中药双黄补对人牙周膜细胞在玉米醇溶蛋白支架材料上黏附生长及增殖的影响

Effect of Shuanghuangbu on the Adhesion Growth and Proliferation of Human Periodontal Ligament Cells on the Zein Stent

目的研究中药双黄补对人牙周膜细胞(human periodontal ligament cells,HPDLCs)在玉米醇溶蛋白(Zein)支架材料上黏附生长及增殖的影响,探讨双黄补-Zein作为一种复合支架材料应用于牙周组织工程的可行性。方法采用组织块法体外培养HPDLCs,水提醇沉法制备双黄补煎液,溶剂浇铸/粒子沥滤法制备Zein支架,制备Zein支架浸提液并用其配制50、100、150、200、500、1000μg/mL浓度的双黄补培养液。分别设不同浓度双黄补组、Zein支架浸提液组及细胞对照组。MTT法检测各组对HPDLCs增殖的影响;根据MTT检测结果,选取最佳浓度(100μg/mL)双黄补作为实验组,10%FBS的DMEM培养液作为对照组,倒置相差显微镜及扫描电镜观察双黄补对HPDLCs在Zein支架材料上细胞形态的影响。结果与Zein支架浸提液组及细胞对照组比较,不同浓度双黄补均能促进HPDLCs的增殖,且以100μg/mL浓度的双黄补促增殖作用最明显(P<0·01);Zein支架浸提液组与细胞对照组比较差异无统计学意义(P>0·05),但其细胞增殖率为正值;形态学观察结果显示HPDLCs在Zein支架上生长良好,且在双黄补作用下Zein支架上的HPDLCs数目较多,形态较好。结论双黄补能促进HPDLCs在Zein支架上的黏附生长及增殖。双黄补-Zein作为一种复合支架材料应用于牙周组织工程具有可行性。

Objective To study the effect of Shuanghuangbu on the adhesion growth and proliferation of human periodontal ligament cells（HPDLCs） on the zein scaffold,and to study the feasibility of Shuanghuangbu zein used in the periodontal tissue engineering as a compound scaffold material.Methods HPDLCs were cultured in vitro using the explants adherent method.Shuanghuangbu Decoction was prepared using water extraction and alcohol precipitation method.Zein scaffold was prepared using solvent casting/corpuscle leaching method,which was dispensed into Shuanghuangbu culture fluid at the concentration of 50,100,150,200,500,1 000 μg/mL,respectively.Shuanghuangbu of different concentrations groups,the zein scaffold leaching liquid group,and the cell control group were also set up.The effect of each group on the proliferation of HPDLCs was detected by MTT.According to results of MTT,the best concentration of Shuanghuangbu（100 μg/mL） was selected as the experiment group.Ten percent FBS of DMEM culture fluid was taken as the control group.The effect of Shuanghuangbu on the morphology of HPDLCs on the zein scaffold was observed using inverted phase contrast microscope and scanning electron microscope.Results Compared with the zein scaffold leaching liquid group and the cell control group,the proliferation of HPDLCs could be promoted by different concentrations of Shuanghuangbu culture fluid,with the most obvious effect shown by Shuanghuangbu at 100 μg/mL（P0.01）.There was no obvious difference between the zein scaffold leaching liquid group and the cell control group（P0.05）,but their cell proliferation rates were positive values.The morphological results demonstrated that HPDLCs successfully grew on the zein scaffold.The number of HPDLCs under the action of Shuanghuangbu was more and the morphous was better.ConclusionsShuanghuangbu could promote the adhesion growth and proliferation of HPDLCs on the zein scaffold.Shuanghuangbu zein was feasible to be used in the periodontal tissue engineering as a compound scaffold materia.