蓝光照射人视网膜色素上皮细胞后通过增加细胞内钙离子浓度促进凋亡的机制
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摘要
目的观察蓝光对人视网膜色素上皮细胞凋亡的影响并探讨其可能机制。方法细胞株分为蓝光照射组和对照组。蓝光照射组,利用35 W白色冷光灯加用蓝色滤光片建立蓝光损伤体外培养的RPE细胞模型,蓝光控制波长范围470~520 nm,光照强度在2000 lx左右,光照时间24~96 h,实验过程中保证光照在细胞培养孵箱内密进行。对照组为常规避光孵箱培养细胞,除无光线照射外,各培养条件相同。利用流式细胞仪检测RPE细胞的凋亡变化,利用流式细胞仪和激光共聚焦显微镜检测RPE细胞内游离钙离子浓度。结果与对照组比较,蓝光照射组的RPE细胞凋亡增加(P<0.05);照射组的RPE细胞内游离钙离子浓度增加(P<0.05);通过特异性抑制L型电压依赖性钙离子通道功能降低RPE细胞内游离钙离子浓度,在一定程度上降低了蓝光引起的RPE细胞凋亡。结论蓝光可以通过增加RPE细胞内游离钙离子浓度促进RPE细胞的凋亡,L型电压依赖性钙离子通道有望成为治疗年龄相关性黄斑变性等视网膜变性疾病的新靶点。
Objective To investigate the influence and mechanism of blue light on the apoptosis of human retinal pigment epithelial cells. Methods Cells were divided into blue light group and control group. 35 W white light lamp with blue filter was used to establish damaged RPE cell model in vitro, blue ray wave length ranged 470-520 nm, and the light intensity was about 2000 lx, light exposure time was set to 24-96 h, in the experimental process, the light was carried out in the cell culture incubation box. The control group cells were cultured in the conventional light avoidance incubator, and the culture conditions were as the same as the blue light group except without light. After exposure to blue light, the apoptosis of human retinal pigment epithelial cells was tested by flow cytometry. And then the intracellular calcium concentration of human retinal pigment epithelial cells was measured by flow cytometry and laser scanning confocal microscope. Results Compared with control group, the apoptosis of human retinal pigment epithelial cells increased in blue light group(P < 0.05), and the intracellular calcium concentration of human retinal pigment epithelial cells increased(P < 0.05). The intracellular calcium concentration of human RPE cells decreased by specific inhibition of L-type voltage-dependent calcium channel, RPE cell apoptosis caused by blue light decreased. Conclusion Blue light promotes apoptosis of human retinal pigment epithelial cells by the up-regulation of intracellular calcium concentration.
Objective To investigate the influence and mechanism of blue light on the apoptosis of human retinal pigment epithelial cells. Methods Cells were divided into blue light group and control group. 35 W white light lamp with blue filter was used to establish damaged RPE cell model in vitro, blue ray wave length ranged 470-520 nm, and the light intensity was about 2000 lx, light exposure time was set to 24-96 h, in the experimental process, the light was carried out in the cell culture incubation box. The control group cells were cultured in the conventional light avoidance incubator, and the culture conditions were as the same as the blue light group except without light. After exposure to blue light, the apoptosis of human retinal pigment epithelial cells was tested by flow cytometry. And then the intracellular calcium concentration of human retinal pigment epithelial cells was measured by flow cytometry and laser scanning confocal microscope. Results Compared with control group, the apoptosis of human retinal pigment epithelial cells increased in blue light group(P < 0.05), and the intracellular calcium concentration of human retinal pigment epithelial cells increased(P < 0.05). The intracellular calcium concentration of human RPE cells decreased by specific inhibition of L-type voltage-dependent calcium channel, RPE cell apoptosis caused by blue light decreased. Conclusion Blue light promotes apoptosis of human retinal pigment epithelial cells by the up-regulation of intracellular calcium concentration.
引文
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[18]黄汉辉,郑师明,荆冬冬,等替米沙坦对血管紧张素Ⅱ所致血管内皮细胞游离钙离子的抑制作用[J].中国医药导报,2015,12(3):11-14.
