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Hyaluronic Acid; Uses, Dosage, Side Effects, Interactions

Hyaluronic Acid is a glucosaminoglycan consisting of the D-glucuronic acid and N-acetyl-D-glucosamine disaccharide units that is a component of connective tissue, skin, vitreous humor, umbilical cord, synovial fluid and the capsule of certain microorganisms contributing to adhesion, elasticity, and viscosity of extracellular substances

Hyaluronic acid also called hyaluronan, is an anionic, nonsulfated glycosaminoglycan distributed widely throughout connective, epithelial, and neural tissues. It is unique among glycosaminoglycans in that it is nonsulfated, forms in the plasma membrane instead of the Golgi apparatus, and can be very large, with its molecular weight often reaching the millions.One of the chief components of the extracellular matrix, hyaluronan contributes significantly to cell proliferation and migration, and may also be involved in the progression of some malignant tumors

Mechanism of Action of Hyaluronic Acid

Osteoarthritis (OA), the most common type of arthritis, is a degenerative joint disease. Oxidative stress is well known to play important roles in cartilage degradation and pathogenesis of OA. The intra-articular injection of hyaluronic acid (IAHA) is accepted as an effective clinical therapy for OA, but we do not yet fully understand the mechanisms underlying the effects of HA on OA chondrocytes under oxidative stress. Here, we show for the first time that IAHA significantly reduces the synovial fluid levels of hydrogen peroxide (H2O2) and superoxide (O2(-)) in patients with knee OA. We also demonstrate that HA suppresses H2O2-induced cell death in human OA chondrocytes. Proteomic approaches (2-DE combined with mass spectrometry) allowed us to identify 13 protein spots corresponding to 12 non-redundant proteins as HA-regulated proteins in OA chondrocytes under oxidative stress. The expression levels of three putative HA-regulated proteins (TALDO, ANXA1, and EF2) in control, H2O2-, HA- and HA/H2O2-treated OA chondrocytes were verified by Western blotting and the results indeed support the notion that HA acts in anti-oxidation, anti-apoptosis, and the promotion of cell survival. Our results collectively demonstrate the utility of proteomic approaches and provide new insights into the chondroprotective effects of HA on OA. In the present study, we show for the first time that IAHA reduces the levels of H2O2 and O2(-) in synovial fluids from OA patients. We used primary cultured human OA chondrocytes as a model, treated cells with H2O2 to partly mimic their physiological conditions under oxidative stress, and examined the protective effects of HA. The proteomic approach allowed us to identify candidate proteins regulated by H2O2 and/or HA in OA chondrocytes. We found that proteins functioning in stress responses, apoptosis and protein synthesis were consistently regulated by HA in chondrocytes under oxidative stress. These novel results contribute to our understanding of the molecular mechanisms underlying HA-mediated chondroprotection.

ICAM-1 is known mainly as a metabolic cell surface receptor for hyaluronic acid, and this protein may be responsible mainly for the clearance of hyaluronic acid from lymph and blood plasma, which accounts for perhaps most of its whole-body turnover. Ligand binding of this receptor, thus, triggers a highly coordinated cascade of events that includes the formation of an endocytotic vesicle, its fusion with primary lysosomes, enzymatic digestion to monosaccharides, active transmembrane transport of these sugars to cell sap, phosphorylation of GlcNAc and enzymatic deacetylation. ICAM-1 may also serve as a cell adhesion molecule, and the binding of hyaluronic acid to ICAM-1 may contribute to the control of ICAM-1-mediated inflammatory activation.

Indications of Hyaluronic Acid

Hyaluronic acid has been used in attempts to treat osteoarthritis of the knee via injecting it into the joint. It has not been proven, however, to generate significant benefit and has potentially severe adverse effects.
Dry, scaly skin such as that caused by atopic dermatitis may be treated with skin lotion containing sodium hyaluronate as its active ingredient.
Hyaluronic acid has been used in various formulations to create artificial tears to treat dry eye.
Used to treat knee pain in patients with joint inflammation (osteoarthritis). It is usually used in patients who have not responded to other treatments such as acetaminophen, exercise, or physical therapy.
