松油烯4醇的抑菌性及促进MC3T3-E1细胞成骨分化作用  

Terpinen-4-ol with antibacterial properties promotes osteogenic differentiation of MC3T3-E1 cells

作　　者：郭鹏达 刘珂珂 段昕 刘朝辉 张云涛[1] Guo Pengda;Liu Keke;Duan Xin;Liu Zhaohui;Zhang Yuntao(Department of Stomatology,Binzhou Medical University Hospital,Binzhou 256600,Shandong Province,China;Department of Stomatology,Zhoukou Central Hospital,Zhoukou 466099,Henan Province,China)

机构地区：[1]滨州医学院附属医院口腔科,山东省滨州市256600 [2]周口市中心医院口腔科,河南省周口市466099

出　　处：《中国组织工程研究》2023年第25期4006-4012,共7页Chinese Journal of Tissue Engineering Research

摘　　要：背景:有研究发现,松油烯4醇对于种植体周围炎相关致病菌表现出明显的抑菌活性,当应用于钛种植体表面时能有效减少表面生物膜形成,降低种植体周围炎的发生,但是有关其生物安全性的研究不多。目的:分析松油烯4醇对金黄色葡萄球菌、变异链球菌的抗菌效果,以及其对MC3T3-E1细胞生长与成骨分化的影响。方法:将松油烯4醇以不同体积分数溶于二甲基亚砜中,检测松油烯4醇溶液对金黄色葡萄球菌和变异链球菌的最低抑菌浓度及最低杀菌浓度,并采用抑菌圈实验检测1/2最低抑菌浓度、最低抑菌浓度及最低杀菌浓度松油烯4醇的抗菌效果。以不同浓度(0%,0.3%,0.2%,0.1%,均为体积分数)松油烯4醇的培养基或钙化诱导液处理MC3T3-E1细胞,采用CCK-8实验检测细胞的增殖活性,罗丹明-鬼笔环肽染色观察细胞骨架,碱性磷酸酶活性实验检测细胞分化能力,茜素红染色观察矿化结节形成,RT-PCR检测Runx2、骨钙素mRNA的表达。结果与结论:①松油烯4醇溶液对变异链球菌的最低抑菌浓度为0.1%、最低杀菌浓度为0.4%,对金黄色葡萄球菌的最低抑菌浓度为0.03%、最低杀菌浓度为0.125%;随着浓度的增加,松油烯4醇溶液对两种细菌的抑菌圈直径增加(P<0.05);②CCK-8实验显示,松油烯4醇可促进MC3T3-E1细胞的增殖,其中以浓度0.2%组最明显;③罗丹明-鬼笔环肽染色显示,松油烯4醇可促进MC3T3-E1细胞的黏附;④碱性磷酸酶活性与茜素红染色显示,松油烯4醇均可促进MC3T3-E1细胞的早期分化与矿化,其中以浓度0.2%组最明显;⑤RT-PCR检测显示,钙化诱导1 d时,0.3%,0.2%,0.1%组Runx2 mRNA的表达高于0%组(P<0.05);钙化培养7,14 d时,0.2%,0.1%组Runx2、骨钙素mRNA的表达高于0.3%,0%组(P<0.05);⑥结果表明,松油烯4醇可以抑制金黄色葡萄球菌及变异链球菌的生长,促进MC3T3-E1细胞的增殖、分化和矿化。BACKGROUND:Te rpinen-4-ol has been found to have a significant antibacterial activity against pathogenic bacte ria associated with peri-implantitis and can reduce the formation of surface biofilm and the occurrence of peri-impla ntitis when applied to the surfa ce of titanium implants;however,the studies regarding its biosafety are lacking.OBJECTIVE:To explore the antibacterial effects of terpinen-4-ol on Staphylococcus aureus and Streptococcus mutans and its effect on the growth and osteogenic differentiation of MC3T3-E1 cells.METHODS:Different concentrations of terpinen-4-ol were dissolved in dimethyl sulfoxide.The minimum inhibito ry concentration and minimum bactericidal concentration of pineolene 4 alcohol solution against Staphylococcus aureus and Streptococcus mutans were tested.The 1/2 minimum inhibitory concentration,minimum inhibitory concentration,and minimum bactericidal concentration of pineolene 4 alcohol were tested by bacteriostatic zone test.MC3T3-E1 cells were treated with different concentrations(0%,0.3%,0.2%,0.1%,v/v)of terpinen-4-ol in medium or calcification-inducing solution.The proliferation activity of cells was detected by CCK-8 assay.Cells were stained with rhodamine-phalloidin to observe cytos keleton.Diffe rentiation ability of cells was detected by alkaline phosphatase activity assay.The formation of mineralized nodules was observed by alizarin red staining.The mRNA expression of Runx2 and osteocalcin was detected by real-time reverse transcri ption-polymerase chain reaction.RESULTS AND CONCLUSION:(1)Minimum inhibito ry concentration and minimum bactericidal concentration of terpinen-4-ol against Streptococcus mutans were 0.1%(v/v)and 0.4%(v/v)respectively and against Staphylococcus aureus were 0.03%(v/v)and 0.125%(v/v)respectively.With the increase of the concentration of terpinen-4-ol,the diameter of bacte riostatic zone was significantly increased(P<0.05).(2)CCK-8 assay showed that terpinen-4-ol promoted the prolife ration of MC3T3-E1 cells,with the most obvious effects in the 0.2

关 键 词：松油烯4醇 金黄色葡萄球菌 变异链球菌 MC3T3-E1细胞 骨钙素OCN RUNX2 

分 类 号：R459.9[医药卫生—治疗学] R313[医药卫生—临床医学] R782.12