Garayoa M, Martínez A, Lee S, Pío R, An W G, Neckers L, Trepel J, Montuenga L M, Ryan H, Johnson R, Gassmann M, Cuttitta F
Department of Cell and Cancer Biology, National Cancer Institute, National Institutes of Health Bethesda, Maryland 20892, USA.
Mol Endocrinol. 2000 Jun;14(6):848-62. doi: 10.1210/mend.14.6.0473.
Little is known about the molecular mechanisms that control adrenomedullin (AM) production in human cancers. We demonstrate here that the expression of AM mRNA in a variety of human tumor cell lines is highly induced in a time-dependent manner by reduced oxygen tension (1% O2) or exposure to hypoxia mimetics such as desferrioxamine mesylate (DFX) or CoCl2. This AM expression seems to be under hypoxia-inducible factor-1 (HIF-1) transcriptional regulation, since HIF-1alpha and HIF-1beta knockout mouse cell lines had an ablated or greatly reduced hypoxia AM mRNA induction. Similarly, inhibition or enhancement of HIF-1 activity in human tumor cells showed an analogous modulation of AM mRNA. Under hypoxic conditions, immunohistochemical analysis of tumor cell lines revealed elevated levels of AM and HIF-1alpha as compared with normoxia, and we also found an increase of immunoreactive AM in the conditioned medium of tumor cells analyzed by RIA. AM mRNA stabilization was shown to be partially responsible for the hypoxic up-regulated expression of AM. In addition, we have identified several putative hypoxia response elements (HREs) in the human AM gene, and reporter studies with selected HREs were capable of enhancing luciferase expression after exposure to DFX. Furthermore, transient coexpression of HIF-1alpha resulted in an augmented transactivation of the reporter gene after DFX treatment. Given that most solid human tumors have focal hypoxic areas and that AM functions as a mitogen, angiogenic factor, and apoptosis-survival factor, our findings implicate the HIF-1/AM link as a possible promotion mechanism of carcinogenesis.
关于控制人类癌症中肾上腺髓质素（AM）产生的分子机制，人们了解甚少。我们在此证明，在多种人类肿瘤细胞系中，低氧张力（1% O₂）或暴露于低氧模拟物（如甲磺酸去铁胺（DFX）或CoCl₂）会以时间依赖性方式高度诱导AM mRNA的表达。这种AM表达似乎受缺氧诱导因子-1（HIF-1）转录调控，因为HIF-1α和HIF-1β基因敲除小鼠细胞系对低氧诱导的AM mRNA表达缺失或大幅降低。同样，在人类肿瘤细胞中抑制或增强HIF-1活性也显示出对AM mRNA的类似调节。在低氧条件下，对肿瘤细胞系的免疫组织化学分析显示，与常氧相比，AM和HIF-1α水平升高，并且我们还通过放射免疫分析发现，在分析的肿瘤细胞条件培养基中免疫反应性AM增加。AM mRNA的稳定化被证明部分负责低氧上调的AM表达。此外，我们在人类AM基因中鉴定出了几个假定的缺氧反应元件（HREs），对选定HREs的报告基因研究能够在暴露于DFX后增强荧光素酶表达。此外，HIF-1α的瞬时共表达导致DFX处理后报告基因的转录激活增强。鉴于大多数人类实体瘤都有局部缺氧区域，且AM作为一种促有丝分裂因子、血管生成因子和凋亡存活因子发挥作用，我们的发现表明HIF-1/AM联系可能是一种致癌促进机制。
