Author ORCID Identifier
Date Available
1-1-2024
Year of Publication
2024
Degree Name
Doctor of Philosophy (PhD)
Document Type
Doctoral Dissertation
College
Engineering
Department/School/Program
Biomedical Engineering
First Advisor
Ashley W. Seifert
Second Advisor
Sheng Tong
Abstract
Chronic inflammation is a major cause of the pathogenesis of musculoskeletal diseases such as fragility, fracture, and nonunion. Studies have shown that modulating the immune phenotype of macrophages from proinflammatory to pro-healing can heal recalcitrant bone defects. Current therapeutic strategies predominantly apply biochemical cues, which often lack target specificity, and controlling their release kinetics in vivo is challenging spatially and temporally. We have developed a magnetic iron-oxide nanocomplexes (MNC)-based therapy for resolving chronic inflammation in the context of promoting fracture healing. Here, we show that MNC internalized macrophages, when coupled with an external magnetic field, can exert an intracellular magnetic force on the cytoskeleton to promote a pro-healing phenotype switch by altering cell shape and actin dynamics. The intracellular magnetic force perturbs actin polymerization, which significantly reduces nuclear to cytoplasm redistribution of MRTF-A and HDAC3, major drivers of inflammatory and osteogenic gene expression, respectively. The reduction in nuclear MRTF-A and HDAC3 levels due to intracellular magnetic force subsequently downregulates Nos2 gene expression, confirmed by quantitative PCR analysis. These findings may facilitate efforts to develop MNC-based resolution-centric therapeutic intervention to direct macrophage phenotype and function towards healing to supplement or replace the currently used anti-inflammatory therapies for fracture healing.
Diabetes is a chronic metabolic disorder that can lead to diabetic myopathy and bone diseases. The etiology of musculoskeletal complications in such metabolic disorders and the interplay between the muscular and osseous systems are not well understood. Exercise training promises to prevent diabetic myopathy and bone disease and offers protection. Although the muscle-bone interaction is largely biomechanical, the muscle secretome has significant implications for bone biology. Uncoupling effects of biophysical and biochemical stimuli on the adaptive response of bone during exercise training may offer therapeutic targets for diabetic bone disease. Here, we have developed an in vitro model to elucidate the effects of mechanical strain on myokine secretion and its impact on bone metabolism decoupled from physical stimuli. We developed bone constructs using cross-linked gelatin, which facilitated osteogenic differentiation of osteoprogenitor cells. Then, muscle constructs were made from fibrin, which enabled myoblast differentiation and myotube formation. We investigated the myokine expression by muscle constructs under strain regimens replicating endurance (END) and high-intensity interval training (HIIT) in hyperglycemic conditions. In monocultures, both regimens induced higher expression of Il15 and Igf1, whereas END supported more myoblast differentiation and myotube maturation than HIIT. When co-cultured with bone constructs, HIIT regimen increased Glut4 expression in muscle constructs more than END, supporting higher glucose uptake. Likewise, the muscle constructs under the HIIT regimen promoted a healthier and more matured bone phenotype than END. Under static conditions, myostatin (Mstn) expression was significantly downregulated in muscle constructs co-cultured with bone constructs compared with monocultures. Together, our in vitro co-culture system allowed orthogonal manipulation of mechanical strain on muscle constructs while facilitating bone-muscle biochemical crosstalk. Such systems can provide an individualized microenvironment that allows decoupled biomechanical manipulation, help identify molecular targets, and develop engineered therapies for metabolic bone disease.
Digital Object Identifier (DOI)
https://doi.org/10.13023/etd.2024.01
Funding Information
- National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIAMS) Award Number R21AR078447
- Small grant award by the UK center for Clinical and Translational Science (CCTS)
- Igniting Research Collaborations grant (IRC)
- Orthopedic Trauma Association (OTA; grant number 6889)
Recommended Citation
Suresh Kumar, Harshini, "DEVELOPING AN IMMUNOMODULATORY STRATEGY USING BIOPHYSICAL CUES TO MODULATE MACROPHAGE PHENOTYPE FOR FRACTURE HEALING AND BONE REGENERATION" (2024). Theses and Dissertations--Biomedical Engineering. 81.
https://uknowledge.uky.edu/cbme_etds/81
