氟染毒对大鼠皮肤成纤维细胞中SOST及DKK1蛋白表达的影响
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摘要
[目的]观察氟染毒对体外培养新生大鼠皮肤成纤维细胞硬骨素(sclerostin,SOST)和Dickkopf-1(DKK1)蛋白表达的影响。[方法]采用原代培养方法,将大鼠皮肤成纤维细胞分为对照组(0 mg/L)和7个染氟剂量组(0.000 1、0.001、0.01、0.1、1、10、20 mg/L),分别在4个染氟时间段(24、48、72、96 h)收集细胞培养上清液,应用ELISA法测定SOST和DKK1蛋白表达水平。[结果]与染氟0 mg/L组相比,SOST表达量在染氟48 h的0.000 1、0.001、0.01 mg/L组,染氟72 h的1、10 mg/L组及染氟96 h的0.001、0.01、0.1、1 mg/L组均明显下降(P<0.05)。与染氟24 h组相比,SOST表达量在染氟48 h的0.01 mg/L组,染氟96 h的0.001、0.01、0.1、1、10 mg/L组均明显下降(P<0.05)。与染氟0 mg/L组相比,DKK1表达量在染氟48 h的0.001 mg/L组,染氟72 h的0.01、0.1、1、10、20 mg/L组及染氟96 h的0.01、0.1、1、10、20 mg/L组均明显下降(P<0.05)。与染氟24 h组相比,DKK1表达量在染氟48 h的0.000 1、0.001、0.01 mg/L组,染氟72 h的0.001、0.01、1、10、20 mg/L及染氟96 h的0.01、0.1、1、20 mg/L组均明显下降(P<0.05)。[结论]氟可导致大鼠皮肤成纤维细胞SOST和DKK1蛋白表达下调。
[Objective] To observe the changes of protein expressions of sclerostin(SOST) and dickkopf-1(DKK1) in fibroblasts of rat skin following fluoride exposure.[Methods] Rat skin fibroblasts were cultured with fluoride at different 0, 0.000 1, 0.001, 0.01, 0.1, 1, 10, and 20 mg/L. The expression levels of SOST and DKK1 in cell culture supernatants were detected at four exposure time points(24, 48, 72, and 96 h) by ELISA. [Results] The expression levels of SOST of the 0.000 1, 0.001, 0.01 mg/L fluoride groups at 48 h, the 1 and 10 mg/L fluoride groups at 72 h, and the 0.001, 0.01, 0.1, and 1 mg/L fluoride groups at 96 h were significantly lower than that of the 0 mg/L fluoride group(P < 0.05). The expression levels of SOST of the 0.01 mg/L fluoride group at 48 h and the 0.001, 0.01, 0.1, 1, and 10 mg/L fluoride groups at 96 h were significantly lower than those of the groups at 24 h(P < 0.05). The expression levels of DKK1 of the 0.001 mg/L fluoride group at 48 h, the 0.01, 0.1, 1, 10, and 20 mg/L fluoride groups at 72 h, and the 0.01, 0.1, 1, 10, and 20 mg/L fluoride groups at 96 h were significantly lower than that of the 0 mg/L fluoride group(P < 0.05). The expression levels of DKK1 of the 0.000 1, 0.001, 0.01 mg/L fluoride groups at 48 h, the 0.001, 0.01, 1, 10, and 20 mg/L fluoride groups at 72 h, and the 0.01, 0.1, 1, and 20 mg/L groups at 96 h were significantly lower than those of the groups at 24 h(P < 0.05).[Conclusion] Fluoride can cause down-regulation of expression of SOST and DKK1 in rat skin fibroblasts.
[Objective] To observe the changes of protein expressions of sclerostin(SOST) and dickkopf-1(DKK1) in fibroblasts of rat skin following fluoride exposure.[Methods] Rat skin fibroblasts were cultured with fluoride at different 0, 0.000 1, 0.001, 0.01, 0.1, 1, 10, and 20 mg/L. The expression levels of SOST and DKK1 in cell culture supernatants were detected at four exposure time points(24, 48, 72, and 96 h) by ELISA. [Results] The expression levels of SOST of the 0.000 1, 0.001, 0.01 mg/L fluoride groups at 48 h, the 1 and 10 mg/L fluoride groups at 72 h, and the 0.001, 0.01, 0.1, and 1 mg/L fluoride groups at 96 h were significantly lower than that of the 0 mg/L fluoride group(P < 0.05). The expression levels of SOST of the 0.01 mg/L fluoride group at 48 h and the 0.001, 0.01, 0.1, 1, and 10 mg/L fluoride groups at 96 h were significantly lower than those of the groups at 24 h(P < 0.05). The expression levels of DKK1 of the 0.001 mg/L fluoride group at 48 h, the 0.01, 0.1, 1, 10, and 20 mg/L fluoride groups at 72 h, and the 0.01, 0.1, 1, 10, and 20 mg/L fluoride groups at 96 h were significantly lower than that of the 0 mg/L fluoride group(P < 0.05). The expression levels of DKK1 of the 0.000 1, 0.001, 0.01 mg/L fluoride groups at 48 h, the 0.001, 0.01, 1, 10, and 20 mg/L fluoride groups at 72 h, and the 0.01, 0.1, 1, and 20 mg/L groups at 96 h were significantly lower than those of the groups at 24 h(P < 0.05).[Conclusion] Fluoride can cause down-regulation of expression of SOST and DKK1 in rat skin fibroblasts.
引文
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[11]Pan L,Shi X,Liu S,et al.Fluoride promotes osteoblastic differentiation through canonical Wnt/β-catenin signaling pathway[J].Toxicol Lett,2014,225(1):34-42.
[2]Shi Y C,Worton L,Estehen L,et al.Effects of continuous activation of vitamin D and Wnt response pathways on osteoblastic proliferation and differentiation[J].Bone,2007,41(1):87-96.
[3]Yamabuki T,Takano A,Hayama S,et al.Dikkopf-1 as a novel serologic and prognostic biomarker for lung and esophageal carcinomas[J].Cancer Res,2007,67(6):2517-2525.
[4]Balemans W,Van Hul W.Human genetics of SOST[J].J Musculoskelet Neuronal Interact,2006,6(4):355-356.
[5]李长城.SOST/DKK1调控Wnt通路在氟性骨损伤发生中作用的实验研究[D].武汉:华中科技大学,2012.
[6]Bourhis E,Wang W,Tam C,et al.Wnt antagonists bind through a short peptide to the firstβ-propeller domain of LRP5/6[J].Structure,2011,19(10):1433-1442.
[7]Bodine P V,Zhao W,Kharode Y P,et al.The Wnt antagonist secreted frizzled-related protein-1 is a negative regulator of trabecular bone formation in adult mice[J].Mol Endocrinol,2004,18(5):1222-1237.
[8]井玲.氟化物刺激成纤维细胞成骨表型表达及其在氟骨症骨周化骨发生机制中的作用[D].长春:吉林大学,2006.
[9]魏海峰.染氟成纤维细胞和成骨细胞IGF-1及b FGF的表达及其意义[D].长春:吉林大学,2007.
[10]刘晓利,李长城,刘克俭,等.氟对成纤维细胞核心结合因子α1及增殖活性量-效关系的影响[J].中国组织工程研究,2012,16(28):5232-5236.
[11]Pan L,Shi X,Liu S,et al.Fluoride promotes osteoblastic differentiation through canonical Wnt/β-catenin signaling pathway[J].Toxicol Lett,2014,225(1):34-42.