臨床検査49巻1号

2005年01月発行

今月の主題 ミトコンドリア病 ミトコンドリア病の臨床検査

ミトコンドリア遺伝子A3243G変異の高感度検出法―Peptide Nucleic Acid-assisted allele-specific Polymerase Chain Reaction法

著者： 浦田美秩代¹ 栢森裕三² 濱﨑直孝³

所属機関： ¹九州大学病院検査部遺伝子検査室 ²九州大学病院検査部 ³九州大学大学院医学研究院臨床分子医学

ページ範囲：P.66 - P.70

〔SUMMARY〕　ミトコンドリア遺伝子変異のなかで，MELASや糖尿病の原因として知られるA3243G変異の検出は診断的な価値が大きい．A3243G変異は同一細胞内に正常型と異常型が共存しており，しかも，検査材料として使用される末梢血白血球ではその変異率が低いために，測定法としては高感度な方法が望まれている．本稿では，2004年にわれわれが開発したA3243G変異の簡便で高感度な定量法を紹介する．〔臨床検査　49：66-70，2005〕

参考文献

１) Umeda N, Suzuki T, Yukawa M, et al：Mitochondria-specific RNA-modifying enzymes responsible for the biosynthesis of the wobble base in mitochondrial tRNAs；implications for the molecular pathogenesis of human mitochondrial diseases. J Biol Chem　2004 (in Press)

２) van den Ouweland JM, Lemkes HHJ, RuitenbeekW, et al：Mutation in mitochondrial tRNALew(UUR) gene in a large pedigree with maternally transmitted type II diabetes mellitus and deafness. Nat Genet　1：368-371, 1992

３) Kadowaki T, Kadowaki H, Mori Y, et al：A subtype of diabetes mellitus associated with a mutation of mitochondrial DNA. N Engl J Med　330：962-968, 1994

４) Katagiri H, Asano T, Ishihara H, et al：Mitochondrial diabetes mellitus：prevalence and clinical characterization of diabetes due to mitochondrial tRNALew(UUR) gene mutation in Japanese patients. Diabetetologia　37：504-510, 1994

５) Tokunaga M, Mita S, Sakuta R, et al：Increased mitochondrial DNA in blood vessels and ragged-red fibers in mitochondrial myopathy, enchephalopathy, lactic acidosis, and stroke-like episodes (MELAS). Ann Neurol　33：275-280, 1993

６) Ohkubo K, Yamano A, Nagashima M, et al：Mitochondrial gene mutation in the tRNALew(UUR) region and diabetes；prevalence and clonical phenotypes in Japan. Clin Chem　47：1641-1648, 2001

７) Gerbitz K-D, van den Ouweland JMW, Maassen JA, et al：Mitochondrial diabetes；a review. Biochim Biophys Acta　1271：253-260, 1995

８) Sue CM, Quigley A, Katsabanis S, et al：Detection of MELAS A3243G point mutation in muscle, blood and hair follicles. J Neurol Sci　161：36-39, 1998

９) Smith ML, Hua X-Y, Marsden DL, et al：Diabetes and mitochondrial encephalomyopathy with lactic acidosis and stroke-like episodes(MELAS): radiolabeled polymerase chain reaction is necessary accurate detection of low percentages of mutation. J Clin Endocrinol Metab　82：2816-2831, 1997

10) Suzuki Y, Goto Y, Taniyama M, et al：Muscle histopathology in diabetes mellitus associated with mitochondrial tRNALeu(UUR) mutation at position 3243. J Neurol Sci　145：49-53, 1997

11) Suzuki Y, Nishimaki K, Taniyama M, et al：Lipoma and opthalmoplegia in mitochondrial diabetes associated with small heteroplasmy level of 3243 tRNA(Leu(UUR)) mutation. Diabetes Res Clin Pract　63：225-229, 2004

12) Urata M, Wakiyama M, Iwase M, et al：New sensitive method for the detection of the A3243G mutation of human mitochondrial deoxyribonucleic acid in diabetes mellitus patients by ligation-mediated polymerase chain reaction. Clin Chem　44：2088-2093, 1998

13) Murdock DG, Christacos NC, Wallace DC：The age-related accumulation of a mitochondrial control region mutation in muscle, but not brain, detected by a sensitive PNA-directed PCR clamping based method. Nucleic Acids Res　28：4350-4355, 2000

14) Urata M, Wada Y, Kim S, et al：High-sensitivity detection of the A3243G mutation of mitochondrial DNA by acombination of allele-specific PCR and PNA-directed PCR clamping. Clin Chem　50：2045-2051, 2004

15) Hancock DK, Schwarz FP, Song F, et al：Design and use of a peptide nucleic acid for detection of the heteroplasmic low-frequency mitochondrial encephalomyopathy, lactic acidosis, and stroke-like episodes (MELAS) mutation in human mitochondrial DNA. Clin Chem　48：2155-2163, 2002

16) Yoneda M, Miyatake T, Attardi G：Complementation of mutant and wild-type human mitochondrial DNAs coexisting since the mutation event and lack of complementation of DNAs introduced separately into a cell within distinct organelles. Mol Cell Biol　14：2699-2712, 1994