The Role And Mechanism Of DDR1a(DDR1a) In Mice With Exprimental Colitis

Objective :To explore the role and mechanism of DDR1 a on DSS-induced acute colitis.Methods:Screening for lentivirus: IEC-6 cells were transfected by three sets of DDR1a-shRNA lentivirus(i.e..DDR1a-Homo-1400,DDR1a-homo-1403 and DDR1a-Homo-1733)provided by the manufacturer,and expressions of DDR1 a were detected by Western blot.Animal experiment:32 SPF female Balb/c mice were randomly divided into 4 groups:normal control group(A),model group(B),treatment group(C),and negative control group(D).Mice in group A were free to drink distilled water,while mice in group B,C and D were fed with 4%DSS for 7 consecutive days to build the model of acute colitis,and continued to drink distilled water for3 days.Mice in group C were treated with DDR1a-shRNA lentivirus on the tail vein on the fourth day,whereas mice in group D were injected with scramble DDR1a-shRNA.All mice were sacrificed on the 11 th day.Disease activity index(DAI)was daily calculated.Colon tissues were removed and stained with haematoxylin and eosin(HE),and the histopathological score(HS)was evaluated.Expressions of DDR1 a and NF-?B p65 proteins were detected by Immunohistochemistry and Western-blot,and levels of TNF-?,IL-6 and IL-8 were detected by ELISA.Cell experiment:Intestinal epithelial cells(IEC-6)were randomly divided into 4 groups:normal control group(A),activation group(B),intervention group(C),and negative control group(D).Cells in intervention group and negative control group were successfully transfected with DDR1a-shRNA and scramble DDR1a-shRNA,respectively.Cells in group B,C and D were incubated with type I collagen for 2h.Expressions of DDR1 a and NF-?B p65 were detected by Western-blot analysis,and levels of TNF-? and IL-10 were detected by ELISA.Results:1.Screening for lentivirusWestern blot results showed that the expression of DDR1 a was down-regulated after transfected with DDR1a-homo-1403(p<0.05).2.Animal experiment1)The DAI score and TDI score of model group were significantly higher than those of normal control group and tteeatment group(p<0.05).2)Immunohistochemistry and Western-blot showed that expressions of DDR1 a and NF-?B p65 in model group were significantly higher than those in normal control group(p<0.05).The levels of DDR1 a and NF-?B p65 in treatment group were lower than those in model group(p<0.05).3)ELISA showed that the levels of TNF-?,IL-6 and IL-8 in model group were significantly higher than in normal control group(p<0.05).In contrast,the levels of TNF-?,IL-6 and IL-8 in treatment group were lower than in model group(p<0.05).3.Cell experiment1)Westernal-blot showed that expressions of DDR1 a and NF-?B p65 were significantly increased in the activiation group,and the expression of NF-?B p65 was decreased after the down-regulation of DDR1 a expression(p<0.05).2)ELISA showed that levels of TNF-?and IL-10 were significantly higher in activiation group than in normal control group and intervention group(p<0.05).Conclusion:1)DDR1a is involved in the pathogenesis of experimental colitis in mice.2)DDR1a may promote excessive release of inflammatory factors by activating NF-?B p65.