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1

Improved adhesion and growth of vascular smooth muscle cells on polycaprolactone nanofibrous membranes modified by amine-rich plasma

Parizek M., Blahova L., Michlicek M., Medalova J., Cernochova P., Bacakova L., Zajickova L.

Engineering of Biomaterials

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2018

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Vol. 21, no. 148

117

2

Black Orlon as promising material for bone tissue engineering

Parizek M., Vetrik M., Hruby M., Lisa V., Bacakova L.

Engineering of Biomaterials

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2014

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Vol. 17, no. 128-129

4--6

3

Improved adhesion and growth of osteoblast-like MG-63 cells in cultures on titanium modified by gold particles

Parizek M., Base T., Hruby M., Lisa V., Bacakova L.

Engineering of Biomaterials

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2013

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Vol. 16, no. 122-123 spec. iss.

77

EN

Metallic materials are important for load-bearing bone implants. The osteointegration of these implants can be improved by appropriate surface modifications. Therefore, we present here a study of the cell growth on titanium surfaces modified with films created from gold microparticles. These particles in the form of microplates or polyhedral microcrystals were deposited on titanium plates from ethanol solutions, dried and annealed with a hydrogen flame. Some samples were additionally modified by polyethylene imine. The materials engendered from these modifications were used to investigate the adhesion and growth of human osteoblast-like MG-63 cells on these surfaces in the DMEM medium with 10% of fetal bovine serum. One day after seeding, the highest number of initially adhered cells was found on the surfaces modified by both types of gold microparticles. This trend was the same three and seven days after seeding. The numbers of cells on pure Ti and Ti modified only with gold particles were significantly higher than on samples which were modified with polyethylene imine. The cell spreading areas projected on the materials were significantly larger in cells on the samples with polyethylene imine modification. However, the shape of these cells was mostly rounded or star-like with thin and long protrusions, while on the materials without polyethylene imine, it was mostly polygonal. The cell proliferation activity was estimated from XTT test, based on the activity of mitochondrial enzymes. This test showed that the proliferation activities of osteoblast-like MG-63 cells of the 3rd and 7th days of the experiment were more pronounced on the samples modified only by gold microparticles. Immunofluorescence showed that the focal adhesion plaques containing vinculin and the fibers containing β-actin were most apparent, more numerous and more brightly stained in cells on Ti modified by gold microplates and gold polyhedral microcrystals, especially in comparison with the corresponding samples modified with polyethylene imine (Fig. 1). Thus, it can be concluded that the modification of titanium samples by both types of gold microparticles enhanced the adhesion and growth of MG 63 cells.

4

The adhesion and growth of human osteoblast-like MG 63 cells in cultures on titanium modified with gold microparticles and polyethylene imine)

Parizek M., Base T., Hruby M., Lisa V., Bacakova L.

Engineering of Biomaterials

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2012

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Vol. 15, no. 116-117 spec. iss.

