內耳畸形之流行病學與遺傳研究

Epidemiological and Genetic Study in Patients with Inner Ear Malformations

吳振吉(Chen-Chi Wu)

指導教授 ： 陳培哲 ； 許權振

國立臺灣大學/醫學院/臨床醫學研究所/碩士(2004年)

https://doi.org/10.6342/NTU.2004.01182

摘要

研究背景及目的
耳聾是常見的疾病，小孩罹患遺傳性耳聾對個人、家庭和醫療體系都是沈重的負擔。由於分子生物學的進步和人類基因體的解碼，最近數年來，醫界對於導致遺傳性聽障的成因有了顯著的進展。目前已經發現約有三十多個基因與非症候群型聽障的發生有關，歐美等國的研究指出，這些基因中，以Cx26基因最為重要，高達40-60%的病患是因為Cx26的突變造成的。然而，國人的研究則顯示，只有約10-15%的病人可找到Cx26的突變。這不禁讓我們思考，是否還有其他重要基因，導致國人遺傳性耳聾。
隨著影像儀器的進步，使得內耳畸形的診斷率大為提昇。先天性聽障的患者中，約有30%合併內耳畸形，而且釵h證據顯示，內耳畸形可由基因突變造成。由於大多數的遺傳性耳聾屬於非症候群型，無其他身體表徵可以辨識，因此以影像檢查的結果作為表現型的分類基準，來研究遺傳性耳聾，不失為一可行的切入點。雖然國外的研究已指出SLC26A4 (PDS)基因的突變會導致內耳畸形，然而對其致病機轉仍無法提供完整的解釋，且鑒於我國目前還欠缺相關資料，因此吾人著手進行本研究，希望能夠藉由建立本土的內耳畸形檢體庫，釐清SLC26A4基因突變之於國人遺傳性耳聾的重要性，探討各種內耳畸形之間的相關性和致病機轉，並找尋是否有其他新基因可能導致內耳畸形。

研究材料與方法
本研究分兩部分執行。
第一部分為內耳畸形之流行病學研究。吾人回顧性分析臺大醫院過去五年來，因為聽力障礙接受顳骨高解析度電腦斷層掃描檢查，並接受完整聽力檢查之18歲以下病人160例。吾人回顧所有病人之完整病歷，並於其中取得「影像學檢查結果」(radiographic result)、「聽障病因」(etiology)、「聽力喪失形式」(hearing loss pattern)、「聽力圖圖形」(audiogram configuration)、以及「聽損程度」(hearing level)等資料。其後，吾人將所有病人之「影像學檢查結果」加以分類後，再比較「有內耳畸形病人」與「無內耳畸形病人」之間，以及各類內耳畸形病人之間，聽障病因及各種聽力學檢查結果是否有異同。
第二部分為內耳畸形之遺傳研究。吾人自兩年前起前瞻性收集內耳畸形患者及其家族成員，以納入本研究。就方法而言，我們依據典型的基因研究之程序，包括定義疾病之表現型、確認病例、收集病例、收集DNA檢體、處理DNA檢體、基因定型(genotyping)、以及資料分析等步驟，檢測患者及其家屬是否帶有SLC26A4基因突變，分析國人內耳畸形患者中帶有SLC26A4基因突變的比率，探研其基因型與表現型之間的相關性，並驗證SLC26A4基因突變之於各種內耳畸形致病機轉之特異性。

