种植体周围炎对成骨细胞影响的体外研究

Effect of peri-implantitis on proliferation,differentiation and maturation of osteoblasts in vitro

目的:利用体外分离培养的种植体周围炎来源和正常组织来源人成骨细胞(osteoblast,OB),观察炎性微环境对其增殖、分化成熟的影响。探讨种植体周围炎对OB功能的影响,期望能为临床种植修复提供相关的理论基础。方法:首先进行成骨细胞原代及传代培养,然后对细胞爬片进行碱性磷酸酶(alkaline phosphatase,ALP)染色,光镜下观察计数ALP阳性OB数,并将细胞爬片用茜素红进行钙结节染色,光镜下观察钙结节,计数并拍照,均用于成骨细胞鉴定;传代第三代细胞,收集细胞上清液分别进行ALP活性检测及应用放射免疫方法测定骨钙素(osteocalcin,OCN)含量。结果:原代培养的细胞呈现OB的形态学特征,ALP染色阳性,钙结节茜素红染色呈橘红色钙化结节,此两项结果证实培养细胞为OB;传代细胞种植体来源的细胞,ALP阳性OB数量减少,钙结节数量及骨钙素数值降低。结论:种植体周围炎引发的炎性微环境降低成骨细胞的增殖分化及成骨功能。

Objective:To observe the effects of inflammatory microenvironment on proliferation,differentiation and maturation of osteoblasts derived from peri-implantitis and human osteoblasts derived from normal tissue;and to provide a theoretical basis for clinical implantation and repair.Methods:The primary culture and subculture of osteoblasts were carried out at first.Then the cell sheets were stained with alkaline phosphatase(ALP).The number of ALP positive osteoblast was counted under the light microscope,and the cell sheets were stained by alizarin red calcium.The calcium nodules were observed and counted for osteoblast identification under the light microscope.The third generations of cells were passed on,and the activity of ALP in cell supernatant was detected and the content of osteocalcin(OCN)was determined by radioimmunoassay.Results:The primary cultivated cells exhibited morphological characteristics of osteoblast,and they were confirmed as osteoblasts with positive ALP staining and red orange calcified nodules stained by alizarin red calcium.The passage osteoblasts derived from peri-implantitis demonstrated decreased number of positive ALP staining,decreased number of calcium nodules and decreased value of osteocalcin.Conclusion:Inflammatory microenvironment induced by peri-implantitis can decrease the proliferation,differentiation and osteogenic function of osteoblasts.