Benzyl Ester Of Hyaluronic Acid Research Articles

Synergy of Hydeal-D® and Hyaluronic Acid for Protecting and Restoring Urothelium: In Vitro Characterization.

Interstitial cystitis (IC) or painful bladder syndrome is a chronic dysfunction due to an inflammatory condition, characterized by bladder pain and urinary frequency. Currently, no gold standard therapy is available since IC does not respond to conventional ones. Given these premises, the aim of this work was the in vitro characterization of biological properties (mucoadhesion and anti-inflammatory activity) of a commercial product (HydealCyst–HydC) based on hyaluronic acid (HA) and the benzyl ester of HA (Hydeal-D®) intended for bladder instillation to restore and/or protect the urothelial layer of glycosamino glycans (GAGs). The in vitro characterization demonstrated that an interaction product is formed between HA and Hydeal-D® that has a role in the rheological behavior and mucoadhesive properties. HA was identified as a key component to form the mucoadhesive joint, while the interaction of HA with Hydeal-D® improved polysaccharide stability and prolonged the activity ex vivo. Moreover, HydC is cytocompatible with urothelial cells (HTB-4) and possesses an anti-inflammatory effect towards these cells by decreasing the secretion of IL-6 and IL-8, which were both increased in patients with IC, and by increasing the secretion of sulfated GAGs. These two findings, along with the resilience properties of the formulation due to mucoadhesion, suggest the active role of HydC in protecting and restoring urothelium homeostasis.

Surgical treatment of the neglected achilles tendon rupture with Hyalonect.

The purpose of this study was to report the management and outcomes of ten patients with chronic Achilles tendon rupture treated with a turndown gastrocnemius-soleus fascial flap wrapped with a surgical mesh (Hyalonect). Ten men with neglected Achilles tendon rupture were treated with a centrally based turndown gastrocnemius fascial flap wrapped with Hyalonect. Hyalonect is a knitted mesh composed of HYAFF, a benzyl ester of hyaluronic acid. The Achilles tendon ruptures were diagnosed more than 1 month after injury. The mean patient age was 41 years. All of the patients had weakness of active plantarflexion. The mean preoperative American Orthopaedic Foot and Ankle Society score was 64.8. The functional outcome was excellent. The mean American Orthopaedic Foot and Ankle Society score was 97.8 at the latest follow-up. There were significant differences between the preoperative and postoperative scores. Ankle range of motion was similar in both ankles. Neither rerupture nor major complication, particularly of wound healing, was observed. For patients with chronic Achilles tendon rupture with a rupture gap of at least 5 cm, surgical repair using a single turndown fascial flap covered with Hyalonect achieved excellent outcomes.

Allogeneic epidermal substitutes in the treatment of chronic diabetic leg and foot ulcers

Aim: Diabetic foot ulcers are the most common cause of nontraumatic lower extremity amputations in the industrialized world. Tissue-engineering products offer a lower extremity salvage strategy when healing does not proceed according to the standard of care. New allogeneic sheets are available for the management of diabetic leg and foot ulcers. Methods: The endpoints of this case series study regard preliminary outcomes of the application of allogeneic keratinocytes composed of benzyl ester of hyaluronic acid to 16 diabetic foot and leg ulcers in 11 patients with type 2 diabetes mellitus. Results: Between 21 and 70 days after cellular therapy, 6 out of 16 lesions were completely healed, reducing the wound dimension by 70% and improving the wound bed score by 52%. Conclusion: The clinical results of the new allogeneic sheets indicate that allogeneic keratinocytes may represent an effective and safe therapy for diabetic foot and leg ulcers in the multidisciplinary approach to this diabetes-related complication.

