He-Ne激光照射对成纤维细胞内[Ca^(2+)]_i浓度影响的流式细胞计测定

Effect on Intracellular Calcium Ion of the Fibroblast by the Heliumneon Laser Erradiated with Flow Cytometric Measurement

目的：ＨｅＮｅ激光照射治疗的机理不明，激光照射引起细胞内Ｃａ２＋水平变化，为治疗机理提供理论依据。方法：ＨｅＮｅ激光照射引起鼠成纤维细胞Ｌ９２９内［Ｃａ２＋］ｉ的变化，用ＨＯ３４２对细胞ＤＮＡ活性染色，Ｆｌｕｏ３ＡＭ对细胞内Ｃａ２＋染色，利用ＦＣＭ同时定量分析细胞ＤＮＡ和细胞内Ｃａ２＋的变化。结果：激光照射１５ｍｉｎ（光剂量１１．８１Ｊ／ｃｍ２后，ＦＣＭ分析可见ＤＮＡ分布直方图右移，表明成纤维细胞进入周期加快，二维点阵图设窗分析，分为低Ｃａ２＋区和高Ｃａ２＋区，与对照组相比，成纤维细胞在周期进程中，在低Ｃａ２＋区，Ｇ１期细胞变化不大，Ｓ期和Ｇ２＋Ｍ期细胞内［Ｃａ２＋］ｉ水平增加，但是在高Ｃａ２＋区，细胞的Ｇ１、Ｓ和Ｇ２＋Ｍ期细胞内［Ｃａ２＋］ｉ均明显增加。结论：许多研究者用化学因子和药物刺激，使细胞随着周期进程，细胞内［Ｃａ２＋］ｉ明显增加，出现复杂变化。本实验结果表明ＨｅＮｅ激光照射成纤维细胞，引起细胞周期进程中细胞内［Ｃａ２＋］ｉ出现明显变化。

Objective: The therapy mechanism has not been known with the HeNe laser irradiation. A change of the intracellular Ca 2+ levels was induced by laser irradiation. In order to offer in accordance with a theory is provided the therapy mechanism. Methods: The change of the intracellular Ca 2+ of the mouce fibroblast cell line L929 was induced by the HeNe lasser irradiation . The cells were stained with both of the Hoechst 33342(a supravitral stain for nuclear DNA) and the Fluo3 AM (a calcium sensitive dye) and analyzed by flow cytometer simultaneously. Results: The DNA distribution histogram by the laser irradiated 15 min (light dose of 11.81 J/cm 2) was found to the right shift, in which indicated to speed into different phases of the cell cycle of the mice fibroblasts. Regions R2R4 and R5R7 in represent the electronic gates used respetively for analysis of the low calcium and the high calcium distribution. In comparsion with the control, the intracellular Ca 2+ levels during progression of the cell cycle of the fibroblasts had no changes at G1 phase in the low calcium region, in which the intracellular Ca 2+ levels increased at S phase and G2+M phase. But the intracellular Ca 2+ levels increased obviously at G1, S and G2+M phases in the high calcium regions. Conclusion: The reseachers by the chemicals and the drugs stimulared the cells, the intracellular Ca 2+ rose steadily and appeared complexly during the cell cycle progression. Based upon these results we propose the changes in the intracellular Ca 2+ of the fibroblasts during the progression of the cell cycle by the HeNe laser irradiation.