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目的观察大鼠牙囊干细胞(rat dental follicle stem cells,rDFCs)在珊瑚羟基磷灰石生物玻璃陶瓷(coralline hydroxyapatite,CHA)上的附着增殖以及成骨因子表达情况,探索二者间生物相容性。方法 10只6 d龄SD大鼠,雌雄各半,处死后剥离双侧下颌骨磨牙区牙囊组织,通过组织块酶消化法获得rDFCs,rDFCs与CHA体外复合培养,扫描电镜(SEM)观察rDFCs与CHA黏附增殖情况,RTPCR法测定rDFCs、rDFCs-CHA及rDFCs成骨诱导组黏附增殖5、7、9 d成骨基因OPN、ALP、Osterix的表达。结果 SEM观察到rDFCs在CHA上能正常黏附、生长、增殖,4 d时细胞开始相互接触,成片状黏附于CHA表面,7 d时,细胞外基质已完全覆盖CHA表面。RT-PCR结果显示rDFCs与CHA共培养5、7、9 d时,CHA促进了成骨基因OPN、ALP、Osterix的表达,rDFCs-CHA组与rDFCs组相比,Osterix基因表达:7 d时前者是后者267倍;OPN基因表达:7 d时是24.7倍,ALP基因表达:5 d时rDFCs-CHA组是rDFCs组的3.1倍(P<0.01)。结论 CHA能促进rDFCs黏附、增殖、成骨分化及表达成骨因子,二者生物相容性良好。
Objective To observe the attachment and proliferation of rat dental follicle stem cells (rDFCs) on coralline hydroxyapatite (CHA) and the expression of osteogenic factors, and to explore their biocompatibility . Methods Ten 6-day-old SD rats were sacrificed in both male and female rats. The dental follicles in bilateral mandibular molars were dissected after sacrificed. RDFCs, rDFCs and CHA were cultured in vitro by tissue enzymatic digestion and examined by scanning electron microscopy (SEM) The adhesion and proliferation of CHA were detected by RT-PCR. The expressions of OPN, ALP and Osterix were detected by RTPCR at 5, 7 and 9 days after rDFCs, rDFCs-CHA and rDFCs osteogenic induction. Results SEM observation showed that rDFCs could adhere, grow and proliferate normally on CHA. After 4 days, cells began to contact with each other, and adhered to CHA surface. At 7 days, the extracellular matrix completely covered the surface of CHA. RT-PCR results showed that CHA promoted the expression of osteogenic genes OPN, ALP and Osterix when rDFCs-CHA co-cultured with CHA for 5, 7 and 9 days. The expression of Osterix gene in rDFCs-CHA group compared with rDFCs group: 267 folds of the latter; OPN gene expression was 24.7 folds at 7 days, and ALP gene expression was 3.1-fold more potent than rDFCs at 5 days (P <0.01). Conclusion CHA can promote rDFCs adhesion, proliferation, osteogenic differentiation and expression of osteogenic factor, both good biocompatibility.