Highly sulfated glycosaminoglycans inhibit aggrecanase degradation of aggrecan by bovine articular cartilage explant cultures.

The catabolism of 35S-labeled aggrecan and loss of tissue glycosaminoglycans was investigated using bovine articular cartilage explant cultures maintained in medium containing 10(-6) M retinoic acid or 40 ng/ml recombinant human interleukin-1alpha (rHuIL-1alpha) and varying concentrations (1-1000 microg/ml) of sulfated glycosaminoglycans (heparin, heparan sulfate, chondroitin 4-sulfate, chondroitin 6-sulfate, dermatan sulfate and keratan sulfate) and calcium pentosan polysulfate (10 microg/ml). In addition, the effect of the sulfated glycosaminoglycans and calcium pentosan polysulfate on the degradation of aggrecan by soluble aggrecanase activity present in conditioned medium was investigated. The degradation of 35S-labeled aggrecan and reduction in tissue levels of aggrecan by articular cartilage explant cultures stimulated with retinoic acid or rHuIL-1alpha was inhibited by heparin and heparan sulfate in a dose-dependent manner and by calcium pentosan polysulfate. In contrast, chondroitin 4-sulfate, chondroitin 6-sulfate, dermatan sulfate and keratan sulfate did not inhibit the degradation of 35S-labeled aggrecan nor suppress the reduction in tissue levels of aggrecan by explant cultures of articular cartilage. Heparin, heparan sulfate and calcium pentosan polysulfate did not adversely affect chondrocyte metabolism as measured by lactate production, incorporation of [35S]-sulfate or [3H]-serine into macromolecules by articular cartilage explant cultures. Furthermore, heparin, heparan sulfate and calcium pentosan polysulfate inhibited the proteolytic degradation of aggrecan by soluble aggrecanase activity. These results suggest that highly sulfated glycosaminoglycans have the potential to influence aggrecan catabolism in articular cartilage and this effect occurs in part through direct inhibition of aggrecanase activity.     

Sodium pentosan polysulfate resulted in cartilage improvement in knee osteoarthritis - An open clinical trial-

Background

Pentosan polysulfate sodium (pentosan) is a semi-synthetic drug manufactured from beech-wood hemicellulose by sulfate esterification of the xylopyranose hydroxyl groups. From in vitro and animal model studies, pentosan has been proposed as a disease modifying osteoarthritis drug (DMOAD). The objective of this study was to assess the efficacy, safety, and patient satisfaction in patients with mild radiographic knee osteoarthritis (OA) findings and OA-associated symptoms and signs.

Methods

Twenty patients were assessed clinically at Nagasaki University Hospital. The radiographic indications of OA were grade 1 to 3 using the Kellgren-Lawrence Grading System (K/L grade). Pentosan used in this study was manufactured and supplied in sterile injectable vials (100 mg/ml) by bene GmbH, Munich, Germany. The study was a single-center, open-label trial. Treatment consisted of 6 weekly subcutaneous injections (sc) of pentosan (2 mg/kg). Patients were clinically assessed at entry and 1 to 8, 11, 15, 24 & 52 weeks post treatment. The results were analyzed using one way ANOVA and Dunnett's method.

Results

Hydrarthroses were reduced quickly in all cases. The clinical assessments, i.e., knee flexion, pain while walking, pain after climbing up and down stairs, etc, were improved significantly and these clinical improvements continued for almost one year. The dose used in this study affected the blood coagulation test, but was within safe levels. Slightly abnormal findings were noted in serum triglycerides.

Conclusions

Pentosan treatment in twenty patients with mild knee OA seemed to provide improvements in clinical assessments and C2C level of cartilage metabolism.

