脱细胞真皮基质对年轻恒牙牙乳头干细胞分化的作用

摘要/Abstract

摘要： 目的：获取年轻恒牙牙乳头组织，进行细胞培养以及与脱细胞真皮基质（acellular dermal matrix, ADM）的复合培养，了解ADM在干细胞成骨、成脂转化中的作用。方法：利用口腔外科手段获取年轻恒牙牙乳头组织，分离培养细胞，对其进行流式细胞分析，细胞分为两组，并通过染色进行观察，利用实时荧光定量聚合酶链反应（real-time polymerase chain reaction, real-time PCR）对特定蛋白的RNA水平进行分析。将牙乳头细胞复合ADM培养，实验组对培养细胞进行成骨与成脂诱导分化，对照组不进行诱导，同样进行染色与real-time PCR分析。结果：牙乳头细胞生长活跃，利用流式细胞分析发现70.3％细胞的间充质干细胞表面标志物Stro-1表达呈阳性，有96％细胞CD105表达呈阳性，经过诱导后，实验组细胞与成骨相关的骨钙素（osteocalcin， OCN)和骨涎蛋白（bone sialoprotein，BSP）的RNA水平，与成脂相关的肝X-受体α（liver X-recepter α, LXRα)、脂蛋白脂酶（lipoprotein lipase，LPL）、过氧化物酶体增殖物激活受体γ（peroxisome proliferator activated receptor γ ，PPAR-γ）和B类清道夫受体1（scavenger receptor class B type 1，SR-B1）的RNA水平均高于未经诱导的对照组细胞。牙乳头细胞复合ADM培养后细胞仍然生长活跃，两者紧密贴附，经诱导分化，实验组OCN与BSP的RNA水平高于对照组（P>0.05），实验组LPL的RNA水平高于对照组（P<0.05）。结论：通过口腔外科获得的年轻恒牙牙乳头组织含有大量具有成骨能力的干细胞，将其与具有胶原支架结构的ADM复合培养，发现ADM可以作为干细胞生长的支架结构，并对干细胞的成骨诱导起到一定作用。

关键词: 牙乳头, 干细胞, 脱细胞真皮, 细胞分化

Abstract: Objective： To get the stem cells from the young permanent tooth apical papillae, and observe the osteogenic differentiation of the cells after cultured with acellular dermal matrix (ADM). Methods: Young permanent tooth apical papillae were obtained by the oral surgeon. The cells from the apical papillae were isolated, cultured and analyzed through a flow cytometer. The cells in the experimental group were induced both osteogenic and adipogenic differentiation. The cells were not induced in the control group.Both groups were evaluated by staining and real-time polymerase chain reaction (real-time PCR) to examine the quantity of RNAs in the experimental group. The cells from apical papillae were also cultured with ADM. These cells were also induced both osteogenic and adipogenic differentiation in the experimental group, and not induced in the control group. The measures of staining and real-time PCR were also carried out. Results: The cells from the apical papillae proliferated in a rapid rate. Of which 70.3% in cultures were positive for Stro-1, and 96% positive for CD105 according to flow cytometric analysis. After induction, the RNA level related to osteogenic and adipogenic differentiation expressed higher in the experimental group than those of the control group without induction obviously, such as osteocalcin (OCN)， bone sialoprotein (BSP), liver X-recepter α (LXRα)， lipoprotein lipase（LPL）， peroxisome proliferator activated receptor γ (PPAR-γ)， and scavenger receptor class B type 1（SR-B1）. The cells cultured with ADM also had a fast proliferation, and grew attached to ADM. After induction, the RNA level of OCN and BSP had a higher expression than the control group (P>0.05), and LPL also expressed higher (P<0.05). Conclusion: The study approved that there were a big amount of stem cells in the young permanent tooth apical papilla obtained by oral surgery, which had significant osteogenic potential. The cells still proliferated well when they were cultured with ADM as a kind of collagen skelecton. The results showed that ADM could be performed as a base to support the stem cells to survive the environment, and it also could play a role in osteogenic differentiation of stem cells from apical papilla.

Key words: Dental papilla, Stem cells, Acellular dermis, Cell differentiation

许向亮，王恩博，崔念晖. 脱细胞真皮基质对年轻恒牙牙乳头干细胞分化的作用[J]. 北京大学学报（医学版）, 2014, 46(1): 12-18.

XU Xiang-liang, WANG En-bo, CUI Nian-hui. Influence of acellular dermal matrix on differentiation of stem cells from young permanent tooth apical papilla[J]. Journal of Peking University(Health Sciences), 2014, 46(1): 12-18.

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