Abstract
Photo-reactive poly(vinyl alcohol) (PRPVA) was synthesized by introduction of phenyl azido groups into poly(vinyl alcohol) (PVA) and applied for surface modification. PRPVA was grafted onto cell culture plate surface homogeneously or in a micropattern. Human mesenchymal stem cells (hMSCs) cultured on cell culture plate surface and PVA-modified surface showed different behaviors. Cells adhered and spread well on cell culture plate surface, while they did not adhere on PVA-grafted surface at all. When hMSCs were cultured on PVA-micropatterned surface, they formed a cell micropattern. Cells formed pellets after cultured on PVA homogeneously modified surface in chondrogenic induction medium for 2 weeks. The pellets were positively stained by hematoxylin/eosin, safranin-O/fast green and toluidin blue, and they were also stained brown by Type II collagen and proteoglycan immunohistological staining. Real-time PCR analysis was conducted to investigate the expression of colI, colII, colX, aggrecan and sox9 mRNA. Results of gene expression were in agreement with those of histological and immunohistological observations. These results indicated that hMSCs cultured on PVA-modified surface performed chondrogenic differentiation, and it was possible to construct scaffold-free cartilage like pellets with PVA-modified surface in vitro.
Highlights
Mesenchymal stem cells (MSCs) are a unique class of multipotent cells; they can be extracted from bone marrow, adipose tissue, umbilical cord tissue and so on
The introduction of azidophenyl group into the side chain of poly(vinyl alcohol) (PVA) was confirmed by 1H-NMR spectrum, as shown in Fig. 1, the azidophenyl proton appeared in the 1H-NMR spectrum at 7.07 and 7.93 ppm, methine and methene protons appeared at 3.9 ppm and 1.5 ppm, respectively
Cells in pellets formed on cell culture plate surface in the same culture condition did not show positive staining for them (Fig. 4h, k, i and l). These results demonstrated that PVA-modified surface was beneficial for Human mesenchymal stem cells (hMSCs) to perform chondrogenic differentiation, and the extent of chondrogenic differentiation for hMSCs was higher compared with that on cell culture plate surface
Summary
Mesenchymal stem cells (MSCs) are a unique class of multipotent cells; they can be extracted from bone marrow, adipose tissue, umbilical cord tissue and so on. Most of studies indicate that high cell density improves chondrogenic differentiation of MSCs [6, 7], so that the micromass or pellet culture with chondrogenic induction supplement is often chosen to confirm the potential of stem cells to differentiate into chondrocytes. Polyallylamine (PAAm), poly(ethylene glycol) (PEG) and poly(acrylic acid) were selected to modify cell culture plate surface and their effects on the chondrogenic differentiation of hMSCs were investigated. Taken what has been discussed above into consideration, and combining excellent film-forming ability of PVA with the property to support chondrogenic differentiation of PEG-modified surface, in this study, another kind of neutral photo-reactive polymer, photoreactive PVA (PRPVA), was designed and fabricated to modify cell culture plate surface; its effect on the adhesion and chondrogenic differentiation of hMSCs was investigated; scaffoldfree cartilage like pellets was constructed on PVA-modified surface
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