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

关于实体瘤固有的核型异质性，我们通过多参数分析评估了同一核上17号染色体的显示数目和核DNA含量，以显示在细胞荧光计量学上不同亚群之间17号染色体数目的变化。在本项关于结直肠癌的研究中，我们利用计算机控制的自动扫描阶段，在同一核上依次收集了这两个值。1)我们调查了8个手术切除的结直肠癌病变，DNA定量分析将其归类为非整倍体，并显示17个异倍体核增加。这些观察表明，17号染色体数目的分布反映了基因组内容的内重复，但并不随细胞周期的不同而变化。2)我们于1996年引入了原位聚合酶链式反应的热循环仪。从样品制备、引物选择、条件设置、信号标记方法等方面对原位聚合酶链式反应的条件进行了评价。3)同时测定细胞核DNA含量是必要的。虽然需要在同一个细胞核上同时检测核DNA含量和特定染色体的数目，但由于FISH程序中所需的变性过程阻碍了对核DNA含量的正确评估，所以我们通常不能获得成功的结果。4)原位PCR方法应用于结直肠癌等实体肿瘤仍然存在问题。

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.