Characterization and biodegradation of chitosan-alginate polyelectrolyte complexes

Polyelectrolyte complexes (PECs) have been the focus of an expanding number of studies for their wide use. This study investigated the characteristics and biodegradation of chitosan-alginate PECs prepared by freeze-drying a precipitate from sufficient mixtures of the two polymers. The analyses of X-ray photoelectron spectroscopy (XPS), Fourier transform infrared spectroscopy (FTIR), X-ray diffraction (XRD) and differential scanning calorimetry (DSC) suggested that the partial protonated amine groups of chitosan reacted with the carboxylate groups of alginate and thus strong PECs were formed. After incubating in lysozyme solution, the PECs showed high ability of enzyme adsorption, and low degradation rate in spite of different degrees of deacetylation of chitosan, due to the strong interaction between chitosan and alginate and the hindrance of closely adsorbed lysozyme.     

Chitosan based polyelectrolyte complexes

Polyelectrolyte complexes (PECs) are formed through the electrostatic interactions between polymers carrying opposite charges. Here are presented results of basic studies on the PECs of chitosan with other polysaccharides such as sodium alginate, carboxymethyl cellulose, polygalacturonic acid and κ‐carrageenan. An extensive study on chitosan/carboxymethyl cellulose membranes, regarding its swelling characteristics and water vapour sorption is offered. Also the interaction of chitosan with polyacrylic acid has been examined from the thermodynamic point of view.     

Comprehensive study on the formation of polyelectrolyte complexes from (quaternized) poly[2‐(dimethylamino)ethyl methacrylate] and poly(2‐acrylamido‐2‐methylpropane sodium sulfonate)

Polyelectrolyte complexes (PECs) have been prepared from well‐defined (quaternized) poly[2‐(dimethylamino)ethyl methacrylate] (PDMAEMA) and high molecular weight poly(2‐acrylamido‐2‐methylpropane sodium sulfonate) (PAMPSNa) after a thorough study of their viscometric properties. The effect of pH and quaternization degree of PDMAEMA on PECs stoichiometry has been examined. PEC‐based materials have been characterized in terms of thermal stability, equilibrium swelling degree, and free/bound water composition. The stoichiometry and swellability of the physically crosslinked hydrogels obtained from fully quaternized PDMAEMA/PAMPSNa complexes do not depend on pH. In contrast, PECs made of non quaternized PDMAEMA and PAMPSNa are highly affected by pH, and could reversibly disintegrate at pH ≥ 9. Partially quaternized PDMAEMA/PAMPSNa PECs exhibit intermediate properties and form stable loose structures in the whole investigated pH range. Finally, stable dispersions of PECs nanoparticles have been successfully produced from dilute solutions of the complementary polyelectrolytes. The nanoparticle average diameter as determined by dynamic light scattering proved to depend on the molar fraction of DMAEMA‐based subunits and on the initial polyelectrolyte concentration. © 2006 Wiley Periodicals, Inc. J Polym Sci Part A: Polym Chem 44: 5468–5479, 2006     

Interaction of hepatocytes with polyelectrolyte complex(I): the effect of nonspecific interaction on adhesion of the cell

The adhesion of hepatocytes to polyelectrolyte complex (PEC), prepared by mixing of aqueous solutions of polycation and polyanion, is discussed. Four PECs, poly((dimethyliminio)ethylene(dimethyliminio)methylene‐1,4‐phenylenemethylene dichloride)–poly(acrylic acid)2X–(PAA), 2X‐poly(acrylic acid–co‐2‐ethylhexylacrylate(COA), 2X‐poly(acrylicacid–co‐butylacrylate) (CBA) and 2X‐poly(acrylic acid–co‐laurylacrylate)(CLA) were prepared. Hydrophobic properties of these PECs increased in the order of 2X–PAA < 2X–COA < 2X–CLA ≤ 2X–CBA. About all the hepatocytes adhered rapidly to various PECs even in the absence of serum, while fewer cells adhered to polystrene (tissue culture grade) dishes. At 37 °C (biological condition) about 70–80% of cells adhered to a collagen‐coated dish, but at 4 °C (nonbiological condition), no cell adhered to it. Nonactive cells (prepared by a single cycle of freezing and thawing) did not adhere to collagen, either. On the contrary about 40% of cells adhered to PEC‐coated dishes even at 4 °C, and nonactive cells also adhered to them. Cytocharosine B and colchicine, which are known as inhibitors of the polymerization of intracellular matrix, did not prevent cell adhesion to PECs. From these results it was suggested that hepatocytes adhered to PEC‐coated dishes mainly through a nonbiological interaction. Copyright © 1999 John Wiley & Sons, Ltd.     

