Polysulfated Glycosaminoglycans Research Articles

Equine Musculoskeletal Pathologies: Clinical Approaches and Therapeutical Perspectives-A Review.

Musculoskeletal injuries such as equine osteoarthritis, osteoarticular defects, tendonitis/desmitis, and muscular disorders are prevalent among sport horses, with a fair prognosis for returning to exercise or previous performance levels. The field of equine medicine has witnessed rapid and fruitful development, resulting in a diverse range of therapeutic options for musculoskeletal problems. Staying abreast of these advancements can be challenging, prompting the need for a comprehensive review of commonly used and recent treatments. The aim is to compile current therapeutic options for managing these injuries, spanning from simple to complex physiotherapy techniques, conservative treatments including steroidal and non-steroidal anti-inflammatory drugs, hyaluronic acid, polysulfated glycosaminoglycans, pentosan polysulfate, and polyacrylamides, to promising regenerative therapies such as hemoderivatives and stem cell-based therapies. Each therapeutic modality is scrutinized for its benefits, limitations, and potential synergistic actions to facilitate their most effective application for the intended healing/regeneration of the injured tissue/organ and subsequent patient recovery. While stem cell-based therapies have emerged as particularly promising for equine musculoskeletal injuries, a multidisciplinary approach is underscored throughout the discussion, emphasizing the importance of considering various therapeutic modalities in tandem.

Prescribing practices of veterinarians in the treatment of chronic musculoskeletal pain in cats.

Despite the high prevalence and increasing awareness of chronic musculoskeletal pain in cats, approved treatment options are completely lacking in the USA, and few other options have sufficient safety and efficacy data. Knowledge of current prescribing practices should inform future research of putative therapies. We aimed to determine which drug and non-drug therapies were being used by general practitioners for the treatment of musculoskeletal pain in cats and to understand demographic influences on prescribing practices. We distributed a survey to 36,676 veterinarians who were members of the Veterinary Information Network in January 2017. Within 3 weeks, 1056 practitioners completed the survey. The survey included demographic and background information, questions on prescribing frequency and dosing regimen of 13 drug and non-drug therapies and questions on preferred medication formulations and dosing frequencies. Descriptive statistics were used, as well as χ2 testing to evaluate relationships between demographic variables and prescription practices. Gabapentin was prescribed most frequently (71% of respondents), followed by joint supplements (67.8%), meloxicam (64.0%), opioids (62.6%), fish oil (62.1%) and polysulfated glycosaminoglycans (61.9%). Years in practice appeared to influence prescribing habits, with practitioners graduated for >20 years prescribing glucocorticoids more frequently than other age groups ( P = 0.0002), whereas recent graduates (<1 year) reported prescribing therapies less frequently across all categories. These results show a contrast between therapies prescribed by practitioners and what is supported by evidenced-based literature. Future research evaluating the safety and efficacy of gabapentin should be prioritised.

HEMORRHAGIC DIATHESIS IN AVIAN SPECIES FOLLOWING INTRAMUSCULAR ADMINISTRATION OF POLYSULFATED GLYCOSAMINOGLYCAN

Polysulfated glycosaminoglycans (PSGAGs) have been used for decades in a variety of species for the management of osteoarthritic pain. However, reports on the use of PSGAGs in avian species are scarce. In domestic cats and dogs, PSGAG injections have caused prolongation of clotting times but are considered to be an efficacious drug with a wide margin of safety. This publication documents four cases of fatal coagulopathies in different avian species (one coraciiforme, two raptors, and one psittacine) following the intramuscular administration of PSGAG. All affected birds received varying dosages and dosing intervals of PSGAG. Three of the four birds experienced fatal hemorrhage into the pectoral muscle, while the fourth bled continuously from the injection site. Only one bird had chronic, severe pre-existing disease; the remainder were being managed for osteoarthritis. This report highlights the importance of species-specific dosing of PSGAG and warrants further investigation into the etiopathogenesis of this process.

