钛颗粒对OPG-RANKL-RANK通路干预的时间依赖性研究

目的 探讨钛颗粒在体外不同时间点对核因子-κB受体活化因子配体-核因子-κB受体活化因子-骨保护素(RANKL-RANK-OPG)通路的干预作用。 方法 MC3T3-E1细胞培养传5代后,移入4个6孔板中进行培养,随机分为A、B、C、D 4组,每板上3孔均加入含0.1mg/L钛颗粒培养液作为实验组,下3孔加入普通培养液作为对照组。A、B、C、D 4组分别在培养后12、24、36、48h提取培养液进行OPG和RANKL蛋白的ELISA检测,同时提取贴壁细胞进行real-time PCR检测,观察OPG和RANKL的mRNA表达水平变化。 结果 MC3T3-E1细胞连续传代后生长良好,形态稳定,经ELISA检测OPG蛋白吸光度(A)值A组10.33±2.44,B组8.96±2.02,C组7.84±1.30,D组7.66±0.63,B、C、D组与对照组相比,差异有统计学意义(P<0.05);RANKL蛋白A值A组6.04±1.60,B组7.51±0.57,C组8.91±0.59,D组10.94±0.58,4组与对照组相比,差异均有统计学意义(P<0.05)。经PCR检测OPG mRNA表达A组0.89±0.07,B组0.81±0.11,C组0.74±0.15,D组0.72±0.16,B、C、D组与对照组相比,差异有统计学意义(P<0.05);RANKL mRNA表达A组1.21±0.21,B组1.22±0.33,C组1.24±0.09,D组1.24±0.04,4组与对照组相比,差异均有统计学意义(P<0.05)。 结论 钛颗粒既能够上调RANKL的表达水平,又会抑制OPG的形成,从而加速假体周围的骨溶解作用。

Study on the Time Dependence of the Effect of Ti Granule on OPG-RANKL-RANK System

Objective To study the Ti granule effect on the RANKL-RANK-OPG system at different point-in-time in vitro. Method MC3T3-E1 cells were passaged to the P5, and then they were inoculated into 4 six-hole cell cultured plates were divided into A, B, C and D groups randomly. The first three holes of each plate were added medium with 0.1mg/L Ti granule, and the second three holes were added ordinary medium. After 12h, 24h, 36h and 48h, the OPG and RANKL protein in group A, B, C and D were respectively tested by ELISA, and the cells of each group were respectively tested for the OPG and RANKL mRNA by real-time PCR. Results With the continuous passaging, the MC3T3-E1cells grew well with stable morphological characteristics. The A value of OPG tested by ELISA was group A 10.33±2.44, B 8.96±2.02, C 7.84±1.30, D 7.66±0.63. Compared with control groups, group B, C and D had statistically significant difference( P<0.05). RANKL protein was 6.04±1.60 in group A, 7.51±0.57 in B, 8.91±0.59 in C, 10.94±0.58 in D. Compared with control groups, group B, C and D had statistically significant difference (P<0.05). The mRNA expression of OPG was 0.89±0.07 in group A, 0.81±0.11 in B, 0.74±0.15 in C, 0.72±0.16 in D. Compared with control groups, all of the groups had statistically significant difference (P<0.05). The RANKL expression was 1.21±0.21 group A, 1.22±0.33 in B, 1.24±0.09 in C,1.24±0.04 in D. Compared with control groups, all of the groups had statistically significant difference(P<0.05). Conclusion Ti granule could not only up-regulate the RANKL expression, but also reduce the production of OPG, so that it accelerate the osteolysis around the prosthesis.