煅烧骨/壳聚糖复合材料对成骨细胞增殖黏附的影响

Effects of calcined bone/chitosan composite materials on proliferation and adhesion in osteoblasts

背景:前期研究制备了不同配比的煅烧骨/壳聚糖复合材料,体外细胞实验显示该复合材料安全无毒。目的:观察煅烧骨/壳聚糖复合材料对成骨细胞增殖与黏附的影响。方法:采用溶液共混法制备煅烧骨与壳聚糖质量比分别为1/2、1/1、2/1的复合材料,将新生大鼠成骨细胞分别接种于3种复合材料上,以单独培养的细胞为对照。采用CCK-8法检测细胞增殖情况,DAPI荧光染色观察细胞在材料上的生长情况,扫描电镜观察细胞在复合材料表面的黏附。结果与结论:①CCK-8检测显示4组成骨细胞增殖良好,在培养的第1-3天增殖较慢,第3天后增殖加快,第3-7天进入对数生长期,第7天后进入平台期,复合材料组细胞增殖与对照组无差异;②培养3 d后的DAPI荧光染色显示,成骨细胞在3种复合材料表面生长良好,组间无明显差异;③扫描电镜显示,培养3d后成骨细胞紧密黏附于3种复合材料表面,伸展完全,胞体丰满并伸出伪足嵌入材料内部;培养7d后细胞铺满材料表面,生长密集,其中质量比为2/1煅烧骨/壳聚糖复合材料表面的细胞最密集;④结果表明,成骨细胞可在煅烧骨/壳聚糖复合材料表面增殖与黏附。

BACKGROUND:The different ratios of calcined bone/chitosan composites were prepared in previous studies,and in vitro cell experiments showed that the composites were safe and non-toxic.OBJECTIVE:To observe the effect of calcined bone/chitosan composite on the proliferation and adhesion of osteoblasts.METHODS:The composite materials with calcined bone and chitosan mass ratios of 1/2,1/1,and 2/1 were prepared by solution blending method.Neonatal rat osteoblasts were inoculated on three kinds of composite materials,and the cells cultured alone were used as control.CCK-8 kit was used to detect cell proliferation.DAPI fluorescence staining was used to observe the growth of cells on the material.Scanning electron microscopy was used to observe the adhesion of cells on the surface of the composite material.RESULTS AND CONCLUSION:(1)The CCK-8 test showed that the osteoblasts of the four components proliferated well,proliferated slowly from the first to third days of culture,accelerated after the third day,entered the logarithmic growth phase from the third to seventh days,and entered the plateau phase after the seventh day.There was no difference in cell proliferation between the composite materials group and the control group.(2)After 3 days of culture,DAPI fluorescent staining showed that osteoblasts grew well on the surface of the three kinds of composites,and there was no significant difference between the groups.(3)Scanning electron microscopy showed that after 3 days of culture,the osteoblasts adhered tightly to the surface of the three kinds of composites,stretched completely;the cell body was full and protruded the pseudopod that was embedded in the material.After 7 days of culture,the cells covered the surface of the material and grew densely,among which the cells on the surface of calcined bone/chitosan composite with a mass ratio of 2/1 were most dense.(4)The results showed that osteoblasts could proliferate and adhere on the surface of the calcined bone/chitosan composite material.