Hyaluronan in morphogenesis

Hyaluronan is a very large polysaccharide that is found in extracellular matrices, at the cell surface and inside cells. This review focuses on the functions of hyaluronan directly associated with the cell surface, where it is commonly present as the essential core of a highly hydrated pericellular matrix that contains several other components (hyaladherins) bound to hyaluronan. Three major molecular characteristics of hyaluronan contribute to its physiological functions: its unique hydrodynamic properties, its interactions with structural extracellular hyaladherins, and its instructive effects on cell signaling and behavior. Recent studies of hyaluronan-deficient mouse embryos illustrate the importance of each of these classes of function of hyaluronan. It is postulated that the morphogenetic effects of hyaluronan are due to its ability to act as a template for assembly of a multi-component, pericellular matrix as well as to its physical properties. This matrix would provide a hydrated environment in which cells are separated from structural barriers to morphogenetic changes and receive signals from hyaluronan itself and from associated factors.     

Hyaluronan in limb morphogenesis

Hyaluronan (HA) is a large glycosaminoglycan that is not only a structural component of extracellular matrices, but also interacts with cell surface receptors to promote cell proliferation, migration, and intracellular signaling. HA is a major component of the extracellular matrix of the distal subapical mesenchymal cells of the developing limb bud that are undergoing proliferation, directed migration, and patterning in response to the apical ectodermal ridge (AER), and has the functional potential to be involved in these processes. Here we show that the HA synthase Has2 is abundantly expressed by the distal subridge mesodermal cells of the chick limb bud and also by the AER itself. Has2 expression and HA production are downregulated in the proximal central core of the limb bud during the formation of the precartilage condensations of the skeletal elements, suggesting that downregulation of HA may be necessary for the close juxtaposition of cells and the resulting cell-cell interactions that trigger cartilage differentiation during condensation. Overexpression of Has2 in the mesoderm of the chick limb bud in vivo results in the formation of shortened and severely malformed limbs that lack one or more skeletal elements. Skeletal elements that do form in limbs overexpressing Has2 are reduced in length, exhibit abnormal morphology, and are positioned inappropriately. We also demonstrate that sustained HA production in micromass cultures of limb mesenchymal cells inhibits formation of precartilage condensations and subsequent chondrogenesis, indicating that downregulation of HA is indeed necessary for formation of the precartilage condensations that trigger cartilage differentiation. Taken together these results suggest involvement of HA in various aspects of limb morphogenesis.     

Hyaluronan.

Hyaluronan (hyaluronic acid) is a high-molecular-mass polysaccharide found in the extracellular matrix, especially of soft connective tissues. It is synthesized in the plasma membrane of fibroblasts and other cells by addition of sugars to the reducing end of the polymer, whereas the nonreducing end protrudes into the pericellular space. The polysaccharide is catabolized locally or carried by lymph to lymph nodes or the general circulation, from where it is cleared by the endothelial cells of the liver sinusoids. The overall turnover rate is surprisingly rapid for a connective tissue matrix component (t1/2 0.5 to a few days). Hyaluronan has been assigned various physiological functions in the intercellular matrix, e.g., in water and plasma protein homeostasis. Hyaluronan production increases in proliferating cells and the polymer may play a role in mitosis. Extensive hyaluronidase-sensitive coats have been identified around mesenchymal cells. They are either anchored firmly in the plasma membrane or bound via hyaluronan-specific binding proteins (receptors). Such receptors have now been identified on many different cells, e.g., the lymphocyte homing receptor CD 44. Interaction between a hyaluronan receptor and extracellular polysaccharide has been connected with locomotion and cell migration. Hyaluronan seems to play an important role during development and differentiation and has other cell regulatory activities. Hyaluronan has also been recognized in clinical medicine. A concentrated solution of hyaluronan (10 mg/ml) has, through its tissue protective and rheological properties, become a device in ophthalmic surgery. Analysis of serum hyaluronan is promising in the diagnosis of liver disease and various inflammatory conditions, e.g., rheumatoid arthritis. Interstitial edema caused by accumulation of hyaluronan may cause dysfunction in various organs.     

