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Dose Dependently (dose + dependently)
Selected AbstractsEstimation of endogenous adenosine activity at adenosine receptors in guinea-pig ileum using a new pharmacological methodACTA PHYSIOLOGICA, Issue 2 2010K. F. Nilsson Abstract Aim:, Adenosine modulates neurotransmission and in the intestine adenosine is continuously released both from nerves and from smooth muscle. The main effect is modulation of contractile activity by inhibition of neurotransmitter release and by direct smooth muscle relaxation. Estimation of adenosine concentration at the receptors is difficult due to metabolic inactivation. We hypothesized that endogenous adenosine concentrations can be calculated by using adenosine receptor antagonist and agonist and dose ratio (DR) equations. Methods:, Plexus-containing guinea-pig ileum longitudinal smooth muscle preparations were made to contract intermittently by electrical field stimulation in organ baths. Schild plot regressions were constructed with 2-chloroadenosine (agonist) and 8-(p -sulfophenyl)theophylline (8-PST; antagonist). In separate experiments the reversing or enhancing effect of 8-PST and the inhibiting effect of 2-chloroadenosine (CADO) were analysed in the absence or presence of an adenosine uptake inhibitor (dilazep), and nucleoside overflow was measured by HPLC. Results:, Using the obtained DR, baseline adenosine concentration was calculated to 28 nm expressed as CADO activity, which increased dose dependently after addition of 10,6 m dilazep to 150 nm (P < 0.05). HPLC measurements yielded a lower fractional increment (80%) in adenosine during dilazep, than found in the pharmacological determination (440%). Conclusion:, Endogenous adenosine is an important modulator of intestinal neuro-effector activity, operating in the linear part of the dose,response curve. Other adenosine-like agonists might contribute to neuromodulation and the derived formulas can be used to calculate endogenous agonist activity, which is markedly affected by nucleoside uptake inhibition. The method described should be suitable for other endogenous signalling molecules in many biological systems. [source] Exenatide: a review from pharmacology to clinical practiceDIABETES OBESITY & METABOLISM, Issue 6 2009R. Gentilella Background:, Exenatide is an incretin mimetic that activates glucagon-like-peptide-1 receptors. It blunts the postprandial rise of plasma glucose by increasing glucose-dependent insulin secretion, suppressing inappropriately high glucagon secretion and delaying gastric emptying. Methods:, In seven clinical trials performed in 2845 adult patients with type 2 diabetes mellitus who were inadequately controlled by a sulphonylurea and/or metformin (glycosylated haemoglobin, HbA1c ,11%), or by thiazolidinediones (with or without metformin) and treated for periods from 16 weeks to 3 years, exenatide (5 ,g b.i.d. s.c. for the first 4 weeks of treatment and 10 ,g b.i.d. s.c. thereafter) reduced HbA1c, fasting and postprandial glucose, and body weight dose dependently, and was similar to insulin glargine and biphasic insulin aspart in reducing HbA1c. Body weight diminished with exenatide, whereas it increased with both insulin preparations. Positive effects on the lipid profile and a reduction in C-reactive protein were also recorded with exenatide. Treatment extensions up to 3 years showed that benefits were maintained in the long term. Adverse events were usually mild to moderate in intensity, and generally the frequency decreased with continued therapy. The most common was nausea (whose incidence may be reduced by gradual dose escalation from 5 ,g b.i.d. to 10 ,g b.i.d.), vomiting, diarrhoea, headache and hypoglycaemia (almost exclusively in patients treated with a sulphonylurea). Results and conclusions:, Exenatide is a new, promising therapeutic option for type 2 diabetic patients inadequately controlled by oral agents, before insulin therapy, offering the added benefits of body weight reduction and tight postprandial glucose control. [source] Dose-dependent long-term effects of Tat in the rat hippocampal formation: A design-based stereological studyHIPPOCAMPUS, Issue 4 2010Sylvia Fitting Abstract The human immunodeficiency virus type 1 (HIV-1) protein transactivator of transcription (Tat) is believed to play a critical role in mediating central nervous system (CNS) pathology in pediatric HIV-1 infection. Long-term neurotoxicity was investigated in a design-based stereology study following intrahippocampal injection of Tat on postnatal day (P)10, a time period that approximates the peak in the rats' rate of brain growth and mimics clinical HIV-1 CNS infection at labor/delivery. The goal was to examine the impact of P10 intrahippocampal Tat injection on the anatomy of the adult hippocampus (5 month) to gain a better understanding about how timing of infection influences the rate of progression of pediatric HIV-1 infection [cf. Fitting et al. (2008a) Hippocampus 18:135,147]. Male P10 Sprague-Dawley rats were bilaterally injected with vehicle or one of three different doses of Tat (5, 25, or 50 ,g). Unbiased stereological estimates were used to quantify total neuron number (Nissl stain) in five major subregions of the rat hippocampus: granular layer (GL), hilus of the dentate gyrus (DGH), cornu ammonis fields (CA)2/3, CA1, and subiculum (SUB). Glial cells (astrocytes and oligodendrocytes) were quantified in the DGH and SUB. No significant reduction of neuron number was noted for any of the five hippocampal subregions, in contrast to the very prominent reductions reported when Tat was administered on P1 [Fitting et al. (2008a) Hippocampus 18:135,147]. However, for glial cells, the number of astrocytes in the DGH and SUB as well as the number of oligodendrocytes in the DGH were linear dose dependently increased as a function of dose of Tat. In conjunction with previous stereological research [Fitting et al., (2008a) Hippocampus 18:135,147], the present data suggest that variability in the progression of pediatric HIV/acquired immunodeficiency syndrome (AIDS) may be better understood with the knowledge of the factor of timing of HIV-1 CNS infection. © 2009 Wiley-Liss, Inc. [source] Gliotoxin, an inhibitor of nuclear factor-kappa B, attenuates peptidoglycan-polysaccharide-induced colitis in ratsINFLAMMATORY BOWEL DISEASES, Issue 3 2002Dr. Leo R. Fitzpatrick Abstract Gliotoxin is a fungal metabolite that has immunosuppressive properties. First, we determined if gliotoxin could inhibit bacterial peptidoglycan,polysaccharide-stimulated tumor necrosis factor-, production, as well as nuclear factor-kappa B (NF-,B), in a rat macrophage (NR8383) cell line. Next, the apoptosis-inducing potential of gliotoxin was also evaluated in this cell line. Finally, we evaluated whether gliotoxin could reduce peptidoglycan,polysaccharide-induced colitis in rats. Gliotoxin (2 mg/kg/day) was dosed from day 14 after the initial intramural colonic injection of peptidoglycan,polysaccharide until day 21. A gross colonic injury score, myeloperoxidase activity, and cytokine levels were all evaluated on day 21. Gliotoxin dose dependently inhibited cytokine production, as well as NF-,B, and also induced apoptosis in the NR8383 cell line. On day 21, gliotoxin significantly reduced gross colonic injury (adhesions, nodules, mucosal lesions) in rats. Gliotoxin-treated rats also had partially normalized biochemical indices of colitis, such as colonic cytokine levels. The colonic level of NF-,B was also partially normalized in gliotoxin treated rats. Gliotoxin also exhibited an antiarthritis effect in peptidoglycan,polysaccharide-treated rats. In summary, gliotoxin effectively attenuated the chronic reactivation phase of peptidoglycan,polysaccharide-induced colitis. This anticolitis effect may be related to the inhibition of NF-,B in Lewis rats. [source] Cigarette smoke extract affects functional activity of MRP1 in bronchial epithelial cellsJOURNAL OF BIOCHEMICAL AND MOLECULAR TOXICOLOGY, Issue 5 2007Margaretha van der Deen Abstract Cigarette smoke is the principal risk factor for development of chronic obstructive pulmonary disease (COPD). Multidrug resistance-associated protein 1 (MRP1) is a member of the ATP-binding cassette (ABC) superfamily of transporters, which transport physiologic and toxic substrates across cell membranes. MRP1 is highly expressed in lung epithelium. This study aims to analyze the effect of cigarette smoke extract (CSE) on MRP1 activity. In the human bronchial epithelial cell line 16HBE14o,, MRP1 function was studied flow cytometrically by cellular retention of carboxyfluorescein (CF) after CSE incubation and MRP1 downregulation by RNA interference (siRNA). Cell survival was measured by the MTT assay. Immunocytochemically, it was shown that 16HBE14o, expressed MRP1 and