基质细胞衍生因子-1对炎症和正常来源的人牙周膜干细胞成骨分化能力的影响

doi:10.7518/hxkq.2019.05.003

华西口腔医学杂志 ›› 2019, Vol. 37 ›› Issue (5): 469-475.doi: 10.7518/hxkq.2019.05.003

基质细胞衍生因子-1对炎症和正常来源的人牙周膜干细胞成骨分化能力的影响

林永盛¹,王丰芝²,雷晓静²,何健民³()

1.西北民族大学甘肃省口腔疾病研究重点实验室（培育基地）, 西北民族大学口腔医学国家民委重点实验室,兰州 730030
2.海南口腔医院口腔内科,海南 570100
3.甘肃省人民医院口腔治疗中心,兰州 730000

收稿日期:2019-04-18 修回日期:2019-08-03 出版日期:2019-10-01 发布日期:2019-10-15
通讯作者: 何健民 E-mail:hjm1121@126.com
作者简介:林永盛,副教授,学士,E-mail：lin_yongsheng@126.com
基金资助:
甘肃省科技厅科技支撑项目(1504FKCA095);中央高校基金(31920150048)

Comparative study with the effect of stromal cell derived factor-1 on osteogenic differentiation of human healthy and inflammatory periodontal ligament stem cells

Lin Yongsheng¹,Wang Fengzhi²,Lei Xiaojing²,He Jianmin³()

1. Key Laboratory of Oral Diseases of Gansu Provincial, Key Laboratory of Stomatology of State Ethnic Affairs Commission, Northwest Minzu University, Lanzhou 730030, China
2.Dept. of Oral Medicine, Hainan Stomatological Hospital, Hainan 570100, China
3.Dept. of Stomatology, Gansu Provincial Hospital, Lanzhou 730000, China

Received:2019-04-18 Revised:2019-08-03 Online:2019-10-01 Published:2019-10-15
Contact: Jianmin He E-mail:hjm1121@126.com
Supported by:
Key Science and Technology Program Supported by Gansu Science and Technology Department(1504FKCA095);Central University Fund(31920150048)

摘要/Abstract

摘要：

目的通过体外培养炎症来源的人牙周膜干细胞（iPDLSCs）和正常来源的人牙周膜干细胞（hPDLSCs）,比较基质细胞衍生因子-1（SDF-1）对于两种来源细胞的成骨分化作用。方法采用组织块酶消化法原代培养iPDLSCs 和hPDLSCs,经有限稀释法纯化,通过流式细胞仪对干细胞表面标记物检测鉴定后,对其进行成骨诱导;MTT法检测并比较SDF-1对两种来源的细胞增殖能力的影响;茜素红染色检测SDF-1作用于两种来源的细胞后钙化骨量的表达;碱性磷酸酶法比较SDF-1作用于两者的成骨分化能力;逆转录聚合酶链反应（RT-PCR）法检测SDF-1作用于两种牙周膜干细胞前后成骨相关基因表达水平的变化。结果两种来源的牙周膜细胞经纯化后均阳性表达干细胞标记物。hPDLSCs较iPDLSCs增殖能力高;两种细胞经SDF-1成骨诱导培养后,成骨相关基因的表达水平均较诱导前明显上调（P<0.05）,SDF-1在50、200 ng·mL ^-1时分别对iPDLSCs和hPDLSCs细胞成骨分化作用最明显（P<0.05）。结论正常来源和炎症来源的人牙周膜干细胞均具有成骨分化能力,SDF-1可增强两种来源的牙周膜干细胞的成骨分化能力。

关键词: 基质细胞衍生因子-1, 炎症, 人牙周膜干细胞, 成骨分化

Abstract:

Objective This study aims to compare the osteogenic differentiation capability of stem cells derived from human inflammatory periodontal ligament tissues (iPDLSCs) with those of stem cells derived from healthy periodontal ligament tissues (hPDLSCs). Both types of tissues were induced by stromal cell derived factor (SDF-1) in vitro. Methods iPDLSCs and hPDLSCs were primarily cultured by tissue digestion method and purified by limited dilution cloning. The cells were passaged and identified by stem cell surface marker expression through flow cytometry. Then, we used thiazolyl blue tetrazolium bromide to detect and compare the proliferation capabilities of the iPDLSCs and hPDLSCs. Express of bone volumes were detected by alizarin red staining after SDF-1 was added to the cells. Using alkaline phosphatase, we evaluated the osteogenic differentiation capability of the cells induced by SDF-1. The expression levels of the osteogenesis-related genes of the cells induced by SDF-1 were determined by reverse transcription-polymerase chain reaction. Results After purification, both iPDLSCs and hPDLSCs expressed stem cell markers. hPDLCSs had a higher proliferation capability than iPDLSCs. Osteogenesis-related genes had higher expression levels in the cells induced by SDF-1 than in those without induction (P<0.05). SDF-1 at 50 and 200 ng·mL ^-1 concentration greatly affected the differentiation capabilities of iPDLSCs and hPDLSCs respectively. Conclusion iPDLSCs and hPDLSCs had osteogenic differentiation capability. The level of osteogenic differentiation in normal and inflamed periodontal ligament stem cells increases after SDF-1 induction.

Key words: stromal cell derived factor-1, inflammation, human periodontal ligament stem cells, osteogenic differentiation

中图分类号:

R780.2

林永盛,王丰芝,雷晓静,何健民. 基质细胞衍生因子-1对炎症和正常来源的人牙周膜干细胞成骨分化能力的影响[J]. 华西口腔医学杂志, 2019, 37(5): 469-475.

Lin Yongsheng,Wang Fengzhi,Lei Xiaojing,He Jianmin. Comparative study with the effect of stromal cell derived factor-1 on osteogenic differentiation of human healthy and inflammatory periodontal ligament stem cells[J]. West China Journal of Stomatology, 2019, 37(5): 469-475.

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参考文献 20

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基因	编号	引物序列
Runx2	NM001015051	F：5’-TGCCAGCTGCATCCTATTT-3’
		R：5’-CCCTAAAGTCACTCGGTATGTG-3’
ColⅠ	BC054498	F：5’-AGAGTGGAGCAGTGGTTACTA-3’
		R：5’-GATACAGGTTTCGCCAGTAGAG-3’
OPN	AB469789	F：5’-CGAGGAGTTGAATGGTGCATA-3’
		R：5’-TCCAGCTGACTCGTTTCATAAC-3’
OCN	NM199173	F：5’-TCACACTCCTCGCCCTATT-3’
		R：5’-CCTCCTGCTTGGACACAAA-3’
BMP-2	NM001200	F：5’-GGGTGGAATGACTGGATTGT-3’
		R：5’-GAGTTCAGATGATCAGCCAGAG-3’
β-actin	NM001101	F：5’-GGGTGGAATGACTGGATTGT-3’
		R：5’-GACAGGATGCAGAAGGAGAT-3’

基质细胞衍生因子-1对炎症和正常来源的人牙周膜干细胞成骨分化能力的影响

Comparative study with the effect of stromal cell derived factor-1 on osteogenic differentiation of human healthy and inflammatory periodontal ligament stem cells

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