Hagerman R A, Fischer S M, Locniskar M F
Division of Nutritional Sciences, University of Texas at Austin 78712-1097, USA.
Mol Carcinog. 1997 Jul;19(3):157-64.
Differentiation of cultured keratinocytes is controlled by the calcium concentration of the medium and is marked by the expression of differentiation-specific keratins. Treatment with 12-O-tetradecanoylphorbol-13-acetate (TPA) alters the normal differentiation program and suppresses keratin (K) 1 expression. Based on reported similarities in the effects of TPA and the arachidonic acid metabolite 12(S)-hydroxyeicosatetraenoic acid (12(S)-HETE), we hypothesized that 12(S)-HETE might suppress K1 expression in mouse keratinocytes. We also investigated the effect of pretreatment with 13(S)-hydroxyoctadecadienoic acid (13(S)-HODE) because others have reported that 13(S)-HODE prevents 12(S)-HETE-induced events. In our study, 100 nM 12(S)-HETE mimicked the effect of 500 nM TPA in suppressing K1 mRNA expression within 24 h of calcium-induced differentiation. Pretreatment with 100 nM 13(S)-HODE blocked the 12(S)-HETE effect but not the TPA effect. A role for protein kinase C (PKC) was suggested for both TPA and 12(S)-HETE based on the loss of response with the PKC inhibitors bryostatin-1 or RO-31-8220. Both TPA and 12(S)-HETE stimulated keratinocyte PKC activity. Pretreatment with 13(S)-HODE blocked the 12(S)-HETE-induced increase in PKC activity. Immunoblotting showed that whereas TPA caused a rapid, partial translocation of the PKC alpha isozyme, it had no effect on the distribution of PKC delta. Conversely, 12(S)-HETE had no effect on the distribution of PKC alpha but caused a complete translocation of PKC delta. Pretreatment with 13(S)-HODE prevented 12(S)-HETE-elicited translocation of PKC delta. We conclude that 12(S)-HETE mimics the effect of TPA on K1 mRNA and that the effect is mediated through different isoforms of PKC.
培养的角质形成细胞的分化受培养基钙浓度的控制，并以分化特异性角蛋白的表达为标志。用12 - O - 十四烷酰佛波醇 - 13 - 乙酸酯（TPA）处理会改变正常的分化程序并抑制角蛋白（K）1的表达。基于已报道的TPA与花生四烯酸代谢物12(S)-羟基二十碳四烯酸（12(S)-HETE）作用的相似性，我们推测12(S)-HETE可能会抑制小鼠角质形成细胞中K1的表达。我们还研究了用13(S)-羟基十八碳二烯酸（13(S)-HODE）预处理的效果，因为其他人曾报道13(S)-HODE可阻止12(S)-HETE诱导的事件。在我们的研究中，100 nM的12(S)-HETE在钙诱导分化的24小时内模拟了500 nM TPA抑制K1 mRNA表达的作用。用100 nM的13(S)-HODE预处理可阻断12(S)-HETE的作用，但不能阻断TPA的作用。基于使用蛋白激酶C（PKC）抑制剂苔藓抑素 - 1或RO - 31 - 8220后反应丧失，提示TPA和12(S)-HETE的作用均与PKC有关。TPA和12(S)-HETE均刺激角质形成细胞的PKC活性。用13(S)-HODE预处理可阻断12(S)-HETE诱导的PKC活性增加。免疫印迹显示，TPA可导致PKCα同工酶快速、部分易位，但对PKCδ的分布没有影响。相反，12(S)-HETE对PKCα的分布没有影响，但会导致PKCδ完全易位。用13(S)-HODE预处理可阻止12(S)-HETE引起的PKCδ易位。我们得出结论，12(S)-HETE模拟了TPA对K1 mRNA的作用，且该作用是通过PKC的不同同工型介导的。
