BACKGROUND: Nanofibrous scaffolds loaded with bioactive nanoparticles are promising materials for bone tissue engineering. METHODS: In this study, composite nanofibrous membranes containing a copolymer of L-lactide and glycolide (PLGA) and diamond nanoparticles were fabricated by an electrospinning technique. PLGA was dissolved in a mixture of methylene chloride and dimethyl formamide (2:3) at a concentration of 2.3 wt%, and nanodiamond (ND) powder was added at a concentration of 0.7 wt% (about 23 wt% in dry PLGA). RESULTS: In the composite scaffolds, the ND particles were either arranged like beads in the central part of the fibers or formed clusters protruding from the fibers. In the PLGA-ND membranes, the fibers were thicker (diameter 270 ± 9 nm) than in pure PLGA meshes (diameter 218 ± 4 nm), but the areas of pores among these fibers were smaller than in pure PLGA samples (0.46 ± 0.02 μm(2) versus 1.28 ± 0.09 μm(2) in pure PLGA samples). The PLGA-ND membranes showed higher mechanical resistance, as demonstrated by rupture tests of load and deflection of rupture probe at failure. Both types of membranes enabled the attachment, spreading, and subsequent proliferation of human osteoblast-like MG-63 cells to a similar extent, although these values were usually lower than on polystyrene dishes. Nevertheless, the cells on both types of membranes were polygonal or spindle-like in shape, and were distributed homogeneously on the samples. From days 1-7 after seeding, their number rose continuously, and at the end of the experiment, these cells were able to create a confluent layer. At the same time, the cell viability, evaluated by a LIVE/DEAD viability/cytotoxicity kit, ranged from 92% to 97% on both types of membranes. In addition, on PLGA-ND membranes, the cells formed well developed talin-containing focal adhesion plaques. As estimated by the determination of tumor necrosis factor-alpha levels in the culture medium and concentration of intercellular adhesion molecule-1, MG-63 cells, and RAW 264.7 macrophages on these membranes did not show considerable inflammatory activity. CONCLUSION: This study shows that nanofibrous PLGA membranes loaded with diamond nanoparticles have interesting potential for use in bone tissue engineering.

• MeSH
• buněčná adheze MeSH
• buněčné linie MeSH
• diamant chemie   MeSH
• kostní náhrady chemie   MeSH
• kyselina mléčná chemie   MeSH
• kyselina polyglykolová chemie   MeSH
• lidé MeSH
• mikrofilamenta metabolismus   MeSH
• mikroskopie elektronová rastrovací MeSH
• myši MeSH
• nanočástice chemie   ultrastruktura   MeSH
• nanomedicína MeSH
• nanovlákna chemie   ultrastruktura   MeSH
• osteoblasty cytologie   imunologie   fyziologie   MeSH
• proliferace buněk MeSH
• testování materiálů MeSH
• tkáňové inženýrství metody   MeSH
• tkáňové podpůrné struktury chemie   MeSH
• transmisní elektronová mikroskopie MeSH
• viabilita buněk MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Diamond is recognized as an attractive material for merging solid-state and biological systems. The advantage of diamond field-effect transistors (FET) is that they are chemically resistant, bio-compatible, and can operate without gate oxides. Solution-gated FETs based on H-terminated nanocrystalline diamond films exhibiting surface conductivity are employed here for studying effects of fetal bovine serum (FBS) proteins and osteoblastic SAOS-2 cells on diamond electronic properties. FBS proteins adsorbed on the diamond FETs permanently decrease diamond conductivity as reflected by the -45 mV shift of the FET transfer characteristics. Cell cultivation for 2 days results in a further shift by another -78 mV. We attribute it to a change of diamond material properties rather than purely to the field-effect. Increase in gate leakage currents (by a factor of 4) indicates that the FBS proteins also decrease the diamond-electrolyte electronic barrier induced by C-H surface dipoles. We propose a model where the proteins replace ions in the very vicinity of the H-terminated diamond surface.

