The effects of CoCl2-induced in vitro hypoxia on aquaporin-4 expression in cultured astrocytes / 中华神经科杂志

ABSTRACT

Objective To investigate the expression of aquaporin-4 (AQP4) in cultured astrocytes after in vitro hypoxia induced by CoCl2. Methods After primary culture and subculture, the astrocytes were placed in a controlled atmosphere culture chamber. Both control group and hypoxia groups were established.These groups were further divided into seven sub-groups according to the different time intervals 15, 30minutes and 1,2, 4, 6, 12 hours, respectively (6 apertures for each group). The shape of the astrocytes in each group was observed with light microscopy and transmission electron microscopy ( TEM ). All groups were examined using in situ hybridization, real time fluorescence quantitative reverse transcriptase polymerase chain reaction, immunocytochemistry and Western blot. The data was analyzed statistically with SPSS 13.0 software. Results There was significant consistency between the AQP4 mRNA and protein ( r =0. 85, P ＜0. 01 ). There was slight positive expression of AQP4 in a few astrocytes of the control groups. In the hypoxia groups, the expression of AQP4 increased within 15 minutes; the increase was most prominent between 1 and 4 hours( mRNA in hypoxia groups 0. 26 ± 0. 04, 0. 31 ± 0. 02, 0. 36 ± 0. 04; control groups0. 06 ±0. 01,0. 09 ±0. 01,0. 08 ±0. 01 )after hypoxia and became less between 6 and 12 hours; There was significant difference in the AQP4 expression between the hypoxia groups and control groups among different time points (t = 16. 51, 18.20, 15.26,all P＜0. 01 ). The corresponding pathological changes were cellular edema, which was most prominent between 1 and 4 hours. Under TEM, increase in size of the nucleolus and swelling of endoplasmic reticulum and mitochondria; these changes became more marked with time.Disruption of a few astrocytes was detected in the hypoxia groups at 12 hours. Conclusions The pathological change of astrocytes is cellular edema following hypoxia. There is a positive relationship between the presence and degree of cellular edema as well as the duration of hypoxia and the up-regulating of AQP4.These results imply that AQP4 expression is an important molecular mechanism of celluar edema of astrocytes.