Cell outer membrane mimetic modification of a cross-linked chitosan surface to improve its hemocompatibility

A novel strategy has been developed to improve the hemocompatibility of chitosan surface by cell outer membrane mimetic structure able to reduce protein adsorption and cell adhesion. Phosphorylcholine dichloride was synthesized and grafted onto a glutaraldehyde-cross-linked chitosan (CS-GA) film surface to prepare phosphorylcholine-coated CS-GA film (CS-GA-PC) through a heterogeneous reaction process. The spectroscopic and contact angle characterization show that a cell outer membrane mimetic structure was formed on the cross-linked chitosan surface, and the significantly improved hemocompatibility of the modified surface was shown by a suppression of 94% on platelet adhesion, a suppression of 60–70% for bovine plasma fibrinogen and bovine serum albumin adsorptions. These results demonstrated that this cell outer membrane mimetic surface modification with phosphorylcholine dichloride is a promising strategy to improve the hemocompatibility of chitosan.     

Mitigated membrane fouling in an SMBR by surface modification

Fouling is a major obstacle in membrane processes applied in membrane bioreactor. To improve the antifouling characteristics of polypropylene hollow fiber microporous membranes (PPHFMMs) in a submerged membrane bioreactor (SMBR), the PPHFMMs were surface modified by NH₃, CO₂ plasma treatment, photoinduced graft polymerization of acrylamide and acrylic acid. The structural and morphological changes on the membrane surface were characterized by X-ray photoelectron spectroscopy (XPS), attenuated total reflection-Fourier transform infrared spectroscopy (FT-IR/ATR) and field emission scanning electron microscopy (FE-SEM). The change of surface wettability was monitored by contact angle measurements. The results of XPS and FT-IR/ATR clearly indicated the successful modification on the membrane surface. The static water contact angle of the modified membrane reduced obviously. The antifouling characteristics of the modified membranes in an SMBR were evaluated. The modified membranes showed better filtration performances in the submerged membrane bioreactor than the unmodified one, and the acrylic acid-grafted membrane presented the best antifouling characteristics. The results demonstrated that (1) the surface carboxyl-containing membranes were better than the surface amido-containing membranes; (2) surface-grafted membranes were better than the plasma-treated membranes.     

Surface modification of poly(tetramethylene adipate-co-terephthalate) membrane via layer-by-layer assembly of chitosan and dextran sulfate polyelectrolyte multiplayer

The improvement of hydrophilicity and hemocompatibility of poly(tetramethylene adipate-co-terephthalate) (PTAT) membrane was developed via polyelectrolyte multilayers (PEMs) immobilization. The polysaccharide PEMs included chitosan (CS, as a positive-charged and antibacterial agent) and dextran sulfate (DS, as a negative-charged and anti-adhesive agent) were successfully prepared using the aminolyzed PTAT membrane in a layer-by-layer (LBL) self-assembly manner. The obtained results showed that the contact angle of as-modified PTAT membranes reached to the steady value after four bilayers of coating, hence suggesting that the full coverage was achieved. It could be found that the PTAT–PEMs membranes with DS as the outmost layer could resist the platelet adhesion and human plasma fibrinogen (HPF) adsorption, thereby prolonging effectively the blood coagulation times. According to L929 fibroblast cell growth inhibition index, the as-prepared PTAT membranes exhibited non-cytotoxic. Overall results demonstrated that such an easy, valid and shape-independent processing should be potential for surface modification of PTAT membrane in the application of hemodialysis devices.     

