The Role of Hyaluronan/Receptor for Hyaluronan-Mediated Motility Interactions in the Modulation of Macrophage Polarization and Cartilage Repair.

Hyaluronan (HA), a negatively charged linear glycosaminoglycan, is a key macromolecular component of the articular cartilage extracellular matrix. The differential effects of HA are determined by a spatially/temporally regulated display of HA receptors, such as CD44 and receptor for hyaluronan-mediated motility (RHAMM). HA signaling through CD44 with RHAMM has been shown to stimulate inflammation and fibrotic processes. This study shows an increased expression of RHAMM in proinflammatory macrophages. Interfering with HA/RHAMM interactions using a 15-mer RHAMM-mimetic, HA-binding peptide, together with high-molecular-weight (HMW) HA reduced the expression and release of inflammatory markers and increased the expression of anti-inflammatory markers in proinflammatory macrophages. HA/RHAMM interactions were interfered in vivo during the regeneration of a full-thickness cartilage defect after microfracture surgery in rabbits using three intra-articular injections of 15-mer RHAMM-mimetic. HA-binding peptide together with HMWHA reduced the number of proinflammatory macrophages and increased the number of anti-inflammatory macrophages in the injured knee joint and greatly improved the repair of the cartilage defect compared with intra-articular injections of HMWHA alone. These findings suggest that HA/RHAMM interactions play a key role in cartilage repair/regeneration via stimulating inflammatory and fibrotic events, including increasing the ratio of proinflammatory/anti-inflammatory macrophages. Interfering with these interactions reduced inflammation and greatly improved cartilage repair.

A qualitative evaluation of an exercise practitioner in an outpatient child and adolescent mental health service.

BACKGROUND: Physical activity interventions can confer a range of physical and mental health benefits among young people with mental disorders. In some contexts, such as Ireland, integrated physical activity is not easily available within child and adolescent mental health services. Therefore, an interagency pilot intervention was established in a child and adolescent mental health service in Ireland with the integration of a novel exercise practitioner into the multidisciplinary mental health team. OBJECTIVE: A qualitative evaluation was conducted to understand the impact of the pilot intervention and to understand issues of implementation that arose throughout. METHODS: In-depth qualitative interviews with service users' parents/guardians (N = 3) and a single focus group with existing service providers (N = 3), framed by the RE-AIM framework were conducted to evaluate the pilot intervention. Data were analysed using thematic analysis to explore themes. RESULTS: Three overarching themes were identified. These were as follows: (i) Making changes toward healthier physical activity behaviours; (ii) An intervention of therapeutic holism; and (iii) The integrated service delivery. CONCLUSIONS: This research provides insight on the value of a novel integrated exercise practitioner in outpatient young persons' mental health services in Ireland, indicating an enhanced and complimentary therapeutic service. These findings will be helpful for integrating Exercise Practitioners in this setting going forward.

Noncovalent hyaluronan crosslinking by TSG-6: Modulation by heparin, heparan sulfate, and PRG4.