Trial Registration

UMIN Clinical Trials Registry (UMIN-CTR) UMIN000002790     

Antithrombotic therapy in patients with known risk factors for thromboembolism

In about 50% of the cases of spontaneous deep vein thrombosis a congenital deficiency of an inhibitor of coagulation or an insufficient fibrinolytic mechanism can be detected. In arterial thromboembolism a connection with hyperactive platelets or with a diminished availability of tissue plasminogen activator can be found in about 70%. However, in these cases the defect which provokes thrombosis is mostly acquired and is connected with hyperlipidemia and/or with atherosclerotic alterations of the vessel wall. A study on patients with thromboembolic tendency and detectable risk factors was carried out. A total of 470 patients could be observed for 2 years under an adequate antithrombotic prophylaxis. The occurrence of thromboembolic episodes 2 years prior to prophylaxis and 2 years under prophylaxis was compared. In venous cases thrombosis could be controlled almost completely by coumarins when the underlying cause was a deficient plasmatic inhibitor. In patients with diminished fibrinolysis there was only a partial effect of oral anticoagulants. A better result could be obtained when pentosan polysulfate was administered. In arterial thromboembolism the results of prophylaxis were less convincing. The efficacy of ASA in patients with an increased platelet function was only moderate. In addition, ASA hat to be discontinued in about 20% of the patients because of gastrointestinal problems. Pentosan polysulfate in patients with a diminished fibrinolytic capacity had a fairly good effect and resulted in a 60% reduction of thromboembolic manifestations. It is shown that an exact diagnosis of the underlying deficiency which is likely to cause thrombosis can also improve the efficacy and the specificity of prophylaxis.     

Pentosan polysulfate protects brain endothelial cells against bacterial lipopolysaccharide-induced damages

Peripheral inflammation can aggravate local brain inflammation and neuronal death. The blood-brain barrier (BBB) is a key player in the event. On a relevant in vitro model of primary rat brain endothelial cells co-cultured with primary rat astroglia cells lipopolysaccharide (LPS)-induced changes in several BBB functions have been investigated. LPS-treatment resulted in a dose- and time-dependent decrease in the integrity of endothelial monolayers: transendothelial electrical resistance dropped, while flux of permeability markers fluorescein and albumin significantly increased. Immunostaining for junctional proteins ZO-1, claudin-5 and beta-catenin was significantly weaker in LPS-treated endothelial cells than in control monolayers. LPS also reduced the intensity and changed the pattern of ZO-1 immunostaining in freshly isolated rat brain microvessels. The activity of P-glycoprotein, an important efflux pump at the BBB, was also inhibited by LPS. At the same time production of reactive oxygen species and nitric oxide was increased in brain endothelial cells treated with LPS. Pentosan polysulfate, a polyanionic polysaccharide could reduce the deleterious effects of LPS on BBB permeability, and P-glycoprotein activity. LPS-stimulated increase in the production of reactive oxygen species and nitric oxide was also decreased by pentosan treatment. The protective effect of pentosan for brain endothelium can be of therapeutical significance in bacterial infections affecting the BBB.     

Sulfated glycans and elevated temperature stimulate PrP(Sc)-dependent cell-free formation of protease-resistant prion protein

A conformational conversion of the normal, protease- sensitive prion protein (PrP-sen or PrP(C)) to a protease-resistant form (PrP-res or PrP(Sc)) is commonly thought to be required in transmissible spongiform encephalopathies (TSEs). Endogenous sulfated glycosaminoglycans are associated with PrP-res deposits in vivo, suggesting that they may facilitate PrP-res formation. On the other hand, certain exogenous sulfated glycans can profoundly inhibit PrP-res accumulation and serve as prophylactic anti-TSE compounds in vivo. To investigate the seemingly paradoxical effects of sulfated glycans on PrP-res formation, we have assayed their direct effects on PrP conversion under physiologically compatible cell-free conditions. Heparan sulfate and pentosan polysulfate stimulated PrP-res formation. Conversion was stimulated further by increased temperature. Both elevated temperature and pentosan polysulfate promoted interspecies PrP conversion. Circular dichroism spectropolarimetry measurements showed that pentosan polysulfate induced a conformational change in PrP-sen that may potentiate its PrP-res-induced conversion. These results show that certain sulfated glycosaminoglycans can directly affect the PrP conversion reaction. Therefore, depending upon the circumstances, sulfated glycans may be either cofactors or inhibitors of this apparently pathogenic process.     

