Blood-brain barrier (BBB) disruption is a common feature in neurodegenerative diseases. However, BBB integrity has not been assessed in spinocerebellar ataxias (SCAs) such as Machado-Joseph disease/SCA type 3 (MJD/SCA3), a genetic disorder, triggered by polyglutamine-expanded ataxin-3. To investigate that, BBB integrity was evaluated in a transgenic mouse model of MJD and in human post-mortem brain tissues. Firstly, we investigated the BBB permeability in MJD mice by: i) assessing the extravasation of the Evans blue (EB) dye and blood-borne proteins (e.g fibrinogen) in the cerebellum by immunofluorescence, and ii) in vivo Dynamic Contrast Enhanced-Magnetic Resonance Imaging (DCE-MRI). The presence of ataxin-3 aggregates in brain blood vessels and the levels of tight junction (TJ)-associated proteins were also explored by immunofluorescence and western blotting. Human brain samples were used to confirm BBB permeability by evaluating fibrinogen extravasation, co-localization of ataxin-3 aggregates with brain blood vessels and neuroinflammation. In the cerebellum of the mouse model of MJD, there was a 5-fold increase in EB accumulation when compared to age-matched controls. Moreover, vascular permeability displayed a 13-fold increase demonstrated by DCE-MRI. These results were validated by the 2-fold increase in fibrinogen extravasation in transgenic animals comparing to controls. Interestingly, mutant ataxin-3 aggregates were detected in cerebellar blood vessels of transgenic mice, accompanied by alterations of TJ-associated proteins in cerebellar endothelial cells, namely a 29% decrease in claudin-5 oligomers and a 10-fold increase in an occludin cleavage fragment. These results were validated in post-mortem brain samples from MJD patients as we detected fibrinogen extravasation across BBB, the presence of ataxin-3 aggregates in blood vessels and associated microgliosis. Altogether, our results prove BBB impairment in MJD/SCA3. These findings contribute for a better understanding of the disease mechanisms and opens the opportunity to treat MJD with medicinal products that in normal conditions would not cross the BBB.

中文翻译：

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在Machado-Joseph疾病/脊髓小脑共济失调类型3中，血脑屏障被破坏：来自转基因小鼠和人类死后样本的证据。

血脑屏障（BBB）破坏是神经退行性疾病的常见特征。但是，尚未在脊髓小脑共济失调（SCA）中评估BBB完整性，例如Machado-Joseph疾病/ SCA 3型（MJD / SCA3）是一种由聚谷氨酰胺扩展的ataxin-3引发的遗传性疾病。为了调查这一点，在MJD的转基因小鼠模型和人类死后脑组织中评估了BBB的完整性。首先，我们通过以下方法研究了MJD小鼠的血脑屏障通透性：i）通过免疫荧光评估小脑伊文思蓝（EB）染料和血源性蛋白质（例如纤维蛋白原）的外渗，和ii）体内动态对比增强型磁性共振成像（DCE-MRI）。还通过免疫荧光和蛋白质印迹探索了脑血管中共青霉素3聚集体的存在和紧密连接（TJ）相关蛋白的水平。通过评估纤维蛋白原外渗，ataxin-3聚集体与脑血管的共定位以及神经炎症，将人脑样本用于确认BBB渗透性。在MJD小鼠模型的小脑中，与年龄匹配的对照组相比，EB积累增加了5倍。此外，DCE-MRI证实血管通透性增加了13倍。与对照相比，转基因动物中纤维蛋白原外渗增加了2倍，从而验证了这些结果。有趣的是，在转基因小鼠的小脑血管中检测到了突变型共青霉素3聚集体，伴随着小脑内皮细胞中TJ相关蛋白的改变，即claudin-5寡聚物减少了29％，而occludin裂解片段增加了10倍。这些结果在MJD患者的死后脑样本中得到了验证，因为我们检测到BBB中纤维蛋白原外渗，血管中ataxin-3聚集体的存在以及相关的小胶质细胞增生。总之，我们的结果证明了MJD / SCA3中的BBB损伤。这些发现有助于人们更好地了解疾病的机制，并为在正常情况下不会超过血脑屏障的药物治疗MJD提供了机会。这些结果在MJD患者的死后大脑样本中得到了验证，因为我们检测到BBB中纤维蛋白原外渗，血管中ataxin-3聚集体的存在以及相关的小胶质细胞增生。总之，我们的结果证明了MJD / SCA3中的BBB损伤。这些发现有助于人们更好地了解疾病的机制，并为在正常情况下不会超过血脑屏障的药物治疗MJD提供了机会。这些结果在MJD患者的死后脑样本中得到了验证，因为我们检测到BBB中纤维蛋白原外渗，血管中ataxin-3聚集体的存在以及相关的小胶质细胞增生。总之，我们的结果证明了MJD / SCA3中的BBB损伤。这些发现有助于人们更好地了解疾病的机制，并为在正常情况下不会超过血脑屏障的药物治疗MJD提供了机会。