breast cancer resistance protein. Coincubation of CSE IC50 (1.53% ± 0.22%) with MK571 further decreased cell survival 31% (p, = 0.018). CSE increased cellular CF retention dose dependently from 1.7-fold at 5% CSE to 10.3-fold at 40% CSE (both p < 0.05). siRNA reduced MRP1 RNA expression with 49% and increased CF accumulation 67% versus control transfected cells. CSE exposure further increased CF retention 24% (p = 0.031). A linear positive relation between MRP1 function and CSE-modulating effects (r = 0.99, p =0.089) was shown in untransfected, control transfected, and MRP1 downregulated 16HBE14o, cells analogous to blocking effects with MRP1 inhibitor MK571 (r = 0.99, p = 0.034). In conclusion, cigarette smoke extract affects MRP1 activity probably competitively in bronchial epithelial cells. Inhibition of MRP1 in turn results in higher CSE toxicity. We propose that MRP1 may be a protective protein for COPD development. © 2007 Wiley Periodicals, Inc. J Biochem Mol Toxicol 21:243,251, 2007; Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/jbt.20187 [source] Dietary vitamin E reduces labile iron in rat tissuesJOURNAL OF BIOCHEMICAL AND MOLECULAR TOXICOLOGY, Issue 5 2005Wissam Ibrahim Abstract Previous studies have shown that dietary vitamin E reduced generation and/or levels of superoxide. As superoxide has potential to release iron from its transport and storage proteins, and labile or available form of iron is capable of catalyzing the formation of reactive hydroxyl radicals, the effect of dietary vitamin E on labile iron pool was studied in rats. One-month-old Sprague-Dawley male and female rats were fed a basal vitamin E-deficient diet supplemented with 0, 20, 200, or 2,000 IU vitamin E/kg diet for 90 days. The levels of labile iron were measured in the liver, kidney, spleen, heart and skeletal muscle. Additionally, the levels of lipid peroxidation products were measured. The results showed that, except for labile iron in the heart of male rats, dietary vitamin E dose dependently reduced the levels of labile iron and lipid peroxidation products in all tissues of male and female rats. The findings suggest that dietary vitamin E may protect against oxidative tissue damage by reducing the generation and/or level of superoxide, which in turn attenuates the release of iron from its protein complexes. © 2005 Wiley Periodicals, Inc. J Biochem Mol Toxicol 19:298,303, 2005; Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/jbt.20094 [source] Intermittently Administered Human Parathyroid Hormone(1,34) Treatment Increases Intracortical Bone Turnover and Porosity Without Reducing Bone Strength in the Humerus of Ovariectomized Cynomolgus MonkeysJOURNAL OF BONE AND MINERAL RESEARCH, Issue 1 2001David B. Burr Abstract Cortical porosity in patients with hyperparathyroidism has raised the concern that intermittent parathyroid hormone (PTH) given to treat osteoporotic patients may weaken cortical bone by increasing its porosity. We hypothesized that treatment of ovariectomized (OVX) cynomolgus monkeys for up to 18 months with recombinant human PTH(1,34) [hPTH(1,34)] LY333334 would significantly increase porosity in the midshaft of the humerus but would not have a significant effect on the strength or stiffness of the humerus. We also hypothesized that withdrawal of PTH for 6 months after a 12-month treatment period would return porosity to control OVX values. OVX female cynomolgus monkeys were given once daily subcutaneous (sc) injections of recombinant hPTH(1,34) LY333334 at 1.0 ,g/kg (PTH1), 5.0 ,g/kg (PTH5), or 0.1 ml/kg per day of phosphate-buffered saline (OVX). Sham OVX animals (sham) were also given vehicle. After 12 months, PTH treatment was withdrawn from half of the monkeys in each treatment group (PTH1-W and PTH5-W), and they were treated for the remaining 6 months with vehicle. Double calcein labels were given before death at 18 months. After death, static and dynamic histomorphometric measurements were made intracortically and on periosteal and endocortical surfaces of sections from the middiaphysis of the left humerus. Bone mechanical properties were measured in the right humeral middiaphysis. PTH dose dependently increased intracortical porosity. However, the increased porosity did not have a significant detrimental effect on the mechanical properties of the bone. Most porosity was concentrated near the endocortical surface where its mechanical effect is