Hickok N J, Uitto J
Department of Dermatology, Jefferson Medical College, Philadelphia, Pennsylvania.
J Invest Dermatol. 1992 Mar;98(3):327-32. doi: 10.1111/1523-1747.ep12499799.
Regulation of ornithine decarboxylase (ODC) gene expression and cell growth by all-trans-retinoic acid in the presence and absence of exogenous putrescine were examined in normal keratinocyte cultures maintained in serum-free medium containing 0.15 mM Ca++. Putrescine and the higher polyamines are negative feedback regulators of ODC synthesis and are essential for cell growth. Human keratinocytes were incubated with and without 1 microM putrescine and the effects of 5 x 10(-7) M retinoic acid on ODC mRNA levels, ODC activity, polyamine levels, and DNA synthetic rates were determined. Northern blot analysis of total RNA isolated from breast reduction keratinocytes treated with retinoic acid up to 24 h showed a time-dependent suppression of ODC mRNA levels that was unaffected by putrescine. ODC activity was suppressed more rapidly in keratinocytes grown in the absence of putrescine; however, at 24 h, ODC activity was suppressed to an equal extent under both culture conditions. The effect of retinoic acid on polyamine levels was determined in the absence of exogenous putrescine. Retinoic acid treatment markedly suppressed putrescine and N1-acetylspermidine levels, whereas spermidine and spermine levels were relatively unaffected. The effect of retinoic acid on DNA synthetic rates, as measured by 3H-thymidine incorporation, was variable. Retinoic acid either stimulated or had little effect on keratinocyte DNA synthetic rates in cells derived from breast reductions and cultured in the absence of putrescine; these effects were not opposed by the presence of exogenous putrescine. In contrast, DNA synthesis in keratinocytes derived from neonatal foreskins was consistently suppressed by retinoic acid, independent of the polyamine status. Our data, therefore, suggest that the effect of retinoic acid on cell growth, as indicated by DNA synthetic rates, does not necessarily parallel its effect on ODC activity and mRNA levels.
在含有0.15 mM Ca++的无血清培养基中培养的正常角质形成细胞中，研究了全反式维甲酸在有和没有外源性腐胺存在的情况下对鸟氨酸脱羧酶（ODC）基因表达和细胞生长的调节作用。腐胺和高级多胺是ODC合成的负反馈调节因子，对细胞生长至关重要。将人角质形成细胞分别在有和没有1 microM腐胺的情况下进行孵育，测定5×10(-7) M维甲酸对ODC mRNA水平、ODC活性、多胺水平和DNA合成速率的影响。对用维甲酸处理长达24小时的缩乳术角质形成细胞分离的总RNA进行Northern印迹分析，结果显示ODC mRNA水平呈时间依赖性抑制，且不受腐胺影响。在没有腐胺的情况下生长的角质形成细胞中，ODC活性被更快地抑制；然而，在24小时时，两种培养条件下ODC活性被抑制的程度相同。在没有外源性腐胺的情况下测定了维甲酸对多胺水平的影响。维甲酸处理显著抑制了腐胺和N1-乙酰亚精胺水平，而亚精胺和精胺水平相对未受影响。用3H-胸腺嘧啶核苷掺入法测定，维甲酸对DNA合成速率的影响是可变的。在没有腐胺的情况下培养的缩乳术来源的细胞中，维甲酸要么刺激角质形成细胞DNA合成速率，要么对其影响很小；外源性腐胺的存在并未对抗这些作用。相比之下，维甲酸始终抑制新生儿包皮来源的角质形成细胞中的DNA合成，与多胺状态无关。因此，我们的数据表明，维甲酸对DNA合成速率所表明的细胞生长的影响不一定与其对ODC活性和mRNA水平的影响平行。
