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Hyaluronic acid (Hyaluronan): A review

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Abstract

Hyaluronic acid (HA) is a high molecular weight biopolysacharide, discovered in 1934, by Karl Meyer and his assistant, John Palmer in the vitreous of bovine eyes. Hyaluronic acid is a naturally occurring biopolymer, which has important biological functions in bacteria and higher animals including humans. It is found in most connective tissues and is particularly concentrated in synovial fluid, the vitreous fluid of the eye, umbilical cords and chicken combs. It is naturally synthesized by a class of integral membrane protein s called hyaluronan synthases, and degraded by a family of enzymes called hyaluronidases. This review describes metabolisms, different physi - ological and pathological functions, basic pharmacological properties, and the clinical use of hyaluronic acid.

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... For SC groups, A inj (0) and PH20(0) were set equal to dose of cetuximab and rHuPH20, respectively. HA(0) was set to a literature value (k out was estimated, and k in was a secondary parameter), and the rest of the initial conditions were fixed to zero (34). ...
... To facilitate model convergence, the baseline concentration of HA in rat skin was fixed to 840 μg/g according to published value (34). Some model-estimated parameters were compared with experimental data from previous studies. ...
... The model combined three major mechanistic components: kinetics of rHuPH20 at SC site; hyaluronan (HA) homeostasis and its disruption by rHuPH20; and cetuximab systemic disposition and the effect of HA disruption on cetuximab SC absorption (Fig. 1). Furthermore, the estimates obtained from the mechanistic model were able to also capture previously published data (28,34,35) (Figs. 5 and 6). ...
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Purpose To evaluate the duration of effect of rHuPH20 on SC absorption of cetuximab and to develop a mechanistic pharmacokinetic model linking the kinetics of rHuPH20 action with hyaluronan (HA) homeostasis and absorption of cetuximab from the SC space. Methods Serum pharmacokinetics of cetuximab was evaluated after IV and SC dosing at 0.4 and 10 mg/kg (control groups). In test groups, SC cetuximab was administered simultaneously with rHuPH20 (Co-Injection) or 12 h after injection of rHuPH20 (Pre-Injection). Mechanistic pharmacokinetic model was developed to simultaneously capture cetuximab kinetics in all groups. Results Administration of rHuPH20 resulted in a faster absorption of cetuximab; the difference between co-injection and pre-injection groups appeared to be dependent on the dose level. The model combined three major components: kinetics of rHuPH20 at SC site; HA homeostasis and its disruption by rHuPH20; and cetuximab systemic disposition and the effect of HA disruption on cetuximab SC absorption. The model provided good description of experimental data obtained in this study and collected previously. Conclusions Proposed model can serve as a potential translational framework for capturing the effect of rHuPH20 across multiple preclinical species and in human studies and can be used for optimization of SC delivery of biotherapeutics.
... Exogenous hyaluronic acid increases hyaluronic acid within cells as well as increases glycoprotein synthesis and decreases the formation and function of inflammatory mediators, matrix metalloproteinases, and behavioral changes in immune cells by preventing the formation of immune complex and polymorphonuclear cells, inhibiting the migration of leukocytes and macrophages, and regulating fibroblast proliferation. Many of the activities of hyaluronic acid depend on its molecular weight [13]. HA can be absorbed in the low molecular weight form and can interact with many cells (involved in the proliferation process as well as with immune cells) such as dendritic cells, macrophages, and osteoclasts [14]. ...
... The reticulated HA might show a superior regenerative ability in comparison with linear HA [35]. Besides, molecular weight and particle size as well considerably influence the HA function [13,36]. HAs with high molecular weights might increase the mRNA expressions of RUNX-2, ALP, and OCN [37], inducing bone formation [30]. ...
Article
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Introduction: There is no study on the effectiveness of hyaluronic acid (HA) placement either with or without absorbable collagen sponge (ACS) in reducing or preventing bisphosphonate-related osteonecrosis of the jaws (BRONJ). This preliminary animal study examined the efficacy of this clinically important treatment. Methods: For simulating BRONJ, zoledronic acid was administered to 40 rats for 5 weeks. Two weeks later, a right first molar was extracted from each rat. The rats were randomized into four groups of socket treatments: control (empty extraction socket) or with sockets filled with ACS, HA, or HA+ACS (n=4×10). After 2 weeks, 5 rats in each group were sacrificed and subjected to histopathologic and histomorphometric evaluation. Eight weeks post-surgically, the rest of rats were euthanized and histologically examined. The Kruskal-Wallis test was used to compare the four treatments at each time point (α=0.05). Results: Six rats were lost overall. In the second week, vascularization was higher in ACS group (P<0.05); osteoclast activity was not different between groups (P>0.05); empty lacunae were the most and fewest in control and HA+ACS groups, respectively (P<0.05); eosinophil infiltration was maximum in HA group (P<0.05); lymphocyte counts were maximum and minimum in the HA+ACS and ACS groups, respectively (P<0.05); the highest and lowest neutrophil counts were seen in ACS and control groups, respectively (P<0.05); and the extent of live bone did not differ between groups (P>0.05). In the eighth week, vascularization was not different in groups (P>0.05); the highest and lowest osteoclast activities were seen in the control and HA+ACS groups, respectively (P<0.05); empty lacunae were the most and fewest in control and HA+ACS, respectively (P<0.05); maximum and minimum numbers of eosinophils were in control and HA+ACS groups, respectively (P<0.05); HA and control groups exhibited the highest and lowest lymphocyte counts, respectively (P<0.05); the lowest and highest neutrophil counts were observed in HA+ACs and control groups, respectively (P<0.05); and the highest and lowest extents of the live bone were observed in HA+ACS and control groups, respectively (P<0.05). Conclusions: Within the limitations of this preliminary animal study, HA and especially HA+ACS seem a proper method for preventing or treating BRONJ.
... As depicted in Figure 1, when both D-glucuronic acid and N-acetyl-D-glucosamine are in the β conformation, the functional moieties in the equatorial positions are sterically stabilised, whereas the hydrogen atoms on each monosaccharide unit occupy the more energetically and sterically unfavourable axial positions, forming the energetically stable conformation of HA [53][54][55]. Each disaccharide unit is inverted respective to the preceding disaccharide unit in the polysaccharide chain, allowing HA to adapt a coiled, yet semirigid, ribbon conformation in solution [56][57][58]. ...
... The biological activity of HA is highly dependent on MW. HA is endogenously produced within the inner plasma membrane via the enzymatic action of three isoenzymes, HA synthases 1, 2 and 3 [56,89]. The MW of the chains produced is controlled by the differential expression of the synthases in response to varying intracellular conditions. ...
Article
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Extensive research is currently being conducted into novel ocular drug delivery systems (ODDS) that are capable of surpassing the limitations associated with conventional intraocular anterior and posterior segment treatments. Nanoformulations, including those synthesised from the natural, hydrophilic glycosaminoglycan, hyaluronic acid (HA), have gained significant traction due to their enhanced intraocular permeation, longer retention times, high physiological stability, and inherent biocompatibility, and biodegradability. However, conventional nanoformulation preparation methods often require large volumes of organic solvent, chemical cross-linkers, and surfactants, which can pose significant toxicity risks. We present a comprehensive, critical review of the use of HA in the field of ophthalmology and ocular drug delivery, with a discussion of the physicochemical and biological properties of HA that render it a suita-ble excipient for drug delivery to both the anterior and posterior segments of the eye. The pivotal focus of this review is a discussion of the formation of HA-based nanoparticles via poly-electrolyte complexation, a mild method of preparation driven primarily by electrostatic interaction between opposing polyelectrolytes. To the best of our knowledge, despite the growing number of publications centred around the development of HA-based polyelectrolyte complexes (HA-PECs) for ocular drug delivery, no review articles have been published in this area. This review aims to bridge the identified gap in the literature by (1) reviewing recent advances in the area of HA-PECs for anterior and posterior ODD, (2) describing the mechanism and thermodynamics of polyelectrolyte complexation, and (3) critically evaluating the intrinsic and extrinsic formulation parameters that must be considered when designing HA-PECs for ocular application.
... 27 Secondly, hyaluronic acid also has a positive contribution to the liquid absorption properties of the fibers due to its excellent hydrophilic and gelation properties. 28 The sodium and calcium ions exchange phenomenon of alginate has also been illustrated in Figure 5. ...
... However, the presence of calcium ions may have suppressed these properties of hyaluronic acid present in the fiber resulting in minor swelling and liquid absorption. 28 Swelling % in saline solution is high as there is a maximum ion exchange of calcium ions with sodium ions. The fibers absorbed an excess amount of solution, resulting in gel formation. ...
Article
Sodium alginate and hyaluronic acid are being used extensively in wound care applications for their exceptional properties, including gelation, biocompatibility, exudate absorption, and the ability to provide a moist environment to the wound bed that accelerates healing. This research work presents the potential of developing silver and zinc ions loaded bioactive fibers for wound care applications with improved absorption, swelling (gelling), and antibacterial properties. The effect of loading silver and zinc on alginate/hyaluronic acid biofiber absorption, swelling (gelation), tensile and antibacterial activity were analyzed. It was found that the addition of silver and zinc salts improved the absorption, tensile, and gelation properties of alginate/hyaluronic acid fibers. Zinc-containing fibers exhibited superior properties to silver-containing fibers. The presence of hyaluronic acid influenced the release of silver and zinc ions in various liquid media with a maximum of 26 g/g absorption was observed which suggested the good wound exudate absorption capacity of the developed fibers. The developed fibers showed good antibacterial activity against Staphylococcus aureus and Escherichia coli strains.
... Hyaluronic acid is remarkably hydrophilic and readily soluble in water. Its rheological properties are uncommon, exhibiting high water-retention capability, usually described as a lubricant [145,146]. Its ubiquitous presence in the mammalian extracellular matrix has made hyaluronic acid a good candidate for functional medical materials that promote wound healing. ...
Article
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Nanotechnology is a powerful tool for engineering functional materials that has the potential to transform textiles into high-performance, value-added products. In recent years, there has been considerable interest in the development of functional textiles using metal nanoparticles (MNPs). The incorporation of MNPs in textiles allows for the obtention of multifunctional properties, such as ultraviolet (UV) protection, self-cleaning, and electrical conductivity, as well as antimicrobial, antistatic, antiwrinkle, and flame retardant properties, without compromising the inherent characteristics of the textile. Environmental sustainability is also one of the main motivations in development and innovation in the textile industry. Thus, the synthesis of MNPs using ecofriendly sources, such as polysaccharides, is of high importance. The main functions of polysaccharides in these processes are the reduction and stabilization of MNPs, as well as the adhesion of MNPs onto fabrics. This review covers the major research attempts to obtain textiles with different functional properties using polysaccharides and MNPs. The main polysaccharides reported include chitosan, alginate, starch, cyclodextrins, and cellulose, with silver, zinc, copper, and titanium being the most explored MNPs. The potential applications of these functionalized textiles are also reported, and they include healthcare (wound dressing, drug release), protection (antimicrobial activity, UV protection, flame retardant), and environmental remediation (catalysts).
... The estimated quantity of HA in the human body is 15 g, of which one-third is replenished daily, with an average half-life varying from 2.3 to 5.5 min. Approximately 7-8 g of this HA is found in the skin; moreover, HA is also located in connective tissue, vitreous humor, synovial fluids, and others (Necas et al., 2008). Hydrolysis of HA is predominantly by the HAase family, which may be categorized based on the producing organism into (i) mammalian HAases, which cleave the β1-4 glycosidic linkage; (ii) leech HAases, which target the β1-3 glycosidic bond; and (iii) bacterial HAase (B-HAase), also known as hyaluronan lyase, which targets β1-4 bond like mammalian enzyme; however, the end product here is unsaturated disaccharides (Hynes & Walton, 2000). ...
Article
Hyaluronic acid (HA) has become a topic of significant interest in drug delivery research due to its excellent properties, including biosafety, biodegradability, and nonimmunogenicity. Moreover, due to its ease of modification, HA can be used to prepare several HA-based nanosystems using various approaches. These approaches involve conjugating/grafting of hydrophobic moieties, polyelectrolytes complexation with cationic polymers, or surface modification of various nanoparticles using HA. These nanoparticles are able to selectively deliver antibacterial drugs or diagnostic molecules into the site of infections. In addition, HA can bind with overexpressed cluster of differentiation 44 (CD44) receptors in macrophages and also can be degraded by a family of enzymes called hyaluronidase (HAase) to release drugs or molecules. By binding with these receptors or being degraded at the infection site by HAase, HA-based nanoparticles allow enhanced and targeted antibacterial delivery. Herein, we present a comprehensive and up-to-date review that highlights various techniques of preparation of HA-based nanoparticles that have been reported in the literature. Furthermore, we also discuss and critically analyze numerous types of HA-based nanoparticles that have been employed in antibacterial delivery to date. This article offers a critical overview of the potential of HA-based nanoparticles to overcome the challenges of conventional antibiotics in the treatment of bacterial infections. Moreover, this review identifies further avenues of research for developing multifunctional and biomimetic HA-based nanoparticles for the treatment, prevention, and/or detection of pathogenic bacteria. This article is categorized under: Therapeutic Approaches and Drug Discovery > Nanomedicine for Infectious Disease Nanotechnology Approaches to Biology > Nanoscale Systems in Biology Therapeutic Approaches and Drug Discovery > Emerging Technologies.
