OBJECTIVE Increased protein phosphatase magnesium-dependent 1A (PPM1A) levels in patients with ankylosing spondylitis regulate osteoblast differentiation in bony ankylosis; however, the potential mechanisms that regulate osteoclast differentiation in relation to abnormal bone formation remain unclear. This study was undertaken to investigate the relationship of PPM1A to osteoclast differentiation by generating conditional gene-knockout (PPM1Afl/fl ;LysM-Cre) mice and evaluating their bone phenotype. METHODS The bone phenotypes of LysM-Cre mice (n = 6) and PPM1Afl/fl ;LysM-Cre mice (n = 6) were assessed by micro-computed tomography. Osteoclast differentiation was induced by culturing bone marrow-derived macrophages in the presence of RANKL and macrophage colony-stimulating factor (M-CSF), and was evaluated by counting tartrate-resistant acid phosphatase-positive multinucleated cells. Levels of messenger RNA for PPM1A, RANK, and osteoclast-specific genes were examined by real-time quantitative polymerase chain reaction, and protein levels were determined by Western blotting. Surface RANK expression was analyzed by fluorescence flow cytometry. RESULTS The PPM1Afl/fl ;LysM-Cre mice displayed reduced bone mass (P < 0.001) and increased osteoclast differentiation (P < 0.001) and osteoclast-specific gene expression (P < 0.05) compared with their LysM-Cre littermates. Mechanistically, reduced PPM1A function in osteoclast precursors in PPM1Afl/fl ;LysM-Cre mice induced osteoclast lineage commitment by up-regulating RANK expression (P < 0.01) via p38 MAPK activation in response to M-CSF. PPM1A expression in macrophages was decreased by Toll-like receptor 4 activation (P < 0.05). The Ankylosing Spondylitis Disease Activity Score was negatively correlated with the expression of PPM1A in peripheral blood mononuclear cells from patients with axial spondyloarthritis (SpA) (γ = -0.7072, P < 0.0001). CONCLUSION The loss of PPM1A function in osteoclast precursors driven by inflammatory signals contributes to osteoclast lineage commitment and differentiation by elevating RANK expression, reflecting a potential role of PPM1A in dynamic bone metabolism in axial SpA.

中文翻译：

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通过细胞内蛋白磷酸酶镁依赖性1A破骨细胞承诺的负调控。

目的：强直性脊柱炎患者中蛋白磷酸酶镁依赖性1A（PPM1A）水平的升高可调节骨性强直病中成骨细胞的分化。然而，调节破骨细胞分化与异常骨形成有关的潜在机制仍不清楚。通过产生条件基因敲除（PPM1Afl / fl; LysM-Cre）小鼠并评估其骨表型来研究PPM1A与破骨细胞分化的关系。方法用计算机X线断层扫描技术评估LysM-Cre小鼠（n = 6）和PPM1Afl / fl； LysM-Cre小鼠（n = 6）的骨表型。通过在RANKL和巨噬细胞集落刺激因子（M-CSF）存在下培养骨髓来源的巨噬细胞来诱导破骨细胞分化，并且通过计数抗酒石酸的酸性磷酸酶阳性的多核细胞来评估。通过实时定量聚合酶链反应检测PPM1A，RANK和破骨细胞特异性基因的信使RNA水平，并通过Western印迹法测定蛋白质水平。通过荧光流式细胞术分析表面RANK表达。结果PPM1Afl / fl; LysM-Cre小鼠与LysM-Cre同窝仔相比，骨量减少（P <0.001），破骨细胞分化增加（P <0.001），破骨细胞特异性基因表达增加（P <0.05）。从机制上讲，PPM1Afl / fl中破骨细胞前体中PPM1A功能的降低； LysM-Cre小鼠通过响应M-CSF的p38 MAPK激活来上调RANK表达（P <0.01），从而诱导破骨细胞谱系定型。Toll样受体4激活可降低巨噬细胞中PPM1A的表达（P <0.05）。强直性脊柱炎疾病活动评分与轴性脊柱关节炎（SpA）患者外周血单个核细胞中PPM1A的表达呈负相关（γ= -0.7072，P <0.0001）。结论炎症信号驱动破骨细胞前体中PPM1A功能的丧失，通过提高RANK表达，促进了破骨细胞谱系的定向和分化，反映了PPM1A在轴向SpA的动态骨代谢中的潜在作用。0001）。结论炎症信号驱动破骨细胞前体中PPM1A功能的丧失，通过提高RANK表达，促进了破骨细胞谱系的定向和分化，反映了PPM1A在轴向SpA的动态骨代谢中的潜在作用。0001）。结论炎症信号驱动破骨细胞前体中PPM1A功能的丧失，通过提高RANK表达，促进了破骨细胞谱系的定向和分化，反映了PPM1A在轴向SpA的动态骨代谢中的潜在作用。