The size, conformation, and organization of the glycosaminoglycan hyaluronan (HA) affect its interactions with soluble and cell surface-bound proteins. HA that is induced to form stable networks has unique biological properties relative to unmodified soluble HA. AlphaLISA assay technology offers a facile and general experimental approach to assay protein-mediated networking of HA in solution. Connections formed between two end-biotinylated 50 kDa HA (bHA) chains can be detected by signal arising from streptavidin-coated donor and acceptor beads being brought into close proximity when the bHA chains are bridged by proteins. We observed that incubation of bHA with the protein TSG-6 (tumor necrosis factor alpha stimulated gene/protein 6, TNFAIP/TSG-6) leads to dimerization or higher order multimerization of HA chains in solution. We compared two different heparin (HP) samples and two heparan sulfate (HS) samples for the ability to disrupt HA crosslinking by TSG-6. Both HP samples had approximately three sulfates per disaccharide, and both were effective in inhibiting HA crosslinking by TSG-6. HS with a relatively high degree of sulfation (1.75 per disaccharide) also inhibited TSG-6 mediated HA networking, while HS with a lower degree of sulfation (0.75 per disaccharide) was less effective. We further identified Proteoglycan 4 (PRG4, lubricin) as a TSG-6 ligand, and found it to inhibit TSG-6-mediated HA crosslinking. The effects of HP, HS, and PRG4 on HA crosslinking by TSG-6 were shown to be due to HP/HS/PRG4 inhibition of HA binding to the Link domain of TSG-6. Using the AlphaLISA platform, we also tested other HA-binding proteins for ability to create HA networks. The G1 domain of versican (VG1) effectively networked bHA in solution but required a higher concentration than TSG-6. Cartilage link protein (HAPLN1) and the HA binding protein segment of aggrecan (HABP, G1-IGD-G2) showed only low and variable magnitude HA networking effects. This study unambiguously demonstrates HA crosslinking in solution by TSG-6 and VG1 proteins, and establishes PRG4, HP and highly sulfated HS as modulators of TSG-6 mediated HA crosslinking.

Selective isolation of hyaluronan by solid phase adsorption to silica.

A solid phase adsorption method for selective isolation of hyaluronan (HA) from biological samples is presented. Following enzymatic degradation of protein, HA can be separated from sulfated glycosaminoglycans, other unsulfated glycosaminoglycans, nucleic acids, and proteolytic fragments by adsorption to amorphous silica at specific salt concentrations. The adsorbed HA can be released from silica using neutral and basic aqueous solutions. HA ranging in size from ∼9 kDa to MDa polymers has been purified by this method from human serum and conditioned medium of cultured cells.

Densification: Hyaluronan Aggregation in Different Human Organs.

Hyaluronan (HA) has complex biological roles that have catalyzed clinical interest in several fields of medicine. In this narrative review, we provide an overview of HA aggregation, also called densification, in human organs. The literature suggests that HA aggregation can occur in the liver, eye, lung, kidney, blood vessel, muscle, fascia, skin, pancreatic cancer and malignant melanoma. In all these organs, aggregation of HA leads to an increase in extracellular matrix viscosity, causing stiffness and organ dysfunction. Fibrosis, in some of these organs, may also occur as a direct consequence of densification in the long term. Specific imaging evaluation, such dynamic ultrasonography, elasto-sonography, elasto-MRI and T1ρ MRI can permit early diagnosis to enable the clinician to organize the treatment plan and avoid further progression of the pathology and dysfunction.

Methods for isolating and analyzing physiological hyaluronan: a review.

The carbohydrate hyaluronan (or hyaluronic acid, HA) is found in all human tissues and biofluids where it has wide-ranging functions in health and disease that are dictated by both its abundance and size. Consequently, hyaluronan evaluation in physiological samples has significant translational potential. Although the analytical tools and techniques for probing other biomolecules such as proteins and nucleic acids have become standard approaches in biochemistry, those available for investigating hyaluronan are less well established. In this review, we survey methods related to the assessment of native hyaluronan in biological specimens, including protocols for separating it from biological matrices and technologies for determining its concentration and molecular weight.

MTADV 5-MER peptide suppresses chronic inflammations as well as autoimmune pathologies and unveils a new potential target-Serum Amyloid A.

