纳米羟基磷灰石颗粒对成骨样细胞 生物活性的尺寸效应

 目的 探索不同尺寸的纳米羟基磷灰石颗粒(nano hydroxyapatite, nHAP)对成骨细胞增殖 和凋亡的生物学效应。 方法采用化学沉淀的方法, 可控制备不同大小的nHAP, 包括20 nm 球形 (20HAP)、40 nm球形(40HAP)和80 nm 球形(80HAP)。通过透射电子显微镜、X-射线衍射仪、红外线光 谱仪等工具分析颗粒大小、形状以及化学组分。将nHAP 颗粒制备成薄膜, 与成骨样细胞MG-63 细胞共 培养5 d, 通过甲基噻唑基四唑(methyl thiazolyl tetrazolium, MTT)计数活细胞数量, 采用流式细胞仪检测 细胞的凋亡情况, 通过透射电子显微镜观察细胞器的变化情况以及nHAP在细胞内的分布。 结果 可控 合成并对20HAP、40HAP 和80HAP进行大小组分分析。薄膜细胞共培养结果显示, 5 天时20HAP 组的 细胞吸光度值高达1.22±0.13, 与空白对照组相比差异无统计学意义(F=6.843, P=0.124)。细胞的凋亡率 随nHAP 尺寸的增加而增大, 与空白对照组相比, 差异有统计学意义(q=4.284、5.487、2.933, P <0.05);各 组细胞凋亡率分别为1.12%±0.51%(空白对照组), 6.68%±1.04%(20HAP 组), 10.08%±2.10%(40HAP 组)和13.61%±2.45%(80HAP 组)。透射电子显微镜结果证实, 20HAP 能够穿透细胞膜而对细胞超微结 构和细胞形态不具有毒性作用。 结论 成骨细胞增殖和凋亡都与nHAP 的尺寸相关, 其中20 HAP的促 进细胞增殖和抑制细胞凋亡作用最佳。

Controllable synthesis of different size hydroxyapatite nanoparticles and their biological effects on osteoblast-like cells

Objective To investigate the size effect of hydroxyapatite nanoparticles on proliferation and apoptosis of osteoblast-like cells. Methods Cetyltrimethylammnonium bromide (CTAB) was used to regulate the size of nano hydroxyapatite (nHAP) particles. All obtained particles were characterized by transmission electron microscopy (TEM), X-ray diffraction, dynamic light scattering and chemical analysis. HAP films were obtained by slowly coating cover glasses with 1% HAP particle suspension. MG-63 cells on three different films (20HAP, 40HAP and 80HAP) were cocultured for up to 5 days. Cell proliferation assay was obtained by methyl thiazolyl tetrazolium (MTT). Cell apoptosis was detected by flow cytometric detection. Cell ultrastructure morphology was observed by TEM observation. Results nHAP with diameter of 20 nm, 40 nm and 80 nm were synthesized and and analyzed. The MG-63 cells were cultured on three different films. The optical density value of cells on 20HAP was 1.22?0.13 after 5 days incubation, and there was no different compared to the control group (F=6.843, P=0.124). Cell number and viability were significantly higher on 20HAP compared to large nHAP after 5 days incubation. The percentage of apoptotic cells increased with increasing nHAP particle size. TEM images showed 20HAP was found in cytoplasm and cell morphology had no changes. Conclusion Both cell proliferation and cell apoptosis are related to the size of the nHAP particles. 20HAP was the most effective on promoting cell growth and inhibiting cell apoptosis.