Song Z C, Li S, Dong J C, Sun M J, Zhang X L, Shu R
a Department of Periodontology , Ninth People's Hospital, Shanghai Jiao Tong University School of Medicine.
b Shanghai Key Laboratory of Stomatology & Shanghai Research Institute of Stomatology, National Clinical Research Center of Stomatology , Shanghai , 200011 , China.
Biotech Histochem. 2017;92(8):606-618. doi: 10.1080/10520295.2017.1370131. Epub 2017 Dec 5.
Hypoxia is a crucial microenvironment for inflamed periodontal tissue and periodontal wound healing. Enamel matrix proteins (EMPs) potentially can promote the formation of new periodontium. The effects of EMPs on periodontal ligament cells under hypoxia, however, remain unclear. We investigated the effects of EMPs on cellular biobehavior and osteogenic differentiation of human periodontal ligament cells (hPDLCs) under hypoxia. Under cobalt chloride (CoCl)-induced hypoxia, cellular biobehavior of hPDLCs, including proliferation, attachment, spreading, and migration with or without EMPs, was evaluated by 3-(4, 5-dimethylthiazol- 2-yl)-2, 5-diphenyl tetrazolium bromide (MTT), cell counting, spreading area measurement and wound scratch assay. The osteogenic activity of hPDLCs was assessed using alkaline phosphatase (ALP) and alizarin red S staining (ARS). The expressions of osteogenic genes including runt related transcription factor 2 (Runx2), ALP, osteocalcin (OCN) and collagen type I (Col-I) were detected using real time quantitative PCR, western blot and immunocytochemistry assays. The biobehavior and osteogenic differentiation of hPDLCs were inhibited significantly under hypoxia. EMPs have no effect on cell proliferation under mimicked hypoxia. EMPs partly reversed the inhibitory effects of hypoxia, however, for other cellular biobehavior including attachment, spreading and migration, and markedly up-regulated osteogenic differentiation activities including ALP, mineralization ability and the expressions of osteogenic genes such as Runx2, ALP, osteocalcin, and collagen type I in hPDLCs under hypoxia. EMPs attenuate the hypoxic injury to cellular biobehavior and osteogenic differentiation in hPDLCs under hypoxia.
缺氧是炎症性牙周组织和牙周伤口愈合的关键微环境。牙釉质基质蛋白（EMPs）可能促进新牙周组织的形成。然而，EMPs在缺氧条件下对牙周膜细胞的影响仍不清楚。我们研究了EMPs在缺氧条件下对人牙周膜细胞（hPDLCs）细胞生物学行为和成骨分化的影响。在氯化钴（CoCl）诱导的缺氧条件下，通过3-（4,5-二甲基噻唑-2-基）-2,5-二苯基四氮唑溴盐（MTT）、细胞计数、铺展面积测量和伤口划痕试验，评估了有无EMPs时hPDLCs的细胞生物学行为，包括增殖、附着、铺展和迁移。使用碱性磷酸酶（ALP）和茜素红S染色（ARS）评估hPDLCs的成骨活性。使用实时定量PCR、蛋白质印迹和免疫细胞化学分析检测包括 runt 相关转录因子 2（Runx2）、ALP、骨钙素（OCN）和 I 型胶原（Col-I）在内的成骨基因的表达。在缺氧条件下，hPDLCs的生物学行为和成骨分化受到显著抑制。在模拟缺氧条件下，EMPs对细胞增殖没有影响。然而，EMPs部分逆转了缺氧对其他细胞生物学行为（包括附着、铺展和迁移）的抑制作用，并显著上调了缺氧条件下hPDLCs的成骨分化活性，包括ALP、矿化能力以及Runx2、ALP、骨钙素和I型胶原等成骨基因的表达。EMPs减轻了缺氧对hPDLCs细胞生物学行为和成骨分化的损伤。
