The modification of biomaterial surfaces has become increasingly relevant in the context of ongoing advancements in tissue engineering applications and the development of tissue-mimicking polymer materials. In this study, we investigated the layer-by-layer (LbL) deposition of polyelectrolyte multilayer protein reservoirs consisting of poly-l-lysine (PLL) and hyaluronic acid (HA) on the hydrophobic surface of poly(glycerol sebacate) (PGS) elastomer. Using the methods of isothermal titration calorimetry and surface plasmon resonance, we systematically investigated the interactions between the polyelectrolytes and evaluated the deposition process in real time, providing insight into the phenomena associated with film assembly. PLL/HA LbL films deposited on PGS showed an exceptional ability to incorporate bone morphogenetic protein-2 (BMP-2) compared to other growth factors tested, thus highlighting the potential of PLL/HA LbL films for osteoregenerative applications. The concentration of HA solution used for film assembly did not affect the thickness and topography of the (PLL/HA)10 films, but had a notable impact on the hydrophilicity of the PGS surface and the BMP-2 release kinetics. The release kinetics were successfully described using the Weibull model and hyperbolic tangent function, underscoring the potential of these less frequently used models to compare the protein release from LbL protein reservoirs.

• MeSH
• kyselina hyaluronová * chemie   MeSH
• nanočástice layer-by-layer MeSH
• polyelektrolyty MeSH
• polylysin * chemie   MeSH
• polymery MeSH
• Publikační typ
• časopisecké články MeSH

Long-term delivery of growth factors and immunomodulatory agents is highly required to support the integrity of tissue in engineering constructs, e.g., formation of vasculature, and to minimize immune response in a recipient. However, for proteins with a net positive charge at the physiological pH, controlled delivery from negatively charged alginate (Alg) platforms is challenging due to electrostatic interactions that can hamper the protein release. In order to regulate such interactions between proteins and the Alg matrix, we propose to complex proteins of interest in this study - CXCL12, FGF-2, VEGF - with polyanionic heparin prior to their encapsulation into Alg microbeads of high content of α-L-guluronic acid units (high-G). This strategy effectively reduced protein interactions with Alg (as shown by model ITC and SPR experiments) and, depending on the protein type, afforded control over the protein release for at least one month. The released proteins retained their in vitro bioactivity: CXCL12 stimulated the migration of Jurkat cells, and FGF-2 and VEGF induced proliferation and maturation of HUVECs. The presence of heparin also intensified protein biological efficiency. The proposed approach for encapsulation of proteins with a positive net charge into high-G Alg hydrogels is promising for controlled long-term protein delivery under in vivo conditions.

• MeSH
• algináty chemie   MeSH
• chemokin CXCL12 chemie   MeSH
• endoteliální buňky pupečníkové žíly (lidské) MeSH
• fibroblastový růstový faktor 2 chemie   MeSH
• heparin chemie   MeSH
• lidé MeSH
• mikrosféry MeSH
• nádorové buněčné linie MeSH
• tkáňové inženýrství MeSH
• vaskulární endoteliální růstový faktor A chemie   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• hodnotící studie MeSH

The clinical transplantation (Tx) of pancreatic islets (PI) is limited by a significant cell loss during 2 days after Tx, and by permanent immunosuppressive therapy. Therefore, there is a need of an alternative Tx technique, which would significantly improve the engraftment, the local immune protection as well as the safe graftectomy. Based on the literature as well as our preliminary data the sufficient oxygen supply is a crucial factor. The cavity created under skin by implantation of a polymer microporous scaffold can provide adequate conditions. VEGF attached to the scaffold can control the ingrowth of connective tissue with capillaries and cytokine CXCL-12 can restrict activity of immunocompetent cells. We will evaluate the effect of VEGF bound to polymer scaffolds on the cavity vascularization and controlled release of CXCL-12 from scaffolds or microparticles on PI survival. The optimized cavity can improve outcomes of allogeneic PI Tx for Type-1 diabetic patients as well as of autologous PI Tx in patients after total pancreatectomy.

