Identification and pharmacological characterization of the prostaglandin FP receptor and FP receptor variant complexes
Identification and pharmacological characterization of the prostaglandin FP receptor and FP receptor variant complexes
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DOI：
10.1038/bjp.2008.142
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发表时间：
2008-07-01
影响因子：
7.3
通讯作者：
Fliri, H.
中科院分区：
文献类型：
--
作者：
Liang, Y.;Woodward, D. F.;Fliri, H.
背景和目的：一种前列胺类似物，比马前列素，已被证明能有效降低眼压，但其确切的作用机制尚不清楚。因此，为了阐明比马前列素这种作用的分子机制，我们重点研究了前列腺素FP受体（FP）和FP受体变体（altFP）复合物的药理学特征。实验方法：通过逆转录聚合酶链反应鉴定FP受体mRNA变异。在共表达FP和altFP4受体变体的HEK293/EBNA细胞中建立FP-altFP4异源二聚体。荧光成像板阅读器用于研究Ca2+的动员。Northern blot检测富半胱氨酸血管生成蛋白61 (Cyr61) mRNA表达上调，western blot检测肌球蛋白轻链（MLC）磷酸化水平。关键结果：在人眼部组织中鉴定出FP受体mRNA的6个剪接变体。免疫沉淀证实，在共表达FP和altFP4受体的HEK293/EBNA细胞中，FP受体与altFP4受体二聚体。在Ca2+动员的动力学研究中，前列腺素F-2 α (PGF(2 α))引起细胞内Ca2+的快速增加，随后是一个稳态阶段。相比之下，bimatoprost引起细胞内Ca2+的立即增加，随后是第二阶段。前列腺素拮抗剂AGN211335选择性和剂量依赖性地抑制了bimatoprost启动的第二阶段Ca2+动员，Cyr61 mRNA上调和MLC磷酸化，但不阻断PGF的作用（2 α）。结论和意义：比马前列素对FP受体缺乏作用，但可能与FP- altfp受体异二聚体相互作用，诱导第二信使信号传导的改变。因此，FP-altFP复合物可能代表了比马前列素药理学的潜在基础。
Background and purpose: A prostamide analogue, bimatoprost, has been shown to be effective in reducing intraocular pressure, but its precise mechanism of action remains unclear. Hence, to elucidate the molecular mechanisms of this effect of bimatoprost, we focused on pharmacologically characterizing prostaglandin FP receptor (FP) and FP receptor variant (altFP) complexes.Experimental approach: FP receptor mRNA variants were identified by reverse transcription-polymerase chain reaction. The FP-altFP4 heterodimers were established in HEK293/EBNA cells co-expressing FP and altFP4 receptor variants. A fluorometric imaging plate reader was used to study Ca2+ mobilization. Upregulation of cysteine-rich angiogenic protein 61 (Cyr61) mRNA was measured by Northern blot analysis, and phosphorylation of myosin light chain (MLC) by western analysis.Key results: Six splicing variants of FP receptor mRNA were identified in human ocular tissues. Immunoprecipitation confirmed that the FP receptor is dimerized with altFP4 receptors in HEK293/EBNA cells co-expressing FP and altFP4 receptors. In the studies of the kinetic profile for Ca2+ mobilization, prostaglandin F-2 alpha (PGF(2 alpha)) elicited a rapid increase in intracellular Ca2+ followed by a steady state phase. In contrast, bimatoprost elicited an immediate increase in intracellular Ca2+ followed by a second phase. The prostamide antagonist, AGN211335, selectively and dose-dependently inhibited the bimatoprost-initiated second phase of Ca2+ mobilization, Cyr61 mRNA upregulation and MLC phosphorylation, but did not block the action of PGF(2 alpha).Conclusion and implications: Bimatoprost lacks effects on the FP receptor but may interact with the FP-altFP receptor heterodimer to induce alterations in second messenger signalling. Hence, FP-altFP complexes may represent the underlying basis of bimatoprost pharmacology.