DIDEOXYNUCLEOSIDE TRANSPORT IN THE BLOOD/BRAIN BARRIER

人类免疫缺陷病毒（HIV）感染在大脑中产生 严重的神经系统并发症 据估计，至少有30%的 所有艾滋病成人和50%的艾滋病儿童将遭受 脑内与HIV感染有关的神经系统疾病。 与艾滋病有关的神经系统并发症的治疗是困难的， 通常用于治疗艾滋病的抗病毒双脱氧核苷不 容易穿过血脑屏障（BBB）。 携带者的存在 3 '-叠氮基-2'，3 '-双脱氧胸苷（AZT）及相关 血脑脊髓液屏障中的双脱氧核苷已经被 有据可查 然而，很少有研究对 脑微血管内存在双脱氧核苷载体 形成血脑屏障的内皮细胞。 在本提案中，承运人 系统介导的运输双脱氧核苷进出 检测脑微血管内皮细胞。 工作 所提出的研究假设是， 系统存在于脑微血管内皮细胞的质膜上 控制双脱氧核苷摄取和流出的细胞。 将鉴定和表征血脑屏障上的双脱氧核苷载体 使用原代培养的牛脑微血管内皮细胞（BBMEC）。 为了检查摄取载体，一系列放射性标记的 双脱氧核苷（AZT，双脱氧肌苷，双脱氧胞嘧啶，和 将检查BBMEC单层中的双脱氧胸苷），以确定：1） 每种化合物的摄取速率，2）摄取的饱和度， 核苷与载体相互作用所需的结构特征， 3)吸收的能量依赖性和4）载体的本地化 内腔（血液侧）和非内腔（脑侧）质膜。 相同系列放射性标记的双脱氧核苷流出BBMEC 将检查单层以确定：1） 双脱氧核苷外排至已知的核苷载体抑制剂，2） 双脱氧核苷相互作用所需的重要结构特征 与外排载体，和3）的位置和分布， BBMEC腔侧和非腔侧的双脱氧核苷载体 单层。 这些研究将严格审查细胞机制 控制双脱氧核苷通过血脑屏障。 通过 根据其特异性表征摄取和外排载体 抑制剂及其能量需求，可以进行比较， 其他细胞中的核苷转运系统。 中获得的信息 这些研究也可用于改善现有的 治疗神经系统疾病的未来抗病毒核苷 与艾滋病有关。

Human immunodeficiency virus (HIV) infections within the brain produce serious neurological complications. It is estimated that at least 30% of all adults with AIDS and 50% of all children with AIDS will suffer central nervous system disorders related to HIV infection within the brain. Treatment of the neurological complications related to AIDS is difficult as anti-viral dideoxynucleosides commonly used in the treatment of AIDS do not readily cross the blood-brain barrier (BBB). The presence of carrier systems for 3'-azido-2',3'-dideoxythymidine (AZT) and related dideoxynucleosides in the blood-cerebral spinal fluid barrier have been well documented. However, few studies have critically evaluated the presence of dideoxynucleoside carriers within the brain microvessel endothelial cells that form the BBB. In the present proposal, carrier systems mediating the transport of dideoxynucleosides both into and out of brain microvessel endothelial cells will be examines. The working hypothesis for the proposed studies are that specific, distinct carrier systems exist on the plasma membrane of the brain microvessel endothelial cells that control the uptake and efflux of dideoxynucleosides. Dideoxynucleoside carriers on the BBB will be identified and characterized using primary cultured bovine brain microvessel endothelial cells (BBMEC). To examine uptake carriers, the accumulation of a series of radiolabeled dideoxynucleosides (AZT, dideoxyinosine, dideoxycytosine, and dideoxythymidine) in BBMEC monolayers will be examined to determine: 1) the uptake rates for each compound, 2) the saturability of uptake and structural features required for nucleoside interactions with the carrier, 3) the energy-dependency of uptake and 4) the localization of carriers on the lumenal (blood side) and ablumenal (brain side) plasma membrane. Efflux of the same series of radiolabeled dideoxynucleosides out of BBMEC monolayers will be examined to determine: 1) the susceptibility of dideoxynucleoside efflux to known nucleoside carrier inhibitors, 2) important structural features required for dideoxynucleoside interactions with the efflux carrier, and 3) the location and distribution of dideoxynucleoside carriers on the lumenal and ablumenal side of BBMEC monolayers. These studies will critically examine the cellular mechanisms controlling the passage of dideoxynucleosides across the BBB. By characterizing uptake and efflux carriers with regard to their specificity of inhibitors and their energy requirements, comparisons can be made to nucleoside transport systems in other cells. The information obtained in these studies can also be used to improve the efficacy of both current and future anti-viral nucleosides in the treatment of neurological disorders associated with AIDS.