Application of all-trans retinoic acid in the preparation of anti-porcine infectious gastroenteritis virus drugs
A technology of all-trans retinoic acid and infectivity, applied in the field of application of all-trans retinoic acid in the preparation of anti-porcine transmissible gastroenteritis virus drugs, can solve the problem of no effective drugs for TGEV infection, and achieve toxicity Less side effects, easy metabolism and absorption effect
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Embodiment 1
[0031] Example 1 Determination of cytotoxicity of all-trans retinoic acid on IPEC-J2 cells
[0032] IPEC-J2 cells were counted and diluted to an appropriate density with DMEM-F12 nutrient solution containing 10% fetal bovine serum (FBS), and then the cells were counted at 1×10 4 The concentration of cells / well was added to a 96-well cell culture plate, 100 μL per well, at 37°C, 5% CO 2 After the cells were grown to 70-80% density, IPECs were treated with different concentrations (0, 1, 10, 20, 40, 60, 80, 100, 200 μM / L) of all-trans retinoic acid. - J2 cells (all-trans retinoic acid was solubilized with dimethyl sulfoxide (DMSO), and the same volume of all-trans retinoic acid in DMSO containing different concentrations of all-trans retinoic acid was added to the medium for each treatment group), each Process 8 replicates. After 48 hours of drug action, 10 μL of CCK8 working solution was added to each well, and the culture was continued for 2 hours. Then, the 96-well plate wa...
Embodiment 2
[0034] Example 2 All-trans retinoic acid inhibits porcine transmissible gastroenteritis virus infection on IPEC-J2 cells (flow cytometry and fluorescence quantitative PCR)
[0035] Transfer IPEC-J2 cells to 1 x 10 5 The concentration of 1 / well was seeded in 12-well plate, placed at 37°C, 5% CO 2 Culture in an incubator, when the cells grow to about 80%, discard the old culture medium, and pretreat IPEC-J2 with all-trans retinoic acid at different concentrations (0, 1, 10, 20, 40, 60, 80 μM / L). 12h after the cells were inoculated with 1MOI (multiplicity of infection) of porcine transmissible gastroenteritis virus in IPEC-J2 cells, 37°C, 5% CO 2Incubate in the incubator for 1 h in the presence of all-trans retinoic acid, discard the old culture medium, wash 3 times with PBS, add all-trans retinoic acid corresponding to the previous concentration to each well, place at 37°C, 5% CO 2 After culturing in the incubator for 36 h, the cell samples were collected.
[0036] The cell s...
Embodiment 3
[0038] Example 3 The effect of all-trans retinoic acid on the infection process of porcine transmissible gastroenteritis virus
[0039] 1. The effect of all-trans retinoic acid on preconditioning of IPEC-J2 cells infected with porcine transmissible gastroenteritis virus
[0040] Transfer IPEC-J2 cells to 1 x 10 5 The concentration of 1 / well was seeded in 12-well plate, placed at 37°C, 5% CO 2 Culture in an incubator, when the cells grow to about 80%, discard the old culture medium, pre-treat IPEC-J2 cells with different concentrations (0, 60, 80 μM / L) of all-trans retinoic acid for 12 h, and wash with PBS for 3 and then inoculated IPEC-J2 cells with 1MOI of porcine transmissible gastroenteritis virus at 37°C, 5% CO 2 After 1 h of incubation in the incubator, wash three times with PBS, replace with DMEM-F12 nutrient solution without all-trans retinoic acid, and place at 37°C, 5% CO. 2 After culturing in the incubator for 36 h, the cell samples were collected.
[0041] The c...
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