Despite the existence of potent anti-inflammatory biological drugs e.g., anti-TNF and anti IL-6 receptor antibodies, for treating chronic inflammatory and autoimmune diseases, these are costly and not specific. Cheaper oral available drugs remain an unmet need. Expression of the acute phase protein Serum Amyloid A (SAA) is dependent on release of pro-inflammatory cytokines IL-1, IL-6 and TNF-α during inflammation. Conversely, SAA induces pro-inflammatory cytokine secretion, including Th17, leading to a pathogenic vicious cycle and chronic inflammation. 5- MER peptide (5-MP) MTADV (methionine-threonine-alanine-aspartic acid-valine), also called Amilo-5MER, was originally derived from a sequence of a pro-inflammatory CD44 variant isolated from synovial fluid of a Rheumatoid Arthritis (RA) patient. This human peptide displays an efficient anti-inflammatory effects to ameliorate pathology and clinical symptoms in mouse models of RA, Inflammatory Bowel Disease (IBD) and Multiple Sclerosis (MS). Bioinformatics and qRT-PCR revealed that 5-MP, administrated to encephalomyelytic mice, up-regulates genes contributing to chronic inflammation resistance. Mass spectrometry of proteins that were pulled down from an RA synovial cell extract with biotinylated 5-MP, showed that it binds SAA. 5-MP disrupted SAA assembly, which is correlated with its pro-inflammatory activity. The peptide MTADV (but not scrambled TMVAD) significantly inhibited the release of pro-inflammatory cytokines IL-6 and IL-1ß from SAA-activated human fibroblasts, THP-1 monocytes and peripheral blood mononuclear cells. 5-MP suppresses the pro-inflammatory IL-6 release from SAA-activated cells, but not from non-activated cells. 5-MP could not display therapeutic activity in rats, which are SAA deficient, but does inhibit inflammations in animal models of IBD and MS, both are SAA-dependent, as shown by others in SAA knockout mice. In conclusion, 5-MP suppresses chronic inflammation in animal models of RA, IBD and MS, which are SAA-dependent, but not in animal models, which are SAA-independent.

Protective Effects of a Hyaluronan-Binding Peptide (P15-1) on Mesenchymal Stem Cells in an Inflammatory Environment.

Mesenchymal stem cells (MSCs) obtained from various sources, including bone marrow, have been proposed as a therapeutic strategy for the improvement of tissue repair/regeneration, including the repair of cartilage defects or lesions. Often the highly inflammatory environment after injury or during diseases, however, greatly diminishes the therapeutic and reparative effectiveness of MSCs. Therefore, the identification of novel factors that can protect MSCs against an inflammatory environment may enhance the effectiveness of these cells in repairing tissues, such as articular cartilage. In this study, we investigated whether a peptide (P15-1) that binds to hyaluronan (HA), a major component of the extracellular matrix of cartilage, protects bone-marrow-derived MSCs (BMSCs) in an inflammatory environment. The results showed that P15-1 reduced the mRNA levels of catabolic and inflammatory markers in interleukin-1beta (IL-1ß)-treated human BMSCs. In addition, P15-1 enhanced the attachment of BMSCs to HA-coated tissue culture dishes and stimulated the chondrogenic differentiation of the multipotential murine C3H/10T1/2 MSC line in a micromass culture. In conclusion, our findings suggest that P15-1 may increase the capacity of BMSCs to repair cartilage via the protection of these cells in an inflammatory environment and the stimulation of their attachment to an HA-containing matrix and chondrogenic differentiation.

Exploring the Experiences of Physical Activity among Key Stakeholders in Rehabilitation and Recovery Mental Health Services.

Physical activity has therapeutic benefits for mental health service users. To date, there is limited evidence that has brought together the experiences of physical activity for service users and key multi-disciplinary service providers that support activity in outpatient settings, particularly in contexts where activity is not well integrated into policy and care structures. Previous research has relied on homogenous samples of either service users or service providers of a specific discipline, and key stakeholders like peer-support workers are under-represented. This research explored and thematically analysed multi-stakeholder (service users, n = 6; and service providers, n = 8) experiences of physical activity in outpatient mental health service in Ireland using phenomenologically influenced qualitative interviews. Two salient themes were identified; 'The challenges of being physically active in recovery' and 'Physical activity is a tool for recovery'. This research presents an account of the experiences of some of these poorly represented stakeholders such as carers, peer-support workers, doctors and nurse management, in addition to other well represented stakeholders.