149--150

EN

Metallic materials are indispensable for construction of surgical implants, particularly those designed for load-bearing application, such as the bone-anchoring parts of big joint replacements. For good osteointegration, long-term function, durability and also mechanical and chemical resistance of the implant, the physical and chemical properties of the material surface are of a great importance. These properties can be favorably influenced by coating the bone-anchoring parts of the implants with appropriate biocompatible and bioactive films. Therefore, in this study, we have investigated the adhesion and growth of human osteoblast-like MG 63 cells in cultures on titanium substrates coated with films made of gold microparticles and/or poly(ethylene imine) (PEI). Gold microparticles were chosen for good biocompatibility of gold and absence of its cytotoxicity, which has been proved by numerous studies performed in vitro and in vivo [1,2]. When deposited on the material surface, these microparticles provide this surface with microstructure, which has been reported to enhance the osteogenic differentiation of bone-derived cells. On the other hand, the material surface microroughness has dual effect on the cell adhesion, spreading and proliferation - some studies reported the enhancement, other the reduction of these events (for a review, see [3,4]). This suggests that not only the size of the microscale irregularities, but also their shape should be taken into account. Therefore, in our study, gold microparticles were used in the form of plates or polyhedral crystals [5]. These microparticles were deposited on square samples of Ti (1x1 cm, thickness 1 mm) and annealed with a hydrogen flame. As for PEI, this polymer has been used as precursor base layer for further functionalization of metallic substrates, particularly with polyelectrolyte multilayer films [6] or biomolecules such as gelatin, hyaluronan or chitosan [7,8]. Other reason for the PEI deposition was creation of an intermediate layer which would compensate the differences in mechanical properties (e.g., hardness, toughness, specific weight) between a metallic implant and bone tissue. PEI was deposited either on pure or on gold microparticle-coated Ti samples. The materials were sterilized with 70% ethanol (1 hour), inserted into 24-well polystyrene plates (well diameter 1.5 cm; TPP, Switzerland) and seeded with human osteoblast-like MG 63 cells (30 000 cells/well, i.e. 17 000 cells/cm2). Each well contained 1.5 ml of a medium DMEM with 10% of fetal bovine serum and 40 /jg/ml of gentamicin. On days 1, 3 and 7 after seeding, the cell number and morphology were evaluated. For evaluating the cell number, the cells were trypsinized and counted in Bürker hemocytometer. For evaluating the cell morphology, i.e. the cell shape and the size of cell spreading area, the cells were fixed with 70% ethanol (-20°C, 10 min) and stained with a combination of fluorescence dyes Texas Red C2-maleimide, which stains the cell membrane and cytoplasm, and Hoechst #33342, which stains the cell nuclei. The microphotographs of cells were taken using an Olympus IX 51 microscope equipped with a DP 70 digital camera, and the cell spreading area was measured on these pictures using a software Atlas (Tescan, Brno, Czech Rep.) One day after seeding, the highest number of initially adhered cells was found on the surface modified by gold polyhedral crystals. This trend was the same on days 3 and 7 after seeding (FIG.1,2). However, the cell number on Ti modified with gold plates was significantly lower than on Ti with polyhedral crystals. Nevertheless, the numbers of cells on Ti samples coated with gold microparticles without PEI were significantly higher than on PEI-coated samples. Also the cell spreading areas were significantly larger on the samples without PEI. The cells on the samples without PEI were mostly polygonal, while the cells on PEI-coated samples were of star-like appearance, i.e. with multiple long protrusions (FIG.2). This is in accordance with findings published by other authors, documenting cytotoxic effects of PEI, particularly that of a high molecular weight [6], which was also used in our study (m.w. 750 kDa). Nevertheless, this cytotoxicity was considerably reduced by further functionalization of PEI with biomolecules, such as gelatin, hyaluronan or chitosan [7,8]. Thus, it can be concluded that the modification of titanium plates by gold microparticles supported the adhesion and growth of MG 63 cells. In this context, the polyhedral crystals were more advantageous than plates. The effects of PEI coatings on cell behavior need further investigation.

5

Application of cellulose-based biomaterials in vascular tissue engineering - a review and our experience

Bacakova L., Novotna K., Parizek M., Havelka P., Sopuch T.

Engineering of Biomaterials

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2012

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Vol. 15, no. 116-117 spec. iss.