Canine Hip Dysplasia: Reviewing the Evidence for Nonsurgical Management

To systematically review the evidence available for nonsurgical management of hip dysplasia (HD). Literature review. Databases (Pubmed, Veterinary Information Network) were searched for clinical studies on nonsurgical management of HD in dogs. The evidence in each study was reviewed and assigned a score (I-IV) based on previously reported levels of evidence. Fourteen articles were identified that met the inclusion criteria, including 3 Level IV, 4 Level III, and 7 Level II studies. Methods of nonsurgical management reviewed included: activity restrictions, weight management, acupuncture, modulation of joint disease by polysulfated glycosaminoglycans, mesenchymal stem cell therapy, and extra corporeal shock wave therapy. Weight management is an effective and important component of managing dogs with HD and associated osteoarthritis. Techniques that modulate the progression of joint disease may also be beneficial for treating dogs with HD. Further studies are needed to investigate other methods of managing HD such as hydrotherapy and physical rehabilitation.

Effect of Prophylactic Intramuscular Administration of Polysulfated Glycosaminoglycan on Developmental and Traumatic Joint Injuries in Thoroughbred Foals

The objective of this study was to determine whether a prophylactic regimen of intramuscular polysulfated glycosaminoglycans (PSGAG), administered to growing Thoroughbred foals, would significantly reduce the incidence or severity of osteochondritis dissecans (OCD) or traumatic joint lesions that had been found in untreated foals (historical controls) from the same farm during the preceding 3 years. The number of PSGAG-treated foals with clinically significant joint lesions was 44.1% lower than the number of foals with OCD/joint injuries that had been found in the untreated historical controls. Surgical intervention of the treated foals declined 87.7% from the incidence on the farm during the preceding 3 years. Although the incidence of OCD lesions of the hock, stifle, and fetlocks in treated foals was not significantly different from that found in the matched controls, the lesions in the PSGAG-treated foals resolved without surgery. These results suggest that PSGAG has the potential to improve the clinical course of osteochondrosis (OC) and OCD. Additional studies are underway to determine whether a shorter or more targeted prophylactic regimen will provide a similar benefit.

Systematic review of clinical trials of treatments for osteoarthritis in dogs

To identify and critically evaluate the quality of evidence of the most commonly used pharmacologic, nutraceutical, and purported slow-acting drugs of osteoarthritis for the management of osteoarthritis in dogs by use of the FDA's evidence-based medicine scoring system. Systematic review. 16 clinical trials. A broad bibliographic search was performed prior to May 2006. Inclusion criteria focused on prospective trials evaluating commonly used medical treatment interventions for the management of osteoarthritis in dogs and published in peer-reviewed journals. The analysis consisted of the following: study design rating, quality factor rating, quantity rating, consistency rating, relevance to disease risk reduction rating, and cumulative strength of evidence ranking. 4 trials evaluating meloxicam were rated as type I. Three trials evaluating carprofen were rated as type I, and 2 trials were rated as type III. One trial evaluating each of the following agents was rated as type 1: etodolac; P54FP; polysulfated glycosaminoglycan; and a combination of chondroitin sulfate, glucosamine hydrochloride, and manganese ascorbate. Two trials evaluating pentosan polysulphate and 2 trails evaluating green-lipped mussels were rated as type I. One trial evaluating hyaluronan was rated as type III. A high level of comfort exists for meloxicam that the claimed relationship is scientifically valid and that its use is clinically efficacious for the treatment of osteoarthritis in dogs. A moderate level of comfort exists for carprofen; etodolac; pentosan polysulphate; green-lipped mussels; P54FP; polysulfated glycosaminoglycans; and a combination of chondroitin sulfate, glucosamine hydrochloride, and manganese ascorbate. An extremely low level of comfort exists for hyaluronan.

