Myeloid Lineage Ablation of Phlpp1 Regulates M-CSF Signaling and Tempers Bone Resorption in Female Mice.
Myeloid Lineage Ablation of Phlpp1 Regulates M-CSF Signaling and Tempers Bone Resorption in Female Mice.
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DOI：
10.3390/ijms22189702
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发表时间：
2021-09-08
影响因子：
5.6
通讯作者：
Bradley EW
中科院分区：
文献类型：
--
作者：
Karkache IY;Damodaran JR;Molstad DHH;Mansky KC;Bradley EW
先前的研究表明，Phlpp1缺乏会改变小梁骨量并增强M-CSF的反应性，但目前尚不清楚Phlpp1对这种作用的细胞类型和需求。为了了解Phlpp1在髓系细胞中的功能，我们将Phlpp1连接的小鼠与含有LysM-Cre的小鼠杂交。12周龄小鼠股骨远端显微计算机断层扫描显示，Phlpp1雌性条件敲除小鼠的骨体积比总体积增加30%，但我们在雄性Phlpp1 cKOLysM小鼠中没有观察到显著变化。胫骨近端骨组织形态测量进一步显示，Phlpp1 cKOLysM雌性小鼠破骨细胞数量升高，但相反，与对照组相比，血清骨吸收标志物水平降低。成骨细胞数量和骨形成血清标志物不变。体外实验证实，Phlpp1消融增加破骨细胞数量和面积，但限制骨吸收。此外，外源性Phlpp1重组抑制破骨细胞数量。剂量反应实验表明，Phlpp1−/−细胞对M-CSF的反应更强，但用Phlpp1重建则消除了这种效应。此外，小分子介导的Phlpp抑制增强了破骨细胞的数量和大小。Phlpp底物（包括Akt、ERK1/2和pkc - ζ）的磷酸化增强伴随着这些观察结果。相反，在Phlpp抑制剂处理的破骨细胞中，肌动蛋白细胞骨架发生破坏。此外，Phlpp抑制降低了体外培养的牛骨片细胞的吸收。我们的研究结果表明，髓系细胞中Phlpp1的缺乏通过限制骨吸收而增加骨量，同时保持破骨细胞数量不变；此外，我们发现Phlpp1抑制破骨细胞生成并控制对M-CSF的反应。
Prior work demonstrated that Phlpp1 deficiency alters trabecular bone mass and enhances M-CSF responsiveness, but the cell types and requirement of Phlpp1 for this effect were unclear. To understand the function of Phlpp1 within myeloid lineage cells, we crossed Phlpp1 floxed mice with mice harboring LysM-Cre. Micro-computed tomography of the distal femur of 12-week-old mice revealed a 30% increase in bone volume per total volume of Phlpp1 female conditional knockouts, but we did not observe significant changes within male Phlpp1 cKOLysM mice. Bone histomorphmetry of the proximal tibia further revealed that Phlpp1 cKOLysM females exhibited elevated osteoclast numbers, but conversely had reduced levels of serum markers of bone resorption as compared to littermate controls. Osteoblast number and serum markers of bone formation were unchanged. In vitro assays confirmed that Phlpp1 ablation enhanced osteoclast number and area, but limited bone resorption. Additionally, reconstitution with exogenous Phlpp1 suppressed osteoclast numbers. Dose response assays demonstrated that Phlpp1−/− cells are more responsive to M-CSF, but reconstitution with Phlpp1 abrogated this effect. Furthermore, small molecule-mediated Phlpp inhibition enhanced osteoclast numbers and size. Enhanced phosphorylation of Phlpp substrates—including Akt, ERK1/2, and PKCζ—accompanied these observations. In contrast, actin cytoskeleton disruption occurred within Phlpp inhibitor treated osteoclasts. Moreover, Phlpp inhibition reduced resorption of cells cultured on bovine bone slices in vitro. Our results demonstrate that Phlpp1 deficiency within myeloid lineage cells enhances bone mass by limiting bone resorption while leaving osteoclast numbers intact; moreover, we show that Phlpp1 represses osteoclastogenesis and controls responses to M-CSF.
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