Exploring peptide‐functionalized alginate scaffolds for engineering cardiac tissue from human embryonic stem cell‐derived cardiomyocytes in serum‐free medium

Engineering human cardiac tissue is a promising solution for myocardial repair of injured hearts and for drug screening. Herein, we examined the capability of chemically defined alginate scaffolds to promote cardiac tissue regeneration from human embryonic stem cell‐derived cardiomyocytes (hESC‐CMs) in serum‐free, chemically defined medium. The cells were single seeded or coseeded with human dermal fibroblasts (HFs) in macroporous scaffolds made from pristine alginate or alginate modified with arginine‐glycine‐aspartate (RGD) peptide and heparin‐binding peptide (HBP). Our results show that the addition of fibroblasts to the 3‐D culture is indispensable for the formation of functional cardiac tissues and that the presence of RGD/HBP attached to the alginate matrix further improves its functionality. The engineered tissue displayed the typical fiber morphology with massive striation. An increase in contraction amplitude and calcium transients with time, together with a decrease in excitation threshold, indicated advancement toward tissue maturation. Our results thus point to the importance of co‐cultivating fibroblasts with hESCs‐CMs in chemically defined peptide‐functionalized alginate scaffolds and culture medium for regenerating functional cardiac tissue in vitro.     

Production of mitogen-contamination free alginates with variable ratios of mannuronic acid to guluronic acid by free flow electrophoresis.

Commercial alginates consisting of variable homopolymeric regions of beta-D-mannuronic acid and alpha-L-guluronic acid, interspaced with regions of alternating blocks, are potent stimulators of macrophages and lymphocytes. Therefore, inflammatory reactions and fibrotic overgrowth of the beads result if Langerhans islets are encapsulated in raw alginate hydrogel beads (cross-linked with divalent cations). The result is random failure of the islets some time after transplantation. Analysis of raw alginates by using free flow electrophoresis demonstrated that commercial alginates contained at least 10-20 fractions (characterized by different electrophoretic mobilities) which showed mitogenic activity. These fractions could be quantitatively separated from the alginic acids by free flow electrophoresis on a preparative scale. The purified alginates cross-linked with Ca2+ ions exhibited no mitogenic reactions as proved by an in vitro assay. In addition, examination of purified Ba2+ alginate beads implanted intraperitoneally in rats or mice for three weeks showed no fibrotic overgrowth in contrast to implants made from unpurified alginate.     

Polypyrrole‐coated alginate/magnetite nanoparticles composite sorbent for the extraction of endocrine‐disrupting compounds

Magnetite nanoparticles incorporated into alginate beads and coated with a polypyrrole adsorbent were prepared (polypyrrole/Fe₃O₄/alginate bead) and used as an effective magnetic solid‐phase extraction sorbent for the extraction and enrichment of endocrine‐disrupting compounds (estriol, β‐estradiol and bisphenol A) in water samples. The determination of the extracted endocrine‐disrupting compounds was performed using high‐performance liquid chromatography with a fluorescence detector. The effect of various parameters on the extraction efficiency of endocrine disrupting compounds were investigated and optimized including the type and amount of sorbent, sample pH, extraction time, stirring speed, and desorption conditions. Under optimum conditions, the calibration curves were linear in the concentration range of 0.5–100.0 μg/L, and the limit of detection was 0.5 μg/L. The developed method showed a high extraction efficiency, the recoveries were in the range of 90.5 ± 4.1 to 98.2 ± 5.5%. The developed sorbent was easy to prepare, was cost‐effective, robust, and provided a good reproducibility (RSDs < 5%), and could be reused 16 times. The developed method was successfully applied for the determination of endocrine‐disrupting compounds in water samples.     

