Effects of apolipoprotein E gene polymorphism on the intracellular Ca2+ concentration of astrocytes in the early stages post injury.

Abstract

The present study aimed to investigate the correlation between apolipoprotein E (APOE) polymorphisms and the intracellular concentration of Ca2+ in astrocytes in the early stages after an injury. The chondroitin sulfate region of three APOE alleles (ε2, ε3 and ε4) was obtained by reverse transcription-polymerase chain reaction (RT-PCR). A recombinant plasmid, pEGFP-N1-APOE, was constructed and identified by sequencing, while astrocytes were isolated from APOE gene-knockout mice and examined using immunocytochemistry. The recombinant plasmid was transfected into the astrocytes using the liposome-mediated method and cell injury models were constructed by a scratch assay. Laser confocal scanning microscopy (LCSM) was used to detect dynamic alterations in intracellular Ca2+ concentration at 12, 24, 48 and 72 h after injury. Compared with the control group, cells transfected with any of the three alleles demonstrated significant increases in the fluorescence intensity of Ca2+ (P<0.05). The fluorescence intensity of Ca2+ was weak at 12 h after injury, with no statistically significant difference detected between any two groups at this time point (P>0.05). However, the fluorescence intensity increased in a time-dependent manner and at 24, 48 and 72 h post injury, the fluorescence intensity of the ε4 allele-containing cells was significantly higher when compared with that of cells harboring the other two alleles (P<0.05). These results indicate that intracellular Ca2+ overloading may contribute to the deterioration of brain cells and poor outcome subsequent to traumatic brain injury in APOE ε4 carriers.