Surface properties of Reactive Yellow 2 immobilised pHEMA and HEMA/chitosan membranes: characterisation of their selectivity to different proteins

Poly(hydroxyethyl methacrylate), pHEMA, and a composite pHEMA/chitosan networks were synthesized in the membrane form via UV initiated photo-polymerisation in the presence of an initiator ,′-azoisobutyronitrile. Reactive Yellow 2 (RY-2) was covalently immobilised as a dye–ligand onto both membranes. The polarity and surface energy of the investigated membranes were determined by contact angle measurement. The incorporation of chitosan in the pHEMA networks produced more hydrophilic surface, as indicated by contact angle analysis. The binding characteristics of lysozyme, γ-globulins, human serum albumin (HSA) and bovine serum albumin (BSA) to pHEMA-RY-2 and pHEMA/chitosan-RY-2 affinity membranes have been investigated from aqueous solution and their dye–ligand free forms were used as control systems. When chitosan was incorporated in the pHEMA network as a cationic polymer led to higher adsorption capacity for the lysozyme. Selective adsorption behaviour was also observed in the case of pHEMA/chitosan-RY-2 membrane for the lysozyme. The non-specific adsorptions of the lysozyme on the pHEMA and pHEMA/chitosan membranes were about 1.9 and 7.2 mg/ml, respectively. These were negligible for all others investigated proteins. The lysozyme adsorption data was analysed using the first-order and the second-order models. The first-order equation in both affinity membrane systems is the most appropriate equation to predict the adsorption capacities of the adsorbents. The adsorption isotherms well fitted the combined Langmuir–Freundlich model. A theoretical analysis has been conducted to estimate the thermodynamic contributions (changes in enthalpy, entropy and Gibbs free energy) for the adsorption of lysozyme to both dye–ligand immobilised membranes. The adsorption capacities of both dye–ligand immobilised membranes increased with increasing the temperature while decreased with increasing the NaCl concentration. Both affinity membranes are stable when subjected to sanitization with sodium hydroxide after repeated separation–elution cycles.     

Preparation and physical properties of chitosan-coated calcium sulphate whiskers

To improve the performance of calcium sulphate whiskers (CSWs) in the papermaking industry, a series of chitosan-coated (CS) calcium sulphate whiskers (CS-CSWs) was prepared, with the aim of effectively reducing their solubility in aqueous solutions. The CS-CSWs were prepared by immersing CSWs in a chitosan acid solution and then cross-linking with alkaline gel via hydrogen-bonding interaction. The CS-CSWs were characterised in terms of water solubility, zeta potential, contact angle, scanning electron microscopy, thermo gravimetric analysis and X-ray photoelectron spectroscopy (XPS). The results indicated that chitosan had a good effect on the surface modification of CSWs. Under optimal conditions, the water solubility of CS-CSWs decreased by 50 %, and the contact angle increased by 63 %. The XPS measurement indicated that the relative thickness of the chitosan coating was 9.8 nm.     

Physico-chemical and thermodynamic aspects of fibroblastic attachment on RGDS-modified chitosan membranes

In the present study, the cell attachment/spreading behaviour of L929 mouse fibroblasts on chitosan membranes was evaluated by using physico-chemical properties. For this purpose chitosan membranes were prepared and then photochemically modified with the cell adhesive peptide RGDS (Arg-Gly-Asp-Ser). The physico-chemical properties of unmodified (CHI) and RGDS-modified chitosan (CHI-RGDS) membranes were evaluated by calculating surface free energy (γ_sv) and interfacial free energy (γ_sw) values using captive bubble contact angle measurements and harmonic mean equation. The cell attachment experiments were performed both in 10% FBS containing and serum-free media with CHI and CHI-RGDS membranes. Eventually, it was not possible to predict a direct relationship between the change in physico-chemical properties and L929 cell attachment behaviour. The experimental results obtained from cell attachment agree with the theoretical prediction for the free energy of adhesion except for the cell attachment on CHI membrane in serum-free medium. Although a negative interfacial free energy of adhesion was calculated for CHI membrane in serum-free medium (ΔF_adh = −2.19 ergs/cm²), the cell attachment was poor (70%) compared to CHI-RGDS (90%) and none of the cells were spread on CHI surface to gain a fibroblastic morphology. Negative energy of adhesion was calculated for CHI and CHI-RGDS in 10% FBS medium, in which 100% of cells were attached on the membranes correlating with the thermodynamic approach. It can be suggested that, adsorption of serum proteins strongly affected the cell attachment meanwhile the presence of biosignal RGDS molecules triggered the cell spreading in serum medium.     

