|dc.description.abstract||Genetic disorders associated with skeletal disease are extremely complex and vary greatly in their clinical phenotypes. Thus, in vivo models that accurately recapitulate these rare bone pathologies are essential for understanding the mechanisms of disease and can serve as tools for evaluating therapeutic treatments while providing insight into normal bone physiology. However, by their nature these rare diseases combined with a deficiency in animal models can significantly delay mechanistic understanding and even therapeutic development. Therefore, the goal of this work was to evaluate murine models of Down syndrome (DS) and develop an appropriate model of human Hypophosphatasia (HPP).
The low bone mass phenotype in DS is defined by low bone turnover due to decreased osteoclast and osteoblast activity, decreasing the utility of anti-resorptive agents in people with DS. Thus, we examined the effects of a known bone anabolic agent – sclerostin antibody (SclAb). Male Ts65Dn and age-matched WT littermate mice (8 weeks old) were treated with 4 weekly i.v. injections of 100 mg/kg SclAb. Analysis by DXA, microCT, and ex vivo bone marrow cultures revealed that SclAb had a significant anabolic effect on both controls and Ts65Dn DS mice that was osteoblast mediated, without significant changes in osteoclast numbers. Additionally, comparative gene profiling by RNAseq of whole femurs from 7 month old male Ts65Dn mice and WT provided insight into the molecular mechanisms underlying this unusual and rare bone phenotype.
Moreover, we successfully generated the first large animal model of a rare human bone disease using CRISPR/Cas9 to introduce a single point mutation in the tissue nonspecific alkaline phosphatase (TNSALP) gene (ALPL) (1077C>G) in sheep, thereby creating HPP. Compared to wild-type (WT) controls, HPP sheep have reduced serum alkaline phosphatase activity, decreased tail vertebral bone size, and metaphyseal flaring, consistent with mineralization deficits observed in human HPP. Oral radiographs and computed tomography revealed thin dentin and wide pulp chambers in incisors, and radiolucency of jaws in HPP vs. WT sheep. Skeletal muscle biopsies reveal aberrant fiber size and mitochondrial cristae structure in HPP vs. WT sheep. These genetically engineered sheep phenocopy HPP and provide a novel large animal platform for the longitudinal study of HPP progression, as well as other rare bone diseases.||en