Establishment An Expression Platform Of OATP1B1388GG And521CC Genetic Polymorphism And The Uptake Study Of Tamoxifen In Vitro

Objective:this study was aim at establishing an expression platform of OATP1B1388GG and521CC genetic polymorphism in vitro, and using this platform to research the study of OATP1B1388GG and521CC genetic polymorphism uptake Tamoxifen (TAM).Methods:this study used molecular biological technique to structure the plasmid of pcDNA3.1(-)OATP1B1and used site-specific mutagenesis to structure the plasmid of poDNA3.1(-)-OATP1B1388GG and521CC. Real-time Polymerase Chain Reaction (PCR) was used to check the CT number of SLCO1B1in HEK293/MCF-7. These plasmids were transient transfection into HEK293/MCF-7, was tested the expression of OATP1B1by Western-blot and tested the expression of SLCO1B1by PCR. The study was divided into six groups:A group was HEK293/MCF-7; B group was HEK293/MCF-7+TAM; C group was HEK293/MCF-7+pcDNA3.1(-)+TAM; D group was HEK293/MCF-7+pcDNA3.1(-)OATP1B1+TAM; E group was HEK293/MCF-7+pcDNA3.1(-)OATP1B1388GG+TAM; F group was HEK293/MCF-7+pcDNA3.1(-)OATP1B1521CC+TAM. In the study, the cell expression platform of pcDNA3.1(-)OATP1B1-HEK293/MCF-7gene polymorphism was joined by10μM、30μM or100μM TAM, and was used HPLC-MS/MS to measure the content TAM after24hours and48hours. At the same time, the cell expression platform was joined by10μM TAM, was used MTT to measure the changes of cell inhibition rate after24hours and48hours, and was used flow cytometry (FCM) to measure the changes of apoptosis and cell cycle after24hours and48hours.Results:1) After24hours or48hours, the content TAM of group B was similarity to group C, which was no statistical significance (P>0.05); the content TAM of group B were remarkably lower than group D, E and F, which was statistically significant,(P<0.05); the content TAM of group D was a little more than group E or F, which was no statistical significance (P>0.05) 2) After24hours or48hours, the changes of cell inhibition rate of group B was similar to group C, which was no statistical significance (P>0.05); the changes of cell inhibition rate of group B were remarkably lower than that of group D, E and F, which was statistically significant (P<0.05); the changes of cell inhibition rate of group D was a little more than group E or F, which was no statistical significance (P>0.05).3) After24hours or48hours, the apoptosis of group B was similar to group C. which was no statistical significance (P>0.05); the apoptosis of group B were evidently lower than that of group D, E and F, which was statistically significant, P<0.05; the apoptosis of group D was a1the more than to group E or F, which was no statistical significance (P>0.05).4) After24hours or48hours, the cell cycle of group B, C, D, E and F markedly arrested cell cycle in G0/G1phase, and decreased S and G2M phase,which compared with that of group A and was statistically significant (P<0.05); the cell cycle of group B was similarity to group C, which was no statistical significance (P>0.05); the cell cycle of group C, D, E and F markedly arrested cell cycle in G0/G1phase, and decreased S and G2M phase, which compared with that of group B and was statistically significant (P<0.05); the cell cycle of group D was a little more than group E or F, which was no statistical significance (P>0.05).Conclusion:The cell platform of OATP1B1genetic polymorphisms has been retinopathy of ptematttrity in vitro, and OATP1B1were reduced by OATP1B1388GG and521CC, but which were no statistical significance.