Department of Anatomy & Cell Biology, Indiana University School of Medicine, Indianapolis, IN, USA; Richard L. Roudebush VA Medical Center, Indianapolis, IN, USA; School of Physical Science & Engineering, Anderson University, Anderson, IN, USA.
Department of Biomedical & Applied Sciences, Indiana University School of Dentistry, Indianapolis, IN, USA.
Bone. 2019 Mar;120:166-175. doi: 10.1016/j.bone.2018.10.007. Epub 2018 Oct 25.
High-bone-mass (HBM)-causing missense mutations in the low density lipoprotein receptor-related protein-5 (Lrp5) are associated with increased osteoanabolic action and protection from disuse- and ovariectomy-induced osteopenia. These mutations (e.g., A214V and G171V) confer resistance to endogenous secreted Lrp5/6 inhibitors, such as sclerostin (SOST) and Dickkopf homolog-1 (DKK1). Cells in the osteoblast lineage are responsive to canonical Wnt stimulation, but recent work has indicated that osteoclasts exhibit both indirect and direct responsiveness to canonical Wnt. Whether Lrp5-HBM receptors, expressed in osteoclasts, might alter osteoclast differentiation, activity, and consequent net bone balance in the skeleton, is not known. To address this, we bred mice harboring heterozygous Lrp5 HBM-causing conditional knock-in alleles to Ctsk-Cre transgenic mice and studied the phenotype using DXA, μCT, histomorphometry, serum assays, and primary cell culture. Mice with HBM alleles induced in Ctsk-expressing cells (TG) exhibited higher bone mass and architectural properties compared to non-transgenic (NTG) counterparts. In vivo and in vitro measurements of osteoclast activity, population density, and differentiation yielded significant reductions in osteoclast-related parameters in female but not male TG mice. Droplet digital PCR performed on osteocyte enriched cortical bone tubes from TG and NTG mice revealed that ~8-17% of the osteocyte population (depending on sex) underwent recombination of the conditional Lrp5 allele in the presence of Ctsk-Cre. Further, bone formation parameters in the midshaft femur cortex show a small but significant increase in anabolic action on the endocortical but not periosteal surface. These findings suggest that Wnt/Lrp5 signaling in osteoclasts affects osteoclastogenesis and activity in female mice, but also that some of the changes in bone mass in TG mice might be due to Cre expression in the osteocyte population.
高骨量（HBM）导致低密度脂蛋白受体相关蛋白 5（Lrp5）的错义突变与增加的成骨作用和防止废用性和卵巢切除引起的骨质疏松有关。这些突变（例如 A214V 和 G171V）使内源性分泌的 Lrp5/6 抑制剂，如硬化蛋白（SOST）和 Dickkopf 同源物-1（DKK1）抵抗。成骨细胞谱系中的细胞对经典 Wnt 刺激有反应，但最近的工作表明破骨细胞对经典 Wnt 表现出间接和直接反应。在破骨细胞中表达的 Lrp5-HBM 受体是否会改变破骨细胞分化、活性以及随后对骨骼中净骨平衡的影响尚不清楚。为了解决这个问题，我们将携带杂合 Lrp5 HBM 致条件敲入等位基因的小鼠与 Ctsk-Cre 转基因小鼠杂交，并使用 DXA、μCT、组织形态计量学、血清测定和原代细胞培养研究表型。在 Ctsk 表达细胞中诱导 HBM 等位基因的小鼠（TG）与非转基因（NTG）对照相比，表现出更高的骨量和骨结构特性。体内和体外的破骨细胞活性、种群密度和分化测量表明，在雌性 TG 小鼠中，破骨细胞相关参数显著降低，但在雄性 TG 小鼠中没有。在 TG 和 NTG 小鼠的骨细胞丰富的皮质骨管上进行的液滴数字 PCR 显示，在存在 Ctsk-Cre 的情况下，约 8-17%的骨细胞群体（取决于性别）经历了条件 Lrp5 等位基因的重组。此外，中股骨皮质的骨形成参数显示，在内皮质而非骨膜表面的骨形成作用有一个小但显著的增加。这些发现表明，破骨细胞中的 Wnt/Lrp5 信号传导影响雌性小鼠的破骨细胞发生和活性，但也表明 TG 小鼠骨量变化的部分原因可能是 Cre 在骨细胞群体中的表达。
