Narrow Results

Chondrocytes in the growth plate undergo a relatively linear differentiation process. The progression of a chondrocyte from the proliferative stage to the hypertrophic stage is governed by complex interactions with the extracellular matrix within which it resides. A network of peptides, ion channels, and second messengers affects the transcription of certain genes that are ultimately translated into peptides which control cellular activity. Much effort has been invested into replicating this environment under in vitro conditions. It has been found that the three-dimensional (3D) cell culture is a more accurate representation of the in vivo environment in comparison to the traditional monolayer culture. It has also been found that a variety of stimuli may be used to induce the proliferation and differentiation of chondrocytes; one such stimulus is the mechanical stimulation of chondrocytes embedded in a 3D Gelfoam sponge. Chondrocytes are obtained from the chicken sternum. After the cells are cultured and cyclically loaded, mRNA levels of various mechanosensitive genes are quantified by real-time reverse transcription-polymerase chain reaction (RT-PCR). Mechanical stimulation has been shown to upregulate the expression of type X collagen mRNA in early hypertrophic chondrocytes. The entire process, beginning with the obtainment of chondrocytes and ending with the quantification and interpretation of gene expression, is detailed in the following chapter.

Effects of porous hydroxyapatite granules (HAGs) on osteogenesis were examined in vitro using cells from adult rabbit femurs. Cultures in the space of 30-600 μm between glass plates with two-dimensionally arranged HAGs on the horizontal bottom glass surface induced thin insular tissue formation within 2 weeks. It was induced earlier and more than that without HAGs. Von Kossa staining-positive tissues were found at 1-2 weeks after their formation, suggesting their mineralization. No cells or several cells in monolayer were included within some of them. Several grew quadrate tissues. Similar thin tissues were formed in inclined and rotated glass tubes with three-dimensionally arranged HAGs. They were almost irregular in shape. By contrast, little extracellular matrix deposition and no mineralization were seen in the spaces over 600 μm and under 30 μm between the glasses with HAGs. These results suggest that osteoblastic cells from adult femurs form mineralized quadrate tissues with or without embedded cells on the horizontal glass in the space of 30-600 μm, while mechanical loading without system induces irregular-shaped tissue formation. Porous HAGs in the space seem to accelerate extracellular matrix accumulation and its mineralization.