Influence of macro-pore bone block on osteoblast behavior in vitro / 中华骨科杂志

ABSTRACT

Objective To observe and identify the impact of a type of macro?pore bone block bioactive glass on osteo?blast in vitro. Methods Extract fluid of new bioactive glass was prepared withα?MEM culture medium as the bioactive medium group. And the concentrations of different ions were detected with Inductively Coupled Plasma?Atomic Emission Spectrometry in bioactive medium group andα?MEM medium group. MC3T3?E1 cells cultured in bioactive medium group were considered as ex?perimental group and cells cultured inα?MEM medium as control group. Giemsa and immunofluorescence staining was performed to detect the cell numbers, the karyoplasmic ratio and the average fluorescence intensity per cell. Cell proliferation and viability in different groups were detected by cell cycle analysis, MTT assay and BrdU assay, respectively. Total RNAs of cells in different groups were extracted and the expressions of ALP, OCN and collagenⅠwere measured by quantitative real time PCR. ALP stain?ing and alizarin red staining were performed to assess the differentiation and mineralization of MC3TC?E1 cells in different groups. Results The concentrations of Si and F were 40.02 ± 0.67 mg/L and 0.02 ± 0.001 mg/L in bioactive medium group, higher than 2.02±0.01 mg/L and 0.00 mg/L inα?MEM solution, and the concentration of Ca was lower than that inα?MEM solution. The con?centration of P and Na had no difference. In Giemsa staining, the cell number in 400 times field under a microscope was 106.0 ± 6.025 in bioactive medium group and 40.20 ± 3.639 inα?MEM medium group. In the immunofluorescence of vinculin, the karyo?plasmic ratio and the expression of vinculin were higher in bioactive medium group (40.85±5.720, 0.050 88±0.021 78) than inα? MEM medium group (21.93 ± 4.137, 0.023 60 ± 0.003 18). In cell cycle analysis, the proportion of cells retained in S and G2/M phase in the bioactive medium group was more than that in theα?MEM medium group after 72 hours of cell culture. In the BrdU and MTT assay, MC3T3?E1 cells in bioactive medium group both showed a higher proliferation rate with statistical significance. In MC3T3?E1 cells cultured with the bioactive medium, the expressions of osteogenesis?related genes were higher than those cultured with ordinaryα?MEM solution;in the ALP staining and alizarin red staining, the expression of ALP and the mineralization rate were higher in bioactive medium group (1.328%±0.015 36%, 2.953%±0.536 3%) than inα?MEM medium group (0.979%±0.030 59%, 1.000%±0.208 1%). Conclusion The bioactive medium promotes cell proliferation and osteoblastic differentiation of MC3T3?E1 cells, and has much more Si ions, which indicates that macro?pore bone block bioactive glass can promote cell proliferation and dif?ferentiation and has promising bioactivity.