Membranes from acrylonitrile-based polymers for selective cultivation of human keratinocytes

A cell carrier made from synthetic material supporting selective growth of keratinocytes is a promising approach to avoid the phenomenon of fibroblast overgrowth during in vitro culture of skin substitutes. Therefore, we investigated polymer membranes made of polyacrylonitrile and copolymers of acrylonitrile and N-vinylpyrrolidone (NVP) for their ability to support selectively the growth of keratinocytes. It was found that a copolymer with an NVP-content of 30% (NVP30) supports growth of human keratinocyte cell line (HaCaT) cells and inhibits fibroblast growth under serum-containing conditions. Cell proliferation of HaCaT cells was measured over 14 days. If both cell types were cultured under serum-free conditions for initial adhesion over 6 h on these NVP30 polymers, they adhered to the same extent. Long-term experiments over 7 days were performed as a coculture of both cell types showing that HaCaT cells had a growth advantage that seems to be related to the paracrine activity of contaminating fibroblasts. As a result, confluent layers of HaCaT cells were obtained with small numbers of remaining fibroblasts. The new poly [acrylonitrile-co(NVP) membranes seem to be a promising culture system for the production of epidermal transplants.     

Study of a (trimethylenecarbonate-co-epsilon-caprolactone) polymer--part 2: in vitro cytocompatibility analysis and in vivo ED1 cell response of a new nerve guide

Future surgical strategies to restore neurological function in peripheral nerve loss may involve replacement of nerve tissue with cultured Schwann cells using biodegradable guiding implants. Random copolymers of trimethylene carbonate and epsilon caprolactone (P(epsilonCL-TMC), 50: 50) have been synthesized by ring opening polymerization using rare earth alkoxides as initiator. Their potential use as nerve guide repairs has been assessed through indirect and direct in vitro biocompatibility tests and in vivo soft tissue response to EDI subclass macrophages. In vitro, we exposed monolayers of human skin fibroblasts and an established continuous cell line (Hela) to liquid extracts (either pure or diluted in the culture medium) of epsilonCL-TMC copolymer including positive (phenol) and negative controls. Then, colorimetric assays (Neutral red and MTT) were performed. The extracts of epsilonCL-TMC induced no significant cytotoxic effect. We also exposed in vitro Schwann cells to pieces of P(epsilonCL-TMC) and P(LA-GA) copolymers. We evaluated cell attachment at 1 and 3 h by measuring the activity of the lysosomal enzyme (N-acetyl-beta-hexosaminidase) and cell proliferation at 1, 3, 6 and 9 days by measuring the cell metabolic activity (MTT assay). Values for attachment slightly decreased between 1 and 3 h but were significantly higher than on agars (negative control). Cells plated on epsilonCL-TMC showed a rate of proliferation comparable with that of normalized controls and higher than on PGA-PLA at day 9. Finally, we evaluated in vivo the soft tissue response after implantation of cylindrical tubes of P(epsilonCL-TMC) and P(LA-GA) copolymers with an immunohistochemistry staining procedure for the newly recruited ED1 macrophages. An image analysis system automatically measured the optical density of labelled positive ED1 cells at 9, 21 and 60 days after implantation. epsilonCL-TMC copolymer showed a mild soft tissue reaction with no adverse chronic inflammatory reaction. These data allowed us to consider this conduit as a potential effective substitute in nerve repair. El sevier Science Ltd. All rights reserved.     

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.     

Effect of argon-plasma treatment on proliferation of human-skin-derived fibroblast on chitosan membrane in vitro

Chitosan is not only a nontoxic, biocompatible, and biodegradable polymer, but has also a chemical structure similar to glycosaminoglycans (GAGs), which promote scarless wound healing of skin. In this study, chitosan membranes were treated with argon plasma to improve their surface hydrophilicity. The results showed that the water contact angles of these surface-treated membranes were significantly reduced from 60.76 to 11.57 degrees . The total surface energy was increased from 41.06 to 67.31 mJ/m(2), with 60-86.95% improvement in the gamma-negative component and a 20% difference in the nonpolar component. Argon-plasma-treated chitosan membranes exhibited excellent attachment, migration, and proliferation of the human-skin-derived fibroblasts (hSFs) compared to the untreated ones. It was found that the duration of argon-plasma treatment influenced the cell proliferation, and the optical densities in MTT assay were enhanced. Argon-plasma treatment improved the surface hydrophilicity of chitosan membranes and promoted the attachment and proliferation of hSFs.     

