Chou Hsin-Chiao, Lin Sung-Yen, Chou Liang-Yin, Ho Mei-Ling, Chuang Shu-Chun, Cheng Tsung-Lin, Kang Lin, Lin Yi-Shan, Wang Yan-Hsiung, Wei Chun-Wang, Chen Chung-Hwan, Wang Chau-Zen
Graduate Institute of Medicine, College of Medicine, Kaohsiung Medical University, Kaohsiung 80708, Taiwan.
Orthopaedic Research Center, Kaohsiung Medical University, Kaohsiung 80708, Taiwan.
Biomedicines. 2022 Sep 2;10(9):2173. doi: 10.3390/biomedicines10092173.
Discoidin domain receptor 1 (DDR1) is a collagen receptor that belongs to the receptor tyrosine kinase family. We have previously shown that DDR1 plays a crucial role during bone development, resulting in dwarfism and a short stature in osteoblast-specific knockout mice (OKO mice). However, the detailed pathophysiological effects of DDR1 on bone development throughout adulthood have remained unclear. This study aims to identify how DDR1 regulates osteoblast and osteocyte functions in vivo and in vitro during bone development in adulthood. The metabolic changes in bone tissues were analyzed using Micro-CT and immunohistochemistry staining (IHC) in vivo; the role of DDR1 in regulating osteoblasts was examined in MC3T3-E1 cells in vitro. The Micro-CT analysis results demonstrated that OKO mice showed a 10% reduction in bone-related parameters from 10 to 14 weeks old and a significant reduction in cortical thickness and diameter compared with flox/flox control mice (FF) mice. These results indicated that DDR1 knockout in OKO mice exhibiting significant bone loss provokes an osteopenic phenotype. The IHC staining revealed a significant decrease in osteogenesis-related genes, including RUNX2, osteocalcin, and osterix. We noted that DDR1 knockout significantly induced osteoblast/osteocyte apoptosis and markedly decreased autophagy activity in vivo. Additionally, the results of the gain- and loss-of-function of the DDR1 assay in MC3T3-E1 cells indicated that DDR1 can regulate the osteoblast differentiation through activating autophagy by regulating the phosphorylation of the mechanistic target of rapamycin (p-mTOR), light chain 3 (LC3), and beclin-1. In conclusion, our study highlights that the ablation of DDR1 results in cancellous bone loss by regulating osteoblast/osteocyte autophagy. These results suggest that DDR1 can act as a potential therapeutic target for managing cancellous bone loss.
盘状结构域受体1（DDR1）是一种属于受体酪氨酸激酶家族的胶原蛋白受体。我们之前已经表明，DDR1在骨骼发育过程中起关键作用，导致成骨细胞特异性敲除小鼠（OKO小鼠）出现侏儒症和身材矮小。然而，DDR1在成年期整个骨骼发育过程中的详细病理生理作用仍不清楚。本研究旨在确定DDR1在成年期骨骼发育过程中如何在体内和体外调节成骨细胞和骨细胞的功能。使用微型计算机断层扫描（Micro-CT）和免疫组织化学染色（IHC）在体内分析骨组织的代谢变化；在体外MC3T3-E1细胞中研究DDR1在调节成骨细胞中的作用。Micro-CT分析结果表明，与flox/flox对照小鼠（FF小鼠）相比，OKO小鼠在10至14周龄时骨相关参数降低了10%，皮质厚度和直径显著减小。这些结果表明，OKO小鼠中DDR1基因敲除导致明显的骨质流失，引发骨质减少的表型。IHC染色显示成骨相关基因，包括RUNX2、骨钙素和osterix显著减少。我们注意到，DDR1基因敲除在体内显著诱导成骨细胞/骨细胞凋亡，并明显降低自噬活性。此外，MC3T3-E1细胞中DDR1功能获得和丧失实验的结果表明，DDR1可以通过调节雷帕霉素机制靶点（p-mTOR）、轻链3（LC3）和beclin-1的磷酸化来激活自噬，从而调节成骨细胞分化。总之，我们的研究强调，DDR1的缺失通过调节成骨细胞/骨细胞自噬导致松质骨流失。这些结果表明，DDR1可以作为治疗松质骨流失的潜在治疗靶点。
