DRUG SCREENING METHOD, DRUGS PROMOTING EXTRACELLULAR MATRIX CROSSLINKING AND THE APPLICATIONS THEREOF

摘要

The present invention discloses a drug screening method, drugs promoting extracellular matrix protein crosslinking and their applications, the said drug screening method, is used to screen out materials promoting the expression of LOXL1 gene, wherein, the said drug screening method contains the following steps: A, Construct the 2nd generation lentiviral vector used to control the ZsGreen expression by human LOXL 1 gene promoter. B, Infect human fibroblasts with the 2nd generation lentiviral vector, and construct the new human fibroblasts which integrate PLOXL1-ZsGreen components. C, Drug screening: Inoculate the said human fibroblasts integrating PLOXL1-ZsGreen components into culture medium. Add the analyte into the cell culture medium containing human fibroblast cells. After culturing, detect the green fluorescence intensity of these fibroblast cells, then decide if the analyte promotes LOXL1 gene expression by checking if the green fluorescence intensity is increased. The method provided by the present invention is easy to operate and widely applicable.

主权项

1. A drug screening method used to screen out the materials promoting the expression of LOXL1 genes, wherein, the said drug screening method comprises the following steps:
A. Construct the 2nd generation lentiviral vector used to control the ZsGreen expression by human LOXL 1 gene promoter: Cloning the human LOXL 1 gene promoter fragment into lentiviral vector, pLVX-ZsGreen, substituting its original CMV promoter, achieving the 2nd generation lentiviral vector, pLenti-PLOXL1-ZsGreen, which contains a component of PLOXL1-ZsGreen, used to control the ZsGreen expression through human LOXL1 gene promoter. B. Infect human fibroblasts with the 2nd generation lentiviral vector, and construct the new human fibroblasts which integrate PLOXL1-ZsGreen components. C. Drug screening: Inoculate the said human fibroblasts integrating PLOXL1-ZsGreen component into culture medium, and incubate overnight. Dissolve the analyte into DMSO and add into the cell culture medium containing human fibroblast cells. After culturing, detect the green fluorescence intensity of these fibroblast cells, then decide if the analyte promotes LOXL1 gene expression by checking if the green fluorescence intensity is increased.