骨髓间充质干细胞在壳聚糖凝胶中体外三维培养的实验研究

目的研究猪骨髓间充质干细胞在壳聚糖/β-甘油磷酸钠凝胶中体外三维培养的生长和增殖情况,探讨壳聚糖/β-甘油磷酸钠凝胶与骨髓间充质干细胞的细胞相容性。方法将间充质干细胞与壳聚糖/β-甘油磷酸钠凝胶混合,体外三维培养3周,采用倒置相差显微镜观察细胞在凝胶内粘附、生长和增殖等情况,复合物行组织形态学观察。结果间充质干细胞能良好地在壳聚糖/β-甘油磷酸钠凝胶内伸展、粘附、生长和增殖,细胞在新形成的组织内保持成活。结论壳聚糖/β-甘油磷酸钠凝胶与间充质干细胞具有良好的细胞相容性,可作为间充质干细胞的载体。     

Ultrathin films of charged polysaccharides assembled alternately with linear polyions

As a means of preparation of biocompatible molecular surfaces, an alternate assembly of charged polysaccharides and oppositely-charged synthetic polymers was conducted. Cationic chitosan was assembled alternately with anionic poly(sodium styrenesulfonate) (PSS) at pH 4. Regular film growth and its dependence on ionic strength were detected by the quartz crystal microbalance (QCM) method. Averaged film thicknesses for the chitosanCPSS layer were 15, 31, 46, and 69 A° , respectively, when 0, 0.25, 0.5, and 1 M of NaCl was contained in aqueous chitosan. Adsorption of chitosan did not reach saturation in 20 min at 0 M NaCl, while the adsorption became saturated within 6 min with 0.25 M NaCl. Anionic sodium chondroitin sulfate was also assembled in alternation with cationic poly(dimethyldiallylammonium chloride) (PDDA) at pH 6.5. The adsorption of chondroitin sulfate was less sensitive to ionic strength. Surface morphology of chitosan–PSS films was investigated by non-contact atomic force microscopy (AFM) observation. Maximum height difference and Ra value for a 1000 × 1000 nm area were 11 and 0.69 nm, respectively, indicating the formation of a molecularly flat surface by alternate layer-by-layer adsorption.     

Balancing mechanical strength with bioactivity in chitosan–calcium phosphate 3D microsphere scaffolds for bone tissue engineering: air- vs. freeze-drying processes

The objective of this study was to evaluate the potential benefit of 3D composite scaffolds composed of chitosan and calcium phosphate for bone tissue engineering. Additionally, incorporation of mechanically weak lyophilized microspheres within those air-dried (AD) was considered for enhanced bioactivity. AD microsphere, alone, and air- and freeze-dried microsphere (FDAD) 3D scaffolds were evaluated in vitro using a 28-day osteogenic culture model with the Saos-2 cell line. Mechanical testing, quantitative microscopy, and lysozyme-driven enzymatic degradation of the scaffolds were also studied. FDAD scaffold showed a higher concentration (p?<?0.01) in cells per scaffold mass vs. AD constructs. Collagen was ～31% greater (p?<?0.01) on FDAD compared to AD scaffolds not evident in microscopy of microsphere surfaces. Alternatively, AD scaffolds demonstrated a superior threefold increase in compressive strength over FDAD (12 vs. 4?MPa) with minimal degradation. Inclusion of FD spheres within the FDAD scaffolds allowed increased cellular activity through improved seeding, proliferation, and extracellular matrix production (as collagen), although mechanical strength was sacrificed through introduction of the less stiff, porous FD spheres.     

Covalently Bonded Chitosan on Graphene Oxide via Redox Reaction

Carbon nanostructures have played an important role in creating a new field of materials based on carbon. Chemical modification of carbon nanostructures through grafting has been a successful step to improve dispersion and compatibility in solvents, with biomolecules and polymers to form nanocomposites. In this sense carbohydrates such as chitosan are extremely valuable because their functional groups play an important role in diversifying the applications of carbon nanomaterials. This paper reports the covalent attachment of chitosan onto graphene oxide, taking advantage of this carbohydrate at the nanometric level. Grafting is an innovative route to modify properties of graphene, a two-dimensional nanometric arrangement, which is one of the most novel and promising nanostructures. Chitosan grafting was achieved by redox reaction using different temperature conditions that impact on the morphology and features of graphene oxide sheets. Transmission Electron Microscopy, Fourier Transform Infrared, Raman and Energy Dispersive spectroscopies were used to study the surface of chitosan-grafted-graphene oxide. Results show a successful modification indicated by the functional groups found in the grafted material. Dispersions of chitosan-grafted-graphene oxide samples in water and hexane revealed different behavior due to the chemical groups attached to the graphene oxide sheet.     

Induction of an in vitro reversible hypometabolism through chitosan-based nanoparticles

Objective: Chitosan-based nanoparticles (NPs) were prepared to promote intracellular sustained delivery of the synthetic delta opioid D-Ala(2)-D-Leu(5)-enkephalin (DADLE), prolonging peptide activity and inducing a safe and reversible hypometabolic state.

Materials and methods: NPs were prepared by combining ionotropic gelation and ultrasonication treatment. NP uptake studies and the effects of encapsulated DADLE on HeLa cells proliferation were tested by transmission electron microscopy (TEM) analysis, by immuno-fluorescence and immuno-cytochemistry.

