Hyaluronan (or hyaluronic acid, HA) is a ubiquitous molecule that plays critical roles in numerous physiological functions in vivo, including tissue hydration, inflammation, and joint lubrication. ...Both the abundance and size distribution of HA in biological fluids are recognized as robust indicators of various pathologies and disease progressions. However, such analyses remain challenging because conventional methods are not sufficiently sensitive, have limited dynamic range, and/or are only semi-quantitative. Here we demonstrate label-free detection and molecular weight discrimination of HA with a solid-state nanopore sensor. We first employ synthetic HA polymers to validate the measurement approach and then use the platform to determine the size distribution of as little as 10 ng of HA extracted directly from synovial fluid in an equine model of osteoarthritis. Our results establish a quantitative method for assessment of a significant molecular biomarker that bridges a gap in the current state of the art.
Glycopolymers, conjugates of synthetic polymers with pendant carbohydrates, are becoming increasingly important to probe the role of carbohydrates in cellular processes and for applications like ...biosensors and drug delivery. A library of glycopolymers bearing different sugar moieties was synthesized by grafting amino-functionalized sugars to poly(acrylic acid) via DMTMM coupling. Primary amines were introduced at the anomeric (C-1) position to a number of unprotected mono-, di-, and trisaccharides using ammonium carbamate and conjugated to poly(acrylic acid) of different molecular weights, synthesized by reversible addition–fragmentation chain transfer (RAFT) polymerization. This approach provides a simple and efficient route for the preparation of glycopolymers that differ only in the identity or degree of substitution of the sugar moiety on the polymer. The binding parameters (k a, k d, and K D) of these new glycopolymers to galectins-1 and -3 were quantified using surface plasmon resonance. The galectins selectively bound only to lactose-containing polymers, and the binding affinity was dependent on the galectin type, degree of sugar substitution and the molecular weight of polymer chains. Binding to both galectin-1 and -3 increased with a higher degree of sugar substitution, and higher molecular weight of the polymer backbone, reaching K D values on the order of 10–11 M.
Cells bend their plasma membranes into highly curved forms to interact with the local environment, but how shape generation is regulated is not fully resolved. Here, we report a synergy between ...shape-generating processes in the cell interior and the external organization and composition of the cell-surface glycocalyx. Mucin biopolymers and long-chain polysaccharides within the glycocalyx can generate entropic forces that favor or disfavor the projection of spherical and finger-like extensions from the cell surface. A polymer brush model of the glycocalyx successfully predicts the effects of polymer size and cell-surface density on membrane morphologies. Specific glycocalyx compositions can also induce plasma membrane instabilities to generate more exotic undulating and pearled membrane structures and drive secretion of extracellular vesicles. Together, our results suggest a fundamental role for the glycocalyx in regulating curved membrane features that serve in communication between cells and with the extracellular matrix.
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•Biopolymers in the glycocalyx can generate forces that bend the plasma membrane•Glycocalyx polymers regulate formation of membrane projections and microvesicles•Both intracellular dynamics and glycocalyx-imposed forces regulate membrane shapes•Polymer theories explain effects of polymer size/density on membrane morphologies
The extracellular glycocalyx can impose forces that bend the plasma membrane and regulate projections and vesicle formation.
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Mucins represent a largely untapped class of polymeric building block for biomaterials, therapeutics, and other biotechnology. Because the mucin polymer backbone is genetically ...encoded, sequence-specific mucins with defined physical and biochemical properties can be fabricated using recombinant technologies. The pendent O-glycans of mucins are increasingly implicated in immunomodulation, suppression of pathogen virulence, and other biochemical activities. Recent advances in engineered cell production systems are enabling the scalable synthesis of recombinant mucins with precisely tuned glycan side chains, offering exciting possibilities to tune the biological functionality of mucin-based products. New metabolic and chemoenzymatic strategies enable further tuning and functionalization of mucin O-glycans, opening new possibilities to expand the chemical diversity and functionality of mucin building blocks. In this review, we discuss these advances, and the opportunities for engineered mucins in biomedical applications ranging from in vitro models to therapeutics.
Inflammation of the synovium, known as synovitis, plays an important role in the pathogenesis of osteoarthritis (OA). Synovitis involves the release of a wide variety of pro‐inflammatory mediators in ...synovial fluid (SF) that damage the articular cartilage extracellular matrix and induce death and apoptosis in chondrocytes. The composition of synovial fluid is dramatically altered by inflammation in OA, with changes to both hyaluronic acid and lubricin, the primary lubricating molecules in SF. However, the relationship between key biochemical markers of joint inflammation and mechanical function of SF is not well understood. Here, we demonstrate the application of a novel analytical framework to measure the effective viscosity for SF lubrication of cartilage, which is distinct from conventional rheological viscosity. Notably, in a well‐established equine model of synovitis, this effective lubricating viscosity decreased by up to 10,000‐fold for synovitis SF compared to a ~4 fold change in conventional viscosity measurements. Further, the effective lubricating viscosity was strongly inversely correlated (r = −0.6 to −0.8) to multiple established biochemical markers of SF inflammation, including white blood cell count, prostaglandin E2 (PGE2), and chemokine ligand (CCLs) concentrations, while conventional measurements of viscosity were poorly correlated to these markers. These findings demonstrate the importance of experimental and analytical approaches to characterize functional lubricating properties of synovial fluid and their relationships to soluble biomarkers to better understand the progression of OA.
