中国组织工程研究 ›› 2020, Vol. 24 ›› Issue (7): 1009-1015.doi: 10.3969/j.issn.2095-4344.2022

• 骨髓干细胞 bone marrow stem cells • 上一篇    下一篇

钛表面锶改性后骨髓基质干细胞黏附、迁移、增殖及成骨相关基因的表达

刘春栋1,申晓青1,张艳丽2,3,张小根1,吴补领2,3   

  1. 1南方医科大学珠江医院口腔科,广东省广州市  510280;2南方医科大学南方医院口腔科,广东省广州市  510515;3南方医科大学口腔医学院,广东省广州市  510515
  • 收稿日期:2019-04-23 修回日期:2019-04-30 接受日期:2019-07-15 出版日期:2020-03-08 发布日期:2020-01-19
  • 通讯作者: 吴补领,博士,教授,主任医师,南方医科大学南方医院口腔科,广东省广州市 510515;南方医科大学口腔医学院,广东省广州市 510515
  • 作者简介:刘春栋,男,1979年生,山东省临清市人,汉族,2015年南方医科大学毕业,博士,主治医师,主要从事口腔种植学相关研究。
  • 基金资助:
    广东省医学科研基金项目(B2016040)

Effects of strontium-modified titanium surfaces on adhesion, migration and proliferation of bone marrow mesenchymal stem cells and expression of bone formation-related genes

Liu Chundong1, Shen Xiaoqing1, Zhang Yanli2, 3, Zhang Xiaogen1, Wu Buling2, 3   

  1. 1Department of Stomatology, Zhujiang Hospital, Southern Medical University, Guangzhou 510280, Guangdong Province, China; 2Department of Stomatology, Nanfang Hospital, Southern Medical University, Guangzhou 510515, Guangdong Province, China; 3College of Stomatology, Southern Medical University, Guangzhou 510515, Guangdong Province, China
  • Received:2019-04-23 Revised:2019-04-30 Accepted:2019-07-15 Online:2020-03-08 Published:2020-01-19
  • Contact: Wu Buling, MD, Professor, Chief physician, Department of Stomatology, Nanfang Hospital, Southern Medical University, Guangzhou 510515, Guangdong Province, China; College of Stomatology, Southern Medical University, Guangzhou 510515, Guangdong Province, China
  • About author:Liu Chundong, MD, Attending physician, Department of Stomatology, Zhujiang Hospital, Southern Medical University, Guangzhou 510280, Guangdong Province, China
  • Supported by:
    the Medical Research Foundation of Guangdong Province, No. B2016040

摘要:

文题释义:

钛表面锶改性:锶是人体必需的一种微量元素,研究表明低剂量的锶具有促进成骨细胞生长同时抑制破骨细胞功能的作用。有学者利用微弧氧化、阳极氧化加水热法、直流磁控共溅射等方法在钛种植体表面涂层内掺入锶元素,但存在步骤复杂、难以统一标准等缺点。该实验依次采用NaOH、醋酸锶、热处理及水化处理成功对纯钛表面进行锶改性,在经过锶改性后可以提高成骨细胞及骨髓基质干细胞的生物学活性。
骨髓基质干细胞:是来源于骨髓中未完全分化的原始细胞,这类细胞可塑性极强,有多向分化潜能和低免疫原性以及很强的自我更新能力。FRIEDENSTEIN等在1966年最早从骨髓中提取出了干细胞,发现在骨髓中含有能分化为其他间质细胞的细胞及成纤维母细胞,且在培养过程中发现该细胞能贴壁呈集落性生长。通过流式细胞术检测间充质干细胞表面标记物以及成骨及成脂向诱导,可以对获得的细胞进行鉴定。

