Regulation of Expression of Renal Organic Anion Transporters OAT1 and OAT3 in a Model of Ischemia/Reperfusion Injury
Regulation of Expression of Renal Organic Anion Transporters OAT1 and OAT3 in a Model of Ischemia/Reperfusion Injury
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DOI：
10.1159/000430328
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发表时间：
2015-08
影响因子：
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通讯作者：
Christina Preising;R. Schneider;M. Bucher;M. Gekle;C. Sauvant
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文献类型：
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作者：
Christina Preising;R. Schneider;M. Bucher;M. Gekle;C. Sauvant
背景资料：最近，我们获得的证据表明，缺血性急性肾损伤（ischemic acute kidney injury，阿基）诱导的rOat 1和rOat 3表达的损害是由考克斯代谢产物介导的，这种抑制可能与肾损伤密切相关。研究方法：（i）通过荧光素摄取研究模型缺血和再灌注（I/R）后近端肾小管细胞对基底外侧有机阴离子的摄取。将来自hOAT 1（SLC 22 A6）和hOAT 3（SCL 22 A8）的推定启动子序列克隆到报告质粒中，转染到HEK细胞中，并且（ii）在模型缺血和再灌注后作为SEAP报告基因测定确定转录活性。在再灌注开始时应用抑制剂或拮抗剂。结果如下：通过使用PKA（H89）和PLC（U 73122）的抑制剂，E前列腺素受体2型（AH 6809）和4型（L161，982）的拮抗剂，我们获得了证据，表明I/R诱导的有机阴离子转运下调是由COX 1代谢产物通过E前列腺素受体4型介导的。通过将布他前列素（EP 2激动剂）或TCS 2510（EP 4激动剂）应用于对照细胞来证实后一种信号传导。简而言之，在建立的报告基因测定中验证了后者信号传导的转录活性。其中，还应用了COX 1（SC 58125）和COX 2（SC 560）的选择性抑制剂。结论：我们的数据表明：（a）COX 1代谢产物通过EP 4受体参与I/R后肾有机阴离子转运（ers）的调节;（B）这是由于相应转运蛋白的转录调节。由于克隆的启动子序列是人源性的，并在人肾上皮细胞系中表达，我们（c）假设I/R后描述的调控机制对人类也有意义。
Background: Recently, we gained evidence that impairment of rOat1 and rOat3 expression induced by ischemic acute kidney injury (AKI) is mediated by COX metabolites and this suppression might be critically involved in renal damage. Methods: (i) Basolateral organic anion uptake into proximal tubular cells after model ischemia and reperfusion (I/R) was investigated by fluorescein uptake. The putative promoter sequences from hOAT1 (SLC22A6) and hOAT3 (SCL22A8) were cloned into a reporter plasmid, transfected into HEK cells and (ii) transcriptional activity was determined after model ischemia and reperfusion as a SEAP reporter gen assay. Inhibitors or antagonists were applied with the beginning of reperfusion. Results: By using inhibitors of PKA (H89) and PLC (U73122), antagonists of E prostanoid receptor type 2 (AH6809) and type 4 (L161,982), we gained evidence that I/R induced down regulation of organic anion transport is mediated by COX1 metabolites via E prostanoid receptor type 4. The latter signaling was confirmed by application of butaprost (EP2 agonist) or TCS2510 (EP4 agonist) to control cells. In brief, the latter signaling was verified for the transcriptional activity in the reporter gen assay established. Therein, selective inhibitors for COX1 (SC58125) and COX2 (SC560) were also applied. Conclusion: Our data show (a) that COX1 metabolites are involved in the regulation of renal organic anion transport(ers) after I/R via the EP4 receptor and (b) that this is due to transcriptional regulation of the respective transporters. As the promoter sequences cloned were of human origin and expressed in a human renal epithelial cell line we (c) hypothesize that the regulatory mechanisms described after I/R is meaningful for humans as well.