Yuji Yin

β-TCP/MCPM-based premixed calcium phosphate cements

Novel premixed calcium phosphate cements (CPCs) were prepared by combining cement liquids comprised of glycerol or polyethylene glycol with CPC powders that consisted of beta-tricalcium phosphate (beta-TCP) and monocalcium phosphate monohydrate (MCPM). Changing the powder to liquid mass ratio enabled the setting time to be regulated, and improved the compressive strength of the CPCs. Although some ratios of the new premixed CPCs had long setting times, these ranged from 12.4 to 27.8 min which is much shorter than the hour or more reported previously for a premixed CPC. The premixed CPCs had tolerable washout resistance before final setting, and the cements had strengths matching that of cancellous bone (5-10 MPa); their maximum compressive strength was up to 12 MPa. The inflammatory response around the premixed CPCs implanted in the subcutaneous tissue in rabbits was more prominent than that of apatite cement. These differences might be due to the much faster resorption rate of the premixed CPCs.

Modulation of mesenchymal stem cells behaviors by chitosan/gelatin/pectin network films

In this article, the chitosan/gelatin/pectin (CGP) network films were prepared to build appropriate physicochemical and mechanical microenvironment for attachment, proliferation, and differentiation of mesenchymal stem cells (MSCs). Results suggested that the hydrophilicity and mechanical character of CGP composites films could be modulated via adjusting the pectin content in the composites. The investigations of attachment and proliferation behaviors of mesenchymal stem cells (MSCs) on the CGP films were carried out. The morphology of cells was observed with hematoxylin/eosin staining (HE) and scanning electron microscope (SEM). The osteogenic differentiation of MSCs was investigated via ALP and polymerase chain reaction (PCR). Results suggested that the CGP films have excellent biocompatibility. MSCs seeded on CGP (0.1) film show higher proliferation capacity compared with other samples. Moreover, osteogenic differentiation of MSCs also depends on the properties of the substrate. The MSCs seeded on CGP (0.5) expressed the highest ALP activity, osteogenic gene expression and mineral formation capacity. These results suggest that the composition of the CGP network films could effectively modulate their physicochemical and mechanical properties and further regulate the cell behaviors of MSCs.

High mechanical strength chitosan-based hydrogels cross-linked with poly(ethylene glycol)/polycaprolactone micelles for the controlled release of drugs/growth factors

High mechanical strength hydrogels without burst release are known to be beneficial to deliver bioactive molecules including drugs and growth factors. Herein, chitosan-based hydrogels are fabricated by the covalent attachment of poly(ethylene glycol) diacrylate (PEGDA) to thiol groups of thiolated natural polymers via Michael addition reaction under physiological conditions. Poly(ethylene glycol-b-caprolactone-b-ethylene glycol) (PECL) micelles bearing double bonds act as both fillers and chemical cross-linkers to mechanically reinforce chitosan-based hydrogels, which is confirmed by the results of rheological behavior and compressive strength measurements. Indomethacin (IMC) and/or basic fibroblast growth factor (bFGF) are/is entrapped into the PECL micelle cross-linked hydrogel network primarily through hydrophobic interaction and specific affinity to thiolated heparin, respectively. After a relatively quick initial release, rather than an initial burst commonly occurring in conventional hydrogels based on drug delivery systems, IMC and/or bFGF are/is released from these PECL micelle cross-linked hydrogels at a slower rate until a steady state is reached. The release rate of IMC and/or bFGF could be readily tuned by varying the micelle amount and the thiolated heparin content in a polymer matrix.

Alginate/PEG based microcarriers with cleavable crosslinkage for expansion and non-invasive harvest of human umbilical cord blood mesenchymal stem cells