Hyalonect in the treatment of pseudarthrosis

The aim of this study was to evaluate union rates and complications of Hyalonect, a knitted mesh composed of HYAFF, a benzyl ester of hyaluronic acid, and a naturally occurring constituent of the extracellular matrix, for the treatment of pseudarthrosis. The study included 11 patients (8 male, 3 female; mean age: 44.6 years; range: 23 to 57 years) operated for pseudarthrosis using Hyalonect. Average time between initial treatment and surgical procedure was 12.9 (range: 8 to 48) months. Pseudarthrosis of the tibia, femur and humerus was present in 4, 2 and 5 patients, respectively. All patients had undergone prior surgery (1 to 6 times). Each patient underwent open reduction and internal fixation. Allograft was applied to the pseudarthrosis area and covered with Hyalonect. Mean follow up period was 31 (range: 12 to 48) months. Union was achieved in all patients after an average of 6 (range: 4 to 8) months. One patient had a discharge for two weeks. Another developed an infection which responded well to appropriate antibiotic treatment. No malunion or implant failure was observed. One patient with pre-existing radial nerve palsy maintained the condition. Use of Hyalonect appears to be a safe method with a positive impact on union in the surgical treatment of pseudarthrosis, particularly in the absence of any infection.

Interaction between a Regenerative Matrix and Wound Bed in Nonhealing Ulcers: Results with 16 Cases

A chronic wound is a wound that is delayed in one of the wound-healing stages and cannot progress any further. A chronic wound leaves the patient at risk of infection and hospitalization. In these case series, 16 patients affected by venous ulcers underwent Hyalomatrix PA grafting for reconstructive surgery. Hyalomatrix PA is a bilayered, sterile, flexible, and conformable three-dimensional matrix made of fibers of HYAFF, a benzyl ester of hyaluronic acid, and a semipermeable silicone membrane. Hyalomatrix PA acts as a substitutive and regenerative permanent matrix able to replace the dermis providing a three-dimensional matrix for cellular invasion and capillary growth. The silicon layer controls water vapor loss avoiding an excessive loss of fluids and acts as a semipermeable barrier to the external agents. In the presented cases, the average area grafted per procedure was 153 cm2. The length of followup ranged from 0.5 to 1 year. The final results were considered to be good in 12 cases, fair in 3 cases, and poor in one case. This study suggests that the combination of wound bed preparation with application of the hyaluronic regenerative matrix can be a valid approach for treatment of partial thickness ulcers.

Morphology and degradation properties of PCL/HYAFF11® composite scaffolds with multi-scale degradation rate

The analysis of scaffold degradation is a promising strategy for understanding the dynamic changes in texture and pore morphology which accompany polymer resorption, and for collecting same fundamental indicators regarding the potential fate of the scaffold in the biological environment. In this study, we investigate the morphology and degradation properties of three composite scaffolds based on poly(ε-caprolactone) (PCL) embedded with benzyl ester of hyaluronic acid (HYAFF11®) phases, and, in turn, different reinforcement systems – i.e., calcium phosphate particles or continuous poly(lactic acid) (PLA) fibres. Scanning electron microscopy (SEM) and μ-tomography supported by digital image analysis enabled a not invasive investigation of the scaffold morphology, providing a quantitative assessment of porosity (which ranged from 63.1 to 82.8), pore sizes (which varied from 170.5 to 230.4μm) and pore interconnectivity. Thermal analyses (DSC and TGA) and Raman spectroscopy demonstrated the multi-scale degradation of the composite with highly tailoring degradation kinetics depending on the component material phases and scaffold architecture changes, due to their conditioning in simulated in vivo environment (i.e., SBF solution). These results demonstrate that the judicious mixing of materials with faster (i.e., HYAFF11) and slower (i.e., PLA and PCL) degradation kinetics, different size and shape (i.e., domains, particles or long fibres), certainly concurs to design a smart composite scaffold with time-controlled degradation which can support the regeneration of a large variety of tissues, from the cartilage to the bone.

