摘要
目的:探究高迁移率蛋白1(HMGB1)在机械应力诱导的人牙周膜细胞(h PDLCs)迁移中的作用及机制。方法:用Real-time PCR和Western blotting方法检测机械应力(10%形变量、频率0.5 Hz)加载不同时间(0、6、12、24、48 h)对h PDLCs中HMGB1和金属基质蛋白酶2、9(MMP2/9)表达的影响。然后,使用siRNA干扰技术下调HMGB1,并分别用Transwell检测h PDLCs迁移力、Real-time PCR检测MMPs表达、Western blotting检测ERK1/2和p-ERK1/2的表达水平。另外,分别使用1μmol/L MMPs抑制剂Batimastat、50 nmol/L ERK1/2信号通路抑制剂GDC-0994预处理h PDLCs 2 h,然后再加载机械应力24 h,并分别检测细胞迁移或MMPs的表达。结果:h PDLCs经机械应力加载后,其HMGB1、MMP2、MMP9 mRNA和蛋白的表达水平均随着加载时间而逐渐增加,24 h时达到峰值,48 h时呈下降趋势;下调HMGB1能够抑制机械应力诱导的h PDLCs迁移及细胞中MMP2、MMP9和p-ERK1/2的表达;Batimastat能够抑制机械应力诱导的细胞迁移;GDC-0994能够抑制机械应力诱导的MMP2和MMP9的表达。结论:HMGB1介导机械应力诱导的h PDLCs迁移,可能是通过激活ERK1/2信号通路、上调MMPs的表达而发挥作用的。
AIM： To study the role and mechanism of high mobility group box protein 1（ HMGB1） in mechanical stress induced the migration of human periodontal ligament cells（ h PDLCs）. METHODS： The expression of HMGB1 and matrix metalloproteinase 2/9（ MMP2/9） was assayed by real-time PCR and western blotting when cells were cultured with mechanism stress with 10% deformation,and 0. 5 Hz sinusoidal wave for 0,6,12,24 and 48 h respectively. Then,HMGB1 expression was downregulated by siRNA,the migration of h PDLCs was examined by transwell,MMPs expression was analyzed by real-time PCR,and the levels of ERK1/2 and p-ERK1/2 were determined by western blotting. Additionally,h PDLCs were pretreated with 1 μmol/L of Batimastat（ a MMPs inhibitor） and50 nmol/L of GDC-0994（ an ERK1/2 inhibitor） respectively for 2 h and then treated with mechanism stress for24 h,the cell migration and MMPs expression was detected,respectively. RESULTS： The mRNA and protein expression of HMGB1,MMP2 and MMP9 were all increased at 6,12 and 24 h in a time-dependent manner,while reduced at 48 h. Downregulation of HMGB1 inhibited mechanical stress-induced the migration of h PDLCs and the expression of MMP2,MMP9 and p-ERK1/2. Batimastat suppressed h PDLCs migration. The expression of MMP2 and MMP9 induced by mechanical stress was dampened by GDC-0994. CONCLUSION： HMGB1 mediates mechanical stress-induced the migration of h PDLCs,which may be through activating ERK1/2 signaling and upregulating MMPs expression.
出处
《牙体牙髓牙周病学杂志》
CAS
2017年第9期499-505,共7页
Chinese Journal of Conservative Dentistry
关键词
HMGB1
机械应力
人牙周膜细胞
迁移
金属基质蛋白酶
HMGB1
mechanical stress
human periodontal ligament cells
migration
matrix metalloproteinase
