DIDEOXYNUCLEOSIDE TRANSPORT IN THE BLOOD/BRAIN BARRIER

人类免疫缺陷病毒(HIV)在大脑内感染会产生 严重的神经系统并发症。据估计，至少30%的 所有艾滋病成人和50%的艾滋病儿童将遭受中央 与大脑内艾滋病毒感染有关的神经系统疾病。 与艾滋病相关的神经系统并发症的治疗很困难，因为 通常用于艾滋病治疗的抗病毒双脱氧核苷不 容易跨越血脑屏障(BBB)。载体的存在 3‘-叠氮-2’，3‘-二脱氧胸苷(AZT)及其相关体系 血-脑脊液屏障中的双脱氧核苷 有据可查。然而，很少有研究对 脑微血管内双脱氧核苷载体的存在 形成血脑屏障的内皮细胞。在本提案中，承运人 调节双脱氧核苷进出的系统 将对脑微血管内皮细胞进行检查。在工作中 拟议研究的假设是特定的、不同的载体 存在于脑微血管内皮细胞质膜上的系统 控制双脱氧核苷摄取和外流的细胞。 血脑屏障上的双脱氧核苷载体将被识别和表征 采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累积 双脱氧核苷(AZT、双脱氧肌苷、双脱氧胞嘧啶和 BBMEC单层中的双脱氧胸苷)将被检测以确定：1) 每种化合物的摄取率，2)摄取的饱和度和 核苷与载体相互作用所需的结构特征， 3)摄取的能量依赖性和4)载流子在 腔(血侧)和腔(脑侧)质膜。 同一系列放射性标记双脱氧核苷从BBMEC外流 将对单层进行检查以确定：1)对 二脱氧核苷外流到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，以及3)位置和分布 双脱氧核苷载体在BBMEC腔内和腔内的分布 单层。这些研究将批判性地检验细胞机制 控制双脱氧核苷通过血脑屏障。通过 关于摄取和外排载体的特异性的特征 对于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。在以下位置获得的信息 这些研究也可以用来提高目前和 未来抗病毒核苷在神经疾病治疗中的应用 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.