Hafizi Redona, Imeri Faik, Wenger Roland H, Huwiler Andrea
Institute of Pharmacology, University of Bern, Inselspital, INO-F, CH-3010 Bern, Switzerland.
Institute of Physiology, University of Zürich, CH-8057 Zürich, Switzerland.
Int J Mol Sci. 2021 Aug 31;22(17):9467. doi: 10.3390/ijms22179467.
Erythropoietin (Epo) is the critical hormone for erythropoiesis. In adults, Epo is mainly produced by a subset of interstitial fibroblasts in the kidney, with minor amounts being produced in the liver and the brain. In this study, we used the immortalized renal interstitial fibroblast cell line FAIK F3-5 to investigate the ability of the bioactive sphingolipid sphingosine 1-phosphate (S1P) to stimulate Epo production and to reveal the mechanism involved. Stimulation of cells with exogenous S1P under normoxic conditions (21% O) led to a dose-dependent increase in Epo mRNA and protein levels and subsequent release of Epo into the medium. S1P also enhanced the stabilization of HIF-2α, a key transcription factor for Epo expression. S1P-stimulated Epo mRNA and protein expression was abolished by HIF-2α mRNA knockdown or by the HIF-2 inhibitor compound 2. Furthermore, the approved S1P receptor modulator FTY720, and its active form FTY720-phosphate, both exerted a similar effect on Epo expression as S1P. The effect of S1P on Epo was antagonized by the selective S1P and S1P antagonists NIBR-0213 and TY-52156, but not by the S1P antagonist JTE-013. Moreover, inhibitors of the classical MAPK/ERK, the p38-MAPK, and inhibitors of protein kinase (PK) C and D all blocked the effect of S1P on Epo expression. Finally, the S1P and FTY720 effects were recapitulated in the Epo-producing human neuroblastoma cell line Kelly, suggesting that S1P receptor-dependent Epo synthesis is of general relevance and not species-specific. In summary, these data suggest that, in renal interstitial fibroblasts, which are the primary source of plasma Epo, S1P receptor activation upregulates Epo under normoxic conditions. This may have a therapeutic impact on disease situations such as chronic kidney disease, where Epo production is impaired, causing anemia, but it may also have therapeutic value as Epo can mediate additional tissue-protective effects in various organs.
促红细胞生成素（Epo）是红细胞生成的关键激素。在成年人中，Epo主要由肾脏间质成纤维细胞的一个亚群产生，肝脏和大脑中也有少量产生。在本研究中，我们使用永生化肾间质成纤维细胞系FAIK F3-5来研究生物活性鞘脂鞘氨醇-1-磷酸（S1P）刺激Epo产生的能力，并揭示其中涉及的机制。在常氧条件（21% O₂）下用外源性S1P刺激细胞，导致Epo mRNA和蛋白水平呈剂量依赖性增加，随后Epo释放到培养基中。S1P还增强了HIF-2α（Epo表达的关键转录因子）的稳定性。通过敲低HIF-2α mRNA或使用HIF-2抑制剂化合物2可消除S1P刺激的Epo mRNA和蛋白表达。此外，已获批的S1P受体调节剂FTY720及其活性形式FTY720-磷酸，对Epo表达的作用与S1P相似。S1P对Epo的作用被选择性S1P₁和S1P₃拮抗剂NIBR-0213和TY-52156拮抗，但不被S1P₂拮抗剂JTE-013拮抗。此外，经典的MAPK/ERK、p38-MAPK抑制剂以及蛋白激酶（PK）C和D的抑制剂均阻断了S1P对Epo表达的作用。最后，在产生Epo的人神经母细胞瘤细胞系Kelly中重现了S1P和FTY720的作用，表明S1P受体依赖性Epo合成具有普遍相关性，而非物种特异性。总之，这些数据表明，在作为血浆Epo主要来源的肾间质成纤维细胞中，S1P受体激活在常氧条件下上调Epo。这可能对诸如慢性肾病等疾病状况具有治疗意义，在慢性肾病中Epo产生受损会导致贫血，但由于Epo可在各种器官中介导额外的组织保护作用，它也可能具有治疗价值。
