Optimum Ho:YAG laser irradiation condition on percutaneous laser disk decompression (PLDD): in-vitro quantitive evaluation of influence on disk cells using three-dimensional cell culture system

摘要

The purpose of this study is to evaluate the influence on disc cells after laser irradiation using three-dimensional (3-D) culture system and to clarify the optimum Ho:YAG laser irradiation condition on percutaneous laser disc decompression (PLDD) therapy. Since the Ho:YAG laser ablation is characterized by water-vapor bubble dynamics with pressure wave, thermal effect on cell metabolism might occur in the intervertebral disc. We studied the disc cell damage on the metabolic point of view to investigate the optimum irradiation parameter of the Ho:YAG laser. We have developed the 3-D cultured disc cell system using agarose gel to investigate laser - disc cell interaction. This culture system provides a highly in vivo-like environment for disc cells in which cell- extracellular matrix interactions appear to be more important than contacts among cells. Intervertebral discs were obtained from Japanese white. The isolated disc cells were seeded in 96-well culture plates at the cell densities of 1 $MUL 10$+6$/ cells/ml, and incubated for 12 days. A pulsed Ho:YAG laser was delivered through a 200 micrometer-core diameter single silica glass fiber. On the agarose gel including the 3-D cultured disc cells, we used the Ho:YAG laser irradiation energy ranging from 40 to 180 mJ/pulse at the fiber end. Cytotoxicity and matrix synthesis after the laser irradiations were evaluated in time course to determine the optimum condition of laser irradiations. It was confirmed that laser irradiation causes necrosis of the cells and additionally produces apoptosis depending on the condition. The ability of matrix synthesis was maintained even after the irradiation, which differed depending on the irradiation conditions. The optimum irradiation conditions seemed related to the preservation of intact area and the acceleration of matrix synthesis in reactive area.