Transplantace (Tx) Langerhansových ostrůvků (LO) v klinické praxi je limitovaná mimo jiné vysokým procentem buněk ztracených během 2 dnů po Tx, a nutností trvalé imunosupresívní léčby. Proto se hledá alternativní způsob Tx, který by významně zvýšil úspěšnost přihojení, umožnil lokální ochranu před rejekcí a bezpečné vyjmutí štěpu v případě komplikací. Podle literárních i našich předběžných výsledků je klíčové dostatečné zásobení kyslíkem. Takové podmínky může zajistit dutina uměle vytvořená v podkoží příjemce pomocí polymerního skeletu s navázanými funkčními molekulami (VEGF, CXCL-12), které umožní řízené prorůstání vaziva s kapilárami a lokálně omezí aktivitu imunokompetentních buněk. Plánujeme posoudit efekt mikroporezní struktury skeletu, VEGF navázaného na skelet a řízeného uvolňování molekuly CXCL-12 ze skeletu nebo mikročástic, které budou součástí suspense při Tx LO, na vaskularizaci dutiny a dlouhodobou funkčnost transplantovaných LO u potkanů. Technika je slibná jak pro allogenní Tx LO pro diabetiky 1. typu, tak zejména pro autoTx LO po totální pankreatektomii.

• MeSH
• krevní oběh MeSH
• krysa rodu rattus MeSH
• lidé MeSH
• modely nemocí na zvířatech MeSH
• remodelace cév MeSH
• subkutánní tkáň MeSH
• tkáňové podpůrné struktury MeSH
• transplantace Langerhansových ostrůvků MeSH
• vstřebatelné implantáty MeSH
• vyvíjení léků MeSH
• zvířata MeSH
• Check Tag
• krysa rodu rattus MeSH
• lidé MeSH
• zvířata MeSH
• Publikační typ
• hodnotící studie MeSH

• Konspekt
• Patologie. Klinická medicína
• NLK Obory
• transplantologie
• angiologie
• NLK Publikační typ
• závěrečné zprávy o řešení grantu AZV MZ ČR

The effectiveness of cell transplantation can be improved by optimization of the transplantation site. For some types of cells that form highly oxygen-demanding tissue, e.g., pancreatic islets, a successful engraftment depends on immediate and sufficient blood supply. This critical point can be avoided when cells are transplanted into a bioengineered pre-vascularized cavity which can be formed using a polymer scaffold. In our study, we tested surface-modified poly(lactide-co-caprolactone) (PLCL) capsular scaffolds containing the pro-angiogenic factor VEGF. After each modification step (i.e., amination and heparinization), the surface properties and morphology of scaffolds were characterized by ATR-FTIR and XPS spectroscopy, and by SEM and AFM. All modifications preserved the gross capsule morphology and maintained the open pore structure. Optimized aminolysis conditions decreased the Mw of PLCL only up to 10% while generating a sufficient number of NH2 groups required for the covalent immobilization of heparin. The heparin layer served as a VEGF reservoir with an in vitro VEGF release for at least four weeks. In vivo studies revealed that to obtain highly vascularized PLCL capsules (a) the optimal VEGF dose for the capsule was 50 μg and (b) the implantation time was four weeks when implanted into the greater omentum of Lewis rats; dense fibrous tissue accompanied by vessels completely infiltrated the scaffold and created sparse granulation tissue within the internal cavity of the capsule. The prepared pre-vascularized pouch enabled the islet graft survival and functioning for at least 50 days after islet transplantation. The proposed construct can be used to create a reliable pre-vascularized pouch for cell transplantation.