small. In PTH5 monkeys, cortical area (Ct.Ar) and cortical thickness (Ct.Th) increased because of a significantly increased endocortical mineralizing surface. After withdrawal of treatment, porosity in PTH1-W animals declined to sham values, but porosity in PTH5-W animals remained significantly elevated compared with OVX and sham. We conclude that intermittently administered PTH(1,34) increases intracortical porosity in a dose-dependent manner but does not reduce the strength or stiffness of cortical bone. [source] Basic Fibroblast Growth Factor Stimulates Vascular Endothelial Growth Factor Release in Osteoblasts: Divergent Regulation by p42/p44 Mitogen-Activated Protein Kinase and p38 Mitogen-Activated Protein KinaseJOURNAL OF BONE AND MINERAL RESEARCH, Issue 12 2000Haruhiko Tokuda Abstract We previously showed that basic fibroblast growth factor (bFGF) activates p38 mitogen-activated protein (MAP) kinase via Ca2+ mobilization, resulting in interleukin-6 (IL-6) synthesis in osteoblast-like MC3T3-E1 cells. In the present study, we investigated the effect of bFGF on the release of vascular endothelial growth factor (VEGF) in these cells. bFGF stimulated VEGF release dose dependently in the range between 10 and 100 ng/ml. SB203580, an inhibitor of p38 MAP kinase, markedly enhanced the bFGF-induced VEGF release. bFGF induced the phosphorylation of both p42/p44 MAP kinase and p38 MAP kinase. PD98059, an inhibitor of upstream kinase of p42/p44 MAP kinase, reduced the VEGF release. SB203580 enhanced the phosphorylation of p42/p44 MAP kinase induced by bFGF. The enhancement by SB203580 of the bFGF-stimulated VEGF release was suppressed by PD98059. The depletion of extracellular Ca2+ by [ethylenebis-(oxyethylenenitrilo)]tetracetic acid (EGTA) or 1,2-bis-(O -aminophinoxy)-ethane- N,N,N,N -tetracetic acid tetracetoxymethyl ester (BAPTA/AM), a chelator of intracellular Ca2+, suppressed the bFGF-induced VEGF release. A23187, a Ca ionophore, or thapsigargin, known to induce Ca2+ release from intracellular Ca2+ store, stimulated the release of VEGF by itself. A23187 induced the phosphorylation of p42/p44 MAP kinase and p38 MAP kinase. PD98059 suppressed the VEGF release induced by A23187. SB203580 had little effect on either A23187-induced VEGF release or the phosphorylation of p42/p44 MAP kinase by A23187. These results strongly suggest that bFGF stimulates VEGF release through p42/p44 MAP kinase in osteoblasts and that the VEGF release is negatively regulated by bFGF-activated p38 MAP kinase. [source] Sonic hedgehog is involved in osteoblast differentiation by cooperating with BMP-2JOURNAL OF CELLULAR PHYSIOLOGY, Issue 2 2002Takahito Yuasa The roles of Sonic hedgehog (Shh) and Bone morphogenetic protein-2 (Bmp-2) in osteoblast differentiation were investigated using in vitro cell systems. Recombinant amino-terminal portion of SHH (rSHH-N) dose dependently stimulated ALP activity in C3H10T1/2 and MC3T3-E1 cells. rSHH-N induced expression of Osteocalcin mRNA in C3H10T1/2 cells. A soluble form of the receptor for type IA BMP receptor antagonized rSHH-N-induced ALP activity in C3H10T1/2 and MC3T3-E1 cells, indicating that BMPs are involved in SHH-induced osteoblast differentiation. Simultaneous supplement with rSHH-N and BMP-2 synergistically induced ALP activity and expression of Osteocalcin mRNA in C3H10T1/2 cells. Pretreatment with rSHH-N for 6 h enhanced the response to BMP-2 by increasing ALP activity in C3H10T1/2 and MC3T3-E1 cells. Stimulatory effects of rSHH-N and additive effects with rSHH-N and BMP-2 on ALP activity were also observed in mouse primary osteoblastic cells. Transplantation of BMP-2 (1 ,g) into muscle of mice induced formation of ectopic bone, whereas transplantation of r-SHH-N (1,5 ,g) failed to generate it. These results indicate that Shh plays important roles in osteoblast differentiation by cooperating with BMP. © 2002 Wiley-Liss, Inc. [source] Focal adhesion kinase mediates human leukocyte histocompatibility antigen class II-induced signaling in gingival fibroblastsJOURNAL OF PERIODONTAL RESEARCH, Issue 6 2007S. Yoshizawa Background and Objective:, The role of human leukocyte antigen class II molecules on nonantigen-presenting cells has been a matter of controversy. We previously reported that human leukocyte antigen class II molecules on human gingival fibroblasts do not present antigens, but transduce signals into the cells by making a complex with