Data from: Gut mucosal cells transfer α-synuclein to the vagus nerve

Tabular raw data corresponding to figure sets used in the study. Figure 1. α-Synuclein expression and seeding activity in SNCAA53T mice. ELISA quantification of human α-synuclein in (C) duodenum (α-synuclein quantification in ng/mg of nodose tissue), (D) nodose ganglia (α-synuclein quantification in pg/mg of nodose tissue), and (E) hindbrain (α-synuclein quantification in ng/mg of nodose tissue), from Snca–/– and SNCAA53T mice. RT-QuIC analysis of (F) duodenum, (G) nodose ganglia, and (H) hindbrain of Snca–/– and SNCAA53T mice. Figure 3. Conditional human α-synuclein expression induces α-synuclein seeding activity in gut organoids. (C) A representative ThT fluorescence profile for these genotypes is provided. (D) Endpoint values were collected after 100 hours of RT-QuIC relative to negative controls. Figure 4. Conditional human α-synuclein expression in gut mucosal cells produces in α-synuclein seeding activity in nodose ganglia. (C) ELISA quantification of human α-synuclein protein in nodose ganglia of nontransgenic, Snca–/–, and SNCAbow mice. (D) A representative ThT fluorescence profile (RT-QuIC) and endpoint analysis of nodose ganglia from nontransgenic (nTg), Snca–/–, and SNCAbow mice at 1 month of age. (E) RT-QuIC analysis of nodose ganglia from 6-month-old nTg, Snca–/–, and SNCAbow mice. Figure 5. Vagotomy spares the nodose ganglia from α-synuclein seeding activity and prevents spread to the hindbrain. (C) ELISA measurements of α-synuclein protein in the gut 3 months after tamoxifen treatment. RT-QuIC analysis of (D and E) vagal nodose ganglia and (F and G) hindbrain analyzed 3 months after tamoxifen treatment. Representative ThT fluorescence profiles are shown in D and F. Epidemiological and histopathological findings have raised the possibility that misfolded α-synuclein protein might spread from the gut to the brain and increase the risk of Parkinson's disease. Although past experimental studies in mouse models have relied on gut injections of exogenous recombinant α-synuclein fibrils to study gut-to-brain α-synuclein transfer, the possible origins of misfolded α-synuclein within the gut have remained elusive. We recently demonstrated that sensory cells of intestinal mucosa express α-synuclein. Here, we employed mouse intestinal organoids expressing human α-synuclein to observe the transfer of α-synuclein protein from epithelial cells in organoids to cocultured nodose neurons devoid of α-synuclein. In mice expressing human α-synuclein, but no mouse α-synuclein, α-synuclein fibril-templating activity emerged in α-synuclein–seeded fibril aggregation assays in intestine, vagus nerve, and dorsal motor nucleus. In newly engineered transgenic mice that restrict pathological human α-synuclein expression to intestinal epithelial cells, α-synuclein fibril-templating activity transfered to the vagus nerve and dorsal motor nucleus. Subdiaphragmatic vagotomy prior to induction of α-synuclein expression in intestinal epithelial cells effectively protected the hindbrain from emergence of α-synuclein fibril-templating activity. Overall, these findings highlight a potential non-neuronal source of fibrillar α-synuclein protein that might arise in gut mucosal cells.本研究配套图表集所对应的表格化原始数据。 图1 SNCAA53T小鼠的α-突触核蛋白表达与播种活性。对Snca–/–与SNCAA53T小鼠的（C）十二指肠（以ng/mg结状组织计的α-突触核蛋白定量值）、（D）结状神经节（以pg/mg结状组织计的α-突触核蛋白定量值）及（E）后脑（以ng/mg结状组织计的α-突触核蛋白定量值）中人源α-突触核蛋白开展酶联免疫吸附试验（ELISA）定量检测。对上述小鼠的（F）十二指肠、（G）结状神经节及（H）后脑进行实时定量Quaking诱导转化试验（RT-QuIC）分析。 图3 条件性人源α-突触核蛋白表达可诱导肠道类器官中的α-突触核蛋白播种活性。（C）给出了对应基因型的代表性硫黄素T（ThT）荧光曲线。（D）收集了相对于阴性对照组、经100小时RT-QuIC反应后的终点检测值。 图4 肠道黏膜细胞中的条件性人源α-突触核蛋白表达可在结状神经节中引发α-突触核蛋白播种活性。（C）对非转基因（nTg）、Snca–/–及SNCAbow小鼠的结状神经节中人源α-突触核蛋白进行ELISA定量检测。（D）展示了1月龄非转基因（nTg）、Snca–/–及SNCAbow小鼠结状神经节的代表性ThT荧光曲线（RT-QuIC）及终点分析结果。（E）对6月龄非转基因（nTg）、Snca–/–及SNCAbow小鼠的结状神经节开展RT-QuIC分析。 图5 迷走神经切断术可使结状神经节免受α-突触核蛋白播种活性侵扰，并阻断其向后脑的扩散。（C）在他莫昔芬处理3个月后，对肠道内的α-突触核蛋白进行ELISA定量检测。对他莫昔芬处理3个月后的（D、E）迷走结状神经节及（F、G）后脑开展RT-QuIC分析。（D）与（F）附代表性ThT荧光曲线。 流行病学与组织病理学研究结果提示，错误折叠的α-突触核蛋白可能从肠道扩散至大脑，并升高帕金森病的发病风险。既往基于小鼠模型的实验研究多通过肠道注射外源性重组α-突触核蛋白纤丝，以探究肠道至大脑的α-突触核蛋白转移过程，但肠道内错误折叠α-突触核蛋白的潜在来源仍未明确。本团队此前已证实，肠黏膜感觉细胞可表达α-突触核蛋白。本研究中，我们利用表达人源α-突触核蛋白的小鼠肠道类器官，观察到类器官上皮细胞中的α-突触核蛋白可转移至共培养的、缺失内源性α-突触核蛋白的结状神经元中。在仅表达人源α-突触核蛋白、不表达小鼠内源α-突触核蛋白的小鼠体内，肠道、迷走神经及背侧运动核的α-突触核蛋白播种聚集实验中均检测到了α-突触核蛋白纤丝模板活性。在新构建的、将病理性人源α-突触核蛋白表达限定于肠上皮细胞的转基因小鼠中，α-突触核蛋白纤丝模板活性可转移至迷走神经与背侧运动核。在肠上皮细胞α-突触核蛋白表达诱导前实施膈下迷走神经切断术，可有效保护后脑免受α-突触核蛋白纤丝模板活性的诱导产生。综上，本研究结果揭示了一种潜在的非神经元源性纤丝状α-突触核蛋白来源，该蛋白可在肠道黏膜细胞中生成。