Three-dimensional hydrogel supports for mesenchymal and neural stem cells (NSCs) are promising materials for tissue engineering applications such as spinal cord repair. This study involves the preparation and characterization of superporous scaffolds based on a copolymer of 2-hydroxyethyl and 2-aminoethyl methacrylate (HEMA and AEMA) crosslinked with ethylene dimethacrylate. Ammonium oxalate is chosen as a suitable porogen because it consists of needle-like crystals, allowing their parallel arrangement in the polymerization mold. The amino group of AEMA is used to immobilize RGDS and SIKVAVS peptide sequences with an N-γ-maleimidobutyryloxy succinimide ester linker. The amount of the peptide on the scaffold is determined using 125 I radiolabeled SIKVAVS. Both RGDS- and SIKVAVS-modified poly(2-hydroxyethyl methacrylate) scaffolds serve as supports for culturing human mesenchymal stem cells (MSCs) and human fetal NSCs. The RGDS sequence is found to be better for MSC and NSC proliferation and growth than SIKVAVS.

• MeSH
• buněčné linie MeSH
• lidé MeSH
• methylmetakryláty chemie   farmakologie   MeSH
• mezenchymální kmenové buňky cytologie   MeSH
• nervové kmenové buňky cytologie   metabolismus   MeSH
• oligopeptidy * chemie   farmakologie   MeSH
• tkáňové inženýrství metody   MeSH
• tkáňové podpůrné struktury chemie   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Superporous poly(2-hydroxyethyl methacrylate-co-2-aminoethyl methacrylate) (P(HEMA-AEMA)) hydrogel scaffolds are designed for in vitro 3D culturing of leukemic B cells. Hydrogel porosity, which influences cell functions and growth, is introduced by adding ammonium oxalate needle-like crystals in the polymerization mixture. To improve cell vitality, cell-adhesive Arg-Gly-Asp-Ser (RGDS) peptide is immobilized on the N-(γ-maleimidobutyryloxy)succinimide-activated P(HEMA-AEMA) hydrogels via reaction of SH with maleimide groups. This modification is especially suitable for the survival of primary chronic lymphocytic leukemia cells (B-CLLs) in 3D cell culture. No other tested stimuli (interleukin-4, CD40 ligand, or shaking) can further improve B-CLL survival or metabolic activity. Both unmodified and RGDS-modified P(HEMA-AEMA) scaffolds serve as a long-term (70 days) 3D culture platforms for HS-5 and M2-10B4 bone marrow stromal cell lines and MEC-1 and HG-3 B-CLL cell lines, although the adherent cells retain their physiological morphologies, preferably on RGDS-modified hydrogels. Moreover, the porosity of hydrogels allows direct cell lysis, followed by efficient DNA isolation from the 3D-cultured cells. P(HEMA-AEMA)-RGDS thus serves as a suitable 3D in vitro leukemia model that enables molecular and metabolic assays and allows imaging of cell morphology, interactions, and migration by confocal microscopy. Such applications can prospectively assist in testing of drugs to treat this frequently recurring or refractory cancer.

• MeSH
• buněčné kultury metody   MeSH
• chronická lymfatická leukemie * MeSH
• hydrogely chemie   MeSH
• lidé MeSH
• mezenchymální kmenové buňky MeSH
• nádorové buněčné linie MeSH
• oligopeptidy MeSH
• poréznost MeSH
• sukcinimidy chemie   MeSH
• tkáňové podpůrné struktury chemie   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH

Polymer layers capable of suppressing protein adsorption from biological media while presenting extracellular matrix-derived peptide motifs offer valuable new options for biomimetic surface engineering. Herein, we provide detailed insights into physicochemical changes induced in a nonfouling poly(ethylene oxide) (PEO) brush/polydopamine (PDA) system by incorporation of adhesion ligand (RGD) peptides. Brushes with high surface chain densities (σ ≥ 0.5 chains·nm-2) and pronounced hydrophilicity (water contact angles ≤ 10°) were prepared by end-tethering of heterobifunctional PEOs ( Mn ≈ 20 000 g·mol-1) to PDA-modified surfaces from a reactive melt. Using alkyne distal end group on the PEO chains, azidopentanoic-bearing peptides were coupled through a copper-catalyzed Huisgen azide-alkyne "click" cycloaddition reaction. The surface concentration of RGD was tuned from complete saturation of the PEO surface with peptides (1.7 × 105 fmol·cm-2) to values which may induce distinct differences in cell adhesion (<6.0 × 102 fmol·cm-2). Infrared reflection-absorption and X-ray photoelectron spectroscopies proved the PDA-PEO layers covalent structure and the immobilization of RGD peptides. The complete reconstruction of experimental electrohydrodynamics data utilizing mean-field theory predictions further verified the attained brush structure of the end-tethered PEO chains which provided hydrodynamic screening of the PDA anchor. Increasing the surface concentration of immobilized RGD peptides led to increased interfacial charging. Supported by simulations, this observation was attributed to the ionization of functional groups in the amino acid sequence and to the pH-dependent adsorption of water ions (OH- > H3O+) from the electrolyte. Despite the distinct differences observed in the electrokinetic analysis of the surfaces bearing different amounts of RGD, it was found that the peptide presence on PEO(20 000)-PDA layers does not have a significant effect on the nonfouling properties of the system. Notably, the presented PEO(20 000)-PDA layers bearing RGD peptides in the surface concentration range 5.9 to 1.7 × 105 fmol·cm-2 reduced the protein adsorption from fetal bovine serum to less than 30 ng·cm-2, that is, values comparable to the ones obtained for pristine PEO(20 000)-PDA layers.000)-PDA layers bearing RGD peptides in the surface concentration range 5.9 to 1.7 x 105 fmol.cm-2 reduced the protein adsorption from fetal bovine serum to less than 30 ng.cm-2, that is, values comparable to the ones obtained for pristine PEO(20 000)-PDA layers.

• MeSH
• adsorpce MeSH
• buněčná adheze MeSH
• fotoelektronová spektroskopie MeSH
• molekulární struktura MeSH
• peptidy * chemie   MeSH
• polyethylenglykoly * chemie   MeSH
• povrchové vlastnosti MeSH

A novel and facile in vitro cell sensing system has been developed with one-step electropolymerization of the conducting polypyrrole(PPy) polymer using RGD peptide as the sole dopant on an indium tin oxide (ITO) surface. The resulted RGD peptide-doped polypyrrole (PPy/RGD) composite film had a robust surface, in which PPy provided a biocompatible matrix for cell growth and a conducting interface for electrical detection, while the RGD peptide entrapped in the PPy matrix conferred the desired biomimetic properties. Using the human lung cancer cell A549 as a model, this system can be used to monitor cell behaviors of proliferation and cytotoxicity.

• Klíčová slova
• RGD peptidy, elektropolymerizace, polypyrrole,
• MeSH
• antitumorózní látky toxicita   MeSH
• biomimetické materiály MeSH
• biosenzitivní techniky * MeSH
• oligopeptidy MeSH
• polymery MeSH
• proliferace buněk MeSH
• pyrroly * MeSH
• skenovací elektrochemická mikroskopie MeSH
• Publikační typ
• práce podpořená grantem MeSH

The cellular adhesion receptor αvβ6-integrin is highly expressed in many cancers, e.g., pancreatic, lung, head-and-neck, cervical, bladder, and esophageal carcinoma. Multimerization of αvβ6-integrin-specific RGD peptides increases the target affinity and retention but affects biodistribution and pharmacokinetics. Amide formation of the terminal carboxylic acid moieties of the square-symmetrical bifunctional chelator DOTPI with 3-azidopropylamine yields derivatives with 4, 3, and 2 terminal azides and zero, 1, and 2 remaining carboxylic acids, respectively, whereby formation of the 2-cis-isomer is preferred according to NMR investigation of the Eu(III)-complexes. Cu(II)-catalyzed alkyne-azide cycloaddition (CuAAC) of the alkyne-functionalized αvβ6-integrin binding peptide cyclo[YRGDLAYp(NMe)K(pent-4-ynoic amide)] (Tyr2) yields the respective di-, tri-, and tetrameric conjugates for Lu-177-labeling. In mice bearing αvβ6-integrin-expressing xenografts of H2009 (human lung adenocarcinoma) cells, the Lu-177-labeled trimer's tumor-to-blood ratio of 112 exceeds that of the tetramer (10.4) and the dimer (54). Co-infusion of gelofusine (succinylated gelatin) reduces the renal uptake of the trimer by 89%, resulting in a 10-fold better tumor-to-kidney ratio, while no improvement of that ratio is observed with arginine/lysine, para-aminohippuric acid (PAH), and hydroxyethyl starch (HES) coinfusions. Since the Lu-177-labeled Tyr2-trimer outperforms the dimer and the tetramer, such trimers are considered the best lead structures for the ongoing development of αvβ6-integrin targeted anticancer theranostics.