Spectroscopic constants and molecular properties of the X2Π and a4Σ− electronic states of the SiF radical

The potential energy curves (PECs) of the X²Π and a⁴Σ^? electronic states of the SiF radical have been studied by an ab initio quantum chemical method. The calculations have been made using the complete active space self‐consistent field (CASSCF) method, which is followed by the valence internally contracted multireference configuration interaction (MRCI) approach in combination with several correlation‐consistent basis sets. The effects on the PECs by the core‐valence correlation and relativistic corrections are included. The way to consider the relativistic correction is to use the third‐order Douglas–Kroll Hamiltonian approximation. The relativistic corrections are made at the level of cc‐pV5Z basis set. The core‐valence correlation corrections are performed using the cc‐pCV5Z basis set. To obtain more reliable results, the PECs determined by the MRCI calculations are also corrected for size‐extensivity errors by means of the Davidson modification (MRCI+Q). These PECs are extrapolated to the complete basis set limit by the total‐energy extrapolation scheme. Using these PECs, the spectroscopic parameters are determined and compared with those reported in the literature. With these PECs obtained by the MRCI+Q/CV+DK+56 calculations, the vibrational levels, inertial rotation, and centrifugal distortion constants of the first 20 vibrational state of each electronic state are calculated when the rotational quantum number J equals zero. Comparison with the Rydberg‐Klein‐Rees (RKR) data shows that the present results are reliable and accurate. The molecular constants of the X²Π and a⁴Σ^? electronic states determined by the MRCI+Q/CV+DK+56 calculations should be good prediction for future laboratory experiment. © 2011 Wiley Periodicals, Inc. Int J Quantum Chem, 2011     

Solar Ultraviolet Radiation Exposure Proxy‐estimated by Sky View Fish‐eye Photography—Potentials and Limitations from an Exploratory Correlation Study

Potentials and limitations of sky view fish‐eye photography and calculation of the percentage of free sky (sky view factor, SVF) as a proxy to estimate solar ultraviolet radiation (UVR) exposure in shade settings are investigated using controlled situations. SVF and measured solar UVR exposure correlated at high mid‐latitude (56.65°N) near autumnal equinox in September. The correlation was enhanced by splitting the sky view images into a south‐ and a north‐half and weighting the south‐half higher to account for the direct sun. Sky view images from eight different settings with SVF‐values 98.3% – 14.9% were compared to exposure measurements by polysulphone film dosimeter badges in the horizontal zenith‐, vertical‐south‐, east‐, west‐ and north‐directions and their combinations. The sky view images were un‐split and un‐weighted or split and the semi‐skies given south/north weights (3.0/1.0) or a higher weight ratio (3.5/0.5). Of all tested combinations split sky view SVFs weighted 3.0/1.0 and compared to horizontal (zenith‐oriented) dosimeters yielded the highest correlation (R² = 0.96). The weight ratio (3.5/0.5) yielded the 2^nd highest correlation (R² = 0.90) both compared to measured horizontal exposure and compared to the horizontal exposure averaged with the vertical‐south‐oriented exposure. SVF from sky view fish‐eye photography may estimate solar UVR exposure in shade settings.     

Synthesis and Characterization of Chitosan‐Based Polyelectrolyte Complexes Doped with Xanthene Dyes

Biocompatible chitosan‐based polyelectrolyte complexes (PECs) doped with xanthene dyes (fluorescein, eosin Y, erythrosin B, rhodamine 6G) were synthesized and characterized by scanning electron microscopy, dynamic light scattering, zeta potential measurements, and absorption and luminescence (including polarized, time‐resolved, and phosphorescence) spectroscopy. The results are discussed in terms of the mechanism and rigidity of dye–PEC binding, the heavy‐atom effect in dyes and PEC stability. Eosin Y is found to be the optimal dopant, providing both a high dye content in PECs and a high quantum yield of fluorescence.     