Background
Proximal sesamoid bone (PSB) fractures are the most common fatal musculoskeletal injury in North American racehorses. Computed tomography has the potential to detect morphological changes ...in bone structure but can be challenging to analyse reliably and quantitatively.
Objectives
To develop a radiomics platform that allows the comparison of features from micro‐CTs (µCT) of PSBs in horses that sustained catastrophic fractures with horses that did not. To compare features calculated with a radiomics approach with features calculated from a previously published study that used quantitative µCT in the same specimens.
Study design
Retrospective study of cadaver specimens of µCT images of PSBs using prospectively applied radiomics.
Methods
Radiomics features were computed on standardised CT datasets to benchmark the software. Features from µCT images of PSBs from eight horses that sustained PSB fracture and eight controls were computed using the contralateral, intact forelimb from horses sustaining PSB fracture (cases, n = 19) and all available forelimbs for controls (n = 30). Two‐hundred and fifteen radiomic features were calculated, and similar or comparable features were compared with those reported in a previous study that used the same specimens.
Results
Morphologic features computed with the radiomics approach, such as volume, minor axis dimensions and anisotropy were highly correlated with previously published data. A high number of imperceptible radiomic features, such as entropy, coarseness and histogram features were also found to be significantly different (P < .01). The extent of the differences in image features for the cases and controls PSBs depends on radiomic calculation settings.
Main limitations
Only datasets obtained from cadaver specimens were included in the study.
Conclusions
A radiomics approach for analysing µCT images of PSBs was able to identify and reproduce differences in image features in cases and controls. Furthermore, radiomics revealed many more imperceptible image features between cases and control PSBs.
Abstract
Background
Non‐septic tenosynovitis is a clinically relevant and often performance limiting musculoskeletal injury in the horse.
Objectives
To review the published literature to determine ...which tendon sheaths are commonly affected by non‐septic tenosynovitis and to describe the most frequently reported pathological lesions, outcomes, and surgical complications in equine non‐septic tenosynovitis.
Study design
Systematic review.
Methods
Literature searches were conducted in July 2021 from the online search engines PubMed, Scopus, Web of Science Core, VetMed Resource and ProQuest Theses & Dissertations. The inclusion criteria followed a participants, interventions, comparisons, outcome and study design (PICOS) approach. For inclusion, studies had to include live equids with non‐septic tenosynovitis of any tendon sheath. Studies were excluded if they only described non‐equine species, if they included data only on non‐tendon sheath structures, or if they included data exclusively on cases of septic or contaminated tendon sheaths. Determination of non‐sepsis relied on the diagnosis of the original authors; however, if non‐sepsis was not explicitly specified, then cases that had a history of contamination of the sheath, a wound near the sheath or a positive bacterial culture were excluded from analysis. Data analysed included the distribution of structures affected by non‐septic tenosynovitis, the most common pathological lesions identified within each sheath, and the most frequently reported surgical complications of non‐septic tenosynovitis. The quality of each study was assessed using a methodological quality analysis.
Results
A total of 85 studies describing non‐septic tenosynovitis in the horse were included. Across all 85 studies, there were a total of 2449 tendon sheaths in 2101 horses reported to be affected by non‐septic tenosynovitis. The digital flexor sheath was the most reported structure to be diagnosed with non‐septic tenosynovitis: 41/85 (48%) studies examined the digital flexor sheath exclusively, followed by the carpal flexor sheath, tarsal flexor sheath, carpal extensor sheaths, tarsal extensor sheaths, and one case of biceps brachii non‐septic tenosynovitis. For most tendon sheaths, the most common pathological lesion was an intrathecal tear of a soft tissue structure, including tears of the deep digital flexor tendon and tears of the manica flexoria. Bilateral disease was most common in the carpal flexor sheath, where distal radial physeal exostoses were the most common pathological lesions. Less common causes of non‐septic tenosynovitis included neoplasia, fracture of a bone adjacent to a tendon sheath, and mineralisation of an intrathecal tendon. The likelihood of return to previous level of athletic function following non‐septic tenosynovitis of most structures was approximately 50%, and the most common complication was persistent effusion following tenoscopy. While iatrogenic infection following surgery was uncommon, it was most likely following tenoscopy of the digital flexor sheath.
