Enhancing cartilage regeneration through spheroid culture and hyaluronic acid microparticles: A promising approach for tissue engineering.

Cell therapy using chondrocytes has shown promise for cartilage regeneration, but maintaining functional characteristics during in vitro culture and ensuring survival after transplantation are challenges. Three-dimensional (3D) cell culture methods, such as spheroid culture, and hydrogels can improve cell survival and functionality. In this study, a new method of culturing spheroids using hyaluronic acid (HA) microparticles was developed. The spheroids mixed with HA microparticles effectively maintained the functional characteristics of chondrocytes during in vitro culture, resulting in improved cell survival and successful cartilage formation in vivo following transplantation. This new method has the potential to improve cell therapy production for cartilage regeneration.

Non-cell adhesive hexanoyl glycol chitosan hydrogels for stable and efficient formation of 3D cell spheroids with tunable size and density.

Three-dimensional (3D) culture systems that provide a more physiologically similar environment than conventional two-dimensional (2D) cultures have been extensively developed. Previously we have provided a facile method for the formation of 3D spheroids using non-adhesive N-hexanoyl glycol chitosan (HGC) hydrogel-coated dishes, but with limitations such as low gel stability and weak mechanical properties. In this study, chemically crosslinked hydrogels were prepared by photocrosslinking of methacrylated HGCs (M-HGCs), and their spheroid-forming abilities were evaluated for long-term 3D cell cultures. The M-HGC hydrogels demonstrated not only enhanced gel stability, but also good spheroid-forming abilities. Furthermore, the M-HGC-coated dishes were effective in generating spheroids of larger size and higher cell density depending on the crosslinking density of the M-HGCs. These results indicate that our hydrogel-coated dish system could be widely applied as an effective technique to produce cell spheroids with customized sizes and densities that are essential for tissue engineering and drug screening.

Preparation and characterization of 3D human glioblastoma spheroids using an N-octanoyl glycol chitosan hydrogel.

The current 2D culture model systems developed for drug screening are not sufficient to reflect the characteristics of in vivo solid tumors. Therefore, more effective in vitro tumor model systems must be developed for translational studies on therapeutic drug screening and testing. Herein, we report a new ultra-low adhesion (ULA) hydrogel for generating 3D cancer cell spheroids as tumor models in vitro. N-octanoyl glycol chitosan (OGC) was synthesized and coated onto the surface of a typical cell culture dish. Cell spheroids were effectively formed on the OGC-coated surface, and phenotypes of the tumor cells were well maintained during culture. More importantly, U373-MG cells cultured on OGC-coated plates were more resistant to doxorubicin than cells cultured on typical plates. Our OGC-based ULA system may offer a convenient method for 3D cell culture to provide enhanced performance in cancer research, drug screening and toxicology.

Synthesis and characterization of thiolated hexanoyl glycol chitosan as a mucoadhesive thermogelling polymer.

BACKGROUND: Mucoadhesive polymers, which may increase the contact time between the polymer and the tissue, have been widely investigated for pharmaceutical formulations. In this study, we developed a new polysaccharide-based mucoadhesive polymer with thermogelling properties. METHODS: Hexanoyl glycol chitosan (HGC), a new thermogelling polymer, was synthesized by the chemical modification of glycol chitosan using hexanoic anhydride. The HGC was further modified to include thiol groups to improve the mucoadhesive property of thermogelling HGC. The degree of thiolation of the thiolated HGCs (SH-HGCs) was controlled in the range of 5-10% by adjusting the feed molar ratio. The structure of the chemically modified polymers was characterized by 1H NMR and ATR-FTIR. The sol-gel transition, mucoadhesiveness, and biocompatibility of the polymers were determined by a tube inverting method, rheological measurements, and in vitro cytotoxicity tests, respectively. RESULTS: The aqueous solution (4 wt%) of HGC with approximately 33% substitution showed a sol-gel transition temperature of approximately 41 °C. SH-HGCs demonstrated lower sol-gel transition temperatures (34 ± 1 and 31 ± 1 °Ð¡) compared to that of HGC due to the introduction of thiol groups. Rheological studies of aqueous mixture solutions of SH-HGCs and mucin showed that SH-HGCs had stronger mucoadhesiveness than HGC due to the interaction between the thiol groups of SH-HGCs and mucin. Additionally, we confirmed that the thermogelling properties might improve the mucoadhesive force of polymers. Several in vitro cytotoxicity tests showed that SH-HGCs showed little toxicity at concentrations of 0.1-1.0 wt%, indicating good biocompatibility of the polymers. CONCLUSIONS: The resultant thiolated hexanoyl glycol chitosans may play a crucial role in mucoadhesive applications in biomedical areas.

Detection of DNA immobilization and hybridization on gold/silver nanostructures using localized surface plasmon resonance.

In this study, we used localized surface plasmon resonances (LSPR) to observe a phenomenon of binding of DNA on Au and Au/Ag nanostructure arrays. Au and Au/Ag nanostructures of various geometric sizes and metal compositions were fabricated by colloidal lithography technique. The immobilization of capture DNA and subsequent hybridization with target DNA on the nanostructures caused the shift of maximum peak in LSPR spectra of the nanostructures. Using the peak shift, the immobilization of capture DNA was clearly verified in a nondestructive manner and hybridization with complementary target DNA was reliably differentiated from the non-specific binding with noncomplementary DNA. This work firmly implies that the LSPR spectra of the nanostructrues can be efficiently utilized to achieve a novel strategy for the detection of DNA on the nanostructures.

Poly(dimethyl siloxane)-based protein chip for simultaneous detection of multiple samples: use of glycidyl methacrylate photopolymer for site-specific protein immobilization.

This paper describes fabrication of a poly(dimethyl siloxane) (PDMS)-based chip to analyze multiple protein interactions utilizing glycidyl methacrylate (GMA) photopolymer for a site-specific immobilization of capture proteins in a closed system. First, using one direction channels of a PDMS mold having cross-channels, GMA micropads were prepared by photopolymerizing GMA solution by 365 nm light irradiation at predetermined positions. After the first mold was replaced with a second mold having higher height or directly without mold changing, capture proteins were allowed to be covalently immobilized onto the surface of the epoxide-activated GMA pads. Following immobilization, poly(ethylene glycol) diacrylate (PEG-DA) precursor was photopolymerized at specific regions to generate plugs for prevention of mixing between different sample injection channels, diminishing the need of a mold changing for sample injections. Final chip was assembled by connecting separated sample injection channels using a connector mold. The viability of this strategy was successfully demonstrated by simultaneous detection of two different antigen-antibody interactions.