Growth of keratinocytes on porous films of poly(3‐hydroxybutyrate) and poly(4‐hydroxybutyrate) blended with hyaluronic acid and chitosan

The objective of this study was to develop novel absorbable films suitable for use as a tissue-engineering scaffold for keratinocytes as a therapy for replacement of damaged skin. Poly(4-hydroxybutyrate) (P(4HB)) and poly (3-hydroxybutyrate) (P(3HB)) were blended with small amounts of the polysaccharides hyaluronic acid (HA), chitosan (CH), pectin and alginic acid, and were solution cast to produce porous films. The resulting composites had favorable mechanical properties, and these films were compared with two commercially available implantable films made of poly(L-lactide-co-D,L-lactide) (PLA copolymer) and HA benzyl ester. Tensile testing demonstrated that a high level of flexibility of P(4HB) was retained in the P(4HB)-polysaccharide composite films, whereas the P(3HB) film and its polysaccharide composites were stiffer and more brittle. The proliferation kinetics of adherent HaCaT keratinocytes on the films was examined in vitro. The porous surface of the P(4HB) and P(3HB) films blended with HA or CH promoted the growth of keratinocytes significantly. The order of maximum cell numbers on these films was P(4HB)/HA > P(4HB)/CH > P(3HB)/HA > P(3HB)/CH > P(3HB)/pectin > P(3HB)/alginic acid. Scanning electron microscopy and confocal laser scanning microscopy revealed differences in cell growth. Cells formed clusters on P(3HB) and its composites, while the cells grew as a confluent layer on P(4HB) and its composites. HaCaT cells formed large numbers of filaments only on P(4HB) films, indicating the excellent biocompatibility of this material. For the nonporous PHB films, the proliferation rate of cells was found to increase with decreasing hydrophobicity in the order: P(4HB) > P(3HB)/P(4HB) blend > P(3HB).

Chemical-Physical Characterization and in vitro Preliminary Biological Assessment of Hyaluronic Acid Benzyl Ester-Hydroxyapatite Composite

HYAFF11 is a biocompatible, biodegradable benzyl ester of hyaluronic acid. However, in order to use it for orthopedic application, its mechanical performance needs to be improved. In this study, a novel composite based on HYAFF11 polymer matrix reinforced with hydroxylapatite (HA) has been developed. Its advantage is having a similar component of the mineral phase of bone resulting in favorable osteoconductive properties. The present study has examined the compressive mechanical and surface chemical-physical properties of the novel HYAFF11-HA composite. Preliminary biological investigations, including pH and cytotoxicity studies of the material extracts, have also been performed using an in vitro primary human osteoblast-like cell model. Moreover, protein, especially fibronectin adsorption has been investigated following incubation in culture medium and human plasma. The results show a grainy surface topography composed mainly of C, P, and Ca, with a Ca/P atomic ratio indicating HA on the composite surface. Mechanical analysis shows an improvement of the compressive properties of HYAFF11 matrix, both in the dry and swollen states, with values in the range of that of spongy bone. No cytotoxic effects and no inhibition of cell proliferation have been observed in the presence of the material extracts with pH values within acceptable ranges for cell vitality. Protein studies reveal a similar pattern, but a higher amount of fibronectin following incubation in human plasma when compared with culture medium. The results show that the novel HYAFF11-HA composite shows a great potential for application in orthopedic fields, especially as vertebral trabecular bone substitute.

Autologous human keratinocytes cultured on membranes composed of benzyl ester of hyaluronic acid for grafting in nonhealing diabetic foot lesions: A pilot study

Diabetic foot complications are the most common cause of nontraumatic lower extremity amputations in the industrialised world. Unsatisfactory healing requires advanced therapeutic strategies, such as the use of skin grafts, which may represent a helpful option for wound coverage. Alternatively, a method using autologous keratinocytes grown to thin sheet grafts is available. The purpose of this pilot study was to investigate the application of autologous human keratinocytes cultured on membranes composed of benzyl ester of hyaluronic acid (Laserskin autograft) to diabetic foot ulcers. We studied 14 patients with type 2 diabetes mellitus and a nonhealing diabetic foot lesion, defined as existing longer than 6 months or with no wound healing apparent for 12 weeks. Between 7 and 64 days after the transplantation (depending on the size of the ulceration), 11/14 of the lesions were completely healed. The transplantation of autologous keratinocytes may allow faster closure of diabetic foot lesions and subsequently reduce length of hospitalization. This method can easily be planned with regard to logistics and time, and furthermore, this therapy option can be carried out by the diabetologist.