128--130

EN

Artificial vascular replacements used in current clinical practice are fabricated from polyethylene terephthalate (PET, e.g. Dacron) orpolyterafluoroethylene (PTFE, e.g. Teflon). Older materials used earlier for constructing vascular prostheses are polyamide (Nylon), polyvinyl alcohol (Ivalon) and polyacrylonitrile (Orlon). New promising materials include polyurethane and a wide range of biodegradable synthetic or nature-derived polymers, which are usually designed as temporary scaffolds for vascular cells forming a new regenerated blood vessel wall (for a review, see [1]). One of the nature-derived polymers is cellulose and its derivatives and composites with other materials. Cellulose is the most abundant biopolymer on Earth. It is a polysaccharide consisting of a linear chain of several hundred to over ten thousand ß(1\to 4) linked D-glucose units [2,3]. Cellulose is the structural component of the primary cell wall of green plants, many forms of algae and the oomycetes. In plant cells, cellulose microfibrils are synthesized at the plasma membrane by hexameric protein complexes, also known as cellulose synthase complexes [4]. Some species of bacteria secrete cellulose to form biofilms. For industrial use, cellulose is mainly obtained from wood pulp and cotton. For tissue engineering applications, bacterial cellulose has been predominantly used, mainly that synthesized by Acetobacterxylinum. Bacterial cellulose is identical to plant cellulose in chemical structure, but it can be produced without contaminant molecules, such as lignin and hemicelluloses, and does not require intensive purification processes. In addition, it is remarkable for its mechanical strength, its ability to be engineered structurally and chemically at nano-, micro-, and macroscales, its biocompatibility and chemical and morphologic controllability [5]. Bacterial cellulose has been used for experimental engineering of bone tissue [6], cartilage [7], skin [8], heart valve [9], and also for urinary reconstruction and diversion [10]. One of the first attempts at vascular tissue engineering was made with cellulose fibers, which were used for constructing three-dimensional vascularized tissue in vitro. These fibers were immobilized with fibronectin in order to improve cell adhesion, and were seeded with bovine coronary artery smooth muscle cells. These cells proliferated on the scaffolds and, after they formed multilayers on the fibers, the fibers were removed by enzymatic digestion using cellulase. The remaining smooth muscle cell aggregates maintained lumens after this procedure, and thus mimicked newly-formed blood vessels [11]. Similarly, three-dimensional nanofibrous scaffolds with micropores made of bacterial cellulose allowed attachment and proliferation of human saphenous vein smooth muscle cells on the surface and also in the inside of the scaffolds [12]. In addition, the mechanical properties of nanofibrous bacterial cellulose scaffolds, evaluated by the shape of the stress-strain response, were reminiscent of the properties of the carotid artery, most probably due to the similarity in architecture of the nanofibril network [13]. The adhesion and growth of vascular endothelial cells was also supported by cellulose-based scaffolds, namely by nanofibrous bacterial cellulose or cellulose acetate scaffolds, especially if these scaffolds were functionalized with RGD-containing oligopeptides, i.e. ligands for integrin adhesion receptors on cells [14, 15], or if they were combined with chitosan [16]. The angiogenic response to bacterial cellulose was also observed under in vivo conditions, i.e. after implantation of these scaffolds in the form of dorsal skinfold chambers into Syrian golden hamsters [17]. Cellulose has also been used for creating tubular structures designed for replacing small-caliber vessels. Hollow-shaped segments of bacterial cellulose were created with a length of 10 mm, an inner diameter of 3.0-3.7 mm and a wall thickness of 0.6 -1.0 mm. These grafts were used to replace the carotid arteries of eight pigs. After a follow-up period of 3 months, seven grafts (87.5%) remained patent, whereas one graft was found to be occluded. All patent grafts developed a single inner layer of endothelium with a basement membrane and a thin layer of collagen, followed by a concentric medial layer containing smooth muscle cells and cellulose, and an outer layer of fibrous cells [18]. Similarly, bacterial cellulose grafts 4 cm in length and 4 mm in internal diameter were implanted bilaterally in the carotid arteries of eight sheep. Although 50% of the grafts occluded within 2 weeks, all patent grafts developed a confluent inner layer of endothelial- like cells [19]. In addition, the mechanical properties of tubular structures created from bacterial cellulose seemed to be advantageous for vascular tissue engineering. For example, these structures exhibited a compliance response similar to that of human saphenous vein [20]. In our experiments, we have concentrated on cellulose-based materials modified with oxidation and/or functionalization with biomolecules. We have prepared fibrous scaffolds made of non-oxidized viscose, dialdehyde cellulose and 6-carboxycellulose with 2.1 wt.% or 6.6 wt.% of -COOH groups. In addition, all these material types were functionalized with arginine, i.e. an amino acid with a basic side chain, or with chitosan, in order to balance (compensate) the relatively acid character of oxidized cellulose molecules. Two groups of samples with and without functionalization were then seeded with vascular smooth muscle cells (VSMC) derived from the rat thoracic aorta by an explantation method [21]. We found that the oxidized cellulose with 2.1 wt.% of-COOH groups was the most appropriate of all the tested materials for colonization with VSMC. The cells on this material achieved an elongated shape, while they were spherical in shape on the other materials. In addition, the numbers of cells obtained in one week after seeding and the concentration of alpha-actin and SM1 and SM2 myosins, measured per mg of protein, were significantly higher on oxidized cellulose with 2.1 wt.% of -COOH groups. Functionalization with arginine and chitosan improved the cell adhesion, but usually only slightly. The most apparent increase in cell number after functionalization was observed on oxidized cellulose with 2.1 wt.% of -COOH groups functionalized with chitosan, and on viscose functionalized with chitosan or arginin. However, the cells on all samples proliferated slowly and with a non-significant increase in cell population densities from day 1 to 7 after seeding. This suggests that cellulose-based materials can be used in applications where high proliferation activity of vascular smooth muscle cells is not desirable. They can therefore be used on vascular prostheses, where excessive VSMC proliferation can lead to the restenosis of the graft. Alternatively, cell proliferation might be enhanced by some other more efficient modification. This would require further research.