• MeSH
• adsorpce MeSH
• biosenzitivní techniky přístrojové vybavení   metody   MeSH
• buněčné linie MeSH
• diamant chemie   MeSH
• elektrochemie metody   MeSH
• elektronické tranzistory MeSH
• ionty MeSH
• krevní proteiny chemie   MeSH
• lidé MeSH
• mikroskopie elektronová rastrovací MeSH
• nanočástice chemie   ultrastruktura   MeSH
• povrchové vlastnosti MeSH
• roztoky MeSH
• skot MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• skot MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

The excellent mechanical, tribological and biochemical properties of diamond coatings are promising for improving orthopedic or stomatology implants. A crucial prerequisite for such applications is an understanding and control of the biological response of the diamond coatings. This study concentrates on the correlation of diamond surface properties with osteoblast behavior. Nanocrystalline diamond (NCD) films (grain size up to 200 nm, surface roughness 20 nm) were deposited on silicon substrates of varying roughnesses (1, 270 and 500 nm) and treated by oxygen plasma to generate a hydrophilic surface. Atomic force microscopy was used for topographical characterization of the films. As a reference surface, tissue culture polystyrene (PS) was used. Scanning electron microscopy and immunofluorescence staining was used to visualize cell morphological features as a function of culture time. Metabolic activity, alkaline phosphatase activity, and calcium and phosphate deposition was also monitored. The results show an enhanced osteoblast adhesion as well as increased differentiation (raised alkaline phosphatase activity and mineral deposition) on NCD surfaces (most significantly on RMS 20 nm) compared to PS. This is attributed mainly to the specific surface topography as well as to the biocompatible properties of diamond. Hence the controlled (topographically structured) diamond coating of various substrates is promising for preparation of better implants, which offer faster colonization by specific cells as well as longer-term stability.

• MeSH
• biokompatibilní materiály chemie   MeSH
• buněčná diferenciace MeSH
• buněčné kultury metody   MeSH
• buněčné linie MeSH
• diamant chemie   MeSH
• lidé MeSH
• molekulární konformace MeSH
• nanostruktury chemie   ultrastruktura   MeSH
• osteoblasty cytologie   fyziologie   MeSH
• osteogeneze fyziologie   MeSH
• povrchové vlastnosti MeSH
• proliferace buněk MeSH
• tkáňové inženýrství metody   MeSH
• velikost buňky MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• práce podpořená grantem MeSH

• Publikační typ
• abstrakty MeSH

In this interdisciplinary project, adipose tissue-derived stem cells (ASCs) will be obtained by liposuction from the fat tissue of patients. Optimal conditions of the liposuction (particularly local anesthesia, negative pressure) will be elaborated in order to obtain the ASCs in high numbers and viability. ASCs will be then differentiated towards osteoblasts, vascular smooth muscle cells, vascular endothelial cells and keratinocytes by appropriate cell culture conditions, namely (i) a cell carrier with suitable physicochemical properties, such as rigidity or deformability, wettability, charge and conductivity, roughness and morphology, surface chemical structure, 2D or 3D structure, (ii) composition of cell culture media, (iii) mechanical stimulation in dynamic bioreactors and (iv) electrical stimulation. The differentiated ASCs will be used for construction of hybrid replacements of the bone tissue, blood vessels and skin. These replacements will contain a material carrier optimal for a given application and a cell component, and will be promising for future clinical applications.

Kmenové buňky získané liposukcí z tukové tkáně pacientů budou v tomto mezioborovém projektu využity k inženýrství bioarteficiální kostní, cévní a kožní tkáně. Nejprve budou vypracovány podmínky liposukce (především lokální anestézie, negativní tlak) optimální pro získání vysokého počtu životaschopných kmenových buněk. Tyto buňky budou dále vhodnými kultivačními podmínkami diferencovány směrem k osteoblastům a cévním hladkým svalovým buňkám, cévním endotelovým buňkám a keratinocytům. Kultivační podmínky budou zahrnovat: (i) nosič buněk z „umělého“ materiálu s vhodnými fyzikálně-chemickými vlastnostmi, jako je např. tuhost, deformabilita, smáčivost, náboj a vodivost, drsnost, morfologie, povrchová chemická struktura, 2D či 3D struktura, (ii) složení kultivačního média, (iii) mechanickou stimulaci v dynamickém bioreaktoru a (iv) elektrickou stimulaci. Diferencované buňky budou využity ke konstrukci hybridních náhrad kostní tkáně, cév a kůže. Tyto konstrukty budou obsahovat materiál nejvhodnější pro danou aplikaci a buněčnou složku, a budou perspektivní pro budoucí klinické aplikace.

• MeSH
• biokompatibilní materiály MeSH
• buněčná diferenciace MeSH
• elektrická stimulace MeSH
• kultivované buňky MeSH
• lipektomie MeSH
• mezenchymální kmenové buňky MeSH
• primární buněčná kultura MeSH
• protézy a implantáty MeSH
• regenerativní lékařství MeSH
• tkáňové inženýrství MeSH

• Konspekt
• Patologie. Klinická medicína
• NLK Obory
• biomedicínské inženýrství
• cytologie, klinická cytologie
• NLK Publikační typ
• závěrečné zprávy o řešení grantu AZV MZ ČR