摘要
目的:探索微模式培养控制和导电基底对人牙周膜干细胞和骨髓间充质干细胞生物电特性的影响.方法:通过光刻技术在盖玻片和导电玻璃两种透明材料表面构造与细胞自身宽度相当的微模式化培养基底,微模式图案分为两种:单种细胞的单格阵列结构、双细胞共培养的交叉指贯通结构.应用全细胞膜片钳技术记录两种细胞在不同培养模式下的静息电位、钾通道电流等生物电特性.实验分9组:培养皿对照组、盖玻片牙周膜干细胞组、盖玻片骨髓间充质干细胞组、盖玻片共培养牙周膜干细胞组、盖玻片共培养骨髓间充质干细胞组、导电玻璃牙周膜干细胞组、导电玻璃骨髓间充质干细胞组、导电玻璃共培养牙周膜干细胞组、导电玻璃共培养骨髓间充质干细胞组.结果:对于牙周膜干细胞,与对照组自由铺展培养模式相比,微模式化形态控制导致细胞膜电容减小,钾通道电流密度增大(P＜0.01),导电基底导致细胞静息膜电位绝对值增大;两种微模式化培养模式相比,双细胞交叉指贯通培养使得细胞静息膜电位绝对值减小.对于骨髓间充质干细胞,与对照组自由铺展培养模式相比,导电基底表面细胞膜电容增大、钾通道电流密度减小和静息膜电位绝对值减小,但双细胞交叉指贯通培养则导致细胞膜电容减小、钾电流密度增大和静息膜电位绝对值增大;与盖玻片共培养模式相比,导电玻璃共培养细胞的钾电流密度显著增大(P＜0.01).结论:导电培养微模式能够调控人牙周膜和骨髓间充质干细胞的生物电特性,从利用内源电场角度,为深入探索两种干细胞促进口腔软硬组织再生修复机制提供新的思路.
Abstract
Objective:To explore the influence of micropatterned culture mode control and conductive substrate on the bioelectrical characteristics of the human periodontal ligamant stem cells(PDLSCs)and bone marrow mesenchymal stem cells(BMSCs).Methods:By using photolithography technology,the micro-patterned culture substrates with the width equivalent to the cell itself were constructed on the surface of two transparent materials,coverslip and ITO conductive glass.The micropatterns were divided into two types:single frame for single-type cell and interconnected interdigitated structure for two co-cultured cells.And the whole-cell patch clamp technique was used to record the bioelectrical characteristics such as resting membrane potential and potassium current of two types of cells under different culture modes.The cells were divided into 9 groups:culture dish as control group,PDLSCs on coverslip,BMSCs on coverslip,PDLSCs on coverslip with co-cultured structure,BMSCs on coverslip with co-cultured structure,PDLSCs on ITO,BMSCs on ITO,PDLSCs on ITO with co-cultured structure,BMSCs on ITO with co-cultured structure.Results:For PDLSCs,compared with the control group,micropatterned morphology control resulted in a decrease in cell membrane capacitance,an increase in potassium channel current density(P＜0.01),conductive substrate increased the absolute resting membrane potential;between the two micro-patterned modes,two cells co-cultured on the interconnected interdigitated structure resulted in a decrease in the absolute value of cell resting membrane potential.For BMSCs,compared with the control,micropatterned morphology control and conductive substrate induced an increase in cell membrane capacitance,a decrease in potassium channel current density,and a decrease in absolute resting membrane potential.However,two cells co-cultured on the interconnected interdigitated structure resulted in a decrease in cell membrane capacitance,an increase in potassium current density,and an increase in absolute resting membrane potential;compared with the coverslip co-cultured mode,the potassium current density of the cells co-cultured on ITO significantly increased(P＜0.01).Conclusion:Conductive culture micromode could change the bioelectric characteristics of human PDLSCs and BMSCs.From the perspective of using the endogenous electric field,it provided new ideas for the in-depth exploration of two stem cells to promote the regeneration and repair mechanism of oral soft and hard tissue.
基金项目
北京市自然科学基金预探索项目资助课题(7163215)
维普
万方数据