ABCG2 Protects Kidney Side Population Cells From Hypoxia/Reoxygenation Injury Through Activation of the MEK/ERK Pathway
ABCG2 Protects Kidney Side Population Cells From Hypoxia/Reoxygenation Injury Through Activation of the MEK/ERK Pathway
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ABCG2 通过激活 MEK/ERK 通路保护肾侧细胞群免受缺氧/复氧损伤
DOI：
10.3727/096368912x657206
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发表时间：
2013-01-01
影响因子：
3.3
通讯作者：
Liu, Shui-Bing
中科院分区：
文献类型：
--
作者：
Liu, Wei-Hui;Liu, Hong-Bao;Liu, Shui-Bing
乳腺癌耐药蛋白1(BCRP1/ABCG2)被用来识别一组细胞中的侧群(SP)，这些SP富含不同组织中的干细胞和祖细胞。在这里，我们研究了细胞外信号调节激酶(ERK)1/2在肾脏SP细胞缺血/缺氧条件下的信号机制中的作用。用Hoechst 33342染料介导的荧光素激活的细胞分选法分离肾SP细胞，然后在缺氧/复氧(H/R)条件下与选择性BCRP1/ABCG2抑制剂维拉帕米共同孵育。H/R处理后检测ABCG2表达、ERK活性、细胞存活率、代谢活性和膜损伤。为探讨ERK 1/2在ABCG2表达和功能中的作用，用重组正义表达载体pcDNA3.1-MEK上调了优先激活ERK的丝裂原活化蛋白激酶(MAPK)/ERK激酶(MEK)的表达，并用MEK特异性抑制剂U0126抑制其表达。我们发现，在体外和体内，低氧均能激活肾脏SP细胞的ERK活性，但对非SP细胞的ERK活性无明显影响。MEK过表达可模拟低氧诱导的ABCG2表达。相反，U0126抑制缺氧和MEK上调ABCG2的表达。此外，H/R诱导SP细胞和非SP细胞的核、代谢和膜损伤显著增加，但这种H/R诱导的细胞毒性在非SP细胞中比SP细胞严重得多。值得注意的是，MEK过表达增强了肾SP细胞的活性，而U0126则抑制了其活性。维拉帕米逆转MEK诱导的肾SP细胞活性。将SP细胞全身应用于肾缺血/再灌注损伤动物体内，可显著改善肾功能，加速细胞分裂反应，减少细胞凋亡。然而，维拉帕米可显著降低SP细胞的这种改善的治疗潜力。总之，这些发现为MEK/ERK-ABCG2通路在保护肾脏SP细胞免受缺血/缺氧损伤中发挥关键作用提供了证据。
Breast cancer resistance protein 1 (BCRP1/ABCG2) is used to identify the side population (SP) within a population of cells, which is enriched for stem and progenitor cells in different tissues. Here, we investigated the role of extracellular signal-regulated kinase (ERK) 1/2 in the signaling mechanisms underlying ischemic/hypoxic conditions in kidney SP cells. Kidney SP cells were isolated using Hoechst 33342 dye-mediated fluorescein-activated cell sorting and then incubated under hypoxia/reoxygenation (H/R) with or without verapamil, a selective BCRP1/ABCG2 inhibitor. ABCG2 expression, ERK activity, cell viability, metabolic activity, and membrane damage were tested after H/R treatment. To evaluate the role of ERK 1/2 on the expression and function of ABCG2, the expression of mitogen-activated protein kinase (MAPK)/ERK kinase (MEK), which preferentially activates ERK, was upregulated by transfection with the recombinant sense expression vector pcDNA3.1-MEK and downregulated by pretreatment with U0126, a specific MEK inhibitor. We found that hypoxia activated ERK activity in the kidney SP cells but not in non-SP cells both in vitro and in vivo. Overexpression of MEK mimicked hypoxia-induced ABCG2 expression. Contrarily, U0126 inhibited hypoxia- and MEK-upregulated ABCG2 expression. Furthermore, H/R induced significant increases in nuclear, metabolic, and membrane damage in both SP cells and non-SP cells; however, this H/R-induced cytotoxicity was much more severe in non-SP cells than in SP cells. Notably, the viability of kidney SP cells was enhanced by MEK overexpression and inhibited by U0126. Verapamil treatment reversed MEK-induced viability of kidney SP cells. When administered systemically into animals with renal ischemia/reperfusion injury, the SP cells significantly improved renal function, accelerated mitogenic response, and reduced cell apoptosis. However, this improved therapeutic potential of SP cells was significantly reduced by pretreatment with verapamil. Collectively, these findings provide evidence for a crucial role for the MEK/ERK-ABCG2 pathway in protecting kidney SP cells from ischemic/hypoxic injury.