... HA produced in mammals has an essential biological function. Therefore, the primary source of HA is animal tissues; however, extensive purification protocols are required to prevent toxin contamination [3,4]. ...
Article
Full-text available
The growing, existing demand for low-cost and high-quality hyaluronic acid (HA) needs an outlook of different possible production strategies from renewable resources with the reduced possibility of cross-infections. Recently, the possibility of producing HA from harmless microorganisms appeared, which offers the opportunity to make HA more economical, without raw material limitations, and environmentally friendly. HA production is mainly reported with Lancefield Streptococci A and C, particularly from S. equi and S. zooepidemicus. Various modes of fermentation such as batch, repeated batch, fed-batch, and continuous culture have been investigated to optimize HA production, particularly from S. zooepidemicus, obtaining a HA yield of 2.5 g L−1 – 7.0 g L−1. Among the different utilized DSP approaches of HA production, recovery with cold ethanol (4°C) and cetylpyridinium chloride is the ideal strategy for lab-scale HA production. On the industrial scale, besides using isopropanol, filtration (0.22 um), ultrafiltration (100 kDa), and activated carbon absorption are employed to obtain HA of low molecular weight and additional ultrafiltration to purify HA of higher MW. Even though mature technologies have already been developed for the industrial production of HA, the projections of increased sales volume and the expansion of application possibilities require new processes to obtain HA with higher productivity, purity, and specific molecular weights. In this review, we have put forth the progress of HA technological research by discussing the microbial biosynthetic aspects, fermentation and downstream strategies, industrial-scale scenarios of HA, and the prospects of HA production to meet the current and ongoing market demands.
... Recently, CD44 was also identified as a typical surface marker for cancer stem cells, being a critical regulator of cancer stemness, self-renewal, tumor initiation, and metastases [203]. A link with the expression of one or more CD44v in individual types of cancer has been proven, for example in: breast cancer (CD44v3, [8][9][10] [204,205], head and neck squamous carcinomas (CD44v3) [206,207], prostate cancer (CD44v6) [208], pancreatic cancer (CD44v4, 5,6,9) [209][210][211], and gastrointestinal cancer (CD44v6) [212,213]. Therefore, CD44v expression is currently being studied intensively, as individual isoforms may serve as valuable diagnostic or prognostic markers, as well as ideal targets for developing clinical therapeutics in many types of cancer [214]. ...
Article
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Hyaluronic acid (HA) has a special position among glycosaminoglycans. As a major component of the extracellular matrix (ECM). This simple, unbranched polysaccharide is involved in the regulation of various biological cell processes, whether under physiological conditions or in cases of cell damage. This review summarizes the history of this molecule’s study, its distinctive metabolic pathway in the body, its unique properties, and current information regarding its interaction partners. Our main goal, however, is to intensively investigate whether this relatively simple polymer may find applications in protecting against ionizing radiation (IR) or for therapy in cases of radiation-induced damage. After exposure to IR, acute and belated damage develops in each tissue depending upon the dose received and the cellular composition of a given organ. A common feature of all organ damage is a distinct change in composition and structure of the ECM. In particular, the important role of HA was shown in lung tissue and the variability of this flexible molecule in the complex mechanism of radiation-induced lung injuries. Moreover, HA is also involved in intermediating cell behavior during morphogenesis and in tissue repair during inflammation, injury, and would healing. The possibility of using the HA polymer to affect or treat radiation tissue damage may point to the missing gaps in the responsible mechanisms in the onset of this disease. Therefore, in this article, we will also focus on obtaining answers from current knowledge and the results of studies as to whether hyaluronic acid can also find application in radiation science.
... Crosslinked hyaluronic acid is a biocompatible, viscoelastic, and hygroscopic compound (163,164). One of the main properties of HA is its capacity to surround itself with water molecules, being a highly hydrophilic polymer, preventing dehydration and maintains skin structure. ...
Thesis
Hyaluronic acid (HA), a major glycosaminoglycan in the extracellular matrix, plays an important role in a number of biological processes such as wound healing, cancer or embryonic development. The catabolism of HA generates fragments that play a role in migration, apoptosis or immune regulation. HA metabolism is a finely regulated process, adapting its functions to its microenvironment. In the event of deregulation of the metabolism, the accumulation of HA fragments seems to be strongly related to the severity of chronic inflammatory pathologies. Nevertheless, due to its rheological and structural properties, HA prevents dehydration and maintains the structure of the skin. Although it is generally considered to be an inert and biocompatible material, rare cases of late side effects have been observed. Thus, in order to improve our understanding of the mechanisms that can lead to late complications, we hypothesized that the rare complications observed could be related to an inflammatory microenvironment. In this thesis work, we evaluated the inflammatory and vascular potential of degraded and non-degraded HA in dermal cells, immune cells and in a mouse model. In an inflammatory microenvironment, adding HA fragments to endothelial cells and dermal fibroblasts does not worsen the inflammatory response, unlike immune cells. This suggests the involvement of M1 macrophages and mature dendritic cells to a lesser extent in the transmission of inflammation to the tissue. In addition, in a mouse model of chronic low-grade inflammation we showed in vivo that injection of HA can lead to microvascular sensitivity in response to acetylcholine. The hypothesis is that this vascular sensitivity is linked to the recruitment of immune cells at the injection site. Furthermore, we demonstrated in vitro that in the absence of immune cells, an inflammatory vascular microenvironment could lead to an alteration of the extracellular matrix. These results show a diversity in cell-cell interactions depending on the inflammatory degree of the microenvironment, which in the presence of degradation fragments of HA (i.e. during aging or lesion) could weaken the skin tissue
... Also, its unique viscoelasticity and limited immunogenicity led to its use in various medical applications, such as osteoarthritis treatment (OA), eye surgery aid, and wound regeneration. 2 We will discuss the use of injectable hyaluronic acid as a dermal filler and its interactions with its primary receptor, CD44, to understand how it can promote rejuvenation through its biological interactions. ...
... Большой интерес на протяжении целого ряда лет сохраняется к ЛИТ с использованием гиалуроновой кислоты -гликозаминогликана, в структуре которого представлены повторяющиеся фрагменты β-1,4-D-глюкуроновой кислоты и β-1,3-N-ацетилглюкозамина, образующие молекулы различной массы до 6 500 и даже до 10 900 кДа [11][12][13]. Гиалуроновая кислота (ГлК) способствует восстановлению механической устойчивости хрящевой ткани и сухожилий, ускорению репаративных процессов за счет повышения синтеза эндогенного гиалуроната, обеспечивает вязкоэластичность, удерживает влагу, осуществляет защитную функцию тканей сустава [14][15][16][17]. ГлК ингибирует экспрессию провоспалительных медиаторов ИЛ-1β и ИЛ-6, матриксных металлопротеиназ, снижает синтез простагландина Е2, пролиферацию синовиальных клеток. ...
Article
The article deals with local injection therapy (LIT) in arthrological practice, the purpose of which is the introduction of a drug into the joint cavity or periarticularly, removal of excess synovial fluid from the joint cavity with subsequent analysis of its composition for differential diagnosis and decision-making on treatment tactics. The most commonly used in LIT are glucocorticoids, hyaluronic acid (GlK), autologous platelet-rich plasma. The method of introducing GLA into the joint cavity is considered by experts as a promising method for the treatment of osteoarthritis, the most common chronic joint disease. GlK-based preparations differ in concentration, molecular weight and production method. The article presents the results of studies using high- or low-molecular drugs GlK in the treatment of patients with osteoarthritis of the knee, hip and small joints. Attention is drawn to the fact that LIT with GlK preparations is used in the treatment of other diseases of the musculoskeletal system. Data are presented, including those of Russian authors, on the effectiveness of GlK in OA of the shoulder joints, in the syndrome of compression of the shoulder rotators. Thus, the restoration of range of motion and a significant reduction in pain in the shoulder area was achieved against the background of the use of 3 to 5 injections of the GlK preparation produced in the Russian Federation by bacterial fermentation. The unique production technology of the GlK preparation makes it possible to reduce the risk of bacterial endotoxins, completely purify the solution from hyaluronidase, increase homogeneity, and preserve the molecular weight. On the basis of the obtained evidence of efficacy and safety, HLA preparations are included by various medical associations and professional communities in Russian and international clinical guidelines for the treatment of OA.
... HA is a repeating disaccharide unit of N-acetylglucosamine and glucuronic acid [52]. The central nervous system (CNS), which includes the brain and spinal cord, contains a high HA level [53]. ...
Article
Full-text available
The biology and physics underlying glioblastoma is not yet completely understood, resulting in the limited efficacy of current clinical therapy. Recent studies have indicated the importance of mechanical stress on the development and malignancy of cancer. Various types of mechanical stress activate adaptive tumor cell responses that include alterations in the extracellular matrix (ECM) which have an impact on tumor malignancy. In this review, we describe and discuss the current knowledge of the effects of ECM alterations and mechanical stress on GBM aggressiveness. Gradual changes in the brain ECM have been connected to the biological and physical alterations of GBM cells. For example, increased expression of several ECM components such as glycosaminoglycans (GAGs), hyaluronic acid (HA), proteoglycans and fibrous proteins result in stiffening of the brain ECM, which alters inter- and intracellular signaling activity. Several mechanosensing signaling pathways have been identified that orchestrate adaptive responses, such as Hippo/YAP, CD44, and actin skeleton signaling, which remodel the cytoskeleton and affect cellular properties such as cell–cell/ECM interactions, growth, and migration/invasion of GBM cells. In vitro, hydrogels are used as a model to mimic the stiffening of the brain ECM and reconstruct its mechanics, which we also discuss. Overall, we provide an overview of the tumor microenvironmental landscape of GBM with a focus on ECM stiffening and its associated adaptive cellular signaling pathways and their possible therapeutic exploitation.
... Hyaluronan synthases (HAS 1, 2 and 3) are suspected to increase cell migration and adhesion, which also affect immune cell infiltration. Their properties influence the early stage of inflammation and modulate it (Necas et al., 2008). Additionally, HASs, IL-6 and MCP-1 seem to play an important role in the pathogenesis of endometrosis, another crucial reproductive problem in mares (Aresu et al., 2012). ...
Article
Full-text available
Endometritis is an important issue decreasing mares’ fertility. In the case of endometritis both, inflammatory cells infiltration and proinflammatory molecules production are regulated by various cellular and gene‐regulatory mechanisms, including the nuclear factor‐κB (NF‐κB) dependent pathway. NF‐κB signaling pathway has been recently studied in the equine endometrium in the context of endometrosis. Thus, this study aimed to determine gene transcription of NF‐κB subunits (RelA; NF‐κB1; NF‐κB2), proinflammatory molecules (MCP‐1; IL‐6), and hyaluronan synthases (HAS 1; HAS 2; HAS 3) in endometritis and compare them with the intensity and type of inflammatory cell infiltration. Endometrial samples, collected post‐mortem from cyclic mares in estrus or diestrus, were classified histologically and examined using quantitative PCR. Transcription NF‐κB subunits genes did not differ with either inflammatory intensity or type of inflammatory cell infiltration. Transcription of MCP‐1 and IL‐6 genes increased with the severity of inflammation, with the involvement of HAS 3 and HAS 2 genes, as opposed to HAS 1 genes. These proinflammatory molecules and hyaluronan synthases in the equine inflamed endometrium do not seem to be regulated by the NF‐κB pathway. Hence, separate signaling pathways for the development and progression of equine endometritis and endometrosis may be suggested.
... Overall, it compromises a major part of the extracellular matrix and can be found in almost all human tissues. Because of its role in vivo, it has been used for various medical applications [43][44][45]. CMC and XG are also polysaccharides, and all three of them are able to form hydrocolloids [46,47], but different from a lot of commonly used hydrogels, they are not crosslinked in our application presented in this paper. Interestingly, although CMC, a cellulose derivative produced from wood, and xanthan gum, a polysaccharide produced by bacteria, do not have a physiological role in mammalians, they still promoted spheroid formation of MSCs. ...