Topical effect of various agents on gelatinase activity in the tear film of normal dogs

To evaluate the topical effect of various agents, currently used in the treatment of melting ulcers, on gelatinase activity present in the tear film of normal dogs. Eight normal adult beagles. Each animal received the following agents: cyclosporine A 1%, N-acetylcysteine 10%, ciprofloxacin 0.3%, ethylenediaminetetraacetic acid (EDTA) 1%, doxycycline 0.001%, polysulfated glycosaminoglycans (PSGAG) 5%, autoserum, and artificial tears during a 48-h period following a Latin square design. Tear samples were collected with micro-capillary pipettes following a corneal surface irrigation of each eye with sterile saline on four different occasions. Basal and total gelatinase activities were evaluated by optical density after processing in a commercial gelatinase activity assay. From the optical density ratio, a semi-quantitative measure of gelatinase activity was obtained. Basal and total activities were measured in all samples. The lowest total gelatinase activity, representing a percent decline in the enzyme activity although not significant, was observed 1 h after the last treatment in seven out of the eight ophthalmic agents; EDTA (68%), ciprofloxacin (76%), cyclosporine A (68%), doxycycline (47%), artificial tears (26%), PSGAG (25%), and N-acetylcysteine (20%). However, only the reduction observed with EDTA 6 h after the last treatment was significantly lower compared to the reduction observed with the artificial tears. This study indicated that only EDTA was able to significantly reduce the gelatinase activity in a persistent manner in the tear film of normal canine eyes. Further studies will be required to evaluate the effect of EDTA under ulcerative conditions and to more accurately ascertain the potential in vivo effect of the other agents.

Uptake and distribution of tritium-labeled polysulfated glycosaminoglycan in serum, urine, and superficial digital flexor tendon of rabbits after intramuscular administration.

To determine concentrations of tritiated polysulfated glycosaminoglycans (3H-PSGAG) in serum, urine, and the superficial digital flexor tendon (SDFT) of rabbits after IM administration and molecular weight of 3H-PSGAG recovered from the SDFT. Twenty-five 12-week-old New Zealand White rabbits. Rabbits were given a single dose of 3H-PSGAG (1.1 mg/kg [70 mCi of specific activity/kg] of body weight) IM. At each of 5 sample times (2, 24, 48, 96, and 192 hours), 5 rabbits were randomly selected and sedated, and blood and urine samples were collected. Rabbits were then euthanatized, and the SDFT were immediately harvested from the hind limbs. Scintillation spectrometry was used to detect concentration of 3H-PSGAG in fluid and tissue samples. Gel-filtration chromatography was used to determine molecular weight of recovered 3H-PSGAG. Mean concentrations of 3H-PSGAG in SDFT, serum, and urine were greatest 2 hours after administration. Tritiated PSGAG could be detected in all samples collected 192 hours after administration. Gel-filtration chromatography confirmed that 3H-PSGAG detected in SDFT samples was high molecular weight PSGAG. Results indicate that PSGAG is distributed to the SDFT, serum, and urine after IM administration in rabbits. Further study is needed to determine whether the same is true in horses and to determine what effect, if any, PSGAG has on inflammation of the SDFT.

Structure–Function Relations of Antithrombin III–Heparin Interactions as Assessed by Biophysical and Biological Assays and Molecular Modeling of Peptide–Pentasaccharide-Docked Complexes