Saccharide anions as inhibitors of the malaria parasite

The asexual erythrocytic stage of Plasmodium falciparum was grown in culture in the presence or absence of glycoconjugate polyanions of varying structure, size and substitutions. Heparin, dextran sulfate, fucoidan and pentosan polysulfate had antimalarial IC50 values between one and 11 μg ml−1. Constituent heparin disaccharides were ineffective against the malaria parasite and desulfation from either the O- or N-substitution sites of heparin or reduction of the uronic acid carboxyl group neutralized the antimalarial response to varying degrees. Immobilization of heparin onto agarose beads still permitted antimalarial activity suggesting that parasite uptake of the glycoconjugate is not required for inhibition. Accordingly, it is concluded that invasion of free parasites into the erythrocytes was inhibited rather than parasite maturation within the red cell. Merozoite surface antigen-1 was apparently prevented from binding to human erythrocytes in the presence of highly sulfated polyanions and, in a dose-dependent fashion, heparin. Abbreviations: MSA-1, merozoite surface antigen-1 This revised version was published online in November 2006 with corrections to the Cover Date.     

Effect of Ultrasound on Structure and Functional Properties of Antithrombin III and Proteins of PPSB Complex

A combination of gel-permeation HPLC, affinity chromatography on heparin-Sepharose, gel electrophoresis, and estimation of inhibitory activity showed that effect of low-frequency ultrasound (26 W/cm(2), 37 degrees C, pH 7.4) on homogeneous antithrombin III was accompanied by formation of aggregates and a latent form of serpin. Heparin and pentosan polysulfate stabilized antithrombin III; this resulted in decrease in ultrasonic-induced formation of the aggregate and latent forms. The influence of ultrasound was not accompanied by significant changes in the contents of non-activated blood coagulation factors in the PPSB complex.     

Assessment of prospective preventive therapies for chronic wasting disease in mule deer

We compared prion infection rates among mule deer (Odocoileus hemionus) receiving pentosan polysulfate, tannic acid, tetracycline HCl, or no treatment 14 days before to 14 days after (dpi) oral inoculation with tonsil tissue homogenate. All deer were infected, but the rapid disease course (230-603 dpi) suggested our challenge was overwhelming.     

Interstitial cystitis: characterization and management of an enigmatic urologic syndrome

The enigmatic urologic condition known as interstitial cystitis has an estimated prevalence of 0.01% to 0.50% of the female population. Its etiology is unknown but may involve microbiologic, immunologic, mucosal, neurogenic, and/or other, as yet undefined, agents. There is no gold standard for the diagnosis of interstitial cystitis; rather, it is a diagnosis of exclusion. It is impossible to provide a purely evidence-based treatment strategy, but review of available evidence suggests that conservative supportive therapy (including diet modification); oral treatment with pentosan polysulfate, amitriptyline, hydroxyzine, or cimetidine; and intravesical treatments with heparinoids, dimethyl sulfoxide, alkalized lidocaine, or bacille Calmette-Guérin may be effective in some patients.     

Calcium pentosan polysulfate directly inhibits enzymatic activity of ADAMTS4 (aggrecanase-1) in osteoarthritic chondrocytes

Aggrecanases that include ADAMTS1, 4, 5, 8, 9 and 15 are considered to play key roles in aggrecan degradation in osteoarthritic cartilage. Here we demonstrate that calcium pentosan polysulfate (CaPPS) directly inhibits the aggrecanase activity of ADAMTS4 without affecting the mRNA expression of the ADAMTS species in interleukin-1alpha-stimulated osteoarthritic chondrocytes. Synthetic peptides corresponding to specific regions of the thrombospondin type 1 repeat, cysteine-rich or spacer domain of ADAMTS4 inhibit the binding to immobilized CaPPS. These data suggest that CaPPS could function as chondroprotective agent for the treatment of osteoarthritis by inhibition of ADAMTS4 through interaction with the C-terminal ancillary domain.     

Clinical experience with heparinoid sodium pentosan polysulfate (SP54) in the treatment of cerebral ischemia

Fifty-five patients suffering from acute cerebral ischaemia were treated with pentosan polysulphate (SP54) intravenously and subsequently orally. The control group consisting of 35 patients received the infusion solution and vasodilator drugs. The statistically significant changes appearing following infusion of SP54 include the following parameters: haematocrit, blood viscosity and cholesterol concentrations. In individual cases the euglobulin lysis time was shortened within two hours after the beginning of the infusion. As demonstrated by the statistically reliable results in patients treated with SP54 the neurological symptoms improved significantly within the first two weeks after the onset of the disease compared to the control groups. Based on case reports, recommendations are given for the indication and contraindication of SP54 treatment in cerebral ischaemia.     