結果
第一部分流行病學的研究結果顯示，160名病例中，有59名(37%)病人可發現內耳畸形。而在總計302隻患耳中，最常見之內耳畸形分別為大前庭導水管(66耳)、Mondini氏發育不全(56耳)、前庭擴大(48耳)、及半規管發育不全(30耳)。此四者常合併出現，有74%的內耳畸形病耳之影像學結果，可單獨或合併以上述四種畸形描述之。約73%的內耳畸形病人，呈現波動性聽力喪失，而上述四類之內耳畸形，更高達九成的病人可追溯到此一病史，明顯高於無內耳畸形病耳的7% (chi-square test, p < 0.05)。內耳畸形病耳之聽力圖形狀，以極重度(54%)、高音漸傾型(27%)及高音急墜型(13%)三者為主，相較於無內耳畸形的病耳，並未發現有「疾病特徵的」(pathognomonic)的聽力圖圖形。而最常見的四種內耳畸形中，聽障的嚴重程度與畸型的種類無關，而與合併畸形的數目有關(ANCOVA, p < 0.05)。由於此四種內耳畸形之盛行率遠較其他種內耳畸形高，且常合併出現，聽力學特徵亦具同質性，於是吾人取此四種畸形之英文字首，而合稱之為「EMVS複合畸形」(EMVS complex)。
至於第二部分之遺傳研究，目前為止共計35個家族計44名內耳畸形患者完成SLC26A4基因突變之檢測。35個家族中，25個家族的初始受試者(proband)其病變主要為上述四種內耳畸形，而其餘10個家族之初始受試者，則以其他罕見內耳畸形為表現，如共同腔(common cavity)、內聽道狹窄、耳蝸發育低下(cochlear hypoplasia)或完全未發育(aplasia)等。在前組25名初始受試者中，計於22位(88%)找到SLC26A4基因突變，而後組10名初始受試者，則未發現任何突變(Fisher’s exact test, p < 0.01)，顯見SLC26A4基因突變僅特異於某些內耳畸形，並初步驗證了吾人於第一部分流行病學研究所提出之假設。吾人共發現7處突變：IVS7-2A>G、1115C>T、1160C>T、1229C>T、1343C>T、2162C>T及2168A>G，其中1160C>T與1343C>T為未曾報告於文獻之新突變，而IVS7-2A>G則是國人最常見之突變，佔所有突變之79%，顯示國人SLC26A4基因之突變光譜(mutation spectrum)，與國外明顯不同。進一步分析基因型與表現型之相聯性，則發現病人之SLC26A4基因型與影像學結果及聽力學結果之間，並無顯著關連。

結論
於第一部分內耳畸形之流行病學研究中，吾人發現，高達37%的聽障病童可發現內耳畸形，其中又以大前庭導水管、Mondini氏發育不全、前庭擴大及半規管發育不全等四類畸形特別常見。由於此四類畸形分別位於四處不同之內耳解剖位置，且經常合併出現，聽力學特徵亦具同質性，因此吾人以為其致病機轉可能雷同。
第二部分內耳畸形之遺傳研究則發現，國人SLC26A4基因之突變光譜與國外大不相同。同時，吾人證實SLC26A4基因突變之於某些特定內耳畸形之特異性，間接驗證了內耳畸形之致病機轉，顯然無法單純由「發育停止」假說來解釋。當然，我們也注意到有些尚未解決的課題，比如SLC26A4基因突變型與表現型之間欠缺相關性，乃至於其他亦會導致內耳畸形基因的尋找等，也都有待吾人進行更深入的研究。

關鍵字

Pendred氏症候群 ； 高解析度電腦斷層掃描 ； Mondini氏發育不全 ； 內耳畸形 ； 聽力學檢查結果 ； 大前庭導水管 ； 臺灣人 ； 發育停止 ； PDS基因 ； 聽障病因 ； SLC26A4基因 ； 耳蝸未完全分隔

並列摘要

PART I. Epidemiological Study in patients with Inner Ear Malformation

Background
With the improvements in imaging technology, congenital bony inner ear malformation, traditionally considered as a rare disease entity, has proven to be a common cause of pediatric deafness. However, a number of unanswered questions remain concerning the pathogenesis of the hearing loss associated with inner ear malformations. Only a careful review of temporal bone images in affected patients with well documented hearing data will clarify these questions.
Objective
As a pilot study for genetic research of inner ear malformation, the first purpose of the study is to establish the epidemiological data in the Taiwanese hearing-impaired population. Also to be investigated are the etiologies and the audiologic data of different types of malformations, which we think might be helpful in elucidating the interrelation between malformations and shedding light on the pathogenesis.
Study design
Retrospective study from 1998 to 2002 at a tertiary-care university hospital.
Methods
One hundred and sixty consecutive children with a total of 302 affected ears undergoing high-resolution CT of the temporal bone for sensorineural hearing loss were enrolled. The image results were correlated with etiologies, hearing loss patterns, hearing levels and audiogram configurations
Results
As many as 37% hearing-impaired children who visited our clinic turned out to demonstrate inner ear malformations on high-resolution CT, and inner ear malformation was present in a total of 114 (38%) ears. The most common malformations were enlarged vestibular aqueduct, incomplete partition of cochlea (Mondini dysplasia), large vestibule, and semicircular canal dysplasia, presenting either as isolated finding or in combination. Eighty-four (74%) ears have abnormalities confined to these four malformations, and only 30 (26%) ears showed other malformations. Patients with complex of enlarged vestibular aqueduct, Mondini dysplasia, large vestibule and semicircular canal dysplasia (EMVS complex) demonstrated a significantly higher incidence of fluctuating hearing loss (93%) and a better hearing level compared to those with other malformations. Homogeneity in audiologic features among these four malformations was also disclosed.
Conclusions
Inner ear malformation proves to be a common finding in the hearing-impaired Taiwanese. By comparing the audiologic data of children with various inner ear malformations, we also identify a distinct clinical entity, the EMVS complex, which is characterized by fluctuating hearing loss and a better hearing level. Based on the homogeneity of audiologic features and previous knowledge about embryogenesis, we propose that malformations belonging to this complex result from a common pathogenetic mechanism.

