Biology subjects

Burt, K. G.

Publications and source records attributed to Burt, K. G..

7 recordsLinked to original sources

bioRxiv2026

Mechano-activation of synovial fibroblasts and macrophages during OA progression in the dynamically stiffening synovial microenvironment

ObjectiveDuring osteoarthritis (OA) progression, the synovial membrane undergoes profound structural and compositional remodeling and fibrosis. We sought to elucidate how evolving synovial microenvironmental mechanics during fibrotic remodeling influence cell behavior and drive the progression of synovial pathology. MethodsSkeletally-mature male C57BL/6J mice were subjected to destabilization of the medial meniscus (DMM). To control for surgical confounders, both sham-operated and unoperated mice were included, with evaluation at 4- and 8-weeks. Synovial micromechanics were quantified via atomic force microscopy (AFM). Single-cell RNA sequencing (scRNA-seq), RNA fluorescence in situ hybridization (FISH), and flow cytometry were employed to investigate cellular heterogeneity, spatial organization, and crosstalk within fibrotic and non-fibrotic synovial niches. ResultsProgressive fibrotic remodeling and marked matrix stiffening were observed in DMM-operated synovium but absent in sham- and un-operated controls. While both sham and DMM joints mounted an acute stromal and immune response to surgery, these changes resolved over time in sham conditions but persisted in DMM synovium. During disease progression, distinct functional subsets of synovial fibroblasts and immune cells emerged, with mechanosignalling pathways and distinct immune cell-fibroblast crosstalk robustly activated within DMM-induced fibrotic microenvironments. ConclusionThis study demonstrates the complex cellular dynamics and crosstalk that differentiate the evolution of the pathological synovial response in the fibrotic DMM condition relative to surgical sham controls. Our findings highlight mechanotransduction as a central mechanism driving OA synovial pathogenesis and underscore the utility of the DMM model as a platform to dissect the molecular underpinnings of synovial fibrosis.

bioengineering↗

bioRxiv2026

CD14-deficiency protects against osteoarthritic subchondral bone sclerosis via enhanced osteoclastogenesis following joint injury

Aberrant bone remodeling is a hallmark of osteoarthritis, the most common arthritis affecting over 27 million US adults. Subchondral bone sclerosis, one sign of aberrant bone remodeling observable by routine x-rays, occurs as the trabeculae thicken, leading to increased bone volume. Toll-like receptors, pattern-recognition receptors of the innate immune system, have been implicated in OA pathogenesis, with TLR ligands, receptors, and co-receptors shown to mediate the severity and progression of OA. We have previously shown that CD14-deficiency protects mice against post-traumatic OA, and specifically reduces subchondral sclerosis post-injury. We hypothesized that depletion of CD14 protects against TLR4-dependent inhibition of osteoclastogenesis and therefore increases OC density in the SCB after injury, mitigating aberrant bone deposition in a murine model of OA. To determine how cellular changes correlate with bone structure derangements post-DMM, we performed MicroCT, Tartrate-resistant acid phosphatase staining, and alkaline phosphatase staining. To establish mechanistic changes in cellular signaling, we isolated WT and CD14-deficient osteoclast precursors and subjected them to LPS, an osteoarthritis-relevant TLR ligand, during differentiation. CD14-deficient mice, as well as WT mice treated with an anti-CD14 monoclonal antibody, show protection from post-injury increases in both bone volume fraction and bone mineral density. CD14-deficient mice had an increased osteoclast presence in the SCB two weeks post-injury, potentially protecting them from increases in bone volume and density. In vitro, CD14-deficient OCPs differentiated faster than WT OCPs, due to reduced Type I Interferon (IFN-I) signaling. In the presence of an LPS challenge, CD14-deficient OCPs were protected against LPS and TLR4-mediated inhibition, likely due to decreased MyD88-dependent TLR4 signaling. This work opens up new potential pathways to therapeutically target aberrant bone remodeling in the setting of joint injury and PTOA. Lay SummaryOsteoarthritis is one of the leading causes of disability worldwide. One of the hallmarks is subchondral sclerosis, or thickening of the bone in and around the joint. In this work, we used a mouse model of osteoarthritis to show that decreasing inflammatory signaling, through removal of CD14, protects against subchondral sclerosis, due to an increased presence of osteoclasts, cells that combat bone thickening. Osteoclasts without CD14 differentiate faster than osteoclasts with CD14, due to decreased Type I Interferon, an inflammatory cytokine. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=194 SRC="FIGDIR/small/705094v1_ufig1.gif" ALT="Figure 1"> View larger version (58K): org.highwire.dtl.DTLVardef@176bdd5org.highwire.dtl.DTLVardef@a914bborg.highwire.dtl.DTLVardef@902748org.highwire.dtl.DTLVardef@2f9b2_HPS_FORMAT_FIGEXP M_FIG C_FIG

immunology↗

bioRxiv2025

CD14 plays a critical role in pain and inflammation across multiple models of post-traumatic osteoarthritis

