Division of Hematology, Cancer and Blood Diseases Institute, Cincinnati Children's Hospital Medical Center, Cincinnati, Ohio; Department of Biochemistry and Molecular Biology, Keck School of Medicine of the University of Southern California, Los Angeles, California.
Am J Hematol. 2014 Mar;89(3):273-81. doi: 10.1002/ajh.23631.
The potassium chloride cotransporters (KCC) family of proteins are widely expressed and are involved in the transepithelial movement of potassium and chloride ions and the regulation of cell volume. KCC activity is high in reticulocytes, and contributes to the dehydration of sickle red blood cells. Because plasma levels of both vascular endothelial growth factor (VEGF) and placental growth factor (PlGF) are elevated in sickle cell individuals, and VEGF has been shown to increase KCC expression in other cells, we hypothesized that VEGF and PlGF influence KCC expression in erythroid cells. Both VEGF and PlGF treatment of human erythroid K562 cells increased both mRNA and protein levels of KCC1, KCC3b, and KCC4. VEGF- and PlGF-mediated cellular signaling involved VEGF-R1 and downstream effectors, specifically, PI-3 kinase, p38 MAP kinase, mTOR, NADPH-oxidase, JNK kinase, and HIF-1α. VEGF and PlGF-mediated transcription of KCC3b and KCC4 involved hypoxia response element (HRE) motifs in their promoters, as demonstrated by promoter analysis, EMSA and ChiP. These results were corroborated in vivo by adenoviral-mediated overexpression of PlGF in normal mice, which led to increased expression of mKCC3 and mKCC4 in erythroid precursors. Our studies show that VEGF and PlGF regulate transcription of KCC3b and KCC4 in erythroid cells via activation of HIF-1α, independent of hypoxia. These studies provide novel therapeutic targets for regulation of cell volume in RBC precursors, and thus, amelioration of dehydration in RBCs in sickle cell disease.
钾氯协同转运蛋白（KCC）家族蛋白广泛表达，参与钾离子和氯离子的跨上皮运动以及细胞体积的调节。KCC 活性在网织红细胞中较高，有助于镰状红细胞的脱水。由于镰状细胞个体的血管内皮生长因子（VEGF）和胎盘生长因子（PlGF）的血浆水平升高，并且 VEGF 已被证明可增加其他细胞中的 KCC 表达，我们假设 VEGF 和 PlGF 影响红细胞中的 KCC 表达。VEGF 和 PlGF 处理人红系 K562 细胞均可增加 KCC1、KCC3b 和 KCC4 的 mRNA 和蛋白水平。VEGF 和 PlGF 介导的细胞信号转导涉及 VEGF-R1 和下游效应物，具体而言，PI-3 激酶、p38 MAP 激酶、mTOR、NADPH 氧化酶、JNK 激酶和 HIF-1α。VEGF 和 PlGF 介导的 KCC3b 和 KCC4 转录涉及其启动子中的缺氧反应元件（HRE）基序，如启动子分析、EMSA 和 ChiP 所示。通过在正常小鼠中过表达 PlGF 的腺病毒介导的体内研究证实了这些结果，这导致红系前体细胞中 mKCC3 和 mKCC4 的表达增加。我们的研究表明，VEGF 和 PlGF 通过激活 HIF-1α调节红细胞中 KCC3b 和 KCC4 的转录，独立于缺氧。这些研究为调节 RBC 前体中的细胞体积提供了新的治疗靶点，从而改善镰状细胞病中 RBC 的脱水。