Hyaluronan: Biosynthesis and signaling

Background

Hyaluronan is a critical component of extracellular matrix with several different roles. Besides the contribution to the tissue hydration, mechanical properties and correct architecture, hyaluronan plays important biological functions interacting with different molecules and receptors.

Scope of review

The review addresses the control of hyaluronan synthesis highlighting the critical role of hyaluronan synthase 2 in this context as well as discussing the recent findings related to covalent modifications which influence the enzyme activity. Moreover, the interactions with specific receptors and hyaluronan are described focusing on the importance of polymer size in the modulation of hyaluronan signaling.

Major conclusions

Due to its biological effects on cells recently described, it is evident how hyaluronan is to be considered not only a passive component of extracellular matrix but also an actor involved in several scenarios of cell behavior.

General significance

The effects of metabolism on the control of hyaluronan synthesis both in healthy and pathologic conditions are critical and still not completely understood. The hyaluronan capacity to bind several receptors triggering specific pathways may represent a valid target for new approach in several therapeutic strategies. This article is part of a Special Issue entitled Matrix-mediated cell behaviour and properties.     

Hyaluronan chain conformation and dynamics

An overview of the present state of research in the field of hyaluronan chain conformational aspects is presented. The relationship between structure and dynamics are illustrated for a series of hyaluronan oligomers. Conformational characteristics of hyaluronan chains are discussed, together with the dynamic chain patterns, evaluated by using a theoretical approach to diffusive polymer dynamics. The dependence of correlation times and NMR relaxation parameters from the chain dimension are investigated. Topological features and dimensional properties are related to the structural determinants by using classical computational methods of molecular mechanics and Monte Carlo simulation.     

Hyaluronan in human malignancies

Hyaluronan, a major macropolysaccharide in the extracellular matrix of connective tissues, is intimately involved in the biology of cancer. Hyaluronan accumulates into the stroma of various human tumors and modulates intracellular signaling pathways, cell proliferation, motility and invasive properties of malignant cells. Experimental and clinicopathological evidence highlights the importance of hyaluronan in tumor growth and metastasis. A high stromal hyaluronan content is associated with poorly differentiated tumors and aggressive clinical behavior in human adenocarcinomas. Instead, the squamous cell carcinomas and malignant melanomas tend to have a reduced hyaluronan content. In addition to the stroma–cancer cell interaction, hyaluronan can influence stromal cell recruitment, tumor angiogenesis and epithelial–mesenchymal transition. Hyaluronan receptors, hyaluronan synthases and hyaluronan degrading enzymes, hyaluronidases, are involved in the modulation of cancer progression, depending on the tumor type. Furthermore, intracellular signaling and angiogenesis are affected by the degradation products of hyaluronan. Hyaluronan has also therapeutic implications since it is involved in multidrug resistance.     

Effects of Ascorbic Acid and Analogs on the Activity of Testicular Hyaluronidase and Hyaluronan Lyase on Hyaluronan

We have evaluated the inhibition of testicular hyaluronidase and hyaluronan lyase by L-ascorbic acid and chemical analogs. We observed that L-ascorbic acid, D-isoascorbic acid and dehydroascorbic acid inhibited both types of enzymes, but showed stronger effects towards hyaluronan lyase. But these compounds were observed to degrade the substrate, hyaluronan, by themselves. Of the other ascorbic acid analogs tested, saccharic acid inhibited hyaluronan lyase, while not affecting the enzymatic activity of testicular hyaluronidase, nor affecting the physic-chemical stability of hyaluronan. This is the first compound, to our knowledge, to be shown to possess such selective inhibition. Therefore, we propose that saccharic acid could serve as a lead compound for the development of potent and selective inhibitors of bacterial hyaluronan lyase or of polysaccharide lyase enzymes in general as we observed this compound to be capable of inhibiting chondroitinase ABC in addition to hyaluronan lyase.     

