Failure to clear Aβ from the brain is partly responsible for Aβ brain accumulation in Alzheimer’s disease (AD). A critical protein for clearing Aβ across the blood-brain barrier is the efflux transporter P-glycoprotein (P-gp). In AD, P-gp levels are reduced, which contributes to impaired Aβ brain clearance. However, the mechanism responsible for decreased P-gp levels is poorly understood and there are no strategies available to protect P-gp. We previously demonstrated in isolated brain capillaries ex vivo that human Aβ40 (hAβ40) triggers P-gp degradation by activating the ubiquitin-proteasome pathway. In this pathway, hAβ40 initiates P-gp ubiquitination, leading to internalization and proteasomal degradation of P-gp, which then results in decreased P-gp protein expression and transport activity levels. Here, we extend this line of research and present results from an in vivo study using a transgenic mouse model of AD (human amyloid precursor protein (hAPP)-overexpressing mice; Tg2576). In our study, hAPP mice were treated with vehicle, nocodazole (NCZ, microtubule inhibitor to block P-gp internalization), or a combination of NCZ and the P-gp inhibitor cyclosporin A (CSA). We determined P-gp protein expression and transport activity levels in isolated mouse brain capillaries and Aβ levels in plasma and brain tissue. Treating hAPP mice with 5 mg/kg NCZ for 14 days increased P-gp levels to levels found in WT mice. Consistent with this, P-gp-mediated hAβ42 transport in brain capillaries was increased in NCZ-treated hAPP mice compared to untreated hAPP mice. Importantly, NCZ treatment significantly lowered hAβ40 and hAβ42 brain levels in hAPP mice, whereas hAβ40 and hAβ42 levels in plasma remained unchanged. These findings provide in vivo evidence that microtubule inhibition maintains P-gp protein expression and transport activity levels, which in turn helps to lower hAβ brain levels in hAPP mice. Thus, protecting P-gp at the blood-brain barrier may provide a novel therapeutic strategy for AD and other Aβ-based pathologies.

中文翻译：

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在阿尔茨海默病小鼠模型中保护血脑屏障处的 P-糖蛋白免于降解

未能从大脑中清除 Aβ 是阿尔茨海默病 (AD) 中 Aβ 脑积聚的部分原因。通过血脑屏障清除 Aβ 的关键蛋白质是外排转运蛋白 P-糖蛋白 (P-gp)。在 AD 中，P-gp 水平降低，这会导致 Aβ 脑清除受损。然而，导致 P-gp 水平降低的机制知之甚少，并且没有可用的策略来保护 P-gp。我们之前在离体的脑毛细血管中证明了人类 Aβ40 (hAβ40) 通过激活泛素-蛋白酶体途径触发 P-gp 降解。在该途径中，hAβ40 启动 P-gp 泛素化，导致 P-gp 的内化和蛋白酶体降解，然后导致 P-gp 蛋白表达和转运活性水平降低。这里，我们扩展了这一研究领域，并展示了使用 AD 转基因小鼠模型（人淀粉样前体蛋白 (hAPP) 过表达小鼠；Tg2576）的体内研究结果。在我们的研究中，hAPP 小鼠用载体、诺考达唑（NCZ，微管抑制剂阻断 P-gp 内化）或 NCZ 和 P-gp 抑制剂环孢菌素 A (CSA) 的组合进行治疗。我们测定了离体小鼠脑毛细血管中的 P-gp 蛋白表达和转运活性水平，以及血浆和脑组织中的 Aβ 水平。用 5 mg/kg NCZ 治疗 hAPP 小鼠 14 天后，P-gp 水平增加到在 WT 小鼠中发现的水平。与此一致，与未治疗的 hAPP 小鼠相比，NCZ 治疗的 hAPP 小鼠的脑毛细血管中 P-gp 介导的 hAβ42 转运增加。重要的是，NCZ 治疗显着降低了 hAPP 小鼠的 hAβ40 和 hAβ42 脑水平，而血浆中的 hAβ40 和 hAβ42 水平保持不变。这些发现提供了体内证据，表明微管抑制维持 P-gp 蛋白表达和转运活性水平，这反过来有助于降低 hAPP 小鼠的 hAβ 脑水平。因此，在血脑屏障保护 P-gp 可能为 AD 和其他基于 Aβ 的病理提供一种新的治疗策略。