Surface Performance and Cytocompatibility Evaluation of Acrylic Acid-Mediated Carboxymethyl Chitosan Coating on Poly(tetrafluoroethylene-co-hexafluoropropylene)

To improve the biocompatibility of poly(tetrafluoroethylene-co-hexafluoropropylene) (FEP) film, a technique based on Ar plasma pretreatment and UV-induced grafting polymerization was used to immobilize carboxymethyl chitosan (CMCS) on the FEP film surfaces. Initially Ar plasma was used to treat FEP film. Then, plasma treated FEP film was modified via UV-induced grafting polymerization with hydrophilic acrylic acid (AAc) monomer. The following immobilization of CMCS on the FEP-pAAc surface was carried out via an amidation reaction. The change of chemical composition and surface morphology of FEP film were characterized by attenuated total reflectance Fourier transform infrared (ATR-FTIR), X-ray photoelectron spectroscopy (XPS), scanning electronic microscopy (SEM) and atomic force microscopy (AFM). Results of water contact angles measurement showed that the hydrophilicity of the surface has improved significantly after surface modification. Furthermore, methyl thiazolyl tetrazolium (MTT) assay and cell morphology analysis confirmed that mouse fibroblasts (L929 cells) attachment and proliferation were improved remarkably on the modified FEP surface. These results suggest that CMCS were successfully employed to surface engineering FEP film, and enhanced its cell biocompatibility. The approach presented here may be exploited for surface modification of biomaterials.     

Electrospinning of zein/chitosan composite fibrous membranes

<正>Zein/chitosan composite fibrous membranes were fabricated from aqueous ethanol solutions by electrospinning. Poly(vinyl pyrrolidone)(PVP) was introduced to facilitate the electrospinning process of zein/chitosan composites.The asspun zein/chitosan/PVP composite fibrous membranes were characterized by scanning electron microscopy(SEM) and tensile tests.SEM images indicated that increasing zein and PVP concentrations led to an increase in average diameters of the composite fibers.In order to improve stability in wet stage and mechanical properties,the composite fibrous membranes were crosslinked by hexamethylene diisocyanate(HDI).The crosslinked composite fibrous membranes showed slight morphological change after immersion in water for 24 h.Mechanical tests revealed that tensile strength and elongation at break of the composite fibrous membranes were increased after crosslinking,whereas Young's modulus was decreased.     

Action of chitosan against Xanthomonas pathogenic bacteria isolated from Euphorbia pulcherrima

The antibacterial activity and mechanism of two kinds of chitosan were investigated against twelve Xanthomonas strains recovered from Euphorbia pulcherrima. Results indicated that both chitosans markedly inhibited bacterial growth based on OD loss. Furthermore, the release of DNA and RNA from three selected strains was increased by both chitosans. However, the release of intracellular proteins was inhibited by both chitosans at different concentration and incubation times, except chitosan A at 0.1 mg/mL for 0.5 h incubation and 0.2 mg/mL for 2.0 h incubation increased the release of proteins, indicating the complexity of the interaction and cell membranes, which was affected by incubation time, bacterial species, chitosan type and concentration. Transmission electron microscopic observations revealed that chitosan caused changes in protoplast concentration and surface morphology. In some cells, the membranes and walls were badly distorted and disrupted, while other cells were enveloped by a thick and compact ribbon-like layer. The contrary influence on cell morphology may explain the differential effect in the release of material. In addition, scanning electron microscope and biofilm formation test revealed that both chitosans removed biofilm biomass. Overall, this study showed that membrane and biofilm play an important role in the antibacterial mechanism of chitosan.     

