論文 - 鳴海　覚志

変異TSH受容体V711FfsX18の機能解析 1塩基欠失によるたんぱく質不安定化シグナルの獲得

鳴海 覚志, 杉澤 千穂, 阿部 清美, 長谷川 奉延

日本内分泌学会雑誌 （(一社)日本内分泌学会）  90 （ 2 ） 528 - 528 2014年09月

ISSN  0029-0661

奇異性123I摂取率高値を認めた両アリル性TSHR変異の一男児例 世界で5例目の非古典型TSH不応症

杉澤 千穂, 鳴海 覚志, 阿部 清美, 長谷川 奉延

日本内分泌学会雑誌 （(一社)日本内分泌学会）  90 （ 2 ） 507 - 507 2014年09月

ISSN  0029-0661

TPO遺伝子異常による甲状腺腫性クレチン症の8歳女児例

福留 啓祐, 坂口 善市, 冨田 理絵, 藤岡 啓介, 阿部 容子, 木下 ゆき子, 市原 朋子, 藤井 笑子, 幸山 洋子, 関口 隆憲, 大原 克明, 鳴海 覚志, 阿部 清美, 長谷川 奉延

日本小児科学会雑誌 （(公社)日本小児科学会）  118 （ 7 ） 1126 - 1126 2014年07月

ISSN  0001-6543

サイログロブリン著明高値、甲状腺腫大を契機に診断したDUOX2異常症の2例

後藤 元秀, 荒木 俊介, 久保 和泰, 川越 倫子, 山本 幸代, 楠原 浩一, 齋藤 玲子, 河田 泰定, 阿部 清美, 鳴海 覚志, 長谷川 奉延

日本小児科学会雑誌 （(公社)日本小児科学会）  118 （ 6 ） 1016 - 1016 2014年06月

ISSN  0001-6543

新生児マススクリーニング(MS)で発見されなかった先天性甲状腺機能低下症の3例

上村 美季, 鳴海 覚志, 梅木 郁美, 箱田 明子, 菅野 潤子, 長谷川 奉延, 藤原 幾磨

日本小児科学会雑誌 （(公社)日本小児科学会）  118 （ 5 ） 875 - 875 2014年05月

ISSN  0001-6543

4世代・世界最大のPAX8変異陽性家系からみえた甲状腺表現型の多様性 高齢PAX8変異保有者の臨床像

西岡 淳子, 阿部 清美, 鳴海 覚志, 長谷川 奉延, 八ツ賀 秀一, 佐々木 孝子, 片山 幸樹, 久野 建夫, 古賀 靖敏

日本内分泌学会雑誌 （(一社)日本内分泌学会）  90 （ 1 ） 296 - 296 2014年04月

ISSN  0029-0661

IGSF1変異による中枢性先天性甲状腺機能低下症の3男児例

阿部 清美, 鳴海 覚志, 杉澤 千穂, 室谷 浩二, 朝倉 由美, 安達 昌功, 沼倉 周彦, 長谷川 奉延

日本内分泌学会雑誌 （(一社)日本内分泌学会）  90 （ 1 ） 271 - 271 2014年04月

ISSN  0029-0661

Identification and functional characterization of two novel NPR2 mutations in Japanese patients with short stature.

Amano N, Mukai T, Ito Y, Narumi S, Tanaka T, Yokoya S, Ogata T, Hasegawa T

The Journal of clinical endocrinology and metabolism （ENDOCRINE SOC）  99 （ 4 ） E713 - 8 2014年04月

査読有り,  ISSN  0021-972X

　概要を見る

Context: C-type natriuretic peptide-natriuretic peptide receptor B (NPR-B) signaling is critical for endochondral ossification, which is responsible for longitudinal growth in limbs and vertebrae. Biallelic NPR2 mutations cause acromesomelic dysplasia, type Maroteaux, which is bone dysplasia characterized by severe short stature and short limbs. A monoallelic NPR2 mutation has been suggested to mildly impair long bone growth.
Objective: The goal of this study was to identify and characterize NPR2 mutations among Japanese patients with short stature.
Subjects and Methods: We enrolled 101 unrelated Japanese patients with short stature. NPR2 and NPPC were sequenced, and the identified variants were characterized in vitro.
Results: In two subjects, we identified two novel heterozygous NPR2 mutations (R110C and Q417E) causing a loss of C-type natriuretic peptide-dependent cGMP generation capacities and having dominant-negative effects. R110C was defective in trafficking from the endoplasmic reticulum to the Golgi apparatus. In contrast, Q417E showed clear cell surface expression.
Conclusions: We identified heterozygous NPR2 mutations in 2% of Japanese patients with short stature. Our in vitro findings indicate that NPR2 mutations have a dominant negative effect, and their dominant-negative mechanisms vary corresponding to the molecular pathogenesis of the mutations.

