口腔医学研究

口腔医学研究 ›› 2019, Vol. 35 ›› Issue (4): 335-339.DOI: 10.13701/j.cnki.kqyxyj.2019.04.008

• 龋病牙髓病学研究 • 上一篇    下一篇

载重组人血管内皮生长因子165的邻苯二酚壳聚糖促进乳牙牙髓干细胞血管形成的研究

杨雪1, 倪世磊2, 王婷婷1, 韩佳岐1, 张鲁鲁1, 高华丽1, 姜秋1*   

  1. 1. 吉林大学口腔医院儿童口腔科 吉林 长春 130021;
    2. 吉林省牙发育及颌骨重塑与再生重点实验室 吉林 长春 130021
  • 收稿日期:2018-09-03 出版日期:2019-04-28 发布日期:2019-04-23
  • 通讯作者: 姜秋,E-mail:jiangqiu1221@163.com
  • 作者简介:杨雪(1991~ ),女,山东临沂人,硕士在读,主要从事儿童口腔医学相关研究。
  • 基金资助:
    吉林省自然科学基金(编号:2014G075)

Angiogenesis of Stem Cells from Human Exfoliated Deciduous Teeth Stimulated by rhVEGF165 Loaded Catechol-chitosan

YANG Xue1, NI Shi-lei2, WANG Ting-ting1, HAN Jia-qi1, ZHANG Lu-lu1, GAO Hua-li1, JIANG Qiu1*   

  1. 1. Department of Pediatric Dentistry, Stomatology Hospital, Jilin University, Changchun 130021, China;
    2. Provincial Key Laboratory of Tooth Development and Jaw Remodeling and Regeneration, Changchun 130021, China.
  • Received:2018-09-03 Online:2019-04-28 Published:2019-04-23

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摘要： 目的: 体外研究载重组人血管内皮生长因子(recombinant human vascular endothelial growth factor,rhVEGF165)的邻苯二酚壳聚糖(catechol-chitosan,Cat-CS)支架材料对乳牙牙髓干细胞(stem cells from pulpof deciduous teeth,SHED)血管形成的影响,为其在年轻恒牙牙髓再生中的应用提供理论基础。方法: 结合酶联合消化法和倒置贴壁法分离培养SHED。经细胞形态、碱性磷酸酶和茜素红染色对SHED进行鉴定,通过噻唑蓝(methyl thiazol tetrazolium,MTT)法检测和对比壳聚糖(chitosan,CS)及Cat-CS的生物相容性,HE染色及扫描电镜(scanning electron microscope,SEM)检测细胞在CS及Cat-CS上的粘附,实时定量聚合酶链反应(real-time quantitative polymerase chain reaction,real-time PCR)检测血管内皮细胞表面标记物CD31、血管内皮生长因子受体2(VEGFR2)、血管内皮钙粘蛋白(VE-cadherin)的表达。结果: SHED具有较强的增殖能力,碱性磷酸酶及茜素红染色阳性;MTT结果显示CS及Cat-CS对细胞无明显毒性;HE染色及SEM观察结果显示相比于CS,Cat-CS更有利于细胞在孔径内的延伸以及在其表面的粘附;与空白对照组比较,CS及Cat-CS能促进SHED表达CD31、VEGFR2和VE-cadherin,Cat-CS与rhVEGF165复合后,这些基因的表达进一步增加(P<0.05)。结论: Cat-CS具有良好的生物相容性,可以作为牙髓再生良好的支架材料。

关键词: 邻苯二酚壳聚糖, 乳牙牙髓干细胞, 牙髓再生

Abstract: Objective: To study the effects of rhVEGF165 loaded catechol chitosan (cat-cs) scaffold on the angiogenesis of stem cells from human exfoliated deciduous teeth (SHED) in young permanent teeth. Methods: SHED was isolated and cultured by enzyme digestion and inverted adherent method. The stemness of SHEDs was identified by cell morphology, alkaline phosphatase, and alizarin red staining. The biocompatibility of CS and Cat-CS was detected by MTT, HE staining, and scanning electron microscope (SEM). The expression of endothelial cell marker CD31, VEGFR2, and VE-cadherin was detected by real-time PCR. Results: SHED had strong reproductive activity. Alkaline phosphatase and alizarin red staining revealed differentiation of SHED. The MTT results showed no obvious toxicity of Cat-CS to SHED. HE staining and SEM showed Cat-CS was more favorable for cell extension and adhesion. Cat-CS could efficiently promote more expression of CD31, VEGFR2, and VE-cadherin than CS. And the expression of these genes is further promoted when Cat-CS is combined with rhVEGF165 (P<0.05). Conclusion: Cat-CS has good biocompatibility and can be used as a good scaffold for pulp regeneration.

Key words: Catechol chitosan, SHED, Pulp regeneration