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Principal Investigator: Alexander John Knights
Organization: UNIVERSITY OF MICHIGAN AT ANN ARBOR
Fiscal Year: 2024
Award: $103,080
Funding agency: National Institute of Arthritis and Musculoskeletal and Skin Diseases
Post-traumatic osteoarthritis (PTOA) is a degenerative joint disease that arises after injury and affects millions
worldwide. There are currently no disease-modifying treatments. PTOA is a complex, multi-tissue joint disease
characterized by pain, cartilage degradation, synovial inflammation and fibrosis, and formation of ectopic bone
growths called osteophytes. The inherent complexity of this disease is a barrier to developing effective
treatments, as little is known about the intricate tissue crosstalk that underlies PTOA progression. Our long-
term goal is to uncover and comprehensively characterize cellular and molecular mechanisms central to key
pathological sequalae of PTOA: synovial fibrosis, inflammation, and osteophyte formation. We will focus on
canonical Wnt/β-catenin (cWnt) signaling. cWnt overactivation has recently been implicated as a driving factor
of arthritis. Our data show that the cWnt signaling agonist R-spondin 2 (Rspo2) is strongly induced in multiple
joint tissues during PTOA, and that Rspo2 alone is sufficient to induce pathological features characteristic of
PTOA. Using single-cell RNA-seq, we profiled synovium of mice with PTOA and found that Rspo2 is produced
by synovial lining fibroblasts. We identified a novel population of pro-fibrotic cells that arise after injury and
express Lgr cell surface receptors for Rspo2. We showed that synovial fibroblasts respond to Rspo2 by
secreting cytokines that in turn activate pro-inflammatory macrophages (known to drive synovial pathology in
PTOA). Single-cell profiling also revealed a novel subset of injury-induced, Lgr-expressing osteochondral
progenitors in synovium, which we propose give rise to osteophytes. We hypothesize that Rspo2-driven cWnt
signaling mediates pathological crosstalk between joint-resident cell types to potentiate PTOA. To test this, our
aims in the K99 phase are to: 1) determine the role of Rspo2-driven cWnt signaling in the emergence and
function of pro-fibrotic synovial cells during PTOA using transgenic reporter mice, multi-omic analyses, and in
vitro differentiation assays, and 2) characterize crosstalk between cWnt-active synovial fibroblasts and pro-
inflammatory macrophages, using knockout mice and crosstalk assays. To extend upon my molecular biology
and immunology expertise, I will receive rigorous technical and conceptual training from my diverse mentorship
committee during the K99 phase, and valuable career guidance. This expert training in bioinformatics; cWnt
signaling; bone, cartilage, and synovial biology; and multi-modal imaging, will be crucial for carrying out my
K99 aims and especially critical for successfully launching my independent career. These skills will be utilized
in my R00 phase to: 3) determine how Rspo2/Lgr signaling promotes osteophyte formation in PTOA, using
tissue-specific deletion and reporter mice, and in vitro differentiation assays. This work will significantly extend
our understanding of cellular and molecular mechanisms that underpin synovial fibrosis, inflammation, and
osteophyte formation in PTOA. These insights will have meaningful, tangible outcomes for human health, by
accelerating development of effective disease-modifying treatments for PTOA sufferers.