Results: DADLE-loaded NPs are produced with suitable characteristics, a satisfactory process yield (55.4%?±?2.4%) and encapsulation efficiency (64.6%?±?2.1%). NPs are effective in inducing a hypometabolic stasis at a 10^?4?M DADLE concentration. Moreover, as seen from the immunofluorescence study, the effect persists through the recovery period (72?h). Indeed, NPs labelled by anti-enkephalin antibody inside cell nucleus reassert that the in vivo release of the peptide can be prolonged with respect to the case of free peptide supply.

Conclusion: The nanoparticulate drug delivery system described seems to be effective in inducing and prolonging a sort of hibernation-like state in the cells.     

Anti‐erosive effects of fluoride and phytosphingosine: an in vitro study

A selection of commercially available products containing stannous fluoride (SnF₂)/sodium fluoride (NaF), SnF₂/amorphous calcium phosphate (ACP), SnF₂/NaF/ACP, tin (Sn)/fluorine (F)/chitosan were compared with phytosphingosine (PHS) with respect to their anti‐erosive properties in vitro. One‐hundred and twenty bovine enamel specimens were immersed in the respective product slurries for 2 min, twice daily. The formulations were diluted with either remineralization solution or artificial saliva. After each treatment, an erosive challenge was performed for 10 min, twice daily, using citric acid, pH 3.4. The specimens were stored in remineralization solution or artificial saliva until the next treatment‐erosion challenge. After 10 d, tissue loss was determined using profilometry. Enamel softening was determined through surface microhardness measurements. Tissue‐loss values (measured in μm and expressed as mean ± SD) for PHS, SnF₂/NaF, SnF₂/ACP, SnF₂/ACP/NaF, and Sn/F/chitosan treatment groups and for the negative‐control group, were, respectively, 35.6 ± 2.8, 15.8 ± 1.8, 22.1 ± 2.0, 22.9 ± 1.8, 16.2 ± 1.2, and 51.2 ± 4.4 in the presence of remineralization solution and 31.7 ± 3.3, 15.6 ± 2.9, 16.5 ± 2.7, 16.8 ± 2.1, 13.1 ± 3.0, and 50.7 ± 2.8 in the presence of artificial saliva. There were no significant differences in surface microhardness measurements between the treatment groups. In conclusion, PHS resulted in a significant reduction of tissue loss compared with the negative control, but in comparison, the toothpastes containing Sn²⁺ and F^? ions were significantly more effective compared with PHS.     

Novel electrochemical xanthine biosensor based on chitosan–polypyrrole–gold nanoparticles hybrid bio-nanocomposite platform

The aim of this study was the electrochemical detection of the adenosine-3-phosphate degradation product, xanthine, using a new xanthine biosensor based on a hybrid bio-nanocomposite platform which has been successfully employed in the evaluation of meat freshness. In the design of the amperometric xanthine biosensor, chitosan–polypyrrole–gold nanoparticles fabricated by an in situ chemical synthesis method on a glassy carbon electrode surface was used to enhance electron transfer and to provide good enzyme affinity. Electrochemical studies were carried out by the modified electrode with immobilized xanthine oxidase on it, after which the biosensor was tested to ascertain the optimization parameters. The Biosensor exhibited a very good linear range of 1–200 μM, low detection limit of 0.25 μM, average response time of 8 seconds, and was not prone to significant interference from uric acid, ascorbic acid, glucose, and sodium benzoate. The resulting bio-nanocomposite xanthine biosensor was tested with fish, beef, and chicken real-sample measurements.     

A temperature-induced and shear-reversible assembly of latanoprost-loaded amphiphilic chitosan colloids: Characterization and in vivo glaucoma treatment

Hydrogels composed of assembled colloids is a material class that is currently receiving much interest and shows great promise for use in biomedical applications. This emerging material class presents unique properties derived from the combination of nanosized domains in the form of colloidal particles with a continuous gel network and an interspersed liquid phase. Here we developed an amphiphilic chitosan-based, thermogelling, shear-reversible colloidal gel system for improved glaucoma treatment and addressed how preparation procedures and loading with the anti-glaucoma drug latanoprost and commonly used preservative benzalkonium chloride influenced the mechanical properties of and drug release from the colloidal gels. The results highlight that incorporated substances and preparation procedures have effects both on mechanical properties and drug release, but that the release of drug loaded in the colloidal carriers is mainly limited by transport out of the carriers, rather than by diffusion within the gel. The developed colloidal chitosan based gels hold outstanding biomedical potential, as confirmed by the ease of preparation and administration, low cytotoxicity in MTT assay, excellent biocompatibility and lowering of intraocular pressure for 40 days in a rabbit glaucoma model. The findings clearly justify further investigations towards clinical use in the treatment of glaucoma. Furthermore, the use of this shear-reversible colloidal gel could easily be extended to localized treatment of a number of critical conditions, from chronic disorders to cancer, potentially resulting in a number of new therapeutics with improved clinical performance.     

Chitosan nanoparticle/PCL nanofiber composite for wound dressing and drug delivery

Many investigations of wound dressings equipped with drug delivery systems have recently been conducted. Chitosan is widely used not only as a material for wound dressing by the efficacy of its own, but also as a nanoparticle for drug delivery. In this study, an electrospun polycaprolactone nanofiber composite with chitosan nanoparticles (ChiNP–PCLNF) was fabricated and then evaluated for its drug release and biocompatibility to skin fibroblasts. ChiNP–PCLNF complexes showed no cytotoxicity and nanoparticles adsorbed by van der Waals force were released into aquatic environments and then penetrated into rat primary fibroblasts. Our studies demonstrate the potential for application of ChiNP–PCLNF as a wound dressing system with drug delivery for skin wound healing without side effects.