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目的探讨金黄色葡萄球菌脂磷壁酸(Staphylococcal lipoteichoic acid,LTA-sa)对破骨细胞分化的影响。方法取RAW264.7细胞根据诱导培养条件不同分为5组,分别为LTA-sa 100 ng/m L组(A组)、LTA-sa200 ng/m L组(B组)、LTA-sa 400 ng/m L组(C组)、100 ng/m L鼠NF-κB受体活化因子配基组(receptor activator of NF-κB ligand,RANKL;作为阳性对照,D组)、空白对照组(E组,等体积PBS)。诱导培养后第5天,A~E组行抗酒石酸酸性磷酸酶(tartrate resistant acid phosphatase,TRAP)染色观察破骨样细胞形成情况并行细胞计数,采用Image-Pro Plus 6.0软件测量COAS(Corning Osteo Assay Surface)24孔板形成的骨吸收凹陷面积,计算其占整个视野面积的比值;同时A、B、C、E组采用MTT法检测细胞增殖情况。结果诱导培养后第5天,各组均可见破骨样细胞及骨吸收凹陷形成,但A~D组所形成的破骨样细胞及骨吸收凹陷面积占整个视野面积的比值均多于E组;且随LTA-sa浓度增加,A、B、C组以上指标逐渐增加,但均低于D组;各组间比较,差异均有统计学意义(P<0.05)。诱导培养后第5天,A、B、C、E组吸光度(A)值比较差异无统计学意义(P>0.05)。结论LTA-sa具有促进RAW264.7细胞向破骨细胞分化形成的作用。
Objective To investigate the effect of Staphylococcal lipoteichoic acid (LTA-sa) on osteoclast differentiation. Methods RAW264.7 cells were divided into 5 groups according to the induction culture conditions: LTA-sa 100 ng / m L group (A group), LTA-sa 200 ng / m L group (group C), 100 ng / m L receptor activator of NF-κB ligand (RANKL; positive control group D), blank control group (group E) , Equal volume of PBS). On the fifth day after induction, the cells were stained with tartrate resistant acid phosphatase (TRAP) in groups A to E for cell counting and COAS (Corning Osteo Assay Surface) 24-well plate to form the bone absorption area of depression, calculate the ratio of the total field area; the same time, A, B, C, E group using MTT assay cell proliferation. Results On the fifth day after induction, the osteoclast-like cells and the bone resorption pits were seen in all the groups. However, the ratio of osteoclast-like cells and bone resorption pits formed in groups A to D to the entire visual field area were more than those in group E (P <0.05). With the increase of LTA-sa concentration, the above indexes of A, B and C group increased gradually, but all of them were lower than that of D group. The differences among the groups were statistically significant (P <0.05). On the fifth day after induction, there was no significant difference in absorbance (A) between A, B, C and E groups (P> 0.05). Conclusion LTA-sa can promote the differentiation of RAW264.7 cells into osteoclasts.