The Effect And Mechanism Of ADAMTSs-1in Mouse’ Pulmonary Fibrosis Induced By Bleomycin

Objective:To investigate the effect of a disintegrin-like and metalloproteinasewith thrombospondin type1motif(ADAMTS-1) on mouse fibroblaststreated with Bleomycin and the potential mechanism.Methods:1. Establish ment of the model of pulmonary fibrosis: mousefibroblasts were cultured. they were dealed with10ug/ml bleomycinfor24hours, collagen expression was observed by Masson staining.2. Mouse fibroblasts were randomly divided into6groups: controlgroup, Bleomycin group, Bleomycin+1μg ADAMTS-1group,Bleomycin+2μg ADAMTS-1group, Bleomycin+4μg ADAMTS-1group, and4μg ADAMTS-1group. To join without fetal bovine serumstarved cells culture medium after24hours, the group1and group6join not containing fetal bovine serum medium for2ml. The othergroups were combined with bleomycin (1mg/ml) and without20ul fetalbovine serum medium for1.98ml. In sterile temperature box24hoursof training,24hours after the group3with1ug ADAMTS;4groupwith2ug ADAMTS;5group with4ug ADAMTS4ug ADAMTS into6groups. Then at37℃,5%CO2cell culture box24hours.3. The cell supernatant was collected in every group, and Theconcentrations of CTGF and COLI were dected by ELISA. Theexpression of CTGF and COLI mRNA and protein was examined byreal-time quantitative PCR and Westem blot,respectively.Results:1. Successful establishment of fibroblast fibrosis model.Compared with the normal mouse lung fibroblasts, bleomycin induced group fibroblast cell morphology degree increased.2. ELISA results showed that bleomycin group (group2)concentration in the supernatant of cell culture medium CTGF, COLⅠ was significantly higher than that in the other groups; inExperiment3,4,5groups with the increase of ADAMTS-1content inthe supernatant of cell culture medium CTGF, COLⅠ concentrationdecreasing; ADAMTS-1group (Group6) cell culture the lowestconcentration in the supernatant of CTGF based, COL I (P<0.05).3. The detection results using real-time quantitative PCR showedthat BLM group (group2) cells in CTGF, COLⅠ mRNA expressionwas the highest; in Experiment3,5groups of4, with the increase ofADAMTS-1content in cells CTGF, COL I mRNA expressiondecreasing; group ADAMTS-1(6) cells in CTGF, COLⅠ mRNA theexpression level was the lowest (P<0.05).4. Western Blot testing results showed that bleomycin group(group2) cells in CTGF, COL Ⅰ protein expression was the highest;in Experiment3,5groups of4, with the increase of ADAMTS-1content in cells CTGF, COL Ⅰ protein expression quantity wasdecreased; the ADAMTS-1group (Group6) cells in CTGF, COL Ⅰprotein expression was the lowest (P<0.05).Conclusion:ADAMTS-1can down regulate the expression of COLI andCTGF in mouse fibroblasts treated with bleomycin, the degree ofdecrease is generally in proportion to its dose.