Research Summary
Research topic: Compared vascularity and healing of fractures at the protein-receptor level in fracture models at different ages to find potential therapeutics that block inhibitors of angiogenesis
Research results: Found that altering the CD47 receptor on the thrombospondin protein (TSP)-CD47 pathway has the potential to improve fracture healing
Patient care application of results: Potential therapeutics that block inhibitors of angiogenesis, allowing for the growth of new blood vessels and the vascularization required for better fracture healing in older patients
Simplified patient care application: Therapeutics that improve bone healing in older patients
Modulating Vascularity
OREF grant recipient compares healing in young and old bones
Jay D. Lenn
Bone is peculiar in its healing capacity after injury as the regenerated tissue is microscopically composed of normal rather than scar tissue. The efficiency of this regenerative process changes, however, as we age. While the mechanism of bone growth is the same in older adults, the healing is delayed. Older adults also are more likely to experience complications from fractures, and delays in healing translate into delays in functional and rehabilitative therapy.
"The burden of fragility fractures is tremendous," stated Jaimo Ahn, MD, PhD,
Harold W. and Helen L. Gehring Research Professor of Orthopaedic Surgery and
associate chair for Education, Orthopaedic Surgery, chief and service co-director, Orthopaedic Trauma at the University of Michigan Health. "Initially my interest was in pediatric orthopaedics, and then I transitioned to trauma. What has stuck with me is the amazing burden of disease in older adults and the amazing power of healing in the young."
This early impression is the foundation of Dr. Ahn's investigations into why bone regeneration is delayed in older adults and, in particular, how age-related changes in vascularity regulation affect healing. OREF funded part of his work with a 2016 Career Development Grant.1
Inhibiting the inhibitors
A critical factor in the regenerative process is vascularization. The growth of new blood vessels at the fracture site is essential for supplying oxygen and nutrients, delivering the precursor cells for cartilage and bone development, and removing debris. The exploration of potential therapies to enhance vascularization is still relatively new, and no therapies are currently in clinical use.
A number of factors regulate the development of new vessels. Some promote angiogenesis, and some inhibit it. One pathway of inhibition is the interaction of extracellular thrombospondin proteins (TSP) with the CD47 receptor. In the OREF-funded study, Dr. Ahn investigated the role of TSP1-CD47 and TSP2-CD47 interactions on vascularization and bone healing in a geriatric mouse model and explored the viability of any of these factors as a therapeutic target.
Dr. Ahn would like to find a therapeutic that blocks inhibitors of angiogenesis. He noted, "The benefit of therapeutically blocking an inhibitor is that it disrupts a naturally occurring negative signal and enables other endogenous signaling proteins to do their work. It's more difficult to boost a positive signaling protein because it's likely regulated and blocked by endogenous inhibitors."
Models of fracture healing
Dr. Ahn investigated the role of the TSP-CD47 signaling pathway in mouse models with a population of 5-month-old mice, the equivalent of human young adulthood, and 25-month-old mice, the approximate equivalent of 80-year-old humans. The two groups enabled Dr. Ahn and his colleagues to compare the vascularity and healing of fractures in young and old subjects.
Mice received a surgically created fracture that was set, and the mice were treated for pain. In order to track the progression of healing over time, mice were euthanized and the bones harvested at different times: presurgery, day of surgery, and 5, 10, 20 and 40 days after surgery.
Various subsets of mice enabled the researchers to focus independently on the role of each of the three factors: TSP1, TSP2, and CD47. In each subset, gene expression for one of the proteins was underexpressed, knocked out, or overexpressed. Dr. Ahn explained, "The significance for studying them independently is to improve our mechanistic knowledge of the signaling pathways and to set the direction for further investigations. Also, although CD47 appears to be the primary receptor, there is a possibility that TSPs could be regulating vascularity through a different receptor."
The researchers employed micro-computerized tomography scans to measure physical properties at the healing site, such as mineral density and bone volume. A mechanical testing apparatus was used to test strength and rigidity of the bone. And additional laboratory tests characterized such elements as cell proliferation, cartilage and bone formation, vascularization, and the presence of precursor cells and growth factors.
Dr. Ahn and his research team’s initial results showed that altering one of the TSPs did not seem to enhance fracture healing in their geriatric model but altering the CD47 receptor might have beneficial effects. “We will continue to investigate the cellular and molecular underpinnings of the TSP-CD47 pathway as it relates to bone healing and regeneration through collaborative grants with the ultimate goal of improving patient care,” Dr. Ahn said.
Bridging the funding grant
This early work provided critical data for further research funded by a National Institutes of Health R01 grant to study the utility of modulating the TSP-CD47 pathway to improve vascularity and bone healing. In collaboration with a multi-PI group coordinated by Kurt Hankenson, DVM, MS, PhD, the team including Dr. Ahn have recently made progress on this work, finding that CD47 is important for mesenchymal stem cell proliferation and fracture repair.
Dr. Ahn noted that the Career Development Grant was particularly critical for his ability to conduct a multi-year study essential to explore bone healing in both young and geriatric animal models.
More generally, he observed that OREF Career Development Grants provide a bridge in the current funding model for developing clinician scientists. He stated, "For many projects there is a gap in the middle where you need multiple years of funding. You don't need it to the scale of a multimillion dollar grant, but you also need more than a one-year early investigator award."
Dr. Ahn commended OREF for supporting clinician scientists through the spectrum of their education, training, and career development. "Really understanding the science, whether it's clinical science or basic science, and making changes to the way we practice based on that science is going to transform the lives of our patients in the future."
Jay D. Lenn is a contributing writer for OREF. He can be reached at communications@oref.org
- Dr. Ahn's 2016 OREF Career Development Grant was underwritten by the Dr. Dane and Mrs. Mary Louise Miller Endowment Fund and the Dr. Zachary B. and Mrs. Kathleen Friedenberg Endowment Fund. At the time of his award, Dr. Ahn was assistant professor of orthopaedic surgery at the University of Pennsylvania.