Article
Full-text available
Mesenchymal stem cells (MSCs) are primary candidates in tissue engineering and stem cell therapies due to their intriguing regenerative and immunomodulatory potential. Their ability to self-assemble into three-dimensional (3D) aggregates further improves some of their therapeutic properties, e.g., differentiation potential, secretion of cytokines, and homing capacity after administration. However, high hydrodynamic shear forces and the resulting mechanical stresses within commercially available dynamic cultivation systems can decrease their regenerative properties. Cells embedded within a polymer matrix, however, lack cell-to-cell interactions found in their physiological environment. Here, we present a “semi scaffold-free” approach to protect the cells from high shear forces by a physical barrier, but still allow formation of a 3D structure with in vivo-like cell-to-cell contacts. We highlight a relatively simple method to create core–shell capsules by inverse gelation. The capsules consist of an outer barrier made from sodium alginate, which allows for nutrient and waste diffusion and an inner compartment for direct cell-cell interactions. Next to capsule characterization, a harvesting procedure was established and viability and proliferation of human adipose-derived MSCs were investigated. In the future, this encapsulation and cultivation technique might be used for MSC-expansion in scalable dynamic bioreactor systems, facilitating downstream procedures, such as cell harvest and differentiation into mature tissue grafts.
... The enhancements of anticancer activities were found when nanoparticles of an anticancer drug were coated with hyaluronic acid [38][39][40]. This may be because the high binding affinity of hyaluronic acid to the CD44 receptor, which overexpresses in tumor cell [39,[41][42][43]. ...
Article
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Novel turmeric rhizome extract nanoparticles (TE-NPs) were developed from fractions of dried turmeric (Curcuma longa Linn.) rhizome. Phytochemical studies, by using HPLC and TLC, of the fractions obtained from ethanol extraction and solvent–solvent extraction showed that turmeric rhizome ethanol extract (EV) and chloroform fraction (CF) were composed mainly of three curcuminoids and turmeric oil. Hexane fraction (HE) was composed mainly of turmeric oil while ethyl acetate fraction (EA) was composed mainly of three curcuminoids. The optimal TE-NPs formulation with particle size of 159.6 ± 1.7 nm and curcumin content of 357.48 ± 8.39 µM was successfully developed from 47-run D-optimal mixture–process variables experimental design. Three regression models of z-average, d50, and d90 could be developed with a reasonable accuracy of prediction (predicted r2 values were in the range of 0.9120–0.9992). An in vitro cytotoxicity study using MTT assay demonstrated that the optimal TE-NPs remarkably exhibited the higher cytotoxic effect on human hepatoma cells, HepG2, when compared with free curcumin. This study is the first to report nanoparticles prepared from turmeric rhizome extract and their cytotoxic activity to hepatic cancer cells compared with pure curcumin. These nanoparticles might serve as a potential delivery system for cancer therapy.
... It can also be called a hyaluronan [11,12]. HA is distributed widely throughout epithelial, connective, and neural tissues [13,14]. It is an endogenous component of human skin [15]. ...
Article
Full-text available
In this paper, surfaces of thin films prepared from blends of collagen, hyaluronic acid, and chitosan and modified by neodymium laser radiation were researched. To evaluate the laser beam effect on the surface structure, scanning electron microscopy (SEM) imaging and infrared spectroscopy (FTIR-ATR) were employed. The results demonstrated that during laser treatment the specimens lost water due to the evaporation process. SEM images revealed some changes in the biopolymer films structure. After laser treatment, the micro-foam formation was observed on the biopolymeric films. The micro-foaming in films based on ternary blends was more extensive than in those made of a single biopolymer. The results of this study indicate that collagen, hyaluronic acid, and chitosan materials can be modified with laser treatment. Such treatment can be used for material modification for potential biomedical purposes.
... HA produced in mammals has an essential biological function. Therefore, the primary source of HA is animal tissues; however, extensive purification protocols are required to prevent toxin contamination [3,4]. ...
Article
Full-text available
The growing, existing demand for low-cost and high-quality hyaluronic acid (HA) needs an outlook of different possible production strategies from renewable resources with the reduced possibility of cross-infections. Recently, the possibility of producing HA from harmless microorganisms appeared, which offers the opportunity to make HA more economical, without raw material limitations, and environmentally friendly. HA production is mainly reported with Lancefield Streptococci A and C, particularly from S. equi and S. zooepidemicus. Various modes of fermentation such as batch, repeated batch, fed-batch, and continuous culture have been investigated to optimize HA production, particularly from S. zooepidemicus, obtaining a HA yield of 2.5 g L −1-7.0 g L −1. Among the different utilized DSP approaches of HA production, recovery with cold ethanol (4°C) and cetylpyridinium chloride is the ideal strategy for lab-scale HA production. On the industrial scale, besides using isopropanol, filtration (0.22 um), ultrafiltration (100 kDa), and activated carbon absorption are employed to obtain HA of low molecular weight and additional ultrafiltration to purify HA of higher MW. Even though mature technologies have already been developed for the industrial production of HA, the projections of increased sales volume and the expansion of application possibilities require new processes to obtain HA with higher productivity, purity, and specific molecular weights. In this review, we have put forth the progress of HA technological research by discussing the microbial biosynthetic aspects, fermentation and downstream strategies, industrial-scale scenarios of HA, and the prospects of HA production to meet the current and ongoing market demands. Marine Sources Microbial FermentaƟon Hyaluronic acid ApplicaƟons Dermatology Urology CosmeƟc ophthalmology OsteoarthriƟs Surgery and Wound Healing ARTICLE HISTORY
... It should be noted that in several studies where good biocompatibility of ε-PL containing composites has been shown, ε-PL concentrations lower than those used in the current study were applied [17,66,69]. On the other hand, HA as a bioactive component for tissue engineering applications is widely investigated and is considered non-cytotoxic, and shows good biocompatibility [70]. However, slight changes in cell viability have been observed in different cell lines when different HA-containing materials were assayed for cytotoxicity [71,72]. ...
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The design of multifunctional hydrogels based on bioactive hyaluronic acid (HA) and antibacterial cationic polymer ɛ-poly-l-lysine (ε-PL) is a promising tool in tissue engineering applications. In the current study, we have designed hyaluronic acid and ɛ-polylysine composite hydrogel systems with antibacterial and cell attractive properties. Two distinct crosslinking approaches were used: the physical crosslinking based on electrostatic attractions and the chemical crosslinking of charged functional groups (-NH2 and -COOH). The impact of the crosslinking strategy on fabricated hydrogel molecular structure, swelling behavior, gel fraction, morphology, porosity, viscoelastic properties, antibacterial activity, and in vitro biocompatibility was evaluated. Both chemically and physically crosslinked HA/ԑ-PL hydrogels demonstrated fast swelling behavior and long-term stability for at least 28 days, as well as similar order of stiffness (10–30 kPa). We demonstrated that physically crosslinked hydrogels inhibited over 99.999% of Gram-negative E. coli, while chemically crosslinking strategy led to the antibacterial efficiency decrease. However, cell viability was significantly improved, confirming the importance of the applied crosslinking approach to the antibacterial activity and in vitro biocompatibility. The distinct differences in the physicochemical and biological properties of the developed materials provide new opportunities to design next-generation functional composite hydrogel systems.
... HA is an anionic polysaccharide extensively employed in DDS development due to its excellent biocompatibility and biodegradability (Necas, Bartosikova, Brauner, & Kolar, 2008). CD44 receptors, which are natural receptors of HA, and hyaluronidase (HAase), which is a family of enzymes catalyzing HA degradation, are both overexpressed in many cancer cells, e.g. in breast and brain tumors (Stern, 2008;Xu, Niu, Yuan, Wu, & Liu, 2020). ...
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A novel kind of thermoresponsive polyelectrolyte complex-based nanogels (PEC-NGs) was elaborated by mixing hyaluronic acid (HA) functionalized with Jeffamine® M-2005 (M2005, a thermoresponsive amine-terminated polyether) and diethylaminoethyl dextran (DEAE-D) or poly-l-lysine (PLL) in water. The presence of M2005 grafts led to PEC-NGs with larger particle size, lower net surface charge and thermoresponsiveness, namely shrinkage with increasing hydrophobicity at higher temperature. Both M2005 grafts and replacing DEAE-D with PLL as polycation allowed PEC-NGs to have higher stability against salinity and better encapsulation of curcumin, most probably through intraparticle hydrophobic interactions, whereas interparticle hydrophobic interactions may facilitate particle aggregation over time. Curcumin encapsulation can be optimized by applying higher temperature during the complexation. Enzymatic degradability of PEC-NGs was also verified through particle size evolution in the presence of hyaluronidase. These results provide new insights into the physicochemical aspect of such systems as promising nanocarriers for drug delivery.
... HA is formed by repeated chains of disaccharide units of N-acetylglucosamine and sodium glucuronate and represents a fundamental component of the extracellular matrix of most tissues, including the skin. 1,2 One of the leading causes of skin aging is the reduction of HA concentration within the connective tissues. The decrease in HA levels contributes to its disassociation with collagen and elastin and the reduced water-binding capacity. ...
... Native HA exists at a high-molecularweight form (10 4 -10 7 Da) in the human connective and epithelial tissues, and it comprises a major component of the extracellular matrix. 8 In the oral cavity, it constitutes a structural part of both mineralized (alveolar bone, cementum) and non-mineralized tissues (periodontal ligament, gingival tissues). 9 Hyaluronic acid participates in many biological activities, such as phagocytosis, cell migration, and adhesion, and exerts anti-inflammatory and anti-bacterial actions demonstrating a significant role in wound repair. ...
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Aim: The main purpose of the present study was to investigate the potential benefit of local use of hyaluronic acid as an adjunct to periodontal therapy, since commercial products of hyaluronic acid (HA), due to its anti-inflammatory and anti-bacterial actions and its significant role in wound repair, have been proposed as adjuncts to either nonsurgical or surgical periodontal therapy. Materials and methods: A total of 19 electronic databases were searched and the appropriate studies were identified with the use of specific eligibility criteria, according to PRISMA guidelines. Two reviewers independently screened and selected the studies and made the data extraction and the assessment of risk of bias, by using the Cochrane risk of bias tool. Results: Out of 3,186 papers, 38 randomized clinical trials (8 related to gingivitis therapy, 20 related to nonsurgical periodontal therapy, and 10 related to surgical periodontal therapy) were finally included in the review. The outcomes were categorized as primary (that answered the focus question) and secondary (regarding additional quality characteristics). The adjunct use of HA combined to all treatment modalities shows improvement of patients' postoperative course, in terms of decreased inflammatory reactions, and changes in periodontal pocket depth and clinical attachment level. No side effects were reported in any of the included studies. Among the secondary outcomes were the variety of HA formulations and chemical forms, the variety in application, follow-up protocol and blinding design, the uneven geographic distribution of the studies, and the low bibliometric characteristics of most studies. Conclusion: Overall and despite the positive effects reported, further research is needed to define the ideal HA compound, formulation, and regimen characteristics for periodontal disease treatment. Clinical significance: The adjunct use of HA may lead in the reduction of the prescription of nonsteroid anti-inflammatory drugs and achieve improved clinical parameters, including periodontal probing depth, periodontal inflammation, and clinical attachment level.
... 100 It can bind to the cell surface receptor CD44 to modulate cell differentiation, migration, and angiogenesis without eliciting any foreign-body response, making it an intriguing material for vascular application. 101 High molecular weight HA inhibits EC proliferation, whereas low molecular weight HA oligosaccharides (0.75−10 kDa) stimulate EC proliferation and migration. HA has also been found to inhibit fibroblast proliferation, which is another action that may assist in the prevention of graft occlusion. ...
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Small-diameter artificial vascular grafts (SDAVG) are used to bypass blood flow in arterial occlusive diseases such as coronary heart or peripheral arterial disease. However, SDAVGs are plagued by restenosis after a short while due to thrombosis and the thickening of the neointimal wall known as intimal hyperplasia (IH). The specific causes of IH have not yet been deduced; however, thrombosis formation due to bioincompatibility as well as a mismatch between the biomechanical properties of the SDAVG and the native artery has been attributed to its initiation. The main challenges that have been faced in fabricating SDAVGs are facilitating rapid re-endothelialization of the luminal surface of the SDAVG and replicating the complex viscoelastic behavior of the arteries. Recent strategies to combat IH formation have been mostly based on imitating the natural structure and function of the native artery (biomimicry). Thus, most recently, developed grafts contain a multilayered structure with a designated function for each layer. This paper reviews the current polymeric, biomimetic SDAVGs in preventing the formation of IH. The materials used in fabrication, challenges, and strategies employed to tackle IH are summarized and discussed, and we focus on the multilayered structure of current SDAVGs. Additionally, the future aspects in this area are pointed out for researchers to consider in their endeavor.