Inflammation is a primary driver of osteoarthritis (OA), and no therapies exist to halt or delay disease progression or substantially ameliorate the chronic pain, inflammation and disability that are characteristic of disease. Soluble CD14 (sCD14), a co-factor that enhances inflammatory toll-like receptor signaling, is present in synovial fluid in patients with OA and positively associates with joint space narrowing, synovial macrophage content, and pain. In this study, we show that increased sCD14 within human synovial fluid correlates with joint effusion volume and increased knee hyperalgesia. Next, we evaluated CD14 as a potential therapeutic target in three pre-clinical models of post traumatic OA (PTOA), using both genetic deficiency and pharmacologic blockade to modulate its activity. We demonstrate that deficiency or blockade of CD14 results in significant protection from increased evoked pain behaviors and from OA driven mobility impairments (i.e. decreased cage activity) across models that differ in severity, and across male and female cohorts. Using flow cytometry, single cell transcriptomics, and spatial proteomics, we further show that CD14-deficiency drastically influences the local synovial inflammatory landscape post-injury, reducing monocyte and macrophage populations and modulating local fibroblast populations. Targeting CD14 via genetic deficiency or therapeutic blockade revealed no substantial protection, but no worsening, of cartilage degeneration. Ultimately, our results provide strong support that targeting synovial inflammation through blockade of CD14 can safely ameliorate OA pain and disability after a pre-disposing injury. One Sentence SummaryThe study demonstrates the key role of CD14 in pain, mobility loss, and inflammation in PTOA, and demonstrates the therapeutic potential of CD14 blockade for OA pain relief.

immunology↗

bioRxiv2024

HMGB1 mediates macrophage recruitment and regional intervertebral disc tissue functional and mechanical property changes following injury.

ObjectiveFrequently evaluated in musculoskeletal disease, damage associated molecular patterns (DAMPs) respond to tissue damage and cellular stress by facilitating an inflammatory response via macrophage activation and broad inflammatory pathway activation. In the context of disc degeneration (DD), high mobility group box 1 (HMGB1), a potent intracellular DAMP, is seen to be increased within severely degenerated human IVDs and to directly mediate inflammatory responses within disc cells in vitro. To further understand how HMGB1 mediated inflammation influences DD, this study evaluated the possible protective effect of an HMGB1 knockout on DD pathology following injury. MethodsUsing a needle puncture injury model in murine caudal IVDs we evaluated DD pathology within an IVD specific Hmgb1 knockout (KO) model. Structural and compositional changes in IVD cellularity, histopathology, disc height, and biomechanics were evaluated in addition to an assessment of disc inflammation via gene expression and macrophage presence throughout the course of degeneration. ResultsHMGB1 expression robustly increased shortly following needle puncture injury and elevated levels were sustained up to 28-days post injury both in injured IVDs and in the IVDs adjacent to the level of injury. IVD specific Hmgb1 KO mice had an increased disc height following injury both at the injured and adjacent to injury level compared to injured WT IVDs. Hmgb1 KO also protected against tissue mechanical property losses at both the injured (dynamic modulus) and adjacent to injury level (dynamic modulus, creep, and equilibrium modulus) compared to injured WT IVDs, however there was no significant effect on histopathologic scores post injury. Hmgb1 KO resulted in alterations in macrophage (F4/80+) recruitment to the IVD post injury in vivo. A lower macrophage migration was also observed in vitro in response to the secretome of an injured Hmgb1 KO IVD compared to injured WT IVDs. Hmgb1 KO had no effect on inflammatory gene expression changes following injury within adjacent to injury level or injury level IVDs. ConclusionOverall findings indicate that HMGB1 is upregulated regionally, at both the injured level and at the level adjacent to injury. Results suggest that HMGB1 plays a role in mediating structural, biomechanical, and inflammatory responses to IVD injury and serves as a potent chemoattractant, mediating macrophage recruitment to the IVD and overall migratory function.

bioengineering↗

bioRxiv2023

Recommendations For a Standardized Approach to Histopathologic Evaluation of Synovial Membrane in Murine Models of Experimental Osteoarthritis

BackgroundSynovial pathology has been linked to osteoarthritis (OA) pain in patients. Microscopic grading systems for synovial changes in human OA have been described, but a standardized approach for murine models of OA is needed. We sought to develop a reproducible approach and set of minimum recommendations for synovial histopathology in mouse models of OA. MethodsCoronal and sagittal sections from male mouse knee joints subjected to destabilization of medial meniscus (DMM) or partial meniscectomy (PMX) were collected as part of other studies. Stains included Hematoxylin and Eosin (H&E), Toluidine Blue (T- Blue) and Safranin O/Fast Green (Saf-O). Four blinded readers graded pathological features (hyperplasia, cellularity, and fibrosis) at specific anatomic locations in the medial and lateral compartments. Inter-reader reliability of each feature was determined. ResultsThere was acceptable to very good agreement between raters. After DMM, increased hyperplasia and cellularity and a trend towards increased fibrosis were observed 6 weeks after DMM in the medial locations, and persisted up to 16 weeks. In the PMX model, cellularity and hyperplasia were evident in both medial and lateral compartments while fibrotic changes were largely seen on the medial side. Synovial changes were consistent from section to section in the mid-joint area mice. H&E, T-blue, and Saf-O stains resulted in comparable reliability. ConclusionsTo allow for a standard evaluation that can be implemented and compared across labs and studies, we recommend using 3 readers to evaluate a minimum set of 3 pathological features at standardized anatomic areas. Pre-defining areas to be scored, and reliability for each pathologic feature should be considered.