A Hyaluronan-binding Peptide (P15-1) Reduces Inflammatory and Catabolic Events in IL-1ß-treated Human Articular Chondrocytes.

Inflammation plays a critical role in osteoarthritis (OA). It stimulates catabolic events in articular chondrocytes and prevents chondrogenic precursor cells from repairing cartilage lesions, leading to accelerated cartilage degradation. Therefore, the identification of novel factors that reduce catabolic events in chondrocytes and enhances chondrogenic differentiation of precursor cells in an inflammatory environment may provide novel therapeutic strategies for the treatment of OA. The goal of this study was to determine whether a hyaluronan (HA)-binding peptide (P15-1), via interacting with high molecular weight (HMW)HA can enhance the anti-inflammatory properties of HMWHA and decrease catabolic events in interleukin-1beta (IL-1ß)-treated human articular chondrocytes. Treatment with P15-1 decreased catabolic events and stimulated anabolic events in articular chondrocytes cultured in an inflammatory environment. P15-1 pre-mixed with HMWHA was more effective in inhibiting catabolic events and stimulating anabolic events than P15-1 or HMWHA alone. Our findings suggest that P15-1 together with HMWHA inhibits catabolic events in articular chondrocytes via the inhibition of p38 mitogen-activated protein kinases (MAPK) and increasing the thickness of the pericellular matrix (PCM) around chondrocytes thereby decreasing catabolic signaling. Finally, conditioned medium from IL-1ß and P15-1-treated human articular chondrocytes was less inhibitory for chondrogenic differentiation of precursor cells than conditioned medium from chondrocytes treated with IL-1ß alone. In conclusion, P15-1 is proposed to function synergistically with HMWHA to enhance the protective microenvironment for chondrocytes and mesenchymal stem cells during inflammation and regeneration.

Extracellular Vesicles Released From Articular Chondrocytes Play a Major Role in Cell-Cell Communication.

The purpose of this investigation was to determine the role of extracellular vesicles (EVs), released from articular chondrocytes in a physiological or pathological state, in cell-cell communication with other articular chondrocytes or chondrocyte precursor cells. The conditioned medium from interleukin-1ß (IL-1ß)-treated human articular chondrocytes stimulated catabolic events and inhibited type II collagen expression in articular chondrocytes to a much greater degree than medium from IL-1ß-treated chondrocytes after complete removal of EVs. The vehicle-treated and IL-1ß-treated human articular chondrocytes released EVs of similar size; however, the number of EVs released by IL-1ß-treated chondrocytes was markedly higher than the number of EVs released from the vehicle-treated cells. Furthermore, our findings demonstrate that similar to medium from IL-1ß-treated chondrocytes containing EVs, EVs isolated from medium of IL-1ß-treated chondrocytes stimulated catabolic events in articular chondrocytes, whereas EVs isolated from the medium of vehicle-treated chondrocytes inhibited catabolic events and increased messenger RNA levels of aggrecan and type II collagen in IL-1ß-treated chondrocytes. Furthermore, the medium containing EVs from vehicle-treated articular chondrocytes or EVs isolated from this medium stimulated chondrogenesis of C3H10T1/2 cells, whereas medium containing EVs from IL-1ß-treated chondrocytes or EVs isolated from this medium inhibited chondrogenesis. Our findings suggest that EVs released by articular chondrocytes play a key role in the communication between joint cells and ultimately in joint homeostasis, maintenance, pathology, and repair. © 2019 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 38:731-739, 2020.

Role of Hyaluronan in Inflammatory Effects on Human Articular Chondrocytes.

