Tong Min, Ding Yunfei, Tai Hsin-Hsiung
Department of Pharmaceutical Sciences, College of Pharmacy, University of Kentucky Lexington, KY 40536-0082, USA.
Carcinogenesis. 2006 Nov;27(11):2170-9. doi: 10.1093/carcin/bgl053. Epub 2006 Apr 22.
Human lung adenocarcinoma cells, A549, possess the capacity of expressing both cyclooxygenase-2 (COX-2) and NAD+-linked 15-hydroxyprostaglandin dehydrogenase (15-PGDH). Resting cells express little COX-2 but significant levels of 15-PGDH. Interleukin (IL) 1beta, tumor necrosis factor-alpha (TNF-alpha) or phorbol ester [phorbol 12-myristate 13-acetate (PMA)] induced the expression of COX-2, as revealed by western blot analysis. Combination of PMA and IL-1beta or TNF-alpha induced synergistically the expression of COX-2. Interestingly, cytokines and cytokine plus PMA-induced expression of COX-2 were accompanied by a downregulation of 15-PGDH. This was evident from both the western blot analysis and activity assay of 15-PGDH. It appears that the higher the expression of COX-2 was induced, the lower the expression of 15-PGDH was found. This was further supported by the observation that overexpression of COX-2 but not COX-1 by adenovirus-mediated approach led to a decrease in 15-PGDH expression, indicating the specificity of COX-2. Furthermore, downregulation of the IL-1beta-induced expression of COX-2 by silencing RNA (siRNA) approach resulted in an increase in the expression of 15-PGDH by COX-2-siRNA but not by COX-1-siRNA, indicating that it was indeed the expression of COX-2 attenuating the expression of 15-PGDH. The IL-1beta-induced reduction of the expression of 15-PGDH was shown not to be mediated by COX-2-derived products since the presence of COX-2 inhibitors did not block the attenuation of the expression of 15-PGDH. Exogenous PGE2 also did not induce the reduction of the expression of 15-PGDH. However, overexpression of 15-PGDH by transfection with recombinant plasmid encoding 15-PGDH or adenovirus-mediated approach attenuated IL-1beta-induced expression of COX-2. On the contrary, downregulation of 15-PGDH expression by 15-PGDH-siRNA or 15-PGDH-antisense approach resulted in an increase in IL-1beta-induced expression of COX-2 but not that of COX-1. In fact, it was further observed that A549 clones expressing different degrees of 15-PGDH showed also different levels of COX-2 expression after IL-1beta induction. The levels of IL-1beta-induced COX-2 expression appeared to correlate inversely with those of 15-PGDH expression in the cells. These results support the contention that COX-2 and 15-PGDH are regulated reciprocally in A549 cells.
人肺腺癌细胞A549具有同时表达环氧化酶-2（COX-2）和NAD⁺连接的15-羟基前列腺素脱氢酶（15-PGDH）的能力。静息细胞中COX-2表达很少，但15-PGDH表达水平较高。白细胞介素（IL）1β、肿瘤坏死因子-α（TNF-α）或佛波酯[佛波醇12-肉豆蔻酸酯13-乙酸酯（PMA）]可诱导COX-2表达，这通过蛋白质免疫印迹分析得以揭示。PMA与IL-1β或TNF-α联合使用可协同诱导COX-2表达。有趣的是，细胞因子以及细胞因子加PMA诱导的COX-2表达伴随着15-PGDH的下调。这从蛋白质免疫印迹分析和15-PGDH活性测定中都很明显。似乎诱导的COX-2表达越高，发现的15-PGDH表达越低。腺病毒介导的方法过表达COX-2而非COX-1导致15-PGDH表达下降，这一观察结果进一步支持了上述观点，表明了COX-2的特异性。此外，通过小干扰RNA（siRNA）方法沉默IL-1β诱导的COX-2表达导致COX-2-siRNA而非COX-1-siRNA介导的15-PGDH表达增加，表明确实是COX-2的表达减弱了15-PGDH的表达。IL-1β诱导的15-PGDH表达降低并非由COX-2衍生产物介导，因为COX-2抑制剂的存在并未阻断15-PGDH表达的减弱。外源性前列腺素E2也未诱导15-PGDH表达的降低。然而，通过转染编码15-PGDH的重组质粒或腺病毒介导的方法过表达15-PGDH可减弱IL-1β诱导的COX-2表达。相反，通过15-PGDH-siRNA或15-PGDH反义方法下调15-PGDH表达导致IL-1β诱导的COX-2表达增加，但COX-1的表达未增加。事实上，进一步观察到，表达不同程度15-PGDH的A549克隆在IL-1β诱导后也显示出不同水平的COX-2表达。细胞中IL-1β诱导的COX-2表达水平似乎与15-PGDH表达水平呈负相关。这些结果支持了COX-2和15-PGDH在A549细胞中相互调节的观点。
