Effect Of Aquaporin-4 On Swelling Of Astrocytes After Oxygen-glucose Deprivation

ObjectiveTo investigate the effect of aquaporin-4(AQP-4)on the swelling of astrocytes(Ast) after oxygen-glucose deprivation(OGD) by observing the expression of AQP-4 during the different phase of the cell swelling and analyzing the association between the expression of AQP-4 and the Ast swelling.Methods1. Ast Culture and identification, and OGD edema model foundation: Extract and purificate Ast from the cerebral cortex of SD rats, which were borned within 2 days, then identify it by using glial fibrillary acidic protein(GFAP) immunofluorescence staining. Divide Ast into two groups: normal group and OGD group. OGD group cells were cultured for 5h OGD in order to establish OGD edema model, and use the optical microscope to investigate the Ast morphological alterations at 0.5, 2, 8, 24h and 48h after 5 h OGD respectively.2. AQP-4 expression Detection: The expression of AQP-4 mRNA and protein of Ast were detected by RT-PCR and immunofluorescence staining at 0.5, 2, 8, 24h and 48h after 5h OGD respectively,and then analyse the correlation between AQP-4 expression and Ast swelling.Resultes1. Ast Culture, identification and OGD edema model foundation:â‘ Optical microscope: Ast had multilateral irregular shapes and slender processes, and grew closely to each other.â‘¡Fluorescence microscope: GFAP immunofluorescence staining showed that the cells were positive for GFAP staining, and the cell purity were above 98%.â‘¢Optical microscope:after 5h OGD, Ast swelling began at 0.5h, aggravated to the peak at 2h,and then it subsided gradually.Until 48h after the OGD, the swelled Ast finally recoverd to its normal morphology.2. AQP-4 expression Detection: RT-PCR and immunofluorescence staining showed that AQP-4 mRNA and protein expression of Ast had a same change trend after 5h OGD. Compared to the normal group, AQP-4 mRNA and protein expression transiently declined at 0.5 h(P<0.05), then gradually increased to the peak at 8 h(P<0.05),and finally returned back to normal level at 48 h after 5 h OGD.Conclusion1. 5h OGD can cause significant Ast swelling. Cell swelling peaked at 2h, and recovered at 48h after 5h OGD.2. AQP-4 expression decreased at the cell swelling form stage(0.5h to 2h after the OGD), which may inhibit water influx into Ast. AQP-4 expression increased at cell swelling recovery stage(8h to 24h after the OGD), which may help swelled Ast eject water, after 5 h OGD.