炎症微环境下芦丁对牙周膜干细胞成骨分化能力的影响

doi:10.19439/j.sjos.2019.04.004

摘要/Abstract

摘要： 目的研究芦丁（rutin）对炎症微环境下牙周膜干细胞成骨分化能力的影响。方法采用有限稀释法分离纯化获得牙周膜干细胞,采用流式细胞术鉴定牙周膜干细胞。以脂多糖（lipopolysaccharide, LPS）刺激牙周膜干细胞,建立体外炎症模型。实验分为4组,第1组使用α-MEM培养基培养牙周膜干细胞,第2组使用含有脂多糖的α-MEM培养基培养牙周膜干细胞,第3组在含有脂多糖的α-MEM培养基加入芦丁培养牙周膜干细胞,第4组使用含有芦丁的α-MEM培养基培养牙周膜干细胞。通过碱性磷酸酶染色、碱性磷酸酶活性测试、茜素红染色、RT-PCR以及蛋白免疫印迹等方法检测成骨分化能力的改变。采用SPSS 17.0软件包对数据进行统计分析。结果 CCK-8和碱性磷酸酶活性测试结果显示,10 μmol/L芦丁对炎症状态下牙周干细胞增殖和成骨分化作用最明显。碱性磷酸酶染色和茜素红染色结果显示,10 μmol/L芦丁可以改善炎症微环境下牙周膜干细胞的成骨分化能力。RT-PCR、蛋白免疫印迹结果显示,芦丁可以增强炎症状态下COL1、ALP、RUNX2等成骨基因和成骨蛋白的表达。结论芦丁可以增强炎症微环境下牙周膜干细胞的成骨分化能力。

关键词: 芦丁, 脂多糖, 牙周膜干细胞, 成骨分化

Abstract: PURPOSE: To explore the effect of rutin on osteogenic differentiation of periodontal ligament stem cells under inflammatory microenvironment. METHODS: Periodontal ligament stem cells (PDLSCs) were obtained by limited dilution method in vitro. PDLSCs were identified by flow cytometery. Lipopolysaccharide(LPS) was used to stimulate human periodontal ligament stem cells to establish an inflammation model in vitro. The experiment was divided into 4 groups: in group 1, only α-MEM was used to culture PDLSCs; in group 2, α-MEM medium containing LPS was used to culture PDLSCs, in group 3, rutin was added to α-MEM medium containing LPS to PDLSCs; and in group 4, α-MEM medium containing rutin was used to culture PDLSCs. Cell counting kit-8 was used to detect cell proliferation activity. Alkaline phosphatase(ALP) staining, ALP activity test, alizarin red staining, RT-PCR, and Western blot were used to detect the changes of osteogenic differentiation ability. The data were analyzed by SPSS 17.0 software package. RESULTS: The results of CCK-8 and ALP activity analysis showed that rutin at 10 μmol/L could significantly promote the proliferation and differentiation of periodontal stem cells under inflammatory state. ALP staining and alizarin red staining proved that (10 μmol/L) rutin could improve osteogenic differentiation of periodontal ligament stem cells under inflammatory microenvironment. RT-PCR and Western blot results showed that rutin could enhance the expression of osteogenic genes and proteins such as COL1, ALP, and RUNX2 under inflammatory state. CONCLUSIONS: Rutin can promote osteogenic differentiation of periodontal ligament stem cells under inflammatory microenvironment.

Key words: Rutin, Lipopolysaccharide, Periodontal ligament stem cells, Osteogenic differentiation

中图分类号:

R781.4

赵斌, 张云鹏, 徐欣. 炎症微环境下芦丁对牙周膜干细胞成骨分化能力的影响[J]. 上海口腔医学, 2019, 28(4): 356-361.

ZHAO Bin, ZHANG Yun-peng, XU Xin. Rutin promotes osteogenic differentiation of periodontal ligament stem cells under inflammatory microenvironment[J]. Shanghai Journal of Stomatology, 2019, 28(4): 356-361.

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