Cartilage tissue engineering with novel nonwoven structured biomaterial based on hyaluronic acid benzyl ester

The aim of this study was to investigate the possibility of using the benzyl ester of hyaluronic acid (HYAFF® 11), a recently developed semisynthetic resorbable material, as a scaffold for the culture of human nasoseptal chondrocytes in tissue-engineering procedures of cartilage reconstruction. Different techniques such as immunohistochemistry, scanning electron microscopy, and confocal laser scanning microscopy were used to study the behavior, morphology, and phenotype expression of the chondrocytes, which were initially expanded and then seeded on the material. The nonwoven cell carrier allowed good viability and adhesivity of the cells without any surface treatment with additional substances. Furthermore, the cultured cells expressed cartilage-specific collagen type II, indicating that they were able to redifferentiate within the scaffold of HYAFF® 11 and were able to retain a chondrocyte phenotype even after a long period of in vitro conditions. Nevertheless, the expression of collagen type I, which was produced by dedifferentiated or incompletely redifferentiated chondrocytes, was noticeable. Additional data were obtained by subcutaneous implantation of samples seeded with human cells in the in vivo model of the athymic nude mouse. The results after 1 month revealed the development of tissue similar to hyaline cartilage. This study is promising for the use of this scaffold for tissue engineering of cartilage replacements. © 1998 John Wiley & Sons, Inc. J Biomed Mater Res, 42, 172–181, 1998.

Cartilage tissue engineering with novel nonwoven structured biomaterial based on hyaluronic acid benzyl ester.

The aim of this study was to investigate the possibility of using the benzyl ester of hyaluronic acid (HYAFF 11), a recently developed semisynthetic resorbable material, as a scaffold for the culture of human nasoseptal chondrocytes in tissue-engineering procedures of cartilage reconstruction. Different techniques such as immunohistochemistry, scanning electron microscopy, and confocal laser scanning microscopy were used to study the behavior, morphology, and phenotype expression of the chondrocytes, which were initially expanded and then seeded on the material. The nonwoven cell carrier allowed good viability and adhesivity of the cells without any surface treatment with additional substances. Furthermore, the cultured cells expressed cartilage-specific collagen type II, indicating that they were able to redifferentiate within the scaffold of HYAFF 11 and were able to retain a chondrocyte phenotype even after a long period of in vitro conditions. Nevertheless, the expression of collagen type I, which was produced by dedifferentiated or incompletely redifferentiated chondrocytes, was noticeable. Additional data were obtained by subcutaneous implantation of samples seeded with human cells in the in vivo model of the athymic nude mouse. The results after 1 month revealed the development of tissue similar to hyaline cartilage. This study is promising for the use of this scaffold for tissue engineering of cartilage replacements.

Selective acetylation reactions of hyaluronic acid benzyl ester derivative

The reaction of hyaluronic acid benzyl ester derivative ( 1) with trimethyl orthoacetate in the presence of an acid catalyst in N,N-dimethylformamide gave the expected 4,6-orthoester 2, along with its hydrolysed products the 6- O- ( 3) and the 4- O- ( 4) acetate hyaluronic acid benzyl esters as minor components. Acid hydrolysis of 2 followed by O-4→O-6 acetyl migration using t-butylamine afforded a mixture which contained the 6-acetate 3 as the major and the 4-acetate 4 as the minor compound. Synthesis of a mixture of 2′,3′,6- ( 6) and 2′,3′,4- ( 7) tri-acetates was achieved by peracetylation of 2 followed by treatment with aqueous acetic acid. Compound 1 on treatment with 2,2-dimethoxypropane in dimethyl sulfoxide in the presence of p-toluenesulfonic acid afforded the corresponding 4,6- O-isopropylidene derivative 8. The 2′,3′-di- O-acetate 10 was obtained from 8 by conventional acetylation followed by de-acetylation using trifluoroacetic acid. Acetylation of 1 with acetic anhydride in a mixture of N,N-dimethylformamide and pyridine gave the expected 2′,3′,4,6-tetraacetate derivative 11. In order to facilitate the NMR assignments of the acetate groups in 3– 11, the 2′,3′-diacetate-4,6-di(trideuterioacetate) 12 and the 2′,3′-di(trideuterioacetate)-4,6-diacetate 13 were prepared. The effect of substituents on conformational changes around the glycosidic linkages has been investigated.

