Effect of Chitosan and Amphiphilic Polymers on the Photosensitizing and Spectral Properties of Rose Bengal.

The influence of chitosan (CS) and amphiphilic polymers (AP: pluronic F108 and polyvinylpyrrolidone (PVP)) on the photocatalytic activity of rose bengal (RB) in a model reaction of tryptophan photo-oxidation in phosphate-buffered saline (PBS) was studied. It was shown that in the presence of CS, the effective rate constant keff of tryptophan photo-oxidation catalyzed by RB in PBS solution decreases by a factor of two. This is due to the ionic interaction of the RB with the chitosan. Rose bengal in a slightly acidic environment (pH 4.5) passes into a neutral lactone form, which sharply reduces the photosensitizing properties of the dye. It was demonstrated that the introduction of AP into a solution containing RB and CS prevents direct interaction between RB and CS. This is evidenced by the presence of photocatalytic activity of the dye in the RB-AP-CS systems, as well as bathochromic shifts of the main absorption bands of the dye, and an increase in the optical density and luminescence intensity of the RB when AP is introduced into a buffer solution containing RB and chitosan. The presence of RB-CS and RB-AP interaction in aqueous and PBS media is confirmed by the increase in the degree of fluorescence anisotropy (r) of these binary systems. In an aqueous solution, the value of r for the RB-F108-CS system decreases by a factor of 3.5 (compared to the value of r for the RB-CS system), which is associated with the localization of the dye in pluronic micelles. In PBS, the fluorescence anisotropy is practically the same for all systems, which is related to the stability of the dye structure in this medium. The presence of interaction between RB and AP in aqueous solutions was confirmed by the proton NMR method. In addition, the formation of RB-F108 macromolecular complexes, which form associates during solution concentration (in particular, during evaporation), was shown by AFM. Such RB-AP-CS systems may be promising for practical application in the treatment of local foci of infections by aPDT.

A Hydrophobic Derivative of Ciprofloxacin as a New Photoinitiator of Two-Photon Polymerization: Synthesis and Usage for the Formation of Biocompatible Polylactide-Based 3D Scaffolds.

A hydrophobic derivative of ciprofloxacin, hexanoylated ciprofloxacin (CPF-hex), has been used as a photoinitiator (PI) for two-photon polymerization (2PP) for the first time. We present, here, the synthesis of CPF-hex and its application for 2PP of methacrylate-terminated star-shaped poly (D,L-lactide), as well a systematic study on the optical, physicochemical and mechanical properties of the photocurable resin and prepared three-dimensional scaffolds. CPF-hex exhibited good solubility in the photocurable resin, high absorption at the two-photon wavelength and a low fluorescence quantum yield = 0.079. Structuring tests showed a relatively broad processing window and revealed the efficiency of CPF-hex as a 2PP PI. The prepared three-dimensional scaffolds showed good thermal stability; thermal decomposition was observed only at 314 °C. In addition, they demonstrated an increase in Young's modulus after the UV post-curing (from 336 ± 79 MPa to 564 ± 183 MPa, which is close to those of a cancellous (trabecular) bone). Moreover, using CPF-hex as a 2PP PI did not compromise the scaffolds' low cytotoxicity, thus they are suitable for potential application in bone tissue regeneration.

Cathepsin D-Managing the Delicate Balance.

Lysosomal proteases play a crucial role in maintaining cell homeostasis. Human cathepsin D manages protein turnover degrading misfolded and aggregated proteins and favors apoptosis in the case of proteostasis disruption. However, when cathepsin D regulation is affected, it can contribute to numerous disorders. The down-regulation of human cathepsin D is associated with neurodegenerative disorders, such as neuronal ceroid lipofuscinosis. On the other hand, its excessive levels outside lysosomes and the cell membrane lead to tumor growth, migration, invasion and angiogenesis. Therefore, targeting cathepsin D could provide significant diagnostic benefits and new avenues of therapy. Herein, we provide a brief overview of cathepsin D structure, regulation, function, and its role in the progression of many diseases and the therapeutic potentialities of natural and synthetic inhibitors and activators of this protease.