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A novel KAL1 mutation is associated with combined pituitary hormone deficiency.

Takagi M, Narumi S, Hamada R, Hasegawa Y, Hasegawa T

Human genome variation （Nature Publishing Group）  1   14011 2014年

ISSN  2054-345X

　概要を見る

Using a next-generation sequencing strategy, we identified a novel KAL1 missense mutation (p.His568Gln) in a patient with combined pituitary hormone deficiency, right microphthalmia, right renal aplasia and severe developmental delay. Our findings will provide additional evidence that KAL1 mutations are associated with hypopituitarism, in addition to luteinizing hormone, and follicle-stimulating hormone deficiencies, and improve our understanding of the phenotypic features and developmental course associated with KAL1 mutations.

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A novel mutation in SOX2 causes hypogonadotropic hypogonadism with mild ocular malformation.

Takagi M, Narumi S, Asakura Y, Muroya K, Hasegawa Y, Adachi M, Hasegawa T

Hormone research in paediatrics （KARGER）  81 （ 2 ） 133 - 8 2014年

査読有り,  ISSN  1663-2818

　概要を見る

Background: Heterozygous SOX2 mutations have been reported to cause isolated hypogonadotropic hypogonadism (HH) in addition to ocular and brain abnormalities. Objective: We report a novel missense SOX2 (Y110C) mutation in an HH patient with mild ocular malformation. Patients: The 20-year-old male was referred because of typical signs of complete hypogonadism, with small intrascrotal testes (2 ml), no pubic hair (P1), and a micropenis. Hormone assays revealed very low plasma testosterone levels and very low levels of plasma gonadotropin. He was found to have retinal detachment in his right eye and surgery was performed at the age of 14 years. Results: Using a next-generation sequencing strategy, we identified a novel heterozygous SOX2 mutation, c. 329A>G (p.Y110C). Y110C SOX2 had reduced transactivation and no dominant negative effect. Subcellular localization revealed no significant difference between wild-type and mutant SOX2. EMSA experiments showed that the Y110C SOX2 abrogated DNA-binding ability. Conclusion: The Y110C mutation affects a critical residue in the SOX2 protein. This study extends our understanding of the phenotypic features, molecular mechanism, and developmental course associated with mutations in SOX2. When multiple genes need to be analyzed for mutations simultaneously, targeted sequence analysis of interesting genomic regions is an attractive approach. (C) 2014 S. Karger AG, Basel

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新生児マス・スクリーニング検査でみつかったTPO異常症の2例

石津 桂, 森川 俊太郎, 鳴海 覚志, 長谷川 奉延, 田島 敏広

日本内分泌学会雑誌 （(一社)日本内分泌学会）  89 （ 3 ） 977 - 977 2013年12月

ISSN  0029-0661

IGSF1遺伝子変異による中枢性甲状腺機能低下症の2家系 家族検索とPRL分泌能

松浦 信夫, 橘田 一輝, 阿部 清美, 小池 明美, 椿 淳子, 鳴海 覚志, 長谷川 奉延, 横田 行史, 大津 成之, 田久保 憲行, 柴山 啓子, 大山 宜秀

日本内分泌学会雑誌 （(一社)日本内分泌学会）  89 （ 2 ） 463 - 463 2013年09月

ISSN  0029-0661

PAX8変異陽性甲状腺機能低下症の世界最大家系 甲状腺表現型の多様性

阿部 清美, 鳴海 覚志, 八ツ賀 秀一, 西岡 淳子, 久野 建夫, 長谷川 奉延

日本内分泌学会雑誌 （(一社)日本内分泌学会）  89 （ 2 ） 474 - 474 2013年09月

ISSN  0029-0661

A report of two novel NR5A1 mutation families: possible clinical phenotype of psychiatric symptoms of anxiety and/or depression.