antigenic peptide T-cell receptor or by stimulating cell surface human leukocyte antigen-DR molecules with human leukocyte antigen-DR antibody (L243), which mimics the formation of the human leukocyte antigen class II,antigenic peptide T-cell receptor complex, resulting in the expression of several cytokines. The aim of this study was to detect human leukocyte antigen class II-associated molecules mediating human leukocyte antigen class II-induced signals into the cells. Material and Methods:, Antibody-based protein-microarray analysis was performed to detect activated signaling molecules in gingival fibroblasts stimulated via human leukocyte antigen class II molecules. Then, we examined if these molecules structurally associate with human leukocyte antigen class II and actually transduce signals into the cells. Results:, Stimulation of human leukocyte antigen class II on gingival fibroblasts by L243 resulted in enhanced phosphorylation of focal adhesion kinase. Focal adhesion kinase was co-immunoprecipitated with human leukocyte antigen-DR by L243. Stimulation of gingival fibroblasts with L243 induced phosphorylation of focal adhesion kinase. Luteolin, a putative focal adhesion kinase inhibitor, suppressed phosphorylation of focal adhesion kinase and dose dependently inhibited human leukocyte antigen class II-induced cytokine production. Conclusion:, Focal adhesion kinase is structurally associated with human leukocyte antigen-DR and mediates human leukocyte antigen class II-induced signals in gingival fibroblasts. [source] Milk basic protein increases alveolar bone formation in rat experimental periodontitisJOURNAL OF PERIODONTAL RESEARCH, Issue 1 2007H. Seto Background and Objective:, It is conceivable that the active components extracted from milk whey protein (i.e. milk basic protein, MBP) stimulate bone formation and suppress bone resorption. Periodontitis is characterized by excessive alveolar bone resorption. We examined whether milk basic protein could recover alveolar bone loss in rat experimental periodontitis. Material and Methods:, A nylon ligature was placed around the cervix of molars in 8-wk-old male Fischer rats for 20 d. Then, the ligature was removed and a powder diet containing 0.2 or 1.0% milk basic protein was provided daily for another 45,90 d. On days 45 and 90, the maxillae were extracted and analyzed using microcomputerized tomography (micro-CT), followed by histological analysis. Results:, Micro-CT images showed that alveolar bone resorption was severely induced around the molar by the 20-d ligature procedure. Treatment with high-dose milk basic protein (1.0%) clearly recovered ligature-induced alveolar bone resorption on days 45 and 90, whereas low-dose milk basic protein (0.2%) did not show such a clear effect. Histological examination clarified that the osteoid thickness of alveolar bone was dose dependently increased by milk basic protein treatment for 90 d. Conclusion:, These findings suggest that a systemic administration of milk basic protein may be effective for the recovery of alveolar bone loss in periodontitis. [source] Anti-angiogenic, antinociceptive and anti-inflammatory activities of Lonicera japonica extractJOURNAL OF PHARMACY AND PHARMACOLOGY: AN INTERNATI ONAL JOURNAL OF PHARMACEUTICAL SCIENCE, Issue 6 2008Hye-Jung Yoo This study aimed to elucidate some novel pharmacological activities of Lonicera japonica (Caprifoliaceae), which is widely used in Oriental folk medicine. The ethanolic extract of L. japonica (LJ) dose dependently inhibited chick chorioallantoic membrane angiogenesis. The antinociceptive activity of LJ was assessed using the acetic acid-induced constriction model in mice. LJ showed anti-inflammatory activity in two in-vivo models: the vascular permeability and air pouch models. LJ suppressed the production of nitric oxide via down-regulation of inducible nitric oxide synthase in lipopolysaccharide-stimulated RAW264.7 macrophage cells. However, LJ was unable to suppress induction of cyclooxygenase-2 in the stimulated macrophage cells. LJ decreased the reactive oxygen species level in the stimulated macrophage cells. In brief, the flowers of L. japonica possess potent anti-angiogenic and antinociceptive activities, in addition to anti-inflammatory activity, which partly supports its therapeutic efficacy. [source] Inhibitory effect of apigenin on benzo(a)pyrene-mediated genotoxicity in Swiss albino miceJOURNAL