Influence of ATP-binding cassette transporter A1 protein in eukaryocyte and its expression on arsenic resistance

Abstract

Objective To examine the expression of ATP-binding cassette transporter A1(ABCA1)in eukaryotie cells and the effect of arsenic resistance after the transfection of eukaryotic expression vector containing ABCA1 gene.Methods HeLa cells were transfected with the recombinant plasmid by lipofectaonmine 2000 (recombinant plasmid group),empty plasmid and untransfected HeLa cell as the control group.The level of the mRNA was examined by real-time PCR,and the expression of ABCA1 protein wag examined by Western blot,the change of cell survival rate was examined by methyl thiazolyl tetrazolium(MTT)after exposure in a series of arsenic [0(contro1),4,8,16,32,64,128 μmol/L]for 48 hours.Results Expression level of ABCA1 mRNA in recombinant plasmid,empty plasmid and untransfeeted groups was(2.09±0.08)×10-4,(0.09±0.02)×10-4,(0.08±0.02)×10-4,there was a significant difference between the groups(F=1499.23,P<0.01).The level of ABCA1 mRNA in recombinant plasmid group was higher than empty plasmid and untransfected group(all P<0.01).Western blot showed that specific protein straps existed at 254×103 in all the three groups,with a similar size to the ABCA1 protein.The amount of the recombinant plasmid group was higher than the other two groups.MTT shows that arsenic concentration at 4,8,16,32,64,128 μmol/L,the survival rates of recombinant plasmid group was(94.8±0.9)%,(86.5 ± 2.6)%, (77.8 ± 2.0)%, (56.0 ± 2.0)%, (23.8 ± 1.7)%, (18.6 ± 0.6)%, higher than that of empty plasmid group[ (85.3 ± 1.1)%, (78.7 ± 0.6)%, (67.8 ± 2.4)%, (43.2 ± 1.5)%, (14.5 ± 1.3)%, (8.0 ± 0.4)%], the difference of survival rate had a statistical signifieance(t = 18.985,6.689,5.922,9.504,9.481,32.634, all P < 0.01). Conclusions ABCA1 protein is over expressed in HeLa cells after transfect ABCA1 gene. ABCA1 protein increases resistance of arsenic in HeLa cells. Key words: ATP-binding cassette transporter A1 gene; Plasmids; Transfection; Eukaryotic cells; Arsenic resistance