Conclusion
Non‐septic tenosynovitis is commonly reported in equine athletes, with intra‐thecal tears of the deep digital flexor tendon, superficial digital flexor tendon and manica flexoria frequently reported. Directions for future research include more thorough assessment of and reporting of complications following non‐septic tenosynovitis and correlation of characteristics of intrathecal pathological lesions with clinical outcomes.
TNF-α-stimulated gene 6 (TSG-6) protein, a TNF-α-responsive hyaladherin, possesses enzymatic activity that can catalyze covalent crosslinks of the polysaccharide hyaluronic acid (HA) to another ...protein to form heavy chain-hyaluronic acid (HC-HA) complexes in pathological conditions such as osteoarthritis (OA). Here, we examined HA synthase and inflammatory gene expression; synovial fluid HA, TNF-α, and viscosity; and TSG-6-mediated HC-HA complex formation in an equine OA model. The objectives of this study were to (1) evaluate the TNF-α-TSG-6-HC-HA signaling pathway across multiple joint tissues, including synovial membrane, cartilage, and synovial fluid, and (2) determine the impact of OA on synovial fluid composition and biophysical properties.
HA and inflammatory cytokine concentrations (TNF-α, IL-1β, CCL2, 3, 5, and 11) were analyzed in synovial fluid from 63 OA and 25 control joints, and HA synthase (HAS1-3), TSG-6, and hyaluronan-degrading enzyme (HYAL2, HEXA) gene expression was measured in synovial membrane and cartilage. HA molecular weight (MW) distributions were determined using agarose gel electrophoresis and solid-state nanopore measurements, and HC-HA complex formation was detected via immunoblotting and immunofluorescence. SEC-MALS was used to evaluate TSG-6-mediated HA crosslinking, and synovial fluid and HA solution viscosities were analyzed using multiple particle-tracking microrheology and microfluidic measurements, respectively.
TNF-α concentrations were greater in OA synovial fluid, and TSG6 expression was upregulated in OA synovial membrane and cartilage. TSG-6-mediated HC-HA complex formation was greater in OA synovial fluid and tissues than controls, and HC-HA was localized to both synovial membrane and superficial zone chondrocytes in OA joints. SEC-MALS demonstrated macromolecular aggregation of low MW HA in the presence of TSG-6 and inter-α-inhibitor with concurrent increases in viscosity.
Synovial fluid TNF-α concentrations, synovial membrane and cartilage TSG6 gene expression, and HC-HA complex formation were increased in equine OA. Despite the ability of TSG-6 to induce macromolecular aggregation of low MW HA with resultant increases in the viscosity of low MW HA solutions in vitro, HA concentration was the primary determinant of synovial fluid viscosity rather than HA MW or HC-HA crosslinking. The TNF-α-TSG-6-HC-HA pathway may represent a potential therapeutic target in OA.
Foals are susceptible to many of the same types of fractures as adult horses, often secondary to external sources of trauma. In addition, some types of fractures are specific to foals and occur ...routinely in horses under 1 year of age. These foal-specific fractures may be due to the unique musculoskeletal properties of the developing animal and may present with distinct clinical signs. Treatment plans and prognoses are tailored specifically to young animals. Common fractures not affecting the long bones in foals are discussed in this article, including osteochondral fragmentation, proximal sesamoid bone fractures/sesamoiditis, and distal phalanx fractures.
Proximal sesamoid bone (PSB) fractures are the most common musculoskeletal injury in race-horses. X-ray CT imaging can detect expressed radiological features in horses that experienced catastrophic ...fractures. Our objective was to assess whether expressed radiomic features in the PSBs of 50 horses can be used to develop machine learning models for predicting PSB fractures. The μCTs of intact contralateral PSBs from 50 horses, 30 of which suffered catastrophic fractures, and 20 controls were studied. From the 129 intact μCT images of PSBs, 102 radiomic features were computed using a variety of voxel resampling dimensions. Decision Trees and Wrapper methods were used to identify the 20 top expressed features, and six machine learning algorithms were developed to model the risk of fracture. The accuracy of all machine learning models ranged from 0.643 to 0.903 with an average of 0.754. On average, Support Vector Machine, Random Forest (RUS Boost), and Log-regression models had higher performance than K-means Nearest Neighbor, Neural Network, and Random Forest (Bagged Trees) models. Model accuracy peaked at 0.5 mm and decreased substantially when the resampling resolution was greater than or equal to 1 mm. We find that, for this in vitro dataset, it is possible to differentiate between unfractured PSBs from case and control horses using μCT images. It may be possible to extend these findings to the assessment of fracture risk in standing horses.
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