6

Human osteoblast-like MG 63 cells in cultures on nanofibrous PLGA membranes loaded with nanodiamonds

Parizek M., Douglas T. E. L., Kromka A., Renzing A., Voss E., Brady M. A., Warnke P. H., Bacakova L.

Engineering of Biomaterials

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2010

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Vol. 13, no. 94

11--13

EN

Using an electrospinning technique, we constructed composite nanofibrous membranes containing a copolymer of L-lactide and glycolide (PLGA, ratio 85:15) and 33 wt% of nanodiamond particles. he number of initially adhering human osteoblast-like MG 63 cells on day 1 after seeding, their spreading and subsequent growth were similar on both types of membranes. However, higher cell numbers on day 3 and 7 after seeding and a larger cell spreading area were found in the cells in the control polystyrene cell culture dishes. Nevertheless, the composite PLGA-ND membranes provided relatively good support for colonization with bone-derived cells; thus this material is promising for bone tissue engineering.

7

Vascular smooth muscle cells in cultures on low density polyethylene modified with plasma discharge and biofunctionalization

Parizek M., Kasalkova N., Bacakova L., Kolarova K., Lisa V., Svorcik V.

Engineering of Biomaterials

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2009

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Vol. 12, no. 89-91

25--28

EN

Low density polyethylene (LDPE) was modified by an Ar plasma discharge and then grafted with glycine (Gly), bovine serum albumin (BSA) or polyethylene glykol (PEG). Some plasma-treated samples and samples grafted with BSA were exposed to a suspension of colloidal carbon particles (C, BSA+C). Pristine LDPE and tissue culture polystyrene dishes (PSC) were used as control samples. The materials were seeded with rat aortic smooth muscle cells and incubated in a medium DMEM with 10% of fetal bovine serum. On day 1 after seeding, the cells on LDPE modified with plasma only, Gly, BSA and BSA+C adhered in similar numbers as on PSC, while the values on non-modified and PEG-modified samples were significantly lower. On day 5, the highest cell numbers were found again on LDPE with Gly, BSA and BSA+C. On day 7, the highest number of cells was found on LDPE modified only with plasma. The latter cells also dis-played the largest cell spreading area. The increased cell colonization was probably due to the formation of oxygen-containing chemical functional groups after plasma irradiation, and also due to positive effects of grafted Gly, BSA and BSA in combination with colloidal C particles.

8

Micro- and nanopatterned surfaces for guided adhesion, growth and phenotypic maturation of cells

Bacakova L., Filova E., Grausova L., Vandrovcova M., Parizek M., Novotna K., Svorcik V., Vacik J., Rypacek F., Kromka A., Heitz J., Shard A.

