Alterations Of The AQP4 Expression In Cultured Neonatal Rat Astrocytes During Hypoxia/reoxygenation And The Regulation Mechanism

Objective To investigate the relationship between the alterations of aquaporin-4 expression and edema of astrocytes, and regulation of aquaporin-4 expression by NF- kB in cultured neonatal rat astrocyte hypoxia/reoxygenation, and use of PDTC(PDTC group), and to explore the molecular mechanism of water movement in the brain and its regulation. Methods The cultured neonatal rat primary astrocytes were randomly assigned to hypoxia group(the astrocytes was exposed to 1%O₂, without FBS and glucose for 3h, 6h, 12h and 24h respectively), reoxygenation group (the astrocytes subjected to 12h hypoxia, then transferred to ordinary culture for 3h, 6h, 12h and 24h respectively), PDTC group(the astrocytes added 100μM PDTC to, then ordinary culture for 3h, 6h, 12h, and 24h reaspectively) and the control group. AQP4 mRNA expression was detected in astrocytes of all groups by real time PCR and AQP4 protein expression by Western Blot andimmunofluorescence . The changes of mitochondria and foot processes of astrocytes were obverved by light microscopy and electron microscopy and NF- k B expression by immunofluorescence .Results (D The AQP4 tnRNA and protein expression was significantly decreased in hypoxia group compared with the relevant control group during hypoxia(from 3h to 24h)(P<0.05) and it was from down-regulation gradually to up-reguation in reoxygenation group and PDTC group. The lowerest level of AQP4 mRNA was 3h in reoxygenation group and 6h in PDTC group respectively(P<0.05), the lowerest level of AQP4 protein expression was in 6h of both reoxygenation group and PDTC group. The AQP4 mRNA and protein expression level was much more higher at 24h of reoxygenation group than that in the control(P<0.05) and there was no difference between the control' s at 24h and PDTC 24h group(P>0.05).@ The mitochondria and foot processes of astrocytes were markedly edematous compare with the control during hypoxia(3h, 12h, 24h), reoxygenation(6h), and PDTC group(3h, 6h). There were no improvement in astrocyte edema at 24h and in the groups of reoxygenation and PDTC group .(3) hypoxia(from3h to 24h), reoxygenation(3h) and PDTC(3 h , 6h)groups presented marked increase in expression of NF- k B compared with the control(P<0.05), and subsequently it was restored in reoxygenation(6h,12h, 24h)and PDTC groups(12h, 24h) (P<0.05), it retured to the level of the control group at 12h group of reoxygenation and PDTC groups respectively(P>0.05). Conclusions The presence of astrocyte edema is thought be related to the decrease in AQP4 expression and AQP4 may play a key role in the reestablishment of the cells osmotic equilibrium under cells edema conditions.These results demonstrate that AQP4 is responsible for modulating of brain water transport, suggesting that AQP4 may be a potential target for drug in treatment of brain edema, and activator of AQP4 may provide a new therapeutic option for reducing brain edema in a wide variety of cerebral disorders.â‘¡ The reduction of AQP4 expression may be associated with increased activity of NF- k B. these results strongly suggest that NF- k B may play importment roles in regulating the expression of AQP4.