Suwanai AS, Ishii T, Haruna H, Yamataka A, Narumi S, Fukuzawa R, Ogata T, Hasegawa T

Clinical endocrinology （WILEY-BLACKWELL）  78 （ 6 ） 957 - 65 2013年06月

査読有り,  ISSN  0300-0664

　概要を見る

Objective NR5A1 or steroidogenic factor 1 is a nuclear receptor that plays important roles in the hypothalamuspituitarysteroidogenic axis. The clinical phenotype of most 46,XY mutation carriers includes disorders of sex development (DSD) without adrenal insufficiency, whereas 46,XX mutation carriers have phenotypes ranging from no symptoms to ovarian insufficiency. Although genetically engineered ventromedial hypothalamus-specific Nr5a1 knockout mice show anxiety behaviour, no psychiatric symptoms have been reported in human NR5A1 mutation carriers. We report clinical and molecular findings for individuals (from two families) with NR5A1 mutations, showing psychiatric symptoms. Design and methods We screened for NR5A1 mutations in a cohort of 34 patients with 46,XY DSD using PCR-based sequencing. Psychiatric symptoms were assessed using mental health assessment tools and structured clinical interviews. Functional properties of detected mutant NR5A1s were studied in silico and in vitro, including three-dimensional (3D) mutation models, subcellular NR5A1 protein localization and transactivation assays. Results We found 2 (46,XY) patients with NR5A1 heterozygous novel mutations (p.D257fs and p.V424del), which were transmitted from their respective mothers. The patients' clinical findings indicated DSD without adrenal insufficiency. Both mothers showed psychiatric symptoms, including excessive anxiety and/or depression. The mother and grandmother of one patient had premature ovarian insufficiency. Functional studies showed altered 3D models of p.V424del and normal subcellular NR5A1 localization and impaired transcriptional activation without dominant-negative effects in both mutations. Conclusions We found 2 (46,XX) NR5A1 mutation carriers with excessive anxiety and/or depression. These results suggest that excessive anxiety and/or depression are possible clinical phenotypes of 46,XX NR5A1 mutations.

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Congenital hypothyroidism caused by a novel mutation of the dual oxidase 2 (DUOX2) gene

Atsuko Yoshizawa-Ogasawara, Sayaka Ogikubo, Mari Satoh, Satoshi Narumi, Tomonobu Hasegawa

JOURNAL OF PEDIATRIC ENDOCRINOLOGY & METABOLISM （WALTER DE GRUYTER GMBH）  26 （ 1-2 ） 45 - 52 2013年02月

査読有り,  ISSN  0334-018X

　概要を見る

The dual oxidase 2 (DUOX2) mutation results in an impairment of the hydrogen peroxidase-generating system and is identified as a dyshormonogenic cause of congenital hypothyroidism (CH). Here, we describe two unrelated Japanese girls with CH due to a novel DUOX2 mutation. They had high serum thyrotropin levels and low free thyroxine/thyroxine concentrations during the neonatal period. A novel missense mutation with a transversion of G to A at position 1462 in exon 12 of the DUOX2 gene that caused a replacement of glycine (G) with arginine (R) at codon 488 of the protein (c.1462G>A, p.[G488R]) was identified. One patient was a compound heterozygote forp. [L479SfsX3] + [G488R]. The other was homozygous for p.[G488R]. This p. G488R substitution occurred in a highly conserved glycine residue of the mammalian DUOX2 protein. The two patients had different haplotypes, suggesting that the p. G488R alleles were the result of independent, recurrent mutations. Later in life, both patients were still euthyroid even after discontinuing thyroid hormone therapy. We conclude that this p. G488R missense mutation in the DUOX2 gene of the patients is associated with thyroid dysfunction that presents during the neonatal period.

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Delayed onset congenital hypothyroidism in a patient with DUOX2 mutations and maternal iodine excess

Toshihiko Kasahara, Satoshi Narumi, Keisuke Okasora, Ryuzo Takaya, Hiroshi Tamai, Tomonobu Hasegawa

AMERICAN JOURNAL OF MEDICAL GENETICS PART A （WILEY）  161A （ 1 ） 214 - 217 2013年01月

査読有り,  ISSN  1552-4825

　概要を見る

Congenital hypothyroidism (CH), one of the most common congenital endocrine disorders, causes irreversible intellectual disability in untreated patients. Today, the vast majority of patients receive early diagnosis and treatment in the context of newborn screening for CH, and achieve satisfactory cognitive development. However, a subset of patients with delayed onset are undetectable by newborn screening, and miss benefit from early intervention. Here, we report on a delayed-onset CH patient that had two contributing factors in the pathogenesis of CH simultaneously, i.e., a genetic defect and iodine excess. The patient was exposed to excessive iodine in utero because her mother consumed massive amounts of seaweed during pregnancy. Surprisingly, the patient had a negative result in newborn screening, but developed overt CH at age 3 months. She received thyroxine supplementation until when normalization of the thyroid function was confirmed at age 3 years (i.e., transient CH). Mutation screening for DUOX2, a causative gene for transient CH, showed biallelic mutations (p.[E327X]?+?[H678R]). This report provides a new example of environmental modification of phenotypes of CH due to a genetic defect, which can potentially distort screening results. (c) 2012 Wiley Periodicals, Inc.

