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牙龈卟啉单胞菌超声提取物对小鼠成骨细胞骨向分化蛋白表达的影响 被引量：1

Effect of sonicated extracts of Porphyromonas gingivalis on osteogenic differentiation of mouseosteoblasts
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摘要 目的研究牙龈卟啉单胞菌（ Porphyromonas gingivalis , Pg）W83超声提取物对小鼠颅顶前骨细胞亚克隆14（MC3T3-E1）骨向分化相关蛋白表达的影响，探讨Pg超声提取物对细胞成骨功能的作用。方法标准厌氧环境下培养PgW83，收集细菌沉淀，离心后制备细菌的超声提取物，然后以0（对照组）、10、100及1000mg／L的质量浓度加人MC3T3-E1的成骨诱导分化培养基中，培养14d后提取细胞总蛋白，蛋白质印迹法检测细胞骨钙蛋白、骨涎蛋白、骨桥蛋白及骨粘连蛋白的表达改变，酶标仪法检测碱性磷酸酶（alkaline phosphatase，ALP）的表达；培养21d后茜素红染色观察MC3T3-E1矿化结节形成情况；以上实验均重复3次，应用SPSS13．0统计软件对各组结果进行单因素方差分析，检验水准为双侧α=0．05。结果Pg超声提取物作用于MC3T3E114d后，100mg／L实验组细胞表达骨粘连蛋白、骨钙蛋白的相对表达量（分别为0．141±0．023、0．625±0．451）均显著低于对照组（均为1．000±0．000）和10mg／L组（分别为1．035±0．133、0．852±0．110）；1000mg／L组细胞表达骨粘连蛋白、骨钙蛋白的相对表达量（分别为0．020±0．003、0．213±0．053）均显著低于对照组、10及100mg／L组（P〈0．05），Pg超声提取物呈剂量依赖性显著下调骨粘连蛋白和骨钙蛋白的表达。1000mg／L高浓度时，Pg超声提取物显著下调骨桥蛋白的表达（蛋白相对表达量为0．572±0．162），与对照组、10及100mg／L实验组相比骨桥蛋白表达（分别为1．000±0．000、1．029±0．135、1．199±0．337）差异均有统计学意义（P〈0．05）。赡超声提取物不改变骨涎蛋白的表达，对照组、10、100及1000mg／L组间骨涎蛋白的相对表达量（分别为1．000±0．000、0．831±0．182、0．897±0．115、0．778±0．235）差异无统计学意义（P〉0．05）。Pg超声提取物抑制MC3T3-E1的ALP活性，其抑制作用随各实验组质量浓度的增加逐渐增强，10、100、1000mg／L质量浓度组ALP活性[分别为（0．0140±0．0011）、（0．0057±0．0013）及（0．0020±0．0008）U／gprot]均显著低于对照组[（0．0275±0．0014）U／gprot]（P〈0．05），同时Pg超声提取物作用于MC3T3-E121d后能明显抑制MC3T3-E1矿化结节的形成。结论Pg超声提取物可以下调小鼠成骨细胞骨向分化蛋白的表达，抑制细胞的成骨功能。 Objective To investigate the effects of sonicated extracts of Porphyromonas gingivalis （Pg） on osteogenic differentiation of mouse osteoblast cell line MC3T3-E1. Methods PgW83 was cultured under standard anaerobic conditions and extracted by sonication. Mouse osteoblast cell line MC3T3-E1 was cultured with various concentrations of the extraction （0,10,100,1000 rag/L）. Western blotting was applied to investigate the expression of osteocalcin（OC）, bone sialoprotein（BSP）, osteopo ntin（OPN）and osteonectin（ON）. The activity of alkaline phosphatase （ALP） was detected by microplate reader after 14 days. Mineralization nodule formation was measured by alizarin red staining after 21 days. Results Compared with the control group, the extracts of Pg decreased OC and ON expression in a dose-dependent manner（ OC relative expression： 1. 000 ± 0. 000,0. 852 ± 0. 110,0. 625 ± 0. 451,0. 213 ± 0. 053 ）, （ ON relative expression： 1. 000 ± 0. 000, 1. 035 ± 0. 133,0. 141 ± 0. 023,0. 020 ± 0. 003 ） （ P 〈 0. 05 ）. Theexpression of OPN was down-regulated significantly in MC3T3-E1 treated with 1000 mg/L extraction （0. 572 ± 0. 162） compared with control group, 10 and 100 mg/L （1. 000 ± 0. 000,1. 029 ± 0. 135,1. 199 ± 0. 337 ） （P 〈 0. 05 ）. The expression of BSP remained unchanged when the cells were cultured with or without extraction （ BSP relative expression ： 1. 000 ± 0. 000,0. 831 ± 0. 182,0. 897 ± 0. 115,0. 778 ± 0. 235 ） （ P 〉 0. 05 ）. Meanwhile, the extracts of Pg decreased ALP activity [ control group： （0. 0275 ± 0. 0014） U/gprot, 10 rag/L： （0. 0140 ±0. 0011 ） U/gprot, 100 mg/L： （0. 0057 ±0. 0013） U/gprot, 1000 mg/L： （0. 0020 ± 0. 0008） U/gprot ] （ P 〈 0. 05 ） and reduced mineralization nodule formation. Conclusions The results suggest that Pg may inhibit osteoblasts＇osteogenic function by down-regulation of osteogenic differentiation related proteins.
作者 张剑英 俞少杰 付云
机构地区 中山大学光华口腔医学院-附属口腔医院牙周科·广东省口腔医学重点实验室
出处 《中华口腔医学杂志》 CAS CSCD 北大核心 2013年第7期398-402,共5页 Chinese Journal of Stomatology
关键词 紫单胞菌 龈 成骨细胞 碱性磷酸酶 骨向分化 Porphyromonas gingivalis Osteoblasts Alkaline phosphatase Osteogenic differentiation
分类号 R78 [医药卫生—口腔医学]
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参考文献20

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共引文献16

同被引文献32

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中华口腔医学杂志
2013年 第7期

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