Thermosensitive composite based on agarose and chitosan saturated with carbon dioxide. Preliminary study of requirements for production of new CSAG bioink.

This study introduces a method for producing printable, thermosensitive bioink formulated from agarose (AG) and carbon dioxide-saturated chitosan (CS) hydrogels. The research identified medium molecular weight chitosan as optimal for bioink production, with a preferred chitosan hydrogel content of 40-60 %. Rheological analysis reveals the bioink's pseudoplastic behavior and a sol-gel phase transition between 27.0 and 31.5 °C. The MMW chitosan-based bioink showed also the most stable extrusion characteristic. The choice of chitosan for the production of bioink was also based on the assessment of the antimicrobial activity of the polymer as a function of its molecular weight and the degree of deacetylation, noting significant cell reduction rates for E. coli and S. aureus of 1.72 and 0.54 for optimal bioink composition, respectively. Cytotoxicity assessments via MTT and LDH tests confirm the bioink's safety for L929, HaCaT, and 46BR.1 N cell lines. Additionally, XTT proliferation assay proved the stimulating effect of the bioink on the proliferation of 46BR.1 N fibroblasts, comparable to that observed with Fetal Bovine Serum (FBS). FTIR spectroscopy confirms the bioink as a physical polymer blend. In conclusion, the CS/AG bioink demonstrates the promising potential for advanced spatial cell cultures in tissue engineering applications including skin regeneration.

Cytocompatibility, antibacterial, and corrosion properties of chitosan/polymethacrylates and chitosan/poly(4-vinylpyridine) smart coatings, electrophoretically deposited on nanosilver-decorated titania nanotubes.

The development of novel implants subjected to surface modification to achieve high osteointegration properties at simultaneous antimicrobial activity is a highly current problem. This study involved different surface treatments of titanium surface, mainly by electrochemical oxidation to produce a nanotubular oxide layer (TNTs), a subsequent electrochemical reduction of silver nitrate and decoration of a nanotubular surface with silver nanoparticles (AgNPs), and finally electrophoretic deposition (EPD) of a composite of chitosan (CS) and either polymethacrylate-based copolymer Eudragit E 100 (EE100) or poly(4-vinylpyridine) (P4VP) coating. The effects of each stage of this multi-step modification were examined in terms of morphology, roughness, wettability, corrosion resistance, coating-substrate adhesion, antibacterial properties, and osteoblast cell adhesion and proliferation. The results showed that the titanium surface formed nanotubes (inner diameter of 97 ± 12 nm, length of 342 ± 36 nm) subsequently covered with silver nanoparticles (with a diameter of 88 ± 8 nm). Further, the silver-decorated nanotubes were tightly coated with biopolymer films. Most of the applied modifications increased both the roughness and the surface contact angle of the samples. The deposition of biopolymer coatings resulted in reduced burst release of silver. The coated samples revealed potent antimicrobial activity against both Gram-positive and Gram-negative bacteria. Total elimination (99.9%) of E. coli was recorded for a sample with CS/P4VP coating. Cytotoxicity results using hFOB 1.19, a human osteoblast cell line, showed that after 3 days the tested modifications did not affect the cellular growth according to the titanium control. The proposed innovative multilayer antibacterial coatings can be successful for titanium implants as effective postoperative anti-inflammation protection.

An influence of molecular weight, deacetylation degree of chitosan xerogels on their antimicrobial activity and cytotoxicity. Comparison of chitosan materials obtained using lactic acid and CO2 saturation.

This paper presents a comparison of the antimicrobial activity and cytotoxicity against L929 cells of chitosan xerogels prepared by dissolving the polymer in a solution of lactic acid (LA) or carbonic acid (CO2) and then freeze-drying. There was no simple relationship between the antimicrobial activity and cytotoxicity of the samples obtained using both techniques (LA and CO2). Chitosan materials obtained by the LA method in a 1:1 dilution were characterized by the highest cytotoxicity against L929 cells (∼20%). For the same diluted samples prepared using the CO2 saturation method, the viability of L929 cells was approximately 2.5 times greater. Some of the tested chitosan materials obtained by the innovative method were characterized by significantly lower antimicrobial activity, for example, reduction of E. coli bacteria for MMW-LA and MMW-CO2 samples by 6.00 and 0.75 logarithmic order, respectively. This clearly indicates that in many applications, the presence of the acid necessary to dissolve chitosan is responsible for the antimicrobial activity of the polymer solution and its products.

Application of 3D- printed hydrogels in wound healing and regenerative medicine.

Hydrogels are three-dimensional polymer networks with hydrophilic properties. The modifiable properties of hydrogels and the structure resembling living tissue allow their versatile application. Therefore, increasing attention is focused on the use of hydrogels as bioinks for three-dimensional (3D) printing in tissue engineering. Bioprinting involves the fabrication of complex structures from several types of materials, cells, and bioactive compounds. Stem cells (SC), such as mesenchymal stromal cells (MSCs) are frequently employed in 3D constructs. SCs have desirable biological properties such as the ability to differentiate into various types of tissue and high proliferative capacity. Encapsulating SCs in 3D hydrogel constructs enhances their reparative abilities and improves the likelihood of reaching target tissues. In addition, created constructs can simulate the tissue environment and mimic biological signals. Importantly, the immunogenicity of scaffolds is minimized through the use of patient-specific cells and the biocompatibility and biodegradability of the employed biopolymers. Regenerative medicine is taking advantage of the aforementioned capabilities in regenerating various tissues- muscle, bones, nerves, heart, skin, and cartilage.

