All ETDs from UAB

Advisory Committee Chair

Selvarangan Ponnazhagan

Advisory Committee Members

Gene P Siegal

Susan Bellis

Shawn R Gilbert

Yi-Ping Li

Document Type

Dissertation

Date of Award

2014

Degree Name by School

Doctor of Philosophy (PhD) Heersink School of Medicine

Abstract

Despite the body's ability to repair bone fractures under normal circumstances, up to 10% of the 7.9 million fractures suffered in the United States each year do not achieve bony union. Bone fractures heal with overlapping phases of inflammation, cell proliferation, and bone remodeling. Osteogenesis and angiogenesis are known to work in concert to control many stages of this process, but when one is impaired it leads to failure of bone healing, referred to as a nonunion. Such nonunion fractures often result from critical-size defects that will not completely heal over the natural lifetime of the animal. Based on this understanding, this dissertation research investigated the role of the immune system on the influence of angiogenesis and osteogenesis during bone fracture. We hypothesized that the immune system not only mediates the initial inflammatory response after fracture, but also influences neovasculature formation, crucial for osteogenesis. Results of this study demonstrated a population of hematopoietic progenitor cells, immature myeloid cells (IMC), significantly increased in number during the initial response to segmental defect, followed by a decrease in their number as the fracture healed. By specifically depleting the IMC population in vivo, we observed delayed healing of segmental defects; while adoptive transfer of IMC increased bone growth in a nonunion model, further signifying the role of IMC in fracture healing. We also demonstrated IMC have the ability to increase endothelial cell migration, invasion, and tube formation, thereby signaling the essential communication between the immune system and angiogenesis as a requirement for proper bone healing. In parallel, we developed a novel therapeutic osteoprogenitor cell-targeted gene therapy to increase angiogenesis and osteogenesis to prevent the formation of nonunion from critical-size defects. Towards this, we employed a directed evolution strategy of adeno-associated virus (AAV) to increase infectivity and transduction of mesenchymal stem cells (MSC), which are commonly used for skeletal regeneration. We developed an enhanced gene transfer system for increased expression of bone morphogenic protein (BMP-2) to enhance bone growth along with hypoxia inducible factor-1α (HIF-1α) to augment angiogenesis. Upon confirmation that our novel gene delivery vehicles out perform their wild-type counterparts and lead to increased transgene expression, we can apply this cell-targeted gene therapy in vivo. MSC transduced with recombinant AAV containing either BMP-2 or HIF-1α will be placed on an absorbable collagen sponge and implanted into the fracture site. Our findings suggest that overexpressing angiogenic and osteogenic genes can lead to an increase in bone growth and prevents the formation of nonunion fractures. The overall findings that IMC increase angiogenesis and that our novel gene and cell therapy has the potential to increase angiogenesis and osteogenesis to augment bone growth, provide great promise for utilizing these approaches to successfully treat nonunion bone defects. The basic understanding of fracture biology and translational approach to the problem of nonunions will lead to novel therapeutic targets in bone related pathologies.

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