Notch Signaling and Bone Fracture Healing

Bone fractures can exhibit delayed or non-union healing. Current treatments have well- documented limitations. Although morphological aspects of fracture healing are well- characterized, molecular mechanisms that regulate the complex progression of healing are poorly understood. Therefore, a need persists for the identification of novel pathways that regulate fracture healing, and for development of therapeutics targeting these pathways to enhance regeneration. Notch signaling regulates bone development, and many aspects of bone development are recapitulated during repair. Notch signaling is also required for repair of other tissues, and enhancing Notch signaling promotes regeneration. Therefore, the objective of this thesis was to determine the role of Notch signaling during bone fracture healing, and to create a translatable therapy targeting the pathway to enhance bone tissue formation. We hypothesized that (i) Notch signaling components are active during bone repair; (ii) inhibition of Notch signaling alters healing; (iii) expression of the Jagged1 ligand in mesenchymal cells regulates bone formation; and (iv) therapeutic delivery of Jagged1 will activate the Notch signaling pathway and promote osteogenesis. We first characterized activation of Notch signaling during tibial fracture and calvarial defect healing, and demonstrated that Notch signaling components are active during both methods of repair with Jagged1 the most highly upregulated ligand. Then we determined the importance of Notch signaling by using a temporally controlled inducible model (Mx1- Cre;dnMAMLf/-) to impair canonical signaling in all cells during tibial fracture and calvarial defect healing, and demonstrated that Notch inhibition alters the temporal progression of events required for healing, including inflammation, cartilage formation, callus vascularization and bone remodeling. Next we deleted Jagged1 in mesenchymal progenitors (Prx1-Cre;Jagged1f/f) or committed osteoblasts (Col2.3-Cre;Jagged1f/f), and determined that Jagged1 promotes bone formation during development. Finally, we developed a biomaterial construct comprised of Jagged1 and a poly(β-amino ester) scaffold, and demonstrated that it activates Notch signaling and enhances osteoblast differentiation. This thesis identified Notch signaling as an important regulator of fracture healing, developed a translatable therapeutic targeting the pathway to improve bone tissue formation. The study design outlined can also serve as a model for the discovery of novel pathways that regulate, and therefore could enhance, bone fracture healing.