Osteoblast and bacterial cell response on RGD peptide-functionalized chitosan coatings electrophoretically deposited from different suspensions on Ti13Nb13Zr alloy.

Metallic materials for long-term load-bearing implants still do not provide high antimicrobial activity while maintaining strong compatibility with bone cells. This study aimed to modify the surface of Ti13Nb13Zr alloy by electrophoretic deposition of a chitosan coating with a covalently attached Arg-Gly-Asp (RGD) peptide. The suspensions for coating deposition were prepared in two different ways either using hydroxyacetic acid or a carbon dioxide saturation process. The coatings were deposited using a voltage of 10 V for 1 min. The prepared coatings were examined using SEM, EDS, FTIR, and XPS techniques. In addition, the wettability of these surfaces, corrosion resistance, adhesion of the coatings to the metallic substrate, and their antimicrobial activity (E. coli, S. aureus) and cytocompatibility properties using the MTT and LDH assays were studied. The coatings produced tightly covered the metallic substrate. Spectroscopic studies confirmed that the peptide did not detach from the chitosan chain during electrophoretic deposition. All tested samples showed high corrosion resistance (corrosion current density measured in nA/cm2 ). The deposited coatings contributed to a significant increase in the antimicrobial activity of the samples against Gram-positive and Gram-negative bacteria (reduction in bacterial counts from 99% to, for CS-RGD-Acid and the S. aureus strain, total killing capacity). MTT and LDH results showed high compatibility with bone cells of the modified surfaces compared to the bare substrate (survival rates above 75% under indirect contact conditions and above 100% under direct contact conditions). However, the adhesion of the coatings was considered weak.

Antibacterial properties of laser-encapsulated titanium oxide nanotubes decorated with nanosilver and covered with chitosan/Eudragit polymers.

To provide antibacterial properties, the titanium samples were subjected to electrochemical oxidation in the fluoride-containing diethylene glycol-based electrolyte to create a titanium oxide nanotubular surface. Afterward, the surface was covered by sputtering with silver 5 nm film, and the tops of the nanotubes were capped using laser treatment, resulting in an appearance of silver nanoparticles (AgNPs) of around 30 nm in diameter on such a modified surface. To ensure a controlled release of the bactericidal substance, the samples were additionally coated with a pH-sensitive chitosan/Eudragit 100 coating, also exhibiting bactericidal properties. The modified titanium samples were characterized using SEM, EDS, AFM, Raman, and XPS techniques. The wettability, corrosion properties, adhesion of the coating to the substrate, the release of AgNPs into solutions simulating body fluids at different pH, and antibacterial properties were further investigated. The obtained composite coatings were hydrophilic, adjacent to the surface, and corrosion-resistant. An increase in the amount of silver released as ions or metallic particles into a simulated body fluid solution at acidic pH was observed for modified samples with the biopolymer coating after three days of exposure avoiding burst effect. The proposed modification was effective against both Gram-positive and Gram-negative bacteria.

A Novel Method of Endotoxins Removal from Chitosan Hydrogel as a Potential Bioink Component Obtained by CO2 Saturation.

The article presents a new approach in the purification of chitosan (CS) hydrogel in order to remove a significant amount of endotoxins without changing its molecular weight and viscosity. Two variants of the method used to purify CS hydrogels from endotoxins were investigated using the PyroGene rFC Enzymatic Cascade assay kit. The effect of the CS purification method was assessed in terms of changes in the dynamic viscosity of its hydrogels, the molecular weight of the polymer, microbiological purity after refrigerated storage and cytotoxicity against L929 cells based on the ISO 10993-5:2009(E) standard. The proposed purification method 1 (M1) allows for the removal of significant amounts of endotoxins: 87.9-97.6% in relation to their initial concentration in the CS hydrogel without affecting the solution viscosity. Moreover, the final solutions were sterile and microbiologically stable during storage. The M1 purification method did not change the morphology of the L929 cells.

A novel method of creating thermoplastic chitosan blends to produce cell scaffolds by FDM additive manufacturing.

Due to its remarkable and promising biological and structural properties, chitosan has been widely studied in several potential applications in the biomedical sector. Attempts are being made to use this polymer and its properties in thermoplastics dedicated to 3D printing in FDM technology. However, chitosan can be processed only from acid solution, which limits its applications. The paper presents a new path for the production of filaments based on unstable chitosan hydrogel obtained by carbon dioxide saturation, as well as synthetic polymers such as polyvinyl alcohol and polycaprolactone. The results confirm that the absence of acid allows formation of thermally stable and printable filaments containing from 5% to 15% of chitosan, capable of reducing S. aureus and E. coli bacteria by 0.41-1.43 in logarithmic scale (56-94%) and 0.28-0.94 in logarithmic scale (36-89%), respectively.