Construction and applications of cell model expressing human organic cation transporter-1

Abstract

The invention provides a construction method and applications of a cell model expressing human organic cation transporter-1. A hOCT1 wild-type gene segment is obtained from a hepatic tissue; two mutant gene segments P341L and M420del can be obtained by specific point mutation; the two segments are connected with a plasmid vector; darby canine kidney cells (MDCK) are transfected; G418 resistance screening is carried out to obtain wild-type cells expressing hOCT1 and cells of two mutants; mRNA level verification is carried out on the cells; and functional verification is carried out by utilizing a hOCT1 substrate and an inhibitor. The cell model provided by the invention can be used for screening the hOCT1 classic substrate and the inhibitor, and predicting the transportation of medicament in human bodies and medicament interaction which possibly happen; and by utilizing the cell model, the influence of gene polymorphism of the transporter on the medicament transportation function can be predicted; and the cell model provides a standard for clinical reasonable medicament administration and individualized medicament administration, and has reasonable design and good repetitiveness.

Description

technical field [0001] The invention belongs to the field of biological technology, and relates to the construction and application of Martin Darby dog ​​kidney epithelial cell (MDCK) model stably expressing wild-type and mutant human organic cation transporter 1 (hOCT1). technical background [0002] Membrane transporters are a class of proteins that are widely distributed on cell membranes and specifically mediate the entry and exit of exogenous and endogenous chemical substances through active or passive mechanisms. They generally have saturation and substrate specificity. Membrane transporters provide a new and promising direction for people to conduct drug research, especially in pharmacodynamics, pharmacokinetics, new drug development, and drug targeting design. The organic cation / anion / zwitterion transporter family (SLC22) plays an important role in the transport and clearance of many endogenous amines and exogenous substances, including many clinically used drugs. T...

AI Technical Summary

Problems solved by technology

The multi-specific organic cation transporter OCT1 (SLC22A1) in this family is a glycoprotein containing 554 amino acids, including 12 transmembrane regions, and the N-terminal and C-terminal are located inside the cell membrane. There is a large extracellular loop of glycosylation between the first transmembrane structures and a phosphorylation extracellular loop between the sixth and seventh transmembrane structures [1] , its three-dimensional structure is still unclear

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