Previous investigations from our laboratory revealed that Notch plays a critical role in cell differentiation, and its expression in immature osteoblast causes osteopenia secondary to an inhibitory effect on osteoblastogenesis. The effect of Notch is cell context-dependent, and in contrast to the effects observed in osteoblasts, when Notch is expressed in osteocytes under the control of the Dentin matrix protein 1 (Dmp1) promoter it causes a pronounced increase in trabecular and cortical bone volume. In cancellous bone, Notch inhibits bone resorption due to a decrease in the number of osteoclasts. In cortical bone Notch enhances mineral apposition rate and the structure maintains a trabecular appearance. Our observations led to the hypothesis that Notch plays a central role in the regulation of osteocyte function, and Notch effects in osteocytes are unique and distinct from those observed in osteoblasts. Notch induces osteoprotegerin and inhibits the expression of the Wnt antagonist sclerostin enhancing the expression of Wnt target genes. In initial experiments, we discovered that expression of Notch in osteocytes prevents the bone loss that follows muscle paralysis, and we postulate that Notch protects bone from the detrimental effects of unloading.
The aim of the proposed research is to understand the function of Notch in osteocytes and define the mechanisms involved. For this purpose, we will study mouse and cellular models expressing Notch in the osteocyte environment, and models of conditional inactivation of Notch1 and Notch2 in osteocytes.
Our specific aims are: 1) To determine the function of Notch in osteocytes in the adult skeleton by transgenic expression of Notch under the control of the Dmp1 promoter using inducible models of expression and by targeted Notch1 and Notch2 conditional deletion in osteocytes. The skeletal phenotype of mice misexpressing Notch will be compared to that of wild type mice and determined by contact radiography, densitometry, micro CT scanning and histomorphometry. In addition, we will analyze the biochemical and biomechanical properties of bone;2) To determine the mechanism of action of Notch in osteocytes. Mechanisms responsible for the effects of Notch will be established, and we will determine whether the Notch canonical signaling pathway is responsible for the effects of Notch in osteocytes and the role of mechanotransduction on canonical Notch signaling in vivo. The impact of Notch on the expression of sclerostin and osteoprotegerin in osteocytes, their contribution to the phenotype observed and levels of transcriptional and post-transcriptional regulation will be examined in in vivo and in vitro models;and 3) To establish whether Notch protects the skeleton from the detrimental effects of unloading, as it does from those of muscle paralysis.

Public Health Relevance

These investigations will clarify the role of Notch in osteocytes and offer new information on mechanisms involved in the function of osteocytes, mechanotransduction in bone and in the pathogenesis and reversal of disuse osteoporosis.

Agency
National Institute of Health (NIH)
Type
Research Project (R01)
Project #
5R01AR063049-02
Application #
8634022
Study Section
Skeletal Biology Structure and Regeneration Study Section (SBSR)
Program Officer
Chen, Faye H
Project Start
Project End
Budget Start
Budget End
Support Year
2
Fiscal Year
2014
Total Cost
Indirect Cost
Name
St. Francis Hospital and Medical Center
Department
Type
DUNS #
City
Hartford
State
CT
Country
United States
Zip Code
06105
Canalis, Ernesto; Zanotti, Stefano; Smerdel-Ramoya, Anna (2014) Connective tissue growth factor is a target of notch signaling in cells of the osteoblastic lineage. Bone 64:273-80
Canalis, Ernesto; Kranz, Lauren; Zanotti, Stefano (2014) Nemo-like kinase regulates postnatal skeletal homeostasis. J Cell Physiol 229:1736-43
Zanotti, Stefano; Kalajzic, Ivo; Aguila, Hector Leonardo et al. (2014) Sex and genetic factors determine osteoblastic differentiation potential of murine bone marrow stromal cells. PLoS One 9:e86757
Zanotti, Stefano; Canalis, Ernesto (2014) Notch1 and Notch2 expression in osteoblast precursors regulates femoral microarchitecture. Bone 62:22-8
Canalis, Ernesto; Adams, Douglas J; Boskey, Adele et al. (2013) Notch signaling in osteocytes differentially regulates cancellous and cortical bone remodeling. J Biol Chem 288:25614-25
Zanotti, S; Canalis, E (2013) Interleukin 6 mediates selected effects of Notch in chondrocytes. Osteoarthritis Cartilage 21:1766-73
Canalis, Ernesto (2013) Wnt signalling in osteoporosis: mechanisms and novel therapeutic approaches. Nat Rev Endocrinol 9:575-83