Bone disorders and deformities are prevalent in children and young adults. Due to lack of effective modalities to regenerate growing bones, these young patients often undergo multiple surgical interventions, posing a significant burden on them, their family and society. During bone growth, chondrocytes and osteoblasts are continuously generated to make bones bigger and stronger. Endogenous bone stem cells that serve as the source of these cells have not been completely understood. Fundamental knowledge of how these bone stem cells coordinate the two processes of endochondral and intramembranous ossification is essential to develop a reliable approach to regenerate growing bones. In this project, the characteristics of distinct types of bone stem cells that actively promote bone growth will be identified. We hypothesize that a subset of resting chondrocytes in the postnatal growth plate behave as growth-associated bone stem cells, and become a source of mesenchymal stromal progenitor cells in bone marrow; these two types of bone stem/progenitor cells concertedly promote proper bone growth and maintenance. Identifying characteristics and molecular regulations of bone stem cells will facilitate our endeavor to reproduce these cells through regenerative engineering.
In Aim1, we will identify molecular mechanisms regulating properties and fates of resting chondrocytes. The working hypothesis is that resting chondrocytes of the postnatal growth plate exhibit unique characteristics as growth-associated bone stem cells, whose properties and fates are regulated by Hedgehog signaling. We will first identify a self-renewing multipotent subpopulation of resting chondrocytes using in vitro colony-forming assays and in vivo transplantation of isolated growth plate cells. We will second identify the unique gene expression profiles of self-renewing colony-forming resting chondrocytes. We will further define roles of Hedgehog signaling in determining self-renewal and differentiation of resting chondrocytes by modulating its signaling components, while simultaneously tracing their behavior both in vivo and in vitro.
In Aim2, we will define formation and fates of bone marrow mesenchymal stromal progenitors in growing bones. The working hypothesis is that growth plate chondrocytes undergo hypertrophy and transform into Cxcl12- abundant reticular (CAR) cells in bone marrow that behave as regional and reactive mesenchymal stromal progenitor cells. We will first define differentiation potentials of CAR cells into osteoblasts and adipocytes in vivo through intermittent PTH administration and a high-fat diet containing rosiglitazone. We will second determine CAR cells' response to injury using a semistabilized tibial fracture model. We will also identify effects of these manipulations on CAR cells' expression levels of key transcription factors that regulate cell fate choice. We will third define the properties of CAR cells as mesenchymal stromal progenitors through in vitro colony- forming assays and in vivo transplantation of isolated bone marrow cells. We will further define roles of ?- catenin signaling as a cell fate determinant of osteoblast versus adipocyte differentiation using its floxed allele.
In this project, we will reveal novel dynamics and molecular regulations of bone stem cells that control bone growth and maintenance in two ossification mechanisms. The impact of this study is that it will provide an essential platform to understand the fundamental properties of bone stem cells that can regenerate functional growing bones. Defining the characteristics of distinct types of bone stem cells will contribute to the development of inherent bone regenerative strategies for clinical and translational applications to treat children and young adults with bone deformities.
|Sakagami, N; Matsushita, Y; Syklawer-Howle, S et al. (2018) Msx2 Marks Spatially Restricted Populations of Mesenchymal Precursors. J Dent Res 97:1260-1267|
|Mizuhashi, Koji; Ono, Wanida; Matsushita, Yuki et al. (2018) Resting zone of the growth plate houses a unique class of skeletal stem cells. Nature 563:254-258|
|Takahashi, Akira; Nagata, Mizuki; Gupta, Aditi et al. (2018) Autocrine regulation of mesenchymal progenitor cell fates orchestrates tooth eruption. Proc Natl Acad Sci U S A :|