Of the greater than 6 million fractures occurring yearly in the US, up to 20% will result in nonunion or delayed union, thereby requiring intervention for bone regeneration. Mesenchymal stem/stromal cells (MSCs) are an attractive cell source for cell-based therapies of bone healing because of their osteogenic potential and robust secretion of proangiogenic trophic factors. Culture dimensionality has a profound impact on a myriad of cell functions. Compared to dissociated MSCs, our recent data demonstrate that MSC spheroids secrete 100- fold higher levels of angiogenic factors and better resist apoptosis while maintaining osteogenic potential. Spheroid formation is a competition between cohesion and adhesion, and optimizing this balance through the entrapment in engineered biomaterials provides an exciting opportunity to instruct the regenerative potential of MSCs after transplantation. Hydrogel properties such as adhesivity, stiffness, and degradation influence the function of entrapped cells and resulting tissue formation. Alginate is a highly cytocompatible natural polymer that is amenable to control of initial mechanical properties through composition and crosslinking, as well as adhesivity by covalently coupling peptide sequences such as Arg-Gly-Asp (RGD) to the polymer backbone that bind cellular receptors. Thus, alginate hydrogels represent an ideal tool to probe the role of substrate properties on spheroid function. Our central hypothesis is that the therapeutic potential of MSC spheroids for bone regeneration can be enhanced using alginate hydrogels with engineered biophysical properties.
Aim 1. Does adhesion ligand density within alginate hydrogels affect the survival, proangiogenic, and osteogenic potential of entrapped MSC spheroids? We will synthesize alginate hydrogels with varying densities of RGD. The influence of increased adhesion versus cohesion on spheroid function will be determined.
Aim 2. Do hydrogel biomechanical properties influence the functional response of entrapped MSC spheroids? Using composite hydrogels with distinct biophysical properties, we will examine the role of substrate stiffness and degradation on survival, proangiogenic and osteogenic potential of entrapped MSC spheroids.
Aim 3. Can MSC spheroids transplanted in RGD-modified hydrogels with optimized biophysical properties accelerate bone formation in a critical-sized calvarial bone defect? We will characterize the capacity of MSC spheroids transplanted in RGD-modified alginate hydrogels to accelerate bone repair in an orthotopic defect compared to dissociated MSCs. The role of implanted cells, as well as quality of bone formation will be assessed using noninvasive imaging modalities. The proposed research is innovative because it exploits the balance of cellular aggregation versus adhesion to drive cell fate using an injectable, biodegradable hydrogel to potentiate the reparative potential of MSCs. This research will provide a new approach to drive bone formation in nonhealing or slow healing bone fractures, and the strategies have potential in enhancing the efficacy of materials-based therapies for tissue repair.

Public Health Relevance

The development of new approaches to potentiate the activity of transplanted cells will provide valuable and necessary options to clinicians treating slow-healing or nonhealing bone defects. We seek to determine if controlling the geometry of transplanted cells using an injectable matrix will enhance the reparative function of progenitor cells.

Agency
National Institute of Health (NIH)
Institute
National Institute of Dental & Craniofacial Research (NIDCR)
Type
Research Project (R01)
Project #
5R01DE025475-05
Application #
9728725
Study Section
Musculoskeletal Tissue Engineering Study Section (MTE)
Program Officer
Wan, Jason
Project Start
2015-07-01
Project End
2020-06-30
Budget Start
2019-07-01
Budget End
2020-06-30
Support Year
5
Fiscal Year
2019
Total Cost
Indirect Cost
Name
University of California Davis
Department
Biomedical Engineering
Type
Biomed Engr/Col Engr/Engr Sta
DUNS #
047120084
City
Davis
State
CA
Country
United States
Zip Code
95618
Vorwald, Charlotte E; Murphy, Kaitlin C; Leach, J Kent (2018) Restoring vasculogenic potential of endothelial cells from diabetic patients through spheroid formation. Cell Mol Bioeng 11:267-278
Ho, Steve S; Hung, Ben P; Heyrani, Nasser et al. (2018) Hypoxic Preconditioning of Mesenchymal Stem Cells with Subsequent Spheroid Formation Accelerates Repair of Segmental Bone Defects. Stem Cells 36:1393-1403
Leach, J Kent; Whitehead, Jacklyn (2018) Materials-Directed Differentiation of Mesenchymal Stem Cells for Tissue Engineering and Regeneration. ACS Biomater Sci Eng 4:1115-1127
Gionet-Gonzales, Marissa A; Leach, J Kent (2018) Engineering principles for guiding spheroid function in the regeneration of bone, cartilage, and skin. Biomed Mater 13:034109
Vorwald, Charlotte E; Ho, Steve S; Whitehead, Jacklyn et al. (2018) High-Throughput Formation of Mesenchymal Stem Cell Spheroids and Entrapment in Alginate Hydrogels. Methods Mol Biol 1758:139-149
Mitra, Debika; Whitehead, Jacklyn; Yasui, Osamu W et al. (2017) Bioreactor culture duration of engineered constructs influences bone formation by mesenchymal stem cells. Biomaterials 146:29-39
Murphy, Kaitlin C; Whitehead, Jacklyn; Falahee, Patrick C et al. (2017) Multifactorial Experimental Design to Optimize the Anti-Inflammatory and Proangiogenic Potential of Mesenchymal Stem Cell Spheroids. Stem Cells 35:1493-1504
Murphy, Kaitlin C; Hung, Ben P; Browne-Bourne, Stephen et al. (2017) Measurement of oxygen tension within mesenchymal stem cell spheroids. J R Soc Interface 14:
Ho, Steve S; Keown, Andrew T; Addison, Bennett et al. (2017) Cell Migration and Bone Formation from Mesenchymal Stem Cell Spheroids in Alginate Hydrogels Are Regulated by Adhesive Ligand Density. Biomacromolecules 18:4331-4340
Murphy, Kaitlin C; Whitehead, Jacklyn; Zhou, Dejie et al. (2017) Engineering fibrin hydrogels to promote the wound healing potential of mesenchymal stem cell spheroids. Acta Biomater 64:176-186

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