背景:锶是一种既刺激骨形成、又抑制骨吸收的元素,纯钛种植体表面锶改性促进骨质疏松状态下种植体骨结合是当前研究的热点,具有非常重要的意义。

目的:探讨改良锶改性纯钛表面对大鼠骨髓基质干细胞黏附、铺展、迁移、增殖及成骨相关基因表达的影响。

方法:将TA2纯钛片打磨清洗后,依次放于5 mol/L NaOH溶液、100 mmol/L醋酸锶溶液内处理,并以600 ℃或者700 ℃进行热处理1 h,去离子水80 ℃水化处理(W)24 h,以纯钛为对照,共分为5组,分别为Ti、Sr600、Sr600W、Sr700、Sr700W组。采用全骨髓培养法获取大鼠骨髓基质干细胞,将改性钛片置于24孔培养板中与细胞悬液培养,CCK-8法检测骨髓基质干细胞的黏附和增殖情况。骨髓基质干细胞与钛片共培养24 h后对细胞肌动蛋白进行染色,观察细胞黏附伸展形态;通过细胞划痕实验和荧光染色,观察骨髓基质干细胞在各组钛片表面迁移的能力;qRT-PCR法检测各组细胞Ⅰ型胶原、Runx2、骨保护蛋白、RANKL、骨桥蛋白的mRNA表达。

结果与结论:锶改性纯钛片可以促进骨髓基质干细胞的铺展、迁移。在5 d和7 d时,锶改性钛片表面细胞增殖较纯钛组均明显升高(P < 0.001)。在5 d时,Sr700组的细胞增殖数目较Sr600组高(P < 0.01)。在14 d时,锶改性纯钛片可以促进骨髓基质干细胞Ⅰ型胶原、Runx2、骨桥蛋白的表达。结果表明,改良锶改性纯钛可以促进大鼠骨髓基质干细胞的黏附、铺展、迁移、增殖及成骨相关基因的表达,水化处理对大鼠骨髓基质干细胞成骨向分化具有促进作用。

ORCID: 0000-0003-2166-2487(刘春栋)

中国组织工程研究杂志出版内容重点:干细胞;骨髓干细胞;造血干细胞;脂肪干细胞;肿瘤干细胞;胚胎干细胞;脐带脐血干细胞;干细胞诱导;干细胞分化;组织工程

关键词: 纯钛, 锶改性, 骨髓基质干细胞, 细胞黏附, 细胞增殖, 成骨相关基因

Abstract:

BACKGROUND: Strontium promotes bone formation, and reduces bone resorption. Strontium-modified titanium implant surface has been the focus of research in implant osseointegration under osteoporotic conditions.

OBJECTIVE: To investigate the cell adhesion, stretch, and migration of advanced strontium-modified titanium surfaces using bone marrow mesenchymal stem cells isolated from rats.

METHODS: Strontium-modified titanium surfaces were produced by sequential treatments with 5 mol/L NaOH, 100 mmol/L strontium acetate, and heated at 600 or 700 °C for 1 hour. After heat treatment, half the samples were soaked in deionized water at 80 °C for 24 hours, then washed and dried. The pure titanium served as control group, and there were five groups. The whole bone marrow adherence method was used to separate bone marrow mesenchymal stem cells from the rats. The modified titanium sheets were placed in 24-well plates and cultured in cell suspension. Cell adhesion and cell proliferation were assessed using cell counting kit-8 assay. After bone marrow mesenchymal stem cells were with the titanium sheet for 24 hours, the actin was stained to observe cell adhesion and stretch. The migration of bone marrow mesenchymal stem cells on different titanium surfaces was investigated using cell scratch test and fluorescence staining. The expression levels of collagen type І, Runx2, osteoprotegerin, RANKL and osteopontin were detected by qRT-PCR.

RESULTS AND CONCLUSION: Strontium-modified titanium could promote the stretch and migration of bone marrow mesenchymal stem cells. The number of proliferative cells in the Sr700 group was significantly higher than that in the Sr600 group at 5 days (P < 0.01). On day 14, strontium-modified titanium promoted the expression of collagen type І, Runx2, and osteopontin. In summary, strontium-modified titanium can promote adhesion, spreading, migration and proliferation of rat bone marrow mesenchymal stem cells. Moreover, hydration treatment can enhance the osteogenic activity of rat bone marrow mesenchymal stem cells. 

Key words: pure titanium, strontium-modified, bone marrow mesenchymal stem cells, cell adhesion, cell proliferation, bone formation-related genes

中图分类号: