Kumar Hemant, Lim Ji-Hong, Kim In-Su, Choi Dong-Kug
(a)Department of Biotechnology, Konkuk University, Chungju-si 380-701, Republic of Korea.
Department of Biomedical chemistry, Konkuk University, Chungju-si 380-701, Republic of Korea.
Brain Res. 2015 Jun 12;1610:33-41. doi: 10.1016/j.brainres.2015.03.046. Epub 2015 Apr 3.
Hypoxia inducible factor(s) (HIF) are transcription factors that respond to a low level of oxygen or hypoxic conditions. The HIF pathway has been poorly studied under neuroinflammatory conditions, and no reports are available on the regulation of HIF-3α. Several studies have established that non-hypoxic stimuli can modulate the HIF pathway in a cell-specific manner. Recent reports suggest that hypoxia elicits inflammation or that inflammation during hypoxia is involved in a wide array of human diseases. In the present study, we used lipopolysaccharide (LPS), a well know inflammatory agent, to characterize the HIF-3α expression pattern and compare it with that of HIF-1α under inflammatory conditions in BV-2 microglial cells. Moreover, we used reactive oxygen species inhibitors (rotenone, diphenyleneiodonium, and N-acetyl-L-cysteine) under inflammatory conditions to determine the role of the functional electron transport chain in the regulation of HIF-3α in BV-2 microglial cells. Additionally, we utilized YC-1, a specific inhibitor of HIF-1α, to determine the role of HIF-3α in inflammatory conditions after inhibiting the HIF-1α pathway. YC-1 inhibited nuclear localization of HIF-1α following treatment with LPS in BV-2 microglia cells. Immunoblot and immunocytochemistry revealed a transient effect on HIF-3α after pre-treating the cells with YC-1. Furthermore, we determined the role of nuclear factor kappa B (NF-κB) in the regulation of HIF-3α using the NF-κB inhibitor PDTC in LPS-stimulated BV-2 microglia cells. PDTC altogether abolished LPS-induced nuclear translocation of HIF-3α with a partial effect on HIF-1α, suggesting that HIF-3α expression under inflammatory conditions may be directly under the control of the NF-κB pathway in BV-2 microglial cells. Interestingly, HIF-3α and HIF-1α exhibited almost similar responses to a variety of activating or inhibiting pharmacological agents. These results provide the first evidence for regulation of HIF-3α under inflammatory conditions in BV-2 microglial cells.
缺氧诱导因子（HIF）是对低氧水平或缺氧条件作出反应的转录因子。在神经炎症条件下，HIF信号通路的研究较少，目前尚无关于HIF-3α调控的报道。多项研究表明，非缺氧刺激可通过细胞特异性方式调节HIF信号通路。最近的报道显示，缺氧会引发炎症，或者缺氧期间的炎症与多种人类疾病有关。在本研究中，我们使用脂多糖（LPS），一种众所周知的炎症因子，来表征BV-2小胶质细胞在炎症条件下HIF-3α的表达模式，并将其与HIF-1α的表达模式进行比较。此外，我们在炎症条件下使用活性氧抑制剂（鱼藤酮、二苯基碘鎓和N-乙酰-L-半胱氨酸）来确定功能性电子传递链在BV-2小胶质细胞中HIF-3α调控中的作用。此外，我们使用HIF-1α的特异性抑制剂YC-1，来确定在抑制HIF-1α信号通路后HIF-3α在炎症条件下的作用。在BV-2小胶质细胞中用LPS处理后，YC-1抑制了HIF-1α的核定位。免疫印迹和免疫细胞化学显示，用YC-1预处理细胞后，HIF-3α出现短暂变化。此外，我们在LPS刺激的BV-2小胶质细胞中使用NF-κB抑制剂PDTC来确定核因子κB（NF-κB）在HIF-3α调控中的作用。PDTC完全消除了LPS诱导的HIF-3α核转位，对HIF-1α有部分影响，这表明在炎症条件下，BV-2小胶质细胞中HIF-3α的表达可能直接受NF-κB信号通路的控制。有趣的是，HIF-3α和HIF-1α对各种激活或抑制性药物的反应几乎相似。这些结果为BV-2小胶质细胞在炎症条件下HIF-3α的调控提供了首个证据。
