The Research On The Characteristic Of Transport Of Two Kinds Of Bile Acids By Oatp1B3in Placenta

Background:Bile acids are synthesized from cholesterol. They take part in fat digestion and absorbtion while they have cytotoxicity. The fetus liver is able to synthesize bile acids early in intra-uterine life, but poor in metabolism. Placenta plays a great role in bile acids transport between the fetus and the mother. Nowdays, eleven kinds of organic anion transporting polypeptides have been found in human. Some researchers pointed out that OATPs are involed in bile acid transport between the fetus and the mother, OATP1B3may take great part in transport function in trophocytes of placenta.Objectives:To investigate the characteristic of transport of two kinds of bile acids by OATP1B3in placenta; To investigate the harmful mechanism to the fetus in ICP; In order to reduce fetal risk and provide a new potential strategies for targeted drug research.Methods:Knock down the expression of OATP1B3by RNAi mediated by lentiviral vector. Detect the interference efficiency by Real-time PCR and Western blot methods. Use MTT method to detect the cytotoxicity of two kinds of bile acids. Establish vitro models with normal Swan cell and cells with OATP1B3knocked down., then test the concentration of two kinds of bile acids by HPLC in order to find out the characteristic of transport.Results:(1) MTT method:GCA and GCDCA in the concentration of lOumol/L or20μmol/L had no cytotoxicity(P>0.05);(2) RNAi mediated by lentivirus:a. mRNA expression decreased94.42%by Real-time PCR method (P<0.05); b. protein expression decreased49.51%by Western blot method (P<0.05);(3) Successfully established vitro monolayer cell models:the average TEER value was31.90±1.47Ω;(4) Bile acids transport:the shape of transport curves between normal group, negative control group and experimental group were almost identical, but different from the curve of blank group. The transport amount of GCA、GCDCA was significantly lower than the normal group and negative control group. After adding GCA/GCDCA into the upper compartment, they were transported into the lower compartment until the concentration of GCA/GCDCA in the lower compartment were higher than the upper compartment, then opposite direction transport took place.Conclusions:(1) GCA and GCDCA with the concentration no more than20μmol/L have no cytotoxicity.(2) RNAi mediated by OATP1B3shRNA lentivirus can specificly inhibit the mRNA and protein expression of OATP1B3.(3) OATP1B3of the trophocyte celline from human placenta plays an important role in transport of GCA and GCDCA.