The long-term goal of the Zayzafoon laboratory is to develop novel therapeutic approaches for building and retaining bone mass in humans. The immediate goal of this proposed work is to unequivocally determine the role of the Calcineurin (Cn) and Nuclear Factor of Activated T Cells (NFAT) signaling pathway in regulating bone mass. Bone is a highly dynamic structure that is constantly renewing through a process called remodeling. Cn and NFAT are key promoters of osteoclast differentiation and bone resorption. In contrast, the role of Cn and NFAT in osteoblast differentiation and bone formation is not well defined. We discovered that the pharmacologic inhibition of Cn by low concentrations of Cyclosporin A (CsA) increased osteoblast differentiation in vitro and bone mass in vivo. This increase in osteoblast differentiation was associated with a similar increase in the expression of Fra-2, an AP-1 family member. To determine whether Cn directly impacts osteoblast differentiation, we utilized Cre-mediated recombinase technology to create mice lacking the calcineurin regulatory subunit (Cnb1) specifically in osteoblasts. Our results demonstrate that a block in the Cn/NFAT signaling pathway, when restricted to osteoblasts, leads to an increase in bone mass by directly increasing osteoblast differentiation and indirectly decreasing osteoclastogenesis. Therefore, we hypothesize that the Cn/NFAT signaling pathway negatively regulates osteoblast differentiation and bone mass by regulating the expression and activation of AP-1 family of transcription factors.
The Specific Aims of this study are: 1. Determine how the disruption of CN/NFAT signaling in osteoblasts affects bone phenotype. 2. Characterize the roles of Cn/NFAT in osteoblasts growth and differentiation. 3. Determine the molecular mechanisms by which Cn/NFAT regulates bone mass. Previous publications characterize the Cn/NFAT pathway as a positive regulator of bone formation. However, the data from mice with osteoblast-specific knockout of this pathway suggest the opposite - that Cn/NFAT is a negative regulator of bone formation. We are dedicated to definitively resolving this issue because of the therapeutic potential of using inhibitors of Cn/NFAT to anabolically increase bone mass. Our preliminary data provide strong evidence that Cn/NFAT is a negative regulator of bone mass. Completion of these aims will enable us to unequivocally determine the roles of Cn/NFAT signaling in osteoblast differentiation and bone mass. The impact of our work may be far reaching, because it will provide better understanding of the mechanisms by which Cn/NFAT regulates osteoblast differentiation and will have important implications in the development of new anabolic drugs targeting Cn/NFAT pathway to treat osteoporosis and bone loss.

Public Health Relevance

We will unequivocally determine the roles of Cn/NFAT signaling in osteoblasts in regulating bone formation and bone mass. The impact of our work may be far reaching, because it will have important implications in the development of new anabolic drugs targeting Cn/NFAT pathway to treat osteoporosis and bone loss. ? ? ? ?

Agency
National Institute of Health (NIH)
Institute
National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIAMS)
Type
Research Project (R01)
Project #
5R01AR053898-02
Application #
7496442
Study Section
Special Emphasis Panel (ZRG1-MOSS-L (04))
Program Officer
Sharrock, William J
Project Start
2007-09-11
Project End
2012-05-31
Budget Start
2008-06-01
Budget End
2009-05-31
Support Year
2
Fiscal Year
2008
Total Cost
$305,515
Indirect Cost
Name
University of Alabama Birmingham
Department
Pathology
Type
Schools of Medicine
DUNS #
063690705
City
Birmingham
State
AL
Country
United States
Zip Code
35294
Ofotokun, Ighovwerha; Titanji, Kehmia; Lahiri, Cecile D et al. (2016) A Single-dose Zoledronic Acid Infusion Prevents Antiretroviral Therapy-induced Bone Loss in Treatment-naive HIV-infected Patients: A Phase IIb Trial. Clin Infect Dis 63:663-671
Daft, Paul G; Yang, Yang; Napierala, Dobrawa et al. (2015) The growth and aggressive behavior of human osteosarcoma is regulated by a CaMKII-controlled autocrine VEGF signaling mechanism. PLoS One 10:e0121568
Ofotokun, Ighovwerha; Titanji, Kehmia; Vikulina, Tatyana et al. (2015) Role of T-cell reconstitution in HIV-1 antiretroviral therapy-induced bone loss. Nat Commun 6:8282
Titanji, Kehmia; Vunnava, Aswani; Sheth, Anandi N et al. (2014) Dysregulated B cell expression of RANKL and OPG correlates with loss of bone mineral density in HIV infection. PLoS Pathog 10:e1004497
Sesler, Cheryl L; Zayzafoon, Majd (2013) NFAT signaling in osteoblasts regulates the hematopoietic niche in the bone microenvironment. Clin Dev Immunol 2013:107321
Daft, Paul G; Yuan, Kaiyu; Warram, Jason M et al. (2013) Alpha-CaMKII plays a critical role in determining the aggressive behavior of human osteosarcoma. Mol Cancer Res 11:349-59
Vikulina, Tatyana; Fan, Xian; Yamaguchi, Masayoshi et al. (2010) Alterations in the immuno-skeletal interface drive bone destruction in HIV-1 transgenic rats. Proc Natl Acad Sci U S A 107:13848-53
Sloofman, Laura G; Verdelis, Kostas; Spevak, Lyudmila et al. (2010) Effect of HIP/ribosomal protein L29 deficiency on mineral properties of murine bones and teeth. Bone 47:93-101
Choo, Min-Kyung; Yeo, Hyeonju; Zayzafoon, Majd (2009) NFATc1 mediates HDAC-dependent transcriptional repression of osteocalcin expression during osteoblast differentiation. Bone 45:579-89
Yeo, Hyeonju; Beck, Lauren H; Thompson, Sunnie R et al. (2007) Conditional disruption of calcineurin B1 in osteoblasts increases bone formation and reduces bone resorption. J Biol Chem 282:35318-27