pathology↗

bioRxiv2023

Nuclear Factor Kappa B Over-Activation in the Intervertebral Disc Leads to Macrophage Recruitment and Severe Disc Degeneration

ObjectiveLow back pain (LBP) is the leading cause of global disability and is thought to be driven primarily by intervertebral disc (IVD) degeneration (DD). Persistent upregulation of catabolic enzymes and inflammatory mediators have been associated with severe cases of DD. Nuclear factor kappa B (NF-{kappa}B) is a master transcription regulator of immune responses and is over expressed during inflammatory-driven musculoskeletal diseases, including DD. However, its role in triggering DD is unknown. Therefore, this study investigated the effect of NF-{kappa}B pathway over-activation on IVD integrity and DD pathology. MethodsUsing skeletally mature mouse model, we genetically targeted IVD cells for canonical NF-{kappa}B pathway activation via expression of a constitutively active form of inhibitor of {kappa}B kinase B (IKK{beta}), and assessed changes in IVD cellularity, structural integrity including histology, disc height, and extracellular matrix (ECM) biochemistry, biomechanics, expression of inflammatory, catabolic, and neurotropic mediators, and changes in macrophage subsets, longitudinally up to 6-months post activation. ResultsProlonged NF-{kappa}B activation led to severe structural degeneration, with a loss of glycosaminoglycan (GAG) content and complete loss of nucleus pulposus (NP) cellularity. Structural and compositional changes decreased IVD height and compressive mechanical properties with prolonged NF-{kappa}B activation. These alterations were accompanied by increases in gene expression of inflammatory molecules (Il1b, Il6, Nos2), chemokines (Mcp1, Mif), catabolic enzymes (Mmp3, Mmp9, Adamts4), and neurotrophic factors (Bdnf, Ngf) within IVD tissue. Increased recruitment of activated F4/80+ macrophages exhibited a greater abundance of pro-inflammatory (CD38+) over inflammatory-resolving (CD206+) macrophage subsets in the IVD, with temporal changes in the relative abundance of macrophage subsets over time, providing evidence for temporal regulation of macrophage polarization in DD in vivo, where macrophages participate in resolving the inflammatory cascade but promote fibrotic transformation of the IVD matrix. We further show that NF-{kappa}B driven secretory factors from IVD cells increase macrophage migration and inflammatory activation, and that the secretome of inflammatory-resolving macrophages mitigates effects of NF-{kappa}B overactivation. ConclusionOverall the observed results suggest prolonged NF-{kappa}B activation can induce severe DD, acting through increases in inflammatory cytokines, chemotactic proteins, catabolic enzymes, and the recruitment and inflammatory activation of a macrophage cell populations, that can be mitigated with inflammatory-resolving macrophage secretome.

bioengineering↗

bioRxiv2023

The subacromial bursa is a key regulator of the rotator cuff and a new therapeutic target for improving repair

Rotator cuff injuries result in over 500,000 surgeries performed annually, an alarmingly high number of which fail. These procedures typically involve repair of the injured tendon and removal of the subacromial bursa. However, recent identification of a resident population of mesenchymal stem cells and inflammatory responsiveness of the bursa to tendinopathy indicate an unexplored biological role of the bursa in the context of rotator cuff disease. Therefore, we aimed to understand the clinical relevance of bursa-tendon crosstalk, characterize the biologic role of the bursa within the shoulder, and test the therapeutic potential for targeting the bursa. Proteomic profiling of patient bursa and tendon samples demonstrated that the bursa is activated by tendon injury. Using a rat to model rotator cuff injury and repair, tenotomy-activated bursa protected the intact tendon adjacent to the injured tendon and maintained the morphology of the underlying bone. The bursa also promoted an early inflammatory response in the injured tendon, initiating key players in wound healing. In vivo results were supported by targeted organ culture studies of the bursa. To examine the potential to therapeutically target the bursa, dexamethasone was delivered to the bursa, prompting a shift in cellular signaling towards resolution of inflammation in the healing tendon. In conclusion, contrary to current clinical practice, the bursa should be retained to the greatest extent possible and provides a new therapeutically target for improving tendon healing outcomes. One Sentence SummaryThe subacromial bursa is activated by rotator cuff injury and regulates the paracrine environment of the shoulder to maintain the properties of the underlying tendon and bone.

bioengineering↗