The trial for chromosomal distribution analysis of in situ PCR and FISH signals as an index of malignamcy in gastro-enterological carcinomas

关于实体瘤的固有核型异质性，我们通过多参数分析评估同一细胞核上17号染色体的显示数目和核DNA含量，以证明细胞荧光测定不同亚群之间17号染色体数目的改变。我们通过使用计算机控制的自动扫描阶段在同一核上连续收集这两个值。1）我们研究了手术切除的结肠直肠癌的8个病变，其在定量DNA分析中被归类为非整倍体，并且还表现出17-异倍体核的增加。这些观察结果表明，17号染色体数目的分布反映了基因组内容的核内复制，但它并不随细胞周期的时相而改变。2）我们于1996年引进了原位PCR热循环仪。对原位PCR方法的样品制备、引物选择、条件设置、信号标记方法等条件进行了评价。3）为了评价一种必要的细胞遗传学电荷，有必要同时评价细胞核DNA含量。虽然核DNA含量和特定染色体的数目应在同一个核上同时进行评估，但通常不能获得成功的结果，因为FISH过程中所需的变性过程阻碍了核DNA含量的正确评估。4）我们仍然存在关于原位PCR方法应用于实体肿瘤如结直肠癌的问题。

Concerning inherent karyotypic heterogeneities of solid tumor, we evaluated both the exhibiting number of chromosome 17 and nuclear DNA content on an identical nucleus by multi-parametric analysis in order to demonstrate the alteration in number of chromosome 17 among cytofluorometrically distinct subpopulations.In the present study concerning colorectal carcinoma, we have collected both of these values sequentially on an identical nucleus by using computer controlled auto-scanning stage.1) We investigated 8 lesions of surgically resected colorectal carcinomas, which were classified as aneuploid in quantitative DNA analysis and also exhibited an increase of 17-aneusomy nuclei. These observations indicate taht the distribution of number of chromosome 17 reflects an endoreduplication of genome content, yet, it does not alter in accordance with the phase of cell cycle.2) We introduced the thermal cycler for in situ PCR in 1996. the conditions of in situ PCR method were evaluated such as sample preparations primer selection, condition setting and labeling mmethod of signal.3) It is necessary to evaluate nuclear DNA content simultaneously in order to assess an essential cytogenetic charge. Althoug both nuclear DNA content and number of a particular chromosome should be evaluated simultaneously on an identical nucleus, we usually could not obtain a successful result, because the denaturation process, which is required in FISH procedure, hinders correct evaluation of nuclear DNA content.4) We still have the problem concerning in situ PCR method applying solid tumor such as colorectal carcinomas.