Wang T T, Lachance S, Delalandre A, Carrière S, Chan J S
University of Montreal, Maisonneuve-Rosemont Hospital, Research Center, Quebec, Canada.
Am J Physiol. 1996 Sep;271(3 Pt 2):R519-27. doi: 10.1152/ajpregu.1996.271.3.R519.
To investigate whether expression of the renal angiotensinogen gene is regulated by dopaminergic receptors, we used opossum kidney (OK 27) cells with a fusion gene containing the 5'- flanking regulatory sequence of the rat angiotensinogen gene fused with a human growth hormone (hGH) gene as a reporter [pOGH, angiotensinogen nucleotide (N) -1498/+18], permanently integrated into their genomes. The level of expression of pOGH (angiotensinogen N-1498/+18) in OK 27 was evaluated by the amount of immunoreactive hGH (ir-hGH) secreted into the culture medium. In the absence of 3-isobutyl-1-methylxanthine (IBMX), addition of dopamine (10(-13) to 10(-5)M) had minimal effect on the expression of the pOGH (angiotensinogen N-1498/+18) in OK 27 cells. In the presence of IBMX, addition of low concentrations (10(-13) and 10(-7) M) of dopamine stimulated the expression of pOGH angiotensinogen N-1498/+18) in OK 27 cells in a dose-dependent manner, whereas high concentrations (i.e., > 10(-7) M) had minimal effect. The stimulatory effect of dopamine on the expression of pOGH (angiotensinogen N-1498/+18) was inhibited by the presence of SCH-23390 (D1-dopaminergic receptor antagonist) and spiperone (D2-dopaminergic receptor antagonist), but not by ketanserin (5 HT2/5HT1c-serotonergic receptor antagonist). Moreover, the stimulatory effect of dopamine was inhibited by the presence of U-73122 (an inhibitor of phospholipase C and phospholipase A2) or staurosporine (an inhibitor of protein kinase C) or (R)-p-adenosine 3',5'-cyclic monophosphorothioate (Rp-cAMP[S]; an inhibitor of cAMP-dependent protein kinase AI and II). Addition of low concentrations (10(-13) to 10(-9)M) of SKF-82958 (D1-dopaminergic receptor agonist) or PPHT (D2-dopaminergic receptor agonist) also stimulated the expression of pOGH (angiotensinogen N-1498/+18). The stimulatory effect of SKF-82958 was inhibited by the presence of SCH-23390 or Rp-cAMP[S], whereas the effect of PPHT was inhibited by the presence of spiperone or staurosporine. These studies demonstrate that the expression of pOGH (angiotensinogen N-1498/+18) in OK 27 cells is modulated by dopaminergic receptor agonists.
为研究肾血管紧张素原基因的表达是否受多巴胺能受体调控，我们使用了负鼠肾（OK 27）细胞，该细胞带有一个融合基因，其包含大鼠血管紧张素原基因5'侧翼调控序列与人类生长激素（hGH）基因融合，作为报告基因[pOGH，血管紧张素原核苷酸（N）-1498 / +18]，并永久整合到其基因组中。通过分泌到培养基中的免疫反应性hGH（ir-hGH）量来评估OK 27中pOGH（血管紧张素原N-1498 / +18）的表达水平。在不存在3-异丁基-1-甲基黄嘌呤（IBMX）的情况下，添加多巴胺（10^(-13)至10^(-5)M）对OK 27细胞中pOGH（血管紧张素原N-1498 / +18）的表达影响极小。在存在IBMX的情况下，添加低浓度（10^(-13)和10^(-7)M）的多巴胺以剂量依赖方式刺激OK 27细胞中pOGH血管紧张素原N-1498 / +18）的表达，而高浓度（即> 10^(-7)M）影响极小。多巴胺对pOGH（血管紧张素原N-1498 / +18）表达的刺激作用被SCH-23390（D1多巴胺能受体拮抗剂）和螺哌隆（D2多巴胺能受体拮抗剂）抑制，但不被酮色林（5 HT2 / 5HT1c血清素能受体拮抗剂）抑制。此外，多巴胺的刺激作用被U-73122（磷脂酶C和磷脂酶A2抑制剂）或星形孢菌素（蛋白激酶C抑制剂）或（R）-p-腺苷3'，5'-环一磷酸硫代酯（Rp-cAMP[S]；cAMP依赖性蛋白激酶AI和II抑制剂）抑制。添加低浓度（10^(-13)至10^(-9)M）的SKF-82958（D1多巴胺能受体激动剂）或PPHT（D2多巴胺能受体激动剂）也刺激了pOGH（血管紧张素原N-1498 / +18）的表达。SKF-82958的刺激作用被SCH-23390或Rp-cAMP[S]抑制，而PPHT的作用被螺哌隆或星形孢菌素抑制。这些研究表明，OK 27细胞中pOGH（血管紧张素原N-1498 / +18）的表达受多巴胺能受体激动剂调节。
