Inhibition of UVB-induced COX-2 expression by apigenin

描述（由申请人提供）：芹菜素是一种非诱变的生物类黄酮，当局部应用于小鼠皮肤时，可以抑制紫外线诱导的皮肤癌，我们目前正在研究其作用的分子机制。我们实验室最近的研究结果表明，抑制环氧化酶-2 （COX-2）途径是芹菜素发挥其化学预防作用的途径之一。我们已经证明，芹菜素是紫外线诱导的角化细胞中环氧合酶-2 （COX-2）转录的特异性抑制剂。自上一篇文章以来，我们还获得了新的证据，证明紫外线诱导角质形成细胞中COX-2的表达需要COX-2启动子区域前200个碱基中的E-box和ATF/CRE转录因子结合位点，并证明NF-IL6位点的作用。这些结果表明，芹菜素抑制紫外线诱导的皮肤癌的一种机制是通过调节这些位点特异性的DNA结合蛋白的功能。包括我们的合作者Aubrey Morrison博士在内的其他实验室最近的报告表明，COX-2的表达在转录后也受到影响COX-2 mRNA稳定性和调节翻译效率的反式作用因子的调节。因此，在这个修订的应用程序中需要验证的假设是，生物类黄酮芹菜素抑制紫外线诱导的皮肤癌的机制之一是通过调节COX-2表达的转录和转录后控制。在Aim #1中，我们将继续努力鉴定uvb诱导的COX-2转录所需的COX-2启动子中的DNA结合蛋白，以及角化细胞和小鼠表皮中芹菜素的抑制作用。在Aim #2中，我们将研究芹菜素治疗角化细胞调节Aim #1中鉴定的DNA结合蛋白功能的机制。在Aim #3中，我们将描述影响经芹菜素处理的角质形成细胞中uvb诱导的COX-2表达的转录后控制机制，使用COX-2 3'-UTR区域来研究芹菜素干扰紫外线诱导的mRNA稳定和/或翻译效率的能力。在Aim #4中，我们将使用SKH-1小鼠和SKH-1 COX-2缺失小鼠来证实芹菜素在体内阻断uvb诱导的COX-2表达和肿瘤发生的能力。

DESCRIPTION (provided by applicant): Apigenin is a nonmutagenic bioflavonoid that inhibits UV-induced skin cancer when topically applied to mouse skin and we are currently investigating its molecular mechanism(s) of action. Recent results from our laboratory indicate that inhibition of the cyclooxygenase-2 (COX-2) pathway is one of the ways that apigenin exerts its chemopreventive effect. We have demonstrated that apigenin is a specific inhibitor of UV-induced cyclooxygenase-2 (COX-2) transcription in UV-induced keratinocytes. We have also obtained new evidence since the previous submission that UV-induction of COX-2 expression in keratinocytes requires the E-box and ATF/CRE transcription factor binding sites in the first 200 bases of the COX-2 promoter region, as well as demonstrating a role for the NF-IL6 site. These results suggest that one mechanism by which apigenin inhibits UV-induced skin carcinogenesis is by modulating the function of DNA binding proteins specific for these sites. Recent reports from other laboratories including our collaborator Dr. Aubrey Morrison indicate that COX-2 expression is regulated post-transcriptionally as well, by trans-acting factors which affect COX-2 mRNA stability and modulate translational efficiency. Therefore the hypothesis to be tested in this revised application is that one of the mechanisms by which the bioflavonoid apigenin inhibits UV-induced skin carcinogenesis is through modulation of transcriptional and post-transcriptional control of COX-2 expression. The following aims will test this hypothesis: In Aim #1we will continue our efforts to identify the DNA binding proteins in the COX-2 promoter required for UVB-induced COX-2 transcription and for inhibition by apigenin in keratinocytes and mouse epidermis in vivo. In Aim #2 we will investigate the mechanism by which apigenin treatment of keratinocytes modulates the function of the DNA binding proteins identified in Aim #1. In Aim #3 we will characterize the post-transcriptional control mechanisms which affect UVB-induced COX-2 expression in keratinocytes treated with apigenin, using regions of the COX-2 3'-UTR to investigate apigenin's ability to interfere with UV-induced mRNA stabilization and/or translational efficiency. In Aim #4 we will use SKH-1 mice and SKH-1 COX-2 null mice to confirm the ability of apigenin to block UVB-induced COX-2 expression and tumorigenesis in vivo.