【作者】 刘娟；

【导师】 杨勇；

【作者基本信息】 南京医科大学 ， 流行病与卫生统计学， 2023， 博士

【摘要】 长岛型掌跖角化症(Nagashima-type palmoplantar keratoderma,NPPK)是一种常染色体隐性遗传的非综合征型掌跖角化症,主要分布于中国、日本和韩国等东亚地区,临床表现为掌跖部位逾越性、弥漫性红斑和角化,部分患者伴有不同程度手足脱屑,多汗,异味,遇水后发白呈现海绵状外观,肘部、膝关节和跟腱处等易摩擦部位的皮肤红斑角化等症状。已经明确长岛型掌跖角化症是由SERPINB7基因的双等位基因失功能性突变所致,其中c.796C>T突变位点是东亚人群最常见的致病突变且被证明为始祖突变,即东亚人群的c.796C>T突变位点来自于同一祖先。始祖突变的存在导致了长岛型掌跖角化症在东亚地区的高发,也导致了东亚人中存在众多c.796C>T突变位点的健康携带者。长岛型掌跖角化症是中国人群中最常见的遗传性掌跖角化症(hereditary palmoplantar keratoderma,HPPK)类型,属于罕见病中的常见病,由于罕见病的特殊属性,因此,我国尚缺乏长岛型掌跖角化症患者的流行病学调查研究。针对长岛型掌跖角化症仍有许多关键问题未有深入的探索,本研究旨在建立中国人群的长岛型掌跖角化症队列,拟探究该病的中国人群分布特征、基因突变谱和临床特征,确定频发突变的类型,开展始祖突变位点的溯源工作,阐明长岛型掌跖角化症基因突变的起源。罕见病的流行病学资料获取困难,因此本研究从我国两家医院(中国医学科学院皮肤病医院和北京大学第一医院)和已经建立的掌跖角化症群体中招募长岛型掌跖角化症患者。门诊患者采用调取就诊记录的方式获取患者信息,随后以电话或问卷调查的方式进行详细信息采集。掌跖角化症群体的患者则采用网络问卷调查的方式收集患者的基本信息,收集的患者符合长岛型掌跖角化症临床诊断但未遗传学确诊的患者开展高通量测序和Sanger测序,临床表现和基因检测均符合长岛型掌跖角化症诊断的患者纳入本研究队列。患者入组后自愿参加有关临床特征的问卷调查和生活质量评价问卷调查,以探究长岛型掌跖角化症的临床特征和疾病负担。对于致病突变为频发突变位点的患者随后开展相应的单倍型研究以明确是否存在始祖效应。后续对于明确为始祖突变的位点,进一步探究其起源,通过筛选始祖突变位点上下游的单核苷酸多态性(Single Nucleotide Polymorphisms,SNPs)并开展Sanger测序,分析患者群体的测序结果,结合公共基因数据库中健康中国人的数据来推算始祖突变起源的年代。本研究最终建立了一个包含234例长岛型掌跖角化症患者的队列,本队列收集的患者主要分布在中国的中东部地区。共检测出14个致病突变位点,其中最常见的四个突变位点为c.796C>T,c.522dup T,c.650＿653del CTGT和c.455G>T,占队列中所有SERPINB7突变位点的97.6%,参照中国最大的遗传数据库(the Chinese Millionome Database,CMDB),推算我国长岛型掌跖角化症的患病率约为0.975/10,000。我们的研究显示长岛型掌跖角化症患者最常见的合并症包含甲癣(患病率为40.0%),湿疹(患病率为36.8%)和足癣(患病率为30.3%),分层分析显示不同年龄组患者的合并症分布情况不同,幼年患者更易合并湿疹,成年患者更易发生浅表真菌感染。持续的临床症状,遗传的风险和手部异常外观和异味所致的社交尴尬导致长岛型掌跖角化症患者的生活质量存在中等程度的影响,类似于中度银屑病对患者生活质量的影响程度。SNPs的基因分型研究显示中国人群最常见的四个突变位点(c.796C>T,c.522dup T,c.650＿653del CTGT和c.455G>T)存在跨越124,344 bp～376,162 bp范围的独特单倍型,证实除c.796C>T突变外,c.522dup T,c.650＿653del CTGT和c.455G>T突变位点均为中国人群的始祖突变位点。参考国际千人基因组数据库(1000 Genome Project Database,1000G数据库)中正常人群的基因频率信息,根据连锁不平衡衰减原理推算各始祖突变发生的时间为2,150～3,160年前,结合人类学的研究和历史事件,推测长岛型掌跖角化症的在亚洲最常见的致病位点c.796C>T最初起源于3,160年前的黄河流域或西辽河流域,随着人群的扩张和农业的发展进一步在中国,日本和韩国等东亚地区传播开来。总之,我们建立了迄今为止国际上最大的长岛型掌跖角化症队列,更新了我国长岛型掌跖角化症的突变分布谱,阐明了疾病的患病率,揭示了长岛型掌跖角化症的一般临床特征和疾病的生活负担,并确定了中国人群的三个新的始祖突变,并开展了突变溯源。我们的研究对于长岛型掌跖角化症的理解和未来长岛型掌跖角化症患者的临床管理、遗传咨询等提供了重要的科学依据和理论基础。更多还原

