24 h. Dextran labeled with fluorescein isothiocyanate (FITC–dextran) was encapsulated in nanogels as a model drug. The encapsulated FITC–dextran exhibited controlled release from the pOrnAA nanogels. The superlow fouling, biomimetic and multifunctional properties of pLysAA and pOrnAA make them promising materials for a wide range of applications, such as implant coating, drug delivery and biosensing.     

The influence of poly(ethylene oxide) grafting via siloxane tethers on protein adsorption

Amphiphilic PEO–silanes (a–c) having siloxane tethers of varying lengths with the general formula α-(EtO)₃Si–(CH₂)₂–oligodimethylsiloxane_n-block-poly(ethylene oxide)₈–OCH₃ [n = 0 (a), n = 4 (b), and n = 13 (c)] were grafted onto silicon wafers and resistance to adsorption of plasma proteins was measured. Distancing the PEO segment from the hydrolyzable triethoxysilane [(EtO)₃Si] grafting group by a oligodimethylsiloxane tether represents a new method of grafting PEO chains to surfaces. Properties of surfaces grafted with a–c were compared to surfaces grafted with a traditional PEO–silane containing a propyl spacer [(EtO)₃Si–(CH₂)₃–poly(ethylene oxide)₈–OCH₃, PEO control]. As the siloxane tether length increased, chain density of PEO–silanes grafted onto oxidized silicon wafers decreased and hydrophobicity of the PEO–silane increased which led to a decrease in surface hydrophilicity. Despite decreased surface hydrophilicity, resistance to the adsorption of bovine serum albumin (BSA) increased in the order: PEO control < a < b ≈ c and to human fibrinogen (HF) increased in the order: PEO control < a < b < c.     

Surface modification with an antithrombin–heparin complex for anticoagulation: Studies on a model surface with gold as substrate

Gold was used as a substrate for immobilization of an antithrombin–heparin (ATH) covalent complex to investigate ATH as a surface modifier to prevent blood coagulation. Three different surface modification methods were used to attach ATH to gold: (i) direct chemisorption; (ii) using dithiobis(succinimidyl propionate) (DSP) as a linker molecule and (iii) using polyethylene oxide (PEO) as a linker/spacer. The ATH-modified surfaces were compared to analogous heparinized surfaces. Water contact angles and X-ray photoelectron spectroscopy confirmed the modifications and provided data on surface properties and possible orientation. Ellipsometry measurements showed that surface coverage of DSP and PEO was high. ATH and heparin densities were quantified using radioiodination and quartz crystal microbalance, respectively. The surface density of ATH was greatest on the DSP surface (0.17 μg cm^?2) and lowest on the PEO (0.05 μg cm^?2). The low uptake on the PEO surface was likely due to the protein resistance of the PEO component. Using radioiodinated antithrombin (AT), it was shown that ATH-immobilized surfaces bound significantly greater amounts from both buffer and plasma than the analogous heparinized surfaces. Immunoblot analysis of proteins adsorbed from plasma demonstrated that surfaces chemisorbed with PEO, whether or not subsequently modified with ATH, inhibited non-specific adsorption. The immunoblot response for AT was stronger on the DSP–ATH than on the heparin surfaces, thus confirming the results from radiolabelling. The ATH surfaces again showed higher selectivity for AT binding than analogous heparin-modified surfaces, indicating the enhanced anticoagulant potential of ATH for biomaterial surface modification.     

Astrocytes specifically remove surface-adsorbed fibrinogen and locally express chondroitin sulfate proteoglycans

Surface-adsorbed fibrinogen (FBG) was recognized by adhering astrocytes, and was removed from the substrates in vitro by a two-phase removal process. The cells removed adsorbed FBG from binary proteins’ surface patterns (FBG + laminin, or FBG + albumin) while leaving the other protein behind. Astrocytes preferentially expressed chondroitin sulfate proteoglycan (CSPG) at the loci of fibrinogen stimuli; however, no differences in overall CSPG production as a function of FBG surface coverage were identified. Removal of FBG by astrocytes was also found to be independent of transforming growth factor type β (TGF-β) receptor based signaling as cells maintained CSPG production in the presence of TGF-β receptor kinase inhibitor, SB 431542. The inhibitor decreased CSPG expression, but did not abolish it entirely. Because blood contact and subsequent FBG adsorption are unavoidable in neural implantations, the results indicate that implant-adsorbed FBG may contribute to reactive astrogliosis around the implant as astrocytes specifically recognize adsorbed FBG.     