The serine proteinase inhibitor antithrombin III (ATIII) is a key regulatory protein of intrinsic blood coagulation. ATIII attains its full biological activity only upon binding polysulfated glycosaminoglycans, such as heparin. Peptide K121–A134, based on the sequence of ATIII in the D helix region, was previously shown by us (Tyler-Crosset al., Protein Sci.3, 620–627, 1994) to encompass part (or all) of the purported high-affinity heparin binding region of ATIII. A series of peptide analogs has now been prepared whose sequences are identical to K121–A134except that single cationic residues of K121–A134have been successively replaced with Ala. In one case, the Arg residue of the reference peptide corresponding to R129of ATIII has been replaced by Gln (R129ΔQ peptide), thus mimicking the naturally occurring mutant protein, ATIII Geneva. The heparin affinity of all peptides was quantitated by isothermal titration calorimetry and by peptide/ATIII competition binding assays. Replacement of any single cationic residue with Ala had a deleterious effect on heparin binding. The greatest reduction in heparin affinity (more than 30-fold) was observed with the R129ΔQ peptide (KD= 1.5 ± 0.06 μMvs 51 ± 2 nMfor the reference peptide, K121–A134). Furthermore, each of the Ala-replacement peptides was a less-effective inhibitor of ATIII–heparin complex formation than the reference peptide. The poorest inhibitor was the R129ΔQ peptide which showed nearly 30% decrease in inhibition potency (60% inhibition at 100 μMpeptide vs 90% inhibition with the reference peptide). The relative heparin affinities of the peptides measured by biological assay were the same as determined by titration calorimetry. Consequently, we modeled the complexes formed between the pentasaccharide unit structure and the R129ΔQ peptide or the reference peptide, K121–A134. In the “docked” complex, the assumed conformation of K121–A134permitted juxtaposition of the cationic residues of the peptide with functional anionic groups of the pentasaccharide known to be involved in binding. A docked complex could also be formed between the R129ΔQ peptide and the pentasaccharide, but misalignment of critical peptide and saccharide functional groups was observed. The structure of the R129ΔQ–pentasaccharide complex was highly irregular because F123and Y131were completely surface exposed, likely yielding an unfavorable structure in aqueous solution. The observations from molecular modeling allow us to suggest that ATIII Geneva displays decreased heparin binding affinity due to its inability to form a productive binding complex in which essential electrostatic contacts are made between suitably juxtaposed saccharide anionic functional groups and cationic amino acid side chains.

IFN-gamma-induced MHC class II gene expression is suppressed in endothelial cells by dextran sulfate.

IFN-gamma-activated endothelial cells actively participate in initiating immune responses by interacting with immunocompetent cells via class II MHC proteins. In this study, dextran sulfate, a synthetic heparin analogue, was shown to selectively inhibit IFN-gamma-induced surface expression of HLA-DR molecules by human umbilical cord vascular endothelial cells, but not other cytokine-induced molecules such as ELAM-1 or ICAM-1. Inhibition occurred regardless of whether dextran sulfate was added 24 h before, at the same time as, or 24 h after IFN-gamma stimulation of cells. In addition, both high (500 kDa) and low (5 kDa) molecular mass dextran sulfate molecules were able to block class II expression, whereas treating cells with naturally occurring polysulfated glycosaminoglycans such as heparin, heparan, and chondroitin sulfate did not produce any suppressive effects. Radiolabeling of cells with [35S]methionine followed by radioimmunoprecipitation using anti-HLA-DR alpha mAb demonstrated that biosynthesis of class II proteins was specifically blocked. RT-PCR and Southern blotting were utilized to examine transcription of the HLA-DR alpha gene and demonstrated an absence of HLA-DR alpha mRNA from dextran sulfate-treated and IFN-gamma-induced cells. Dextran sulfate also prevented transcription of the gene encoding CIITA, a transactivator protein required for IFN-gamma-inducible expression of class II genes. Thus, dextran sulfate apparently inhibited this step or an earlier one in the intracellular signaling pathway for IFN-gamma in human endothelial cells, subsequent to IFN-gamma binding to its cell surface receptor.

Hyaluronidase induces murine L929 fibrosarcoma cells resistant to tumor necrosis factor and Fas cytotoxicity in the presence of actinomycin D.

