NFAT5 deficiency increases the severity of neuronal death in ischemic condition

Abstract

Previously, we have shown that over expression of endothelin-1 (ET-1) in astrocytes (GET-1) exhibited more severe neurological deficits, increased brain swelling with water accumulation and aquaporin 4 (AQP 4) after transient middle cerebral artery occlusion (MCAO). Unexpectedly, ipsilateral side of GET-1 brains expressed decreased NFAT5/tonEBP/OREBP (tonicity responsive element binding protein) levels when compared to NTg brains after transient MCAO, suggesting that NFAT 5 may play a role in ischemic injury. NFAT5, a transcription factor, regulates edema related genes such as AQP 4. To further confirm whether NFAT5 is regulated by ischemia, we transfected neurons (SHY5Y) with osmotic responsive element (ORE)-linked to luciferase. The increased luciferase activity was observed in neurons treated with 1 or 3 hr of ischemia. In addition, immunocytochemical analysis with NFAT5 antibodies showed that translocation of NFAT5 from cytoplasm to nucleus after ischemic challenge. To understand the role of NFAT5 in ischemia, neurons from the homozygous NFAT5 knockout (NFAT5 -/-) embryos at day 14.5 were cultured and challenged them with 3 hr ischemia. The neurons from NFAT5 -/- mice released a significantly higher LDH level compared to that of NFAT5 +/+, suggesting that activation of NFAT5 protects neurons against ischemic injury. To further examine the in vivo role of NFAT5 in ischemic stress, NFAT5 +/- mice were subjected to transient MCAO since the NFAT5 -/- mice die during embryonic stage. The NFAT5 +/- mice displayed more severe neurological score, larger infarct size and volume and brain swelling compared to NFAT5 +/+ mice. Our present data suggest that activation and translocation of NFAT5 to nucleus under ischemic condition may contribute to protection of neurons against the ischemic injury.