... One of them is to use hyaluronic acid (HA), a natural agonist of CD44. It is a critical component of the extracellular matrix and it is degraded through enzymatic reactions [19] . So, it is bio-compatible. ...
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Nanocarriers have been developed in order to protect drugs or to improve drugs efficiency by reaching the damaged tissue and avoiding systemic and local toxicity. By using HSP90 inhibitors, some cancer drug resistances have been overcome and the loading into nanocarriers of such drugs has shown an increase of their activities. This review will present some advantages of HSP90 inhibitors to treat resistant tumors; especially those targeting the mitochondrial protein TRAP1. We will also focus on the targeting of the primary tumors, cancer stem cells and metastatic cells.
... HA is a negatively-charged polysaccharide, naturally existing in the vitreous body, joint, and skin. However, CHI is the only cationic natural polysaccharide [40,41]. Considering the advantages of the hydration property of polysaccharides, Lin et al. [42] constructed a hydrogel-like polyelectrolyte multilayer coating composed of HA and CHI components on the surface of the silicone IOL using the electrostatic layer-by-layer (LBL) self-assembly method. ...
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Posterior capsular opacification (PCO) is the most common complication resulting from cataract surgery and limits the long-term postoperative visual outcome. Using Nd:YAG laser-assisted posterior capsulotomy for the clinical treatment of symptomatic PCO increases the risks of complications, such as glaucoma, retinal diseases, uveitis, and intraocular lens (IOL) pitting. Therefore, finding how to prevent PCO development is the subject of active investigations. As a replacement organ, the IOL is implanted into the lens capsule after cataract surgery, but it is also associated with the occurrence of PCO. Using IOL as a medium for PCO prophylaxis is a more facile and efficient method that has demonstrated various clinical application prospects. Thus, scientists have conducted a lot of research on new intraocular lens fabrication methods, such as optimizing IOL materials and design, and IOL surface modification (including plasma/ultraviolet/ozone treatment, chemical grafting, drug loading, coating modification, and layer-by-layer self-assembly methods). This paper summarizes the research progress for different types of intraocular lenses prepared by different surface modifications, including anti-biofouling IOLs, enhanced-adhesion IOLs, micro-patterned IOLs, photothermal IOLs, photodynamic IOLs, and drug-loading IOLs. These modified intraocular lenses inhibit PCO development by reducing the residual intraoperative lens epithelial cells or by regulating the cellular behavior of lens epithelial cells. In the future, more works are needed to improve the biosecurity and therapeutic efficacy of these modified IOLs.
... SARS-CoV-2 has been demonstrated to infect diverse cell types in different tissues [150,[332][333][334], including lungs, kidneys, brain, and heart. Interestingly, hyaluronan is widely distributed in all parts of the body and exercises a myriad of biological functions in different cell types [335]. Particularly, HIS-SARS2 can induce hyaluronan production via stimulating HAS2 expression in diverse cells associated with lung, blood vessel, and kidney [17], implying the inflammation trigged by hyaluronan may be the cause of multiple organ injury. ...
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COVID-19 caused by SARS-CoV-2 has created formidable damage to public health and market economy. Currently, SARS-CoV-2 variants has exacerbated the transmission from person-to-person. Even after a great deal of investigation on COVID-19, SARS-CoV-2 is still rampaging globally, emphasizing the urgent need to reformulate effective prevention and treatment strategies. Here, we review the latest research progress of COVID-19 and provide distinct perspectives on the mechanism and management of COVID-19. Specially, we highlight the significance of Human Identical Sequences (HIS), hyaluronan, and hymecromone (“Three-H”) for the understanding and intervention of COVID-19. Firstly, HIS activate inflammation-related genes to influence COVID-19 progress through NamiRNA-Enhancer network. Accumulation of hyaluronan induced by HIS-mediated HAS2 upregulation is a substantial basis for clinical manifestations of COVID-19, especially in lymphocytopenia and pulmonary ground-glass opacity. Secondly, detection of plasma hyaluronan can be effective for evaluating the progression and severity of COVID-19. Thirdly, spike glycoprotein of SARS-CoV-2 may bind to hyaluronan and further serve as an allergen to stimulate allergic reaction, causing sudden adverse effects after vaccination or the aggravation of COVID-19. Finally, antisense oligonucleotides of HIS or inhibitors of hyaluronan synthesis (hymecromone) or antiallergic agents could be promising therapeutic agents for COVID-19. Collectively, Three-H could hold the key to understand the pathogenic mechanism and create effective therapeutic strategies for COVID-19.
Chapter
Tissue engineering and stem cell research have made great progresses in overcoming the problems caused by tissue damage in the body and the side effects of traditional therapies. Although a wide range of techniques are used in the field of tissue engineering, tissue and stem cell engineering represents a viable alternative source for the treatment of complex lesions in various tissues. The most appropriate methods for replacing damaged tissues require various techniques that face many problems. Generally, the final stage of organ failure is treated with tissue transplantation, but the results are usually not completely satisfactory due to the effects of immunosuppression, increasing the number of failed transplants, and decreasing the number of organ donors. Hence, regenerative medicine and tissue engineering are working to understand the mechanisms of tissue regeneration, and to find a way to regenerate damaged tissues. Stem cells are considered as one of the important components in tissue engineering, which due to the polymer structure of tissue engineering scaffolds can repair or regenerate the desired tissue. One of the newest stem cells introduced is induced multipotent stem cells (iPSCs), which are derived from epithelial cells and are multipotent. iPSC lines can differentiate into most cells in the body, just like embryonic stem cells (ESCs). These cells also have the potential to become multifunctional clinical and research tools for understanding and modeling disease, developing and screening selective drugs, and delivering alternative-cell therapy to support regenerative medicine. These cells have attracted the attention of many scientists because they are taken from the patient in terms of nonrejection as well as ethical issues in ESCs. This chapter will review the application of iPSCs in various areas of tissue engineering that have been reported so far.
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With the definition of the 3R principle by Russel and Burch in 1959, the search for an adequate substitute for animal testing has become one of the most important tasks and challenges of this time, not only from an ethical, but also from a scientific, economic, and legal point of view. Microtissue‐based in vitro model systems offer a valuable approach to address this issue by accounting for the complexity of natural tissues in a simplified manner. To increase the functionality of these model systems and thus make their use as a substitute for animal testing more likely in the future, the fundamentals need to be continuously improved. Corresponding requirements exist in the development of multifunctional, hydrogel‐based materials, whose properties are considered in this review under the aspects of processability, adaptivity, biocompatibility, and stability/degradability.
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With approximately 250,000 marine species, the ocean is a vast reservoir of biodiversity and an abundant biological resource of natural polymers. The wide variety, renewable nature, tunable physicochemical and structural behavior and appealing biological properties make these marine biopolymers particularly attractive to the scientific community and numerous industrial sectors. As raw materials, they offer novel opportunities for the development of bio-based materials in response to recent demands for biodegradable plastic materials to lower plastic pollution in marine ecosystems. The biodegradation of marine biopolymers and biopolymer-based materials depends on marine environmental conditions such as temperature, pH and in particular microbial population. Marine microorganisms producing biopolymer-degrading enzymes (i.e., hydrolases, lyases, oxidoreductases) are well studied, nonetheless the biodegradation processes of marine biopolymers-based materials in the marine/aquatic environment need further investigation. This review describes various biodegradation parameters and mechanisms of the degradation of marine biopolymers in the marine environment. It also puts emphasis on the marine microorganisms and the corresponding enzymes that catalyze the degradation of different marine biopolymers. Finally, it focuses on the few studies on biodegradation of emerging bio-based materials in aquatic ecosystems.
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The development of blood-interacting surfaces is critical to fabricate biomaterials for medical use, such as prostheses, implants, biosensors, and membranes. For instance, thrombosis is one of the leading clinical problems when polymer-based materials interact with blood. To overcome this limitation is necessary to develop strategies that limit platelets adhesion and activation. In this work, hyaluronan (HA)/chitosan (Chi) based-films, recently reported in the literature as platforms for tumor cell capture, were developed and, subsequently, functionalized with sulfated chitosan (ChiS) using a layer-by-layer technique. ChiS, when compared to native Chi, presents the unique abilities to confer anti-thrombogenic properties, to reduce protein adsorption, and also to limit calcification. Film physicochemical characterization was carried out using FTIR and XPS for chemical composition assessment, AFM for the surface morphology, and contact angle for hydrophilicity evaluation. The deposition of ChiS monolayer promoted a decrease in both roughness and hydrophilicity of the HA/Chi films. In addition, the appearance of sulfur in the chemical composition of ChiS-functionalized films confirmed the film modification. Biological assay indicated that the incorporation of sulfated groups limited platelet adhesion, mainly because a significant reduction of platelets adhesion to ChiS-functionalized films was observed compared to HA/Chi films. On balance, this work provides a new insight for the development of novel antithrombogenic biomaterials, opening up new possibilities for devising blood-interaction surfaces.
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Background: Tendinopathies are overuse tendon injuries showing load-dependant pain, stiffness, weakness of movement in the affected area, and impairment in the movements. The scientific interest on the role of Hyaluronic Acid (HA) for the management of tendinopathies has been increased due to its anti-inflammatory and lubricative properties. Objective: To collect evidence regarding the effectiveness and safety of HA injections in reducing pain in patients affected by tendinopathies. Methods: A scientific literature search was conducted using the PubMed, Medline and PEDro electronic databases. The databases were searched since their inception until July 2021. The search was limited to English language articles. Different combinations of the terms and MeSH terms "tendinopathy", "tendinosis", "tendinitis", "hyaluronic acid", "hyaluronate", "infiltration", "hyaluronic injections", "viscosupplementation" connected with various boolean operators were used for other electronic databases. Results: One hundred and one records were identified from the selected databases plus three additional papers identified by the authors through other sources. After removing duplicated papers and title/abstract screening, 19 studies were included in our review (eight papers on shoulder, three on elbow, four on hand, one on knee, and three on ankle). Conclusion: The results showed that none of the studies report severe adverse effects and most of them support the use of HA injections in tendinopathies, with a special attention to pain reduction and functional assessment. Further studies are warranted to better investigate effects and methods of administration of HA in tendinopathies.
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Konjac glucomannan (KGM) hydrogel has favorable gel-forming abilities, but its insufficient swelling capacity and poor control release characteristics limit its application. Therefore, in this study, oxidized hyaluronic acid (OHA) was used to improve the properties of KGM hydrogel. The influence of OHA on the structure and properties of KGM hydrogels was evaluated. The results show that the swelling capacity and rheological properties of the composite hydrogels increased with OHA concentration, which might be attributed to the hydrogen bond between the KGM and OHA, resulting in a compact three-dimensional gel network structure. Furthermore, epigallocatechin gallate (EGCG) was efficiently loaded into the KGM/OHA composite hydrogels and liberated in a sustained pattern. The cumulative EGCG release rate of the KGM/OHA hydrogels was enhanced by the increasing addition of OHA. The results show that the release rate of composite hydrogel can be controlled by the content of OHA. These results suggest that OHA has the potential to improve the properties and control release characteristics of KGM hydrogels.
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Breast cancer is the most common cancer in women with highest mortality rate wherein, mastectomy, lumpectomy, radiation therapy, hormone therapy, chemotherapy, and biological therapy are some of the current therapeutic alternatives for breast cancer. The treatments, however, are linked to short and long-term side effects like cardiac problems, anaemia, physiological distress, myelosuppression, peripheral neuropathy and thrombosis. Hence, the transpapillary route is a promising route for delivering drugs directly to the underlying breast tissues for the treatment of breast cancer by achieving higher drug levels at the tumor site. The aim of the present study was to check the potential of transpapillary route for artesunate-loaded microneedles, solid lipid nanoparticles (SLN)-loaded microneedles and layer-by-layer (LBL)-coated SLN-loaded microneedles against breast cancer. SLNs were prepared by hot-melt emulsification method and LBL-coating on SLNs was achieved by 1 mg/1 mL of chitosan and 2 mg/mL of hyaluronic acid. The particle sizes of selected formulations of SLNs and LBL-coated SLNs were found to be 213.72±2.53 nm and 319.39±1.25 nm, respectively. Zeta potentials of selected formulations SLNs and LBL-coated SLNs were found to be -27.32 ± 4.45 mV and -28.41 ± 3.67 mV, respectively. Three formulations of microneedles were prepared for comparative evaluation using artesunate, SLNs, LBL-coated SLNs. Microneedles were evaluated for FTIR, DSC, mechanical insertion property, surface morphology, in-vitro release, ex-vivo release and cell line studies. The cell line study and ex-vivo release profile of LBL-coated SLN microneedles showed a reduction in growth of cell line MCF 7 by 7.92 ± 1.54% and higher ex-vivo release (84.75 ± 2.02%) in comparison to plain drug. Thus, the LBL-coated SLNs microneedles of artesunate proved as an effective alternative against breast cancer cell line.
