The development of a functional attachment between tendon and bone (the "enthesis") is critical for transmitting muscle forces to bone for joint motion. The formation of this attachment is driven by mechanical and biologic factors. Murine models of have demonstrated that removal of muscle load dramatically impairs the development of the enthesis. Furthermore, deletion of scleraxis (Scx), a transcription factor necessary for the development of force-transmitting tendons, leads to defects in enthesis formation. However, the mechanisms driving the mechanosensitivity of enthesis development are not known. Our global hypothesis is that Scx necessary for enthesis development and is driven by mechanical cues during.
The first aim of this study is to determine the temporal influence of Scx on neonatal enthesis maturation using an inducible knockout murine model. Expression of Scx will be limited to development up to embryonic day 15.5, post-natal day 1, or post-natal day 7 and developmental adaptations and maturation of the enthesis will be assessed throughout juvenile development. It is expected that deletion of Scx during embryonic development will have a greater effect on enthesis maturation compared to post-natal deletion, and molecular and functional outcomes will be correlated with duration of Scx expression during maturation. These findings will establish the importance of Scx in enthesis development as well as determine its temporal role in post- natal musculoskeletal development.
The second aim of this study is to determine the necessity of mechanical loading on Scx expression during development. Using in vivo and in vitro methods, we plan to mimic unloading and overloading, respectively, as well as assess the mechanosensitivity of Scx and its role in maturation and function of the enthesis. Using previously validated models of in vivo unloading and tissue-engineered methods for in vitro dynamic loading, we will track Scx-expressing cells as well as potential differentiation of ASCs into tenocytes. The effect of perturbed loading on the molecular and morphological response of Scx-expressing cells will be correlated with adaptations in structure and function using mechanical tests. These findings will determine the role of mechanical loading in Scx expression as well as the necessity of Scx in mechanically- induced tenogenesis. Lastly, the potency of Scx in load-induced tenogenesis following modulation with growth factors such as BMP-12 will be confirmed using BMP-12 siRNA. Findings from these studies will establish the importance of Scx on post-natal enthesis development, the mechanoresponsiveness of Scx in multi-tiered experimental models, and ultimately lead to new approaches for investigating musculoskeletal development and tissue engineered strategies for mechanosensitive orthopaedic tissues.

Public Health Relevance

Mechanical loading influences musculoskeletal development, maturation, and disease. Understanding the key molecular and mechanical cues associated with the development of tendon- to-bone attachments will lead to new rehabilitation approaches, gene therapies, and tissue engineering strategies to treat tendinopathies and re-attach tendon to bone.

Agency
National Institute of Health (NIH)
Institute
National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIAMS)
Type
Postdoctoral Individual National Research Service Award (F32)
Project #
5F32AR064652-02
Application #
8820068
Study Section
Special Emphasis Panel (ZRG1-F10B-S (20))
Program Officer
Tyree, Bernadette
Project Start
2013-05-28
Project End
2016-05-27
Budget Start
2014-05-28
Budget End
2015-05-27
Support Year
2
Fiscal Year
2014
Total Cost
$55,094
Indirect Cost
Name
Washington University
Department
Orthopedics
Type
Schools of Medicine
DUNS #
068552207
City
Saint Louis
State
MO
Country
United States
Zip Code
63130
Zelzer, Elazar; Blitz, Einat; Killian, Megan L et al. (2014) Tendon-to-bone attachment: from development to maturity. Birth Defects Res C Embryo Today 102:101-12