Study of iPS cells-derived neural and glial cells from subjects with Monge's disease

摘要 这项建议的总体目标是更好地了解慢性山区大脑是如何受到影响的 疾病(CMS)或蒙格氏病。大脑是CMS的主要靶点之一，表现为 常见的中枢神经系统症状，如头痛、头晕、睡眠障碍和精神错乱。磁共振成像 研究发现CMS患者脑内存在脑水肿和多个缺血灶，提示 脑细胞损伤/死亡发生在CMS中，可能导致上述神经症状。因此， 了解CMS中脑细胞对低氧的易感性对于开发治疗CMS至关重要 不育系的策略。最近，我们从CMS患者和非CMS健康高地人身上获得了皮肤活检。 并将皮肤细胞重新编程为诱导多能干细胞(IPSCs)。IPSC当时是 分化为神经前体细胞，并进一步分化为星形胶质细胞和神经元。然后我们检查了酸- 基础调控机制及其对缺氧/缺血应激的耐受性/脆弱性 星形胶质细胞/神经元。我们的初步数据显示，来自CMS的星形胶质细胞表现出更高的酸性 挤压速率和CMS星形胶质细胞/神经元更容易受到缺氧/缺血挑战 CMS细胞。因此，我们假设CMS患者的星形胶质细胞/神经元已经改变了酸碱。 使神经元/星形胶质细胞更容易受到缺氧影响的调节机制。我们将对此进行测试 1)检测NHE1和NBCe1在CMS和CMS中的表达和活性 常氧和低氧条件下非CMS星形胶质细胞/神经元；2)研究NHE1和NHE1的作用 NBCe1在CMS星形胶质细胞/神经元缺氧后细胞死亡中的作用我们将使用H+和Na+成像系统 结合NHE1和NBCe1的基因敲除和药理抑制来确定其 在CMS星形胶质细胞/神经元对低氧应激耐受/易感性中的作用。我们研究的一个新特点是 直接来源于CMS和非CMS高原人群的体外CMS模型的建立。它是 从我们的研究中，我们很可能会发现一些有助于我们设计更好的CMS的发现 治疗策略。

ABSTRACT The overall goal of this proposal is to better understand how the brain is affected in Chronic Mountain Sickness (CMS) or Monge's disease. The brain is one of the major targets of CMS, as manifested by the frequently seen CNS symptoms such as headache, dizziness, sleep disturbance, and mental confusion. MRI studies have revealed cerebral edema and multiple ischemic foci in the brain of CMS patients, suggesting that brain cell damage/death occurs in CMS which may lead to the above neurological symptoms. Therefore, understanding the vulnerability of brain cells to hypoxia in CMS is critical for developing the therapeutic strategy of CMS. Recently, we obtained skin biopsies from CMS patients and non-CMS healthy highlanders and re-programmed the skin cells into induced pluripotent stem cells (iPSCs). The iPSCs were then differentiated into neuroprogenitor cells and further into astrocytes and neurons. We then examined the acid- base regulatory mechanisms and the tolerance/vulnerability to hypoxia/ischemia stress in these astrocytes/neurons. Our preliminary data show that astrocytes derived from CMS exhibited increased acid extrusion rate, and CMS astrocytes/neurons were more vulnerable to hypoxic/ischemic challenges than non- CMS cells. Therefore, we hypothesize that astrocytes/neurons in CMS patients have altered acid-base regulatory mechanisms which render neurons/astrocytes more vulnerable to hypoxia. We will test this hypothesis via two specific aims: 1) examine the expression and activities of NHE1 and NBCe1 in CMS and non-CMS astrocytes/neurons under normoxia and hypoxia conditions; 2) investigate the role of NHE1 and NBCe1 in the cell death of CMS astrocytes/neurons following hypoxia. We will use H+ and Na+ imaging system together with the genetic knockdown and pharmacological inhibition of NHE1 and NBCe1 to determine their role in the tolerance/vulnerability of CMS astrocytes/neurons to hypoxic stress. A novel feature of our study is the establishment of an in vitro CMS model that was directly derived from CMS and non-CMS highlanders. It is highly likely from our studies that we will make a number of discoveries that will help us design better CMS therapeutic strategies.