阿司匹林对小鼠骨髓基质细胞生物活性的影响

Effect of aspirin on cell biological activities in murine bone marrow stromal cells

目的 研究不同浓度阿司匹林对小鼠骨髓基质细胞ST2增殖活性、碱性磷酸酶（alkaline phosphatase,ALP）活性、细胞周期及凋亡的影响,为临床应用阿司匹林修复骨缺损尤其是牙槽骨吸收提供参考.方法 用含不同浓度[0（空白对照组）、1、10、100及1 000 μmol/L]阿司匹林的培养液培养细胞1、2、3、5、7d后,采用甲基噻唑基四唑（methyl thiazolyl tetrazolium,MTT）法检测细胞增殖情况;药物作用1、3、7d后按试剂盒说明方法检测细胞的ALP活性;药物作用48 h后采用流式细胞术检测其对细胞周期及凋亡的影响.结果 与空白对照组相比,MTT结果显示低浓度阿司匹林（1、10 μmol/L）可以促进细胞增殖,1、10 μmol/L阿司匹林处理细胞7d后的吸光度值分别为0.413±0.010和0.387±0.017,均显著高于空白对照组（0.313±0.012）（P＜0.01）,1、10 μmol/L阿司匹林可提高细胞的ALP活性,二者处理细胞7d后ALP水平分别为（6.0±0.3）和（7.7±0.4） μmol· min-1·g-1,均显著高于空白对照组[（4.3±0.9） μmol·min-1·g-1]（P＜0.05）,而高浓度（1 000 μmol/L）阿司匹林对细胞增殖和ALP活性有抑制作用,其吸光度值和ALP活性分别为0.267±0.016及（4.3±1.3）μmol· min-1·g-1（P＜0.05）.流式细胞术检测结果显示,1 000 μmol/L阿司匹林可抑制细胞在静止期（G0/G1）,0和1 000μmol/L药物组对应的静止期细胞百分数分别为（75.00±0.90）％及（78.55±1.35）％,二者差异有统计学意义（P＜0.05）,不同浓度阿司匹林均可减少细胞凋亡,0、1、10、100和1 000 μmol/L药物组相应的凋亡细胞百分比分别为（11.50±0.90）％、（5.30±0.10）％、（5.50±0.10）％、（4.90±0.90）％和（7.95±0.25）％,不同浓度药物组与空白对照组相比差异均有统计学意义（P＜0.05）.结论 阿司匹林对骨髓基质细胞的生物活性有促进作用,在适宜的浓度范围内可以促进成骨.

Objective To determine the effect of aspirin on cell proliferation,alkaline phosphatase （ALP） activity,cell cycle and apoptosis in murine bone marrow stromal cells,so as to explore an appropriate dose range to improve bone regeneration in periodontal treatment.Methods ST2 cells were stimulated with aspirin（concentrations of 1,10,100 and 1 000 μmol/L） for 1,2,3,5 and 7 d.Cell proliferation was measured by methyl thiazolyl tetrazolium（MTT） assay.After ST2 cells were treated for 1,3 and 7 d,ALP activity was measured by ALP kit,cell cycle and apoptosis were measured by flow cytometry（FCM） after treated for 48 h.Results MTT assays showed that variousdoses of aspirin have different effects on the cell growth.Briefly,lower concentrations（1,10 μmol/L） of aspirin promoted the cell growth,the A value of 0,1 and 10 μmol/L aspirin 7-day-treated cells were 0.313±0.012,0.413±0.010 and 0.387±0.017 respectively（P 〈 0.01 vs control）,and so did the ALP level（[4.3±0.9],[6.0±0.3] and [7.7±0.4] μmol· min-1· g-1,P 〈 0.05 vs control）,while higher concentrations,especially 1 000 μmol/L of aspirin mightinhibitthe cell growth with time going,A value and ALP level were 0.267±0.016,（4.3±1.3） μmol· min-1· g-1 respectively（P 〈 0.05 vs control）.Cell cycle analysis revealed no changes in comparison to control cells after treatment with 1 or 10 μmol/L aspirin,but it was observed that cell mitosis from S phase to G2/M phase proceeded at higher concentrations of 100 μmol/L aspirin,and the cell cycle in phase G0/G1 arrested at 1 000 μmol/L.Parallel apoptosis/necrosis studies showed that the percentage of cells in apoptosis decreased dramatically at all doses of aspirin,the apoptosis rates of ST2 cells responded to 0,1,10,100 and 1 000 μmol/L aspirin were （11.50±0.90）%,（5.30±0.10）%,（5.50±0.10）%,（4.90±0.90）% and （7.95±0.25）% respectively（P 〈 0.05 vs control）.Conclusions This study demonstrated that lower dosage of aspirin can promote ST2 cells growth,osteogenic activity and inhibit its apoptosis.Aspirin maybe used for the bone reconstruction with a proper concentration.