Pentosan Polysulfate: Oral Versus Subcutaneous Injection in Mucopolysaccharidosis Type I Dogs

BackgroundWe previously demonstrated the therapeutic benefits of pentosan polysulfate (PPS) in a rat model of mucopolysaccharidosis (MPS) type VI. Reduction of inflammation, reduction of glycosaminoglycan (GAG) storage, and improvement in the skeletal phenotype were shown. Herein, we evaluate the long-term safety and therapeutic effects of PPS in a large animal model of a different MPS type, MPS I dogs. We focused on the arterial phenotype since this is one of the most consistent and clinically significant features of the model.ConclusionsPPS treatment led to reductions of pro-inflammatory cytokines and GAG storage in urine and tissues of MPS I dogs, which were most evident after subQ administration. SubQ administration also led to significant cytokine reductions in the CSF. Both treatment groups exhibited markedly reduced carotid and aortic inflammation, increased vessel integrity, and improved histopathology. We conclude that PPS may be a safe and useful therapy for MPS I, either as an adjunct or as a stand-alone treatment that reduces inflammation and GAG storage.     

Dysregulation of CXCL9 and reduced tumor growth in Egr-1 deficient mice

Advances in the treatment of chronic lymphocytic leukemia (CLL) have improved initial overall response (OR) rates, complete response (CR) rates and progression free survival (PFS). Despite these advances, CLL remains incurable with standard therapies. Thus, there remains a need for more effective therapies in both the upfront and relapsed setting, particularly for patients with high-risk cytogenetic abnormalities such as del(11q22) and del(17p13). The 2008 American Society of Hematology (ASH) Annual Meeting featured several presentations which highlighted the ongoing clinical advances in CLL. The benefit of adding rituximab to purine analog therapy in the upfront setting was demonstrated by a large randomized study which showed that the addition of rituximab to fludarabine and cyclophosphamide (FCR) significantly improved OR, CR and PFS. The improvement in PFS directly resulted from an improved ability to eliminate minimal residual disease (MRD) in the peripheral blood, highlighting the importance of MRD eradication. However, a multi-center study suggested that the high CR rates to chemoimmunotherapy regimens such as FCR obtained in academic centers may not be reproducible when the same regimens are given in the community setting. The immunomodulatory drug lenalidomide is active in relapsed high-risk CLL, but two studies of lenalidomide in previously untreated CLL patients failed to achieve a CR and were associated with significant tumor lysis, tumor flare and hematologic toxicity. In the relapsed setting, a combination study of the bifunctional alkylator bendamustine and rituximab (BR) demonstrated a high OR rate in patients with del(11q22) and del(17p13), indicating that further studies to define's bendamustine activity are warranted in high-risk CLL. Similarly, the CDK inhibitor flavopiridol demonstrated significant clinical activity and durable remissions in heavily treated, refractory CLL patients with high-risk cytogenetic features and bulky lymphadenopathy. The monoclonal anti-CD20 antibody ofatumumab appeared to be superior to rituximab in relapsed CLL patients with bulky nodal disease or high-risk cytogenetic features. Ongoing studies of these agents and other novel therapeutic agents in clinical development hold forth the promise that treatment options for CLL patients will continue to expand and improve.     

Comparative effects of cyproterone acetate or a long-acting LHRH agonist in polycystic ovarian disease

A randomized cross-over study was done to compare the therapeutic efficacy of cyproterone acetate (CPA, 50 mg/day orally) and a depot preparation of the LHRH superagonist (D-Trp6 LHRH 3 mg i.m. once a month) in 10 patients with polycystic ovarian disease (PCO). The two treatment periods were separated by 6 months. Both treatments resulted in marked clinical improvement. In response to CPA treatment, basal plasma gonadotropin, estradiol, estrone, testosterone and androstenedione levels significantly decreased. In response to D-Trp6 LHRH, both basal and stimulated gonadotropin levels were completely suppressed after 3 weeks of treatment. After initial elevation on day 2, plasma ovarian steroid levels fell into the castrate range, without any change in dehydroepiandrosterone sulfate levels. Urinary 3 alpha-androstanediol excretion decreased significantly. In patients with PCO, LHRH-A induced more complete gonadotropin inhibition than did CPA. However, following cessation of either therapy, the disease rapidly recurred.     

