Analysis of Skeletal Phenotypes of Matrix-Gla Protein (MGP) and Twisted Gastrulation (Twsg1) Deficient Mice.

Abstract

Matrix Gla-Protein (MGP) and Twisted Gastrulation (Twsg) are secreted proteins that reside in the extracellular matrix. MGP is a potent inhibitor of mineralization in both softand hard- tissues. MGP-/- mice exhibit numerous abnormalities, the most severe being aortic calcification that leads to death by approximately two months of age. Despite extensive studies using cartilage and cardiovascular tissues in these mice, little is known about how MGP deficiency affects bone development, specifically the impact on osteoblasts and osteoclasts. Twisted Gastrulation is a modulator of BMP signaling in many tissues. Twsg1 deficient mice exhibit several phenotypes including craniofacial, salivary gland defects, and skeletal defects. Twsg1 has also been implicated in embryo patterning and cartilage development but to date, there has been no study of the bone in these mice. Preliminary analysis using faxitron (x-ray) analysis of MGP and Twsg1 wild type and homozygous deficient mice revealed reduced bone and an osteopenic bone phenotype. The goal of my thesis project was to characterize 1) the in vivo (both dynamic and static) phenotype of Twsg-/- mice and 2) both in vivo and in vitro (static) skeletal phenotype of MGP-/- mice. Using microcomputer tomography (μCT), both static, and in the case of Twsg1, dynamic, histomorphometric analysis was performed to characterize the skeletal phenotype in the animals. In both mouse models there was a significant decrease in both cortical and trabecular bone present. Primary osteoblasts and osteoclasts were used to further characterize the in vitro phenotype in the MGP-/- mouse model. Primary osteoblasts showed premature mineralization and differentiation. Bone sialoprotein (BSP) and osteoclacin (OCN) mRNA levels were both elevated. Surprisingly, osteoclast differentiation also showed enhancement with increased number and larger multinuclear TRAP+ osteoclasts. They also showed increases in DC-STAMP and TRAP mRNA levels compared to the wild type animals. The data collected in this study was used to evaluate the skeletal phenotype of the MGP and Twsg1-knockout mouse models to better understand the relationship of osteoporosis, BMP signaling, and mineral metabolism.