Actinomycin D (ActD) enhances the potency of tumor necrosis factor-alpha (TNF-alpha) in killing cancer cells. However, it is determined in this study that murine L929 fibrosarcoma cells, when pretreated with bovine testicular hyaluronidase for 12-24 h, became resistant to the cytotoxic effect of TNF-alpha in the presence of DNA intercalators, such as ActD, doxorubicin, and daunorubicin. Monoclonal anti-Fas antibody-mediated apoptosis in the presence of ActD was also blocked in hyaluronidase-pretreated L929 cells. Hyaluronidase failed to up- or downregulate the expression of apoptosis regulatory proteins, including Bcl-2, Bcl-xL, ICH-1, and TIAR, suggesting that these proteins were not involved in the hyaluronidase-induced resistance to TNF/ActD. A semisynthetic polysulfated hyaluronic acid (HA) inhibited the increased TNF/ActD resistance, whereas unmodified HA, dextran sulfate, and naturally polysulfated glycosaminoglycans had no effect. Evidence is provided here that the induced resistance is related to serum fetuin and a novel intracellular 35-kDa TNF-binding protein (intra TBP). Under serum-free conditions, L929 became refractory to TNF/ActD cytotoxicity and hyaluronidase reversed the resistance. Exogenous fetuin increased L929 cell spreading and proliferation, and restored hyaluronidase-induction of TNF/ActD resistance in these serum-starved cells. Hyaluronidase failed to reduce the expression of TNF-receptors and their binding of TNF-alpha. However, binding and Western-blotting analyses revealed that hyaluronidase downregulated the intra-TBP. Overall, these observations suggest that serum fetuin and intraTBP are involved in the hyaluronidase induction of TNF/ ActD resistance.

Synthetic polysulfated hyaluronic acid is a potent inhibitor for tumor necrosis factor production

Based on the premise that naturally occurring glycosaminoglycans could serve as building blocks for synthesizing nontoxic drugs for suppression of tumor necrosis factor (TNF) production by inflammatory cells, we have chemically modified hyaluronic acid (HA) and tested its effects in blocking TNF-alpha and TNF-beta production in vitro. HA was chosen mainly for its structural simplicity, nonimmunogenicity, and readiness for chemical modifications. When HA was chemically polysulfated to a sulfate/hexosamine molar ratio of 3.9, the sulfated HAs was shown to be a potent inhibitor of TNF-alpha production in lipopolysaccharide (LPS)- or interferon-gamma-activated THP-1 cells. For example, a concentration of HAs as low as 10 ng/ml reduced TNF-alpha production in LPS-activated THP-1 cells more than 50%, whereas achieving a similar extent of reduction required 50 micrograms/ml native HA. By decreasing the extent of polysulfation, the inhibitory effect of HAs on TNF-alpha production was diminished. Other chemical modifications, including deacetylation, thiolation, or reduction of the carboxylic groups, could not increase the efficacy of HA in suppression of TNF-alpha production. Naturally polysulfated glycosaminoglycans, such as chondroitin sulfates, keratan sulfate, heparan sulfate, and heparin, failed to inhibit TNF-alpha production. HAs also restricted TNF-beta (lymphotoxin) secretion in an Epstein-Barr virus-transformed B cell line, Roha-9, which constitutively produces TNF-beta. HAs had no inhibitory effect on the proliferation of THP-1 or Roha-9 cells, which would account for the reduced TNF-alpha or TNF-beta production. Furthermore, time-course metabolic labeling studies revealed that HAs could not restrict overall protein synthesis and secretion in THP-1 cells. However, HAs increased complement C1q secretion in THP-1 in a dose-dependent manner, but it had no effect on biosynthesis of complement C1 inhibitor, factor D, and Fc gamma receptor type II (Fc gamma RII). These results indicate that HA, selectively restricts the production of TNF-alpha, TNF-beta, and probably several other protein species.

Diagnosis and Treatment of the Navicular Syndrome in Horses

Navicular syndrome can be treated in a variety of ways. This is related to the fact that it has a variety of causes. Prognostically, most horses will improve with treatment. One can expect about 50 per cent of the horses to become useably sound for 1 year, no matter what treatment is used. The disease is progressive, and affected horses eventually will need to be retired because of lameness. The author's therapeutic approach is to utilize shoeing as the primary therapy. Shoeing is performed to correct structural problems and to ensure that shoeing is physiologically sound. Nonsteroidal anti-inflammatory drugs are not used unless radical changes have been made in the shoeing. In cases of confirmed distal interphalangeal joint synovitis, either sodium hyaluronate or polysulfated glycosaminoglycans will be used in conjunction with shoeing. In cases where decreased circulation is documented, isoxsuprine hydrochloride will be administered if shoeing alone has not improved the horse within 6 weeks. If therapy does not improve the horse within 6 to 12 weeks, palmar digital neurectomy is recommended.