Hyaluronan, a common thread

Hyaluronan, natures simplest, but still exceptionally versatile glycosaminoglycan, is currently the focus of attention across a wide front of research; from cell biology, morphogenesis, matrix organization, pathobiology to tissue engineering. This macromolecule has entangled me in a number of puzzling and challenging projects over the past 3 decades. These entertaining encounters are outlined in this retrospective.     

Hyaluronan in human and rat muscle spindles

 Previous data on the composition of the periaxial fluid of muscle spindles have relied on indirect histochemical methods. We used a biotinylated hyaluronan-binding protein as a specific probe for the detection of hyaluronan in sections of human and rat limb muscles. Hyaluronan was identified in the axial and periaxial space of the muscle spindles as well as in the endoneurium and in the space in between individual axons. Hyaluronan was also present in the innermost layer of the spindle capsule in the A region and in all layers of the capsule in the B region. Accepted: 12 February 1998     

Hyaluronan promotes the malignant phenotype

Hyaluronan is a high-molecular-weight, negatively charged polysaccharide with unusual physical and interactive properties. Hyaluronan is localized in the extracellular matrix, at the cell surface, and inside cells. Its tissue distribution is ubiquitous, but it is particularly concentrated in pericellular matrices surrounding proliferating and migrating cells. Hyaluronan contributes to cell behavior in at least three ways. Its unique physical properties influence the biomechanical properties of extracellular and pericellular matrices; it is a template for assembly of other pericellular macromolecules; and it interacts directly with cell surface receptors that transduce intracellular signals. Experimental studies in animal models have documented a crucial role for hyaluronan in tumor growth and metastasis. Cellular manipulations have shown that hyaluronan promotes anchorage-independent growth and invasiveness, hallmarks of the malignant phenotype.     

Hyaluronan matrices in pathobiological processes

Hyaluronan matrices are ubiquitous in normal and pathological biological processes. This remarkable diversity is related to their unique mechanism of synthesis by hyaluronan synthases. These enzymes are normally activated in the plasma membrane and utilize cytosolic substrates directly to form these large polyanionic glycosaminoglycans, which are extruded directly into the extracellular space. The extracellular matrices that are formed interact with cell surface receptors, notably CD44, that often dictate the biological processes, as described in the accompanying minireviews of this series. This article focuses on the discovery in recent studies that many cell stress responses initiate the synthesis of a monocyte-adhesive hyaluronan extracellular matrix, which forms a central focus for subsequent inflammatory processes that are modulated by the dialogue between the matrix and the inflammatory cells. The mechanisms involve active hyaluronan synthases at the cell membrane when cell stresses occur at physiological levels of glucose. However, dividing cells at hyperglycemic levels of glucose initiate the synthesis of hyaluronan in intracellular compartments, which induces endoplasmic reticulum stress and autophagy, processes that probably contribute greatly to diabetic pathologies.     

Rab10-mediated Endocytosis of the Hyaluronan Synthase HAS3 Regulates Hyaluronan Synthesis and Cell Adhesion to Collagen

Hyaluronan synthases (HAS1–3) are unique in that they are active only when located in the plasma membrane, where they extrude the growing hyaluronan (HA) directly into cell surface and extracellular space. Therefore, traffic of HAS to/from the plasma membrane is crucial for the synthesis of HA. In this study, we have identified Rab10 GTPase as the first protein known to be involved in the control of this traffic. Rab10 colocalized with HAS3 in intracellular vesicular structures and was co-immunoprecipitated with HAS3 from isolated endosomal vesicles. Rab10 silencing increased the plasma membrane residence of HAS3, resulting in a significant increase of HA secretion and an enlarged cell surface HA coat, whereas Rab10 overexpression suppressed HA synthesis. Rab10 silencing blocked the retrograde traffic of HAS3 from the plasma membrane to early endosomes. The cell surface HA coat impaired cell adhesion to type I collagen, as indicated by recovery of adhesion following hyaluronidase treatment. The data indicate a novel function for Rab10 in reducing cell surface HAS3, suppressing HA synthesis, and facilitating cell adhesion to type I collagen. These are processes important in tissue injury, inflammation, and malignant growth.