Development of a stable chemically defined surface for the culture of human keratinocytes under serum-free conditions for clinical use

Within the field of tissue engineering there is a need to develop new approaches to achieve effective wound closure in patients with extensive skin loss or chronic ulcers. This article exploits the well-known interdependency of epithelial keratinocytes and stromal fibroblasts in conjunction with plasma surface technology. The aim was to produce a chemically defined surface, which with the aid of a feeder layer of lethally irradiated dermal fibroblasts would improve the attachment and proliferation of the keratinocyte cell from which subconfluent cells can be transferred to wound bed models. Plasma copolymers of acrylic acid/octa-1,7-diene have been prepared and characterized by X-ray photoelectron spectroscopy. The fibroblasts and keratinocytes were cultured on plasma polymer-coated 24-well plates. Cell attachment and proliferation were assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide-eluted stain assay (MTT-ESTA) and DNA assay. Attachment and proliferation of both cell types on plasma polymer surfaces were compared with tissue culture plastic and collagen I, plus a negative control of a pure hydrocarbon layer. A pure acrylic acid surface, fabricated at a power of 10 W and containing 9.2% carboxylate groups, was found to promote both fibroblast and keratinocyte attachment and proliferation and permit the serum-free coculture of keratinocytes and irradiated fibroblasts.     

The different behaviors of skeletal muscle cells and chondrocytes on PEGT/PBT block copolymers are related to the surface properties of the substrate

The attachment, proliferation, morphology, and differentiation of two cell types-skeletal muscle cells and chondrocytes-were investigated on different compositions of poly(ethylene glycol) and poly(butylene terephthalate) segmented block copolymers. Four weight percentages (40, 55, 60, and 70%) and two different molecular weights (300 and 1000 Da) of poly(ethylene glycol) were tested. Varying the weight percentage and molecular weight of poly(ethylene glycol) resulted in different behaviors for skeletal muscle cells and chondrocytes. The attachment of skeletal muscle was the highest (similar to tissue culture polystyrene) when copolymers containing 55 wt % of poly(ethylene glycol) were used, regardless of the poly(ethylene glycol) molecular weight. Maximum proliferation and differentiation of skeletal muscle cells was achieved when copolymers containing 55 wt % and 300 Da molecular weight of poly(ethylene glycol) were used. In contrast, the weight percentage and molecular weight of poly(ethylene glycol) had no significant effect on chondrocyte attachment and proliferation; the attached chondrocytes retained a differentiated phenotype only when a 70 wt % of poly(ethylene glycol) was used. Cell behavior was correlated with the surface properties of the copolymer films, as indicated by contact-angle measurements. These results suggest that an optimized wt % and molecular weight of poly(ethylene glycol) will be useful depending on the specific cell type.     

Cytocompatibility of poly(acrylonitrile-co-N-vinyl-2-pyrrolidone) membranes with human endothelial cells and macrophages

Polyacrylonitrile modified with N-vinyl-2-pyrrolidone (NVP) shows good hemocompatibility. This work, which aims to evaluate the cytocompatibility of membranes fabricated from poly(acrylonitrile-co-N-vinyl-2-pyrrolidone) (PANCNVP), studied the adhesion of macrophages and endothelial cell (EC) cultures. It was found that PANCNVP membranes with higher NVP content decreased the adhesion of both macrophages and ECs. Compared with polyacrylonitrile and tissue culture polystyrene control, however, these PANCNVP membranes promoted the proliferation of ECs. Furthermore, the viability of ECs cultured on the PANCNVP membrane surfaces was also relatively competitive. Both static and dynamic water contact angle measurements were conducted to explain the nature of cell adhesion to the PANCNVP membranes. On the basis of these results and the phenomena of water swelling and water states reported previously, it was presumed that the coexistence of large amounts of bound water and free water induced by NVP moieties are responsible for the lower adhesion and better function of cells adhering to the PANCNVP membranes.     

