Studies of genetic altarations in pcecancerous and background lesions by tissue microdissection.

通过组织显微切割研究原癌和背景病变的遗传变异。

• 批准号: 08670233
• 负责人: NOGUCHI Masayuki
• 金额: $ 1.41万
• 依托单位: University of Tsukuba
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 1996
• 资助国家: 日本
• 起止时间: 1996 至 1997
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-08670233/
• 关键词: tissue microdissection; PCR-LOH; AP-PCR

First, the quality and quantity of the DNA extracted from formalin fixxed and microdissected tissue fragments were examined. Two to three ng DNA could be extracted from 200 to 400 cells which were scrached from 3 mum-thick formalin fixed materials. Theoretically, one nucleus is thought to contain 12 to 15 pg DNA,this estimation are consistent with the former results. Templates extracted from 10 to 20 cells could be successfully amplified by polymerase chain reaction.Using extracted DNAs by this tissue microdissection method, the following analyzes were done :(1) Small sized pulmonary adenocarcinomas, including 40 of early stage adenocarcinomas and 30 of early advanced adenocarcinomas were examined for loss of heterozygosity (LOH) on 2p, 3p, 9p, 17p, 17q using microsatellite markers. The frequencies of LOH were 19.8% in in early stage adenocar cinomas and 26.8% in early advanced adenocarcinomas. These results indicated that multiple allelic loss are one of the characteristic abnormalities of very early stage of pulmonary adenocarcinogenesis.(2) DNA fingerprints qenerated by a single arbitrary primer were compared between normal and tumor tissues of the same individuals which were fixed with methanol and microdissected. Loss of sequence of chromosome 7 was detected in 41.7% of adenocarcinomas and that of chromosome 22 in 84.6% of small cell carcinomas. Gains of sequences in chromosome 1,8,13 were detected in over 40% of adenocarcinomas and chromosome 2 in 63.3% of squamous cell carcinomas. LOH of chromosome 22 in small cell carcinomas were confirmed by microsatellite PCR analysis and suggested that LOH on chromosome 22q13.3 is very frequent event in small cell carcinoma.

首先，检测了从福尔马林固定和显微解剖的组织片段中提取的DNA的质量和数量。从3个厚的福尔马林固定材料上刮下的200~400个细胞中可提取2~3 ng DNA。理论上认为一个核含有12-15pg的DNA，这一估计与以前的结果是一致的。(1)采用微卫星技术对40例早期肺腺癌和30例早期晚期肺腺癌的2p、3p、9p、17p、17q基因进行杂合性缺失(LOH)检测。LOH频率在早期腺癌为19.8%，在早期晚期腺癌为26.8%。这些结果表明，多重等位基因缺失是肺腺癌发生的早期特征之一。(2)比较了相同个体的正常组织和肿瘤组织经甲醇固定后的DNA指纹图谱。41.7%的腺癌存在7号染色体序列缺失，84.6%的小细胞癌存在22号染色体序列缺失。在40%以上的腺癌和63.3%的鳞癌中，分别检测到1、8、13号染色体序列的扩增。微卫星聚合酶链式反应证实了小细胞癌中22号染色体的杂合性缺失，提示22号染色体q13.3的杂合性缺失在小细胞癌中是非常常见的事件。

项目成果

Cho JH.Noguchi, M. et al.: "Loss of heterozygosity of multiple tumor suppressor genes in human gastric cancers by polymerase chain reaction." Lab Invest. 74. 835-841 (1996)

Cho JH.Noguchi, M. 等人：“通过聚合酶链反应导致人类胃癌中多个肿瘤抑制基因杂合性的丧失。”

Sekine, I.Noguchi, M. et al.: "Roentgenographically occult small cell lung Cancer. Case report and reviah-of the literature." Mayo Clin Proc. 71. 481-484 (1996)

Sekine, I.Noguchi, M. 等人：“X 线造影隐匿性小细胞肺癌。病例报告和文献回顾。”

Muta H,Noguchi M,et al.: "Clinical implications of microsatellite instabillity in colorectal cancer." Cancer. 77. 265-270 (1996)

Muta H、Noguchi M 等人：“结直肠癌中微卫星不稳定性的临床意义。”

Noguchi M,et al.: "Modified formalin and methanol fixation methods for molecular biogical and morphological analyzes." Pathol Int. 47. 685-691 (1997)

Noguchi M 等人：“用于分子生物学和形态学分析的改良福尔马林和甲醇固定方法。”

Yoshioka H,Noguchi M,et al.: "Analysis of loss of heterozygosity in small adenocarcinomas of the lung." Jpn J Clin Oncol. (in press).

Yoshioka H、Noguchi M 等人：“肺小腺癌杂合性丢失的分析”。