The Role of Osteoblasts in Phenotypic Variability of Dominant Osteogenesis Imperfecta: Evidence from Patients and Murine Models

One of the hallmarks of Osteogenesis Imperfecta (OI) is phenotypic variability among individuals with the same mutation. The aim of our study is to investigate the under-explored role of osteoblast differentiation in OI phenotypic variability by using human and murine OI osteoblasts. This is the first comparative study of osteoblasts from OI patients vs. healthy pediatric controls. We investigated osteoblasts carrying <i>COL1A1</i> substitutions Gly352Ser and Gly589Ser, each expressed in two unrelated patients differing in phenotypic severity. Osteoblasts from type III OI patients with both mutations deposited significantly less mineral vs. type IV. RNA-Seq showed osteoblasts from type IV OI patients with different mutations had downregulated mitochondrial pathways, while osteoblasts from type III OI patients showed downregulation of extracellular matrix pathways. Puromycin assay demonstrated osteoblast protein synthesis was significantly upregulated in type III vs. type IV OI patients. UPR PERK and BiP were reduced in osteoblasts with Gly352Ser from type III and IV OI patients and in osteoblasts with Gly589Ser from a type III OI patient, while both proteins were increased in Gly589Ser osteoblasts from the type IV patient. Additionally, in a murine comparative study, <i>Col1a1 Gly349Ser</i>, called Brtl Ser, showed a much more severe skeletal phenotype than Brtl Cys. Brtl Ser calvarial osteoblasts had reduced collagen secretion and folding with abnormal dermal collagen fibrils vs. wildtype. Also, Brtl Ser osteoblasts showed condensed actin filaments but a similar mineral deposition as Brtl Cys. Electron microscopy revealed elongated mitochondria with cristae dropout in patient and mutant murine osteoblasts. Our study yielded novel insights highlighting osteoblast differentiation, mineralization, and a potential role of mitochondria in OI pathology and phenotypic variability.