Sodium alginate/magnesium oxide nanocomposite scaffolds for bone tissue engineering

A simple 2‐step method, consisting of film casting and polyvinyl alcohol leaching, is proposed to prepare magnesium oxide (MO) nanoparticle‐reinforced sodium alginate scaffolds with right properties for bone tissue engineering. The cytocompatibility of the as‐prepared scaffolds was also evaluated using the 3‐(4,5‐dimethylthiazol‐2‐yl)‐2,5‐diphenyltetrazolium‐bromide yellow tetrazole assay test, wherein chondrocyte cells had been considered as target cells. According to the results, the ensuing sodium alginate nanocomposites, containing 4‐wt% MO nanoparticles, demonstrated the highest physical and mechanical properties after leaching step. The Young modulus of sodium alginate/4‐wt% MO was improved about 44%, in comparison with that of the pure alginate sample. Furthermore, incorporating MO nanoparticles up to 4 wt% controlled the liquid uptake capacity of scaffolds vis‐à‐vis the resultant pure sodium alginate sample. Moreover, with increasing the nanoparticle content, the antibacterial properties of scaffolds enhanced, but their degradation rates under in vitro conditions tapered off. With the introduction of 3‐ and 4‐wt% MO, the average diameter of the bacterial zone of the scaffold samples reduced to less than 10 mm², suggesting an insensitive antimicrobial performance, compared with the pure sodium alginate and the samples with 1‐ and 2‐wt% MO content, which exhibit antimicrobial sensitivity. 3‐(4,5‐Dimethylthiazol‐2‐yl)‐2,5‐diphenyltetrazolium‐bromide assay test also revealed the cultivated chondrocyte cells on the 4‐wt% MO nanoparticle‐reinforced scaffold possessed better interaction as well as appropriate cell attachment and proliferation than the pristine sodium alginate sample.     

Chinese Oak Tasar Silkworm Antheraea pernyi Silk Proteins: Current Strategies and Future Perspectives for Biomedical Applications

Chinese nonmulberry temperate oak tasar/tussah, Antheraea pernyi (Ap) silk is a natural biopolymer that has attracted considerable attention as a biomaterial. The proteinaceous components of Ap silk proteins, namely fibroin and sericin may represent an alternative over mulberry Bombyx mori silk proteins. In fact, the silk fibroin (SF) of Ap is rich in Arginyl‐Glycyl‐Aspartic acid (RGD) peptides, which facilitate the adhesion and proliferation of various cell types. The possibility of processing Ap silk proteins into different distinct 2D‐ and 3D‐based matrices is described in earlier studies, such as membranes, nanofibers, scaffolds, and micro/nanoparticles, contributing to a different rate of degradation, mechanical properties, and biological performance useful for various biomedical applications. This review summarizes the current advances and developments on nonmulberry Chinese oak tasar silk protein (fibroin and sericin)‐based biomaterials and their potential uses in tissue engineering, regenerative medicine, and therapeutic delivery strategies.     

Involvement of excited triplet state in the photodissociation of cyclobutane

The potential energy curves (PECs) of the ground state and the low‐lying excited states for the photodissociation of cyclobutane have been calculated at the multi‐reference configuration interaction with singlet and doublet excitation (MRCISD) and the multi‐reference second order perturbation theory (MRPT2). Firstly, the PECs are constructed following a reaction path determined by semiclassical dynamics simulation, which suggests that the lowest triplet state of tetramethylene is involved in the photodissociation of cyclobutane. Then, the adiabatic PECs are calculated for the breaking processes of C1? C3 and C2? C4 bond respectively. The singlet‐triplet PECs' intersections have been found in the two breaking C? C bond processes. During the breaking process of the second C2? C4 bond, a local minimum has been found on the PEC of the lowest triplet state, which gives us some insight to reinterpret the experimental observed diradical intermediate as being trapped in its triplet state. © 2007 Wiley Periodicals, Inc. Int J Quantum Chem, 2008     

Polyelectrolyte Complexes Formed by Calf Thymus DNA and Aliphatic Ionenes: Unexpected Change in Stability upon Variation of Chain Length of Ionenes of Different Charge Density

