IMPROVED DIAGNOSIS OF TUMORS IN COMPANION ANIMALS USING PCR METHOD

研究了erbBl、B2、B3、B4和c-kit原癌基因在犬和猫乳腺肿瘤标本中的表达，以评估其作为肿瘤标志物的潜在价值。通过比较几种哺乳动物cDNA序列的同源性，合成了用于逆转录-聚合酶链反应（RT-PCR）方法的引物。在11种乳腺肿瘤（腺癌、良恶性混合肿瘤）中检测到c-kit转录本的表达。在除肥大细胞瘤外的其他组织的12例肿瘤中，只有5例样品检测到低水平的RT-PCR产物，7例肿瘤未观察到明显扩增。erbB家族在正常乳腺组织中不表达，而在肿瘤组织中多次表达。在三种猫乳腺肿瘤中，也观察到erbB基因的频繁和多重表达。虽然在两个样本中检测到c-kit转录本，但在观察到所有erbB基因表达的一个样本中不明显。这些结果表明，erbB家族和c-kit转录本的表达是犬和猫乳腺肿瘤诊断的有用标记物。对猫乳腺腺癌细胞系的染色体分析表明，FMT-3和FMT4细胞在特定的标记染色体中显示出特定染色体的单体或完全等位基因缺失。在检查的所有猫肿瘤中均未检测到禁止素的RT-PCR产物。用荧光原位杂交法在猫B4染色体长臂上定位了禁止素基因。由于两种细胞系的核型均为B4三体，因此对该基因在FMT-3细胞中的染色体定位进行了研究。一对B4在细胞中显示出明显的双线杂交信号。

The expression of erbBl, B2, B3, B4 and c-kit proto-oncogene in canine and feline mammary tumor specimens was investigated to evaluate its potential usefulness as tumor markers. By comparing the homology among the nucleotide sequences of cDNA reported for several mammalian species, pairs of primers were synthesized for the reverse transcriptase-polymerase chain reaction (RT-PCR) method. The expression of c-kit transcript was detected in 11 mammary tumors (adenocarcinomas, benign and malignant mixed tumors). Of the 12 tumors originated from various tissues except mastocytomas, only low level of RT-PCR products iwre detected in 5 samples, whereas no apparent amplification was observed in 7 tumors. Any of erbB family was not expressed in normal mammary tissue, while the multiple expression was frequently observed in tumor samples. In three feline mammAry tumors, the frequent and multiple expression of erbB genes was also observed. Although c-kit transcript was detected in two samples, it was not apparent in one specimen in which the expressions of all of erbB genes were observed. These results indicate that the expression of erbB family and c-kit transcript is useful markers for the diagnosis of both canine and feline mammary tumors.The chromosome analysis of cell lines derived from feline mammary adenocarcinoma demonstrated that FMT-3 and FMT4 cells showed monosomy or complete alleic deletion of particular chromosomes which were included in specific marker chromosomes. In none of the feline tumors examined, RT-PCR product of prohibitin was detected. Feline prohibitin gene was mapped at the long arm of chromosome B4 by fluorescence in situ hybridization method. Because the karyotypes of both cell lines showed B4 trisomy, the chromosomal localization of the gene in FMT-3 cells was investigated. A couple of B4 in the cells was shown to have apparent doublet signals of hybridization.