Hyaluronic acid may also be used in plastic surgery to reduce wrinkles on the face or as a filler in other parts of the body. May be used in ophthalmology to assist in the extraction of cataracts, the implantation of intraocular lenses, corneal transplants, glaucoma filtration, retinal attachment and in the treatment of dry eyes. Finally, hyaluronic acid is also used to coat the bladder lining in treating interstitial cystitis.

Therapeutic Indication of Hyaluronic Acid

Adjunct in treatment of noninfectious synovitis. Osteoarthritis in dogs and horses.
Osteoarthritis (OA) is the most common form of chronic arthritis worldwide. The etiology of pain in osteoarthritis is multifactorial and includes mechanical and inflammatory processes.
The use of intra-articular viscosupplementation in the nonoperative management of patients with osteoarthritis has become quite popular. Recent clinical data have demonstrated that the anti-inflammatory and chondroprotective actions of hyaluronic acid viscosupplementation reduce pain, from 4 to 14 weeks after injection, while improving patient function.
Viscosupplements are comparable in efficacy to systemic forms of active intervention, with more local reactions but fewer systemic adverse events, and hyaluronic acid has more prolonged effects than IA corticosteroids. Although several randomized controlled trials have established the efficacy of this treatment modality, additional high-quality randomized control studies with appropriate comparison are still required to clearly define the role of intra-articular hyaluronic acid injections in the treatment of osteoarthritis.
We review the basic science and development of viscosupplementation and discuss the mounting evidence in support of its efficacy and safety profile.
While many of the commonly used conservative treatments for knee osteoarthritis (OA) have been recognized to be effective, there is still insufficient evidence available. Among the pharmacological treatments for knee OA, oral non-steroidal anti-inflammatory drugs (NSAIDs) act rapidly and are recommended for the management of OA. However, frequent and serious adverse effects of NSAIDs have been recognized.
Intra-articular injections of hyaluronic acid(IA-HA) for the treatment of knee OA have been shown to reduce pain and improve joint function. However, there has been no qualified direct comparison study of the efficacy and safety between IA-HA and NSAIDs for patients with knee OA. The aim of this study was to clarify the efficacy and safety of early-phase IA-HA in comparison to those of NSAIDs for patients with knee OA.
This multicenter, randomized, open-label, parallel-group, non-inferiority comparison study with an oral NSAID involved a total of 200 patients with knee OA. An independent, computer-generated randomization sequence was used to randomly assign patients in a 1:1 ratio to NSAIDs three times per day for five weeks (n=100) or IA-HA once a week for five weeks (n=100). The primary endpoint was the percentage change in the patient-oriented outcome measure for knee OA, the Japanese Knee Osteoarthritis Measure (JKOM) score. All patients were questioned regarding any adverse events during treatment. The full analysis set (FAS) was used for analysis. The margin of non-inferiority was 10%. The analyses of primary endpoint included 98 patients in the IA-HA group and 86 patients in the NSAID group. The difference in the percentage changes of the JKOM score between the two intervention arms (IA-HA; -34.7% (P<0.001), NSAID; -32.2% (P<0.001)) was -2.5% (95% confidence interval (CI): -14.0 to 9.1), indicating IA-HA was not inferior to NSAID. The frequency of both withdrawal and adverse events in the IA-HA group were significantly lower than those in the NSAID group (P=0.026 and 0.004, respectively). The early efficacy of IA-HA is suggested to be not inferior to that of NSAIDs, and that the safety of the early phase of IA-HA is superior to that of NSAIDs for patients with knee OA.
The introduction of therapeutic genes in target tissues is considered as a novel tool for the treatment of several diseases. We have developed nanoparticles consisting of SLNs, protamine (P) and hyaluronic acid (HA) as a carrier for gene therapy. Stable complexes positively charged and with a particle size ranging from 240 nm to 340 nm were obtained. Transfection studies in ARPE-19 and HEK-293 cells showed the versatility of vectors to efficiently transfect cells with different division rate, widening the potential applications of SLN-based vectors.
In ARPE 19cells, the incorporation of P and HA-induced almost a 7-fold increase in the transfection capacity of SLNs. The CD44 inhibition studies suggested the participation of this receptor in the internalization of the vectors in this cell line. The intracellular disposition of DNA showed that the HA is able to modulate the high degree of condensation of DNA due to the protamine inside the cells; an important fact, if the vector is uptaken via non-degradative endocytosis. Besides, the therapeutic plasmid which encodes the protein retinoschisin was employed achieving a positive transfection in ARPE-19 cells, showing a promising application of this new non-viral system for the treatment of X-linked juvenile retinoschisis by gene therapy.
