The goal of the competitive renewal is to test the hypothesis that in the hypoxic niche of the intervertebral disc, notch signaling is required for maintenance of proliferation and differentiation of cell populations. The objective of the first Specific Aim is to characterize the expression of notch and notch ligands and to investigate the importance of notch signaling in proliferation and differentiation of disc cells;to assess notch and notch ligand expression in tissue isolated from patients with degenerative disc disease. Using loss and gain of function studies we will determine the importance of the notch system in terms of cell proliferation and acquisition of the mature phenotype. By evaluating the localization of notch receptors and ligands we will gain insight into the interaction between different cell types within the disc. The goal of the second Specific Aim is to ascertain if in the hypoxic microenvironment notch and HIF signaling co-regulate expression of selective notch receptors, ligands and notch target genes in disc cells;in addition, we will determine if notch target genes Hey1 and Hey2 influence HIF function. In parallel, the interaction between notch and HIF will be assessed to evaluate the involvement of notch-ICD in the regulation of HIF-1 target genes. A final goal is to examine proliferative cells isolated from degenerate human discs for the expression and activities of notch signaling components. In the third Aim we will use a notochord specific knockout mice to investigate the importance of notch signaling in governing cell proliferation, survival and function during disc development and to delineate the role of notch in modulating HIF-1 function in the disc. Since we will also delete HIF-1, these studies will reveal the importance of this gene in regulating notch activity. The goal of the last Specific Aim is to develop an animal model to learn if inactivation of notch activity promotes a loss of cellularity, a decrease in number of differentiated cells and eventual degeneration of the intervertebral disc. We will inject rabbit intervertebral discs with lentivirus encoding psen1/psen2-SiRNA to suppress expression of endogenous notch activity. We will measure the activities of components of the notch signaling system, HIF function, the activities and numbers of proliferative cells and assess the degenerative status of the disc itself. These studies will validate and further extend the investigations described above, and provide new information on the critical role of notch signaling in terms of proliferation, survival, and differentiation of disc cells. Results of the investigation should provide a new understanding of how cell proliferation is regulated in the disc and provide a strategy for future use of pharmacological agents, and molecular genetic or tissue engineering approaches for effecting tissue repair utilizing endogenous disc cells.

Public Health Relevance

Lower back pain is closely linked to degenerative disc disease, a problem that afflicts most of the adult population. One approach to enhancing disc health is to promote the activity of stem cells that are resident in the disc. The goal of the proposal is to understand the mechanism that controls the fate and function of these progenitor cells in both the normal and diseased intervertebral disc.

Agency
National Institute of Health (NIH)
Institute
National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIAMS)
Type
Research Project (R01)
Project #
5R01AR050087-09
Application #
8468642
Study Section
Musculoskeletal Tissue Engineering Study Section (MTE)
Program Officer
Wang, Fei
Project Start
2005-02-01
Project End
2014-03-31
Budget Start
2013-04-01
Budget End
2014-03-31
Support Year
9
Fiscal Year
2013
Total Cost
$316,382
Indirect Cost
$86,358
Name
Thomas Jefferson University
Department
Orthopedics
Type
Schools of Medicine
DUNS #
053284659
City
Philadelphia
State
PA
Country
United States
Zip Code
19107
Tran, Cassie M; Schoepflin, Zachary R; Markova, Dessislava Z et al. (2014) CCN2 suppresses catabolic effects of interleukin-1? through ?5?1 and ?V?3 integrins in nucleus pulposus cells: implications in intervertebral disc degeneration. J Biol Chem 289:7374-87
Hirose, Yuichiro; Johnson, Zariel I; Schoepflin, Zachary R et al. (2014) FIH-1-Mint3 axis does not control HIF-1 transcriptional activity in nucleus pulposus cells. J Biol Chem 289:20594-605
Merceron, Christophe; Mangiavini, Laura; Robling, Alexander et al. (2014) Loss of HIF-1? in the notochord results in cell death and complete disappearance of the nucleus pulposus. PLoS One 9:e110768
Wang, Xin; Wang, Hua; Yang, Hao et al. (2014) Tumor necrosis factor-?- and interleukin-1?-dependent matrix metalloproteinase-3 expression in nucleus pulposus cells requires cooperative signaling via syndecan 4 and mitogen-activated protein kinase-NF-?B axis: implications in inflammatory disc disease. Am J Pathol 184:2560-72
Fujita, Nobuyuki; Hirose, Yuichiro; Tran, Cassie M et al. (2014) HIF-1-PHD2 axis controls expression of syndecan 4 in nucleus pulposus cells. FASEB J 28:2455-65
Shapiro, Irving M; Layfield, Robert; Lotz, Martin et al. (2014) Boning up on autophagy: the role of autophagy in skeletal biology. Autophagy 10:7-19
Risbud, Makarand V; Shapiro, Irving M (2014) Role of cytokines in intervertebral disc degeneration: pain and disc content. Nat Rev Rheumatol 10:44-56
Tran, Cassie M; Shapiro, Irving M; Risbud, Makarand V (2013) Molecular regulation of CCN2 in the intervertebral disc: lessons learned from other connective tissues. Matrix Biol 32:298-306
Wang, Jianru; Tian, Ye; Phillips, Kate L E et al. (2013) Tumor necrosis factor ?- and interleukin-1?-dependent induction of CCL3 expression by nucleus pulposus cells promotes macrophage migration through CCR1. Arthritis Rheum 65:832-42
Wang, Hua; Tian, Ye; Wang, Jianru et al. (2013) Inflammatory cytokines induce NOTCH signaling in nucleus pulposus cells: implications in intervertebral disc degeneration. J Biol Chem 288:16761-74

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