Hyaluronan (HA) fragments have been proposed to elicit defensive or pro-inflammatory responses in many cell types. For articular chondrocytes in an inflammatory environment, studies have failed to reach consensus on the endogenous production or effects of added HA fragments. The present study was undertaken to resolve this discrepancy. Cultured primary human articular chondrocytes were exposed to the inflammatory cytokine IL-1ß, and then tested for changes in HA content/size in conditioned medium, and for the expression of genes important in HA binding/signaling or metabolism, and in other catabolic/anabolic responses. Changes in gene expression caused by enzymatic degradation of endogenous HA, or addition of exogenous HA fragments, were examined. IL-1ß increased the mRNA levels for HA synthases HAS2/HAS3 and for the HA-binding proteins CD44 and TSG-6. mRNA levels for TLR4 and RHAMM were very low and were little affected by IL-1ß. mRNA levels for catabolic markers were increased, while type II collagen (α1(II)) and aggrecan were decreased. HA concentration in the conditioned medium was increased, but the HA was not degraded. Treatment with recombinant hyaluronidase or addition of low endotoxin HA fragments did not elicit pro-inflammatory responses. Our findings showed that HA fragments were not produced by IL-1ß-stimulated human articular chondrocytes in the absence of other sources of reactive oxygen or nitrogen species, and that exogenous HA fragments from oligosaccharides up to about 40 kDa in molecular mass were not pro-inflammatory agents for human articular chondrocytes, probably due to low expression of TLR4 and RHAMM in these cells.

Methods for Hyaluronan Molecular Mass Determination by Agarose Gel Electrophoresis.

The average molecular mass of hyaluronan (HA) in most healthy biological fluids and tissues is usually about 6000-8000 kDa, but the biosynthetic mechanism results in a polydisperse mixture of sizes. Subsequent enzymatic degradation, or the action of reactive oxygen and nitrogen species, can further increase polydispersity and decrease the average size. Fragmented HA can be a biomarker of inflammation. In addition, reductions in HA size are associated with tissue remodeling and repair processes. Some cell-surface receptor proteins have been reported to have HA-binding affinities that are size specific, and participate in activation of signaling cascades controlling multiple aspects of cell behavior. Here we describe simple agarose gel electrophoresis protocols for the determination of the molecular mass distribution of HA isolated from tissues and fluids.

Human pericardial proteoglycan 4 (lubricin): Implications for postcardiotomy intrathoracic adhesion formation.

OBJECTIVE: Intrapericardial fibrous adhesions increase the risk of sternal reentry. Proteoglycan 4/lubricin (PRG4) is a mucin-like glycoprotein that lubricates tissue compartments and prevents inflammation. We characterized PRG4 expression in human pericardium and examined its effects in vitro on human cardiac myofibroblast fibrotic activity and in vivo as a measure of its therapeutic potential to prevent adhesions. METHODS: Full-length PRG4 expression was determined using Western blot analysis and amplified luminescent proximity homogeneous assay in human pericardial tissues obtained at cardiotomy. The in vitro effects of PRG4 were investigated on human cardiac myofibroblasts for cell adhesion, collagen gel contraction, and cell-mediated extracellular matrix remodeling. The influence of PRG4 on pericardial homeostasis was determined in a chronic porcine animal model. RESULTS: PRG4 is expressed in human pericardial fluid and colocalized with pericardial mesothelial cells. Recombinant human PRG4 prevented human cardiac myofibroblast attachment and reduced myofibroblast activity assessed using collagen gel contraction assay (64.6% ± 8.1% vs 47.1% ± 6.8%; P = .02). Using a microgel assay, human cardiac myofibroblast mediated collagen fiber remodeling was attenuated by PRG4 (1.17 ± 0.03 vs 0.90 ± 0.05; P = .002). In vivo, removal of pericardial fluid alone induced severe intrapericardial adhesion formation, tissue thickening, and inflammatory fluid collections. Restoration of intrapericardial PRG4 was protective against fibrous adhesions and preserved the pericardial space. CONCLUSIONS: For the first time, we show that PRG4 is expressed in human pericardial fluid and regulates local fibrotic myofibroblast activity. Loss of PRG4-enriched pericardial fluid after cardiotomy might induce adhesion formation. Therapeutic restoration of intrapericardial PRG4 might prevent fibrous/inflammatory adhesions and reduce the risk of sternal reentry.

