Department of Veterinary Clinical Sciences, College of Veterinary Medicine, Washington State University, Pullman, Washington 99164-6610, USA.
Pharmacogenet Genomics. 2011 Feb;21(2):66-75. doi: 10.1097/FPC.0b013e3283425f44.
Distribution of fluoroquinolones to the retina is normally restricted by ABCG2 at the blood-retinal barrier. As the cat develops a species-specific adverse reaction to photoreactive fluoroquinolones, our goal was to investigate ABCG2 as a candidate gene for fluoroquinolone-induced retinal degeneration and blindness in cats.
Feline ABCG2 was sequenced and the consensus amino acid sequence was compared with that of 10 other mammalian species. Expression of ABCG2 in feline retina was assessed by immunoblot. cDNA constructs for feline and human ABCG2 were constructed in a pcDNA3 expression vector and expressed in HEK-293 cells, and ABCG2 expression was analyzed by western blot and immunofluorescence. Mitoxantrone and BODIPY-prazosin efflux measured by flow cytometry and a phototoxicity assay were used to assess feline and human ABCG2 function.
Four feline-specific (compared with 10 other mammalian species) amino acid changes in conserved regions of ABCG2 were identified. Expression of ABCG2 on plasma membranes was confirmed in feline retina and in cells transfected with human and feline ABCG2, although some intracellular expression of feline ABCG2 was detected by immunofluorescence. Function of feline ABCG2, compared with human ABCG2, was found to be deficient as determined by flow cytometric measurement of mitoxantrone and BODIPY-prazosin efflux and enrofloxacin-induced phototoxicity assays.
Feline-specific amino acid changes in ABCG2 cause a functional defect of the transport protein in cats. This functional defect may be owing, in part, to defective cellular localization of feline ABCG2. Regardless, dysfunction of ABCG2 at the blood-retinal barrier likely results in accumulation of photoreactive fluoroquinolones in feline retina. Exposure of the retina to light would then generate reactive oxygen species that would cause the characteristic retinal degeneration and blindness documented in some cats receiving high doses of some fluoroquinolones. Pharmacological inhibition of ABCG2 in other species might result in retinal damage if fluoroquinolones are concurrently administered.
ABCG2 通常会限制血视网膜屏障对氟喹诺酮类药物的摄取。由于猫对光反应性氟喹诺酮类药物会产生一种特有的不良反应，我们的目标是研究 ABCG2 是否为猫氟喹诺酮类药物诱导的视网膜变性和失明的候选基因。
对猫的 ABCG2 进行测序，并将共识氨基酸序列与其他 10 种哺乳动物的序列进行比较。通过免疫印迹评估 ABCG2 在猫视网膜中的表达。构建猫和人 ABCG2 的 cDNA 构建体，并在 pcDNA3 表达载体中表达，通过 Western blot 和免疫荧光分析 ABCG2 的表达。通过流式细胞术和光毒性测定来评估米托蒽醌和 BODIPY-普萘洛尔的外排，以评估猫和人 ABCG2 的功能。
在 ABCG2 的保守区域发现了 4 个与其他 10 种哺乳动物物种相比具有独特性的氨基酸变化。在猫的视网膜和转染了人源和猫源 ABCG2 的细胞中均证实了 ABCG2 质膜表达，尽管通过免疫荧光检测到一些猫源 ABCG2 的细胞内表达。通过流式细胞术测量米托蒽醌和 BODIPY-普萘洛尔的外排以及恩诺沙星诱导的光毒性测定，发现与人类 ABCG2 相比，猫源 ABCG2 的功能存在缺陷。
ABCG2 中的猫特有的氨基酸变化导致了该转运蛋白在猫中的功能缺陷。这种功能缺陷可能部分归因于猫源 ABCG2 的细胞内定位缺陷。无论如何，血视网膜屏障处的 ABCG2 功能障碍可能导致光反应性氟喹诺酮类药物在猫视网膜中的积累。然后，暴露于光线下的视网膜会产生活性氧，从而导致一些接受高剂量某些氟喹诺酮类药物的猫出现的特征性视网膜变性和失明。如果同时给予氟喹诺酮类药物，其他物种的 ABCG2 药物抑制可能会导致视网膜损伤。
