Tim-3对单核细胞诱导破骨样细胞生成和骨吸收功能的影响

Effects of Tim-3 on osteoclast-like cell formation and bone resorption induced by peripheral blood mononuclear cells

目的探讨T细胞免疫球蛋白及粘蛋白结构域3(T cells immunoglobulin domain and mucin domain 3,Tim-3)对单核细胞诱导破骨样细胞生成和骨吸收功能的影响。方法流式细胞术检测RA患者和体检健康者外周血单核细胞上Tim-3的表达水平;利用人外周血单核细胞体外诱导破骨样细胞;实时定量PCR法检测破骨样细胞形成过程中RANK、CTSK和MMP9 mRNA表达水平;瑞氏染色和肌动蛋白染色观察细胞形态,TRAP染色计数细胞形成数量;采用骨吸收功能试验检测破骨样细胞骨吸收陷窝的数量和面积。结果 RA患者外周血单核细胞上Tim-3表达水平[(77.31±10.66)%]明显高于健康人对照组[(51.72±16.69)%],差异有统计学意义(t=7.593,P<0.01),不同Tim-3水平的单核细胞在诱导为破骨样细胞时, Tim-3高水平组骨吸收陷窝面积[(1.054±0.085)S/mm^2]明显低于中间水平组[(1.889±0.053)S/mm^2]和低水平组[(2.763±0.066)S/mm^2],差异有统计学意义(F=9.318,P<0.05)。结论 Tim-3可能对破骨样细胞的骨吸收功能具有负向调控作用。

Objective To investigate the effects of Tim-3 on osteoclast-like cell(OLC) formation and bone resorption induced by peripheral blood mononuclear cells(PBMCs). Methods The expression levels of Tim-3 in of rheumatoid arthritis(RA) patients and healthy controls were detected by flow cytometry. The OLCs were induced by human PBMCs in vitro. The expression levels of tartrate-resistant acid phosphatase(TRAP), cathepsin K(CTSK) and matrix metalloproteinase 9(MMP9) mRNAs in the formation of OLCs were detected by real-time quantitative PCR. The morphology of OLCs was observed by Wright′s staining and G-actin staining, and the number of OLCs was counted by TRAP staining. The number and area of bone resorption pits in OLCs were detected by the Corning Osteo Assay Surface. Results The expression levels of Tim-3 in PBMCs of RA patients([77.31±10.66]%) were significantly higher than that of healthy controls([51.72±16.69]%, t=7.593, P<0.01). When PBMCs with different Tim-3 levels were induced into OLCs, the area of bone resorption pits in the high Tim-3 level group([1.054±0.085] S/mm^2) were significantly lower than those in the intermediate Tim-3 level group([1.889±0.053] S/mm^2) and the low Tim-3 level group([2.763±0.066] S/mm^2, F=9.318, P<0.05). Conclusion Tim-3 may negatively regulate the bone resorption of OLCs.