OF PHARMACY AND PHARMACOLOGY: AN INTERNATI ONAL JOURNAL OF PHARMACEUTICAL SCIENCE, Issue 12 2006Tajdar Husain Khan Apigenin, a bioflavonoid, is abundantly present in fruits and vegetables and possesses potential chemopreventive properties against a wide variety of chronic diseases. In this study we investigated the anti-genotoxic effects of apigenin against a known genotoxicant, benzo(a)pyrene (B(a)P) (125 mg kg,1 orally) toxicity in Swiss albino mice. B(a)P administration led to induction of cytochrome P-450 (CYP), aryl hydrocarbon hydroxylase (AHH) and DNA strand breaks (P < 0.001), which was suppressed by apigenin (2.5 and 5 mg kg,1 orally) dose dependently (P < 0.001). B(a)P-induced depletion in the level of reduced glutathione (GSH), quinone reductase (QR) and glutathione-S-transferase (GST) was also shown to be restored by apigenin pre-treatment (P < 0.001). A simultaneous significant and dose-dependent reduction was noted in DNA strand breaks and in-vivo DNA damage (P < 0.001), which gives some insight into restoration of DNA integrity in modulator groups. These results strongly support the protective nature of apigenin against B(a)P-induced toxicity. [source] Effects of recombinant human activated protein C on the coagulation system: a study with rotational thromboelastometryACTA ANAESTHESIOLOGICA SCANDINAVICA, Issue 9 2008C. U. NILSSON Background: Recombinant human activated protein C (rhAPC) is an anticoagulant that can be used for treatment of patients with severe sepsis. The use of rhAPC is accompanied by an increased risk of severe bleeding. Rotational thromboelastometry is a method for measuring the status of the coagulation. The aim of the study was to investigate whether rotational thromboelastometry could be used for monitoring the effects of rhAPC on the coagulation. Methods: Whole blood was mixed in vitro with concentrations of rhAPC ranging from 0 to 75 ng/ml and analysed with rotational thromboelastometry. Results: The parameter Coagulation Time was significantly prolonged by increasing the concentrations of rhAPC (P=0.002). Other parameters were not significantly affected. Conclusion: rhAPC dose dependently affects the early humoral parts of the coagulation, while platelet function and fibrinogen to fibrin conversion seem virtually unaffected. [source] Growth and Hematological Changes of Rockfish, Sebastes schlegeli (Hilgendorf) Exposed to Dietary Cu and CdJOURNAL OF THE WORLD AQUACULTURE SOCIETY, Issue 2 2005Ju-Chan Kang Cd and Cu toxicological effect on growth and hematological parameters was investigated in juvenile rockfish Sebasres schlegeli after sub-chronic dietary Cd (0, 0.5, 5, 25, and 125 mg/kg) and Cu exposure (0, 50, 125, 250, and 500 mg/kg) for 60 d. In the experiment of dietary Cd exposure, weight and length SGR of the rockfish were significantly different from control, and a significant inverse relationship was observed between weight gain and the exposure concentration of dietary Cd at 25, 125 mg/kg (P > 0.05). Hematwrit and hemoglobin decreased significantly and were dose dependently in all Cd exposure. Glucose in serum was also increased significantly (P < 0.05). The concentration of total protein in serum was significantly lower than control at 5, 25, and 125 mgkg. No differences were observed in serum calcium concentration. Magnesium concentration in serum was increased signillcantly with dietary Cd concentration. In the experiment of dietary Cu exposure, Cu was inhibited weight gain and growth rate. No differences were observed in hematocrit, hemoglobin and RBCs compared to control. Contents of total protein, glucose, and Ca in serum remained stable. Mg concentration in serum was increased significantly at 500 mg/kg. [source] Two subpopulations of thrombin-activated platelets differ in their binding of the components of the intrinsic factor X-activating complexJOURNAL OF THROMBOSIS AND HAEMOSTASIS, Issue 11 2005M. A. PANTELEEV Summary., Binding of fluorescein-labeled coagulation factors IXa, VIII, X, and allophycocyanin-labeled annexin V to thrombin-activated platelets was studied using flow cytometry. Upon activation, two platelet subpopulations were detected, which differed by 1,2 orders of magnitude in the binding of the coagulation factors and by 2,3 orders of magnitude in the binding of annexin V. The percentage of the high-binding platelets increased dose dependently of thrombin concentration. At 100 nm of