人牙髓细胞与PLGA微球材料复合构建后的形态学观察  

Morphological Observation of Human Dental Pulp Cells after Compound Construction with PLGA Microsphere Material

作　　者：王冠华 邹慧儒 连小丽 代晓华 颜艳 张林朴 WANG Guanhua;ZOU Huiru;LIAN Xiaoli(Tianjin Key Laboratory of Oral and Maxillofacial Function Reconstruction&Tianjin Stomatological Hospital&School of Medicine,Nankai University,Tianjin City 300041)

机构地区：[1]天津市口腔功能重建重点实验室、天津市口腔医院、南开大学医学院,300041

出　　处：《医学理论与实践》2023年第12期1988-1990,1987,共4页The Journal of Medical Theory and Practice

基　　金：天津市卫生健康委科技人才培育项目(RC20105)。

摘　　要：目的:观察不同浓度的人牙髓细胞(HDPCs)在可注射聚乳酸/羟基乙酸(PLGA)微球支架上的黏附生长,通过形态学观察进一步探讨PLGA微球作为组织工程支架材料用于牙髓组织再生的可行性。方法:取第4代HDPCs以3种不同浓度(2×10^(5)细胞/ml、1×10^(5)细胞/ml、5×10^(4)细胞/ml)接种于PLGA微球,在不同的时间点应用倒置显微镜下观察细胞与材料复合生长的情况,体外培养8周后,终止培养,常规固定、石蜡包埋,组织学切片,分别应用免疫组织化学染色检测牙本质涎磷蛋白(DSPP)、牙本质基质蛋白(DMP-1)、Ⅰ型胶原(COLⅠ)三种蛋白表达情况,并对阳性率进行统计。同时应用扫描电镜观察超微结构。结果:通过显微镜特异性观察细胞在材料表面的黏附状况,结果显示随着时间的推移细胞的生长状况整体较为良好。免疫组化结果显示牙髓组织与HDPCs-微球复合体三种蛋白表达均为阳性,将细胞最高浓度组三种蛋白的阳性率进行统计学分析,结果显示无统计学差异(P>0.05)。在培养8周之后通过显微镜观察细胞和微球之间的融合度较高,但微球局部有凹陷。通过形态学观察细胞与PLGA微球材料复合生长良好,结合前期定量检测结果显示PLGA微球有利于牙髓细胞黏附生长。结论:支架材料PLGA微球能够应用在组织工程化牙髓—牙本质复合体的构建,但下一步需要对微球材料进行改性,希望能够更加全面的将其应用于组织工程牙髓—牙本质复合体的构建之中,这也在临床中有极其重要的应用价值和实践意义。Objective:To observe the adhesion growth of human dental pulp cells(HDPCs)on the scaffold of injectable polylactic acid/acetic acid/hydroxyacetic acid(PLGA)microsphere,and to further explore the feasibility of PLGA microsphere as tissue engineering scaffold material for dental pulp tissue regeneration by morphological observation.Methods:The 4th generation HDPCs were seeded on PLGA microspheres at 3 different concentrations(2×10^(5),1×10^(5),and 5×10^(4) cells/ml),and the composite growth of cells and materials was observed under an inverted microscope at different time points.After 8 weeks of culture in vitro,the culture was suspended,routinely fixed,embedded in paraffin,and histological sections were performed.Immunohistochemical staining was used to detect the expression of dentin sialophosphoprotein(DSPP),dentin matrix protein(DMP-1)and collagen typeⅠ(COLⅠ),and the positive rate was statistically analyzed.The ultrastructure was observed by scanning electron microscope.Results:The adhesion of cells to the material surface was observed under the microscope,and the results showed that the growth of cells was good over time.Immunohistochemical results showed that the expression of the three proteins in dental pulp tissue and HDPCs-microspheres complex was positive,and the positive rates of the three proteins in the highest cell concentration group were statistically analyzed(P>0.05).After 8 weeks of culture,the fusion degree between the cells and the microspheres was high,but the microspheres had a local depression.According to the morphological observation,the cells and PLGA microspheres grew well.Combined with the previous quantitative detection results,PLGA microspheres were conducive to the adhesion and growth of dental pulp cells.Conclusion:PLGA microspheres can be used in the construction of tissue engineered pulp-dentin complex.However,the modification of PLGA microspheres is needed in the next step.It is hoped that PLGA microspheres can be more comprehensively applied in the construction of tissue

关 键 词：人牙髓细胞 PLGA微球材料 牙髓—牙本质复合体 牙本质涎磷蛋白 牙本质基质蛋白 Ⅰ型胶原 

分 类 号：R781.3[医药卫生—口腔医学]