Chapter
This chapter describes briefly the processes of wound healing and the application of stem cell therapy for various wound healing. The current therapeutic outcomes suggest that a functional niche is needed to deliver stem cells toward improved cells engraftment, viability, and function. Hydrogels can be used as such delivery vehicles and thus, their components, means of fabrication, and their promising in wound healing are also discussed. In particular, injectable hydrogels formed by in situ cross‐linking transited from an aqueous mixture of gel precursors to a solid gel are in the focus. The injectable hydrogels are an attractive delivery niche for drugs and cells due to their tuneable mechanical and degradation properties that allow transplanting and localizing cells to a desired anatomic site. To design a device for an efficient healing outcome, key bioactive factors can be incorporated into the hydrogel scaffold. Advanced material science and techniques show great potential to create or mimic extracellular matrix niches, which will ultimately benefit the wound healing process.
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Soft‐tissue augmentation has gained much popularity in recent years. Hyaluronic acid (HA) based dermal fillers; a non‐permanent injectable device, can restore volume loss, fill fine lines and wrinkles and add curves and contours. HA based dermal fillers entered the non‐surgical treatment market in the late 1990's, however there is a lack of data and literature comparing the range of products and detailing the complexities of these products and how it relates to tissue performance. Measuring the physico‐chemical properties of these dermal fillers provide key parameters to predict their performance after injection into the body. This article reviews the currently reported methods and parameters used to characterize dermal fillers. The review of these methods and data from the literature provides a useful guide to clinicians and injectors in selecting the optimal product suitable for the needs of each patient This article is protected by copyright. All rights reserved.
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Hyaluronan (HA) is an essential carbohydrate in vertebrates that is a potentially robust bioindicator due to its critical roles in diverse physiological functions in health and disease. The intricate size-dependent function that exists for HA and its low abundance in most biological fluids have highlighted the need for sensitive technologies to provide accurate and quantitative assessments of polysaccharide molecular weight and concentration. We have demonstrated that solid state (SS-) nanopore technology can be exploited for this purpose, given its molecular sensitivity and analytical capacity, but there remains a need to further understand the impacts of experimental variables on the SS-nanopore signal for optimal interpretation of results. Here, we use model quasi-monodisperse HA polymers to determine the dependence of HA signal characteristics on a range of SS-nanopore measurement conditions, including applied voltage, pore diameter, and ionic buffer asymmetry. Our results identify important factors for improving the signal-to-noise ratio, resolution, and sensitivity of HA analysis with SS-nanopores.
Chapter
Bone tissue engineering (BTE) approaches are focused on repairing fractured or damaged bones. Exciting technologies have been developed to repair and regenerate damaged tissues and organs by the implantation of biopolymers’ composite scaffolds. The hierarchical nature of bone is an inspiration for nanostructured biomaterials, and nanostructured composite scaffolds are gaining special consideration for BTE. These scaffolds are fabricated from biopolymers and bioactive nanofillers and have biodegradability, good mechanical properties, and ease of formation characteristics necessary for osteogenesis. Biopolymers resemble extracellular matrix (ECM) with different levels of biodegradability and better biological qualities, and fillers like hydroxyapatite play a vital role in enhancing biocompatibility, restoration, and osteoconduction. Biopolymers in microspheres, nanofibers, tough foams, and hydrogels can be used to create a wide range of scaffolds. This review will discuss biopolymer-based nanocomposite materials for their design and properties. A detailed review and discussion of the current status and surface engineering of biopolymers and their applications in BTE and their feasibility of production and future applications have also been presented.
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After vitrectomy, the ideal vitreous substitute should be implanted to maintain the normal function of the eye. However, the existing materials (such as silicone oil, air, perfluorocarbons, etc.) still have some shortcomings and cannot fully meet the clinical needs. In this study, thiolated hyaluronic acid (SH-HA) was prepared based on hyaluronic acid. The SH-HA hydrogel was formed by a simple transformation of the sulfhydryl group to the disulfide bond, which had high transparency, controllable swelling property, suitable mechanical strength, excellent biocompatibility and similar physical and chemical properties to natural vitreous. SH-HA hydrogel was filled into the eyes of experimental rabbits to replace their own vitreous after vitrectomy. During the 90 days follow-up period, SH-HA hydrogel showed excellent intraocular compatibility, maintained normal intraocular pressure (IOP), and no cataract, endophthalmitis, retinal detachment and other complications were observed. In general, SH-HA hydrogel has great potential as a vitreous substitute.
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Diverse hydrogel formulations have been investigated for topical and subcutaneous drug delivery. Among the various biopolymers used for hydrogel formulations, hyaluronic acid (HA) has been widely applied due to its nontoxicity, excellent biocompatibility and rheological properties. However, formulations of HA-based hydrogel drug delivery system have several challenges to overcome, including their initial burst drug release and lack of mechanical strength. Thus, diverse strategies have been investigated to control the drug release from HA-based hydrogels. This review introduces recent strategies to modulate HA-based hydrogels for the control of drug release profiles. Recent approaches that will be covered in this review include, (1) both chemical and non-covalent cross-linking methods to modulate the physical properties of hydrogels, (2) ionic cross-lining and host–guest interaction-based hydrogels, and (3) incorporation of amorphous microprecipitated bulk drug powder and drug-encapsulated nano/microparticles in hydrogel networks. These new techniques are being intensively studied to decrease burst release and to sustain the release of drugs. HA-based hydrogel formulations are useful platforms in various biomedical areas, including controlled drug delivery, tissue engineering and cosmetic fillers. Various physical and chemical strategies are being studied to overcome their limitations and enhance the versatility of clinical applications.
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Microneedles are a rapidly developing method for the transdermal delivery of therapeutic compounds. All types of microneedles, whether solid, hollow, coated, or dissolving function by penetrating the stratum corneum layer of the skin producing a microchannel through which therapeutic agents may be delivered. To date, coated and hollow microneedles have been the most successful, despite suffering from issues such as poor drug loading capabilities and blocked pores. Dissolving microneedles, on the other hand, have superior drug loading as well as other positive attributes that make it an ideal delivery system, including simple methods of fabrication and disposal, and abundantly available materials. Indeed, dissolvable microneedles can even be fabricated entirely from the therapeutic agent itself thus eliminating the requirement for additional excipients. This focused review presents the recent developments and trends of dissolving microneedles as well as potential future directions. The advantages, and disadvantages of dissolving microneedles as well as fabrication materials and methods are discussed. The potential applications of dissolving microneedles as a drug delivery system in different therapeutic areas in both research literature and clinical trials is highlighted. Applications including the delivery of cosmetics, vaccine delivery, diagnosis and monitoring, cancer, pain and inflammation, diabetes, hair and scalp disorders and inflammatory skin diseases are presented. The current trends observed in the microneedle landscape with particular emphasis on contemporary clinical trials and commercial successes as well as barriers impeding microneedle development and commercialisation are also discussed.
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The skin serves as the major organ in the targeted transdermal drug delivery system for many compounds. The microneedle acts as a novel technique to deliver drugs across the different layers of the skin, including the major barrier stratum corneum, in an effective manner. A microneedle array patch comprises dozens to hundreds of micron-sized needles with numerous structures and advantages resulting from their special and smart designs. Microneedle approach is much more advanced than conventional transdermal delivery pathways due to several benefits like minimally invasive, painless, self-administrable, and enhanced patient compliance. The microneedles are classified into hollow, solid, coated, dissolving, and hydrogel. Several polymers are used to fabricate microneedle, such as natural, semi-synthetic, synthetic, biodegradable, and swellable polymers. Researchers in the preparation of microneedles also explored the combinations of polymers. The safety of the polymer used in microneedle is a crucial aspect to prevent toxicity in vivo. Thus, this review aims to provide a detailed review of microneedles and mainly focus on the various polymers used in the fabrication of microneedles.
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Hyaluronic acids (hyaluronans, HAs) are glycosaminoglycans produced in the bodies of Anguilliforme and Elopiforme leptocephali, and HA is thought to serve as a metabolic energy source during planktonic life-stages. To examine this hypothesis, we investigated the dynamics of HA during early growth of the Japanese eel (Anguilla japonica), including during metamorphosis. From histochemical observations in the fully grown leptocephalus, HA occupied approximately 40-50% of the cross-sectional area and muscle tissue occupied less than 20%. However, the HA and water content are at a maximum during leptocephalus, decreasing during metamorphosis. We found that during leptocephalus, HA is actively accumulated in the body and plays a role in specific density adjustment, facilitating planktonic life. It was thought that after metamorphosis the role of HA in facilitating floating would end, and it would be metabolized to glucose and/or triglyceride.
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Natural polysaccharides are promising candidates for the component of polymer materials for sustainable societies. Here, the preparation of composite films using high- and low-molecular weight (MW) of hyaluronic acid (H-HYA and L-HYA), carboxymethylated HYA (CM-HYA), and high- and low-MW of chitosan (H–CHI and L-CHI) was performed to clarify the effects of the polysaccharide species on formability, mechanical properties, and water-responsive properties of the resulting films. The films prepared using CM-HYA and H–CHI (CM-HYA/H–CHI films) exhibited the highest maximum stress due to more electrostatic interactions between COO⁻ in CM-HYA and NH3⁺ in H–CHI. Swelling ratio of CM-HYA/H–CHI films was the lowest both in water and phosphate buffer saline (PBS). Meanwhile, CM-HYA/H–CHI films exhibited the highest moisture absorption ratio and moisture retention capacities, indicating water absorption ability of CM-HYA could be maintained even in the film state. Accordingly, CM-HYA/H–CHI films may be advantageous for applications such as moisture-retaining sheets and biomaterials.
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Hyaluronic acid (HA) is a major component of the skin, contributing to tissue hydration and biomechanical properties. As HA content in the skin decreases with age, formulas containing HA are widely used in cosmetics and HA injections in aesthetic procedures to reduce the signs of aging. To prove the beneficial effects of these treatments, efficient quantification of HA levels in the skin is necessary, but remains difficult. A new analytical method has been developed based on matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) to quantify HA content in cross sections of human skin explants. A standardized and reproducible chemical entity (3 dimeric motifs or 6-mer) quantifiable by MALDI-MSI was produced by enzymatic hydrolysis using a specific hyaluronidase (H1136) in HA solution. This enzymatic digestion was carried out on skin sections before laser desorption, enabling the detection of HA. Histological coloration allowed us to localize the epidermis and the dermis on skin sections and, by comparison with the MALDI molecular image, to calculate the relative HA concentrations in these tissue areas. Skin explants were treated topically using a formula containing HA or its placebo, and the HA distribution profiles were compared with those obtained from untreated explants. A significant increase in HA was shown in each skin layer following topical application of the formula containing HA versus placebo and untreated samples (average of 126±40% and 92±40%, respectively). The MALDI-MSI technique enabled the quantification and localization of all HA macromolecules (endogenous and exogenous) on skin sections and could be useful for determining the efficacy of new cosmetic products designed to fight the signs of aging.
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The lack of vascularization in the white-red and white zone of the meniscus causes these zones of tissue to have low self-healing capacity in case of injury and accelerate osteoarthritis (OA). In this study, we have developed hybrid constructs using polycaprolactone (PCL) and decellularized meniscus extracellular matrix (DMECM) surface modified by gelatin (G), hyaluronic acid (HU) and selenium (Se) nanoparticles (PCL/DMECM/G/HU/Se), following by the cross-linking of the bio-polymeric surface. Material characterization has been performed on the fabricated scaffold using scanning electron microscopy (SEM), Fourier transforms infrared (FTIR) spectroscopy, swelling and degradation analyses, and mechanical tests. In Vitro, investigations have been conducted by C28/I2 human chondrocyte culture into the scaffold and evaluated the cytotoxicity and cell/scaffold interaction. For the in vivo study, the scaffolds were transplanted into the defect sites of female New Zealand white rabbits. Good regeneration was observed after two months. We have concluded that the designed PCL/DMECM/G/HU construct can be a promising candidate as a meniscus tissue engineering scaffold to facilitate healing.