Electrical stimulation enhances viability of human cutaneous fibroblasts on conductive biodegradable substrates

Electrically conductive biodegradable polymer membranes were prepared by mixing conductive polypyrrole particles with poly(L-lactic acid) solution followed by solution casting and solvent evaporation. Multi-well electrical cell culture plates were fabricated to host electrically stimulated cell culture and monitor parameters. Human cutaneous fibroblasts were cultured on conductive membranes with or without electrical stimulation (ES). Cell count, MTT, Hoechst staining, and SEM were performed to characterize the cells. The membranes supported the adhesion and proliferation of the fibroblasts in both the presence and absence of ES. In the presence of direct electrical field strength of 100 mV/mm, cell viability on the PPy/PLLA membranes at 2 and 24 h was 2.2- and 4.0-fold (p < 0.05) respectively of that on the same membranes without ES. Direct electrical current ranging from 2.5 to 250 microA/mm had no effect on the viability of cells cultured on the gold-coated Petri dish. Electrical field applied to conductive biodegradable polymer surfaces is therefore an effective approach to upregulate the mitochondrial activity of human skin fibroblasts.     

Novel adhesion prevention membrane based on a bioresorbable copoly(ester-ether) comprised of poly-L-lactide and Pluronic: in vitro and in vivo evaluations.

Block copolymers consisting of poly(L-lactide) (PLLA) and poly(oxyethylene-co-oxypropylene), with various compositions, were synthesized and characterized in vitro and in vivo for their application as postoperative adhesion prevention membranes. It was found that the flexibility and degradability of the cast films of the block copolymers grew with increasing Pluronic F68 [PN; poly(oxyethylene-co-oxypropylene] composition. The receding contact angle of the copolymer films against water became lower than that of the PLLA film, because the surface was predominantly covered with more hydrophilic PN segments in a wet state. This surface property significantly affects the cell attachment property of the copolymer films, and the fibroblasts cultured on the films exhibit a spheroid-like morphology. The copolymer films subcutaneously implanted in the back of rats induced milder tissue responses compared with PLLA homopolymers, because of the increased surface hydrophilicity in the former. In vivo evaluation using a uterus horn model in rats revealed that the performance of these copolymer films as an adhesion-prevention membrane is comparable to that of a conventionally utilized membrane of oxidized regenerated cellulose. These results indicate that the copolymer films are biocompatible materials with controllable mechanical properties and biodegradability as adhesion-prevention membranes.     

Adsorbed poly(ethyleneoxide)-poly(propyleneoxide) copolymers on synthetic surfaces: spectroscopy and microscopy of polymer structures and effects on adhesion of skin-borne bacteria

Poly(ethyleneoxide)-copoly(propyleneoxide) (PEO-PPO) polymer coatings were evaluated for their resistance to the attachment of the marker organism Serratia marcescens and the skin-borne bacteria Staphylococcus epidermidis. The copolymers were adsorbed onto poly(styrene) films-chosen as simplified physicochemical models of skin surfaces-and their surface characteristics probed by contact angle goniometry, attenuated total reflectance-Fourier transform infrared (ATR-FTIR), atomic force microscopy (AFM), and X-ray photoelectron spectroscopy (XPS). These functional surfaces were then presented to microbial cultures, bacterial attachment was assessed by fluorescence microscopy and AFM, and the structures of the polymer films examined again spectroscopically. Surface characterization data suggest that the adsorbed copolymer was partially retained at the surface and resisted bacterial attachment for 24 h. Quantitative evaluation of cell attachment was carried out by scintillation counting of (14)C-labeled microorganisms in conjunction with plate counts. The results show that a densely packed layer of PEO-PPO copolymer can reduce attachment of skin commensals by an order of magnitude, even when the coating is applied by a simple adsorptive process. The work supports the hypothesis that adhesion of microorganisms to biological substrates can be reduced if a pretreatment with an appropriate copolymer can be effected in vivo.     