Surface Characterization and Antimicrobial Activity of Chitosan-Deposited DBD Plasma-Modified Woven PET Surface

In this work, a woven PET with an antimicrobial activity was prepared by depositing chitosan on its surface. Firstly, the hydrophilic property of the PET surface was achieved by a plasma treatment using dielectric barrier discharge (DBD). The hydrophilic property of the PET surface was characterized by wickability and contact angle measurements. The XPS analysis revealed an increment of oxygen-containing polar groups, such as C–O and O–C=O, on the PET surface after the plasma treatment, resulting in an enhanced hydrophilic property. The plasma-treated PET specimen was further deposited with chitosan by immersing in a chitosan acetate aqueous solution. The effects of temperature, chitosan concentration, and number of rinses on the amount of deposited chitosan on the PET surface were investigated. The disappearance of the above-mentioned polar groups from the PET surface was clearly observed after the chitosan deposition, indicating the involvement of these functional groups in interacting with the chitosan. The chitosan-deposited plasma-treated woven PET possessed an exceptionally high antimicrobial activity against both E. coli (gram-negative bacteria) and S. aureus (gram-positive bacteria).     

The effect of direct-current discharge treatment on the surface properties of a poly(1-(trimethylsilyl)-1-propyne) membrane

The effect of treatment in low-pressure direct-current discharge on the surface properties of poly(1-trimethylsilyl)-1-propyne) membranes was investigated by the goniometric technique. It was shown that the treatment of the membranes at the anode makes it possible to significantly improve their contact properties (to increase the hydrophilicity of the surface). The structure of the membranes before and after plasma treatment was examined by means of Fourier-transform IR reflection spectroscopy. In addition, the sorption of ethanol onto the plasma-treated and untreated membranes was studied with the use of this technique in the transmission mode and it was assumed that it proceeds via different mechanisms, a behavior that can be due to a change in the free volume of the polymer or the formation of a surface charge on the membrane as a result of plasma treatment.     

Cytocompatibility and antibacterial activity of a PHBV membrane with surface-immobilized water-soluble chitosan and chondroitin-6-sulfate

A water-soluble chitosan (WSC)/chondroitin-6-sulfate (ChS) polyelectrolyte complex (PEC) is covalently immobilized onto the surface of poly(3-hydroxybutyric acid-co-3-hydroxyvaleric acid) (PHBV) membranes via ozone-induced oxidation and poly(acrylic acid) (PAA) graft polymerization. To characterize the modified membranes, X-ray photoelectron spectroscopy (XPS) and water contact angle measurements are performed. It is shown that by coupling WSC as a spacer, the amount of ChS immobilized can be significantly increased. The water contact angle decreases with the amount of PAA, WSC, and ChS immobilized, which indicates the improving hydrophilicity. After WSC- and PEC-immobilization modification, the PHBV membranes possess antibacterial activity against S. aureus, E. coli, P. aeruginosa, and Methicilin resistant Staphylococus aureus (MRSA). According to the L929 fibroblast cell growth inhibition index, the as-prepared PHBV membranes are non-cytotoxic. In addition, the in-vitro evaluation of L929 fibroblast attachment, proliferation, and viability of PEC-immobilized PHBV membranes are ascertained to be superior to those of immobilized WSC or ChS alone. The overall results demonstrate that WSC/ChS PEC immobilization can not only improve the hydrophilicity and cytocompatibility of the PHBV membrane, but also endows antibacterial activity. [GRAPH: SEE TEXT] The bacterial survival ratio of as-prepared PHBV membranes (n=3).     

