Differential regulation of HIF-3α in LPS-induced BV-2 microglial cells: Comparison and characterization with HIF-1α.

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.