Tissue engineering of fish skin: behavior of fish cells on poly(ethylene glycol terephthalate)/poly(butylene terephthalate) copolymers in relation to the composition of the polymer substrate as an initial step in constructing a robotic/living tissue hybrid

This study presents the development of a biosynthetic fish skin to be used on aquatic robots that can emulate fish. Smoothness of the external surface is desired in improving high propulsive efficiency and maneuvering agility of autonomous underwater vehicles such as the RoboTuna (Triantafyllou, M., and Triantafyllou, G. Sci. Am. 272, 64, 1995). An initial step was to determine the seeding density and select a polymer for the scaffolds. The attachment and proliferation of chinook salmon embryo (CHSE-214) and brown bullhead (BB) cells were studied on different compositions of a poly(ethylene glycol terephthalate) (PEGT) and poly(butylene terephthalate) (PBT) copolymer (Polyactive). Polymer films were used, cast of three different compositions of PEGT/PBT (weight ratios of 55/45, 60/40, and 70/30) and two different molecular masses of PEGT (300 and 1000 Da). When a 55 wt% and a 300-Da molecular mass form of PEGT was used, maximum attachment and proliferation of CHSE-214 and BB cells were achieved. Histological studies and immunostaining indicate the presence of collagen and cytokeratins in the extracellular matrix formed after 14 days of culture. Porous scaffolds of PEGT/PBT copolymers were also used for three-dimensional tissue engineering of fish skin, using BB cells. Overall, our results indicate that fish cells can attach, proliferate, and express fish skin components on dense and porous Polyactive scaffolds.     

Modulation of marrow stromal cell function using poly(D,L-lactic acid)-block-poly(ethylene glycol)-monomethyl ether surfaces.

The adhesion of marrow stromal osteoblasts and the adsorption of fetal bovine serum (FBS) proteins to end-capped poly(D,L-lactic acid) 50:50 (PLA50) of molecular weight 17,000 (PLA5017), non-end-capped PLA50 of molecular weight 11,000 (PLA5011h), and a diblock copolymer made of poly(ethylene glycol)-monomethyl ether of molecular weight 5,000 and PLA50 of molecular weight 20,000 (Me. PEG5-PLA20) were investigated. Cell attachment and proliferation on both PLA50 polymers were equally good. The block copolymer did not allow the proliferation of cells. However, the attached cells were highly differentiated and metabolically active in contrast to the cells on PLA50. Moreover, surface analysis studies using electron spectroscopy revealed that FBS proteins adsorbed well from aqueous solutions to the PLA50 surfaces while they adsorbed substantially less to the block copolymer. These results suggest that Me.PEG-PLA block copolymers may be used to regulate protein adsorption and, therefore, cell adhesion by varying the block composition of the copolymer.     

Study of Comicellization of Diblock Copolymers in a Selective Solvent

Summary: The size, aggregation number and other quantities which characterize the formation of micelles in a bidisperse solution of block copolymers which are chemically identical but have different block asymmetries has been studied in a selective solvent using LS and SANS techniques. Experimental results for the comicellization of two chemically identical block copolymers (high molecular weight, PS‐PI, and a low molecular weight dPS‐PI with a deuterated PS block) and two slightly incompatible copolymers (high molecular weight PS‐HPI, and dPS‐PI) in decane, a selective solvent for PI, are reported. Individually, each of the PS‐HPI and PS‐PI copolymers formed micelles with a PS core, which were stable over the entire temperature range covered by the experiments (25–70 °C). The small copolymer, dPS‐PI, formed micelles with a highly swollen core at low temperatures and was molecularly dispersed above the critical micelle temperature (47 °C). Comicellization was observed for both pairs of copolymers studied over a broad range of mixing ratios. The mixed micelles were comprised of large copolymer (PS‐HPI or PS‐PI) and a portion of the small copolymer (dPS‐PI). The amount of adsorbed dPS‐PI copolymer increased with increasing weight fraction of this copolymer and decreased with increasing temperature. The critical comicellization temperature was estimated as 70 and 60 °C for the comicelles based on PS‐HPI and PS‐PI copolymers, respectively. The SANS data suggest that the small copolymer does not form a shell coating the PS core of the initial micelle, but is instead incorporated into the core where it forms a layer of mixed dPS and PS chains.