Porous microcarriers are increasingly used to expand and harvest stem cells. Generally, the cells are harvested via proteolytic enzyme treatment, which always leads to damages to stem cells. To address this disadvantage, a series of alginate/PEG (AL/PEG) semi-interpenetrating network microcarriers are prepared in this study. In this AL/PEG system, the chemically cross-linked alginate networks are formed via the reaction between carboxylic acid group of alginate and di-terminated amine groups of cystamine. PEG is introduced to modulate the degradation of microcarriers, which does not participate in this cross-linked reaction, while it interpenetrates in alginate network via physical interactions. In addition, chitosan are coated on the surface of AL/PEG to improve the mechanical strength via the electrostatic interactions. Biocompatible fibronectin are also coated on these microcarriers to modulate the biological behaviors of cells seeded in microcarriers. Results suggest that the size of AL/PEG microcarriers can be modulated via adjusting the contents and molecular weight of PEG. Moreover, the microcarriers are designed to be degraded with cleavage of disulfide crosslinkage. By changing the type and concentration of reductant, the ratio of AL to PEG, and the magnitude of chitosan coating, the degradation ability of AL/PEG microcarriers can be well controlled. In addition, AL/PEG microcarriers can support the attachment and proliferation of human umbilical cord blood mesenchymal stem cells (hUCB-MSCs). More importantly, the expanded hUCB-MSCs can be detached from microcarriers after addition of reductant, which indeed reduce the cell damage caused by proteolytic enzyme treatment. Therefore, it is convinced that AL/PEG based microcarriers will be a promising candidate for large-scale expansion of hUCB-MSCs.

Chemically crosslinked alginate porous microcarriers modified with bioactive molecule for expansion of human hepatocellular carcinoma cells.

Microcarrier is an essential matrix for the large-scale culture of anchorage-dependent cells. In this study, chemical cross-linked alginate porous microcarriers (AMC) were prepared using microemulsion and freeze-drying technology. Moreover, chitosan was coated on the surface of microcarriers (AMC-CS) via electrostatic interactions to improve the mechanical strength. The size of AMC can be modulated through adjusting the concentration of alginate, amount of dispersant and stirring rate. The surface chemical characteristics and morphology of AMC-CS were evaluated by Fourier transformed infrared, X-ray photoelectron spectroscopy, and scanning electron microscope. Fibronectin (Fn) or heparin/basic fibroblast growth factor (bFGF) was then immobilized on the surface of microcarriers via layer-by-layer technology to improve the cytocompatibility. Our data suggested that the size of AMC can be accurately modulated from 90 μm to 900 μm with a narrow size distribution. Micropore structures of AMC-CS were relatively disordered and the pore size ranged between 20 μm and 100 μm. Using AMC after modified with Fn or bFGF as the cell expansion microcarriers, we showed that the proliferation rates of HepG2 cells increased significantly, reaching to more than 30-fold of cell expansion after 10 days of culture, with minor cellular damage caused by the microcarriers. Moreover, the AMC microcarriers modified with Fn or bFGF can increase albumin secretion of HepG2. We suggest that our new modified AMC-based microcarriers will be an attractive candidate for the large-scale cell culture of therapeutic cells.

Degradation in vitro and in vivo of β-TCP/MCPM-based premixed calcium phosphate cement

Premixed calcium phosphate cements (CPCs) have been developed to shorten the surgical time of conventional CPCs. However, there is lack of investigation on degradation behavior of premixed CPCs in vitro and in vivo. In this study, the premixed CPCs are prepared by mixing glycerol or polyethylene glycol (PEG) with the CPC power (β-tricalcium phosphate (β-TCP) and monocalcium phosphate monohydrate (MCPM)), and their degradation performances including the microstructure, chemical composition and mechanical properties are systematically evaluated both in vitro and in vivo (subcutaneous implantations in rabbits). When the premixed CPCs aged in PBS or FBS, results show weight loss of the specimens, decreased pH value and increased calcium ion concentration of aging media. Meanwhile, the setting products convert from dicalcium phosphate dihydrate (DCPD) to dicalcium phosphate anhydrous (DCPA), and no hydroxyapatite deposit. The specimen size and the molecular weight of non-aqueous solvent can modulate the setting product of premixed CPCs. For the larger specimens, DCPA is the main setting product, for the smaller ones, the composite contained DCPD and DCPA. With the decrease of the molecular weight of the non-aqueous solvent PEG, the setting product change from both DCPD and DCPA to DCPA due to the quicker exchange rate of PEG with water. After a period of subcutaneous implantation, the surface of the grafts obviously disintegrated with the formation of porous structures, but their internal morphology do not obviously change.