The formation of polyelectrolyte complexes (PECs) between aliphatic 2,4‐ and 2,8‐ionenes and calf thymus DNA in water–salt solutions is monitored by a fluorescence‐quenching technique using an ethidium bromide probe. As expected, the tolerance of PEC to dissociation in water–salt solutions increased with an increase in both the number of charged groups, i. e., degree of polymerization of the ionenes (DP), and in the charge density of their chains. However, the latter tendency weakened markedly upon the lengthening of the ionenes chains and ceased to hold at DP ≈ 25. Comparison of the stability of PECs formed by DNA and poly(methacrylate) anion, respectively, with the ionenes, disclosed that relatively short ionene chains formed noticeably less stable DNA‐containing PECs, whereas upon lengthening of the ionenes the difference in the stability of the PEC successively decreased and evened out at DP ≈ 25, in the case of 2,8‐ionene. All these findings imply that in such systems the role of charge–charge conformity of the partners in DNA‐containing PECs is evident only for rather short ionenes. The unexpected changes in PEC stability revealed suggest that the short chains of the ionenes are of special interest for the design of self‐assembling polyelectrolyte systems with controllable stability attractive for biomedical applications and bioseparation.     

Development of Gelatin‐Alginate Hydrogels for Burn Wound Treatment

Hydrogels are interesting as wound dressing for burn wounds to maintain a moist environment. Especially gelatin and alginate based wound dressings show strong potential. Both polymers are modified by introducing photocrosslinkable functionalities and combined to hydrogel films (gel‐MA/alg‐MA). In one protocol gel‐MA films are incubated in alg‐MA solutions and crosslinked afterward into double networks. Another protocol involves blending both and subsequent photocrosslinking. The introduction of alginate into the gelatin matrix results in phase separation with polysaccharide microdomains in a protein matrix. Addition of alg(‐MA) to gel‐MA leads to an increased swelling compared to 100% gelatin and similar to the commercial Aquacel Ag dressing. In vitro tests show better cell adhesion for films which have a lower alginate content and also have superior mechanical properties. The hydrogel dressings exhibit good biocompatibility with adaptable cell attachment properties. An adequate gelatin‐alginate ratio should allow application of the materials as wound dressings for several days without tissue ingrowth.     

A High Cell‐Bearing Capacity Multibore Hollow Fiber Device for Macroencapsulation of Islets of Langerhans

Macroencapsulation of islets of Langerhans is a promising strategy for transplantation of insulin‐producing cells in the absence of immunosuppression to treat type 1 diabetes. Hollow fiber membranes are of interest there because they offer a large surface‐to‐volume ratio and can potentially be retrieved or refilled. However, current available fibers have limitations in exchange of nutrients, oxygen, and delivery of insulin potentially impacting graft survival. Here, multibore hollow fibers for islets encapsulation are designed and tested. They consist of seven bores and are prepared using nondegradable polymers with high mechanical stability and low cell adhesion properties. Human islets encapsulated there have a glucose induced insulin response (GIIS) similar to nonencapsulated islets. During 7 d of cell culture in vitro, the GIIS increases with graded doses of islets demonstrating the suitability of the microenvironment for islet survival. Moreover, first implantation studies in mice demonstrate device material biocompatibility with minimal tissue responses. Besides, formation of new blood vessels close to the implanted device is observed, an important requirement for maintaining islet viability and fast exchange of glucose and insulin. The results indicate that the developed fibers have high islet bearing capacity and can potentially be applied for a clinically applicable bioartificial pancreas.     

Biofabrication of Cell‐Loaded 3D Spider Silk Constructs

Biofabrication is an emerging and rapidly expanding field of research in which additive manufacturing techniques in combination with cell printing are exploited to generate hierarchical tissue‐like structures. Materials that combine printability with cytocompatibility, so called bioinks, are currently the biggest bottleneck. Since recombinant spider silk proteins are non‐immunogenic, cytocompatible, and exhibit physical crosslinking, their potential as a new bioink system was evaluated. Cell‐loaded spider silk constructs can be printed by robotic dispensing without the need for crosslinking additives or thickeners for mechanical stabilization. Cells are able to adhere and proliferate with good viability over at least one week in such spider silk scaffolds. Introduction of a cell‐binding motif to the spider silk protein further enables fine‐tuned control over cell–material interactions. Spider silk hydrogels are thus a highly attractive novel bioink for biofabrication.     