Temperature‐sensitive chitosan membranes as a substrate for cell adhesion and cell sheet detachment

A series of temperature‐sensitive poly(CSA‐co‐NIPAAm) membranes that were suitable for cell culture and confluent cell sheets detachment were prepared. The membranes with thermo‐responsive surface properties were synthesized by the copolymerization of acrylic acid‐derivatized chitosan (CSA) and N‐isopropylacrylamide (NIPAAm) in aqueous solution. Characterization of the membranes were carried out by means of the Fourier transform infrared (FTIR), X‐ray photoelectron spectroscopy (XPS), scanning electron microscope (SEM), and water contact‐angle (WCA) measurements. The adhesion and detachment of mouse fibroblast (L929) cells on these membranes have been investigated. The study showed that poly(CSA‐co‐NIPAAm) membranes could not only enhance fibroblasts attachment but also harvest confluent cell sheets by simply lowering the temperature. Furthermore, the detached cells retained high viability and could proliferate again after transferred to a new culture surface. Copyright © 2011 John Wiley & Sons, Ltd.     

Blood compatibility of thermoplastic polyurethane membrane immobilized with water-soluble chitosan/dextran sulfate

Water-soluble chitosan (WSC)/dextran sulfate (DS) was immobilized onto the surface of thermoplastic polyurethane (TPU) membrane after ozone-induced graft polymerization of poly(acrylic acid) (PAA). The surface was characterized with contact angle measurement and X-ray photoelectron spectroscopy (XPS). The adsorption of human plasma fibrinogen (HPF) followed the Langmuir adsorption isotherm. The results showed that the surface density of peroxides generated and poly(acrylic acid) (PAA) grafted reached the maximum value at 20 min of ozone treatment. It was found that the WSC- and DS-immobilized amount increased with pH and the molecular weight of WSC. The membrane/water interfacial free energy increased with PAA-grafting and WSC/DS-immobilization, indicating the increasing wettability of TPU membrane. The adsorption of HPF on TPU-WSC/DS membranes could be effectively curtailed and exhibited unfavorable adsorption. Moreover, WSC/DS immobilization could effectively reduce platelet adhesion and prolong the blood coagulation time, thereby membrane improving blood compatibility of TPU membrane. In addition, the in vitro cytotoxicity test of PEC modification was non-cytotoxic according to much low growth inhibition of L929 fibroblasts. Furthermore, TPU-WSC/DS membranes exhibited higher cell viability than native TPU membrane.     

Preparation and Characterization of Chitosan Composite Membranes Crosslinked by Carboxyl-capped Poly（ethylene glycol）

In this study a series of chemically crosslinked chitosan/poly（ethylene glycol） （CS/PEG） composite membranes were prepared with PEG as a crosslinking reagent other than an additional blend. First, carboxyl-eapped poly（ethylene glycol） （HOOC-PEG-COOH） was synthesized. Dense CS/PEG composite membranes were then prepared by casting/evaporation of CS and HOOC-PEG-COOH mixture in acetic acid solution. Chitosan was chemically crosslinked due to the amidation between the carboxyl in HOOC-PEG-COOH and the amino in chitosan under heating, as confirmed by FTIR analysis. The hydrophilicity, water-resistance and mechanical properties of pure and crosslinked chitosan membranes were characterized, respectively. The results of water contact angle and water absorption showed that the hydrophilicity of chitosan membranes could be significantly improved, while no significant difference of weight loss between pure chitosan membranes and crosslinked ones was detected, indicating that composite membranes with amidation crosslinking possess excellent water resistanance ability. Moreover, the tensile strength of chitosan membranes could be significantly enhanced with the addition of certain amount of HOOC-PEG-COOH crosslinker, while the elongation at break didn＇t degrade at the same time. Additionally, the results of swelling behaviors in water at different pH suggested that the composite membranes were pH sensitive.     

ZSM-5分子筛填充壳聚糖膜的制备与性能研究

采用液相共混的方法制备了ZSM-5分子筛填充壳聚糖膜.扫描电镜表征表明分子筛在膜中分散均匀,膜表面没有明显缺陷.考察了填充膜在碳酸二甲酯/甲醇混合液中的溶胀和吸附行为,探讨了填充膜中分子筛含量及操作温度对渗透汽化膜分离性能的影响.结果表明膜优先吸附甲醇,其分离性能主要由溶解过程控制;随着膜中分子筛含量的增加,膜的溶胀度增大,渗透通量大幅度提高;渗透通量与操作温度符合Arrhenius关系式.与壳聚糖均质膜相比,ZSM-5分子筛填充壳聚糖膜对甲醇和碳酸二甲酯混合物具有更好的分离效果.     