Engineering of Biomaterials

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2009

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Vol. 12, no. 89-91

18--21

EN

Micropatterned surfaces were created by UV light-irradiation of polytetrafluoroethylene through a metallic mask, by successive plasma polymerization of acrylic acid and 1,7-octadiene, or by creation of prominences and grooves by deposition of fullerenes C60 through a metallic mask. All these surface types were capable of inducing regionally-selective adhesion, proliferation and phenotypic maturation of vascular endothelial cells, vascular smooth muscle cells or human bone-derived MG 63 cells. Nanopatterned surfaces created by tethering GRGDSG oligopeptides through polyethylene oxide chains on a polymeric surface promoted spreading, formation of focal adhesion plaques and DNA synthesis in vascular smooth muscle cells. Surfaces nanopatterned with nanocrystalline diamond gave good support for the adhesion, growth and metabolic activity of osteoblast-like MG 63 cells.

9

The adhesion and growth of vascular smooth muscle cells in cultures on carboranethiol-modified gold films

Parizek M., Base T., Londesborough M. G. S., Lisa V., Bacakova L.

Engineering of Biomaterials

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2008

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Vol. 11, no. 81-84

117--119

EN

Metal surfaces have become important over the last decade for potential surgical implants, and within this context we present here a study of the cell growth on modified gold surfaces. Gold films, deposited on glass plates and annealed with a hydrogen flame, were modified with four different carboranethiol derivatives: 1-(HS)-1,2-C2B10H11 (A), 1,2-(HS)2-1,2-C2B10H10 (B), 9,12-(HS)2-1,2-C2B10H10 (C) and 1,12-(HS)2-1,12- C2B10H10 (D). The materials engendered from these modifications were used to investigate the adhesion and growth of rat aortic smooth muscle cells cultured on these surfaces in a DMEM medium with 10% of fetal bovine serum. One day after seeding, the highest number of initially adhered cells was found on the surface of a bare gold film. However, three days after seeding, the number of cells on carboranethiol-modified gold samples B, C and D was significantly higher than the number on a bare gold film. After seven days, the number of cells on a bare gold film and on gold films modified with derivatives A, B and D was very similar, but the surface of a gold film modified with derivative C exhibited a significantly smaller number of cells. This may be explained by the exposure of the CH vertices of the carborane cluster, which are more acidic than the BH vertices exposed toward the cells in either A or B.

10

The adhesion and growth of vascular smooth muscle cells in cultures on carboranethiol-modified gold films

Parizek M., Base T., Londesborough M. G. S., Lisa V., Bacakova L.

Engineering of Biomaterials

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2008

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Vol. 11, no. 75

6--8

EN

Metal surfaces have become important over the last decade for potential surgical implants, and within this context we present here a study of the cell growth on modified gold surfaces. Gold films, deposited on glass plates and annealed with a hydrogen flame, were modified with four different carboranethiol derivatives: 1-(HS)-1,2-C2 B10H11(A), 1,2-(HS)2-1,2-C2B10H10(B), 9,12-(HS)2-1,2-C2B10H10(C) and 1,12-(HS)2-1,12- C2B10H10(D). The materials engendered from these modifications were used to investigate the adhesion and growth of rat aortic smooth muscle cells cultured on these surfaces in a DMEM medium with 10% of fetal bovine serum. One day after seeding, the highest number of initially adhered cells was found on the surface of a bare gold film. However, three days after seeding, the number of cells on carboranethiol-modified gold samples B, C and D was significantly higher than the number on a bare gold film. After seven days, the number of cells on a bare gold film and on gold films modified with derivatives A, B and D was very similar, but the surface of a gold film modified with derivative C exhibited a significantly smaller number of cells. This may be explained by the exposure of the CH vertices of the carborane cluster, which are more acidic than the BH vertices exposed toward the cells in either A or B.

11

Włókniste podłoża dla inżynierii tkankowej kości: hodowle komórek MG 63 w warunkach statycznych i dynamicznych

Buczyńska J., Pamuła E., Błażewicz S., Bacakova L., Parizek M., Chlupac J., Mikołajczyk T., Boguń M., Dobrzyński P.

