Tan Zhijia, Shek Hiu Tung, Li Zeluan, Xia Linjian, He Yanni, Chen Peikai, Wong Janus Siu Him, Gao Bo, Chan Danny, To Michael Kai Tsun
Department of Orthopaedics and Traumatology, The University of Hong Kong-Shenzhen Hospital, Shenzhen 518053, China.
Shenzhen Clinical Research Centre for Rare Diseases, The University of Hong Kong-Shenzhen Hospital, Shenzhen 518053, China.
J Bone Miner Res. 2025 May 24;40(5):577-590. doi: 10.1093/jbmr/zjaf022.
Osteogenesis imperfecta (OI) type V is typically characterized by radial head dislocation, calcification of interosseous membrane, and hyperplastic callus. It is caused by the c.-14C>T mutation in the 5' UTR of IFITM5 gene, adding 5 amino acids (MALEP) to the N-terminal of IFITM5 protein. Previous studies have suggested a neomorphic function of the MALEP-IFITM5 protein. However, the underlying mechanisms remain unclear due to embryonic lethality in previous mouse models. Therefore, we developed an inducible mouse model (Ifitm5flox c.-14C>T) that could be induced by Cre expressed at different developmental stages to explore the pathogenic effects of the neomorphic MALEP-IFITM5. The mutant Ifitm5 allele could be regulated by the endogenous regulatory elements after Cre recombination, maintaining its spatiotemporal expression pattern and physiological level. Specifically, Prx1-Cre; Ifitm5flox c.-14C>T mutant mice were born with fractures in all limbs, showing impaired ossification and enhanced chondrogenesis associated with increased SOX9 abundance. Analyses of single-cell RNA sequencing data revealed arrested osteogenesis in Prx1-Cre; Ifitm5flox c.-14C>T mouse. A major population of cells expressing both osteogenic and chondrogenic signature genes was identified in the mutant mouse. Reduced expression of SP7 and SOST in the cortical regions of mutant mice confirmed delayed osteocyte maturation and compromised osteogenesis. Elevated bone marrow adipocytes were found in the adult mutant mice. Ectopic chondrogenesis and SOX9 expression were also observed in the perichondrium regions of Col1a1-Cre; Ifitm5flox c.-14C>T and Ocn-Cre; Ifitm5flox c.-14C>T mutant mice. The inducible Ifitm5flox c.-14C>T mouse model and integrated single-cell transcriptomic analyses elucidated that ectopic expression of SOX9 and disrupted homeostatic balance among osteogenesis, chondrogenesis, and adipogenesis may contribute to the pathogenesis caused by MALEP-IFITM5, helping to gain deeper insights into the molecular mechanisms of type V OI.
V型成骨不全症（OI）的典型特征是桡骨头脱位、骨间膜钙化和骨痂增生。它是由IFITM5基因5'UTR中的c.-14C>T突变引起的，该突变在IFITM5蛋白的N端添加了5个氨基酸（MALEP）。先前的研究表明MALEP-IFITM5蛋白具有新功能。然而，由于先前小鼠模型中的胚胎致死性，其潜在机制仍不清楚。因此，我们开发了一种可诱导的小鼠模型（Ifitm5flox c.-14C>T），该模型可由在不同发育阶段表达的Cre诱导，以探索新功能的MALEP-IFITM5的致病作用。Cre重组后，突变的Ifitm5等位基因可由内源性调控元件调控，维持其时空表达模式和生理水平。具体而言，Prx1-Cre; Ifitm5flox c.-14C>T突变小鼠出生时四肢均有骨折，表现出骨化受损和软骨生成增强，同时SOX9丰度增加。单细胞RNA测序数据分析显示Prx1-Cre; Ifitm5flox c.-14C>T小鼠的成骨停滞。在突变小鼠中鉴定出大量同时表达成骨和软骨生成特征基因的细胞群。突变小鼠皮质区域中SP7和SOST的表达降低，证实骨细胞成熟延迟和成骨受损。在成年突变小鼠中发现骨髓脂肪细胞增多。在Col1a1-Cre; Ifitm5flox c.-14C>T和Ocn-Cre; Ifitm5flox c.-14C>T突变小鼠的软骨膜区域也观察到异位软骨生成和SOX9表达。可诱导的Ifitm5flox c.-14C>T小鼠模型和整合的单细胞转录组分析表明，SOX9的异位表达以及成骨、软骨生成和脂肪生成之间的稳态平衡破坏可能导致MALEP-IFITM5引起的发病机制，有助于更深入地了解V型OI的分子机制。
