PLGA-无机盐复合生物材料对牙胚细胞和牙髓干细胞增殖分化的影响
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摘要
由牙周病、龋齿、外伤或是遗传引起的牙齿缺失给大多数人的生活带来很大的不便。能够取代缺失的牙齿的、具有生物活性的组织工程牙齿将给目前的临床治疗带来新的技术。为了使具有生物活性的组织工程牙齿成为可能,找出适合用于进行组织工程牙齿塑形的生物支架材料,本研究比较了四种生物支架材料即聚乳酸-聚乙醇酸(PLGA)、聚乳酸-聚乙醇酸-磷酸三钙(PLGA/TCP)、聚乳酸-聚乙醇酸-羟基磷灰石(PLGA/HA)、聚乳酸-聚乙醇酸-碳酸钙-羟基磷灰石(PLGA/CDHA)对小鼠磨牙牙胚细胞和人牙髓干细胞增殖分化的影响,主要内容及结论如下:
1、利用细胞活性MTT检测方法,发现四种生物支架材料对Hela细胞的毒性为零级;对人牙髓干细胞(DPSCs)都有毒性,其中PLGA/CDHA的毒性最大;
2、牙胚细胞-生物支架材料复合物在小鼠肾囊膜下生长6周,细胞在PLGA/HA中增殖最多,在PLGA中增殖最少;DPSCs-生物支架材料复合物在裸鼠肾囊膜下生长5周,200um孔径PLGA/HA、320um孔径PLGA/CDHA和320um孔径PLGA/TCP的增殖率相当,PLGA增殖率最低的;
3、检测肾囊膜下生长不同时期的牙胚细胞-生物支架材料复合物和DPSCs-生物支架材料复合物的钙盐沉淀情况,发现PLGA/TCP和PLGA/CDHA的钙盐形成情况较好,PLGA/HA次之,PLGA最差;
4、细胞吸附性检测发现PLGA/HA吸附性最好,而且6周后吸附了牙胚细胞的复合物PLGA/CDHA和纯PLGA的外部有形成结构不完整的牙齿;
5、牙胚组织-生物支架材料复合物在小鼠肾囊膜下6周后均能成牙。
综合所有实验我们得出结论:PLGA/HA和PLGA/TCP有利于细胞增殖,PLGA/TCP和PLGA/CDHA有利于细胞分化,PLGA既不利于细胞增殖也不利于细胞分化。
Tooth loss due to periodontal disease, dental caries, trauma, or a variety of genetic disorders continues to affect most people adversely at some time in the lives. A biological tooth substitute that could replace lost teeth would provide a vital alternative to currently available clinical treatments.To pursue biological tooth and suit to its shape, I compared four different biomaterials,namely PLGA, PLGA/HA, PLGA/TCP, PLGA/CDHA,on the proliferation and differentiation of mouse molar tooth germ cells and postnatal human dental pulp stem cells(DPSCs). The content and results were as follows:
A. by cell activity (MTT assay),the results showed that to Hela cells all the cell cytotoxicity degree of biomaterials were zero degree.To DPSCs,the four different biomaterials were deleterious,and PLGA/CDHA was the most;
B.after tooth germ cells-biomaterial composites cultured on the renal sac of mice for six weeks,we found the proliferation of cells inside the PLGA/HA was highest of all,and the PLGA was the lowest;while the DPSCs-biomaterial composites, the proliferation of cells inside the PLGA/HA(200um),PLGA/CDHA(320um),PLGA/TCP(320um)was almost the same, and the PLGA was the lowest;
C.von kossa staining of tooth germ cells-biomaterials and DPSCs-biomaterials composites cultured for different time displayed a significant amount of mineral in PLGA/TCPand PLGA/CDHA composites;
D. the adsorption of cells of PLGA/HA was the best of the four biomaterials. And we found half-baked tooth structure in the cells-PLGA/CDHA and cells-PLGA composites cultured on the renal sac of mice for six weeks;
E. we could find integrated tooth structure in the tooth germ cells-biomaterials composites cultured on the renal sac of mice for six weeks.
In summary, PLGA/HA and PLGA/TCP were useful for cells to proliferate, PLGA/TCPand PLGA/CDHA are useful for cells to differentiate,while PLGA was not only unuseful for cells to proliferate but also unuseful for cells to differentiate.