Study of cellular dynamics on polarized CoCrMo alloy using time-lapse live-cell imaging

The physico-chemical processes and phenomena occurring at the interface of metallic biomedical implants and the body dictate their successful integration in vivo. Changes in the surface potential and the associated redox reactions at metallic implants can significantly influence several aspects of biomaterial/cell interactions such as cell adhesion and survival in vitro. Accordingly, there is a voltage viability range (voltages which do not compromise cellular viability of the cells cultured on the polarized metal) for metallic implants. We report on cellular dynamics (size, polarity, movement) and temporal changes in the number and total area of focal adhesion complexes in transiently transfected MC3T3-E1 pre-osteoblasts cultured on CoCrMo alloy surfaces polarized at the cathodic and anodic edges of its voltage viability range (?400 and +500 mV (Ag/AgCl), respectively). Nucleus dynamics (size, circularity, movement) and the release of reactive oxygen species (ROS) were also studied on the polarized metal at ?1000, ?400 and +500 mV (Ag/AgCl). Our results show that at ?400 mV, where reduction reactions dominate, a gradual loss of adhesion occurs over 24 h while cells shrink in size during this time. At +500 mV, where oxidation reactions dominate (i.e. metal ions form, including Cr⁶⁺), cells become non-viable after 5 h without showing any significant changes in adhesion behavior right before cell death. Nucleus size of cells at ?1000 mV decreased sharply within 15 min after polarization, which rendered the cells completely non-viable. No significant amount of ROS release by cells was detected on the polarized CoCrMo at any of these voltages.     

Volumetric interpretation of protein adsorption: Kinetics of protein-adsorption competition from binary solution

The standard solution-depletion method is implemented with SDS-gel electrophoresis as a multiplexing, separation-and-quantification tool to measure competition between two proteins (i and j) for adsorption to the same hydrophobic adsorbent particles (either octyl sepharose or silanized glass) immersed in binary-protein solutions. Adsorption kinetics reveals an unanticipated slow protein-size-dependent competition that controls steady-state adsorption selectivity. Two sequential pseudo-steady-state adsorption regimes (State 1 and State 2) are frequently observed depending on i, j solution concentrations. State 1 and State 2 are connected by a smooth transition, giving rise to sigmoidally-shaped adsorption-kinetic profiles with a downward inflection near 60 min of solution/adsorbent contact. Mass ratio of adsorbed i, j proteins (m_i/m_j) remains nearly constant between States 1 and 2, even though both m_i and m_j decrease in the transition between states. State 2 is shown to be stable for 24 h of continuous-adsorbent contact with stagnant solution whereas State 2 is eliminated by continuous mixing of adsorbent with solution. In sharp contrast to binary-competition results, adsorption to hydrophobic adsorbent particles from single-protein solutions (pure i or j) exhibits no detectable kinetics within the timeframe of experiment from either stagnant or continuously mixed solution, quickly achieving a single steady-state value in proportion to solution concentration. Comparison of binary competition between dissimilarly-sized protein pairs chosen to span a broad molecular-weight (MW) range demonstrates that selectivity between i and j scales with MW ratio that is proportional to protein-volume ratio (ubiquitin, Ub, MW = 10.7 kDa; human serum albumin, HSA, MW = 66.3 kDa; prothrombin, FII, 72 kDa; immunoglobulin G, IgG, MW = 160 kDa; fibrinogen, Fib, MW = 341 kDa). Results are interpreted in terms of a kinetic model of adsorption that has protein molecules rapidly diffusing into an inflating interphase that is spontaneously formed by bringing a protein solution into contact with a physical surface (State 1). State 2 follows by rearrangement of proteins within this interphase to achieve the maximum interphase concentration (dictated by energetics of interphase dehydration) within the thinnest (lowest volume) interphase possible by ejection of interphase water and initially-adsorbed proteins. Implications for understanding biocompatibility are discussed using a computational example relevant to the problem of blood–plasma coagulation.     

