Role of Werner's Syndrome Protein in Cigarette Smoke-Induced Cellular Senescence

描述(由申请人提供)： 这项申请是为了支持Toru Nyunoya博士发展成为一名独立的研究员和一名有成就的内科科学家。加里·亨宁哈克博士将作为主要导师支持他，以确保他的职业发展计划取得成功。在该奖项期间，职业发展的具体领域包括课程，包括细胞生物学、遗传学和生物统计学。这一应用的重点是研究香烟烟雾诱导成纤维细胞衰老的机制。这些研究与慢性阻塞性肺疾病(COPD)的发病机制直接相关，COPD与细胞衰老、加速衰老和伤口修复受损有关。Werner综合征(WS)是一种由编码RecQ解旋酶家族成员(WRN蛋白)的Werner综合征基因功能丧失突变引起的遗传性疾病，也会加速衰老。然而，到目前为止，还没有研究探索香烟烟雾导致的加速衰老和WS之间的潜在联系。候选假说是暴露在香烟烟雾中通过下调WRN蛋白诱导成纤维细胞衰老。在目标1中，Nyunoya博士将确定导致香烟烟雾诱导肺成纤维细胞和慢性阻塞性肺疾病患者循环纤维细胞衰老的分子途径(S)。香烟烟雾诱导的WRN蛋白表达的变化将在这些不同的人类模型中进行检测。为了对抗香烟烟雾的影响，利用WRN质粒在培养成纤维细胞中过表达WRN蛋白。WRN蛋白与已知的衰老诱导途径如p53和p16之间的潜在联系将通过使用短发夹状RNA进行敲除来探索。在目标2中，Nyunoya博士将在伤口愈合的体外模型中确定香烟烟雾诱导的细胞衰老的作用。具体地说，p53、p16和WRN蛋白在香烟烟雾诱导的迁移功能障碍和细胞外基质调节中的作用将通过shRNA下调p53和p16或WRN蛋白的过度表达来确定。在Nyunoya博士的整个职业发展过程中，他将会见细胞生物学、衰老、生物统计学和研究设计方面的顾问。这种多学科的培训环境将促进他的职业发展，并确保他发展成为一名独立的研究员。阐明香烟烟雾诱导成纤维细胞衰老的机制将有助于我们理解COPD的发病机制，并为未来的治疗方法提供方向。(摘要结束)

DESCRIPTION (provided by applicant): This application is to support the development of Dr. Toru Nyunoya into an independent researcher and an accomplished physician-scientist. Dr. Gary Hunninghake will support him as primary mentor to ensure the success of his career development plan. Specific areas of career development during this award include coursework, including cell biology, genetics, and biostatistics. The focus of this application is the investigation of mechanisms of cigarette smoke-induced fibroblast senescence. These studies directly relate to the pathogenesis of chronic obstructive pulmonary disease (COPD), which has been linked to cellular senescence, accelerated aging and impaired wound repair. Werner's syndrome (WS), a genetic disorder caused by loss-of-function mutations in the Werner's syndrome gene encoding a member of RecQ helicase family (WRN protein) also accelerates aging. However, to date, no study has explored the potential link between cigarette smoke-induced accelerated aging and WS. The candidate hypothesis is that exposure to cigarette smoke induces fibroblast senescence via WRN protein downregulation. In Aim 1, Dr. Nyunoya will identify the molecular pathway(s) responsible for cigarette smoke-induced cellular senescence in lung fibroblasts and in circulating fibrocytes from patients with COPD. Cigarette smoke-induced alterations in WRN protein expression will be examined in these different human models. To counter the cigarette smoke effect, WRN protein will be overexpressed using WRN plasmid transfection in cultured fibroblasts. A potential link between WRN protein and known senescence-inducing pathways such as p53 and p16 will be explored by knockdown using short hairpin RNA. In Aim 2, Dr. Nyunoya will determine the role of cigarette smoke-induced cellular senescence in an in vitro model of wound healing. Specifically, the role of p53, p16 and WRN protein in cigarette smoke-induced migration dysfunction and extracellular matrix regulation will be determined by shRNA knockdown of p53 and p16 or overexpression of WRN protein. Throughout Dr. Nyunoya's career development, he will meet with advisors in cell biology, aging, biostatistics and study design. This multidisciplinary training environment will enhance his career development and ensure that he evolves into an independent researcher. Elucidating the mechanisms of cigarette smoke-induced fibroblast senescence will contribute to our understanding of COPD pathogenesis and generate direction for future treatment modalities. (End of Abstract)