Transcriptomic profiling of medulla and cervical spinal cord to study respiratory neuropathology in Spinocerebellar ataxia type 7

Spinocerebellar ataxia type 7 (SCA7) is an autosomal dominant neurological disorder caused by a deleterious CAG repeat expansion in the coding region of the ataxin-7 gene. The number of CAG repeats determines the severity and age of onset of the disease. Infantile onset SCA7 leads to severe clinical manifestations including dysphagia, aspiration pneumonia and respiratory distress, but the exact cause of respiratory impairment remains unclear. Using the infantile SCA7 mouse model the SCA7266Q/5Q mouse, we examined the impact of mutant ataxin-7 on the respiratory control centers. Specifically, we examined the effect of pathological poly-Q-ataxin-7 expression on hypoglossal (XII) and phrenic motor units. Further, we identified the transcript profile of the medulla and cervical spinal cord and, investigated the XII and phrenic nerve structure as well as the neuromuscular junctions in the diaphragm and tongue. SCA-7 astrocytes showed significant intranuclear and cytoplasmic inclusions of ataxin-7 in the XII and putative phrenic motor nuclei. Transcriptomic analysis revealed dysregulation of genes involved in amino acid and neurotransmitter transportation, and myelination. Additionally, SCA7 mice demonstrated blunted efferent output of the XII nerve and demyelination in both XII and phrenic nerves. Finally, there was an increased number of NMJ clusters with higher expression of synaptic markers in SCA7 mice compared to WT controls. Thus, pathological ataxin-7 expression disrupts myelination and neurotransmitter transportation, and impairs glial cell function in the respiratory control centers. These pre-clinical findings elucidate the underlying pathophysiology responsible for dysphagia, aspiration and respiratory failure in infantile SCA7. The medulla and cervical spinal cord tissues are harvested and flash frozen from SCA7 and wild-type C57Bl/6J mice at different age groups- p8, 5 weeks, and 9-weeks of age. RNA was extracted using Qiagen miRNeasy kit.脊髓小脑共济失调7型（Spinocerebellar ataxia type 7, SCA7）是一类常染色体显性神经系统疾病，由共济失调蛋白7（ataxin-7）基因编码区发生致病性CAG重复扩增所引发。CAG重复序列的拷贝数决定了该疾病的严重程度与发病年龄。婴儿起病型SCA7可出现包括吞咽困难、吸入性肺炎及呼吸窘迫在内的严重临床表现，但呼吸功能损伤的确切机制尚未明确。 本研究采用婴儿型SCA7小鼠模型——SCA7266Q/5Q小鼠，探究了突变型共济失调蛋白7对呼吸调控中枢的影响。具体而言，我们分析了病理性聚谷氨酰胺共济失调蛋白7（poly-Q-ataxin-7）表达对舌下神经（hypoglossal, XII）及膈运动单位的作用。此外，我们鉴定了延髓与颈髓的转录本表达谱，并对舌下神经、膈神经的结构，以及膈肌与舌部的神经肌肉接头（neuromuscular junctions, NMJ）进行了检测。 SCA7星形胶质细胞在舌下神经核及疑似膈运动神经核中，可见显著的共济失调蛋白7核内与胞浆包涵体。转录组分析显示，参与氨基酸与神经递质转运、髓鞘形成的基因存在表达失调现象。此外，SCA7小鼠的舌下神经传出输出功能减弱，且舌下神经与膈神经均出现脱髓鞘改变。相较于野生型（WT）对照组，SCA7小鼠的神经肌肉接头簇数量增多，且突触标志物的表达水平更高。 综上，病理性共济失调蛋白7表达会破坏呼吸调控中枢的髓鞘形成与神经递质转运功能，并损伤神经胶质细胞功能。上述临床前研究结果阐明了婴儿起病型SCA7患者出现吞咽困难、吸入性肺炎及呼吸衰竭的潜在病理生理机制。 本研究采集了不同年龄组（出生后第8天、5周龄及9周龄）的SCA7小鼠与野生型C57Bl/6J小鼠的延髓及颈髓组织，并进行快速冷冻。总RNA提取采用凯杰（Qiagen）miRNeasy试剂盒完成。