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An in-depth understanding of the effect of physicochemical properties of nanocarriers on their cellular uptake and fate is crucial for the development of novel delivery systems. In this study, well-defined hydrophobic carboxylated poly(3-hydroxypropionate)-based comb polymers were synthesized. Two oligo(3-hydroxypropionate) (HPn) of different degrees of polymerization (DP; 5 and 9) bearing α-vinyl end-groups were obtained by an hydrogen transfer polymerization (HTP)-liquid/liquid extraction strategy. 2-Carboxyethyl acrylate (CEA), representing the DP 1 analogue of HPn, was also included in the study. (Macro)monomers were polymerized via reversible addition-fragmentation chain-transfer (RAFT) polymerization and fully characterized by 1H NMR spectroscopy and size exclusion chromatography. All polymers were non-hemolytic and non-cytotoxic against NIH/3T3 cells. Detailed cellular association and uptake studies of Cy5-labeled polymers by flow cytometry and confocal laser scanning microscopy (CLSM) revealed that the carboxylated water-soluble PCEA, the polymer with the shortest side chain, efficiently targets mitochondria. However, increasing the side-chain DP led to a change in the intracellular fate. P(HP5) was trafficked to both mitochondria and lysosomes, while P(HP9) was exclusively found in lysosomes. Importantly, FLIM-FRET investigation of P(HP5) provided initial insight into the mitochondria subcompartment location of Cy5-labeled carboxylated polymers. Moreover, intracellular uptake mechanism studies were performed. Blocking scavenger receptors by dextran sulfate or cooling cells to 4 °C significantly affected the cell association of hydrophobic carboxylated polymers with an insignificant response to membrane-potential inhibitors. In contrast, water-soluble carboxylated polymers' cellular association was substantially inhibited in cells treated with compounds depleting the mitochondrial potential (ΔΨ). Overall, this study highlights hydrophobicity as a valuable means to tune the cellular interaction of carboxylated polymers and thus will inform the design of future drug carriers based on Cy5-modified carboxylated polymers.
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Hyaluronan (HA) is a glycosaminoglycan constituent of extracellular matrix. In its native form HA exists as a high molecular weight polymer, but during inflammation lower molecular weight fragments accumulate. We have identified a collection of inflammatory genes induced in macrophages by HA fragments but not by high molecular weight HA. These include several members of the chemokine gene family: macrophage inflammatory protein-1alpha, macrophage inflammatory protein-1beta, cytokine responsive gene-2, monocyte chemoattractant protein-1, and regulated on activation, normal T cell expressed and secreted. HA fragments as small as hexamers are capable of inducing expression of these genes in a mouse alveolar macrophage cell line, and monoclonal antibody to the HA receptor CD44 completely blocks binding of fluorescein-labeled HA to these cells and significantly inhibits HA-induced gene expression. We also investigated the ability of HA fragments to induce chemokine gene expression in human alveolar macrophages from patients with idiopathic pulmonary fibrosis and found that interleukin-8 mRNA is markedly induced. These data support the hypothesis that HA fragments generated during inflammation induce the expression of macrophage genes which are important in the development and maintenance of the inflammatory response.
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CD44 is expressed in various isoforms on numerous cell types and tissues during embryogenesis and in the mature organism. CD44 may also be involved in tumor growth. To study the multiple roles of CD44, we abolished expression of all known isoforms of CD44 in mice by targeting exons encoding the invariant N-terminus region of the molecule. Surprisingly, mice were born in Mendelian ratio without any obvious developmental or neurological deficits. Hematological impairment was evidenced by altered tissue distribution of myeloid progenitors with increased levels of colony-forming unit-granulocyte-macrophage (CFU-GM) in bone marrow and reduced numbers of CFU-GM in spleen. Fetal liver colony-forming unit-spleen and granulocyte colony-stimulating factor mobilization assays, together with reduced CFU-GM in peripheral blood, suggested that progenitor egress from bone marrow was defective. In what was either a compensatory response to CD44 deficiency or an immunoregulatory defect, mice also developed exaggerated granuloma responses to Cryotosporidium parvum infection. Finally, tumor studies showed that SV40-transformed CD44-deficient fibroblasts were highly tumorigenic in nude mice, whereas reintroduction of CD44s expression into these fibroblasts resulted in a dramatic inhibition of tumor growth.
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The extracellular matrix glycosaminoglycan hyaluronan (HA) is an abundant component of skin and mesenchymal tissues where it facilitates cell migration during wound healing, inflammation, and embryonic morphogenesis. Both during normal tissue homeostasis and particularly after tissue injury, HA is mobilized from these sites through lymphatic vessels to the lymph nodes where it is degraded before entering the circulation for rapid uptake by the liver. Currently, however, the identities of HA binding molecules which control this pathway are unknown. Here we describe the first such molecule, LYVE-1, which we have identified as a major receptor for HA on the lymph vessel wall. The deduced amino acid sequence of LYVE-1 predicts a 322-residue type I integral membrane polypeptide 41% similar to the CD44 HA receptor with a 212-residue extracellular domain containing a single Link module the prototypic HA binding domain of the Link protein superfamily. Like CD44, the LYVE-1 molecule binds both soluble and immobilized HA. However, unlike CD44, the LYVE-1 molecule colocalizes with HA on the luminal face of the lymph vessel wall and is completely absent from blood vessels. Hence, LYVE-1 is the first lymph-specific HA receptor to be characterized and is a uniquely powerful marker for lymph vessels themselves.
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Cell adhesion molecules are considered to be pivotal elements required for proper embryo development. The transmembrane glycoprotein CD44, which is expressed in numerous splice variants on the surface of many different cell types and tissues, has been suggested to be involved in several physiological processes such as cell-cell interactions, signal transduction, and lymphocyte homing and trafficking during embryogenesis and in the adult organism. Some splice variants are thought to play an important role in tumor progression. To investigate the physiological roles of CD44 in vivo, we abolished expression of all isoforms of CD44 in mice by targeted insertion of a lacZ/neo cassette into the reading frame of the leader peptide. CD44-deficient mice are viable without obvious developmental defects and show no overt abnormalities as adults. However, CD44-deficient lymphocytes exhibit impaired entry into the adult thymus, although lymphocyte development is apparently unaltered. Our data indicate that all splice variants of CD44 are dispensable for embryonic development and implicate a critical function for CD44 in lymphocyte recirculation.
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We identified hyaluronan synthase-2 (Has2) as a likely source of hyaluronan (HA) during embryonic development, and we used gene targeting to study its function in vivo. Has2(-/-) embryos lack HA, exhibit severe cardiac and vascular abnormalities, and die during midgestation (E9.5-10). Heart explants from Has2(-/-) embryos lack the characteristic transformation of cardiac endothelial cells into mesenchyme, an essential developmental event that depends on receptor-mediated intracellular signaling. This defect is reproduced by expression of a dominant-negative Ras in wild-type heart explants, and is reversed in Has2(-/-) explants by gene rescue, by administering exogenous HA, or by expressing activated Ras. Conversely, transformation in Has2(-/-) explants mediated by exogenous HA is inhibited by dominant-negative Ras. Collectively, our results demonstrate the importance of HA in mammalian embryogenesis and the pivotal role of Has2 during mammalian development. They also reveal a previously unrecognized pathway for cell migration and invasion that is HA-dependent and involves Ras activation.
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Hyaluronan has been implicated in biological processes such as cell adhesion, migration and proliferation. Traditionally, it was thought to be associated with the extracellular matrix, but, hyaluronan may also have unimagined roles inside the cell. Investigation of hyaluronan synthesis and degradation, the identification of new receptors and binding proteins, and the elucidation of hyaluronan-dependent signaling pathways are providing novel insights into the true biological functions of this fascinating molecule.
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Current therapies for chronic inflammatory diseases typically act through the nonspecific downregulation of immune cell activation. However, it is becoming increasingly evident that parenchymal cells are also active participants in the inflammatory process. Future prospects for the treatment of inflammation should therefore include the targeting of specific inflammatory pathways in both immune cells and parenchymal cells. CD44, a cell-adhesion molecule that is ubiquitously expressed on leukocytes and parenchymal cells, has been implicated, together with its ligand hyaluronan (HA), in several inflammatory diseases. The mechanisms of action of CD44-HA interactions in inflammation might provide potential targets for therapy.
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Hyaluronan synthase 3 (HAS3) is responsible for the production of both secreted and cell-associated forms of hyaluronan and is the most active of the three isoforms of this enzyme in adults. In this study, the cDNA for human HAS3 was cloned and characterized. The open reading frame consisted of 1659 bp coding for 553 amino acids with a deduced molecular weight of about 63,000 and isoelectric pH of 8.70. The sequence of human HAS3 displayed a 53% identity to HAS1 and a 67% identity to HAS2. It also contained a signal peptide and six potential transmembrane domains, suggesting that it was associated with the plasma membrane. To evaluate the physiological role of human HAS3, expression vectors for this protein were transfected into TSU cells (a prostate cancer cell line), and the phenotypic changes in these cells were examined. The enhanced expression of hyaluronan in the transfected cells was demonstrated by dot blot analysis and ELISA. These cells were found to differ from their vector-transfected counterparts with respect to the following: (a) they grew at a faster rate in high (but not low) density cultures; (b) conditioned media from these cells stimulated the proliferation and migration of endothelial cells; (c) when placed on the chorioallantoic membrane of chicken embryos, these cells formed large, dispersed xenografts, whereas the control transfectants formed compact masses; and (d) when injected s.c. into nude mice, the xenografts formed by HAS3 transfectants were bigger than those formed by control transfectants. Histological examination of these xenografts revealed the presence of extracellular hyaluronan that could act as conduits for the diffusion of nutrients. In addition, they had a greater number of blood vessels. However, the HAS3-transfected TSU cells did not display increased metastatic properties as judged by their ability to form lung masses after i.v. injection. These results suggested that the HAS3-induced overexpression of hyaluronan enhanced tumor cell growth, extracellular matrix deposition, and angiogenesis but was not sufficient to induce metastatic behavior in TSU cells.
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A symptomatic relief by hyaluronic acid (HA, MW: 3.5 x 10(6)), which is synthesized by Streptococcus spp, was investigated in experimental ovine osteoarthrosis. Bilateral osteoarthrosis (OA) of the temporo-mandibular joints (TMJs) was induced by perforating discs and by scrapping subchondral condylar surface. HA was intra-articularly injected into the left joints of 6 sheep on 7, 10, 14, 17 and 21 days after the operation and physiological saline as the control was injected into the contralateral (right) joints on the same day. Three sheep were killed at I month post-operation (MPO) and the remaining three sheep were killed at 3 MPO. Various responses such as proliferation of fibrous tissue, denudation, erosion, osteophyte formation, subcortical cyst formation and ankylosis were observed radiographically and histopathologically. The treatment of HA ameliorated the degenerative changes and lowered the osteoarthrotic score in the left joints at I MPO (9.96 vs 5.81) and 3 MPO (10.86 vs 5.29) compared to the right joints. These results indicate that a repeated intra-articular injection of HA inhibits the progression of OA in ovine TMJs by inducing the development of articular cartilage and by reducing the proliferation of fibrotic tissue.
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Other articles in this series concentrate on normal physiological and cellular functions of hyaluronan. In this article we discuss the influences of hyaluronan on disease progression. Alterations in hyaluronan metabolism, distribution, and function have been documented in many diseases, e.g. arthritis, immune and inflammatory disorders, pulmonary and vascular diseases, and cancer (see Refs. 1 and 2). In this article we will concentrate on cancer and vascular disease because our knowledge in these areas has advanced rapidly over the past several years and because work in these areas has highlighted the importance of hyaluronan-cell interactions in cell behavior.
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Hyaluronidase is a hyaluronic acid-degrading endoglycosidase that is present in many toxins and the levels of which are elevated in cancer. Increased concentration of HYAL1-type hyaluronidase correlates with tumor progression and is a marker for grade (G) 2 or 3 bladder cancer. Using bladder tissues and cells, prostate cancer cells, and kidney tissues and performing reverse transcription-PCR, cDNA cloning, DNA sequencing, and in vitro translation, we identified splice variants of HYAL1 and HYAL3. HYAL1v1 variant lacks a 30-amino acid (aa) sequence (301-330) present in HYAL1 protein. HYAL1v1, HYAL1v2 (aa 183-435 present in HYAL1 wild type), HYAL1v3 (aa 1-207), HYAL1v4 (aa 260-435), and HYAL1v5 (aa 340-435) are enzymatically inactive and are expressed in normal tissues/cells and G1 bladder tumor tissues. However, HYAL1 wild type is expressed in G2/G3 tumors and in invasive tumor cells. Stable transfection and HYAL1v1-specific antibody confirmed that the HYAL1 sequence from aa 301 to 330 is critical for hyaluronidase activity. All tumor cells and tissues mainly express HYAL3 variants. HYAL3v1 lacks a 30-aa sequence (299-328) present in HYAL3 protein, that is homologous to the 30-aa HYAL1 sequence. HYAL3v1, HYAL3v2 (aa 251-417 present in HYAL3 wild type), and HYAL3v3 (aa 251-417, but lacking aa 299-328), are enzymatically inactive. Although splicing of a single independent exon generates HYAL1v1 and HYAL3v1, internal exon splicing generates the other HYAL1/HYAL3 variants. These results demonstrate that alternative mRNA splicing controls cellular expression of enzymatically active hyaluronidase and may explain the elevated hyaluronidase levels in bladder/prostate cancer.