• MeSH
• antigeny nádorové * metabolismus   MeSH
• chelátory * chemie   MeSH
• integriny * metabolismus   MeSH
• lidé MeSH
• lutecium * chemie   MeSH
• myši nahé MeSH
• myši MeSH
• nádorové buněčné linie MeSH
• nádory farmakoterapie   MeSH
• oligopeptidy * chemie   farmakokinetika   MeSH
• radiofarmaka farmakokinetika   chemie   terapeutické užití   MeSH
• radionuklidy * chemie   MeSH
• syntetická chemie okamžité shody MeSH
• teranostická nanomedicína metody   MeSH
• tkáňová distribuce MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

The surface of poly(L-lactide) (PLLA) films deposited on glass coverslips was modified with poly(DL-lactide) (PDLLA), or 1:4 mixtures of PDLLA and PDLLA-b-PEO block copolymers, in which either none, 5% or 20% of the copolymer molecules carried a synthetic extracellular matrix-derived ligand for integrin adhesion receptors, the GRGDSG oligopeptide, attached to the end of the PEO chain. The materials, perspective for vascular tissue engineering, were seeded with rat aortic smooth muscle cells (11,000 cells/cm(2)) and the adhesion, spreading, DNA synthesis and proliferation of these cells was followed on inert and bioactive surfaces. In 24-h-old cultures in serum-supplemented media, the number of cells adhering to the PDLLA-b-PEO copolymer was almost eight times lower than that on the control PDLLA surface. On the surfaces containing 5% and 20% GRGDSG-PEO-b-PDLLA copolymer, the number of cells increased 6- and 3-fold respectively, compared to the PDLLA-b-PEO copolymer alone. On PDLLA-b-PEO copolymer alone, the cells were typically round and non-spread, whereas on GRGDSG-modified surfaces the cell spreading areas approached those found on PDLLA, reaching values of 991 microm(2) and 611 microm(2) for 5% and 20% GRGDSG respectively, compared to 958 microm(2) for PDLLA. The cells on GRGDSG-grafted copolymers were able to form vinculin-containing focal adhesion plaques, to synthesize DNA and even proliferate in a serum-free medium, which indicates specific binding to the GRGDSG sequences through their adhesion receptors.

• MeSH
• biokompatibilní potahované materiály aplikace a dávkování   chemie   MeSH
• buněčná adheze účinky léků   MeSH
• buněčné kultury metody   MeSH
• financování organizované MeSH
• krysa rodu rattus MeSH
• kultivované buňky MeSH
• myocyty hladké svaloviny cytologie   fyziologie   účinky léků   MeSH
• oligopeptidy aplikace a dávkování   chemie   MeSH
• polyestery chemie   MeSH
• potkani Wistar MeSH
• povrchové vlastnosti MeSH
• proliferace buněk účinky léků   MeSH
• svaly hladké cévní cytologie   fyziologie   účinky léků   MeSH
• testování materiálů MeSH
• tkáňové inženýrství metody   MeSH
• viabilita buněk účinky léků   MeSH
• zvířata MeSH
• Check Tag
• krysa rodu rattus MeSH
• mužské pohlaví MeSH
• zvířata MeSH

To tailor cell-surface interactions, precise and controlled attachment of cell-adhesive motifs is required, while any background non-specific cell and protein adhesion has to be blocked effectively. Herein, a versatile and highly reproducible antifouling surface modification based on "clickable" groups and hierarchically structured diblock copolymer brushes for the controlled attachment of cells is reported. The polymer brush architecture combines an antifouling bottom block of poly(2-hydroxyethyl methacrylate) poly(HEMA) and an ultrathin azide-bearing top block, which can participate in well-established "click" reactions including the highly selective copper-catalyzed alkyne-azide cycloaddition (CuAAC) reaction under mild conditions. This straightforward approach allows the rapid conjugation of a cell-adhesive, alkyne-bearing cyclic RGD peptide motif, enabling subsequent specific attachment of NIH 3T3 fibroblasts, their extensive proliferation and confluent cell sheet formation after 48 h of incubation. The generally applicable strategy presented in this report can be employed for surface functionalization with diverse alkyne-bearing biological moieties via CuAAC or copper-free alkyne-azide cycloaddition protocols, making it a versatile functionalization approach and a promising tool for tissue engineering, biomaterial implant design, and other applications that require surfaces supporting highly specific cell attachment.

