[1]张跃红,饶志波,刘娟,等. 低浓度β-淀粉样蛋白抑制小鼠视网膜色素上皮细胞P-糖蛋白的功能性表达[J].眼科新进展,2015,35(9):801-805.[doi:10.13389/j.cnki.rao.2015.0220]
　ZHANG Yue-Hong,RAO Zhi-Bo,LIU Juan,et al. Low concentration of amyloid β-protein inhibiting functional expression of P-glycoprotein in mouse retinal pigment epithelial cells[J].Recent Advances in Ophthalmology,2015,35(9):801-805.[doi:10.13389/j.cnki.rao.2015.0220]
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 低浓度β-淀粉样蛋白抑制小鼠视网膜色素上皮细胞P-糖蛋白的功能性表达
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《眼科新进展》[ISSN:1003-5141/CN:41-1105/R]

卷:
35卷
期数:
2015年9期
页码:
801-805
栏目:
实验研究
出版日期:
2015-09-05

文章信息/Info

Title:
 Low concentration of amyloid β-protein inhibiting functional expression of P-glycoprotein in mouse retinal pigment epithelial cells
作者:
 张跃红; 饶志波; 刘娟; 唐浩英; 卢敏
 510000　广东省广州市，广州医科大学附属广州市第一人民医院眼科
Author(s):
 ZHANG Yue-Hong ; RAO Zhi-Bo ;  LIU Juan ; TANG Hao-Ying ;  LU Min
 Department of Ophthalmology, the First People ’ s Hospital of Guangzhou ,Affiliated Guangzhou Medical College, Guangzhou 510000 , Guangdong Province . China
关键词:
 ; β淀粉样蛋白; P-糖蛋白; 视网膜色素上皮; 年龄相关性黄斑变性
Keywords:
 ; amyloid β-protein;  P-glycoprotein;  retinal pigment epithelium ;  age-related macular degeneration
DOI:
10.13389/j.cnki.rao.2015.0220
文献标志码:
A
摘要:
 目的　探讨低浓度β淀粉样蛋白（ａｍｙｌｏｉｄβ-ｐｒｏｔｅｉｎ，Ａβ）对原代培养小鼠视网膜色素上皮（ｒｅｔｉｎａｌｐｉｇｍｅｎｔｅｐｉｔｈｅｌｉｕｍ，ＲＰＥ）细胞Ｐ-糖蛋白（Ｐ-ｇｌｙｃｏｐｒｏｔｅｉｎ，Ｐ-ｇｐ）表达和活性的影响。方法　应用０．３μｍｏｌ?Ｌ－１Ａβ１-４２处理ＲＰＥ细胞４ｈ、８ｈ、１２ｈ和２４ｈ后，采用实时定量ｒｅａｌ-ｔｉｍｅＰＣＲ和Ｗｅｓｔｅｒｎｂｌｏｔｔｉｎｇ检测Ｐ-ｇｐ在基因和蛋白水平的表达；分别在细胞培养液中加入孕烯醇酮Ｘ受体原形配体１６-α-氰基孕烯诺龙（ｐｒｅｇｎｅｎｏｌｏｎｅ１６-ａｌｐｈａ-ｃａｒ-ｂｏｎｉｔｒｉｌｅ，ＰＣＮ）和Ｐ-ｇｐ特异性抑制剂ＰＳＣ８３３后，孵育罗丹明１２３，然后用流式细胞仪检测各组细胞内罗丹明１２３的荧光强度来验证Ａβ１-４２对ＲＰＥ细胞Ｐ-ｇｐ活性的影响。结果　Ａβ１-４２处理细胞４ｈ、８ｈ、１２ｈ和２４ｈ后ｍｄｒ１表达量分别减至空白对照组的（７１．２５±８．３５）％（Ｐ＞０．０５）、（６１．３８±７．３７）％（Ｐ＜０．０５）、（４７．５１±８．９７）％（Ｐ＜０．０１）和（４１．２３±９．０６）％（Ｐ＜０．０１）；Ｐ-ｇｐ表达量在４个时间点分别减至空白对照组的（８１．７１±５．２７）％（Ｐ＞０．０５）、（５３．６４±４．５８）％（Ｐ＜０．０５）、（５１．２９±６．０２）％（Ｐ＜０．０５）和（４９．８２±７．２４）％（Ｐ＜０．０５）。罗丹明１２３积蓄实验结果显示，应用ＰＣＮ后，细胞内罗丹明１２３的荧光强度随着时间延长而减少，而应用ＰＳＣ８３３后，细胞内罗丹明１２３的荧光强度随着时间延长而增加。结论　低浓度Ａβ１-４２处理小鼠ＲＰＥ细胞，可抑制Ｐ-ｇｐ的表达与活性。Ｐ-ｇｐ可能是干预早期年龄相关性黄斑变性中Ａβ清除的潜在靶点。
Abstract:
 Objective To investigate the effects of low concentration of amyloid β-protein (Aβ) in primary cultured mouse retinal pigment epithelium ( RPE) cells on the expression and activity of P-glycoprotein ( P-gp) . Methods Cells were treated by 0. 3 Vmol . L-l Aβ1-42 for 4 hours ,8 hours.12 hours and 24 hours. The expressions of P-gp in the transcription and translation level were measured by relative quantification real-time polymerase chain reaction and Western blotting. Following the incubation of the original ligand of pregnant X receptor pregnenolone 16-alpha-carbonitrile ( PCN ) and P-gp specific inhibitor PSC833 .rhodamine 123 was added to the medium. The effects of Aβ1-42 0n P-gp activity in RPE cells was verified through the fluorescence intensity of intracellular thodanune 123 by flow cytometery. Results Compared with the control group ,mdrl levels at 4 hours .8 hours .12 hours and 24 hours after treatment with Aβ1-42 were reduced to (71. 25 + 8. 35 ) % (P > 0. 05 ) . (61. 38 + 7. 37 )% (P < 0. 05 ) . ( 47. 51 + 8. 97 ) % (P < 0. 01 ) and ( 41. 23 + 9. 06 ) % ( P < 0. 01 ) . respectively , while P-gp levels at four time points were reduced to ( 81. 71 + 5. 27 ) % (P > 0. 05 ) . ( 53. 64 + 4. 58 ) % (P < 0. 05 ) , (51. 29 +6. 02) % (P < 0. 05 ) and ( 49. 82 + 7. 24 ) % (P < 0. 05 ) , respectively. The results of flow cytometery showed that the application of PCN led to a decrease of fluorescence intensity of intracellular thodamine 123 with prolonged time ,while the fluorescence intensity of intracellular thodamine 123 increased with the prolonged time after incubating with PSC833. Conclusion Aβ1-42 can inhibit the expression and activity of P-gp in cultured primary mouse RPE cells. P-gp in outer blood retinal barrier might be a potential target in the intervention of Aβ clearance in the early age-related macular degeneration.

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备注/Memo

备注/Memo:
 国家自然科学基金资助（编号：８１２００７０９）
更新日期/Last Update: 2015-08-31