     

Films of dextran-graft-polybutylmethacrylate to enhance endothelialization of materials

We have synthesized new structures obtained from amphiphilic copolymers of dextran and polybutylmethacrylate with the aim of endothelialization of biomaterials. Grafting of butylmethacrylate onto dextran has been carried out using ceric ammonium nitrate as initiator. Three copolymers were obtained (11, 30 and 37 wt.% dextran) and homogeneous thin films were successfully prepared. In contrast to dextran, the resulting films were stable in water, and copolymers characterized by Fourier transform infrared spectroscopy, differential scanning calorimetry and dynamic mechanical analysis showed evidence of hybrid properties between the parent homopolymers. Surfaces of films were smooth when analyzed by atomic force microscopy (roughness 2 ± 1 nm) but greatly differed in their hydrophilicity by increasing the dextran content (water contact angle from 99° to 57°). In contrast to polybutylmethacrylate, where the proliferation of vascular smooth muscle cells (VSMCs) was excellent but that of endothelial cells was very low, the copolymer containing 11% of dextran was excellent for endothelial cells but very limited for VSMCs. An in vitro wound assay demonstrated that copolymer with 11% dextran is even more favorable for endothelial cell migration than tissue-culture polystyrene. Increasing the dextran content in the copolymers decreased the proliferation for both vascular cell types. Altogether, these results show that transparent and water-insoluble films made from copolymers of dextran and butylmethacrylate copolymers with an appropriate composition could enhance endothelial cell proliferation and migration. Therefore, a potential benefit of this approach is the availability of surfaces with tunable properties for the endothelialization of materials.     

Cell interactions with perfluoropolyether-based network copolymers

We have investigated the potential of several polymers based on perfluoropolyether (PFPE) macromonomers for use in biomaterial applications. Polymer networks were synthesised from the PFPE macromonomers of increasing chain length and the adhesion and proliferation of corneal, vascular and bone cells was evaluated on these polymers. The polymer surfaces were quite hydrophobic, having sessile air-water contact angles ranging between 96 and 125 degrees. However, these polymers supported the attachment and growth of bovine corneal epithelial and endothelial cells and fibroblasts at 60-100% of the rate of cell growth on the culture substratum, TCPS. Furthermore, the PFPE polymers supported the attachment and growth of vascular endothelial cells (from human umbilical artery) and human bone-derived cells over a 7 day period at an equal level to TCPS. The relationship between the macromonomer chain length (n = 1 to 4) and the ability of the resulting PFPE homopolymer to support the overgrowth of corneal epithelial tissue was also evaluated. The PFPE-containing polymers supported corneal epithelial tissue overgrowth, with the most effective having a performance equivalent to that of TCPS. In addition to these homopolymers, copolymers comprising of PFPE and N,N-dimethylaminoethyl methacrylate (DMAEMA) were also synthesised. Surprisingly, the addition of DMAEMA to the PFPE polymer network lead to a reduction in the growth and attachment of corneal epithelial cells and fibroblasts. These results indicate that PFPE-based materials show a potential for use in the development of biomaterials in the ocular, vascular and orthopaedic areas.     

Cellular responses to electrospun membranes made from blends of PLLGA with PEG and PLLGA-b-PEG

Control of cellular responses is crucial for the use of electrospun membranes in biomedical applications, including tissue engineering or biomedical devices. However, it is still unclear whether adhesion and proliferation of fibroblasts is stimulated or inhibited on polyethylene glycol (PEG)-modified electrospun membranes. In this study, poly(L-lactide-co-glycolide) (PLLGA)-PEG copolymer and pure PEG were blended with PLLGA, and then electrospun onto nonwoven membranes. The effects of blending of PLLGA-PEG or pure PEG on the adsorption of proteins, and further on the adhesion and proliferation of L929 fibroblasts on the electrospun membranes were investigated. Addition of PLLGA-PEG or PEG significantly improved the hydrophilicity of the electrospun membranes. Pure PEG had no obvious effects on the growth of L929 fibroblasts; in contrast, PLLGA-PEG significantly inhibited the adsorption of proteins and the proliferations of the cells on the electrospun membranes. In response to diminished protein adsorption, mRNA expression of genes related to cell adhesion and migration was up-regulated. The limited effects of pure PEG were probably caused by its preferential dissolution, whereas membrane-confined PLLGA-PEG displayed excellent performance on the inhibition of protein adsorption and cell proliferation. ? 2012 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 100A:2897-2904, 2012.     