Fibrin glue increases the cell survival and the transduced gene product secretion of the ceiling culture-derived adipocytes transplanted in mice

The development of clinically applicable scaffolds is important for the application of cell transplantation in various human diseases. The aims of this study are to evaluate fibrin glue in a novel protein replacement therapy using proliferative adipocytes and to develop a mouse model system to monitor the delivery of the transgene product into the blood and the fate of the transduced cells after transplantation. Proliferative adipocytes from mouse adipose tissue were transduced by a retroviral vector harboring the human lecithin-cholesterol acyltransferase (lcat) gene, and were subcutaneously transplanted into mice combined with fibrin glue. The lcat gene transduction efficiency and the subsequent secretion of the product in mouse adipocytes were enhanced using a protamine concentration of 500 μg/ml. Adipogenesis induction did not significantly affect the lcat gene-transduced cell survival after transplantation. Immunohistochemistry showed the ectopic enzyme production to persist for 28 days in the subcutaneously transplanted gene- transduced adipocytes. The increased viability of transplanted cells with fibrin glue was accompanied with the decrease in apoptotic cell death. The immunodetectable serum LCAT levels in mice implanted with the fibrin glue were comparable with those observed in mice implanted with Matrigel, indicating that the transplanted lcat gene-transduced adipocytes survived and functioned in the transplanted spaces with fibrin glue as well as with Matrigel for 28 days. Thus, this in vivo system using fibrin is expected to serve as a good model to further improve the transplanted cell/scaffold conditions for the stable and durable cell-based replacement of defective proteins in patients with LCAT deficiency.     

Tm3+‐Sensitized NIR‐II Fluorescent Nanocrystals for In Vivo Information Storage and Decoding

In vivo fluorescence imaging in the second near‐infrared window (NIR‐II) affords deep‐tissue penetration and high spatial resolution. Herein, we present a new type of Tm³⁺‐sensitized lanthanide nanocrystals with both excitation (1208 nm) and emission (1525 nm) located in the NIR‐II window for in vivo optical information storage and decoding. Taking advantage of the tunable fluorescence lifetimes, the optical multiplexed encoding capacity is enhanced accordingly. Micro‐devices with QR codes featuring the NIR‐II fluorescence‐lifetime multiplexed encoding were implanted into mice and were successfully decoded through time‐gated fluorescence imaging technology.     

In vitro and in vivo evaluation of silk fibroin-hardystonite-gentamicin nanofibrous scaffold for tissue engineering applications

Designing advanced biomaterials with regenerative and drug delivering functionalities remains a challenge in the field of tissue engineering. In this paper we present the design, development, and a use case of an electrospun nano-biocomposite scaffold composed of silk fibroin (SF), hardystonite (HT), and gentamicin (GEN). The fabricated SF nanofiber scaffolds provide mechanical support while HT acts as a bioactive and drug carrier, on which GEN is loaded as an antibacterial agent. Antibacterial zone of inhibition (ZOI) results indicate that the inclusion of 3–6 wt% GEN significantly improves the antibacterial performance of the scaffolds against Gram-negative Escherichia coli (E. coli) and Gram-positive Staphylococcus aureus (S. aureus) bacteria, with an initial burst release of 10–20% and 72–85% total release over 7 days. The release rate of stimulatory silicon ions from SF-HT scaffolds reached 94.53±5 ppm after 7 days. Cell studies using osteoblasts show that the addition of HT significantly improved the cytocompatibility of the scaffolds. Angiogenesis, in vivo biocompatibility, tissue vascularization, and translatability of the scaffolds were studied via subcutaneous implantation in a rodent model over 4-weeks. When implanted subcutaneously, the GEN-loaded scaffold promoted angiogenesis and collagen formation, which suggests that the scaffold may be highly beneficial for further bone tissue engineering applications.