Effect of drug hydrophilicity and membrane hydration on diffusion in hyaluronic acid ester membranes

The diffusion of solutes in hydrated polymer membranes may be influenced by solute hydrophilicity and membrane hydration. These effects were examined using a panel of six neutral drug molecules and hyaluronic acid benzyl ester membranes with varying degrees of esterification. The natural logarithm of diffusivity was found to be a function of membrane hydration for all drugs and membranes studied. A linear relationship between the natural logarithm of diffusivity and the inverse of membrane hydration was observed for the hydrophilic drugs, indicating consistency with Yasuda's free volume theory. Non-linear relationships were observed for hydrophobic drugs, suggesting deviation from the theory. A simple mathematical model was developed, based on parallel diffusion in hydrophilic and hydrophobic domains, and was found to adequately describe the diffusion characteristics of both hydrophilic and hydrophobic solutes.

Prevention of ovariectomy osteopenia in rats after vaginal administration of Hyaff 11 microspheres containing salmon calcitonin.

The benzyl ester of hyaluronic acid (Hyaff 11) is a highly mucoadhesive polymer and can be processed into microspheres that can effectively deliver incorporated drugs by closely adhering to the mucosal surface and by protecting the drug from enzymatic inactivation. In this study, Hyaff 11 microspheres have been investigated as novel delivery system for the vaginal administration of salmon calcitonin (sCT) to ovariectomized rats. Moreover, this particular animal model has been used to evaluate the biological activity of the polypeptide after vaginal and I.M. administration by measuring the skeletal effect of sCT. A total of 66 female rats were used: 6 rats served as the basal group; 12 were subjected to sham operation and left without pharmacological treatment; 48 underwent bilateral ovariectomy and these were divided at random into four groups of 12 animals. One group was not submitted to pharmacological treatment; another was treated daily with sCT I.M.; in the other two groups, the hormone was incorporated into Hyaff 11 microspheres and was daily administered by vaginal route as dry powder or dispersed in pessaries. Sixty days after surgery, rats were sacrificed and the proximal third of the right tibia was removed and prepared for histomorphometry. Treatment with sCT, independent of the administration route, largely prevented the bone volume loss of the nontreated ovariectomized group. Prevention mainly depended on the maintenance of the trabecular number and on the increase of the trabecular thickness. Independent of the administration procedure, sCT greatly reduced the increase of tartrate-resistant acid phosphatase-positive poly- and, particularly, mononucleated osteoclasts found in the nontreated ovariectomized group.(ABSTRACT TRUNCATED AT 250 WORDS)

Ocular sustained delivery of prednisolone using hyaluronic acid benzyl ester films

Hyaluronic acid benzyl ester films were studied for ocular delivery of prednisolone. The four polymers used were HYAFF 11 p25 (25% benzyl ester, 75% sodium salt), HYAFF 11 p50 (50% benzyl ester, 50% sodium salt), HYAFF 11 p75 (75% benzyl ester, 25% sodium salt) and HYAFF 11 (100% benzyl ester). The polymer with the lowest degree of esterification, HYAFF 11 p25, was the most hydrophilic and released drug faster than those with higher degrees of esterification, HYAFF 11 and HYAFF 11 p75, which are generally less hydrophilic. Tear fluid prednisolone concentrations were measured in rabbits after administration of the test films. Areas under the tear fluid concentration vs time curves (AUC 0–8 h) were calculated for all the dosage forms, from the time of dosing to 8 h post-dosing. The HYAFF 11 p25 films provided higher initial concentrations which rapidly declined below 30 μg/ml, 2 h post-dosing. Concentrations for the HYAFF 11 p75 film dropped below 30 μg/ml, 3 h post-dosing. The HYAFF 11 films provided the best results with sustained concentrations between 45 and 72 μg/ml for the 8 h study period. The results show that sustained delivery of prednisolone to the eye may be achieved with the use of hyaluronic acid esters.