【Abstract】 Nagashima-type palmoplantar keratoderma(NPPK)is a hereditary palmoplantar keratoderma that is prevalent in East Asian populations,such as China,Japan,and Korea.NPPK is an autosomal recessive genodermatosis that is characterized by the presence of transgradient diffuse erythema and hyperkeratotic lesions on the palms and soles,which may extend to the dorsal surfaces of the palms,feet,elbows,knees,and the Achilles tendon area.Some patients can also experience peeling/scaling,hyperhidrosis,a whitish spongy appearance after exposure to water,and odor of the palms and soles,among other symptoms.The genetic basis of NPPK has been established as biallelic loss-of-function mutations in the SERPINB7 gene.The recurrent mutation c.796C>T is the most common mutation site among East Asian populations and has been verified as a founder mutation.In other words,this nonsense mutation,c.796C>T,is prevalent in Chinese,Japanese,and Korean NPPK patients and probably descended from the same ancestor.The existence of the founder mutation is the reason for the high incidence of NPPK in East Asia,and has also led to many healthy carriers of the c.796C>T mutation in East Asians.NPPK is the most prevalent hereditary palmoplantar keratoderma(HPPK)in China.However,the epidemiological data on the Chinese NPPK population is limited,and crucial questions about NPPK are yet to be answered.Therefore,the aim of the current study is to establish a Chinese NPPK cohort and investigate the clinical and genetic characteristics,demographic distribution,and disease burden of NPPK in China.Additionally,this study endeavors to identify any potential founder mutations among Chinese NPPK patients and evaluate their origins.Epidemiological data on rare diseases are often lacking,which poses a challenge for conducting comprehensive studies.In order to address this gap,the current study enrolled NPPK patients,including NPPK patients who had visited dermatology departments in Chinese Academy of Medical Sciences and Peking University First Hospital,and NPPK patients from a “PPK patient group” established by a NPPK patient in 2015.The information of outpatients was obtained from medical records,and detailed information of NPPK patients was collected through online questionnaires,which were followed by telephone consultations with dermatologists.To establish a cohort of NPPK patients,individuals from the online PPK patient group with a suspected diagnosis of NPPK underwent Next-generation sequencing and Sanger sequencing.Those who met both the clinical and genetic diagnostic criteria of NPPK were ultimately included in the study.Patients with typical clinical phenotype and definitive genetic diagnosis were requested to complete questionnaires to explore the clinical features and the quality of life(QOL)of NPPK patients,which were used to evaluate the clinical features and the burden of the disease.Haplotype analysis was conducted in patients who harbored the four recurrent SERPINB7 mutations,to determine whether these mutations were founder mutations or hotspot mutations.To explore the origin of the confirmed founder mutations,genotyping of Single Nucleotide Polymorphisms(SNPs)flanking the founder mutation sites was performed using haploview software.The results of the sequencing were recorded and examined,and compared with ethnically matched controls in the public database to estimate the age of each founder mutation.The study ultimately recruited 234 Chinese patients with NPPK,who were distributed throughout China,with a higher incidence in mid-eastern China.A total of14 pathogenic mutations were identified in SERPINB7 from the cohort,with the top four recurrent mutations being c.796C>T,c.522 dup T,c.650＿653del CTGT,and c.455G>T,accounting for 97.6% of Chinese NPPK patients.The estimated prevalence of NPPK in China was found to be 0.975/10,000 based on Chinese databases(the Chinese Millionome Database,CMDB).The study also found that the most common comorbidities in NPPK patients were onychomycosis(40.0%),eczema(36.8%),and tinea pedis(30.3%).The analysis by age showed differences in phenotypes between pediatric and adult NPPK patients.Atopic eczema was more prevalent in the pediatric group compared with the adult group,while the adult group showed higher incidences of superficial fungal infections such as tinea pedis and onychomycosis.The lifelong manifestations,hereditary risk,and social embarrassment due to odor or visible lesions have lasting effects on the NPPK patients and their guardians,which are associated with a moderate effect on the quality of life in patients,which similar to the moderate psoriasis.Genotyping results of the tag SNPs indicated that the four major SERPINB7 mutations,c.796C>T,c.522 dup T,c.650＿653del CTGT and c.455G>T,in Chinese NPPK patients exist a unique haplotype spanning 124,344 bp to 376,162 bp region.Haplotype analysis further indicates that all four mutations are founder mutations in the Chinese population.Based on the decay of linkage disequilibrium(LD)between the mutations and surrounding genetic markers,the time of occurrence of these founder mutations in the Chinese population was estimated to date back to2150～3,160 years ago,consistent with anthropological and historical research suggesting the c.796C>T mutation originated in the Yellow River or West Liao River region 3,160 years ago and subsequently spread to China,Korea,Japan,and other parts of East Asia through the dispersal of agriculture and migration.This study presents the largest cohort of NPPK patients to date and expands the mutation spectrum of SERPINB7 while assessing the prevalence of the disease.The findings provide insights into the clinical presentations and impact of NPPK on QOL and confirm the existence of three newly identified founder mutations in SERPINB7 in the Chinese population.Furthermore,this is the first study to explore the time of origin of several founder mutations,which improves the current understanding of NPPK and informs evidence-based recommendations for clinical management,diagnosis,and genetic counseling for NPPK patients.更多还原