[19]孙中洋,李东韬,张舒.成骨细胞钾、钙离子通道的研究进展[J].解放军医学院学报,2014,35(2):186-189.
[20]黄鹤飞,陈颖,蔡维艳,等.芍药苷对大鼠背根神经无细胞内Ca2+的影响[J].中国实验动物学报,2016,24(1):25-30.
[21]唐晓琼,邓华聪,万青,等.2型糖尿病患者细胞内钙离子浓度变化的研究[J].重庆医科大学学报,2002,27(2):133-135.
[2]梁山,梁云,申国彦.维生素A缺乏易致老年黄斑变性的实验研究[J].中国医药导报,2008,5(24):32-33.
[3]Yam JC,Kwok AK.Ultraviolet light and ocular diseases[J].Int Ophthalmol,2014,34(2):383-400.
[4]蔡莉,易敬林.视网膜色素上皮细胞损伤机制研究进展[J].眼科新进展,2012,32(9):898-900.
[5]蔡善君,严密,张军军.蓝光诱导体外培养的人视网膜色素上皮细胞凋亡[J].中华眼底病杂志,2005,21(6):384-387.
[6]Bhosale G,Sharpe JA,Sundier SY,et al.Calcium signaling as a mediator of cell energy demand and a trigger to cell death[J].Ann NY Acad Sci,2015,1350:107-116.
[7]Ayaz O,Howlett SE.Testosterone modulates cardiac contraction and calcium homeostasis:cellular and molecular mechanisms[J].Biol Sex Differ,2015,6:9.
[8]李上海,梁伟钧,王苗.辛伐他汀对缺血再灌注损伤下内皮祖细胞凋亡的影响[J].中国医药导报,2015,12(19):11-14.
[9]武晓,刘凤娟,王鹏飞,等.巴豆生物碱对肺腺癌细胞凋亡的影响及机制研究[J].中国医药导报,2016,13(1):17-20.
[10]Sun Z,Cao X,Zhang Z,et al.Simulated microgravity inhibits L-type calcium channel currents partially by the up-regulation of mi R-103 in MC3T3-E1 osteoblasts[J].Scientific Reports,2015,5:8077.
[11]Zarbin M.Cell-Based Therapy for Degenerative Retinal Disease[J].Trends Mol Med,2016,22(2):115-134.
[12]Han S,Lu Q,Wang N.Apr3 accelerates the senescence of human retinal pigment epithelial cells[J].Mol Med Rep,2016,13(4):3121-3126.
[13]赵小祺,王春光,王小荣,等.缺氧诱发细胞凋亡的机制[J].中国老年学杂志,2010,30(7):1012-1014.
[14]齐娜,廖迎,张贵林.心肌缺血再灌注损伤及药物治疗研究进展[J].华夏医学,2007,20(1):170-172.
[15]Kistamas K,Szentandrassy N,Hegyi B,et al.Changes in intracellular calcium concentration influence beat-tobeat variability of action potential duration in canine ventricular myocytes[J].J Physiol Pharmacol,2015,66(1):73-81.
[16]Dong XJ,Luo XD,Xiong L,et al.Effects of energy controllable steep pulses on intracellular calcium concentration and cell membrane potential[J].Eur Rev Med Pharmacol Sci,2014,18(5):680-688.
[17]王健,庞军涛.不同剂量辛伐他汀对慢性心房重构犬心房肌细胞内游离钙离子浓度的影响[J].中国医药导报,2014,11(3):36-38.
[18]黄汉辉,郑师明,荆冬冬,等替米沙坦对血管紧张素Ⅱ所致血管内皮细胞游离钙离子的抑制作用[J].中国医药导报,2015,12(3):11-14.
[19]孙中洋,李东韬,张舒.成骨细胞钾、钙离子通道的研究进展[J].解放军医学院学报,2014,35(2):186-189.
[20]黄鹤飞,陈颖,蔡维艳,等.芍药苷对大鼠背根神经无细胞内Ca2+的影响[J].中国实验动物学报,2016,24(1):25-30.
[21]唐晓琼,邓华聪,万青,等.2型糖尿病患者细胞内钙离子浓度变化的研究[J].重庆医科大学学报,2002,27(2):133-135.