Identification of affected cellular targets, mechanisms and signalling pathways in mouse and cell models for spinal muscular atrophy with respiratory distress type 1 (SMARD1).

鉴定小鼠和细胞模型中受影响的细胞靶标、机制和信号通路，以治疗 1 型呼吸窘迫型脊髓性肌萎缩症 (SMARD1)。

• 批准号: 268785760
• 负责人: Privatdozentin Dr. Sibylle Jablonka
• 金额: --
• 依托单位: Institut für Klinische Neurobiologie
• 依托单位国家: 德国
• 项目类别: Research Grants
• 财政年份: 2014
• 资助国家: 德国
• 起止时间: 2013-12-31 至 2017-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/268785760?language=en
• 关键词: Identification; affected; cellular; targets; mechanisms

Muscle atrophy, and diaphragmatic palsy are the clinical characteristics of spinal muscular atrophy with respiratory distress type 1 (SMARD1), and are well represented in the neuromuscular degeneration (Nmd2J) mouse, modeling the juvenile form of SMARD1. Both in humans and mice mutations in the IGHMBP2 gene lead to motoneuron degeneration. We could previously demonstrate that treatment with a polyethylene glycol-coupled variant of IGF1 (PEG-IGF1) improves motor functions accompanied by reduced fiber degeneration in the gastrocnemius muscle and the diaphragm of the Nmd2J mouse, but has no beneficial effect on motoneuron survival. IGHMBP2/Ighmbp2 is a ribosome associated ATPase/helicase supposed to be involved in ribosomal events/orchestration and/or translational events. These data raised the question which affected cell autonomous disease mechanisms and signalling pathways contribute to dysfunction and loss of Ighmbp2 deficient motoneurons and muscle fibers. An analysis of primary Ighmbp2 deficient motoneurons and spinal cord tissue from Nmd2J mice exhibited differentiation deficits with excitability failures corresponding to reduced N-type specific voltage-gated calcium channel (Cav2.2) accumulation and a dysregulation in the expression of transient receptor potential channels (TRPCs). In order to discover affected cellular mechanisms and signalling pathways, we will start a detailed morphological and functional analysis of primary Ighmbp2 deficient motoneurons. Primary Ighmbp2 deficient mouse motoneurons under specific culture conditions will be subjected to a Fluorescence-Recovery after Photobleaching (FRAP) and a Calcium Imaging study. We finally want to find out which specific signalling pathways are affected in Ighmbp2 deficient motoneurons leading to functional failures. Simultaneously to the in vitro study, we will analysed in detail Nmd2J mice on a TRPC5 and Nav1.9 knock-out background to discover whether these ion channels are modifiers of the Nmd2J phenotype. This indirect identification of a cellular target will be completed by microarray studies in order to identify other affected cellular targets on RNA and protein levels. Furthermore, a re-differentiation approach with skin fibroblast from SMARD1 patients is planned. The re-differentiated human motoneurons will be functional and morphological compared with primary mouse motoneurons to verify their potency as a reliable tool for the further analysis of functional deficits in Ighmbp2 mutated motoneurons. In a last workpage we want to analyse in detail the cell autonomous mechanisms which lead to myopathic changes of diaphragm and gastrocnemius muscle in the SMARD1 mouse. By means of our project we would like to understand affected cellular mechanisms in Ighmbp2 deficient motoneurons and muscles in order to identify signalling pathways which could be stimulated and thus bypass the affected cellular mechanisms.

肌肉萎缩和膈肌麻痹是脊髓性肌萎缩伴呼吸窘迫1型（SMARD1）的临床特征，在神经肌肉变性（Nmd2J）小鼠中有很好的表现，模拟了SMARD1的幼年型。在人类和小鼠中，IGHMBP2基因的突变都会导致运动神经元变性。我们之前可以证明，用聚乙二醇偶联的IGF1变体（PEG-IGF1）治疗可以改善Nmd2J小鼠的运动功能，同时减少腓骨肌和膈肌的纤维变性，但对运动神经元的存活没有有益的影响。IGHMBP2/ IGHMBP2是一种核糖体相关的atp酶/解旋酶，被认为参与核糖体事件/协调和/或翻译事件。这些数据提出了一个问题，即受影响的细胞自主疾病机制和信号通路导致Ighmbp2缺陷运动神经元和肌纤维的功能障碍和丧失。对Nmd2J小鼠原发性Ighmbp2缺陷运动神经元和脊髓组织的分析显示分化缺陷和兴奋性失败，这与n型特异性电压门控钙通道（Cav2.2）积累减少和瞬时受体电位通道（trpc）表达失调有关。为了发现受影响的细胞机制和信号通路，我们将开始对原发性Ighmbp2缺陷运动神经元进行详细的形态学和功能分析。在特定培养条件下，原发性Ighmbp2缺陷小鼠运动神经元将进行光漂白后荧光恢复（FRAP）和钙成像研究。我们最后想要找出哪些特定的信号通路在Ighmbp2缺陷运动神经元中受到影响，从而导致功能失效。在体外研究的同时，我们将详细分析TRPC5和Nav1.9敲除背景下的Nmd2J小鼠，以发现这些离子通道是否是Nmd2J表型的修饰因子。这种细胞靶点的间接鉴定将通过微阵列研究来完成，以便在RNA和蛋白质水平上鉴定其他受影响的细胞靶点。此外，计划使用来自SMARD1患者的皮肤成纤维细胞进行再分化。再分化的人类运动神经元将与原代小鼠运动神经元进行功能和形态比较，以验证其作为进一步分析Ighmbp2突变运动神经元功能缺陷的可靠工具的效力。在最后一篇文章中，我们想详细分析导致SMARD1小鼠膈肌和腓肠肌肌病改变的细胞自主机制。通过我们的项目，我们希望了解Ighmbp2缺陷运动神经元和肌肉中受影响的细胞机制，以确定可以刺激的信号通路，从而绕过受影响的细胞机制。

项目成果

Impaired Local Translation of β-actin mRNA in Ighmbp2-Deficient Motoneurons: Implications for Spinal Muscular Atrophy with respiratory Distress (SMARD1)

Ighmbp2 缺陷运动神经元中 β 肌动蛋白 mRNA 的局部翻译受损：对脊髓性肌萎缩伴呼吸窘迫的影响 (SMARD1)

• DOI: 10.1016/j.neuroscience.2018.06.019
• 发表时间: 2018
• 期刊: Neuroscience
• 影响因子: 3.3
• 作者: Surrey V;Zöller C;Lork AA;Moradi M;Balk S;Dombert B;Saal-Bauernschubert L;Briese M;Appenzeller S;Fischer U;Jablonka S
• 通讯作者: Jablonka S

Disease mechanisms in spinal muscular atrophy with respiratory distress type 1 (SMARD1): what about motoneurons?

脊髓性肌萎缩症伴呼吸窘迫 1 型 (SMARD1) 的疾病机制：运动神经元又如何？

• DOI: 10.18103/mra.v6i2.1689
• 发表时间: 2018
• 期刊: Medical research archives
• 作者: Surrey V;Jablonka S
• 通讯作者: Jablonka S