Exploring the role of neuromuscular excitability in sudden infant death

婴儿猝死综合征是指看似健康的婴儿意外死亡，尽管对死亡现场和情况进行了彻底的调查，进行了全面的尸检，并对现有信息进行了多专业审查，但仍无法解释死亡原因。它是高收入国家新生儿后死亡的主要原因。英国每周有5名儿童死于小岛屿发展中国家。目前关于死因的证据表明，“三重风险”汇聚在一起：脆弱的婴儿、关键的发育期和外部应激源，如发烧。电信号控制肌肉的兴奋性，即肌肉收缩和放松的能力。多个蛋白质负责产生和控制这些信号。这些基因的突变可能导致临床疾病，如先天性肌无力。所有这些疾病往往会产生间歇性的肌肉症状，但在两者之间，个人看起来很健康。由于它们影响的是肌肉的电路而不是结构，它们也不会在死后留下印记。这些疾病可能会损害呼吸，并以与小岛屿发展中国家难以区分的方式致命。SCN4A基因编码肌肉兴奋性最关键的蛋白质之一，一种名为Nav1.4的离子通道。SCN4A基因突变引起的临床疾病会导致呼吸肌损伤，包括危及生命的呼吸暂停(呼吸停止)。基于我在婴儿身上所做的观察，我与我的研究伙伴贝尔教授合作，获得了新的遗传和功能证据，证明在某些情况下存在类似的SCN4A基因变异(SID)。我现在假设，像Nav1.4这样的离子通道，以及其他对骨骼肌兴奋性至关重要的蛋白质，可能共同导致了更大比例的SIDS病例(导致脆弱的婴儿风险)，并且正常的肌肉发育可能会改变这些变异在早期生命中的临床严重性。严重程度和风险可能随着年龄的增长而降低，即关键的发育期。这项建议的目的是：1.确定编码影响骨骼肌兴奋性蛋白的罕见基因变体的频率，在约700例SID病例和英国生物库对照中2。确定由这些相同基因的变异引起的神经肌肉兴奋性疾病的家庭中SID的频率。描述人类骨骼肌中肌肉纤维类型的正常发育变化4.使用计算机模型来确定组合的遗传、发育和外源性变量对肌肉兴奋性的生理后果--“三重风险”我将使用来自已死于SIDs的婴儿的C.700个DNA样本的完整外显子组测序数据，在候选基因列表(对肌肉兴奋性至关重要的基因)中筛选预测会影响蛋白质功能的非常罕见或新的变种。这些基因变异在小岛屿发展中国家病例中被发现的频率将与英国生物库对照测序数据进行比较，以确定它们是否在小岛屿发展中国家病例中更常见。我还将使用这些基因突变导致临床疾病的家庭的临床数据来确定婴儿猝死的发生率，以及这种情况是否比普通人群中更常见。肌肉由快纤维和慢纤维两种亚型组成。我认为，肌肉中的快速纤维越多，基因变异的影响就越严重，婴儿肌肉中的快速纤维会比年龄较大的儿童更多。一种对单个骨骼肌切片中不同纤维类型应用不同颜色抗体的全自动方案将用于0至16岁的人类对照活组织检查，以确定不同年龄段每种纤维类型的比例。一个肌肉纤维的计算机模型将被用来确定发育变量对正常和突变(带有基因变体)的肌肉纤维功能和表现的单独和综合影响。

Sudden infant death syndrome (SIDS) is the unexpected death of a seemingly healthy infant that remains unexplained despite a thorough investigation of the scene and circumstances of death, a comprehensive post-mortem examination, and a multi-professional review of the available information. It is the leading cause of post-neonatal infant death in high income countries. Five children in the UK die each week from SIDS. Current evidence as to the cause of death proposes convergence of "a triple risk": a vulnerable infant, a critical developmental period and an external stressor e.g. fever. Electrical signals control a muscle's excitability i.e. it's ability to contract and relax. Multiple proteins are responsible for generating and controlling these signals. Mutations in these genes can cause clinical diseases e.g. congenital myasthenia. All of these diseases tend to produce episodic muscle symptoms but in between individuals appear healthy. As they affect a muscle's electrical circuit rather than structure they also leave no post-mortem hallmark. These diseases may impair breathing and be fatal in a manner that is indistinguishable from SIDS. The SCN4A gene codes for one of the most essential proteins for muscle excitability, an ion channel called Nav1.4. Clinical disorders due to mutations in the SCN4A gene cause breathing muscle impairment including life threatening apnoeas (cessation of breathing) in infants. Based on observations I have made in infants I collaborated with my research partner Prof Behr and obtained new genetic and functional evidence that qualitatively similar SCN4A gene variants occur in some cases of (SIDS). I now hypothesise that ion channels like Nav1.4, and other proteins critical for skeletal muscle excitability may collectively account for a larger proportion of SIDS cases (contributing to the vulnerable infant risk) and that normal muscle development may change the clinical severity of these variants in early life. A severity and risk that may get less with age i.e. a critical developmental period. This proposal aims to: 1. Determine the frequency of rare gene variants encoding proteins that influence skeletal muscle excitability in c.700 SIDS cases and UK biobank controls2. Determine the frequency of SIDS amongst families with neuromuscular excitability diseases caused by variants in these same genes3. Characterise the normal developmental changes of muscle fibre types in human skeletal muscle 4. Use a computer model to determine the physiological consequences of combined genetic, developmental and exogenous variables on muscle excitability - the "triple risk"I will use whole exome sequencing data from c.700 DNA samples of infants who have died from SIDS and screen for very rare or new variants in a candidate gene list (genes critical for muscle excitability) that are predicted to affect protein function. The frequency that these gene variants are found in the SIDS cases will be compared with the UK biobank control sequencing data to see if they are more common in SIDS cases. I will also use clinical data from families with mutations in these genes causing clinical disease to ascertain the incidence of sudden infant deaths and whether this is more common than among the general population. Muscles are made up of subtypes of fast and slow fibres. I think the more fast fibres a muscle has the more severe the effect of the gene variants will be and that infants' muscle will have more fast fibres than older children. A fully automated protocol applying different coloured antibodies to the different fibre types in a single skeletal muscle section will be used on human control biopsies from age 0 to 16years to determine the proportion of each fibre type at different ages. A computer model of a muscle fibre will be used to ascertain the individual and combined effects of developmental variables on normal and mutant (with a gene variant) muscle fibre function and performance.