Effect of Chitosan on the Activity of Water-Soluble and Hydrophobic Porphyrin Photosensitizers Solubilized by Amphiphilic Polymers.

In this work, we studied the photocatalytic activity of photosensitizers (PSs) of various natures solubilized with polyvinylpyrrolidone (PVP) and ternary block copolymer ethylene and propylene oxide Pluronic F127 (F127) in a model reaction of tryptophan photo-oxidation in water in the presence of chitosan (CT). Water-soluble compounds (dimegin and trisodium salt of chlorin e6 (Ce6)) and hydrophobic porphyrins (tetraphenylporphyrin (TPP) and its fluorine derivative (TPPF20)) were used as PSs. It was shown that the use of chitosan (Mw ~100 kDa) makes it possible to obtain a system whose activity is comparable to that of the photosensitizer-amphiphilic polymer systems. Thus, the previously observed drop in the photosensitizing activity of PS in the presence of a polysaccharide and amphiphilic polymers (AP) was absent in this case. At the same time, chitosan had practically no inhibitory effect on hydrophobic porphyrins solubilized by Pluronic F127.

Photocurable Methacrylate Derivatives of Polylactide: A Two-Stage Synthesis in Supercritical Carbon Dioxide and 3D Laser Structuring.

A two-stage polylactide modification was performed in the supercritical carbon dioxide medium using the urethane formation reaction. The modification resulted in the synthesis of polymerizable methacrylate derivatives of polylactide for application in the spatial 3D structuring by laser stereolithography. The use of the supercritical carbon dioxide medium allowed us to obtain for the first time polymerizable oligomer-polymer systems in the form of dry powders convenient for further application in the preparation of polymer compositions for photocuring. The photocuring of the modified polymers was performed by laser stereolithography and two-photon crosslinking. Using nanoindentation, we found that Young's modulus of the cured compositions corresponded to the standard characteristics of implants applied in regenerative medicine. As shown by thermogravimetric analysis, the degree of crosslinking and, hence, the local stiffness of scaffolds were determined by the amount of the crosslinking agent and the photocuring regime. No cytotoxicity was observed for the structures.

Fibrin-based Bioinks: New Tricks from an Old Dog.

For the past 10 years, the main efforts of most bioprinting research teams have focused on creating new bioink formulations, rather than inventing new printing set-up concepts. New tissue-specific bioinks with good printability, shape fidelity, and biocompatibility are based on "old" (well-known) biomaterials, particularly fibrin. While the interest in fibrin-based bioinks is constantly growing, it is essential to provide a framework of material's properties and trends. This review aims to describe the fibrin properties and application in three-dimensional bioprinting and provide a view on further development of fibrin-based bioinks.

Is it possible to combine photodynamic therapy and application of dinitrosyl iron complexes in the wound treatment?

We have studied the effect of interactions between dinitrosyl iron complexes with thiol-containing ligands (DNIC-TL) and diglucamine salt of chlorine e6 (photoditazine, PD) on the rate of photosensitized oxidation of a model organic substrate - tryptophan - in the presence and absence of an amphiphilic polymer, Pluronic F127, as well as on the DNIC-TL and PD photostability. Using EPR and UV spectroscopy, we determined the rate constants for photodegradation of mono- and dinuclear DNIC-TL and PD, respectively. The presence of the photosensitizer and Pluronic F127 has been shown to have a negligible effect on the rate of photodestruction of mono- and dinuclear DNIC-TL, taking into account the changing DNIC-TL and PD concentrations in the photoexcitation conditions. At the same time, in the DNIC-TL presence, the rate of PD photodestruction increases, however, addition of Pluronic F127 leads to a decrease in the rate constant of PD photodestruction. The latter circumstance creates an opportunity for a simultaneous application of DNIC-TL and photodynamic therapy in the wound treatment without losing the PDT efficiency. Indeed, photodynamic therapy in combination with DNIC-TL facilitated skin wound healing in laboratory rats. As shown by a morphological study, application of the DNIC-TL-PD-F127 complex with the subsequent photoactivation was beneficial in reducing inflammation and stimulating regenerative processes.

