Despite extensive research, targeted delivery of substances to the brain still poses a great challenge due to the selectivity of the blood-brain barrier (BBB). Most molecules require either carrier- or receptor-mediated transport systems to reach the central nervous system (CNS). These transport systems form attractive routes for the delivery of therapeutics into the CNS, yet the number of known brain endothelium-enriched receptors allowing the transport of large molecules into the brain is scarce. Therefore, to identify novel BBB targets, we combined transcriptomic analysis of human and murine brain endothelium and performed a complex screening of BBB-enriched genes according to established selection criteria. As a result, we propose the high-affinity cationic amino acid transporter 1 (SLC7A1) as a novel candidate for transport of large molecules across the BBB. Using RNA sequencing and in situ hybridization assays, we demonstrated elevated gene expression in both human and mouse brain endothelium. Moreover, we confirmed SLC7A1 protein expression in brain vasculature of both young and aged mice. To assess the potential of SLC7A1 as a transporter for larger proteins, we performed internalization and transcytosis studies using a radiolabelled or fluorophore-labelled anti-SLC7A1 antibody. Our results showed that SLC7A1 internalised a SLC7A1-specific antibody in human colorectal carcinoma (HCT116) cells. Moreover, transcytosis studies in both immortalised human brain endothelial (hCMEC/D3) cells and primary mouse brain endothelial cells clearly demonstrated that SLC7A1 effectively transported the SLC7A1-specific antibody from luminal to abluminal side. Therefore, here in this study, we present for the first time the SLC7A1 as a novel candidate for transport of larger molecules across the BBB.

中文翻译：

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溶质载体SLC7A1可能充当血脑屏障的蛋白质转运蛋白

尽管进行了广泛的研究，但由于血脑屏障（BBB）的选择性，将物质定向输送到大脑仍然面临着巨大的挑战。大多数分子需要载体或受体介导的运输系统才能到达中枢神经系统 (CNS)。这些转运系统形成了将治疗药物输送到中枢神经系统的有吸引力的途径，但已知的允许将大分子转运到大脑的脑内皮富集受体的数量却很少。因此，为了确定新的 BBB 靶标，我们结合了人和小鼠脑内皮的转录组分析，并根据既定的选择标准对富含 BBB 的基因进行了复杂的筛选。因此，我们提出高亲和力阳离子氨基酸转运蛋白 1 (SLC7A1) 作为跨 BBB 转运大分子的新候选者。通过 RNA 测序和原位杂交测定，我们证明人和小鼠脑内皮细胞中的基因表达均升高。此外，我们证实了年轻和老年小鼠脑血管系统中 SLC7A1 蛋白的表达。为了评估 SLC7A1 作为较大蛋白质转运蛋白的潜力，我们使用放射性标记或荧光团标记的抗 SLC7A1 抗体进行了内化和转胞吞作用研究。我们的结果表明，SLC7A1 在人结直肠癌 (HCT116) 细胞中内化了 SLC7A1 特异性抗体。此外，永生化人脑内皮 (hCMEC/D3) 细胞和原代小鼠脑内皮细胞的转胞吞作用研究清楚地表明，SLC7A1 有效地将 SLC7A1 特异性抗体从腔侧转运至腔侧。因此，在这项研究中，我们首次将 SLC7A1 作为跨 BBB 运输较大分子的新候选者。