A competitive alphascreen assay for detection of hyaluronan.

A method for specific quantification of hyaluronan (HA) concentration using AlphaScreen® (Amplified Luminescent Proximity Homogeneous Assay) technology is described. Two types of hydrogel-coated and chromophore-loaded latex nanobeads are employed. The proximity of the beads in solution is detected by excitation of the donor bead leading to the production of singlet oxygen, and chemiluminescence from the acceptor bead upon exposure to singlet oxygen. In the HA assay, the donor bead is modified with streptavidin, and binds biotin-labeled HA. The acceptor bead is modified with Ni(II), and is used to bind a specific recombinant HA-binding protein (such as HABP; aggrecan G1-IGD-G2) with a His-tag. Competitive inhibition of the HA-HABP interaction by free unlabeled HA in solution is used for quantification. The assay is specific for HA, and not dependent on HA molecular mass above the decasaccharide. HA can be quantified over a concentration range of approximately 30-1600 ng/mL using 2.5 µL of sample, for a detectable mass range of approximately 0.08-4 ng HA. This sensitivity of the AlphaScreen assay is greater than existing ELISA-like methods, due to the small volume requirements. HA can be detected in biological fluids using the AlphaScreen assay, after removal of bound proteins from HA and dilution or removal of other interfering proteins and lipids.

Hyaluronan Isolation from Mouse Mammary Gland.

The glycosaminoglycan hyaluronan (HA) is a key component of the extracellular matrix. The molecular weight of HA is heterogeneous and can reach from several million to several hundred daltons. The effect of HA on cell behavior is size dependent; fragmented HA acts as a danger signal, stimulates cell migration and proliferation and is proinflammatory, native high molecular weight HA suppresses inflammation. Therefore, it is important to analyze HA size distribution when studying the role of HA in tissue homeostasis and pathology. This protocol describes isolation of HA from mouse mammary glands but can also be applied to other tissues. The quality of the isolated HA is sufficient to analyze size distribution by gel electrophoresis ( Calabro et al., 2000 ).

Mouse Mammary Gland Whole Mount Preparation and Analysis.

The mammary gland undergoes extensive remodeling during pregnancy and is also subject to neoplastic processes both of which result in histological changes of the gland epithelial structure. Since the mammary tree is a complex three-dimensional structure a method is needed that provides an overview of the entire gland. Whole mounts provide this information, are inexpensive and do not require specialized equipment. This protocol describes mammary gland isolation, whole mount preparation and analysis. Mammary gland tissue, which is removed postmortem, is stained with Carmine Alum, a nuclear stain, allowing detection of epithelial structures embedded in the adipose tissue of the mammary fat pad. Stained mammary glands are imaged by light microscopy or embedded and sectioned for histological examination. Image analysis software such as Image J can be used to quantify extensity of branching complexity, epithelial structure remodeling or hyperplastic changes.

Hyaluronan and Hyaluronan Fragments.

The glycosaminoglycan hyaluronan (HA) is a key component of the microenvironment surrounding cells. In healthy tissues, HA molecules have extremely high molecular mass and consequently large hydrodynamic volumes. Tethered to the cell surface by clustered receptor proteins, HA molecules crowd each other, as well as other macromolecular species. This leads to severe nonideality in physical properties of the biomatrix, because steric exclusion leads to an increase in effective concentration of the macromolecules. The excluded volume depends on both polymer concentration and hydrodynamic volume/molecular mass. The biomechanical properties of the extracellular matrix, tissue hydration, receptor clustering, and receptor-ligand interactions are strongly affected by the presence of HA and by its molecular mass. In inflammation, reactive oxygen and nitrogen species fragment the HA chains. Depending on the rate of chain degradation relative to the rates of new synthesis and removal of damaged chains, short fragments of the HA molecules can be present at significant levels. Not only are the physical properties of the extracellular matrix affected, but the HA fragments decluster their primary receptors and act as endogenous danger signals. Bioanalytical methods to isolate and quantify HA fragments have been developed to determine profiles of HA content and size in healthy and diseased biological fluids and tissues. These methods have potential use in medical diagnostic tests. Therapeutic agents that modulate signaling by HA fragments show promise in wound healing and tissue repair without fibrosis.