Author Correction: Optimization of Photosensitized Tryptophan Oxidation in the Presence of Dimegin-Polyvinylpyrrolidone-Chitosan Systems.

A correction to this article has been published and is linked from the HTML and PDF versions of this paper. The error has not been fixed in the paper.

Hydrogel-assisted neuroregeneration approaches towards brain injury therapy: A state-of-the-art review.

Recent years have witnessed the development of an enormous variety of hydrogel-based systems for neuroregeneration. Formed from hydrophilic polymers and comprised of up to 90% of water, these three-dimensional networks are promising tools for brain tissue regeneration. They can assist structural and functional restoration of damaged tissues by providing mechanical support and navigating cell fate. Hydrogels also show the potential for brain injury therapy due to their broadly tunable physical, chemical, and biological properties. Hydrogel polymers, which have been extensively implemented in recent brain injury repair studies, include hyaluronic acid, collagen type I, alginate, chitosan, methylcellulose, Matrigel, fibrin, gellan gum, self-assembling peptides and proteins, poly(ethylene glycol), methacrylates, and methacrylamides. When viewed as tools for neuroregeneration, hydrogels can be divided into: (1) hydrogels suitable for brain injury therapy, (2) hydrogels that do not meet basic therapeutic requirements and (3) promising hydrogels which meet the criteria for further investigations. Our analysis shows that fibrin, collagen I and self-assembling peptide-based hydrogels display very attractive properties for neuroregeneration.

Optimization of Photosensitized Tryptophan Oxidation in the Presence of Dimegin-Polyvinylpyrrolidone-Chitosan Systems.

By the example of a model process of tryptophan photooxidation in the aqueous medium in the presence of a three-component photosensitizing complex (porphyrin photosensitizer-polyvinylpyrrolidone- chitosan, PPS-PVP-CT) in the temperature range of 20-40 °Ð¡, we have demonstrated a possibility of modification of such a process by selecting different molar ratios of the components in the reaction mixture. The actual objective of this selection is the formation of a certain PPS-PVP-CT composition in which PVP macromolecules would coordinate with PPS molecules and at the same time practically block the complex binding of PPS molecules with chitosan macromolecules. Such blocking allows utilization of the bactericidal properties of chitosan to a greater extent, since chitosan is known to depress the PPS photosensitizing activity in PPS-PVP-CT complexes when using those in photodynamic therapy (PDT). The optimal composition of photosensitizing complexes appears to be dependent on the temperature at which the PDT sessions are performed. We have analyzed the correlations of the effective rate constants of tryptophan photooxidation with the photophysical characteristics of the formed complexes.

Early Effects of Ionizing Radiation on the Collagen Hierarchical Structure of Bladder and Rectum Visualized by Atomic Force Microscopy.

Radiation therapy, widely used in the treatment of a variety of malignancies in the pelvic area, is associated with inevitable damage to the surrounding healthy tissues. We have applied atomic force microscopy (AFM) to track the early damaging effects of ionizing radiation on the collagen structures in the experimental animals' bladder and rectum. The first signs of the low-dose radiation (2 Gy) effect were detected by AFM as early as 1 week postirradiation. The observed changes were consistent with initial radiation destruction of the protein matrix. The alterations in the collagen fibers' packing 1 month postirradiation were indicative of the onset of fibrotic processes. The destructive effect of higher radiation doses was probed 1 day posttreatment. The severity of the radiation damage was proportional to the dose, from relatively minor changes in the collagen packing at 8 Gy to the growing collagen matrix destruction at higher doses and complete three-dimensional collagen network restructuring towards fibrotic-type architecture at the dose of 22 Gy. The AFM study appeared superior to the optical microscopy-based studies in its sensitivity to early radiation damage of tissues, providing valuable additional information on the onset and development of the collagen matrix destruction and remodeling.

Photodynamic therapy of mouse tumor model using chlorin e6- polyvinyl alcohol complex.