PART II. Genetic Study in Patients with Inner Ear Malformation

Background
The traditional hypothesis concerning the pathogenesis of inner ear malformations holds that various types of malformations represent different stages of developmental arrest during embryogenesis. Among the various types of inner ear malformations, enlarged vestibular aqueduct (EVA) and Mondini’s dysplasia are by far the most common findings on HRCT, and have been recently linked to mutations in the SLC26A4 (PDS) gene. However, a clear and complete correlation between the SLC26A4 genotypes and the phenotypes still cannot be deciphered. Whether SLC26A4 gene mutations are present in patients with malformations other than EVA and Mondini’s dysplasia is also a question of great interest, because it would undoubtedly yield insight into the pathogenesis of the various types of inner ear malformations.
Objective
This first purpose of the study is to determine the mutation spectrum of the SLC26A6 gene among Taiwanese hearing-impaired patients associated with inner ear malformations. Secondly, the correlation between the genotypes and the phenotypes is investigated, with a special emphasis placed on comparison of the severity of inner ear malformations and the hearing levels, which was scarcely documented in the literature. In order to testify the hypothesis of developmental arrest, we also survey SLC26A4 gene mutations in patients with various types of inner ear malformations, including those with EVA and/or Mondini’s dysplasia, as well as those with other malformations.
Study design
Prospective study from 2002 to 2004 at a tertiary-care university hospital.
Methods
Thirty-five families in which at least one family member demonstrated both hearing impairment and inner ear malformation on HRCT were recruited in the study. Mutations of the SLC26A4 gene were screened in the probands of these families, as well as in 30 hearing-impaired patients without inner ear malformations and 50 unaffected subjects with normal hearing for comparison. The radiological and audiological results of the probands were then compared according to the genotypes.
Results
In 25 families, the probands showed EVA and/or Mondini’s dysplasia as the main temporal bone abnormalities, whereas the probands in the remaining 10 families revealed other types of malformations. A total of 7 mutated SLC26A4 alleles, including 6 missense mutations (A372V, A387V, T410M, S448L, T721M and H723R) and 1 splice site mutation (IVS7-2A>G), were detected. All the mutated alleles segregated the malformation of EVA and Mondini’s dysplasia, whereas no mutated allele was found in the 10 probands with other types of malformations, confirming the specificity of the SLC26A4 mutations in the pathogenesis of EVA and Mondini’s dysplasia.
Among the 7 mutations, the splice site mutation IVS7-2A>G, surprisingly comprised 30 (79%) of the 38 mutated alleles, demonstrating a unique SLC26A4 mutation spectrum in Taiwanese and suggesting a founder effect. Meanwhile, analyses of the radiological findings and audiological results showed no obvious correlation between the genotypes and phenotypes.
Conclusion
Although the radiological findings of the various types of inner ear malformations to some extent resemble each other, and together appear to follow a continuum of morphological changes, their pathogenetic mechanisms are essentially different. The mutation spectrum of the SLC26A4 gene in Taiwanese is different from those in the Caucasoid and the Japanese series, and may be representative of those in Southeastern Asian populations. The lack of genotype-phenotype correlation and the evidence that only one mutated allele was detected in certain affected patients suggest interplay of other genetic or environmental factors with the PDS gene in the pathogenesis.

並列關鍵字

Audiologic findings ； Inner ear malformation ； IVS7-2A>G ； EMVS complex ； Pendred syndrome ； Developmental arrest ； Incomplete partition of cochlea ； Etiology ； SLC26A4 gene ； Taiwanese ； PDS gene ； Enlarged vestibular aqueduct ； High-resolution computed tomography

參考文獻

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