Engineering of Biomaterials

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2007

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Vol. 10, no. 65-66

1-6

PL

Resorbowalne włókna z kopolimeru L-laktydu z glikolidem (PLG) i PLG z hydroksyapatytem rozprowadzonym w ich objętości (PLG-HAP) zostały otrzymane metodą formowania z roztworu. Włókna zostały przetworzone w trójwymiarowe podłoża za pomocą metody łączenia włókien. Mikrostrukturę otrzymanych podłoży scharakteryzowano za pomocą mikroskopu stereoskopowego. Wykazano, że podłoża miały różną porowatość, wielkość i orientację pojedynczych włókien. Oddziaływanie włóknistych podłoży z komórkami kostnymi MG 63 było badane in vitro w warunkach statycznych i dynamicznych. Liczba komórek i ich morfologia były oceniane po 3 i 7 dniach od założenia hodowli. Badania wykazały że liczba komórek na materiałach włóknistych rosła wraz z czasem prowadzenia hodowli, chociaż była znacznie niższa niż na płaskiej powierzchni kontrolnej (polistyren do celów kultur komórkowych). W dynamicznych warunkach hodowli obserwowano różną proliferację komórek w zależności od rodzaju użytego podłoża: na PLG występował spadek, zaś na PLG-HAP istotny wzrost liczby komórek. Wyniki sugerują, że obecność cząstek hydroksyapatytu rozprowadzonych w objętości włókien polimerowych poprawia adhezje i proliferacje osteoblastów.

EN

Resorbable poly(L-lactide-co-glycolide) fibres (PLG) and poly(L-lactide-co-glycolide) fibres containing hydroxyapatite nanoparticles in volume of PLG fibres (PLG-HAP) were manufactured by solution spinning process. The resultant fibres were processed into three-dimensional scaffolds using fibre bounding method. The microstructure of resorbable scaffolds was characterized by stereomicroscope. The results show that the scaffolds have different fibrous architecture including porosity, size and arrangement of individual fibres. The interaction of fibrous scaffolds with osteoblast-like MG 63 cells was tested in vitro in static and dynamic cell culture conditions. The number of adhering cells and their morphology were evaluated on days 3 and 7 after seeding. It was found that cell number increased with the cultivation time, although it was significantly lower than on control polystyrene dish (TCPS). During dynamic cultivation the number of cells decreased on PLG scaffolds, whereas on PLG-HAP scaffolds it increased. These results suggest that presence of hydroxyapatite distributed within the whole volume of resorbable polymer fibres promoted adhesion and proliferation of osteoblasts.

12

Modyfikacja powierzchni kopolimeru glikolidu z L-laktydem dla zastosowania w inżynierii tkankowej kości

Kaczmarczyk M., Pamuła E., Bacakova L., Parizek M., Dobrzyński P.

Engineering of Biomaterials

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2007

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Vol. 10, no. 62

12-17

PL

Praca dotyczy modyfikacji powierzchni kopolimeru glikolidu z L-laktydem w 0,1M NaOH przez różne okresy czasu tj. od 2 do 24 h. W pracy scharakteryzowano topografię i budowę chemiczną powierzchni folii polimerowych w funkcji czasu modyfikacji. Zbadano też wpływ zmodyfikowanej powierzchni na jej własności biologiczne in vitro w kontakcie z osteoblastami. Badania wykazały, że zastosowana metoda modyfikacji powoduje zmiany w topografii i wzrost chropowatości powierzchni, ale nie wpływa na skład chemiczny, zwilżalność wodą i masę cząsteczkową polimeru. Najlepszy wzrost osteoblastów obserwowano na folii modyfikowanej przez 6h w 0,1M NaOH (o chropowatości ok. 60nm), natomiast na foliach o chropowatości niższej i wyższej adhezja i zdolność do proliferacji komórek były istotnie niższe.

EN

The study was focused on surface modifications of poly(glycolide-co-L-lactide) films (PGLA) after their exposure to 0.1 M NaOH for 2 to 24 h. Topography and surface chemical structure of the films were characterized. The influence of the modified surface on biological properties in contact with human osteoblast-like cells in vitro was evaluated. The results showed that the modification in NaOH caused topographical changes, such as the increase in surface roughness, without affecting the surface chemical composition, wettability and molecular mass. The best growth of osteoblast-like cells was observed on PGLA films modified in 0.1M NaOH for 6 h (average surface roughness of about 60 nm), whereas on the films with lower or higher roughness, the cell adhesion and proliferation activity were lower.