The use of polymeric carriers to deliver hydrophobic photosensitizers has been widely discussed as a way to improve both fluorescence diagnostic and photodynamic therapy (PDT) of cancers; however, the photophysical and pharmacokinetic parameters, as well as the PDT activity, of such modifications have, until now, only been poorly investigated. The purpose of the present study was to explore the efficacy of PDT with the formulation of the photosensitizer chlorin e6 (Ce6) in combination with polyvinyl alcohol (PVA) in comparison with Ce6 alone and with the clinical drug, Photodithazine in a mouse tumor model. We also investigated the photoactivity of the Ce6-PVA in a model reaction of tryptophan oxidation, analyzed the polymer-Ce6 interaction using fluorescence spectroscopy and atomic-force microscopy, and tested the phototoxicity in vitro. Using fluorescence imaging in vivo we found that injection to mice of Ce6 in a formulation with PVA resulted in a higher tumor-to-normal ratio and greater photobleaching when compared with either the use of Ce6 alone, or with the effects of Photodithazine. Tumor growth study and histological examination of CT26 tumors revealed fast, reproducible tumor regression and more advanced necrosis after PDT with Ce6-PVA. The higher photoactivity of the Ce6-PVA complex was confirmed in a model reaction of tryptophan oxidation and in cultured cells. Therefore, encapsulation of Ce6 in PVA represents a promising strategy for further increasing the selectivity and efficacy of PDT.

Photocatalytic Properties of Tetraphenylporphyrins Immobilized on Calcium Alginate Aerogels.

We have prepared photocatalytic systems based on tetraphenylporphyrins (TPP) immobilized on calcium alginate solid gels in the conditions of thermal drying on air (xerogel), freeze drying in vacuum (cryogel) and supercritical drying in the supercritical carbon dioxide (scCO2) medium (aerogel). As a test reaction to measure the prepared systems' efficiency, we studied tryptophan photooxidation in the aqueous medium. We have demonstrated that the systems with aerogel as a carrier exhibited the highest photocatalytic efficiency. In that case, the rate constant for the test substrate (tryptophan) oxidation exceeds the corresponding rate constants of similar systems based on xerogel and cryogel by more than 5 times. Moreover, the aerogel-based photocatalytic systems demonstrated enhanced functional stability and a possibility of multiple use of such a catalyst in tryptophan oxidation. Based on the data of small-angle X-ray scattering and thermooxidative destruction, we have made a conclusion about the relationship between the high photocatalytic activity of aerogel-immobilized TPP and formation of a developed porous aerogel structure in the conditions of drying in the scCO2 medium, which is stabilized due to formation of additional complex bonds of calcium ions with fragments of glycoside rings.

Quantitative evaluation of atherosclerotic plaques using cross-polarization optical coherence tomography, nonlinear, and atomic force microscopy.

A combination of approaches to the image analysis in cross-polarization optical coherence tomography (CP OCT) and high-resolution imaging by nonlinear microscopy and atomic force microscopy (AFM) at the different stages of atherosclerotic plaque development is studied. This combination allowed us to qualitatively and quantitatively assess the disorganization of collagen in the atherosclerotic arterial tissue (reduction and increase of CP backscatter), at the fiber (change of the geometric distribution of fibers in the second-harmonic generation microscopy images) and fibrillar (violation of packing and different nature of a basket-weave network of fibrils in the AFM images) organization levels. The calculated CP channel-related parameters are shown to have a statistically significant difference between stable and unstable (also called vulnerable) plaques, and hence, CP OCT could be a potentially powerful, minimally invasive method for vulnerable plaques detection.

Chitosan impregnation with biologically active tryaryl imidazoles in supercritical carbon dioxide.