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.     

Attachment,proliferation and collagen type I mRNA expression of human gingival fibroblasts on different biodegradable membranes

Abstract

The purpose of this study was to investigate adhesion, proliferation and type I collagen (COL I) mRNA expression of gingival fibroblasts on different membranes used in periodontal applications. Collagen (C), acellular dermal matrix (ADM) and polylactic acid; polyglycolic acid; lactide/glycolide copolymer (PLGA) biodegradable membranes were combined with gingival fibroblasts in culture and incubated for 48?h. Cell adhesion was examined with scanning electron and confocal microscopy. MTT assay was used to measure proliferation. COL I mRNA expression was assessed using quantitative-polymerase chain reaction (QPCR). The PLGA group exhibited the lowest cell survival on day 5 and 10, and lowest cell proliferation on days 5, 10 and 14. While cell proliferation was similar in C and ADM groups, the C membrane showed a slightly greater increase in viable cells to day 10. Confocal and scanning electron microscopy confirmed the results of proliferation and MTT assays. The highest COL I mRNA expression was noted in the PLGA membrane group when compared to the C (p?<?0.01) and ADM (p?<?0.05) membrane groups. These data revealed that adherence and proliferation of primary gingival fibroblasts on collagen-based C and ADM membranes is better than that seen with PLGA membranes, and thus may be preferable in the treatment of gingival recession defects.     

Hydroxyethyl methacrylate-methyl methacrylate (HEMA-MMA) copolymers for cell microencapsulation: effect of HEMA purity

Thermoplastic copolymers of 2-hydroxyethyl methacrylate (HEMA) and methyl methacrylate (MMA) (molar ratio: 75/25 HEMA-MMA) were synthesized using HEMA containing different amounts of ethylene glycol dimethacrylate (EGDMA) to investigate their suitability for cell microencapsulation. Pure HEMA (0.0% EGDMA) was obtained with preparative chromatography to prepare a linear copolymer. Microcapsules (with a diameter of 300-400 microm) were readily made with the copolymers by interfacial precipitation. Smaller and more transparent capsules were obtained using the copolymer prepared from purer HEMA. Chinese hamster ovary (CHO) fibroblasts, as model cells, were microencapsulated in the linear copolymer. The CHO cells survived the microencapsulation process and the metabolic activity of the encapsulated cells increased within the 14 days observation period.     

Co-culture of bone marrow fibroblasts and endothelial cells on modified polycaprolactone substrates for enhanced potentials in bone tissue engineering

The creation of a vascularized bed makes the survival of seeded cells on 3-dimensional scaffolds much more likely. However, relying purely on random capillary ingrowth into the porous scaffolds from the host may compromise vascularization of a scaffold. One solution is to transplant cells capable of differentiating into new blood vessels into the scaffolds to accelerate the creation of a vascularized scaffold. Because endothelial cells are the key cells involved in blood vessel formation, the present study was designed to investigate the development of a biomaterial surface that supports endothelial cell attachment and proliferation. The subsequent effects of the material surface modifications on the differentiation and proliferation of human bone marrow-derived fibroblasts (HBMFs) when grown in co-culture with a human bone marrow endothelial cell line (HBMEC-60) were studied. Endothelialization studies showed that the gelatin-coated and hydroxyapatite-coated substrates were superior for HBMEC-60 attachment and proliferation to hydrolyzed-only or untreated polycaprolactone substrates. Co-culture studies showed that the presence of the HBMEC-60 specifically enhanced HBMF cell proliferation and differentiation and that this effect was not observed with co-culture with skin fibroblasts. It is concluded that the co-culture of endothelial cells with HBMFs could be a promising culture system for bone tissue- engineering applications.