Purpose
The purpose of this study is to track chondrocytic cells interaction with notochordal cells by an in vitro biologic assays which measure chondrocytic cells activities using robotic time-lapsed microscopic imaging system.
Overview of Literature: Although chondrocytic cells and notochordal cells play critical roles in homeostasis of the intervertebral disc, their real-time interaction has not been studied.
Materials and Methods
Chondrocytic cells and notochordal cells purified from rabbit nucleus pulposus were isolated after serial filtration. Both cells were plated in 24 well-culture plates and then were co-cultured for timelapsed cell tracking study. After numerous locations of each well were selected for time-lapsed analysis, both cells were tracked by visible imaging every 10 minutes over 5 days. Biochemical information were done to compare matrix protein expression by gene expression (collagen II and aggrecan) using reverse transcription polymerase chain reaction (RT-PCR).
Results
In time-lapsed cell tracking analysis, mobility of chondrocytic cells were shown significantly faster than vacuolated notochordal cells although both cells were similar in cytoplasmic area. Chondrocytic cells cocultured with notochordal cells frequently interacted with neighbor vacuolated notochordal cells and increased mitotic fraction and velocity, and decreased population doubling time (PDT) compared with chondrocyte-like cells alone, which are consistent with results of collagen II expression. Aggrecan expression levels did not show any significance.
Conclusion
These results are the first direct evidences of chondrocytic cells activities with notochordal cells in vitro 2-D culture using both individual cell tracking and cell population tracking in real-time with high-throughput experimental capability. This novel system provides ability to quantify numerous and unique temporal changes in chondrotic cells and notochordal cells, and essential information to indentify supporting function of notochordal cells in intervertebral disc, suggesting that this system has the potential to use in disc homeostasis research.