The presented paper is focused on impregnation of chitosan and its derivatives with a biologically active triaryl imidazole model compound ((2-2-hydroxyphenyl)-4.5-diphenyl-1H-imidazole) in the supercritical carbon dioxide medium. Since initial chitosan represents a polycation-exchange resin and does not swell in supercritical carbon dioxide, the impregnation was carried out in the presence of water (0.15-3.0 vol%). The maximum 2-2-hydroxyphenyl)-4.5-diphenyl-1H-imidazole concentration in a chitosan film was achieved at the ~5 × 10(-3) g/cm(3) water content in the reactor. We also used hydroxy carboxylic acid derivatives of chitosan and its copolymer with polylactide as matrices for introduction of hydrophobic 2-2-hydroxyphenyl)-4.5-diphenyl-1H-imidazole. We have shown that unmodified chitosan contains the greatest amount of 2-2-hydroxyphenyl)-4.5-diphenyl-1H-imidazole, as compared with its hydrophobic derivatives. The kinetics of 2-2-hydroxyphenyl)-4.5-diphenyl-1H-imidazole diffusion from a chitosan matrix was studied in acidified water with pH 1.6. We found that the complete release of 2-2-hydroxyphenyl)-4.5-diphenyl-1H-imidazole into the aqueous phase from unmodified chitosan films occurred in 48 h, while its complete release from chitosan modified with hydroxy carboxylic acids occurred in 5 min or less.

Atomic Force Microscopy Study of Atherosclerosis Progression in Arterial Walls.

Cardiovascular disease remains the leading cause of mortality worldwide. Here we suggest a novel approach for tracking atherosclerosis progression based on the use of atomic force microscopy (AFM). Using AFM, we studied cross-sections of coronary arteries with the following types of lesions: Type II-thickened intima; Type III-thickened intima with a lipid streak; Type IV-fibrotic layer over a lipid core; Type Va-unstable fibrotic layer over a lipid core; Type Vc-very thick fibrotic layer. AFM imaging revealed that the fibrotic layer of an atherosclerotic plaque is represented by a basket-weave network of collagen fibers and a subscale network of fibrils that become looser with atherosclerosis progression. In an unstable plaque (Type Va), packing of the collagen fibers and fibrils becomes even less uniform than that at the previous stages, while a stable fibrotic plaque (Vc) has significantly tighter packing. Such alterations of the collagen network morphology apparently, led to deterioration of the Type Va plaque mechanical properties, that, in turn, resulted in its instability and propensity to rupture. Thus, AFM may serve as a useful tool for tracking atherosclerosis progression in the arterial wall tissue.

Collagen structure deterioration in the skin of patients with pelvic organ prolapse determined by atomic force microscopy.

We used atomic force microscopy (AFM) to diagnose pathological changes in the extracellular matrix (ECM) of skin connective tissue in patients with pelvic organ prolapse (POP). POP is a common condition affecting women that considerably decreases the patients' quality of life. Deviations from normal morphology of the skin ECM from patients with POP occur including packing and arrangement of individual collagen fibers and arrangement of collagen fibrils. The nanoindentation study revealed significant deterioration of the mechanical properties of collagen fibril bundles in the skin of POP patients as compared with the skin of healthy subjects. Changes in the skin ECM appeared to correlate well with changes in the ECM of the pelvic ligament tissue associated with POP. AFM data on the ECM structure of normal and pathologically altered connective tissue were in agreement with results of the standard histological study on the same clinical specimens. Thus, AFM and related techniques may serve as independent or complementary diagnostic tools for tracking POP-related pathological changes of connective tissue.

Specific features of early stage of the wound healing process occurring against the background of photodynamic therapy using fotoditazin photosensitizer-amphiphilic polymer complexes.

There is a growing demand on the studies of the wound healing potentials of photodynamic therapy. Here we analyze the effects of Fotoditazin, an e6 chlorine derivative, and its complexes with amphiphilic polymers, on the early stage of wound healing in a rat model. A skin excision wound model with prevented contraction was developed in male albino rats divided into eight groups according to the treatment mode. All animals received injections of one of the studied compositions into their wound beds and underwent low-intensity laser irradiation or stayed un-irradiated. The clinical monitoring and histological examination of the wounds were performed. It has been found that all the Fotoditazin formulations have significant effects on the early stage of wound healing. The superposition of the inflammation and regeneration was the main difference between groups. The aqueous solution of Fotoditazin alone induced a significant capillary hemorrhage, while its combinations with amphiphilic polymers did not. The best clinical and morphological results were obtained for the Fotoditazin-Pluronic F127 composition. Compositions of Fotoditazin and amphiphilic polymers, especially Pluronic F127, probably, have a great potential for therapy of wounds. Their effects can be attributed to the increased regeneration and suppressed reactions changes at the early stages of repair.