Growth Mechanism of Esophageal Carcinoma Using Protein Free Culture (Relation of Oncogenes and Cell Growth)

描述了癌基因和抑制基因与细胞培养的关系。首先，通过与c-myc、c-erbB、hst-1、k-ras、b-FGF、CyclinD1的Southern和Northern杂交，分析了23株人食管鳞状细胞癌（ESC）组织的基因扩增。除了CyclinD1表达在无蛋白环境中有轻微的细胞系建立潜力外，这些基因没有显示出建立细胞系的潜力。其次，我们进行等位基因分析，探讨肿瘤抑制基因是否参与食管癌的发生。在信息性病例中，3p、5q、6p、8p、9p、9q、11p、13q、17p、17q、18q和19q染色体上的杂合性缺失发生率高达30%。其中，5q、13q、17p和18q上的LOH被认为涉及APC、RB、p53和DCC。然而，18q染色体的缺失分析显示，通常丢失的区域不包括DCC位点。对60株原代ESC细胞和20株培养ESC细胞株进行APC基因突变筛选，结果表明，尽管5q染色体上LOH频率较高，但APC基因未发生突变。第三，我们通过所有编码外显子和非内含子连接筛选29个人ESC细胞系p53基因突变。22个细胞系（76%）发现突变。我们还检查了65个新鲜的肿瘤样本，我们试图从所有这些样本中建立细胞系，并在26个样本（40%）中检测到突变。新鲜肿瘤中p53基因的表达与细胞系的建立无明显相关性。我们得出结论，p53突变在建立培养的人类ESC细胞系中并不重要，但在体外提供了生长优势。在无蛋白环境下，p53突变与细胞生长也没有关系。我们还研究了ESC中p53突变、小鼠双分钟2 （MDM2）扩增和人乳头瘤病毒（HPV）感染之间的关系。在大多数肿瘤中，MDM2基因扩增或HPV感染与p53突变无关。在细胞培养中，MDM2基因扩增只与细胞培养有显著关系。在患者预后方面，CyclinD1基因扩增与患者预后不良显著相关，我们还发现p53突变和/或MDM2扩增组患者的生存期明显缩短。我们目前正在研究侵袭性和细胞膜流动性与无蛋白培养的关系。少

The relation of oncogenes and suppressor genes to cell culture was described.First, gene amplification was analyzed in 23 cell lines derived from human esophageal squamous cell carcinoma (ESC) tissues by Southern and Northern hybridizations to c-myc, c-erbB,hst-1, k-ras, b-FGF,CyclinD1. These genes showed no potential for establishing cell lines, with the exception of CyclinD1 expression which demonstated a slight potential to establish cell lines in a protein free environment.Secondly, we performed allelotype analysis to investigate whether tumor suppressor genes are involved in esophageal cancer. Frequent loss of heterozygosity (LOH) of 30%>of the informative cases was observed on chromosomes 3p, 5q, 6p, 8p, 9p, 9q, 11p, 13q, 17p, 17q, 18q, and 19q. Among these, LOH on 5q, 13q, 17p, and 18q was considered to involve the APC,RB,p53, and DCC.However, deletion analysis of chromosome 18q revealed that the region commonly lost did not include the DCC locus. Screening of 60 primary ESC tum … More ors and 20 cultured ESC cell lines for the mutation of the APC gene revealed that there was not any mutation despite the high frequency of LOH on chromosome 5q.Thirdly, we screened 29 human ESC cell lines for mutations of the p53 gene through all coding exons and non-intron junctions. Mutations were found in 22 cell lines (76%) . We also examined 65 fresh tumor samples, from all of which we tried to establish cell lines, and detected mutations in 26 samples (40%) . There was no significant correlation between the status of the p53 gene in fresh tumors and the establishment of cell lines. We conclude that p53 mutation is not important in establishing cultured human ESC cell lines but provides a growth advantage in vitro. Neither was a relationship observed between p53 mutation and cell growth in protein free environment.We also examined relationships between p53 mutation, murine double minute 2 (MDM2) amplification, and human papillomavirus (HPV) infection in ESC.In most tumors, amplification of the MDM2 gene or infection of HPV was not associated with p53 mutations. In cell culture, MDM2 gene amplification only had a significant relation to cell culture.With regard to patients prognosis, CyclinD1 gene amplification was significantly correlated to poor prognosis in patients and we also found that a group of patients with p53 mutation and/or MDM2 amplification exhibited a significantly shorter survival period.We are now studying the relationship of invasive character and cell membrane fluidity to protein free culture. Less