| Objective: To explore the influence of ADAMTS-1on fibroblast proliferationand the expression of collagen type I in vitro, and investigate its possible molecularmechanism.Methods: Mouse fibroblasts were cultured in vitro, and stimulated byADAMTS-1. Our experiments were divided into two parts. The first part aimed to studythe effects of ADAMTS-1on fibroblast proliferation and collagen type I expression.Depending on different concentrations, they were divided into four groups: normalcontrol group, low concentration group(1nM), medium concentration group(10nM), andhigh concentration group(20nM). After24hours, the fibroblast proliferation wereobserved by thiazolyl blue tetrazolium bromide test, and collagen type I expression wasmeasured by Masson staining, reverse transcription-polymerase chain reaction andWestern blot. Mouse fibroblasts were treated with20nM ADAMTS-1for6hours,12hours,24hours respectively. Collagen type I expression was detected by reversetranscription-polymerase chain reaction. The second part aimed to explore its possiblemechanism. The Mouse fibroblasts were administered with different concentrations ofADAMTS-1(0nM,1nM,10nM,20nM) for24hours. The expression of TGF-β1, AKTand pAKT were measured by reverse transcription-polymerase chain reaction andWestern blot.Results:1. the growth of mouse fibroblastsThe growth rate of the fibroblasts gradually slowed with the increase ofADAMTS-1concentrations at the same time. Though fibroblasts grown in differentconcentration with time, the growth trend gradually slowed with the increase of ADAMTS-1concentrations.2. The expression of collagen in mouse fibroblastsCompared with the normal control group, the level of collagen type I decreasedsignificantly in low concentration group, medium concentration group and highconcentration group, especially in high concentration group (P<0.05). The mean densityof collagen were89.54±11.02,57.71±10.26,45.68±3.50and34.19±3.68in normalcontrol group, low concentration group, medium concentration group, and highconcentration group. The mRNA expression of COLI were （12.59±4.01）%,（11.91±4.05）%,（7.02±2.32）%and（3.28±1.13）%in normal control group, lowconcentration group, medium concentration group, and high concentration grouprespectively. The protein expression of COL I in normal control group, low concentrationgroup, medium concentration group, and high concentration group respectively were（90.95.±12.25）%,（56.99±4.74）%（,37.39±6.32）%,（18.19±4.05）%. Compared with6hours, COLI mRNA level was decreased at12hours and24hours（P<0.05）. The COLImRNA expression were （65.63±1.89）%,（42.41±4.19）%,（28.39±3.16）%at6hours,12hours,24hours.3. The influence on TGF-β1in mice fibroblastsCompared with the normal control group, the level of TGF-β1decreasedsignificantly in other three groups, especially in high concentration group (P<0.05). TheTGF-β1mRNA expression were（35.58±2.00）%,（19.65±1.96）%,（17.02±1.60）%,（9.02±0.93）%in normal control group, low concentration group, medium concentrationgroup, and high concentration group respectively. The TGF-β1protein expression were（90.07±2.19）%,(68.63±7.3)%,(51.75±2.72)%,(31.24±5.11)%in normal control group,low concentration group, medium concentration group, and high concentration group.4. The influence on PI3K/AKT in mice fibroblastsThe AKT expression had no significant differences（P>0.05）,whereas the pAKTexpression decreased （P<0.05）, The pAKT protein expression were (72.32±4.41)%,(50.20±6.47)%,(39.77±2.58)%,(29.24±5.08)%in normal control group, lowconcentration group, medium concentration group, and high concentration group respectively.5. The correlation of COLI and TGF-β1The expression of COLI were positive correlation with the expression ofTGF-β1(P<0.01).Conclusion:1. ADAMTS-1may inhibit the growth of fibroblasts.2. ADAMTS-1can down regulate the expression of collagen type I in fibroblast.3. ADAMTS-1can degrade collagen type I by inhibiting TGF-β1and itsdownstream signaling pathways PI3K/AKT. |
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