摘要
目的研究水溶性氮酮对体外培养的兔角膜内皮细胞活性与超微结构的影响,确定水溶性氮酮应用于眼部的安全浓度。方法采用消化法培养兔角膜内皮细胞,取第3或第4代细胞以1×104个.L-1的浓度接种,共分8组,其中一组设为对照组,不给予水溶性氮酮,其他7组细胞中水溶性氮酮浓度分别为0.001,0.005,0.010,0.050,0.100,0.500,1.000 g.L-1,24 h后采用MTT法测定细胞活性。另取上述培养的细胞,按1×104个.L-1的浓度接种到载玻片上,分别加入浓度为0.010,0.100,0.500 g.L-1的水溶性氮酮作用10 m in,观察细胞膜超微结构变化。另采用相同浓度的水溶性氮酮作用于体外培养的兔角膜内皮细胞10 m in,观察细胞膜超微结构的变化。结果与对照组比较,浓度≤0.100 g.L-1的水溶性氮酮作用24 h后对兔角膜内皮细胞的活性没有明显影响,浓度≥0.500 g.L-1的水溶性氮酮作用24 h可以引起兔角膜内皮细胞活性下降(P<0.01);经浓度≤0.100 g.L-1水溶性氮酮作用10 m in,兔角膜内皮细胞细胞膜有裂隙形成,但细胞器等结构无明显损害,浓度≥0.500 g.L-1水溶性氮酮可导致兔角膜内皮细胞凋亡。结论浓度<0.100 g.L-1水溶性氮酮可增加角膜内皮细胞对药物的通透性,并有望用于角膜内皮细胞基因转染研究。
Objective To study the effects of water soluble azone on the viability and uhrastructure of corneal endothelial cells of the rabbit cultured in vitro in order to determine the safe concentration of the drug in its clinical application to ophthalmologic patients. Methods Corneal endothelial cells of the rabbit were cultured in vitro with the digestion method. Cells of the 3^rd or 4^th generation were inoculated to 96-well plates with a concentration of 1 ×10^4cells · L^-1. The cells were then divided into 8 groups one of which serving as control with no addition of water soluble azone. To the remaining 7 groups of cells water soluble azone was added to bring about the drug concentrations of 0. 001,0. 005,0. 010,0.050,0. 100,0.500 and 1. 000 g · L^-1 ,respectively. Viability of the cells were determined with the MTY method 24 b later. Cells of the 3^rd or the 4^th generation in a concentration of 1× 10^4 cells· L^-1 described above were pipetted onto sterile slides. Water soluble azone in concentration of 0.010,0. 100 and 0.500 g· L^-1 was added to these cells, acting for 10 min. The ultrastructure of the cells was then examined under the scanning electron microscope. Besides, rabbit corneal endothelial cells in primary culture were subjected to the action of water soluble azone in the same concentrations described above for 10 min followed by electron microscopic examination as well. Results The viability of cultured rabbit corneal endothelial cells in their 3^rd or 4^th generation was not significantly affected by water soluble azone in concentrations ≤0. 100 g· L^-1 acting for 24 h. The drug in concentrations ≥0. 500 g · L^-1, however, was shown to cause a reduction of the cell viability after acting for 24 h（P〈0.01 ）. The 10 min action of water soluble azone in concentrations≤0.100 g · L^-1 was shown to result in slit formations on the memborane of the rabbit corneal endothelial cells but no significant changes in the organelles as shown by electron microscopy. Water soluble azone in concentrations≥0.500 g·L^-1 , however, led to apoptosis of most of the cultured rabbit corneal endothelial cells. Conclusion Water soluable azone at concentrations no higher than 0.100 g·L^-1 was shown to have no harmful effects on the viability and ultrastmcture of cultured rabbit corneal endothelial cells and may increase the permeability of the cell membrane to the drug. Water soluble azone may also be used in the research work of zene transfection of corneal endothelial cells.
出处
《医药导报》
CAS
2006年第8期732-734,共3页
Herald of Medicine
基金
湖北省自然科学基金资助(基金编号:2003ABA150)
关键词
氮酮
水溶性
角膜内皮细胞
兔
超微结构
Azone, water soluble
Corneal endothelial cell
Rabbit
Ultrastructure