Growth control by cell to cell contact.

Control of cell growth by cell to cell contact is reviewed with particular emphasis on two systems--contact inhibition of growth observed with Swiss 3T3 cells and the mitogenic stimulation of Schwann cells by dorsal root ganglia neurites. In both cases the biological effect can be reproduced by the addition of surface membranes to the corresponding cells. In the case of contact inhibition of 3T3 cells, biological activity appears to correlate with membrane binding to the cells. An octylglucoside extract of 3T3 plasma membranes retains the biological activity (growth inhibition) of the original membranes.     

Penetration of pores in membranes by plasma polymer forming species

Penetration of low‐temperature plasma polymer forming species through a microporous polycarbonate membrane was indicated by advancing contact angle measurement and scanning electron microscopoy of the membrane surface. No penetration of plasma polymer forming species was found in nylon and poly(vinylidene fluoride) membranes. This is attributed to the tortuous pore shapes of the nylon and poly‐(vinylidene fluoride) membranes compared to the straight cylindrical pore shape of the polycarbonate membrane.     

Various approaches to modify biomaterial surfaces for improving hemocompatibility

In this paper, the mechanism of thrombus formation on the surface of polymeric materials and the various approaches of modifying biomaterial surfaces to improve their hemocompatibility are reviewed. Moreover, the blood compatibility of the cellulose membrane grafted with O-butyrylchitosan (OBCS) by using a radiation grafting technique was studied. Surface analysis of grafted cellulose membrane was verified by attenuated total reflection Fourier transform infrared spectroscopy (ATR-FTIR) and electron spectroscopy for chemical analysis (ESCA), which confirmed that OBCS was successfully grafted onto the cellulose membrane surfaces. Blood compatibility of the grafted cellulose membranes was evaluated by platelet rich plasma (PRP) contacting experiments and protein adsorption experiments using blank cellulose membranes as the control. The blood compatibility of OBCS grafted cellulose membranes is better than that of blank cellulose membranes. These results suggest that the photocrosslinkable chitosan developed here has the potential of serving in blood-contacting applications in medical use.     

Chitosan based surfactant polymers designed to improve blood compatibility on biomaterials

We developed chitosan based surfactant polymers that could be used to modify the surface of existing biomaterials in order to improve their blood compatibility. These polymers consist of a chitosan backbone, PEG side chains to repel non-specific protein adsorption, and hexanal side chains to facilitate adsorption and proper orientation onto a hydrophobic substrate via hydrophobic interactions. Since chitosan is a polycationic polymer, and it is thrombogenic, the surface charge was altered to determine the role of this charge in the hemocompatibility of chitosan. Charge had a notable effect on platelet adhesion. The platelet adhesion was greatest on the positively charged surface, and decreased by almost 50% with the neutralization of this charge. A chitosan surface containing the negatively charged SO(3)(-) exhibited the fewest number of adherent platelets of all surfaces tested. Coagulation activation was not altered by the neutralization of the positive charge, but a marked increase of approximately 5-6 min in the plasma recalcification time (PRT) was displayed with the addition of the negatively charged species. Polyethylene (PE) surfaces were modified with the chitosan surfactant resulting in a significant improvement in blood compatibility, which correlated to the increasing PEG content within the polymer. Adsorption of the chitosan surfactants onto PE resulted in approximately an 85-96% decrease in the number of adherent platelets. The surfactant polymers also reduced surface induced coagulation activation, which was indicated by the PEG density dependent increase in PRTs. These results indicate that surface modification with our chitosan based surfactant polymers successfully improves blood compatibility. Moreover, the inclusion of either negatively charged SO(3)(-) groups or a high density of large water-soluble PEG side chains produces a surface that may be suitable for cardiovascular applications.