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A study of the etiology of transient congenital hypothyroidism in Niigata Prefecture, Japan.

Nagasaki K, Narumi S, Abe K, Asami T, Sato H, Ogawa Y, Kikuchi T, Hasegawa T, Saitoh A

International journal of pediatric endocrinology 2013年

査読有り

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Quantitative and sensitive detection of GNAS mutations causing mccune-albright syndrome with next generation sequencing.

Narumi S, Matsuo K, Ishii T, Tanahashi Y, Hasegawa T

PloS one （PUBLIC LIBRARY SCIENCE）  8 （ 3 ） e60525 2013年

査読有り,  ISSN  1932-6203

　概要を見る

Somatic activating GNAS mutations cause McCune-Albright syndrome (MAS). Owing to low mutation abundance, mutant-specific enrichment procedures, such as the peptide nucleic acid (PNA) method, are required to detect mutations in peripheral blood. Next generation sequencing (NGS) can analyze millions of PCR amplicons independently, thus it is expected to detect low-abundance GNAS mutations quantitatively. In the present study, we aimed to develop an NGS-based method to detect low-abundance somatic GNAS mutations. PCR amplicons encompassing exons 8 and 9 of GNAS, in which most activating mutations occur, were sequenced on the MiSeq instrument. As expected, our NGS-based method could sequence the GNAS locus with very high read depth (approximately 100,000) and low error rate. A serial dilution study with use of cloned mutant and wildtype DNA samples showed a linear correlation between dilution and measured mutation abundance, indicating the reliability of quantification of the mutation. Using the serially diluted samples, the detection limits of three mutation detection methods (the PNA method, NGS, and combinatory use of PNA and NGS [PNA-NGS]) were determined. The lowest detectable mutation abundance was 1% for the PNA method, 0.03% for NGS and 0.01% for PNA-NGS. Finally, we analyzed 16 MAS patient-derived leukocytic DNA samples with the three methods, and compared the mutation detection rate of them. Mutation detection rate of the PNA method, NGS and PNA-NGS in 16 patient-derived peripheral blood samples were 56%, 63% and 75%, respectively. In conclusion, NGS can detect somatic activating GNAS mutations quantitatively and sensitively from peripheral blood samples. At present, the PNA-NGS method is likely the most sensitive method to detect low-abundance GNAS mutation.

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Functional characterization of four novel PAX8 mutations causing congenital hypothyroidism: new evidence for haploinsufficiency as a disease mechanism.

Narumi S, Araki S, Hori N, Muroya K, Yamamoto Y, Asakura Y, Adachi M, Hasegawa T

European journal of endocrinology （BIOSCIENTIFICA LTD）  167 （ 5 ） 625 - 32 2012年11月

査読有り,  ISSN  0804-4643

　概要を見る

Background: Individuals carrying a heterozygous inactivating PAX8 mutation are affected by congenital hypothyroidism (CH), although heterozygous Pax8 knockout mice are not. It has remained unclear whether CH in PAX8 mutation carriers is caused by haploinsufficiency or a dominant negative mechanism.
Objective: To report clinical and molecular findings of four novel PAX8 mutations, including one early-truncating frameshift mutation.
Subjects and methods: Four probands were CH patients. Two had family history of congenital or childhood hypothyroidism. Three probands were diagnosed in the frame of newborn screening for CH, while one had a negative result in screening but was diagnosed subsequently. Three had thyroid hypoplasia and one had a slightly small thyroid with low echogenicity. For these probands and their family members, we sequenced PAX8 using a standard PCR-based method. Pathogenicity of identified mutations was verified in vitro.
Results: We found four novel heterozygous PAX8 mutations in the four probands: L16P, F20S, D46SfsX24, and R133Q. Family studies showed four additional mutation carriers, who were confirmed to have high serum TSH levels. Expression experiments revealed that three mutations (L16P, F20S, and R133Q) had defects in target DNA binding, while D46fs had protein instability that was rescued by the proteasome inhibitor MG132. All four mutations had reduced transactivation on the thyroglobulin promoter, supporting that they were inactivating mutations.
Conclusion: D46fs is the first PAX8 mutation with confirmed protein instability. Our clinical and in vitro findings together suggest that pure PAX8 haploinsufficiency can cause CH in humans.

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両アレル性DUOX2変異例におけるヨード有機化能の検討

長崎 啓祐, 鳴海 覚志, 阿部 清美, 佐藤 英利, 小川 洋平, 浅見 直, 長谷川 奉延, 菊池 透

日本内分泌学会雑誌 （(一社)日本内分泌学会）  88 （ 2 ） 497 - 497 2012年09月

ISSN  0029-0661