Date of Award

2017

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Graduate Group

Cell & Molecular Biology

First Advisor

Jaimo Ahn

Second Advisor

Kurt D. Hankenson

Abstract

The R-spondin family of proteins are Wnt agonists, and the complete embryonic disruption of Rspo2 results in skeletal developmental defects that recapitulate the phenotype observed with Lrp5/6 deficiency. Previous work has shown that Rspo2 is both highly expressed in Wnt stimulated pre-osteoblasts and that its over-expression induces osteoblast differentiation in the same cells, supporting its role as a positive autocrine regulator of osteoblastogenesis. However, the role of Rspo2 in the autocrine regulation of osteoblastogenesis in postnatal bone is under explored. Here, we show that limb-bud progenitor cells from Rspo2 knockout mice undergo reduced mineralization during osteoblastogenesis in vitro, and have a corresponding alteration in their osteogenic gene expression profile. The first Rspo2 conditional knockout (Rspo2floxed) mouse was generated, and Rspo2 was knocked out in osteoblasts by crossing to the OsteocalcinCre mouse line (Ocn-Cre+ Rspo2f/f) to address the autocrine role of Rspo2 in vivo. OcnCre+Rspo2f/f male and female mice at 1, 3, and 6 months were examined. Ocn-Cre+Rspo2f/f mice are decreased in overall body size compared to their littermates, and they have decreased trabecular bone mass due primarily to decreased trabecular thickness. Histomorphometric analysis revealed a similar number of osteoblasts and mineralizing surface per bone surface with a simultaneous decrease in mineral apposition and bone formation rates, suggesting that a decreased mineralization capacity rather than an overall reduced number of osteoblasts is the etiology of the reduced bone volume. In following with this, BM-MSC from Ocn-Cre+Rspo2f/f mice undergo less mineralization in vitro. Overall, Rspo2 loss reduces osteoblastogenesis and mineralization both in vitro and in vivo, leading to reduced bone mass.

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