ABSTRACT
Objectives To study the effect of VIM in Enterovirus 71 (EV71) infection of (human brain microvascular endothelial cells (HBMEC) and elaborating the mechanism of EV71 infection in the nervous system. Methods Knocked out the VIM by CRISPR technology , the differences in EV71 absorption , replication , release between wild VIM and VIM knocked-out (VIM-KO) HBMEC were detected by fluorescence quantitative PCR. Results 4 ℃ absorption experiment conformed that EV71 adsorption in VIM- KO is 40% less than in the normal HBMEC. After EV71 infect HBMEC for 48 h (48 h p. i.), the quantitative PCR result showed intracellular viral RNA in VIM-KO was only 1/12 of that in the normal HBMEC. Also the extracellular viral RNA was quantified, and the number of cells in VIM-KO had been reduced 1.4 times compared with the normal HBMEC. Conclusions Once VIM knocking out, EV71 attachment has been obviously reduced. Meanwhile, the level of viral RNA replication and release are decreased compared with the normal HBMEC. VIM may be an attachment receptor of EV71 in HBMEC , when the virus invades HBMEC with the binding of VIM. Moreover , VIM plays an important role in the replication and release of EV71.
ABSTRACT
AIM:TostudythechangeofradiosensitivityofU251cellsaftertreatedwithsodiumdichloroacetate ( DCA) and further to explore the possible mechanism .METHODS: The U251 cells were divided into 4 groups: control group, DCA group, X-ray irradiation without DCA pretreatment ( IR) group and X-ray irradiation with DCA pretreatment ( DIR) group.MTT assay was applied to determine the cell viability .The intracellular reactive oxygen species ( ROS) were detected by DHE fluorescence .The expression level of Bcl-2 was assessed by Western blot .The percentage of apoptosis of cells was determined by flow cytometry .RESULTS:No difference between control group and DCA group in cell viability (P>0.05) was observed.However, the cell viability of both IR group and DIR group was markedly reduced compared with control group ( P<0.05).Furthermore, the viability of DIR group was significantly decreased compared with IR group ( P<0.05 ) .The percentage of ROS positive cells was obviously increased in DIR group compared with IR group (P<0.05).The expression level of Bcl-2 was sharply decreased in DIR group (P<0.05) and the percentage of apoptosis of cells was significantly elevated ( P<0.05) in DIR group compared with IR group .CONCLUSION:The better antitu-mor effect was obtained by improving the radiosensitivity through pretreating the cells with DCA , and the possible mecha-nism was down-regulation of the Bcl-2 expression by developing the intracellular ROS .