Marigo A, Andreis P G, Romano F, Cavatton G, Armato U
Dev Biol Stand. 1985;60:371-91.
A single exposure to a low concentration (10-10 mol/L) of tumor promoters (like 12-0-tetradecanoylphorbol-13-acetate (TPA), phenobarbital, and nafenopin) or of hormones (as epidermal growth factor (EGF), glucagon and insulin) or of drugs (like imidazole and indomethacin) stimulated the 24-h flow into DNA synthesis and mitosis of primary neonatal rat hepatocytes incubated in high-calcium (1.8 mmol/L) Eagle's MEM-FBS medium. However, only tumor promoters acted as enhancers of hepatocytic DNA synthesis when a low-calcium (0.1 mmol/L) FBS-MEM medium was used. The tumor promoters' activity was completely suppressed by the simultaneous (or nearly such) addition of low doses (from 25.0 to .25 micrograms/ml; activity, from 100 to 1.0 unit/ml) of exogenous bovine liver Cu,Zn-superoxide dismutase (SOD), whatever the medium's calcium concentration. By contrast, SOD did not inhibit the growth stimulation elicited by hormones and drugs in hepatocytes exposed to the high-calcium FBS-MEM medium. Moreover, several tumor promoters (namely TPA, phenobarbital, nafenopin, saccharin, teleocidin, benzoyl peroxide, BHT, DDT, lindane, clofibrate, and melittin) stimulated DNA synthesis even when the hepatocytes were incubated in the serumless HiWoBa2000 medium, whatever its calcium concentration. In this synthetic medium, the tumor promoter's stimulatory activity was again completely written off by the simultaneous administration of exogenous (superoxide dismunate) SOD. These results disclose the existence of two quite different mechanisms by which neonatal rat hepatocyte growth can be stimulated: (i) the physiological-pharmacological extracellular calcium-dependent SOD-insensitive machinery, mediating the effects of EGF, glucagon, insulin, imidazole, and indomethacin; and (ii) the pathological extracellular calcium-independent SOD-suppressible mechanism operated by agents belonging to the tumor promoters class and involving, as a critical step, the generation of superoxide anions at the surface of the hepatocyte's plasma membrane. The present results also indicate that primary cultures of neonatal rat hepatocytes may constitute a useful tool for promptly and safely identifying compounds endowed with tumor promoters' capabilities.
单次暴露于低浓度（10⁻¹⁰摩尔/升）的肿瘤启动子（如12 - O - 十四烷酰佛波醇 - 13 - 乙酸酯（TPA）、苯巴比妥和萘酚平）、激素（如表皮生长因子（EGF）、胰高血糖素和胰岛素）或药物（如咪唑和吲哚美辛），会刺激在高钙（1.8毫摩尔/升）伊格尔氏MEM - FBS培养基中培养的原代新生大鼠肝细胞进入DNA合成和有丝分裂的24小时进程。然而，当使用低钙（0.1毫摩尔/升）FBS - MEM培养基时，只有肿瘤启动子可作为肝细胞DNA合成的增强剂。无论培养基的钙浓度如何，同时（或几乎同时）添加低剂量（25.0至0.25微克/毫升；活性，100至1.0单位/毫升）的外源性牛肝铜锌超氧化物歧化酶（SOD），肿瘤启动子的活性会被完全抑制。相比之下，SOD不会抑制在高钙FBS - MEM培养基中培养的肝细胞中激素和药物引发的生长刺激。此外，即使肝细胞在无血清的HiWoBa2000培养基中培养，无论其钙浓度如何，几种肿瘤启动子（即TPA、苯巴比妥、萘酚平、糖精、远藤霉素、过氧化苯甲酰、丁基羟基甲苯、滴滴涕、林丹、氯贝丁酯和蜂毒肽）都会刺激DNA合成。在这种合成培养基中，同时给予外源性（超氧化物歧化酶）SOD，肿瘤启动子的刺激活性会再次完全消除。这些结果揭示了两种截然不同的刺激新生大鼠肝细胞生长的机制：（i）生理 - 药理细胞外钙依赖性SOD不敏感机制，介导EGF、胰高血糖素、胰岛素、咪唑和吲哚美辛的作用；（ii）病理细胞外钙非依赖性SOD可抑制机制，由属于肿瘤启动子类别的物质运作，并且作为关键步骤，涉及在肝细胞质膜表面产生超氧阴离子。目前的结果还表明，原代新生大鼠肝细胞培养物可能构成一种有用的工具，用于快速、安全地鉴定具有肿瘤启动子能力的化合物。
