Action target spot of glucocorticoid and application thereof

[0147] FOX O3a is a target of glucocorticoid action

[0148] Materials and methods

[0149] 6-8 weeks old Balb / C mice were purchased from the SPF animal room of Shanghai Institute of Biochemistry and Cells, Chinese Academy of Sciences

[0150] The siRNA of FoxO3a (5'GGAUGACGUCCAGGAUGAUTT 3' (forward translation strand SEQ ID NO: 2) and 5'AUCAUCCUGGACGUCAUCCTT 3' (reverse translation strand SEQ ID NO: 3) was purchased from GenePharma

[0151] Dexamethasone (DXM), purchased from sigma-aldrich company

[0152] method 1

[0153] Experimental group (group 3): Lymphocytes were obtained from the spleen of 6-8 week Balb / C mice, cultured in RPMI1640 medium and 10% fetal bovine serum, in a 6-well plate of lymphocyte suspension, about 10 per well 6 Dexamethasone was added to the cells at a concentration of 4 μM, 1 μM, and 0.1 μM;

[0154] Control group: ethanol solution without adding dexamethasone;

[0155] The cells were harvested after 6 hours, and the expression changes of FoxO

[0166] Sensitivity of cells to glucocorticoids in vitro

[0167] Materials and methods

[0168] Myeloma cell line SP2 / O was purchased from the cell bank of Shanghai Institute of Biochemistry and Cell, Chinese Academy of Sciences

[0169] Chronic lymphocytic leukemia cell line L1210 was purchased from the cell bank of Shanghai Institute of Biochemistry and Cell, Chinese Academy of Sciences

[0170] Gastric cancer cell line MFC was purchased from the cell bank of Shanghai Institute of Biochemistry and Cell, Chinese Academy of Sciences

[0171] Breast cancer cell line 4T1 was purchased from the cell bank of Shanghai Institute of Biochemistry and Cell, Chinese Academy of Sciences

[0172] Test group (4 groups):

[0173] SP2 / O group: culture SP2 / O in RPMI 1640 medium and 10% fetal bovine serum,

[0174] MFC group: MFC was cultured in RPMI1640 medium and 10% fetal bovine serum,

[0175] 4T1 group: 4T1 was cultured in RPMI1640 medium and 10% fetal bovine serum,

[0176] L121

[0185] Screening for glucocorticoids

[0186] Select the more sensitive tumor cell line SP2 / O, and culture SP2 / O in RPMI1640 medium and 10% fetal bovine serum,

[0187] Add the hormone to be screened, dexamethasone, prednisone, methylprednisolone, and hydrocortisone at 4 μM each to the selected tumor cell culture dish.

[0188] After 8-24 hours, the cells were collected for western blot detection, and the phospho-FoxO3a / FoxO3a ratio was selected to be low, preferably 0.2-0.3. The lower the value, the more sensitive the cells were to the hormone, and the stronger the anti-inflammatory activity of the hormone.