Davenas E, Rouleau A, Morisset S, Arrang J M
Laboratoire de Neurobiologie et Pharmacologie Moléculaire, Centre de Psychiatrie et Neurosciences de l'INSERM, 2 ter rue d'Alésia, 75014 Paris, France.
J Pharmacol Exp Ther. 2008 Aug;326(2):406-13. doi: 10.1124/jpet.107.135368. Epub 2008 May 12.
Previous studies have suggested that histamine (HA) acts as an autocrine growth factor. We have explored the modulation of cell proliferation by HA using McA-RH7777 hepatoma cells. High L-histidine decarboxylase (HDC) expression and HA synthesis were found in McA-RH7777 cells. Whereas extracellular HA reached submicromolar concentrations, intracellular levels were very low, indicating that HA was secreted by the cells. McA-RH7777 cells also express H3-receptor (H3R) transcripts and proteins. Reverse transcriptase-polymerase chain reaction analysis detected only transcripts for the long isoform. Immunocytochemistry performed with a selective H3R antibody showed that most cells were immunoreactive. H3R binding sites (Bmax approximately 30 fmol/mg protein) were identified when [125I] iodoproxyfan binding was displaced by the agonist imetit. High-affinity binding also occurred at cytochrome P450 enzymes. This binding was not inhibited by HA, H3R agonists, or by a nonimidazole H3R antagonist but was displaced by imidazole H3R antagonists or by ketoconazole, a imidazole-containing cytochrome inhibitor. HA inhibited proliferation of McA-RH7777 hepatoma cells. The absence of uptake system, its much higher potency at H3Rs, and its low intracellular levels suggested that HA interacted with H3Rs rather than cytochromes. In agreement, both imidazole H3R antagonists, a nonimidazole H3R antagonist, and the HDC inhibitor alpha-monofluoromethyl histidine increased cell proliferation (up to approximately 60%), revealing a H3R-mediated inhibition by endogenous HA. Moreover, exogenous HA inhibited the increase induced by alpha-FMH or H3R antagonists with a nanomolar potency. In conclusion, our findings show that HA regulates proliferation of McA-RH7777 hepatoma cells by interacting with autoinhibitory H3Rs.
先前的研究表明,组胺(HA)作为一种自分泌生长因子发挥作用。我们利用McA-RH7777肝癌细胞探索了HA对细胞增殖的调节作用。在McA-RH7777细胞中发现了高L-组氨酸脱羧酶(HDC)表达和HA合成。虽然细胞外HA浓度达到亚微摩尔水平,但细胞内水平非常低,表明HA是由细胞分泌的。McA-RH7777细胞还表达H3受体(H3R)转录本和蛋白质。逆转录聚合酶链反应分析仅检测到长异构体的转录本。用选择性H3R抗体进行的免疫细胞化学显示,大多数细胞具有免疫反应性。当激动剂碘甲磺苯酰胺使[125I]碘甲磺苯酰胺结合被取代时,鉴定出H3R结合位点(Bmax约为30 fmol/mg蛋白质)。在细胞色素P450酶处也发生高亲和力结合。这种结合不受HA、H3R激动剂或非咪唑类H3R拮抗剂的抑制,但被咪唑类H3R拮抗剂或酮康唑(一种含咪唑的细胞色素抑制剂)取代。HA抑制McA-RH7777肝癌细胞的增殖。由于缺乏摄取系统、其在H3R上的效力高得多以及细胞内水平低,表明HA与H3R相互作用而非与细胞色素相互作用。一致的是,咪唑类H3R拮抗剂、非咪唑类H3R拮抗剂和HDC抑制剂α-单氟甲基组氨酸均增加细胞增殖(高达约60%),揭示了内源性HA通过H3R介导的抑制作用。此外,外源性HA以纳摩尔效力抑制α-FMH或H3R拮抗剂诱导的增殖增加。总之,我们的研究结果表明,HA通过与自抑制性H3R相互作用来调节McA-RH7777肝癌细胞的增殖。