Terms: <ATAC sequencing><ATAC-seq><ATACseq><Acceleration><Actins><Affect><Agonist><Arthritis><Assay><Assay for Transposase-Accessible Chromatin using sequencing><Automobile Driving><B cell differentiation factor><B cell stimulating factor 2><B-Cell Differentiation Factor><B-Cell Differentiation Factor-2><B-Cell Stimulatory Factor-2><BCDF><BSF-2><BSF2><Beta Cadherin-Associated Protein><Beta-1 Catenin><Binding><Bio-Informatics><Bioassay><Bioinformatics><Biological Assay><Biology><Body Tissues><Bone Formation><Bone Growth><Bone Spur><CUL-2><Career Counseling><Career Guidance><Cartilage><Cartilaginous Tissue><Cell Body><Cell Communication and Signaling><Cell Nucleus><Cell Signaling><Cell Surface Receptors><Cells><Characteristics><Chondrocytes><Chronic><Clinical Research><Clinical Study><Complex><Coupled><DNA Molecular Biology><Data><Degenerative Arthritis><Degenerative polyarthritis><Development><Differentiation in cell culture><Disease><Disorder><Fibroblasts><Fibrosis><G Protein-Complex Receptor><G Protein-Coupled Receptor Genes><G-Protein-Coupled Receptors><GPCR><Goals><HPGF><Health><Hepatocyte-Stimulating Factor><Human><Hybridoma Growth Factor><Hypertrophy><IFN-beta 2><IFNB2><IL-6><IL6 Protein><Immune><Immunes><In Vitro><In vitro cell differentiation><Inflammation><Inflammatory><Injections><Injury><Interleukin-6><Intracellular Communication and Signaling><Involuntary Muscle><Joint Diseases><Joints><KO mice><Kinetics><Knee><Knock-out Mice><Knockout Mice><Knowledge acquisition><Label><Leucine-Rich Repeat><Link><Long term disability><MGI-2><Macrophage><Macrophage Activation><Mediating><Membrana Synovialis Capsulae Articularis><Mentorship><Mice><Mice Mammals><Modern Man><Molecular><Molecular Biology><Molecular Immunology><Molecular Interaction><Multimodal Imaging><Murine><Mus><Musculoskeletal><Myeloid Differentiation-Inducing Protein><Myofibroblast><Mφ><Nucleus><Null Mouse><Occupational Guidance><Osteoarthritis><Osteoarthrosis><Osteoblasts><Osteogenesis><Outcome><PRO2286><Pain><Painful><Pathogenesis><Pathogenicity><Pathologic><Pathology><Patient outcome><Patient-Centered Outcomes><Patient-Focused Outcomes><Phase><Plasmacytoma Growth Factor><Population><Position><Positioning Attribute><Process><Proteins><Proteoglycan><RNA Seq><RNA sequencing><RNAseq><Receptor Protein><Reporter><Reporter Genes><Research><Right-Handed Beta-Alpha Superhelix><Role><Science><Secondary to><Severities><Signal Pathway><Signal Transduction><Signal Transduction Systems><Signaling><Single-Nucleus Sequencing><Smooth Muscle><Synovia><Synovial Cell><Synovial Fluid><Synovial Membrane><Synovitis><Synovium><Technical Expertise><Testing><Tissues><Training><Transgenic Organisms><Traumatic Arthritis><Traumatic Arthropathy><Vocational Counseling><Vocational Guidance><Work><arthritic><arthropathic><arthropathies><arthropathy><articular cartilage><assay for transposase accessible chromatin followed by sequencing><assay for transposase accessible chromatin seq><assay for transposase accessible chromatin sequencing><assay for transposase-accessible chromatin with sequencing><beta catenin><biological signal transduction><bone><bone tissue formation><career><career counselor><cartilage degeneration><cartilage degradation><cell type><cytokine><degenerative joint disease><developmental><differentiation in culture><differentiation in vitro><driving><effective therapy><effective treatment><executive coaching><hypertrophic arthritis><improved><in vitro cellular differentiation><induced Cre><inducible Cre><inflamed synovial tissue><inflamed synovium><injured><injuries><insight><interferon beta 2><joint damage><joint disorder><joint function><joint injury><joint trauma><mineralization><multi-modal imaging><multi-modality imaging><multimodality imaging><multiomics><multiple omics><neutralizing antibody><novel><osteoarthritic><osteochondral><osteochondral tissue><osteogenic><osteophyte><pain sensitivity><panomics><patient oriented outcomes><pharmacologic><post-traumatic osteoarthritis><pre-clinical study><preclinical study><progenitor><receptor><sNuc-Seq><scRNA-seq><single cell RNA-seq><single cell RNAseq><single cell expression profiling><single cell transcriptomic profiling><single nucleus RNA-sequencing><single nucleus seq><single-cell RNA sequencing><single-nucleus RNA-seq><skill acquisition><skill development><skills><snRNA sequencing><snRNA-seq><social role><synovial inflammation><technical skills><transcriptome sequencing><transcriptomic sequencing><transgenic><β-catenin>