• MeSH
• alkyny chemie   farmakologie   MeSH
• antiinfekční látky chemická syntéza   farmakologie   MeSH
• azidy chemie   farmakologie   MeSH
• biokompatibilní materiály chemická syntéza   farmakologie   MeSH
• buňky NIH 3T3 MeSH
• cykloadiční reakce MeSH
• katalýza MeSH
• myši MeSH
• oligopeptidy chemie   MeSH
• polyhydroxyethylmethakrylát chemie   MeSH
• proliferace buněk účinky léků   MeSH
• syntetická chemie okamžité shody MeSH
• tkáňové inženýrství MeSH
• tkáňové podpůrné struktury * MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Cyclic pentapeptides containing the amino acid sequence arginine-glycine-aspartic (RGD) have been widely applied to target αvβ3 integrin, which is upregulated in various tumors during tumor-induced angiogenesis. Multimeric cyclic RGD peptides have been reported to be advantageous over monomeric counterparts for angiogenesis imaging. Here, we prepared mono-, di-, and trimeric cyclic arginine-glycine-aspartic-D-phenylalanine-lysine (c (RGDfK)) derivatives by conjugation with the natural chelator fusarinine C (FSC) using click chemistry based on copper (I)-catalyzed azide-alkyne cycloaddition (CuAAC). The αvβ3 binding properties of 68Ga-labeled mono-, di-, and trimeric c(RGDfK) peptides were evaluated in vitro as well as in vivo and compared with the references monomeric [68Ga]GaNODAGA-c(RGDfK) and trimeric [68Ga]GaFSC(suc-c(RGDfK))3. All 68Ga-labeled c(RGDfK) peptides displayed hydrophilicity (logD = -2.96 to -3.80), low protein binding and were stable in phosphate buffered-saline (PBS) and serum up to 2 h. In vitro internalization assays with human melanoma M21 (αvβ3-positive) and M21-L (αvβ3-negative) cell lines showed specific uptake of all derivatives and increased in the series: mono- < di- < trimeric peptide. The highest tumor uptake, tumor-to-background ratios, and image contrast were found for the dimeric [68Ga]GaMAFC(c(RGDfK)aza)2. In conclusion, we developed a novel strategy for direct, straight forward preparation of mono-, di-, and trimeric c(RGDfK) conjugates based on the FSC scaffold. Interestingly, the best αvβ3 imaging properties were found for the dimeric [68Ga]GaMAFC(c(RGDfK)aza)2.

• MeSH
• alkyny chemie   MeSH
• azidy chemie   MeSH
• cyklické peptidy chemie   farmakokinetika   MeSH
• izotopové značení MeSH
• měď chemie   MeSH
• myši MeSH
• nádorové buněčné linie MeSH
• PET/CT MeSH
• polymerizace MeSH
• radioizotopy galia chemie   MeSH
• siderofory chemie   MeSH
• syntetická chemie okamžité shody MeSH
• tkáňová distribuce MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• srovnávací studie MeSH

• MeSH
• peptidy farmakologie   MeSH
• techniky in vitro MeSH
• virové nemoci prevence a kontrola   MeSH
• virus slintavky a kulhavky MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH

In this study, fibrous scaffolds based on poly(γ-benzyl-l-glutamate) (PBLG) were investigated in terms of the chondrogenic differentiation potential of human tooth germ stem cells (HTGSCs). Through the solution-assisted bonding of the fibres, fully connected scaffolds with pore sizes in the range 20-400 µm were prepared. Biomimetic modification of the PBLG scaffolds was achieved by a two-step reaction procedure: first, aminolysis of the PBLG fibres' surface layers was performed, which resulted in an increase in the hydrophilicity of the fibrous scaffolds after the introduction of N5 -hydroxyethyl-l-glutamine units; and second, modification with the short peptide sequence azidopentanoyl-GGGRGDSGGGY-NH2 , using the 'click' reaction on the previously modified scaffold with 2-propynyl side-chains, was performed. Radio-assay of the 125 I-labelled peptide was used to evaluate the RGD density in the fibrous scaffolds (which varied in the range 10-3 -10 pm/cm2 ). All the PBLG scaffolds, especially with density 90 ± 20 fm/cm2 and 200 ± 100 fm/cm2 RGD, were found to be potentially suitable for growth and chondrogenic differentiation of HTGSCs. Copyright © 2015 John Wiley & Sons, Ltd.

• MeSH
• benzylové sloučeniny chemická syntéza   chemie   farmakologie   MeSH
• chrupavka účinky léků   fyziologie   MeSH
• dítě MeSH
• glutamáty chemická syntéza   chemie   farmakologie   MeSH
• glykosaminoglykany metabolismus   MeSH
• kmenové buňky cytologie   účinky léků   MeSH
• kultivované buňky MeSH
• lidé MeSH
• magnetická rezonanční spektroskopie MeSH
• mladiství MeSH
• peptidy farmakologie   MeSH
• povrchové vlastnosti MeSH
• proliferace buněk účinky léků   MeSH
• syntetická chemie okamžité shody MeSH
• tkáňové inženýrství metody   MeSH
• tkáňové podpůrné struktury chemie   MeSH
• viabilita buněk účinky léků   MeSH
• zubní zárodek cytologie   MeSH
• Check Tag
• dítě MeSH
• lidé MeSH
• mladiství MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH