DDRGK1 is required for the proper development and maintenance of the growth plate cartilage.

Loss-of-function mutations in DDRGK1 have been shown to cause Shohat type spondyloepimetaphyseal dysplasia (SEMD). In zebrafish, loss of function of ddrgk1 leads to defects in early cartilage development. Ddrgk1-/- mice show delayed mesenchymal condensation in the limb buds and early embryonic lethality. Mechanistically, Ddrgk1 interacts with Sox9 and reduces ubiquitin-mediated proteasomal degradation of Sox9 protein. To investigate the cartilage-specific role of DDRGK1, conditional knockout mice were generated by intercrossing Prx1-Cre transgenic mice with Ddrgkfl/fl mice to delete its expression in limb mesenchymal cells. Mutant mice showed progressive severe shortening of the limbs and joint abnormalities. The growth plate showed disorganization with shortened proliferative zone and enlarged hypertrophic zone. In correlation with these findings, Sox9 and Col2a1 protein levels were decreased, while Col10a1 expression was expanded. These data demonstrate the importance of Ddrgk1 during growth plate development. In contrast, deletion of Ddrgk1 with the osteoblast-specific Osteocalcin-Cre and Leptin receptor-Cre lines did not show bone phenotypes, suggesting that the effect on limb development is cartilage-specific. To evaluate the role of DDRGK1 in cartilage postnatal homeostasis, inducible Agc1-CreERT2; Ddrgklfl/fl mice were generated. Mice in which Ddrgk1 was deleted at 3 months of age showed disorganized growth plate, with significant reduction in proteoglycan deposition. These data demonstrate a postnatal requirement for Ddrgk1 in maintaining normal growth plate morphology. Together, these findings highlight the physiological role of Ddrgk1 in the development and maintenance of the growth plate cartilage. Furthermore, these genetic mouse models recapitulate the clinical phenotype of short stature and joint abnormalities observed in patients with Shohat type SEMD.

Localized chondro-ossification underlies joint dysfunction and motor deficits in the Fkbp10 mouse model of osteogenesis imperfecta.

Osteogenesis imperfecta (OI) is a genetic disorder that features wide-ranging defects in both skeletal and nonskeletal tissues. Previously, we and others reported that loss-of-function mutations in FK506 Binding Protein 10 (FKBP10) lead to skeletal deformities in conjunction with joint contractures. However, the pathogenic mechanisms underlying joint dysfunction in OI are poorly understood. In this study, we have generated a mouse model in which Fkbp10 is conditionally deleted in tendons and ligaments. Fkbp10 removal substantially reduced telopeptide lysyl hydroxylation of type I procollagen and collagen cross-linking in tendons. These biochemical alterations resulting from Fkbp10 ablation were associated with a site-specific induction of fibrosis, inflammation, and ectopic chondrogenesis followed by joint deformities in postnatal mice. We found that the ectopic chondrogenesis coincided with enhanced Gli1 expression, indicating dysregulated Hedgehog (Hh) signaling. Importantly, genetic inhibition of the Hh pathway attenuated ectopic chondrogenesis and joint deformities in Fkbp10 mutants. Furthermore, Hh inhibition restored alterations in gait parameters caused by Fkbp10 loss. Taken together, we identified a previously unappreciated role of Fkbp10 in tendons and ligaments and pathogenic mechanisms driving OI joint dysfunction.

Fracture Healing in Collagen-Related Preclinical Models of Osteogenesis Imperfecta.

Osteogenesis imperfecta (OI) is a genetic bone dysplasia characterized by bone deformities and fractures caused by low bone mass and impaired bone quality. OI is a genetically heterogeneous disorder that most commonly arises from dominant mutations in genes encoding type I collagen (COL1A1 and COL1A2). In addition, OI is recessively inherited with the majority of cases resulting from mutations in prolyl-3-hydroxylation complex members, which includes cartilage-associated protein (CRTAP). OI patients are at an increased risk of fracture throughout their lifetimes. However, non-union or delayed healing has been reported in 24% of fractures and 52% of osteotomies. Additionally, refractures typically go unreported, making the frequency of refractures in OI patients unknown. Thus, there is an unmet need to better understand the mechanisms by which OI affects fracture healing. Using an open tibial fracture model, our study demonstrates delayed healing in both Col1a2 G610c/+ and Crtap -/- OI mouse models (dominant and recessive OI, respectively) that is associated with reduced callus size and predicted strength. Callus cartilage distribution and chondrocyte maturation were altered in OI, suggesting accelerated cartilage differentiation. Importantly, we determined that healed fractured tibia in female OI mice are biomechanically weaker when compared with the contralateral unfractured bone, suggesting that abnormal OI fracture healing OI may prime future refracture at the same location. We have previously shown upregulated TGF-ß signaling in OI and we confirm this in the context of fracture healing. Interestingly, treatment of Crtap -/- mice with the anti-TGF-ß antibody 1D11 resulted in further reduced callus size and predicted strength, highlighting the importance of investigating dose response in treatment strategies. These data provide valuable insight into the effect of the extracellular matrix (ECM) on fracture healing, a poorly understood mechanism, and support the need for prevention of primary fractures to decrease incidence of refracture and deformity in OI patients. © 2020 American Society for Bone and Mineral Research.