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The medical literature was reviewed from 1968-2002 using Medline and the key words "intra-articular" and "osteoarthritis" to determine the various intra-articular therapies used in the treatment of osteoarthritis. Corticosteroids and hyaluronic acid are the most frequently used intra-articular therapies in osteoarthritis. Other intra-articular substances such as orgotein, radiation synovectomy, dextrose prolotherapy, silicone, saline lavage, saline injection without lavage, analgesic agents, non-steroidal anti-inflammatory drugs, glucosamine, somatostatin, sodium pentosan polysulfate, chloroquine, mucopolysaccharide polysulfuric acid ester, lactic acid solution, and thiotepa cytostatica have been investigated as potentially therapeutic in the treatment of arthritic joints. Despite the lack of strong, convincing, and reproducible evidence that any of the intra-articular therapies significantly alters the progression of osteoarthritis, corticosteroids and hyaluronic acid are widely used in patients who have failed other therapeutic modalities for lack of efficacy or toxicity. As a practical approach for a knee with effusion, steroid injections should be considered while the presence of symptomatic "dry" knees may favour the hyaluronic acid approach. The virtual absence of serious side effects, coupled with the perceived benefits, make these approaches attractive.
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Tissues must quickly recognize injury to respond to the rapid pace of microbial growth. In skin, dermal microvascular endothelial cells must also react to danger signals from the surrounding tissue and immediately participate by initiating the wound repair process. Components of the extracellular matrix such as hyaluronan are rapidly broken down into smaller molecular weight oligosaccharides in a wound, and these can activate a variety of biological processes. This study set out to determine if hyaluronan fragments released following injury can stimulate endothelial cells and what mechanism is responsible for this response. Using genechip microarray analysis, a response to hyaluronan fragments was detected in endothelial cells with the most significant increase observed for the chemokine IL-8. This observation was verified with qualitative reverse transcriptase-PCR and ELISA in human endothelial cell culture, and in a mouse model by observing serum levels of MIP-2 and KC following hyaluronan fragment administration in vivo. Activation was TLR4-dependent, as shown by use of TLR4 blocking antibody and TLR4-deficient mice, but not due to the presence of undetected contaminants as shown by inactivation following digestion with the hyaluronan-degrading enzyme chondroitinase ABC or incubation with the hyaluronan-specific blocking peptide Pep-1. Inactivation of LPS activity failed to diminish the action of hyaluronan fragments. These observations suggest that endogenous components of the extracellular matrix can stimulate endothelia to trigger recognition of injury in the initial stages of the wound defense and repair response.
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A variety of obstacles have hindered the ultrastructural localization of hyaluronan (HA). These include a lack of adequate fixation techniques to prevent the loss of HA, the lack of highly sensitive and specific probes, and a lack of accessibility due to the masking of HA by HA-binding macromolecules such as proteoglycans and glycoproteins. Despite these problems, a number of studies, both biochemical and histochemical, have been published indicating that HA is not restricted to the extracellular milieu, but is also present intracellularly. This review focuses on the possible functions of intracellular HA, its potential relationships to extracellular HA structures, and implications for inflammatory processes.
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The availability of elastoviscous solutions of highly purified hyaluronan has created two new therapeutic methods in human and veterinary medicine: viscosurgery and viscosupplementation. Viscosurgical tools and implants are widely used in ophthalmology and have been suggested for use in otology. Visco-supplementation of joint fluid using elastoviscous hyaluronan solutions is widely used in the treatment of equine traumatic arthritis. It was also suggested for use in idiopathic osteoarthritis in humans, but this application has not received wide acceptance. Cross-linked forms of hyaluronan have been developed and given the generic name of hylans. Water-insoluble soft gels of hylans are ideally suitable as viscosurgical implants to prevent postoperative adhesions and to control scar formation. Hylan solutions are being used in arthroscopic viscosurgery. Hylan devices in various forms (gels, tubes, membranes) have been used in animal studies for matrix engineering, the purpose of which is to control and direct tissue regeneration and augmentation.
Article
Toole BP (Tufts University School of Medicine, Boston, MA, USA). Hyaluronan in morphogenesis (Minisymposium: Hyaluronan). J Intern Med 1997; 242: 35–40. Pericellular matrices surrounding migrating and proliferating cells in the developing embryo, in regenerating tissues and in other dynamic cellular events, such as tumour cell invasion, are enriched in hyaluronan. In addition to contributing to the unique structure of the pericellular matrix, hyaluronan interacts with cell surface receptors, such as RHAMM and CD44. During morphogenesis, these interactions of hyaluronan with the cell surface are important in several ways. First, hyaluronan–CD44 interactions have been shown to mediate endocytic removal of hyaluronan at critical stages of embryonic development. Secondly, hyaluronan provides an appropriately hydrated, pericellular milieu that facilitates cellular invasion. Thirdly, in-vitro studies suggest strongly that interactions of hyaluronan with RHAMM or CD44 are involved in cell movement and proliferation, which are critical events in morphogenesis.
Article
Hyaluronan, a negatively charged, high-molecular weight polysaccharide is found in all tissues and body fluids of higher animals and most abundantly in soft connective tissues. The total amount in an adult human has been estimated to be 11–17 g. Its turnover has earlier been studied by following the fate of endogenously labelled polysaccharide or injected highly concentrated unlabelled polymer. More recently, hyaluronan biosynthetically labelled with 14C or 3H has been injected into various compartments or tissues and its disappearance followed. These experiments have demonstrated a fairly rapid turnover in many localities ( in the order of one day). This is partly due to lymphatic removal of hyaluronan from the tissues and subsequent degradation in lymph nodes and liver. Recent work with [125I]tyramine cellobiose-labelled hyaluronan has furthermore shown that the polysaccharide can also be endocytosed and degraded locally in the tissues. Thus the catabolism of hyaluronan takes place both by local degradation and drainage via the lymphatic system.
Article
Circulating hyaluronan (or hyaluronic acid or hyaluronate) is efficiently sequestered by receptor-mediated endocytosis mainly in sinusoidal liver endothelial cells (LECs). The receptor recognizes hyaluronan, chondroitin sulfate and dermatan sulfate, but not other ligands. Binding affinity increases with increasing chain length of the polysaccharide: Kd varies between 4.6·10−6 for an octassacharide and 9·10−12 for hyaluronan of Mr 6.4·106. Ligands bound by the receptor are associated with coated pits. Internalization takes less than 1 min, after which the ligand appears in small coated vesicles and shortly thereafter in small uncoated vesicles. The ligand resides in larger macropinocytic vesicles before being transferred to lysosomes. Lysosomal degradation of hyaluronan yields glucuronic acid and f-acetylglucosamine of which the latter is deacetylated by N-acetylglucosamine-6-phosphate deacetylase. The remaining pyranose rings are degraded to lactate. Both lactate and acetate are released from LECs and oxidized by the hepatocytes to CO2 and water.
Article
The normal systemic kinetics of hyaluronan (HYA) is well established in several species including man. The removal of HYA from the circulation is very efficient, with a half-life of 2–6 min and a total normal turnover of 10–100 mg/day in the adult human. The main uptake from the blood takes place in the liver endothelial cells. Evidence for a role of the kidney in the elimination of HYA is, however, accumulating. Recently published data suggest that the elimination kinetics of HYA from the systemic circulation may be influenced by a number of factors, such as saturation of the elimination caused by an increased lymphatic input of HYA to the circulation, alteration of the blood flow over the eliminating organ and competition with other macromolecular substances such as chondroitin sulphate or proteoglycans. Many of these factors may be operative during different disease states, and may therefore partly explain the observed differences between normal and pathological HYA kinetics.
Article
The highest concentrations of hyaluronan occur in synovial fluid, vitreous body, skin and certain specialized tissues such as umbilical cord and rooster comb, during fetal development, and in tissue repair and regeneration. The largest amounts are found in the intercellular matrix of skin and musculoskeletal tissues. Turnover in the bloodstream is normally in the range of 0.3-1.0 microgram min-1/kg body weight. Circulating hyaluronan is mostly derived from lymph. Lymph nodes may nevertheless extract as much as 80-90% from peripheral lymph before it can reach the bloodstream. Turnover in peripheral tissues may be effected by degradation in situ, or by transfer into lymph by diffusion or hydrodynamic forces. Hyaluronan is firmly bound in specific association with cells or binding proteins but much of it exists in freely mobilized compartments with a half-life of two days or less, and it is metabolized after transport elsewhere. Metabolic degradation of hyaluronan is principally intracellular and relies on uptake by a receptor which, in contrast with other hyaluronan-binding structures, also binds chondroitin sulphate. It is suggested that this dual specificity may be primarily associated with metabolic degradation of hyaluronan. Uptake and metabolism are primarily effected in liver and lymph node by endothelial cells lining the sinusoids of each. Further studies indicate that in lymph nodes and in spleen, macrophage-like cells intertwined with the endothelial cells also take up hyaluronan. The metabolic cycle from polymer to monosaccharides, acetate and beyond can be completed in vivo within 10 minutes.
Article
The availability of elastoviscous solutions of highly purified hyaluronan has created two new therapeutic methods in human and veterinary medicine: viscosurgery and viscosupplementation. Viscosurgical tools and implants are widely used in ophthalmology and have been suggested for use in otology. Visco-supplementation of joint fluid using elastoviscous hyaluronan solutions is widely used in the treatment of equine traumatic arthritis. It was also suggested for use in idiopathic osteoarthritis in humans, but this application has not received wide acceptance. Cross-linked forms of hyaluronan have been developed and given the generic name of hylans. Water-insoluble soft gels of hylans are ideally suitable as viscosurgical implants to prevent postoperative adhesions and to control scar formation. Hylan solutions are being used in arthroscopic viscosurgery. Hylan devices in various forms (gels, tubes, membranes) have been used in animal studies for matrix engineering, the purpose of which is to control and direct tissue regeneration and augmentation.
Article
Hepatic and renal clearance of the amino-terminal propeptide of type III procollagen (PIIINP) and of the glycosaminoglycan, hyaluronan (HA) were investigated in a catheterization study of seven healthy anesthetized pigs. Two assays were used, in order to distinguish between the metabolism of different PIIINP-related antigens. One was the PIIINP RIA Kit, which measures the intact propeptide. The other was the PIIINP Fab assay, in which the antibody has an equal affinity to the intact propeptide and to smaller fragments, of which the latter constitutes most of the antigenic activity in serum. Hepatic and gastrointestinal extraction were evaluated from measurements of serum concentrations in the artery, the portal vein and the hepatic vein. We found a significant hepatic extraction of the intact propeptide (extraction ratio 0.14) and of HA (extraction ratio 0.23), but not of smaller PIIINP fragments. No gastrointestinal extraction of any of the tested substances could be demonstrated. Only smaller PIIINP fragments (such as the col 1 fragment) were extracted by the kidneys (the extraction ratio in the PIIINP Fab assay was 0.19). The renal extraction ratio of HA was 0.14. The amounts of PIIINP fragments and of HA extracted by the kidneys were 50- and 3-times the amounts found in urine, respectively, indicating that the col 1 fragment and HA are degraded in the kidneys in addition to urinary excretion. Our results suggest a dynamic turnover of connective tissue-related components with a fast catabolism of circulating components in liver and kidneys.