Hyaluronan modulates growth factor induced mammary gland branching in a size dependent manner.

Mammary gland morphogenesis begins during fetal development but expansion of the mammary tree occurs postnatally in response to hormones, growth factors and extracellular matrix. Hyaluronan (HA) is an extracellular matrix polysaccharide that has been shown to modulate growth factor-induced branching in culture. Neither the physiological relevance of HA to mammary gland morphogenesis nor the role that HA receptors play in these responses are currently well understood. We show that HA synthase (HAS2) is expressed in both ductal epithelia and stromal cells but HA primarily accumulates in the stroma. HA accumulation and expression of the HA receptors CD44 and RHAMM are highest during gestation when gland remodeling, lateral branch infilling and lobulo-alveoli formation is active. Molecular weight analyses show that approximately 98% of HA at all stages of morphogenesis is >300kDa. Low levels of 7-114kDa HA fragments are also detected and in particular the accumulation of 7-21kDa HA fragments are significantly higher during gestation than other morphogenetic stages (p<0.05). Using these in vivo results as a guide, in culture analyses of mammary epithelial cell lines (EpH4 and NMuMG) were performed to determine the roles of high molecular weight, 7-21kDa (10kDa MWavg) and HA receptors in EGF-induced branching morphogenesis. Results of these assays show that while HA synthesis is required for branching and 10kDa HA fragments strongly stimulate branching, the activity of HA decreases with increasing molecular weight and 500kDa HA strongly inhibits this morphogenetic process. The response to 10kDa HA requires RHAMM function and genetic deletion of RHAMM transiently blunts lateral branching in vivo. Collectively, these results reveal distinct roles for HA polymer size in modulating growth factor induced mammary gland branching and implicates these polymers in both the expansion and sculpting of the mammary tree during gestation.

Viscoelastic Properties of Hyaluronan in Physiological Conditions.

Hyaluronan (HA) is a high molecular weight glycosaminoglycan of the extracellular matrix (ECM), which is particularly abundant in soft connective tissues. Solutions of HA can be highly viscous with non-Newtonian flow properties. These properties affect the movement of HA-containing fluid layers within and underlying the deep fascia. Changes in the concentration, molecular weight, or even covalent modification of HA in inflammatory conditions, as well as changes in binding interactions with other macromolecules, can have dramatic effects on the sliding movement of fascia. The high molecular weight and the semi-flexible chain of HA are key factors leading to the high viscosity of dilute solutions, and real HA solutions show additional nonideality and greatly increased viscosity due to mutual macromolecular crowding. The shear rate dependence of the viscosity, and the viscoelasticity of HA solutions, depend on the relaxation time of the molecule, which in turn depends on the HA concentration and molecular weight. Temperature can also have an effect on these properties. High viscosity can additionally affect the lubricating function of HA solutions. Immobility can increase the concentration of HA, increase the viscosity, and reduce lubrication and gliding of the layers of connective tissue and muscle. Over time, these changes can alter both muscle structure and function. Inflammation can further increase the viscosity of HA-containing fluids if the HA is modified via covalent attachment of heavy chains derived from Inter-α-Inhibitor. Hyaluronidase hydrolyzes HA, thus reducing its molecular weight, lowering the viscosity of the extracellular matrix fluid and making outflow easier. It can also disrupt any aggregates or gel-like structures that result from HA being modified. Hyaluronidase is used medically primarily as a dispersion agent, but may also be useful in conditions where altered viscosity of the fascia is desired, such as in the treatment of muscle stiffness.