Hepatoxicity Of Psoralen And Its Effect On Bile Acid Synthesis And Transport In Liver

Objective:Psoralen is the main active ingredient of Fructus Psoraleae.In recent years,more and more studies have been reported on the hepatotoxic effects of psoralen,but the specific mechanism is still unclear.The purpose of this study was to investigate the hepatotoxicity of psoralen and its effect on the synthesis and transport of bile acids in the liver.Methods:In this study,rats were selected as test subjects.After the administration of psoralen by gavage for different days,the relevant indexes were detected and analyzed to determine the hepatotoxic effect of psoralen in vivo.1.Animal drug administration: at the beginning of the experiment,48 Sprague-Dawley rats were randomly divided into 4 groups: control group,Psoralen 1 d group,Psoralen 3 d group and Psoralen 7 d group,with 12 rats in each group.Then,with C57/BL6 J mice as the experimental object,80 mice were randomly divided into 4 groups: control group,Psoralen 3d group,Psoralen 7 d group,and Psoralen 14 d group,with 20 mice in each group,half male and half female.The drug group was given psoralen solution,and the control group was given the same volume of solvent,gavage once a day.After the last administration by gavage,the patient fasted for 12 h and serum and liver samples were collected.2.Detection of liver injury indicators: the changes of body weight,liver weight and liver coefficient before and after administration were compared to determine the degree of liver injury caused by psoralen;Serum biochemical indexes such as ALT,AST and TBA were measured to evaluate the changes of liver function in mice.Liver homogenization was used to detect TBA and other biochemical indexes in the liver to determine whether the liver was cholestasis.Histopathological changes of liver were observed by H&E staining.3.To explore liver damage mechanism : the Real-time PCR detection of bile acid transport related(NTCP,BSEP,MRP2,etc.)gene expression,Western blot determination of bile acid synthesis transport related(CYP7A1,NTCP,BSEP,MRP2 etc.)protein expression,around the bile acid synthesis,transport and other aspects discussion scurfpea fruit element for the mechanism of intrahepatic bile acid,cholesterol balance disorders.4.A method for simultaneous determination of 24 bile acids in mouse liver by LC-MS/MS was established,and the content of bile acids in mouse Control group and psoralen on 14 d group was determined to analyze the effect of psoralen on intrahepatic bile acid metabolism.Results:1.Compared with the control group,after 3 days of psoralen administration,the liver weight was significantly increased,the liver coefficient was significantly increased,and the serum ALT and AST were also significantly increased.The liver histopathology showed coagulative necrosis,bile duct destruction or hyperplasia of liver cells after 7 days of psoralen administration.Real-time PCR and Western Blot results indicated that psoralen significantly inhibited bile acid exfoliating proteins such as BSEP,MRP2 and OST?,and significantly increased the content of NTCP.2.After 3,7 or 14 days of administration of psoralen,the mice all had different degrees of liver damage compared with the control group,which was also reflected in the results of liver coefficient,liver function changes and H&E staining of the liver.At the same time,it was found that the content of TBA in the liver homogenate of male mice was significantly higher than that in the control group after 3 days administration.Western Blot results showed that psoralen increased cholesterol synthase HMGCR,inhibited bile acid synthase CYP7A1,CYP27A1,etc.,and significantly inhibited bile acid effluents BSEP,OST?,etc.Moreover,the liver damage of male mice was greater than that of female mice after psoralen was given to them.3.An analytical method was successfully established for the determination of 24 bile acids in mouse liver by LC-MS/MS.It was found that: compared with the control group,the ALCA content increased significantly after 14 days of administration,the content of DCA,MCA and APCA decreased significantly,and there was no significant change in other bile acids.On the 14 th day of administration,the UCA content of male mice significantly decreased,while the content of Mo CA,UDCA,HDCA,?-MCA,12-DHCA,UCA,ALCA,CA,TUDCA,TDCA,?-TMCA,and TCA all increased to different degrees,which was significantly higher than that of the control group.Conclusion:1.The intragastric administration of psoralen for 3 days can cause obvious hepatotoxicity in rats,change the expression of intrahepatic bile acid transporter genes and proteins,and destroy the balance of intrahepatic bile acid transport.2.Psoralen can affect the expression of cholesterol-bile acid synthesis and transportation-related proteins in the liver of mice,increase the content of bile acid in liver cells,and show obvious liver damage,and the peak time of liver damage degree in male and female mice is inconsistent.3.The total content of 24 bile acids measured in the liver of male mice after 14 days of administration by gavage of psoralen increased significantly,while that of female mice decreased significantly.4.In this study,psoralen was determined to affect the synthesis,transport and metabolism of intrahepatic bile acid,and it was speculated that psoralen affected the balance of intrahepatic bile acid synthesis and transport,thus showing a series of indications of liver injury.