13

Improved adhesion and growth of vascular smooth muscle cells in cultures on polyethylene modified by plasma discharge

Parizek M., Kasalkova N., Bacakova L., Kolarova K., Lisa V., Svorcik V.

Engineering of Biomaterials

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2007

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Vol. 10, no. 67-68

1-4

EN

The attractiveness of synthetic polymers for cell colonization can be affected by physical and chemical modification of the polymer surface. In this study, high density polyethylene (HDPE, m.w. 0.952g/cm3) and low density polyethylene (LDPE, m.w. 0.922g/cm3) were modified by an Ar plasma discharge using Balzers SCD 050 device (exposure time 10, 50, 150 and 400 seconds, discharge power 1.7W). The material was then seeded with rat aortic smooth muscle cells (RASMC; passages 8 to 9, 17 000 cells/cm3) and incubated in a DMEM medium with 10% of fetal calf serum. On day 1 after seeding, the number of initially adhered cells was significantly higher on all modified HDPE and LDPE samples. On day 2, this difference persisted in HDPE, whereas in LDPE only the values on the samples modified by 150 and 400 seconds were significantly higher. On the 5th and 7th day, there were no significant differences in cell number among all LDPE samples. However, on the HDPE foils, significant differences were still apparent on the samples modified for 400 seconds. The cell spreading areas measured on day 1 after seeding were significantly larger on all modified LDPE samples, and, on day 2, on the HDPE samples exposed for 150s. The increased cell colonization was probably due to the formation of oxygen-containing chemical functional groups in the polymer. These results suggest that the responsiveness of the cell to the changes in physiochemical surface properties was more pronounced in HDPE than in LDPE. On both types of polyethylene, the most appropriate exposure time for the enhancement of cell adhesion and growth seemed to be 150 and 400 seconds.

14

Vascular smooth muscle cells in cultures on synthetic polymers with adhesive microdomains

Parizek M., Bacakova L., Lisa V., Kubova O., Svorcik V., Heitz J.

Inżynieria Biomateriałów

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2006

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R. 9, nr 58-60

7-10

EN

Polyethylene terephtalate was modified by UV light irradiation produced by a Xe2-excimer lamp for 10, 20 or 30 min in an acetylene atmosphere. For creation of microdomains for selective cell adhesion, a contact nickel mask (apertures of the diameter of 500 m, centre-to-centre distance 2 mm) was used. The material was then seeded with rat aortic smooth muscle cells (passage 3, 17 000 cells/cm square). After 1, 3 and 7 days of cultivation, the cells were homogeneously distributed on the samples without any preference of the irradiated microdomains. Moreover, on day 1, the number of initially adhered cells was similar on all tested samples. However, on day 3, the number of cells on the irradiated samples was significantly higher than that on control unmodified PET and increased proportionally to the time of exposure to UV light. On day 7 after seeding, however, the cell number on the unmodified PET exceeded the value on all irradiated samples. In the second set of experiments, polyethylene (PE) was irradiated by Ar+ ions in order to create the adhesive microdomains (dose 10\12-10\14 ions/cm square, energy 150 and 15 keV, contact mask with holes of 100 m diameter and distance 200 m). The highest selectivity of the adhesion and growth of rat aortic smooth muscle cells (89% of all cells) was found on the microdomains created at the energy of 150 keV and the dose of 3x10\12 ions Ar+. The lowest selectivity (30%) occurred on samples irradiated with 150 keV Ar+ ions of the dose of 3x10\14 ions/cm square. Therefore, both methods seem to be suitable for modification of materials with highly hydrophobic surfaces in purpose to increase the cell colonization, for example when constructing bioartificial vascular replacements. The second method can be also used for the creation of domains for a regionally selective adhesion and growth of cells on biomaterials.