Continuous Stressing of Mouse Interparietal Suture Fibroblasts in vitro

Abstract

The morphological and biochemical response of sutural fibroblasts in vitro to continuous force was examined. Cells from mouse interparietal sutures were grown and subcultured on glass slides. Titanium disks coated with collagen were allowed to attach to the cellular multilayers. Four of the glass slides were then placed at an angle of 75° for a period of three days so that continuous stress would be created, while four others were left flat. Also, four glass slides were left flat with no disk. Following the incubation period, the dishes were labeled with ¹⁴C-glycine for 15 h. The cells and medium were then collected for collagen extraction followed by SDS-polyacrylamide gel electrophoresis. Dried gels impregnated with fluor were exposed to x-ray films that were then scanned densitometrically for collagen types I and III. It was found that the proportion of newly-synthesized type III collagen increased significantly with the application of continuous stress. A second set of experimental and control glass slides was fixed in glutaraldehyde and post-fixed in osmium tetroxide. Following critical-point drying and coating, the glass slides were examined under a scanning electron microscope. The scanning images showed the formation of a ligament-like structure between the disk and the glass slide. Moreover, mitotic activity, as evidenced by spheroidal cells, was stimulated in the areas previously adjacent to the disc, which had since moved away. This system offers a standardized continuous force system that can stress cells in a ligament-like structure and thus provides an in vitro model analogous to clinical orthodontic and orthopedic stress.

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