Warman, M. | Search bioRxiv

bioRxiv2025

Enhancing Wnt signaling lowers fracture incidence in a severe mouse model of Osteogenesis Imperfecta

Increased risk of fracture is a hallmark feature in the skeletal disorder Osteogenesis Imperfecta (OI). One therapy in clinical trials for OI is an antibody that neutralizes sclerostin (a Wnt signaling inhibitor) that is already FDA approved for patients with osteoporosis. In addition to increasing bone mass and lowering incident fractures in patients with osteoporosis, anti-sclerostin antibody increased bone mass and skeletal strength (measured ex vivo) in several preclinical models of OI. Because in vivo fracture incidence is the primary outcome measure in human OI trials, we examined the effect of enhancing Wnt signaling in a preclinical mouse model of moderate-severe, autosomal dominant OI (the Col1a1Aga2/+ mouse). We genetically enhanced Wnt signaling using the Lrp5A214V allele, which makes the Wnt co-receptor LRP5 resistant to inhibition by sclerostin. By crossing Col1a1Aga2/+ sires with Lrp5A214V/+ dams, we generated pups with OI alone and OI with enhanced Wnt signaling. We radiographically examined these animals for fracture at 5, 9, and 13 weeks of age. We observed that enhanced Wnt signaling significantly reduced the number of fractures in this model by 30% at all 3 ages (e.g., 5.4 {+/-} 1.7 fractures compared to 7.8 {+/-} 1.8 fractures at 5-weeks-old, p < 0.001). These data indicate enhancing Wnt signaling lowers incident fractures in a pre-clinical model of OI and suggest lower fracture incidence should achieved by giving anti-sclerostin antibody to patients with OI. These data also suggest that bone mass could be used as a surrogate marker for lowering fracture risk in patients with OI.

genetics↗

bioRxiv2019

Single Cell RNA Sequencing of Calvarial and Long Bone Endocortical Cells

Single cell RNA-seq (scRNA-seq) is emerging as a powerful technology to examine transcriptomes of individual cells. We determined whether scRNA-seq could be used to detect the effect of environmental and pharmacologic perturbations on osteoblasts. We began with a commonly used in vitro system in which freshly isolated neonatal mouse calvarial cells are expanded and induced to produce a mineralized matrix. We used scRNA-seq to compare the relative cell type abundances and the transcriptomes of freshly isolated cells to those that had been cultured for 12 days in vitro. We observed that the percentage of macrophage-like cells increased from 6% in freshly isolated calvarial cells to 34% in cultured cells. We also found that Bglap transcripts were abundant in freshly isolated osteoblasts but nearly undetectable in the cultured calvarial cells. Thus, scRNA-seq revealed significant differences between heterogeneity of cells in vivo and in vitro. We next performed scRNA-seq on freshly recovered long bone endocortical cells from mice that received either vehicle or Sclerostin-neutralizing antibody for 1 week. Bone anabolism-associated transcripts were also not significantly increased in immature and mature osteoblasts recovered from Sclerostin-neutralizing antibody treated mice; this is likely a consequence of being underpowered to detect modest changes in gene expression, since only 7% of the sequenced endocortical cells were osteoblasts, and a limited portion of their transcriptomes were sampled. We conclude that scRNA-seq can detect changes in cell abundance, identity, and gene expression in skeletally derived cells. In order to detect modest changes in osteoblast gene expression at the single cell level in the appendicular skeleton, larger numbers of osteoblasts from endocortical bone are required.

cell biology↗