Review of pathogenesis and treatment of degenerative joint disease.

Treatment of degenerative joint disease (osteoarthritis) is discussed in relationship to pathogenesis of five clinical entities, defined to facilitate discussion:-1) occurring in high motion joints and associated with synovitis; 2) associated with low motion joints; 3) "non-progressive" articular cartilage erosion; 4) secondary to other identified problems (intra-articular fractures, ligamentous damage, wounds, septic arthritis, osteochondrosis); and 5) chondromalacia of the patella. In addition to direct damage to articular cartilage, synovitis and capsulitis and depletion of matrical glycosaminoglycans and proteoglycans play important roles. The natural healing response in damaged articular cartilage is inadequate. The principles of treatment are divided into: 1) prevention or treatment of primary causes. These include the effects of track surface, shoeing, and the appropriate resolution of intra-articular fractures, septic arthritis and osteochondritis dissecans. 2) treatment of active soft tissue disease contributing to articular cartilage degeneration, including rest, physical therapy, synovectomy and administration of anti-inflammatory drugs, sodium hyaluronate and polysulfated glycosaminoglycans. 3) attempts at treatment of articular cartilage loss or degeneration, including articular cartilage curettage, subchondral bone drilling, osteophyte removal, articular cartilage grafting and arthrodesis.

Identification of binding site for heparin and other polysulfated glycosaminoglycans on human thrombocytes.

Heparin and other polysulfated glycosaminoglycans (GAGPS) can evoke an immune response when complexed to the surface of platelets in vivo. We have previously reported on the detection and characteristics of such antibodies leading to thrombocytopenia. Here we report data concerning the biochemical characterization of the heparin-/GAGPS-binding site on human thrombocytes. By means of heparin affinity chromatography, GAGPS chromatography and subsequent 'Western blotting' of specifically eluted proteins we were able to detect two proteins of apparent molecular mass of 207 and 170 kilodaltons, which could be reduced to 57 and 142 kilodaltons, respectively. The 170-kilodalton glycoprotein was stained by the carbohydrate specific PAS-reagent and by surface labeling with 3H. The second molecule binding to heparin (GAGPS)-Sepharose was detected by platelet surface labeling with 125I, protein-specific staining Coomassie brilliant blue and PAS staining. Binding studies using enzyme-labeling techniques with five monoclonal antibodies (MCA) against platelet surface proteins GPI, GPIIb/IIIa complex, GPIIIa and CRI (C3b receptor) revealed specific interaction of the 170-kilodalton heparin-binding glycoprotein with an MCA to GPI, suggesting identity or immunological cross-reactivity of these two moieties. No specific reactions were observed between the 207-kilodalton glycoprotein and any of the MCA tested. For further evaluation of the antigenicity of separated platelet proteins, an ELISA in microtiter plates was developed, and a cellular sandwich ELISA for serological detection of antibodies against whole native thrombocytes or GAGPS-derivatized proteins was set up.(ABSTRACT TRUNCATED AT 250 WORDS)

Detection of antibodies interacting with glycosaminoglycan polysulfate in patients treated with heparin or other polysulfated glycosaminoglycans.

A new method has been developed for the demonstration of the antibody nature of a factor occurring in some patients during treatment with heparin or other polysulfated glycosaminoglycans (GAGPS). So far, antibodies reacting with GAGPS, except heparin, have not been described. We used heparin- or GAGPS-coated normal blood group O thrombocytes for the detection of suspected antibodies. After addition of patients' plasma or serum to a suspension of purified heparin- or GAGPS-coated platelets, agglutination was determined microscopically. In indirect immunofluorescence, positive staining of agglutinated platelets was observed with fluorescein isothiocyanate anti-Ig and anti-IgG conjugates, but not with anti-IgA or anti-IgM, with one exception in which IgM antibodies were also involved. Binding of complement was also shown using a tetramethylrhodamin isothiocyanate-labelled anti-C3 conjugate in double-staining experiments. Fibronectin could be excluded as a possible factor responsible for thrombocyte agglutination.