The primary aim of our current study was to utilize hyaluronic acid (HA) to decorate reconstituted high density lipoprotein (rHDL) loaded with lovastatin (LT), termed as HA-LT-rHDL, in order to investigate whether coating HA could efficiently evade from the undesired uptake of LT-rHDL in liver mediated by scavenger receptor class B type I (SR-BI) and then greatly accumulate LT-rHDL in atherosclerotic lesions via strong HA affinity to CD44 up-regulated at inflammatory sites such as atherosclerotic lesions, thus exerting enhanced atheroprotective efficacy. In vitro characterizations indicated the successful HA decoration onto the surface of LT-rHDL, which could be indirectly verified by the increased particle size, enhanced negative surface charge and reduced in vitro drug release rate after HA decoration. Compared with rHDL without HA, HA decoration endowed rHDL with better atherosclerotic lesions targeting efficiency and lower liver accumulation, proved by results from ex vivo imaging and tissue distribution. Furthermore, atheroprotective efficacy in model animal showed that HALT-rHDL had the best potent efficacy than other LT preparations, which was demonstrated by the fewest atherosclerotic lesions sizes, the most minimum mean intima-media thickness (MIT), the lowest macrophage infiltration and expression of matrix metalloproteinase-9 (MMP-9), respectively. Above results demonstrated that the newly designed HA-LT-rHDL could decrease the non-targeted uptake by the liver and deliver a large amount of drug into atherosclerotic lesions so as to efficiently suppress the advancement of atherosclerosis.
The objective of this study was to develop mucoadhesive polymeric excipients for vaginal drug delivery systems. Hyaluronic acid was thiolated and subsequently preactivated with 6-mercapto nicotinamide (HA-CYS-MNA) to enhance stability and mucoadhesive properties on vaginal mucosa. After determination of the thiol group content, disintegration studies and in vitro mucoadhesion studies (rotating cylinder and tensile) were performed. Furthermore, swelling behavior and cytotoxicity studies were performed in comparison with corresponding polymers. Both, disintegration and in vitro mucoadhesive studies revealed that modifying HA-CYS with MNA resulted in higher stability (3.6-fold prolonged disintegration time compared to unmodified hyaluronic acid) and prolonged mucoadhesion time. MTT assay and LDH revealed no toxicity for the polymeric excipients and safe for their use. Disintegration and swelling results conducted more pronounced stability of the preactivated thiomers compared to corresponding unmodified ones. According to these results, preactivated hyaluronic acid might be a useful tool for vaginal delivery systems
To evaluate if the hyaluronic acid (HA)-bound spermatozoa surpassed conventional microscopy-selected spermatozoa in the status of sperm DNA integrity by acridine orange (AO) fluorescence staining. Spermatozoa obtained from couples with indication for the intracytoplasmic sperm injection (ICSI) procedure due to male infertility (n = 34) and control males with normal sperm parameters (n = 12) were analyzed using AO fluorescence staining after density-gradient centrifugation (DGC), polyvinylpyrrolidone (PVP)-microscopic selection, and HA-binding selection to determine sperm DNA integrity. Percentages of DNA intact spermatozoa with green fluorescence were significantly higher in both PVP-microscopic selected spermatozoa (82.1 +/- 24.0%) and HA-bound spermatozoa (83.9 +/- 21.1%) than in spermatozoa prepared by DGC (66.8 +/- 24.0%). However, there was no significant difference between the PVP-sperm and HA-sperm groups. When the percentage of green fluorescent spermatozoa prepared by DGC fell initially below 68%, both PVP-microscopic and HA-binding selection failed to select over 90% spermatozoa with intact DNA for ICSI in the male infertility group. Compared to control males with normal sperm parameters (99.3 +/- 1.8%), the proportion of green fluorescence sperm after HA-binding selection from couples with male infertility (83.9 +/- 21.1%) did not reach the range of > 99% reported by Yagci et al. The percentages of DNA intact spermatozoa between the PVP-sperm and HA-sperm groups were not significantly different. In an ICSI procedure, a well-trained embryologist will have the same ability to choose sperm with intact DNA by conventional microscopic selection as with HA-bound spermatozoa selection.
For the effective diagnosis and therapy of atherosclerosis, there is a pressing need to develop the carrier which can specifically deliver the agents to the pathological site. Since the representative hallmark of atherosclerosis in its pathogenic process is the over-expression of the receptors for hyaluronic acid (HA) such as stabilin-2 and CD44, we herein investigated the potential of HA nanoparticles (HA-NPs) as the carrier for active targeting atherosclerosis. From in vitro cellular uptake tests, it was revealed that HA-NPs were selectively taken up by the cells over-expressing stabilin-2 or CD44. On the other hand, the cellular uptake of HA-NPs was drastically reduced when the cells were pre-treated with excess amount of free HA, implying that HA-NPs were taken up by the receptor-mediated endocytosis. Following systemic administration of Cy5.5-labeled NPs into the ApoE-deficient mice as the animal model, the atherosclerotic legion was assessed at 24 post-injection by using the optical imaging system. Interestingly, the fluorescent signal of the atherosclerotic lesion by HA-NPs was much stronger than that of the normal aorta. Three-dimensional z-stack images of an atherosclerotic plaque indicated the even distribution of HA-NPs in the atherosclerotic lesion. It was demonstrated by immunohistochemistry that HA-NPs were co-localized with the HA receptors including stabilising-2 and CD44. In addition, the amount of HA-NPs, accumulated in the atherosclerotic lesion, was much higher than that of HGC-NPs, known to reach the atherosclerotic lesion by the passive targeting mechanism. Overall, it was evident that HA-NPs could effectively reach the atherosclerotic lesion via the active targeting mechanism after systemic administration, implying their high potential as the carrier for diagnosis and therapy of atherosclerosis.
The cluster of differentiation-44 (CD44) is a ubiquitously present glycoprotein on the surface of mammalian cells that plays a significant role in a number of biological functions. Since the discovery that the receptor is over-expressed in a variety of solid tumors, such as pancreatic, breast and lung cancer, many studies have focused on methods for targeting CD44 in an attempt to improve drug delivery and discrimination between healthy and malignant tissue, while reducing residual toxicity and off-target accumulation. In this review, we describe CD44 receptor biology and its involvement in the different stages of tumor growth and metastasis, as well as methods currently used for targeting the receptor. Hyaluronic acid, the primary CD44 binding molecule, has proved a significant ally in developing nanocarriers that demonstrate preferential tumor accumulation and increased cell uptake. We outline a number of research approaches from the current literature that take advantage of hyaluronic acid’s targeting ability and describe the possible advantages for each approach. The value of CD44 targeting can be easily appreciated from the number of different approaches that have reached clinical trials.
Vitrectomy requires the substitution of the natural vitreous, as well as tamponading of retinal breaks. Clinically available alternatives such as gas and silicone oil have side effects such as inflammation, secondary glaucoma, cataract, and a need for head posturing. In this study, a hydrogel of cross-linked sodium hyaluronic acid (Healaflow) is evaluated for use as a novel vitreous substitute. A combined 25-20-gauge pars plana vitrectomy with a posterior vitreous detachment was performed in the right eye of twelve pigmented rabbits, with subsequent injection of approximately 1 ml Healaflow. Clinical evaluation, measurement of intraocular pressure (IOP), and full-field ERG were performed postoperatively. The rabbits were sacrificed at different time points between 42 and 105 days. After enucleation, the eyes were examined macroscopically, photographed, and prepared for histological examination with routine microscopy and immunohistochemistry. Hella flow was successfully used with standard surgical procedures and remained translucent but did lose most of its viscosity during the postoperative period. One rabbit was lost due to unrelated causes. In two eyes iatrogenic partial retinal detachments were seen, and in two eyes significant cataract developed due to intra-operative complications. ERG-recordings revealed no toxic effect on rod or cone function. Routine microscopy and immunohistochemistry demonstrated normal morphology with some Muller cell activation (up-regulation of glial acidic fibrillary protein, GFAP) compared to unoperated eyes and no significant DNA-fragmentation (TUNEL-assay). Hella flow did not affect retinal morphology or function negatively during long-term use as a vitreous substitute, making it highly interesting in this setting. An estimated retention time of a few weeks suggests the potential for use as a short-term tamponade. Future work will include an increased ratio of cross-linking to prolong the structural integrity of the gel.[Barth H et al; Graefes Arch Clin Exp Ophthalmol (2016)\
Although the disulfide bond crosslinked hyaluronic acid hydrogels have been reported by many research groups, the major researches were focused on effectively forming hydrogels. However, few researchers paid attention to the potential significance of controlling the hydrogel formation and degradation, improving biocompatibility, reducing the toxicity of exogenous and providing convenience to the clinical operations later on. In this research, the novel controllable self-crosslinking smart hydrogels with in-situ gelation property were prepared by a single component, the thiolated hyaluronic acid derivative (HA-SH), and applied as a three-dimensional scaffold to mimic native extracellular matrix (ECM) for the culture of fibroblasts cells (L929) and chondrocytes. A series of HA-SH hydrogels were prepared depending on different degrees of thiol substitution (ranging from 10 to 60%) and molecule weights of HA (0.1, 0.3 and 1.0MDa). The gelation time, swelling property and smart degradation behavior of HA-SH hydrogel were evaluated. The results showed that the gelation and degradation time of hydrogels could be controlled by adjusting the composition of HA-SH polymers. The storage modulus of HA-SH hydrogels obtained by dynamic modulus analysis (DMA) could be up to 44.6kPa. In addition, HA-SH hydrogels were investigated as a three-dimensional scaffold for the culture of fibroblasts cells (L929) and chondrocytes cells in vitro and as an injectable hydrogel for delivering chondrocytes cells in vivo. These results illustrated that HA-SH hydrogels with controllable gelation process, intelligent degradation behavior, excellent biocompatibility and convenient operational characteristics supplied potential clinical application capacity for tissue engineering and regenerative medicine.
Hyaluronic acid (HA)-based doxorubicin (DOX) nanoparticles (HA-NPs) were fabricated via ion-pairing between positively charged DOX and negatively charged HA, which displayed near-spherical shapes with an average size distribution of 180.2nm (PDI=0.184). Next, HA-NPs were encapsulated in liposomal carriers to afford HA-based DOX liposomes (HA-LPs), which also showed near-spherical morphology with an average size of 130.5nm (PDI=0.201). HA-NPs and HA-LPs displayed desirable sustained-release profiles compared to free DOX, and moreover, HA-LPs were proven to prevent premature release of DOX from HA-NPs. Cell-based studies demonstrated HA-NPs and HA-LPs were selectively taken up by CD44(+) tumor cells, and DOX was released intracellularly to target the cell nuclei. Both HA-NPs and HA-LPs showed comparable levels of penetration efficiency in tumor spheroids. In vivo studies revealed that HA-NPs and HA-LPs significantly prolonged the blood circulation time of DOX, decreased accumulation in the normal tissues and enriched drugs into the tumors. Furthermore, HA-NPs and HA-LPs greatly enhanced the therapeutic efficacy of DOX in tumor-bearing mice and minimized systemic toxicity against vital organs. In sum, HA-NPs and HA-LPs represent promising nanocarriers for CD44(+) tumor-targeted delivery.
During the last decades, therapies targeting cervical cancer have been considerably improved. Surgery and radiotherapy (RT) represents the main common therapeutic approach in cervical cancer. In order to minimize the side effects of radiotherapy approach, several protocols have been developed such as brachytherapy (BRT). Among the side effects associated with RT, the vaginal atrophy is the most important and common one. Vaginal atrophy, in turn, leads to additional alterations like inflammation, associated to relevant symptoms such as itching, burning, and dyspareunia. All these alterations heavily affect the quality of women’s lives. The aim of our study was to evaluate the toxicity induced by RT on vaginal mucosa, and the adjuvant action of a product containing LMWHA, vitamin A, and Vitamin E. The introduction of adjuvant therapies may have likely had a relevant place in providing that result. A prospective randomized study was designed. From October 2006 to October 2008, 45 women with a mean age 38+/-6 years were enrolled. After surgery, all patients were treated with 4 weeks of RT and 4 weeks of BRT, concomitantly with chemotherapy. They were randomly assigned in two groups: 23 women were treated with two suppositories (Santos) per day for 4 months. For the first two months, the preventive treatment was simultaneous to RT and BRT. Instead, the control groups for composed of 22 patients and they did not undergo any treatment during RT. To evaluate the efficacy of Saints treatment three biopsies were performed. At the second biopsy, after the BRT therapy, the treated group showed a statistically significant improvement (P<0.05 vs. control) on inflammation, cell atypia, fibrosis, mucositis, and bleeding. At the third biopsy, two months after BRT, further statistically improvement was observed for all RT/BRT associated side effects. The treatment showed an efficacy also in terms of pain severity. Our data suggest that low molecular weight HA shows good performances in treating RT-damaged tissue and plays a key role in all steps of the healing process. Indeed the results show that women exposed to RT treatments and simultaneously treated with Saints had an optimal resolution of vaginal atrophy and related symptoms.
Several studies have reported that sodium hyaluronate is able to improve both symptoms and signs in patients with dry eye but none have demonstrated an improvement of conjunctival epithelial cell abnormalities of the ocular surface. The aim of this study was to explore the effect of sodium hyaluronate-containing eye drops on the ocular surface of patients with the dry eye during long-term treatment. A randomized double-blind study was undertaken in 86 patients with medium to the severe dry eye (that is, rose bengal and/or fluorescein test score of at least 3, tear film break up time <10 seconds, or Schirmer’s test <5.5 mm). Patients were treated with either preservative-free sodium hyaluronate or saline for 3 months at a dose of one drop 4-8 times a day. Bulbar impression cytology, slit lamp examinations, and subjective symptoms were evaluated after 1, 2, and 3 months. Impression cytology was considered the primary efficacy parameter of the study. Results: The efficacy analysis was performed on a total of 44 patients who were able to fully adhere to the protocol. After 3 months of treatment, sodium hyaluronate improved impression cytology score (p = 0.024 v baseline). At the same time also the difference with respect to placebo was statistically significant (p = 0.036). Study medication was well tolerated and no treatment-related adverse events occurred during the study. Sodium hyaluronate may effectively improve ocular surface damage associated with dry eye syndrome.
In the present clinical trial, the efficacy of a new nasal cream containing hyaluronic acid (Rhinogen) on mucosal wound healing has been evaluated in comparison to an ointment (H.E.C.), which is commonly prescribed for this disorder in Switzerland. A total of 56 patients recovering from the surgical operation of the nasal cavities participated in this study. In both treatment groups (Rhinogen n = 27 patients, H.E.C. n = 29 patients) respiration and the condition of the nasal mucosa clearly improved. The statistical comparison between the two treatments showed a significant difference in favor of Rhinogen. With regard to the improvement in respiration, the Rhinogen-treated group showed a faster and greater progress than did the H.E.C.-treated group. Furthermore, hyaluronic acid prevented extensive crust formation during the first week of wound healing. The analysis of the efficacy of the treatments, judged by both the patients and the investigator, showed the overall superiority of Rhinogen (patients: p = 0.0041, investigator: p = 0.0023) after 6 weeks of treatment. Furthermore, Rhinogen scored significantly better than H.E.C. with respect to the organoleptic parameters of smell and sensation of cooling. Both treatments were well tolerated. No adverse reactions were reported or observed for Rhinogen, whereas three patients in the H.E.C.-treated group complained of a sore throat and burning sensation when the ointment flowed down into the pharynx. In conclusion, this study confirms the therapeutic benefit of hyaluronic acid in mucosal wound healing.
is indicated for lip augmentation and for mid-to-deep dermal implantation for the correction of moderate to severe facial wrinkles and folds, such as nasolabial folds.

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Drug Interactions of Hyaluronic Acid

Extreme loss of body water
Anticoagulants (eg, warfarin), inhibitors of platelet aggregation (eg, clopidogrel), thrombolytic agents (eg, alteplase, streptokinase): Administer hyaluronic acid with caution in patients who have undergone therapy with these agents in the preceding 3 weeks because bruising or bleeding may occur at the injection sites.
Do not use concomitantly with disinfectants containing quaternary ammonium salts because hyaluronic acid can precipitate in their presence.
Ultraviolet B (UVB) radiation induces inflammation in the skin specifically at the site of exposure. We unexpectedly found that UVB-induced inflammation was not induced in gp91phox-depleted mice. To test whether gp91phox is directly involved in UVB-induced inflammation, neutrophil- and hyaluronic acid-depleted mice were also irradiated and examined for their response. Hyaluronic acid-depleted mice showed strongly inhibited UVB-induced inflammation, but the neutrophil-depleted mice did not exhibit any suppressed UVB-induced inflammation. To elucidate the pathway by which UVB irradiation-induced inflammation, we examined the expression of the nucleotide-binding domain, leucine-rich-containing family, pyrin domain-containing-3 (NLRP3) and caspase-1 in the mouse skin. An increase in the expression of NLRP3 and caspase-1 was seen following the UVB irradiation of C57BL mice; however, the UVB-irradiated gp91phox-knockout (gp91phox(-/-)) mice did not have this increase in expression. Furthermore, the plasma IL-1beta level increased after the UVB irradiation in C57BL mice, but there was no change in the gp91phox(-/-) mice. These results clearly indicate that nicotinamide adenine dinucleotide phosphate oxidase is activated by gp91phox, which is expressed on the surface in response to the increased expression of hyaluronic acid induced by UVB irradiation, and as result, the generation of reactive oxygen species (ROS) increases. This ROS activate NLRP3, and NLRP3 leads to the production of caspase-1, which subsequently increases IL-1beta, thereby finally inducing inflammation. It is thought that this system may play an important role in the damage and aging of the skin, and further studies are necessary to confirm these finding.
Hyaluronic acid (HA), a glycosaminoglycan with high molecular weight, has been reported to promote cell proliferation and serves as an important extracellular matrix component. The aim of this study was to in vitro investigate whether HA is able to reduce reactive oxygen species (ROS)-induced heart ischemia-reperfusion injury and activate the cardiomyocyte’s damage surveillance systems. Accordingly, Rattus cardiomyocyte line, H9C2, was treated with H(2)O(2) as a heart ischemia-reperfusion model followed by incubation with low molecular weight hyaluronan (LMW-HA, 100 kDa) or high molecular weight hyaluronan (HMW-HA, 1000 kDa) and proteomic analysis was performed to investigate the physiologic protection of HA in H(2)O(2)-induced ischemia-reperfusion in cardiomyocyte. Our data demonstrated that HA treatment does protect cardiomyocyte in the ROS-induced ischemia-reperfusion model and the molecular weight of HA is a crucial factor. HMW-HA has been shown to significantly facilitate cell migration and wound healing via cytoskeletal rearrangement. Additionally, 2D-DIGE combined MALDI-TOF/TOF analysis showed that HMW-HA might modulate biosynthetic pathways, cell migration, cell outgrowth and protein folding to stimulate wound healing as well as prevent these ischemia-reperfusion-damaged cardiomyocytes from cell death. To our knowledge, we report for the first time the cell repair mechanism of HMW-HA against ischemia-reperfusion-damage in cardiomyocytes based on cell biology and proteomic analysis.
Within ultraviolet radiation, ultraviolet B (UVB) is the most energetic and damaging to humans. At the protein level, UVB irradiation downregulates the expression of antioxidant enzymes leading to the accumulation of reactive oxygen species (ROS). Due to lacking of a global analysis of UVB-modulated corneal proteome, we investigate in vitro the mechanism of UVB-induced corneal damage to determine whether hyaluronic acid (HA) is able to reduce UVB irradiation-induced injury in human corneal epithelial cells. Accordingly, human corneal epithelial cell lines (HCE-2) were irradiated with UVB, followed by incubation with low molecular weight HA (LMW-HA, 100 kDa) or high molecular weight HA (HMW-HA, 1,000 kDa) to investigate the physiologic protection of HMW-HA in UVB-induced corneal injury, and to perform a global proteomic analysis. The data demonstrated that HA treatment protects corneal epithelial cells in the UVB-induced wound model and that the molecular weight of HA is a crucial factor. Only HMW-HA significantly reduces the UVB-induced cytotoxic effects in corneal cells and increases cell migration and wound-healing ability. In addition, the proteomic analysis showed that HMW-HA might modulate cytoskeleton regulation, signal transduction, biosynthesis, redox regulation, and protein folding to stimulate wound healing and to prevent these UVB-damaged cells from cell death. Further studies evidenced membrane-associated progesterone receptor component 1 (NPR) and malate dehydrogenase (MDH2) play essential roles in protecting corneal cells from UVB irradiation. This study reports on UVB-modulated cellular proteins that might play an important role in UVB-induced corneal cell injury and show HMW-HA to be a potential substance for protecting corneal cells from UVB-induced injury.
Virtually all patients receiving radio- and chemotherapy for cancer develop oral mucositis, a severe and highly debilitating condition. The onset of mucositis is thought to involve the production of reactive oxygen species (ROS) in the submucosa. Here we investigated a possible protective effect of a commercial formulation of hyaluronic acid (HA) enriched with amino acids (Glucosamine) against the damage induced by oxidative stress both in vitro and in vivo. Transient exposure of normal human oral fibroblasts to hydrogen peroxide (H(2)O(2)) led to irreversible senescence, as demonstrated by a sustained increase in the levels of p16(INK4A) and SA-betaGal. Conditioned media from senescent fibroblasts induced detrimental effects on keratinocytes, as shown by reduced metabolic activity and migration capability. Pre-treatment with Glucosamine prevented H(2)O(2) -induced, but not TGF-beta-induced, fibroblast senescence with a concomitant reduction of fibroblast-induced loss of keratinocyte vitality and functional activity. Finally, data from a case-series of patients undergoing radio/chemotherapy strongly suggested that prophylactic use of the hyaluronic acid-based compound in the form of a spray may be effective in preventing the onset of oral mucositis.
Hyaluronic acid (HA) hydrogels are interesting delivery systems for topical applications. Besides moisturizing the skin and improving wound healing, HA facilitates topical drug absorption and is highly compatible with labile biomacromolecules. Hence, in this study we investigated the influence of HA hydrogels with different molecular weights (5 kDa, 100 kDa, 1 MDa) on the skin absorption of the model protein bovine serum albumin (BSA) using fluorescence lifetime imaging microscopy (FLIM). To elucidate the interactions of HA with the stratum corneum and the skin absorption of HA itself, we combined FLIM and Fourier-transform infrared (FTIR) spectroscopy. Our results revealed distinct formulation and skin-dependent effects. In barrier deficient (tape-stripped) skin, BSA alone penetrated into dermal layers. When BSA and HA were applied together, however, penetration was restricted to the epidermis. In normal skin, penetration enhancement of BSA into the epidermis was observed when applying low molecular weight HA (5 kDa). Fluorescence resonance energy transfer analysis indicated close interactions between HA and BSA under these conditions. FTIR spectroscopic analysis of HA interactions with stratum corneum constituents showed an alpha-helix to beta-sheet interconversion of keratin in the stratum corneum, increased skin hydration, and intense interactions between 100 kDa HA and the skin lipids resulting in a more disordered arrangement of the latter. In conclusion, HA hydrogels restricted the delivery of biomacromolecules to the stratum corneum and viable epidermis in barrier deficient skin, and therefore seem to be potential topical drug vehicles. In contrast, HA acted as an enhancer for delivery in normal skin, probably mediated by a combination of cotransport, increased skin hydration, and modifications of the stratum corneum properties.
Successful anticancer chemotherapy requires targeting tumors efficiently and further potential to eliminate cancer stem cell (CSC) subpopulations. Since CD44 is present on many types of CSCs, and it binds especially to hyaluronic acid (HA), we tested whether coating solid lipid nanoparticles with hyaluronan (HA-SLNs) would allow targeted delivery of paclitaxel (PTX) to CD44-overexpressing B16F10 melanoma cells. First, we developed a model system based on melanoma stem-like cells for experiments in vitro and in mouse xenografts, and we showed that cells expressing high levels of CD44 (CD44(+)) displayed a strong CSC phenotype while cells expressing low levels of CD44 (CD44(-)) did not. This phenotype included sphere and colony formation, a higher proportion of side population cells, expression of CSC-related markers (ALDH, CD133, Oct-4) and tumorigenicity in vivo. Next, we showed that administering PTX-loaded HA-SLNs led to the efficient intracellular delivery of PTX and induced substantial apoptosis in CD44(+) cells in vitro. In the B16F10-CD44(+) lung metastasis model, PTX-loaded HA-SLNs targeted the tumor-bearing lung tissues well and subsequently exhibited significant antitumor effects with a relatively low dose of PTX, which provided significant survival benefit without evidence of adverse events.