Osteogenic differentiation of human mesenchymal stem cells on chargeable polymer-modified surfaces.
ABSTRACT Polystyrene cell-culture plates modified with positively charged polyallylamine (PAAm) and negatively charged poly(acrylic acid) (PAAc) and unmodified plate were used for the culture of human mesenchymal stem cells (MSCs) to study the effect of surface electrostatic properties on their osteogenic differentiation. All of these surfaces supported cell adhesion and proliferation. However, the cells adhered, spread, and proliferated somewhat more quickly on the PAAm-modified surface than they did on the PAAc-modified and control surfaces. Osteogenic differentiation was examined by alkaline phosphatase (ALP) staining, alizarin red S staining, and gene-expression analysis. The MSCs cultured on the three kinds of surfaces in the presence of dexamethasone were positively stained with ALP and alizarin red S staining, while the cells cultured without dexamethasone were not positively stained. Gene-expression analyses using real-time PCR indicated that MSCs cultured on these surfaces in the presence of dexamethasone expressed osteogenic marker genes, encoding ALP, osteocalcin, bone sialoprotein, osteopontin, and type I collagen. These results indicate that the positively charged, negatively charged, and unmodified surfaces supported osteogenic differentiation, and that their effect required the synergistic effect of dexamethasone.
Article: Morphological and transcriptomic comparison of adipose and bone marrow derived porcine stem cells[show abstract] [hide abstract]
ABSTRACT: In the present study we provided a morphological and transcriptomic comparison of adult porcine adipose-derived stem cells (ADSC) and bone marrow-derived stem cells (BMSC) as they differentiated in vitro towards the os-teogenic and adipogenic lineages for up to 4 weeks. The long term goal of this comparison is to assess the possibility of using ADSC as a potential alternative to BMSC as a source of autologous adult stem cells in human therapies. Our data indicated that ADSC can differentiate into osteocytes and adipocytes similar to BMSC but with some differ-ences. During the osteogenic differentiation both cell types went through morphological changes; however, while ADSC formed predominately osteogenic islands (nodules) in the culture dish, BMSC formed a continuous osteogenic sheet of small nodules. Transcriptomic analysis revealed that both cell types responded to the osteogenic induction. However, BGLAP mRNA expression did not increase in ADSC suggesting, together with the percentage area stained observed for Alizarin Red and von Kossa in ADSC, a lesser mineralization of bone matrix in this cell type compared to BMSC. During the adipogenic induction ADSC as well as BMSC were able to achieve the morphological and transcriptome changes characteristic of the adipogenic lineage. After 7 days of differentiation the expression patterns of DGAT2 and ADFP became greater in ADSC versus BMSC, which agreed with the larger lipid droplets formation observed in the ADSC by Oil Red O staining. Our findings represent an important step towards the assessment of using ADSC as an alternative to BMSC in therapeutic applications.The Open Tissue Engineering and Regenerative Medicine Journal 01/2009; 2:20-33.
Article: Tubular perfusion system for the long-term dynamic culture of human mesenchymal stem cells.[show abstract] [hide abstract]
ABSTRACT: In vitro culture techniques must be improved to increase the feasibility of cell-based tissue engineering strategies. To enhance nutrient transport we have developed a novel bioreactor, the tubular perfusion system (TPS), to culture human mesenchymal stem cells (hMSCs) in three-dimensional scaffolds. This system utilizes an elegant design to create a more effective environment for cell culture. In our design, hMSCs in the TPS bioreactor are encapsulated in alginate beads that are tightly packed in a tubular growth chamber. The medium is perfused by a peristaltic pump through the growth chamber and around the tightly packed scaffolds enhancing nutrient transfer while exposing the cells to shear stress. Results demonstrate that bioreactor culture supports early osteoblastic differentiation of hMSCs as shown by alkaline phosphatase gene expression. After 14 and 28 days of culture significant increases in the gene expression levels of osteocalcin, osteopontin, and bone morphogenetic protein-2 were observed with bioreactor culture, and expression of these markers was shown to increase with media flow rate. These results demonstrate the TPS bioreactor as an effective means to culture hMSCs and provide insight to the effect of long-term shear stresses on differentiating hMSCs.Tissue Engineering Part C Methods 10/2010; 17(3):337-48. · 4.64 Impact Factor
Article: Dependence of Spreading and Differentiation of Mesenchymal Stem Cells on Micropatterned Surface Area[show abstract] [hide abstract]
ABSTRACT: Micropatterning technology is a highly advantageous approach for directly assessing and comparing the effects of different factors on stem cell functions. In this study, poly(vinyl alcohol)-(PVA-) micropatterned polystyrene surfaces were prepared using photoreactive PVA and ultraviolet photolithography with a photomask. The micropatterned surface was suitable for single-cell array formation and long-term cell culture due to the nanometer thickness of nonadhesive PVA layer. Different degrees of cell spreading with the same cell shape were established by adjusting the sizes of circular, cell-adhesive polystyrene micropatterns. Cell spreading and differentiation of mesenchymal stem cells (MSCs) on the micropatterns were investigated at the single-cell level. The assembly and organization of the cytoskeleton were regulated by the degree of cell spreading. Individual MSCs on large circular micropatterns exhibited a more highly ordered arrangement of actin filaments than did those on the small circular micropatterns. Furthermore, the differentiation of MSCs was dependent on the degree of cell spreading. Increased cell spreading facilitated the osteogenic differentiation but suppressed the adipogenic differentiation of MSCs. This micropatterning method is valuable for stem cell research in tissue engineering and regenerative medicine.Hindawi Publishing Corporation Journal of Nanomaterials. 01/2011; 9.