• MeSH
• bioinženýrství * MeSH
• experimentální diabetes mellitus chemicky indukované   metabolismus   patologie   MeSH
• fyziologická neovaskularizace * MeSH
• injekce intraperitoneální MeSH
• krevní glukóza analýza   MeSH
• krysa rodu rattus MeSH
• molekulární struktura MeSH
• polyestery chemie   metabolismus   MeSH
• potkani inbrední LEW MeSH
• streptozocin aplikace a dávkování   MeSH
• tobolky chemie   metabolismus   MeSH
• transplantace Langerhansových ostrůvků * MeSH
• vaskulární endoteliální růstové faktory chemie   metabolismus   MeSH
• velikost částic MeSH
• zvířata MeSH
• Check Tag
• krysa rodu rattus MeSH
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

Polyelectrolyte layer-by-layer (LbL) films that disintegrate under physiological conditions are intensively studied as coatings to enable the release of bioactive components. Herein, we report on the interactions and pH-stability of LbL films composed of chitosan (CH) or N-(2-hydroxypropyl)-3-trimethylammonium chitosan chloride (CMCH) and tannic acid (TA), employed to guarantee the film disintegration. The self-assembly of TA with CH and CMCH at pH 5 and with CMCH at pH 7.4 were proven by turbidimetric, surface plasmon resonance and UV-Vis analyses. The LbL films exhibited pH-dependent properties; CMCH/TA films prepared at pH 7.4 showed exponential growth as well as a higher layer thickness and surface roughness, whereas films prepared at pH 5 grew linearly and were smoother. The film stability varied with the pH used for film assembly; CH/TA films assembled at pH 5 were unstable at pH 8.5, whereas CMCH/TA films assembled at pH 7.4 disintegrated at pH 4. All films exhibited a similar disassembly at pH 7.4. The coatings reduced the adhesion of E. coli and S. aureus by approximately 80%. CMCH-terminated CMCH/TA films were more resistant to bacterial adhesion, whereas CH-terminated CH/TA films demonstrated stronger killing activity. The prepared pH-triggered decomposable LbL films could be used as degradable coatings that allow the release of therapeutics for biomedical applications and also prevent bacterial adhesion.

• MeSH
• antibakteriální látky farmakologie   MeSH
• biokompatibilní materiály chemie   farmakologie   MeSH
• chitosan chemie   MeSH
• Escherichia coli účinky léků   MeSH
• film jako téma MeSH
• koncentrace vodíkových iontů MeSH
• Staphylococcus aureus účinky léků   MeSH
• taniny chemie   MeSH
• Publikační typ
• časopisecké články MeSH

Growth factors are powerful molecules that regulate cellular growth, proliferation, healing, and cellular differentiation. A delivery matrix that incorporates growth factors with high loading efficiencies, controls their release, and maintains bioactivity would be a powerful tool for regenerative medicine. Alginate has several unique properties that make it an excellent platform for the delivery of proteins. Mild gelling conditions can minimize the risk of protein denaturation; moreover, alginate can serve as protection from degradation until protein release. Various modifications have been proposed to tune alginate binding and release proteins, simultaneously adjusting alginate degradability, mechanical stiffness, swelling, gelation properties and cell affinity. The primary objective of this article is to review the literature related to recent advances in the application of alginate matrices in protein delivery in regenerative medicine. A special emphasis is put on the relevance of delivery of growth factors and chemokine.

• MeSH
• algináty aplikace a dávkování   chemie   metabolismus   MeSH
• cytokiny aplikace a dávkování   chemie   metabolismus   MeSH
• lidé MeSH
• mezibuněčné signální peptidy a proteiny aplikace a dávkování   chemie   metabolismus   MeSH
• nosiče léků aplikace a dávkování   chemie   metabolismus   MeSH
• proliferace buněk účinky léků   fyziologie   MeSH
• systémy cílené aplikace léků metody   MeSH
• uvolňování léčiv fyziologie   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• přehledy MeSH