Structure-activity studies of heparan mimetic polyanions for anti-prion therapies

Polysulfated molecules, as the family of heparan mimetics (HMs) and pentosan polysulfate, are considered among the more promising drugs used in experimental models of prion diseases. Regardless of their therapeutic potential, structure-function studies on these polyanions are still missing. Here, we report the syntheses of a library of HMs of different molecular sizes, containing various sulfation and carboxylation levels, and substituted or not by different hydrophobic cores. The HMs capacities to inhibit the accumulation of PrPres in chronically infected cells (ScGT1-7) and their PrPc binding abilities were examined. Our results showed that an optimal size and sulfation degree are needed for optimum activity, that incorporation of hydrophobic moieties increases compounds efficacy and that the presence of carboxymethyl moieties decreases it. These structural features should be considered on the modelling of polyanionic compounds for optimum anti-prion activities and for advancing in the understanding the mechanisms involved in their biological actions.     

Sulfated polysaccharides inhibit the catabolism and loss of both large and small proteoglycans in explant cultures of tendon

This study investigated the effects of two highly sulfated polysaccharides, calcium pentosan polysulfate and heparin, on the loss of newly synthesized proteoglycans from the matrix of explant cultures of bovine tendon. The tensional region of deep flexor tendon was incubated with [35S]sulfate for 6 h and then placed in culture for up to 15 days. The amount of radiolabel associated with proteoglycans lost to the medium and retained in the matrix was determined for each day in culture. It was shown that both sulfated polysaccharides at concentrations of 1000 microg x mL(-1) inhibited the loss of 35S-labeled large and small proteoglycans from the matrix and concomitant with this was a retention of chemical levels of proteoglycans in the explant cultures. In other explant cultures that were maintained in culture in the presence of both agents for more than 5 days after incubation with [35S]sulfate, inhibition of the intracellular catabolic pathway was evident, indicating that these highly sulfated polysaccharides also interfered with the intracellular uptake of small proteoglycans by tendon cells.     

Augmented extracellular ATP signaling in bladder urothelial cells from patients with interstitial cystitis

Interstitial cystitis (IC) is an idiopathic hypersensory condition of the bladder associated with increased urinary ATP and increased stretch-activated ATP release by bladder urothelial cells (BUCs), suggesting augmented purinergic signaling in the bladder. To test this theory further, monolayers of cultured BUCs derived from bladder biopsies obtained from patients with IC and control patients were stimulated with 10-30 microM ATP with subsequent measurement of extracellular ATP levels using the luciferin-luciferase assay. Stimulation with 30 microM ATP resulted in IC supernatant containing several-fold more ATP than control BUCs initially, followed by a slower decrease in ATP levels. This difference in ATP levels was not completely due to activity of cellular ecto-ATPase, because blockade with ARL67156 did not normalize the difference. Exposure to hypotonic solutions resulted in similar extracellular ATP concentrations in IC and control BUCs, but there was a slower decrease in ATP levels in IC supernatants. Treatment of IC BUCs with 10-40 microM suramin, a nonspecific P2 receptor antagonist, significantly attenuated the IC BUC response to extracellular ATP, restoring IC BUCs to a control phenotype. Pretreatment of IC BUCs with 20 ng/ml of heparin-binding EGF-like growth factor (HB-EGF), which previously has been shown to be decreased in IC urine specimens, also restored IC BUCs to a control phenotype with respect to response to ATP stimulation. In conclusion, IC BUCs have augmented extracellular ATP signaling that could be blocked by suramin and HB-EGF. These findings suggest the possible development of future novel therapeutic techniques.     

