ANALYSIS OF CHROMOSOMAL AND CANCER RELATED GENE ABNORMALITIES IN ATYPICAL EPITHELIUM IN THE RESPIRATORY SYSTEM

使用经支气管活检（TBB）标本进行分子生物学分析，以证实以下假设；(1)在呼吸系统如支气管、肺泡上皮中，染色体及癌相关基因异常的发生与上皮异型性程度一致。(2)在癌症患者中发现的上皮异型性在分子生物学上不同于在非癌症患者中发现的。根据细胞异型程度将TBB标本分为增生、化生和不典型增生。显微解剖法分离非典型上皮后，提取DNA进行分析。评估3pl4.2 （FHIT）、3p21.3、8p22、9p22、18q21.1的杂合性损失（LOH）。此外，比较p53、PCNA和p21的表达与细胞凋亡程度的关系。3p21.3和9p22的LOH发生在肺癌发生的早期，与细胞异型性程度显著相关。8p22和18q21.1位点的LOH多见于肺癌发生的中晚期。3pl4.2 （FHIT）的LOH与细胞异型性程度无关，在吸烟者中选择性发现。有趣的是，这些LOH常见于肺癌和特发性肺纤维化（IPF）患者，而不常见于其他良性肺部疾病患者。结果提示，IPF患者肺癌的频繁发生，至少部分反映了IPF患者非典型上皮细胞染色体异常的频繁发生。先前，我们报道上皮非典型上皮经常表达野生型p53蛋白（Wakamatsu et al ., Am J respiratory Cell Mol Biol, 1999）。在本研究中，我们发现p53表达与PNCA表达显著相关，但与p21表达和细胞凋亡无关。从体外实验中，我们发现吸烟相关的化学致癌物如苯并[a]芘诱导p53蛋白的表达，但在其表达后立即泛素化p21蛋白，导致细胞周期阻滞。这一结果阐明了呼吸系统非典型上皮中（野生型）p53表达与p21表达无关的原因。这些结果表明，呼吸系统非典型上皮中存在多种染色体异常，并且肺癌和IPF患者的细胞异型中染色体异常的频率比其他良性疾病患者更常见。少

Molecular biological analysis was performed using specimens derived from transbronchial biopsy (TBB) to confirm the following hypotheses;(1)Chromosomal and cancer related gene abnormalities occur in concordance with the degree of epithelial atypia in respiratory system such as bronchial and alveolar epithelium.(2)Epithelial atypia found in cancer patients is molecularbiologically different from that found in non-cancerous patients.TBB specimens were classified according to the degree of their cellular atypia into hyperplasia, metaplasia, and dysplasia. After isolation of atypical epithelia by microdissection method, DNA was extracted and analyzed. Loss of heterozygosity (LOH) in 3pl4.2 (FHIT), 3p21.3, 8p22, 9p22, 18q21.1 were evaluated. In addition, the expression of p53, PCNA and p21 was compared to the degree of apoptosis.LOH in 3p21.3 and 9p22 occurred in early stage of lung carcinogenesis and was significantly related to the degree of cellular atypia. LOH in 8p22 and 18q21.1 occurr … More ed in middle or late stage of lung carcinogenesis. LOH of 3pl4.2 (FHIT) was not related to the degree of cellular atypia, and found selectively in smokers. Interestingly, these LOH were frequently found in patients with lung cancer and with idiopathic pulmonary fibrosis (IPF), and infrequently found in patients with other benign lung diseases. The results suggest that the frequent occurrence of lung cancer in patients with IPF, at least in part, reflects the frequent occurrence of chromosomal abnormalities in atypical epithelium in IPF.Previously, we reported that epithelial atypical epithelium frequently express wild-type p53 protein (Wakamatsu et al, Am J Respir Cell Mol Biol, 1999). In the present study, we found that p53 expression significantly related to PNCA expression, but not to p21 expression or to apoptosis. From the in vitro experiments, we showed that smoking related chemical carcinogens such as benzo[a]pyrene induce the expression of p53 protein, but that ubiquitinate p21 protein immediately after its expression resulting in the blockade of cell cycle arrest. This result elucidate the reason why the expression of (wild-type) p53 is not related to the expression of p21 in atypical epithelium in the respiratory system.These results indicated that various chromosomal abnormalities are found in atypical epithelium in the respiratory system, and that the frequency of chromosomal abnormalities are more common in cellular atypia found in patients with lung cancer and IPF than those with other benign diseases. Less