The study monitored in vitro early response of connective tissue cells and immunocompetent cells to enosseal implant materials coated by different blood components (serum, activated plasma, and plasma/platelets) to evaluate human osteoblast proliferation and synthetic activity and inflammatory response presented as a cytokine profile of peripheral blood mononuclear cells (PBMCs) under conditions imitating the situation upon implantation. The cells were cultivated on coated Ti-plasma-sprayed (Ti-PS), Ti-etched (Ti-Etch), Ti-hydroxyapatite (Ti-HA), and ZrO2 surfaces. The plasma/platelets coating supported osteoblast proliferation only on osteoconductive Ti-HA and Ti-Etch whereas activated plasma enhanced proliferation on all surfaces. Differentiation (BAP) and IL-8 production remained unchanged or decreased irrespective of the coating and surface; only the serum and plasma/platelets-coated ZrO2 exhibited higher BAP and IL-8 expression. RANKL production increased on serum and activated plasma coatings. PBMCs produced especially cytokines playing role in inflammatory phase of wound healing, that is, IL-6, GRO-α, GRO, ENA-78, IL-8, GM-CSF, EGF, and MCP-1. Cytokine profiles were comparable for all tested surfaces; only ENA-78, IL-8, GM-CSF, and MCP-1 expression depended on materials and coatings. The activated plasma coating led to uniformed surfaces and represented a favorable treatment especially for bioinert Ti-PS and ZrO2 whereas all coatings had no distinctive effect on bioactive Ti-HA and Ti-Etch.

• MeSH
• biokompatibilní potahované materiály škodlivé účinky   chemie   MeSH
• buněčné linie MeSH
• chemokin CCL2 metabolismus   MeSH
• chemokin CXCL1 metabolismus   MeSH
• chemokin CXCL5 metabolismus   MeSH
• cytokiny metabolismus   MeSH
• epidermální růstový faktor metabolismus   MeSH
• faktor stimulující granulocyto-makrofágové kolonie metabolismus   MeSH
• hojení ran účinky léků   MeSH
• interleukin-6 metabolismus   MeSH
• interleukin-8 metabolismus   MeSH
• leukocyty mononukleární účinky léků   metabolismus   MeSH
• lidé MeSH
• osteoblasty účinky léků   metabolismus   MeSH
• proliferace buněk účinky léků   MeSH
• titan škodlivé účinky   chemie   MeSH
• zánět metabolismus   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH

Understanding the behavior of single proteins at the polyelectrolyte multilayer film/solution interface is of prime importance for the designing of bio-functionalized surface coatings. In the present paper, we study the adsorption of the model proteins, albumin and lysozyme, as well as basic fibroblast growth factor (FGF-2) on a polysaccharide multilayer film composed of quaternized chitosan and heparin. Several analytical methods were used to describe the formation of the polysaccharide film and its interactions with the proteins. Both albumin and lysozyme adsorbed on quaternized chitosan/heparin films, however this process strongly depended on the terminating polysaccharide. Protein adsorption was driven mainly by electrostatic interactions between protein and the terminal layer of the film. The effective binding of FGF-2 by the heparin-terminated film suggested that other interactions could also contribute to the adsorption process. We believe that this FGF-2-presenting polysaccharide film may serve as a biofunctional surface coating for biologically-related applications.

• MeSH
• adsorpce MeSH
• biokompatibilní potahované materiály * MeSH
• chitosan chemie   MeSH
• heparin chemie   MeSH
• proteiny chemie   MeSH
• Publikační typ
• časopisecké články MeSH

The proposed project is focused on improvement of the biological fixation of titanium implants into the bone. The source materials used in the study will be titanium surfaces with an appropriated surface roughness (by sand-blasting, etching). Then, the hydrogel carrier based on alginates enriched with the nanosized calcium phosphates as a source of Ca2+ and (PO4)3- ions needed for formation of bone callus by adhered osteoblasts will be chemically bonded directly to the surface. The sequential degradation of the carrier will provide a simple and consequent release of minerals from the implant surface to the bone-to-implant interface, what is necessary for later successful mineralization of osteoid produced by mature osteoblasts. The effects of titanium surface modifications on adhesion, growth, differentiation and synthetic activity of human osteogenic cells will be evaluated. The extent of activation of mononuclear cells, i.e. an important part of the immune system, will be investigated.

