| Compatibility of prescriptions is the main way of preventing diseases in traditional Chinese medicine.Clinical practice has proved that there are great advantages in efficacy or safety of prescriptions compared with unit drug use.Traditional Chinese medicine prescriptions are not simple additions of flavors,but "Masters under control" optimized by the compatibility theory of "Qiqinghehe".Therefore,it is of great significance to study the compatibility law of prescriptions and elaborate the mechanism of synergism and toxicity reduction for clarifying the material basis of prescriptions,improving the quality standard of prescriptions and rational drug use in clinic.Organic anion transporters(OATs)are transporters secreted by renal tubules,which can transport many organic anions(OAs),including some toxicants,drugs and their metabolites through the kidney,thus maintaining the internal environment of the body.Studies have shown that once inhibited by some drugs,food or poisons entering the human body,kidney organic anion transporters may interfere with the normal excretion of other OATs substrates in the body,resulting in abnormal human excretion,normal physiological activities affected,and even toxic reactions.Yu-ping-feng Prescription(YPF)is composed of Astragalus mongholicus(AM),Largehead Atractylodes Rhizome(RA)and Divaricate Saposhnikovia Root(SD).It isan over-the-counter drug with the functions of benefiting Qi and firming the surface,preventing sweat and tonifying the spleen and stomach.Its main indications include allergic rhinitis and recurrent upper respiratory tract infections.Diseases such as spontaneous sweating.Objective(1)Animal level: The effect of YPF and decoction on the content of OAT1/3protein and mRNA in rat kidney tissue was observed by Western blot and RT-PCR.(2)Cell level: To investigate the expression of OAT1/3 protein and mRNA in human embryonic kidney cells(HEK-293)by YPF decoction,water decoction of each prescription and single active ingredient,so as to explore the effect of YPF on the activity and expression of OAT1/3 protein,and compare the effects of each component on the activity and expression of OAT1/3 protein.MethodAnimal experiment: YPF and each single decoction were given to rats for a long time to observe the activity and expression of OAT1/3 protein in kidney.Rats were randomly divided into 5 groups,6 rats in each group,namely saline group(NE group),YPF group,AM group,RA group and SD group.The rats were given orally for 21 days.The rats in NE group were replaced by 0.9% saline.On the 21 st day,rats in each group were treated and kidney tissues were collected.Western blot and RT-PCR were used to detect the expression of OAT1/3 protein and mRNA in renal tissue.Cell experiment:(1)MTT method was used to select the best concentration of HEK-293 in vitro.MTT method was used to detect different concentrations of YPF decoction,AM decoction,RA decoction,SD decoction and single active ingredient,,Prim-O-Glucosylcifumigin(POG),5-O-methylvisammioside(OM),Atractylenolide I(AT-I).and Atractylenolide III(AT-III)were incubated for 24 hours to detect their effects on the survival rate of HEK-293 cells,and MTT method was used to detect the effects of YPF on the survival rate of HEK-293 cells for 48 h and 72 h.(2)The expression of OAT1/3 protein and mRNA in HEK-293 cells was detected by Western blot and RT-PCR.Result1.YPF decoction can inhibit the expression of genes and proteins of OAT1 and OAT3 in kidney and the excretion of their substrates,which may be one of the main reasons for the changes of pharmacokinetic parameters of astragaloside IV in rats.2.YPF and SD decoction could reduce the expression level of OAT1/3 protein and mRNA in HEK-293 cells in varying degrees.RA decoction decreased the expression level of OAT1 protein and mRNA in HEK-293 cells,but increased the expression level of OAT3 protein and mRNA.3.AT-I(30?g/ml),POG(30?g/ml)and OM(50?g/ml)could significantly reduce the expression of OAT1/3 protein and mRNA in HEK-293 cells.AT-III(30?g/ml)decreased the expression of OAT1 protein and mRNA in HEK-293 cells,but significantly increased the expression of OAT3 protein and mRNA.Conclusion1.YPF decoction could inhibit the expression and activity of OAT1 and OAT3 proteins and the excretion of their substrates,which may be one of the main reasons for the elevated concentration of astragaloside IV in vivo.2.The inhibitory effect of YPF decoction on OAT1 and OAT3 protein of HEK-293 cells was time-dependent and dose-dependent.3.The inhibitory effect of some active ingredients in SD decoction may be one ofthe main reasons for the inhibition of OAT1 and OAT3 proteins in YPF decoction.There are also some components in RA that can inhibit the expression of OAT1 protein and enhance the expression of OAT3,while the AM decoction has no inhibitory effect on the expression of OAT1 and OAT3 protein.This result validates the theory of group ministers and envoys,and provides information for elucidating the mechanism of pharmacokinetic compatibility. |
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