Effect of apolipoprotein E polymorphisms on intracellular Ca2+ concentration in the early stage after astrocyte injury / 中华创伤杂志

ABSTRACT

Objective To investigate the correlation between apolipoprotein E (proteinapoE;geneAPOE) polymorphisms and intracellular Ca2 + concentration in the early stage after astrocyte injury.Methods ( 1 ) The CDS region of three APOE alleles was obtained by using reverse transcription polymerase chain reaction (RT-PCR). Then, the recombinant plasmid pEGFP-N1-APOE was constructed and identified by sequencing. (2) Astrocytes were separated from APOE gene-knockout mice for immunocytochemical identification. The recombinant plasmid was transfected into the astrocytes with liposome-mediated method to screen the cell lines that could stably express APOE information. (3) Cell injury models were set up by scarification. Laser scanning confocal microscope (LCSM) was used to detect the dynamic changes of intracellular Ca2+ at 12, 24, 48 and 72 hours postinjury. Results Compared with the control group ( before injury ), every allele showed significant changes of fluorescence intensity of Ca2 + ( P ＜0.05). At 12 hours after injury, the fluorescence intensity of Ca2+ was weak, with no statistical difference between three groups ( P ＞ 0. 05 ). At 24,48 and 72 hours postinjury, the fluorescence intensity was increased progressively, with significant higher intensity in ε4 group than the other two groups (P ＜0.05 ). Conclusions The concentration of intracellular Ca2+ in the astrocytes carrying APOEε4 allele is higher than that of those carrying APOEε2 and ε3 alleles, indicating that APOEε4 carriers may activate Ca2+ channel and lead to aggravation and poor prognosis of acute injury.