1、利用细胞活性MTT检测方法,发现四种生物支架材料对Hela细胞的毒性为零级;对人牙髓干细胞(DPSCs)都有毒性,其中PLGA/CDHA的毒性最大;
2、牙胚细胞-生物支架材料复合物在小鼠肾囊膜下生长6周,细胞在PLGA/HA中增殖最多,在PLGA中增殖最少;DPSCs-生物支架材料复合物在裸鼠肾囊膜下生长5周,200um孔径PLGA/HA、320um孔径PLGA/CDHA和320um孔径PLGA/TCP的增殖率相当,PLGA增殖率最低的;
3、检测肾囊膜下生长不同时期的牙胚细胞-生物支架材料复合物和DPSCs-生物支架材料复合物的钙盐沉淀情况,发现PLGA/TCP和PLGA/CDHA的钙盐形成情况较好,PLGA/HA次之,PLGA最差;
4、细胞吸附性检测发现PLGA/HA吸附性最好,而且6周后吸附了牙胚细胞的复合物PLGA/CDHA和纯PLGA的外部有形成结构不完整的牙齿;
5、牙胚组织-生物支架材料复合物在小鼠肾囊膜下6周后均能成牙。
综合所有实验我们得出结论:PLGA/HA和PLGA/TCP有利于细胞增殖,PLGA/TCP和PLGA/CDHA有利于细胞分化,PLGA既不利于细胞增殖也不利于细胞分化。
Tooth loss due to periodontal disease, dental caries, trauma, or a variety of genetic disorders continues to affect most people adversely at some time in the lives. A biological tooth substitute that could replace lost teeth would provide a vital alternative to currently available clinical treatments.To pursue biological tooth and suit to its shape, I compared four different biomaterials,namely PLGA, PLGA/HA, PLGA/TCP, PLGA/CDHA,on the proliferation and differentiation of mouse molar tooth germ cells and postnatal human dental pulp stem cells(DPSCs). The content and results were as follows:
A. by cell activity (MTT assay),the results showed that to Hela cells all the cell cytotoxicity degree of biomaterials were zero degree.To DPSCs,the four different biomaterials were deleterious,and PLGA/CDHA was the most;
B.after tooth germ cells-biomaterial composites cultured on the renal sac of mice for six weeks,we found the proliferation of cells inside the PLGA/HA was highest of all,and the PLGA was the lowest;while the DPSCs-biomaterial composites, the proliferation of cells inside the PLGA/HA(200um),PLGA/CDHA(320um),PLGA/TCP(320um)was almost the same, and the PLGA was the lowest;
C.von kossa staining of tooth germ cells-biomaterials and DPSCs-biomaterials composites cultured for different time displayed a significant amount of mineral in PLGA/TCPand PLGA/CDHA composites;
D. the adsorption of cells of PLGA/HA was the best of the four biomaterials. And we found half-baked tooth structure in the cells-PLGA/CDHA and cells-PLGA composites cultured on the renal sac of mice for six weeks;
E. we could find integrated tooth structure in the tooth germ cells-biomaterials composites cultured on the renal sac of mice for six weeks.
In summary, PLGA/HA and PLGA/TCP were useful for cells to proliferate, PLGA/TCPand PLGA/CDHA are useful for cells to differentiate,while PLGA was not only unuseful for cells to proliferate but also unuseful for cells to differentiate.
引文
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2 Nerem RM. Tissue engineering: the hope, the hype, and the future. Tissue Eng, 2006,12:1143-50.
3 Vacanti CA. History of tissue engineering and a glimpse into its future. Tissue Eng, 2006,12:1137-42.
4 Langer R, Vacanti JP. Tissue engineering. Science, 1993,260:920-6.
5 Robert M. Nerem,Athanassios Sambanis. Tissue Engineering: From Biology to Biological Substitutes. Tissue Engineering, 1995,1:3-13.
6 Yilin Cao. The building and development of tissue engineering sciences. Journal of Tissue Engineering and Reconstructive Surgery, 2005,01:5-8.
7 Bonassar LJ, Vacanti CA. Tissue engineering: the first decade and beyond. J Cell Biochem Suppl, 1998,30-31:297-303.
8 Bayne SC. Dental biomaterials: where are we and where are we going?. J Dent Educ, 2005,69:571-85.
9 van der Kooy D, Weiss S. Why stem cells?. Science, 2000,287:1439-41.
10 Gronthos S, Brahim J, Li W, et al. Stem cell properties of human dental pulp stem cells. J Dent Res, 2002,81:531-5.
11 Rolletschek A, Blyszczuk P, Wobus AM. Embryonic stem cell-derived cardiac, neuronal and pancreatic cells as model systems to study toxicological effects. Toxicol Lett, 2004,149:361-9.
12 Reubinoff BE, Pera MF, Fong CY, et al. Embryonic stem cell lines from human blastocysts: somatic differentiation in vitro. Nat Biotechnol, 2000,18:399-404.
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