Modulation of endothelial cells growth induced by heparin

We have previously shown that heparin was bound and internalized by cultured human endothelial cells. In this study, we have investigated the effect of heparin on endothelial cells growth. We found that heparin inhibited 3H-thymidine uptake as well as actual cell growth in a dose-dependent manner in the presence of low concentrations of human serum. Inhibition was maximal at 1% serum concentration and was abolished at 10%. Chasing experiments supported the role of membrane-bound heparin in this inhibition. Low molecular weight heparin fractions, or pentosan polysulfate, were equally effective in inhibiting 3H-thymidine uptake. On the other hand, the simultaneous addition of heparin and ECGS was synergic in stimulating 3H-thymidine uptake. These results suggest a modulatory role of heparin in endothelial cells growth.     

Effects of glycemic control on target organ responses to epinephrine in type 1 diabetes

Severe hypoglycemia occurs in intensively treated patients with type 1 diabetes mellitus (T1DM) due in part to deficient epinephrine counterregulatory responses. Previously, we have found that T1DM patients demonstrated a spectrum of altered responses to epinephrine at a variety of target organs compared with nondiabetic healthy subjects. What is not known is whether intensive glycemic control further modifies target organ responses in individuals with T1DM. Therefore, the aim of this study is to assess whether there is tissue specific (liver, muscle, adipose tissue, pancreas and cardiovascular) resistance to epinephrine in intensively controlled (IC) T1DM compared with those with conventional control (CC). Eight IC patients (age 33 +/- 4 yr, BMI 24 +/- 2 kg/m2, Hb A1C 6.7 +/- 0.1%), and 11 CC patients (age 35 +/- 3 yr, BMI 25 +/- 1 kg/m2, Hb A1C 9.6 +/- 0.1%) underwent two separate randomized, single-blind, 2-h hyperinsulinemic euglycemic clamp studies with (EPI) and without (NO EPI) epinephrine infusion. Epinephrine levels during EPI were similar in all groups (5,197 +/- 344 pmol/l). Glucose (5.3 +/- 0.1 mmol/l) and insulin levels (515 +/- 44 pmol/l) were similar in all groups during the glucose clamps. Endogenous glucose production (EGP) and glucose uptake (R(d)) were determined using [3-H3]glucose. Muscle biopsy was performed at the end of each study. IC had a significantly reduced EGP and R(d) responses to EPI compared with CC. Glucagon responses to EPI were similarly blunted in both IC and CC. Free fatty acid and glycerol response to EPI was greater in CC compared with IC. There was a significantly greater systolic blood pressure response to EPI in CC. We conclude that, despite similar epinephrine, insulin, and glucose levels, intensively treated T1DM patients had reduced cardiovascular, skeletal muscle, hepatic, and adipose target organ responses to EPI compared with conventionally treated T1DM patients.     

The efficacy of psychotherapy and pharmacotherapy in treating depressive and anxiety disorders: a meta‐analysis of direct comparisons

Although psychotherapy and antidepressant medication are efficacious in the treatment of depressive and anxiety disorders, it is not known whether they are equally efficacious for all types of disorders, and whether all types of psychotherapy and antidepressants are equally efficacious for each disorder. We conducted a meta-analysis of studies in which psychotherapy and antidepressant medication were directly compared in the treatment of depressive and anxiety disorders. Systematic searches in bibliographical databases resulted in 67 randomized trials, including 5,993 patients that met inclusion criteria, 40 studies focusing on depressive disorders and 27 focusing on anxiety disorders. The overall effect size indicating the difference between psychotherapy and pharmacotherapy after treatment in all disorders was g=0.02 (95% CI: −0.07 to 0.10), which was not statistically significant. Pharmacotherapy was significantly more efficacious than psychotherapy in dysthymia (g=0.30), and psychotherapy was significantly more efficacious than pharmacotherapy in obsessive-compulsive disorder (g=0.64). Furthermore, pharmacotherapy was significantly more efficacious than non-directive counseling (g=0.33), and psychotherapy was significantly more efficacious than pharmacotherapy with tricyclic antidepressants (g=0.21). These results remained significant when we controlled for other characteristics of the studies in multivariate meta-regression analysis, except for the differential effects in dysthymia, which were no longer statistically significant.