Synthesis,characterization and surface modification of low moduli poly(ether carbonate urethane)ureas for soft tissue engineering

Flexible scaffolds are of great interest in engineering functional and mechano-active soft tissues as such scaffolds might allow mechanical stimuli to transfer effectively from the scaffolds to cells during tissue development. Towards this end, we have developed a family of flexible poly(ether carbonate urethane)ureas (PECUUs) with a triblock copolymer poly(trimethylene carbonate)–poly(ethylene oxide)–poly(trimethylene carbonate) (PTMC–PEO–PTMC) or pentablock copolymers PTMC–PEO–PPO–PEO–PTMC (PPO, polypropylene oxide) as soft segments, linked by 1,4-diisocyanatobutane and putrescine. All of the PECUUs had low glass transition temperatures (<?46 °C). The PTMC–PEO–PTMC-containing PECUUs had low tensile strength and breaking strain. Replacing PEO with the similar length PEO–PPO–PEO resulted in highly flexible and soft PECUUs possessing breaking strains of 362–711%, tensile strengths of 8–18 MPa and moduli of 5.5–7.4 MPa at room temperature in air. Under aqueous conditions at 37 °C, these polymers remained flexible while their moduli were decreased to 3.4–4.0 MPa. PECUUs based on PTMC–PEO–PPO–PEO–PTMC were thermosensitive as the water content at 37 °C was lower than that at 4 °C. PECUU using PTMC–PEO–PTMC as a soft segment showed 30% weight loss over 6 weeks in PBS at 37 °C, while that using PTMC–PEO–PPO–PEO–PTMC as a soft segment had weight loss <6%. Degradation products were found to lack cytotoxicity. The mechanical stresses and moduli of PECUUs based on PTMC–PEO–PPO–PEO–PTMC were unchanged during the degradation. To enhance cell adhesion, PECUUs were surface modified with Arg-Gly-Asp-Ser (RGDS). Smooth muscle cell adhesion was 114% of tissue culture polystyrene for unmodified PECUU and >180% for RGDS-modified PECUUs, with cell viability on both surfaces increasing during culture. These low moduli polyurethanes may find applications in engineering cardiovascular or other soft tissues.     

Electrostatic control of protein adsorption on UV-photofunctionalized titanium

Ultraviolet (UV)-photofunctionalization of titanium to enable the establishment of a nearly complete bone–implant contact was reported recently. However, the underlying mechanism for this is unknown. We hypothesized that UV-treated titanium surfaces acquire distinct electrostatic properties that may play important roles in determining the bioactivity of these surfaces. The objective of this study was to determine the protein adsorption capability of UV-treated titanium surfaces under various electrostatic environments. The amount of albumin adsorbed on UV-treated and untreated titanium disks was evaluated under different pH conditions above and below the isoelectric points of albumin and titanium. The effects of additional treatment with various ionic solutions were also examined. Albumin adsorption on UV-treated surfaces at pH 7.0 was considerably greater (6-fold after 3 h of incubation and 2.5-fold after 24 h) than that to UV-untreated surfaces. UV-enhanced albumin adsorption was abrogated at pH 3.0 or when these titanium surfaces were treated with anions, while maintaining UV-induced superhydrophilicity. Albumin adsorption on UV-untreated titanium surfaces increased after treating these surfaces with divalent cations but not after treating them with monovalent cations. These results indicated that UV-treated titanium surfaces are electropositively charged as opposed to electronegatively charged UV-untreated titanium surfaces. This distinct UV-induced electrostatic property predominantly regulates the protein adsorption capability of titanium, superseding the effect of hydrophilic status, and converts titanium surfaces from bioinert to bioactive. As a result, direct titanium–protein interactions take place exclusively on UV-treated titanium surfaces without the aid of bridging ions.