Marchok A C, Rhoton J C, Griesemer R A, Nettesheim P
Cancer Res. 1977 Jun;37(6):1811-21.
Heterotopic tracheal transplants of rats were exposed in vivo to 150 or 640 microng of 7, 12-dimethylbenz(a)anthracene (DMBA) delivered over 2 weeks, and the epithelium was studied in vitro in an attempt to identify changes in growth behavior during early phases of neoplastic development. Explants were made from the exposed tracheas, as well as from a series of controls, and the rate of epithelial outgrowth from the explants was measured. Secondly, the capacity of the outgrowths to survive as primary cultures and their ability to survive multiple in vitro passages were studied. During the initial planting, the rate of outgrowth was by far the greatest from the explants preexposed to 150 microng DMBA. Outgrowth from explants preexposed to 640 microng DMBA was sparse during the first planting but increased markedly during repeated planting when insulin- and hydrocortisone-supplemented medium was used. Establishment of outgrowth during repeated planting of carcinogen-exposed tracheal pieces was clearly hormone dependent. Control explants did not exhibit this hormone dependency. Primary cultures could be established from only three of six explants preexposed to 150 microng DMBA. These explants had been initiated in insulin- and hydrocortisone-supplemented medium. The primary cultures were successfully subcultured. Primary cultures were also established from five of five explants preexposed to 640 microng DMBA and cultured in hormone-supplemented medium. At least one cell line was obtained from each of the explants. To establish and maintain primary cultures from control tracheas required an enriched medium containing amino acid supplements, sodium pyruvate, and putrescine, as well as the hormone supplements. However, such cultures could not be subcultured. The primary cultures from the carcinogen-exposed explants and the subsequently developed cell lines all exhibited morphological characteristics of keratinizing squamous epithelium. These characteristics include: epithelioid cell morphology, multilayering and sloughing of orangeophilic squamous cells, and the presence of keratohyalin granules. These experiments demonstrate a markedly increased in vitro growth capacity of tracheal epithelium after a short in vivo exposure to carcinogen.
将大鼠异位气管移植体在体内暴露于以两周时间给予的150或640微克的7,12 - 二甲基苯并(a)蒽（DMBA），并在体外研究上皮组织，以试图确定肿瘤发生早期阶段生长行为的变化。从暴露的气管以及一系列对照中制备外植体，并测量外植体上皮细胞的生长速率。其次，研究了这些生长物作为原代培养物存活的能力以及它们在多次体外传代中存活的能力。在最初接种时，预先暴露于150微克DMBA的外植体的生长速率迄今为止是最大的。预先暴露于640微克DMBA的外植体在首次接种时生长稀疏，但在使用补充胰岛素和氢化可的松的培养基进行重复接种时显著增加。致癌物暴露的气管片段在重复接种时生长物的形成明显依赖激素。对照外植体未表现出这种激素依赖性。仅从预先暴露于150微克DMBA的六个外植体中的三个能够建立原代培养物。这些外植体是在补充胰岛素和氢化可的松的培养基中起始培养的。原代培养物成功地进行了传代培养。也从预先暴露于640微克DMBA并在补充激素的培养基中培养的五个外植体中的五个建立了原代培养物。从每个外植体至少获得了一个细胞系。从对照气管建立和维持原代培养物需要一种富含氨基酸补充剂、丙酮酸钠、腐胺以及激素补充剂的培养基。然而，这样的培养物不能传代培养。致癌物暴露的外植体的原代培养物以及随后形成的细胞系均表现出角质化鳞状上皮的形态学特征。这些特征包括：上皮样细胞形态、嗜酸性鳞状细胞的多层化和脱落，以及透明角质颗粒的存在。这些实验表明，气管上皮在体内短时间暴露于致癌物后，其体外生长能力显著增强。
