Control of Breathing & Glycogen Storage Disease

描述（由申请方提供）：II型糖原累积病（GSDII）由酸性α-葡糖苷酶（GAA）（一种降解糖原的溶酶体酶）缺乏引起。GSDII导致婴儿心肺衰竭，以及青少年和成人的进行性呼吸衰竭。虽然呼吸衰竭已被归因于肌肉病理，尸检病例报告显示颈椎糖原积累，并建议运动神经元病理。我们已经观察到，传出膈放电，平均吸气气流，和每分钟通气量代谢率（VE/VCO 2）的比率在小鼠GSDII模型，GAA-/-“敲除”小鼠钝化。此外，具有正常骨骼肌收缩性和GAA表达但中枢神经系统（CNS）中没有GAA的选择性敲除小鼠也表现出膈输出和通气的显著减少。因此，CNS GAA缺乏与呼吸运动输出受损相关，GSDII中的呼吸功能不全可能反映神经和肌肉病理。初步数据显示GAA-/-小鼠中广泛的颈脊髓糖原积累，特别是在逆行识别的膈运动神经元内，也涉及神经机制。确定呼吸功能不全的潜在机制很重要，因为静脉内酶替代（目前的临床GSDII治疗）不靶向CNS，因为GAA不能穿过血脑屏障。靶向肌肉和中枢神经系统的一种有前途的方法是重组腺相关病毒（rAAV）基因治疗。我们的初步数据表明，rAAV有效地阻断膈运动神经元，并可以通过椎管内或胸腔内注射。此外，在胸内注射包装有GAA基因的rAAV（rAAV-GAA疗法）后一个月，GAA-/-小鼠的通气显著增强。这些实验代表了呼吸生理学（Fuller）、基因治疗（Byrne）和神经解剖学（Reier）实验室之间的独特合作。我们提出了三个假设：1）选择性和完全GAA-/-敲除小鼠对膈肌的神经驱动和通气都减弱; 2）这些小鼠中的呼吸缺陷与膈运动神经元中的糖原积累平行发生，和3）脊柱内和胸内rAAV-GAA递送可以改善选择性和GAA-/-中的脊髓糖原积累并增强呼吸运动输出。敲除小鼠。

DESCRIPTION (provided by applicant): Glycogen storage disease type II (GSDII) results from deficiency of acid a-glucosidase (GAA), a lysosomal enzyme that degrades glycogen. GSDII causes cardiorespiratory failure in infants, and progressive respiratory failure in juveniles and adults. Although respiratory failure has been attributed to muscle pathology, autopsy case reports show cervical spinal glycogen accumulation and suggest motoneuron pathology. We have observed that efferent phrenic discharge, mean inspiratory airflow, and the ratio of minute ventilation to metabolic rate (VE/VCO2) are blunted in a murine GSDII model, the GAA-/- "knockout" mouse. In addition, a selective knockout mouse with normal skeletal muscle contractility and GAA expression, but no GAA in the central nervous system (CNS), also exhibits marked reductions in phrenic output and ventilation. Accordingly, CNS GAA deficiency is associated with impaired respiratory motor output, and respiratory insufficiency in GSDII may reflect both a neural and a muscular pathology. A neural mechanism is also implicated by preliminary data showing extensive cervical spinal glycogen accumulation in GAA-/- mice, particularly within retrogradely identified phrenic motoneurons. Determining the mechanisms underlying respiratory insufficiency is important because i.v. enzyme replacement (the current clinical GSDII therapy) does not target the CNS as GAA cannot cross the blood brain barrier. A promising method for targeting both muscle and the CNS is recombinant adeno-associated virus (rAAV) gene therapy. Our preliminary data indicate that rAAV effectively transfects phrenic motoneurons, and can be delivered by intraspinal or intrathoracic injection. Further, ventilation is significantly enhanced in GAA-/- mice one month following intrathoracic injection of rAAV packaged with the GAA gene (rAAV-GAA therapy). These experiments represent a unique collaboration between laboratories specializing in respiratory physiology (Fuller), gene therapy (Byrne), and neuroanatomy (Reier). We propose to test three hypotheses: 1) neural drive to the diaphragm and ventilation are attenuated in both the selective and full GAA-/- knockout mice; 2) respiratory deficits in these mice occur in parallel with glycogen accumulation in phrenic motoneurons, and 3) intraspinal and intrathoracic rAAV-GAA delivery can ameliorate spinal glycogen accumulation and enhance respiratory motor output in the selective and GAA-/- knockout mice, respectively.