Navrhovaný projekt se zabývá usnadněním a podporou biologické fixace implantátu do kosti. Na vhodně drsném titanovém povrchu (pískování, leptání) bude chemicky zakotven hydrogelový nosič na bázi alginátu, který bude obsahovat fosfáty vápníku v nanoformě jako zdroj Ca2+ a (PO4)3- iontů potřebných pro vznik a tvorbu primárního kostního svalku adherovanými osteoblasty jako i pro jeho pozdější remodelaci. Postupná degradace nosiče pak poskytne jednoduché pozvolné uvolňování zdrojových minerálů z povrchu implantátu na rozhraní kost-implantát, což je nezbytné pro pozdější úspěšnou mineralizaci osteoidu. Bude sledován účinek povrchových modifikací na adhezi, proliferaci, diferenciaci a syntetickou aktivitu lidských osteogenních buněk v in vitro podmínkách. Bude zkoumán stupeň aktivace mononukleárních buněk jako důležité složky imunitního systému.

• MeSH
• adhezivita MeSH
• algináty MeSH
• biokompatibilní potahované materiály MeSH
• biomimetika MeSH
• experimentální implantáty MeSH
• fosforečnany vápenaté MeSH
• osteom osteoidní MeSH
• regenerace kostí MeSH
• rozhraní kost/implantát MeSH
• techniky in vitro MeSH
• titan MeSH
• transplantace buněk MeSH
• transplantace kostí MeSH

• Konspekt
• Ortopedie. Chirurgie. Oftalmologie
• NLK Obory
• ortopedie
• biochemie
• NLK Publikační typ
• závěrečné zprávy o řešení grantu IGA MZ ČR

Tissue engineering benefits from novel materials with precisely tunable physical, chemical and mechanical properties over a broad range. Here we report a practical approach to prepare Bombyx mori silk fibroin hydrogels using the principle of non-solvent induced phase separation (NIPS). A combination of reconstituted silk fibroin (RSF) and methanol (non-solvent), with a final concentration of 2.5% w/v and 12.5% v/v respectively, maintained at 22 °C temperature turned into a hydrogel within 10 hours. Freeze-drying of this gel gave a foam with a porosity of 88%, a water uptake capacity of 89% and a swelling index of 8.6. The gelation kinetics and the loss tangent of the gels were investigated by rheometry. The changes in the morphology of the porous foams were visualized by SEM. The changes in RSF chemical composition and the relative fraction of its secondary structural elements were analyzed by ATR-FTIR along with Fourier self-deconvolution. And, the changes in the glass transition temperature, specific heat capacity and the relative fraction of crystallinity of RSF were determined by TM-DSC. Data suggested that RSF-water-methanol behaved as a polymer-solvent-non-solvent ternary phase system, wherein the demixing of the water-methanol phases altered the thermodynamic equilibrium of RSF-water phases and resulted in the desolvation and eventual separation of the RSF phase. Systematic analysis revealed that both gelation time and the properties of hydrogels and porous foams could be controlled by the ratios of RSF and non-solvent concentration as well as by the type of non-solvent and incubation temperature. Due to the unique properties we envisage that the herein prepared NIPS induced RSF hydrogels and porous foams can possibly be used for the encapsulation of cells and/or for the controlled release of both hydrophilic and hydrophobic drugs.

• MeSH
• biokompatibilní materiály chemie   MeSH
• bourec chemie   MeSH
• fibroiny chemie   MeSH
• hydrofobní a hydrofilní interakce MeSH
• hydrogely chemie   MeSH
• poréznost MeSH
• rozpouštědla chemie   MeSH
• teplota MeSH
• tkáňové inženýrství MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH