Exogenous Hyaluronic Acid Research Articles

Efficacy of intra-articular hyaluronic acid injections in the rehabilitation programme of Knee Osteoarthritis

Abstract Knee osteoarthritis (KOA) is a disease of wear and tear, leading to disability and ultimate surgical treatment. KOA has multiple etiologies and is considered a disorder of physiological processes that is manifested by progressive lesions of articular cartilage caused by chondrolysis, synovial membrane inflammation and subchondral osteophytes production. Modalities that limit the progression of the KOA and rehabilitation are a constant interest for clinicians and medical researchers. Various physical therapies and pharmacological strategies are currently used for pain management, with no clear benefit in terms of inhibiting disease progression. In this context, the use of exogenous hyaluronic acid (HA) (that is a vital component of the synovial fluid and involved in lubricating all synovial joints) has been explored as viable alternative to alleviate knee pain and temporarily restore knee function. In the following, we will review the use of HA injections associated with the physiotherapeutic recovery plan in the treatment of KOA. The therapeutic effects of the HA is still a matter of debate in the field, with no consensus being reached so far and thus, difficult to evaluate. However, much research has to be done to understand the therapeutic efficacy of HA alone or in combination with physical therapies and the role that other factors may play in unraveling its beneficial effects on the KOA. Key words: knee, osteoarthritis, hyaluronic acid, physiotherapy, rehabilitation,

Use of Exogenous Hyaluronic Acid for the Treatment of Patellar Chondropathy- A Six-Month Randomized Controlled Trial.

Objective To evaluate the clinical and radiological benefits of intra-articular exogenous hyaluronic acid for the treatment of chondral patellar injury. Method Randomized clinical trial with 70 patients divided into 2 groups: those submitted to physical therapy for 3 months, and those submitted to physical therapy associated with the intra-articular administration of 2 mL of hyaluronic acid for the same period, who had anterior knee pain and patellar cartilage injury of grades II or III with no significant bone abnormalities. The functional scores and the characteristics of the physical and imaging exams were evaluated before and 3 and 6 months after the treatment. Result The average age of the patients was 32 ± 7.6 years. Patients from the hyaluronic acid group had better Kujala et al and Lysholm scores, and lower pain scores after 3 and 6 months of treatment when compared to the control group. The incidence of positive Clarke maneuver was lower in the treated group, but there was no difference in the magnetic resonance imaging classification. Conclusion Patients with patellar chondropathy of grades II or III treated with hyaluronic acid and physical therapy had less pain (visual analogue scale, VAS), and better functional results in the Lysholm and Kujala et al questionnaires after 3 and 6 months of treatment compared to patients undergoing physical therapy alone. In addition, the number of cases with a negative Clarke maneuver was larger in the treated group after 6 months of treatment.

Abstract 119: The influence of CD44/HA signaling on CML survival and resistance to tyrosine kinase inhibitor treatment

Abstract The development of tyrosine kinase inhibitors (TKIs) such as imatinib has greatly improved the prognosis and survival of patients diagnosed with chronic myeloid leukemia (CML). However, many CML patients relapse due to the presence of minimal residual disease and the development of TKI resistance. It is unknown exactly what causes the relapse, but based on research from our laboratory we hypothesize that hyaluronic acid (HA) produced by fibroblasts in the microenvironment of the bone marrow provide for a protective environment preventing the tumor cells from therapy-induced cell death or senescence. The purpose of this study is to determine the influence that HA and its binding to its receptor (CD44) has on CML survival and resistance development. Initially, we examined the effect that stromal cell derived and/or exogenously added HA has on CML resistance to imatinib. Cell survival, apoptosis, HA production, and signaling events elicited by the interactions of HA with CD44 was examined using methods such as ELISA, western blotting, and MTT analysis. Furthermore, we examined the potential therapeutic use of compounds known to alter HA production and/or signaling for the treatment of CML. Our study demonstrates increased cell survival when the tumor cells are co-cultured with fibroblasts or have exogenous HA added to them after imatinib treatment. Additionally, we show that knocking out hyaluronic acid synthases reduces this protective effect. This data supports our hypothesis and we believe it could help lead to improved therapeutic treatment for CML. Note: This abstract was not presented at the meeting. Citation Format: Kylie Humphries, Olga Uchakina, Bryan Hostetler, Robert McKallip. The influence of CD44/HA signaling on CML survival and resistance to tyrosine kinase inhibitor treatment [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 119.

Effect of diclofenac etalhyaluronate (SI-613) on the production of high molecular weight sodium hyaluronate in human synoviocytes

BackgroundWe have reported that a single intra-articular injection of diclofenac etalhyaluronate (SI-613) exerted a potent and long-lasting analgesic effect in experimental arthritis models. In the present study, we investigated the effect of SI-613 on the production of high molecular weight hyaluronic acid (HMW-HA) in synoviocytes from osteoarthritis (OA) patients and compared its efficacy with that of hyaluronic acid (HA).MethodsWe compared the effect of SI-613, HA, and diclofenac sodium (DF-Na) on high molecular weight HA production by human synoviocytes.ResultsSI-613 and exogenous HA induced the production of high molecular weight HA in synoviocytes from OA patients, whereas DF-Na had no effect. The molecular weight of newly produced HA was about 1000 kDa in the HA-treated synoviocytes and much higher than 2400 kDa in the SI-613-treated cells. The effect of the mixture of HA and DF-Na was similar to that of HA alone in that the molecular weight of newly produced HA was around 1000 kDa. SI-613 significantly suppressed hyaluronidase 2 (HYAL2) mRNA expression and significantly enhanced hyaluronan synthase 2 (HAS2) mRNA expression. HA had no effect on the expression levels of HYAL and HAS.ConclusionThe present results clearly demonstrate that SI-613 induces the production of high molecular weight HA in synoviocytes from OA patients, suggesting the long-lasting analgesic and disease modifying effect of SI-613 for OA. Taken together with the anti-inflammatory and analgesic effects we recently reported for the intra-articular administration of SI-613 to experimental animal models, SI-613 holds great promise for the treatment of knee osteoarthritis.

Hyaluronic Acid Accelerates Re-epithelialization and Alters Protein Expression in a Human Wound Model.

Hyaluronic acid (HA), a large glycosaminoglycan involved in proliferation, migration, and tissue repair, is suggested to be an important factor for keratinocyte activation and re-epithelialization. The experimental hypothesis of this study was that HA accelerates re-epithelialization, and we aimed to investigate the effect of exogenous intradermal HA during deep dermal, incisional wound healing in vivo in humans, the primary endpoint being re-epithelialization. A total of 8 standardized deep dermal incisional wounds (depth 1.6 mm, width 1.8 mm) per subject were induced in 10 healthy volunteers. Two of the wound sites per subject were pretreated with injections of HA and 2 with saline solution. At 2 time points (24 hours and 14 days), 2 biopsies for each treatment group (one for histology and one for proteomics) were taken. Skin erythema was measured at 24-hour intervals for 14 days as a surrogate measurement of inflammation. At 24 hours, 8 of 9 wounds pretreated with HA showed complete re-epithelization, whereas none of the wounds pretreated with saline had re-epithelized. Wounds pretreated with HA also showed a 10-fold regulation of 8 identified proteins involved in wound healing compared to wounds treated with saline solution. No difference in inflammation, as measured as erythema, could be seen between any of the groups. We conclude that HA accelerates re-epithelialization and stimulates an altered protein expression in vivo in human deep dermal incisional skin wounds, but has no effect on the inflammation process as measured by erythema.

Exogenous hyaluronic acid and chondroitin sulfate crosslinking treatment for increasing the amount and stability of glycosaminoglycans in bioprosthetic heart valves.

Glutaraldehyde (GLUT) crosslinked bioprosthetic heart valves (BHVs) might fail due to progressive degradation and calcification. GLUT cannot stabilize glycosaminoglycans (GAGs), which are important for BHVs' life time. In this current study we developed a new BHVs preparation strategy using exogenous hyaluronic acid (HA)/chondroitin sulfate (CS) supplement and sodium trimetaphosphate (STP) crosslinking method. Exogenous HA and CS provide additional GAGs for pericardiums. STP could link two GAGs by reacting with hydroxyl groups in GAGs' repeating polysaccharides units. The feeding ratios of HA/CS were optimized. The GAGs content and long-term stability in vitro, biocompatibility, the in vivo GAGs stability and anti-calcification potential of GLUT/HA/CS and STP treated pericardiums were characterized. We demonstrated that GLUT/HA/CS and STP treated pericardiums had sufficiently increased GAGs' amount and stability and decreased calcification. This new exogenous hyaluronic acid/chondroitin sulfate supplement and sodium trimetaphosphate crosslinking strategy would be a promising method to make BHVs with better structural stability and anti-calcification properties.

Hyaluronic acid-CD44 interactions promote BMP4/7-dependent Id1/3 expression in melanoma cells

BMP4/7-dependent expression of inhibitor of differentiation/DNA binding (Id) proteins 1 and 3 has been implicated in tumor progression and poor prognosis of malignant melanoma patients. Hyaluronic acid (HA), a pericellular matrix component, supports BMP7 signalling in murine chondrocytes through its receptor CD44. However, its role in regulating BMP signalling in melanoma is not clear. In this study we found that depletion of endogenously-produced HA by hyaluronidase treatment or by inhibition of HA synthesis by 4-methylumbelliferone (4-MU) resulted in reduced BMP4/7-dependent Id1/3 protein expression in mouse melanoma B16-F10 and Ret cells. Conversely, exogenous HA treatment increased BMP4/7-dependent Id1/3 protein expression. Knockdown of CD44 reduced BMP4/7-dependent Id1/3 protein expression, and attenuated the ability of exogenous HA to stimulate Id1 and Id3 expression in response to BMP. Co-IP experiments demonstrated that CD44 can physically associate with the BMP type II receptor (BMPR) ACVR2B. Importantly, we found that coordinate expression of Id1 or Id3 with HA synthases HAS2, HAS3, and CD44 is associated with reduced overall survival of cutaneous melanoma patients. Our results suggest that HA-CD44 interactions with BMPR promote BMP4/7-dependent Id1/3 protein expression in melanoma, contributing to reduced survival in melanoma patients.

The therapeutic effect of aucubin-supplemented hyaluronic acid on interleukin-1beta-stimulated human articular chondrocytes

BackgroundInjection of exogenous hyaluronic acid (HA) into the joint capsule improves symptoms of early stage osteoarthritis (OA). However, reactive oxygen species degrade HA into small oligosaccharides that can elicit pro-inflammatory responses. Likewise, disturbance of the antioxidant enzyme system and generation of oxidative stress by pro-inflammatory cytokines worsen knee OA. Accordingly, we proposed the use of aucubin, an antioxidant and anti-inflammatory compound, as a versatile adjuvant to HA for treating OA. MethodsPrimary human chondrocytes were cultured in media supplemented with aucubin in a series of concentrations (0, 0.01, 0.1, 1, and 10 μg/ml) to study dose-dependent toxicity. We then evaluated the therapeutic effects of HA (100 μg/ml) supplemented with aucubin (10 μg/ml) on interleukin-1 beta (IL-1β, 10 ng/ml)-stimulated chondrocytes. ResultsThe use of aucubin did not change cell viability or alter lactate dehydrogenase release to normal chondrocytes. Although the proliferation and sulfated glycosaminoglycan production were not affected, aucubin partially restored the hypertrophic transformation of chondrocytes. Relative to treatment with HA or aucubin alone, real-time PCR revealed that aucubin-supplemented HA down-regulated the mRNA levels of tumor necrosis factor-alpha (TNF-α), corrected collagen type 1 and aggrecan, and up-regulated tissue inhibitor of metalloproteinase 1. Moreover, ELISA testing also showed a reduced TNF-α production. Although superoxide dismutases activity was still distributed, aucubin restored total antioxidant capacity of IL-1β-stimulated chondrocytes. Western blotting further showed that aucubin inhibited cyclooxygenase-2 and regulated the nuclear factor (erythroid-derived 2)-like 2 pathway. ConclusionAucubin can enhance the anti-catabolic and anti-inflammatory effects of HA on OA chondrocytes.

A hyaluronic acid binding peptide-polymer system for treating osteoarthritis

Hyaluronic acid (HA) is found naturally in synovial fluid and is utilized therapeutically to treat osteoarthritis (OA). Here, we employed a peptide-polymer cartilage coating platform to localize HA to the cartilage surface for the purpose of treating post traumatic osteoarthritis. The objective of this study was to increase efficacy of the peptide-polymer platform in reducing OA progression in a mouse model of post-traumatic OA without exogenous HA supplementation. The peptide-polymer is composed of an HA-binding peptide (HABP) conjugated to a heterobifunctional poly (ethylene glycol) (PEG) chain and a collagen binding peptide (COLBP). We created a library of different peptide-polymers and characterized their HA binding properties in vitro using quartz crystal microbalance (QCM-D) and isothermal calorimetry (ITC). The peptide polymers were further tested in vivo in an anterior cruciate ligament transection (ACLT) murine model of post traumatic OA. The peptide-polymer with the highest affinity to HA as tested by QCM-D (∼4-fold greater binding compared to other peptides tested) and by ITC (∼3.8-fold) was HABP2-8-arm PEG-COLBP. Biotin tagging demonstrated that HABP2-8-arm PEG-COLBP localizes to both cartilage defects and synovium. In vivo, HABP2-8-arm PEG-COLBP treatment and the clinical HA comparator Orthovisc lowered levels of inflammatory genes including IL-6, IL-1B, and MMP13 compared to saline treated animals and increased aggrecan expression in young mice. HABP2-8-arm PEG-COLBP and Orthovisc also reduced pain as measured by incapacitance and hotplate testing. Cartilage degeneration as measured by OARSI scoring was also reduced by HABP2-8-arm PEG-COLBP and Orthovisc. In aged mice, HABP2-8-arm PEG-COLBP therapeutic efficacy was similar to its efficacy in young mice, but Orthovisc was less efficacious and did not significantly improve OARSI scoring. These results demonstrate that HABP2-8-arm PEG-COLBP is effective at reducing PTOA progression.

Hyaluronic Acid in the Third Millennium.

Since its first isolation in 1934, hyaluronic acid (HA) has been studied across a variety of research areas. This unbranched glycosaminoglycan consisting of repeating disaccharide units of N-acetyl-d-glucosamine and d-glucuronic acid is almost ubiquitous in humans and in other vertebrates. HA is involved in many key processes, including cell signaling, wound reparation, tissue regeneration, morphogenesis, matrix organization and pathobiology, and has unique physico-chemical properties, such as biocompatibility, biodegradability, mucoadhesivity, hygroscopicity and viscoelasticity. For these reasons, exogenous HA has been investigated as a drug delivery system and treatment in cancer, ophthalmology, arthrology, pneumology, rhinology, urology, aesthetic medicine and cosmetics. To improve and customize its properties and applications, HA can be subjected to chemical modifications: conjugation and crosslinking. The present review gives an overview regarding HA, describing its history, physico-chemical, structural and hydrodynamic properties and biology (occurrence, biosynthesis (by hyaluronan synthases), degradation (by hyaluronidases and oxidative stress), roles, mechanisms of action and receptors). Furthermore, both conventional and recently emerging methods developed for the industrial production of HA and its chemical derivatization are presented. Finally, the medical, pharmaceutical and cosmetic applications of HA and its derivatives are reviewed, reporting examples of HA-based products that currently are on the market or are undergoing further investigations.

Long-term Functional Impact of Viscosupplementation Versus True Placebo in Symptomatic Hip Osteoarthritis; A Randomized Control Trial

INTRODUCTION: Degenerative hip osteoarthritis (OA) is a common progressive disorder causing disability. The injection of exogenous hyaluronic acid (HA), or viscosupplementation (VS), can potentially help restore the properties of synovial fluid. There is little literature available evaluating the long-term effects and the functional impact of VS in hip OA. PURPOSE: To determine if a single intra-articular injection of a high-molecular weight (HMW) VS would improve function and decrease pain in persons suffering from hip OA. METHODS: A double-blinded randomized control trial was conducted at a University Hospital Center in Canada. Patients were randomly allocated to either the treatment group, an ultrasound guided single intra-articular injection of a HMW HA, or the placebo group, a single extra-articular injection of local anesthetic. Participants underwent evaluations at 2 weeks prior to the injection (T0), and at 1 month (T1), 3 months (T2) and 6 months (T3) post injection. Patients completed two questionnaires; the Hip Disability and Osteoarthritis Outcome Score (HOOS) and the 36-Item Short Form Survey (SF-36). Gait biomechanics were evaluated in a lab. RESULTS: Between May 2014 and September 2017, 38 participants were evaluated in this study over the course of 6 months. In the treatment group, N = 19 and in the placebo group, N = 18. The mean age at the time of injection was 55. On the HOOS symptom subscale, the placebo group worsened from T0 to T3 by 6.29% compared to the treatment group. The VS group improved their pain subtotal from T0 to T2 by 4.73%. The control group worsened by 1.22% during that same time and continued to deteriorate by 6.09% at T3. There were also improvements in the activities of daily living subscale from T0 to T3, with the treatment group improving by 5.29% while the placebo group worsened by 5.15%. The most important change occurred in the sports and recreational subscale of the HOOS. Between T0 and T3, the placebo group worsened by 7.611 points (- 17.82%). The treatment group improved by 6.67%. CONCLUSION: Our preliminary results suggest that a HMW VS hip injection for degenerative OA, when compared to true placebo, may lead to long-term improvements in pain relief, increase in function and in activity participation. NIH Clinical Trials Registry: NCT02086474

Metabolic engineering to enhance heterologous production of hyaluronic acid in Bacillus subtilis

Hyaluronic acid (HA) is a high-value biopolymer that is produced in large scales using attenuated strains ofgroup C streptococci. However, due to the pathogenicity and fastidious nature of these bacteria, the development of bioprocesses for HA production centered on robust ‘Generally Recognized as Safe (GRAS)’ organisms, such as Bacillus subtilis, is of increased interest. Here, we report metabolic engineering of novel B. subtilis strains in which the carbon flux has been partially diverted from central metabolism, i.e. the pentose phosphate pathway (PPP) and glycolysis, into HA biosynthesis. First, an improved base strain of B. subtilis was engineered for more effective HA production with less susceptibility to catabolite repression when expressing genes from a xylose-inducible promoter. Subsequently, Clustered Regularly Interspaced Palindromic Repeats interference (CRISPRi) was applied to reduce the expression of individual pfkA or zwf in the base strain, leading to substantial improvements to the HA titer with a concomitant decrease in the molecular weight (MW). On the other hand, multiplexed repression of both pfkA and zwf expression resulted in increases to the HA titer of up to 108% and enhancements to the MW, compared to the base strain. Moreover, the addition of exogenous HA monomers, i.e. glucuronic acid (GlcUA) and N-acetyl-glucosamine (GlcNAc), to B. subtilis cultures markedly improved the HA MW but decreased the HA titer, providing insights into the mechanism of HA biosynthesis by streptococcal hyaluronan synthase (SeHAS) in B. subtilis. Our study demonstrates the successful application of metabolic engineering strategies to establish B. subtilis as an effective platform for high-level HA production.

Hyaluronic Acid in the Treatment of Knee Osteoarthritis: Review

Knee osteoarthritis (OA) is a progressive disease in which the cartilage (cushioning between joints) wears away. As the cartilage wears away, it becomes frayed and rough, and the protective space between the bones decreases. This causes rubbing of bones, resulting in pain, swelling, stiffness, decreased ability to move and formation of bone spurs. Osteoarthritis develops slowly and the pain it causes worsens over time. Although there is no cure, there are many treatment options available to help manage pain and keep people staying active. Intra-articular (IA) hyaluronic acid (HA) injection is one of the treatment options. Hyaluronic acid is a polysaccharide, an anionic, non-sulfated glycosaminoglycan and is a naturally occurring substance found in the synovial fluid surrounding joints. It acts as a lubricant to enable bones to move smoothly over each other and as a shock absorber for joint loads. People with knee osteoarthritis have lower concentration and altered volume of hyaluronic acid than normal in their joints. Due to lower concentration and volume, exogenous hyaluronic acid may be injected into the knee joint. Treatment with exogenous hyaluronic acid contributes to restore the elastic and viscous properties of the synovial fluid, pain reduction and functional improvement. It is indicated for patients who have failed to respond adequately to conservative non-pharmacological therapy and simple analgesic.

Technology rationale for the development of CellularMatrix® A-CP-HA Kit, certified medical device allowing the combination of platelet rich plasma and hyaluronic acid for the treatment of osteoarthritis

The composition of synovial fluid of patients suffering from osteoarthritis (OA) is altered with reduced concentration and molecular weight of Hyaluronic Acid (HA) and augmentation of catabolic enzymes and inflammatory markers. Intra-Articular (IA) injections of exogenous HA aim to restore the rheological properties of the synovial fluid in the osteoarthritic joints. However, clinical studies have shown that the benefit of HA injections lasts only for around 6 months and that many patients don’t respond well to repeated course of HA treatment. On the other side, platelet rich plasma (PRP) IA injections have been shown to reduce pain and improve joint mobility, probably by modulating the expression of catabolic enzymes and inflammatory markers. As injections of HA and PRP use different pathway to alleviate symptoms in OA patients, the concept of combining these two treatments has emerged recently. In vitro evaluations and preliminary studies have demonstrated the therapeutic potential of this new therapeutic approach for OA treatment. To meet the needs of medical practitioners that want to treat their patients with this new treatment option, an innovative medical device, the CellularMatrix A-CP-HA Kit, has been specifically designed. This device is the first certified medical device that allows the combination of PRP and HA in manner compliant with medical devices regulation and good clinical practice.

High molecular weight hyaluronic acid: a two-pronged protectant against infection of the urogenital tract?

ObjectivesRecurrent urinary tract infections are associated with uropathogenic Escherichia coli (UPEC) ascending and infecting the urinary tract. Antibiotics provide only symptomatic relief, not prevent recurrence. Clinical evidence suggests that intravesical glycosaminoglycan therapy, such as hyaluronic acid (HA), helps reduce UTI recurrence. This has been investigated here using in vitro systems modelling the urogenital tract tissues.Methods RT4 bladder cells were preconditioned with high molecular weight HA (> 1500 kDa) at 2 mg mL−1 and challenged with UPEC to analyse barrier protection and bacterial adherence. Untreated and HA‐preconditioned VK2 E6/E7 vaginal cells were challenged with E. coli flagellin (50 ng mL−1) to mimic bacterial challenge, and media analysed for lipocalin‐2, human β‐defensin 2 and interleukin‐8 by ELISA. Experiments were repeated after siRNA knockdown of Toll‐like receptors 2, 4 and 5, and CD44 to investigate signalling.ResultsMicroscopic analyses showed reduced bacterial adherence and urothelial disruption with HA, suggesting that HA functions as a barrier protecting the epithelium from bacterial infection. Cells treated with HA and flagellin simultaneously produced more of the host antimicrobial peptide LCN2 and pro‐inflammatory IL‐8 (P < 0.05) compared to the no HA/flagellin challenges. Increased gene expression of DEFB4 (P < 0.05), but not the hBD2 peptide, was observed in the HA/flagellin‐challenged cells.ConclusionThese data suggest that exogenous HA has potential to protect the urogenital epithelia from UPEC infection via a two‐pronged approach that involves the physical enhancement of the epithelial barrier and augmentation of its innate immune response.

Abstract 2886: CD44/SMURF1 signaling maintains cancer stem cell-like invasive cells in head and neck squamous cell carcinoma

Abstract Head and neck squamous cell carcinoma (HNSCC) is highly invasive and resistant to therapies, where treatments often result in high rates of failure and disease recurrence. A subpopulation of tumor-initiating cancer stem cells (CSC) is thought to be responsible for metastatic invasion and drug resistance in many types of cancer, including HNSCC. CD44 is a known CSC marker in HNSCC but its role in maintaining CSC populations is not well understood. We previously reported that SMAD specific E3 ubiquitin protein ligase 1 (SMURF1) inhibition of bone morphogenetic protein (BMP) signaling is essential for maintaining a CD44-high CSC-like population in HNSCC. In this study, we sought to determine how CD44, a receptor for hyaluronic acid (HA) that constitutes a major component of the tumor stroma, is involved in the regulation of SMURF1 and the maintenance of a HNSCC CSC phenotype. CSC-like cells were enriched from HNSCC cell lines as CD44-high cells. CD44 signaling was stimulated by exogenous HA treatment and inhibited by CD44 knockout (KO) using an inducible CRISPR/Cas9 system. We then assessed changes in SMURF1 protein level, BMP signaling, transwell migration, Matrigel colony formation, and invasion through three-dimensional organotypic culture (OTC) following CD44 modulation. CD44-high cells were found to have increased extracellular HA production. Treatment with exogenous HA reduced BMP signaling, as determined by a reduction in phospho-SMAD1/5/8 levels, and increased transwell migration of CD44-high cells. CD44-KO reduced SMURF1 protein expression and inhibited Matrigel colony formation of an invasive and recurrent HNSCC-derived CD44-high cells but only partially reduced colony formation of a less-invasive cell line derived from a primary HNSCC. Knockout of CD44 expression showed a slight reduction in transwell migration of invasive cells. CD44-high cells also recapitulated an invasive and CSC-like growth pattern in OTC assays. In contrast, CD44-KO inhibited OTC invasion and the epithelial-to-mesenchymal transition (EMT) phenotype, resulting in increased apical epithelial growth and differentiation. CD44 reconstitution appeared to restore the invasive and EMT phenotype. Based on our current findings, CD44 may be crucial for maintaining an anchorage-independent and invasive phenotype of HNSCC but plays a minor role in carcinoma cell migration. Citation Format: Ali Khammanivong, Raj Gopalakrishnan, Erin B. Dickerson. CD44/SMURF1 signaling maintains cancer stem cell-like invasive cells in head and neck squamous cell carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 2886. doi:10.1158/1538-7445.AM2017-2886

Abstract 4898: eIF4E reshapes the surface of migrating AML cells through regulating Hyaluronic Acid synthesis &amp; CD44 expression

Abstract Formation of cell surface protrusions has long been associated with tumor cell migration, metastasis &amp; bone marrow homing &amp; retention. It was shown that increased expression of endogenous or exogenous hyaluronic acid synthesizing enzyme 3 (HAS3) induces growth of microvillus-like cell surface protrusions. Some of the biological signals triggered by HA are dependent on its recognition by the cell surface receptor CD44. In our studies we observe that overexpression of the eukaryotic translation initiation factor 4E (eIF4E) increases formation of cell surface protrusions resembling those produced by HAS3 upregulation. eIF4E regulates mRNA export &amp; translation through binding to the m7G cap of mRNA in the nucleus &amp; the cytoplasm, respectively. Strikingly, analysis of our two complimentary screens (immunoprecipitation of mRNAs bound to eIF4E in the nucleus, and mRNA export assay with ribavirin, an established inhibitor of eIF4E) that we carried out to identify candidate eIF4E mRNA export targets, showed that nearly all the enzymes involved in HA synthesis as well as CD44. Here, we hypothesize that eIF4E regulates HA synthesis &amp; CD44 overexpression to reshape the surface of cancer cells producing HA-rich microvillus-like pseudopods. Following validation of the positive hits, we used immunofluorescence staining with biotinylated HA binding protein (HABP) to determine the effect of eIF4E expression on protrusion formation. Our data revealed that eIF4E overexpression increased formation of HA-rich cell surface protrusions &amp; was correlated with increased invasiveness of eIF4E overexpressing cells in in vitro matrigel assay, compared to respective controls. Colocalization studies using confocal microscopy indicated that protrusions positively stained for CD44. Further, pharmacological inhibition with ribavirin and genetic knockdown of eIF4E (or HAS3/CD44 in the context of eIF4E overexpression) abrogated protrusion formation and decreased the invasion capacity of eIF4E overexpressing cells. Together, these data indicate that eIF4E modulates protrusion formation &amp; tumor invasion through coordinately regulating the export of RNAs in the CD44-HA regulon. To establish the clinical relevance of our findings in AML, we examined primary AML specimens with high eIF4E levels for expression of HA synthesizing enzymes, CD44 &amp; we directly measured HA using confocal methods. This comparison showed that these pathways are dysregulated in AML patients &amp; that their inhibition correlates with response to eIF4E inhibition by ribavirin. While HA has long been considered as a component of the tumor microenvironment required for the migration of cancer cells though interaction with surface CD44, our results provide a novel role for eIF4E in the motility &amp; bone marrow homing. These findings offer a rationale for potentially inhibiting tumor cell metastasis through the combinatorial inhibition of eIF4E and CD44 or HAS3. Citation Format: Hiba Zahreddine, Biljana Kraljacic-Culjkovic, Valbona Cali, Mark Lauer, Lucy Skrabanek, Nabilah Khan, Ronald Midura, Leandro Cerchietti, Vincent Hascall, Craig Jordan. eIF4E reshapes the surface of migrating AML cells through regulating Hyaluronic Acid synthesis &amp; CD44 expression [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4898. doi:10.1158/1538-7445.AM2017-4898

The direct action of hyaluronic acid on human U-937 and HL-60 cells — modification of native and model membranes

AbstractThe aim of the study was to investigate the effect of application of exogenous hyaluronic acid (HA) of various molecular weights on the cells of human immune system. Two cell lines HL-60 and U-937 with various ability to differentiation were chosen. HL-60 cells were differentiated to macrophages and granulocytes, whereas U-937 only to macrophages. For all investigated cell systems the most cytotoxic effect, indicated as a decrease of cell viability, was found at HA dose equal to 200 mg/L. However, greater effect was observed for differentiated cells and at longer exposure to HA. The possibility of HA interaction with both specific receptors and membrane lipids was tested by determination of biotin-labelled HA binding to cell surface and analysis of physicochemical parameters of model membranes.

Hypoxia Potentiates Anabolic Effects of Exogenous Hyaluronic Acid in Rat Articular Cartilage.

Hyaluronic acid (HA) is used clinically to treat osteoarthritis (OA), but its pharmacological effects under hypoxic conditions remain unclear. Articular chondrocytes in patients with OA are exposed to a hypoxic environment. This study investigated whether hypoxia could potentiate the anabolic effects of exogenous HA in rat articular cartilage and whether these mechanisms involved HA receptors. HA under hypoxic conditions significantly enhanced the expression of extracellular matrix genes and proteins in explant culture, as shown by real-time reverse transcription-polymerase chain reaction (RT-PCR), Western blotting, and dimethylmethylene blue (DMMB) assays. Staining with Safranin-O and immunohistochemical staining with antibody to type II collagen were also enhanced in pellet culture. The expression of CD44 was increased by hypoxia and significantly suppressed by transfection with siRNAs targeting hypoxia-inducible factor 1 alpha (siHIF-1α). These findings indicate that hypoxia potentiates the anabolic effects of exogenous HA by a mechanism in which HIF-1α positively regulates the expression of CD44, enhancing the binding affinity for exogenous HA. The anabolic effects of exogenous HA may increase as OA progresses.

Viscosupplementation with intra-articular hyaluronic acid for hip disorders. A systematic review and meta-analysis.

Hip joint diseases are common in adult population and their prevalence increases with age. Osteoarthritis, rheumatoid arthritis and femoroacetabular impingement are the most common chronic diseases in the hip joint. Viscosupplementation with exogenous hyaluronic acid (HA) is one of the most widely used conservative treatment aiming to improve synovial fluid properties and to decrease pain. There is no global consensus on the type of HA, method of injection and frequency, or on its efficacy in hip joint. We selected published data in English in the PubMed and Google Scholar electronic databases up to March 2016 about hyaluronic acid injections in hip disorders. 26 articles were included following the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines. There is a lack of standardization of HA injections for hip conditions. Our results suggest that this is the best conservative therapy before surgery and it can act on pain relief and function however there is no evidence to prove its ability to modify the morphological structure of the pathological hip and the natural history of the disease. There are few data about the use of HA in other hip disorders rather than osteoarthritis. The most relevant evidence seems to show the utility of HA injections in improving synovial inflammation, but only a few studies have been conducted. I.