Article
The plasma clearance, tissue distribution and metabolism of hyaluronic acid were studied with a high average molecular weight [3H]acetyl-labelled hyaluronic acid synthesized in synovial cell cultures. After intravenous injection in the rabbit the label disappeared from the plasma with a half-life of 2.5--4.5 min, which corresponds to a normal hyaluronic acid clearance of approx. 10 mg/day per kg body weight. Injection of unlabelled hyaluronic acid 15 min after the tracer failed to reverse its absorption. Clearance of labelled polymer was retarded by prior injection of excess unlabelled hyaluronic acid. The maximum clearance capacity was estimated in these circumstances to be about 30 mg/day per kg body wt. The injected material was concentrated in the liver and spleen. As much as 88% of the label was absorbed by the liver, where it was found almost entirely in non-parenchymal cells. Degradation was rapid and complete, since volatile material, presumably 3H2O, appeared in the plasma within 20 min. Undegraded [3H]hyaluronic acid, small labelled residues and 3H2O were detected in the liver, but there was little evidence of intermediate oligosaccharides. No metabolite except 3H2O was recognized in plasma or urine. Two-thirds of the radioactivity was retained in the body water 24 h later, and small amounts were found in liver lipids. Radioactivity did not decline in the spleen as rapidly as in the liver. The upper molecular weight limit for renal excretion was about 25 000. Renal excretion played a negligible part in clearance. It is concluded that hyaluronic acid is removed from the plasma and degraded quickly by an efficient extrarenal system with a high reserve capacity, sited mainly in the liver.
Article
Previous investigations suggest that the corneal endothelium has specific binding sites for hyaluronan (HYA). In the present study, biochemical and immunological techniques were used to characterize these binding sites and to compare them with the liver endothelial cell (LEC) HYA receptor. Affinity chromatography of solubilised, 125I-labelled corneal endothelial cell surface proteins on immobilised HYA proved that there were molecules that were strongly bound to the polysaccharide. A part of these molecules formed a 100-kDa band when analysed by autoradiography after SDS polyacrylamide electrophoresis (PAGE). A specific antibody against the rat LEC HYA receptor was used for immunohistochemical studies of monkey and human corneas. There was a specific staining of the corneal endothelium of both species, and hyaluronan treatment before isolation of the human eyes reduced the staining intensity. Hyaluronidase treatment of the tissue sections before receptor staining strikingly increased the specific staining of the corneal endothelial cells (CEC). Immunoblotting of human corneal proteins, separated by SDS-PAGE, showed staining at 200, 150-160 and 55 kDa. Uptake experiments of tritiated HYA in cultured monkey CEC showed only a slight increase in cell associated radioactivity over 2-6 hr. The results make it unlikely that the corneal endothelial receptor, like its liver endothelial counterpart, is actively involved in receptor-mediated endocytosis. Our studies suggest that CEC carry receptors for HYA that are immunologically similar to the LEC receptors. CEC receptors might act as binding structures increasing the concentration of HYA close to the CEC as a protection of these vulnerable cells from physicochemical damage.
Article
Hyaluronan is an important constituent of the extracellular matrix. This polysaccharide can be hydrolyzed by various hyaluronidases that are widely distributed in nature. The structure of some bacterial and animal enzymes of this type has recently been elucidated. It could be shown that the hyaluronidases from bee and hornet venom and the PH-20 hyaluronidase present on mammalian spermatozoa are homologous proteins.
Article
The purpose of this study was to test the effect of repeated injections of hyaluronic acid (HA) on the sheep model of osteoarthrotic temporomandibular joint (TMJ) disease. Bilateral osteoarthrosis (OA) was induced in the TMJs of six sheep. HA was injected into one joint on 7, 10, 14, 17, and 21 days postoperatively. Normal saline was injected into the contralateral joint. Three sheep were killed at 1 month and 3 at 3 months. The joints were removed and examined macroscopically and histologically. A special scoring system was applied following the modified Mankin's score to evaluate the histologic changes. The control group showed severe osteoarthrotic changes in the condyle, deviation in form from normal morphology, and marked marrow fibrosis. The HA-treated group showed less deviation from normal condylar morphology. The histologic scores at 1 month were HA 12.6, control 24.2 (P < .001), and at 3 months were HA 6.9, control 18.9 (P < .001). There was a significant difference in osteoarthrotic changes between HA-treated and control TMJs, with the HA-treated TMJs having less severe changes. Repeated intraarticular injections of HA into a sheep TMJ with experimentally induced OA minimizes the extent of osteoarthrotic change when compared with the control joint. Thus, HA may have a role in preventing the progression of TMJ OA.
Article
CD44 is the main receptor for the extracellular polysaccharide hyaluronan (HA). We have recently shown that CD44 is strongly induced on renal tubular epithelial cells (TEC) in autoimmune renal injury and that HA accumulates in the renal interstitium (Kidney Int 1996; 50: 156-163 and Nephrol Dial Transplant 1997; 12: 1344-1353). The functional significance of enhanced tubular CD44 expression and its interaction with HA are not known. The purpose of the present study was to characterize renal tubular CD44 expression and CD44-mediated HA binding in vitro and to investigate the growth modulating effects in response to HA binding by TEC. RT-PCR analysis, flow cytometry, confocal microscopy and Western blotting were used to examine cell surface and soluble CD44 expression by cultured TEC, using SV40-transformed mouse cortical tubular (MCT) cells. HA binding characteristics were examined by flow cytometry and effects of HA on TEC cell growth by [3H]thymidine incorporation. By RT-PCR analysis MCT cells expressed predominantly the standard form of CD44 mRNA, whereas the expression of variant forms was very weak. Confocal microscopy showed that CD44 was expressed basolaterally and apically on MCT cells with strong staining on microvilli. Shedding of CD44 from MCT cells could be induced with crosslinking of anti-CD44 mAbs or with PMA stimulation. MCT cells constitutively bound HA and this binding could be modulated with anti-CD44 mAbs. Soluble and plate-bound HA markedly inhibited MCT cell growth. CD44 is a regulated HA receptor on MCT cells which can be shed into the cellular environment. Upon binding of HA, CD44 functions as a growth inhibitory cell surface protein in MCT cells. We speculate that the interaction of CD44 with HA may have important regulatory effects on cell proliferation in tubulointerstitial renal diseases.
Article
Hyaluronan (HA) has long been implicated in malignant transformation and tumor progression. However, due to the lack of molecular tools to directly manipulate production of HA, which does not require a core protein for its synthesis, our understanding of the role of HA in tumor cells has been largely circumstantial. In this study, we genetically manipulated the production of HA by transfection of a mammalian HA synthase Has2 into human HT1080 cells and examined the malignant phenotype of transfected cells. We found that increased production of HA promotes anchorage-independent growth and tumorigenicity of the cells. Has2-transfected cells formed greater numbers of colonies in semisolid medium. Tumors in nude mice derived from Has2-transfected cells grew more rapidly and were 2-4 times larger than those derived from control cells at termination of experiments. Histological and biochemical analyses of tumors revealed no significant differences in cell density and tissue structures between them, indicating that the larger size of the tumors was due to enhanced cell proliferation, not to increased accumulation of tumor stroma or increased angiogenesis. These results demonstrate that HA production by tumor cells per se promotes proliferation of these cells in tissues and provides direct evidence for the role of HA in tumorigenicity.
Article
Hyaluronan is a major carbohydrate component of the extracellular matrix and can be found in skin, joints, eyes and most other organs and tissues. It has a simple, repeated disaccharide linear copolymer structure that is completely conserved throughout a large span of the evolutionary tree, indicating a fundamental biological importance. Amongst extracellular matrix molecules, it has unique hygroscopic, rheological and viscoelastic properties. Hyaluronan binds to many other extracellular matrix molecules, binds specifically to cell bodies through cell surface receptors, and has a unique mode of synthesis in which the molecule is extruded immediately into the extracellular space upon formation. Through its complex interactions with matrix components and cells, hyaluronan has multifaceted roles in biology utilizing both its physicochemical and biological properties. These biological roles range from a purely structural function in the extracellular matrix to developmental regulation through effects of cellular behavior via control of the tissue macro- and microenvironments, as well as through direct receptor mediated effects on gene expression. Hyaluronan is also thought to have important biological roles in skin wound healing, by virtue of its presence in high amounts in skin. Hyaluronan content in skin is further elevated transiently in granulation tissue during the wound healing process. In this review, the general physicochemical and biological properties of hyaluronan, and how these properties may be utilized in the various processes of wound healing: inflammation, granulation and reepithelization, are presented.
Article
Hyaluronic acid (HA) is an important constituent of the extracellular matrix; its bacterial degradation has been postulated to contribute to the spread of certain streptococci through tissue. Pneumococci and other streptococci produce hyaluronate lyase, an enzyme which depolymerizes HA, thus hyaluronate lyase might contribute directly to bacterial invasion. Although two different mechanisms for lyase action have been proposed, there was no crystallographic evidence to support those mechanisms. Here, we report the high-resolution crystal structure of Streptococcus pneumoniae hyaluronate lyase in the presence of HA disaccharide product, which ultimately provides the first crystallographic evidence for the binding of HA to hyaluronate lyase. This structural complex revealed a key interaction between the Streptococcus peneumoniae hyaluronate lyase protein and the product, and supports our previously proposed novel catalytic mechanism for HA degradation based on the native Streptococcus peneumoniae hyaluronate lyase structure. The information provided by this complex structure will likely be useful in the development of antimicrobial pharmaceutical agents.
Article
The interactions of hyaluronan (HA) with proteins are important in extracellular matrix integrity and leukocyte migration and are usually mediated by a domain termed a Link module. Although the tertiary structure of a Link module has been determined, the molecular basis of HA-protein interactions remains poorly understood. Isothermal titration calorimetry was used to characterize the interaction of the Link module from human TSG-6 (Link_TSG6) with HA oligosaccharides of defined length (HA(4)-HA(16)). All oligomers bound (except HA(4)) with K(d) values ranging from 0.2-0.5 microM at 25 degrees C. The reaction is exothermic with a favourable entropy and the thermodynamic profile is similar to those of other glycosaminoglycan-protein interactions. The HA(8) recognition site on Link_TSG6 was localized by comparing nuclear magnetic resonance (NMR) spectra from a 1:1 complex with free protein. Residues perturbed on HA binding include both amino acids that are likely to be directly involved in the interaction (i.e., Lys11, Tyr59, Asn67, Phe70, Lys72 and Tyr78) and those affected by a ligand-induced conformational change in the beta4/beta5 loop. The sidechain of Asn67 becomes more rigid in the complex suggesting that it is in close proximity to the binding site. In TSG-6 a single Link module is sufficient for a high-affinity interaction with HA. The HA-binding surface on Link_TSG6 is found in a similar position to that suggested previously for CD44, indicating that its location might be conserved across the Link module superfamily. Here we find no evidence for the involvement of linear sequence motifs in HA binding.
Article
Mutual interaction between the metanephric mesenchyme (MM) and the ureteric bud (UB) in the developing kidney leads to branching morphogenesis and the formation of the ureteric tree. A UB-derived cell line, stimulated by conditioned medium derived from an embryonic MM cell line (or, similarly, by 10% fetal calf serum), forms branching tubules under three-dimensional culture conditions (H. Sakurai et al., 1997, Proc. Natl. Acad. Sci. USA 94, 6279-6284). The formation of branching tubules in this simple in vitro system for early nephrogenesis is highly sensitive to the matrix environment, a key component of which is the glycosaminoglycan hyaluronan (HA). Consistent with this, we found that HA in the extracellular environment markedly stimulated the formation of cellular processes and multicellular cords (early steps in branching morphogenesis) and also acted as a cell survival factor. Inhibition of HA binding to the cells by addition of blocking antibodies to CD44, the principal cell surface receptor for HA, or degradation of HA by the addition of Streptomyces hyaluronidase resulted in decreased cell survival and diminished morphogenesis, indicating that the HA-CD44 axis plays a central role in in vitro branching morphogenesis. Analysis of the expression of a large number of genes displayed on a cDNA array revealed that significant changes in gene expression in cells undergoing morphogenesis in the presence of HA were limited to a small subset of genes regulating apoptosis, proliferation, and morphogenesis. This included upregulation by HA of its receptor, CD44, which was found to largely localize to the tips of branching cellular processes. In the embryonic kidney, HA was found near the developing ureteric tree and CD44 was expressed basolaterally in UB-derived structures. In addition, both UB and MM appear to express HA synthase, suggesting their ability to secrete HA. We propose that HA promotes branching morphogenesis by creating a positive feedback loop that results in (1) enhanced interaction of HA-CD44 at branching tips (possibly leading to localization of HA binding morphoregulatory factors at the tips) and (2) an activated transcriptional program favoring cell survival/proliferation and migration/morphogenesis of cells through matrix by the expression of key morphoregulatory molecules. Furthermore, since HA, hyaluronidase, and CD44 have been functionally implicated in branching morphogenesis in this model, and since HA, CD44, and HA synthase are all expressed in an appropriate spatiotemporal fashion in the developing kidney, we propose that these molecules may, together, constitute a morphoregulatory pathway that plays a key role in sequential cycles of branching morphogenesis in the UB.