thrombin, platelets with elevated binding capability constituted ,4% of total platelets and were responsible for the binding of ,50% of the total bound factor. Binding of factors to the high-binding subpopulation was calcium-dependent and specific as evidenced by experiments in the presence of excess unlabeled factor. The percentage of the high-binding platelets was not affected by echistatin, a potent aggregation inhibitor, confirming that the high-binding platelets were not platelet aggregates. Despite the difference in the coagulation factors binding, the subpopulations were indistinguishable by the expression of general platelet marker CD42b and activation markers PAC1 (an epitope of glycoprotein IIb/IIIa) and CD62P (P-selectin). Dual-labeling binding studies involving coagulation factors (IXa, VIII, or X) and annexin V demonstrated that the high-binding platelet subpopulation was identical for all coagulation factors and for annexin V. The high-binding subpopulation had lower mean forward and side scatters compared with the low-binding subpopulation (,80% and ,60%, respectively). In its turn, the high-binding subpopulation was not homogeneous and included two subpopulations with different scatter values. We conclude that activation by thrombin induces the formation of two distinct subpopulations of platelets different in their binding of the components of the intrinsic fX-activating complex, which may have certain physiological or pathological significance. [source] Interstitial Cystitis and the Therapeutic Effect of Suplatast TosilateLUTS, Issue 2009Yukio HAYASHI Painful bladder syndrome (PBS)/interstitial cystitis (IC) can be a chronic and debilitating disease characterized by urinary urgency, frequency, and bladder pain, which are often very difficult to treat, regardless of currently-proposed treatments. Suplatast tosilate (IPD-1151T) is an immunoregulator that suppresses Th2 cytokine production, immunoglobulin E (IgE) synthesis, chemical mediator release from mast cells, and eosinophilic recruitment. In a preliminary, open-label clinical study of IPD-1151T in 14 women with IC, treatment with IPD-1151T significantly increased bladder capacity and decreased urinary urgency, urinary frequency, and lower abdominal pain, as measured by the IC symptom index, in patients with non-ulcerative IC. A concomitant reduction in immunological parameters (eosinophils, IgE, and urine T cells) was observed. Also, in basic experimental studies using hydrochloric acid-induced chronic cystitis rats, the oral administration of IPD-1151T (0.1,100 mg/kg/day) for 7 days after the induction of cystitis dose dependently increased the intercontraction intervals and micturition volume. In addition, the infiltration of mast cells and eosinophils into the bladder was suppressed by IPD-1151T. These findings suggest that IPD-1151T could be a new medicine for treating debilitating symptoms, such as bladder pain and urinary frequency in PBS/IC. [source] The Receptors and Role of Angiotensin II in Knee Joint Blood Flow Regulation and Role of Nitric Oxide in Modulation of Their FunctionMICROCIRCULATION, Issue 5 2003H. NAJAFIPOUR ABSTRACT Objectives: Angiotensin-converting enzyme (ACE) upregulation in the stroma cells of arthritis rheumatoid joints may produce a higher tissue concentration of angiotensin II (angII), which is a vasoconstrictor and mitogen factor that causes local hypoxia and synovial proliferation. No study in the literature has examined the role of angII in joint blood flow (JBF) regulation and the potential effect of ACE inhibitors on JBF. Methods: The study was performed on 20 Dutch white rabbits to examine the JBF response to angII, angII receptor subtypes, and the role of nitric oxide (NO) in angII effects in knee joint blood vessels. Drugs were administered locally through retrograde saphenous artery cannulation. Joint vascular resistance (JVR) was calculated by dividing the arterial blood pressure by the JBF. Results: AngII increased JVR dose dependently. The angII type 1 (AT1) receptor antagonist losartan did not change the basal JVR but completely blocked the effect of angII on JVR. N, -nitro-L-arginin methyl ester (L-NAME) increased JVR by a mean (±SEM) of 25.8 ± 8.7% (p < 0.05) but did not affect the joint vessel response to angII and losartan. Conclusions: AngII receptors are from the AT1 subtype in normal joint blood vessels, but angII plays no significant role in JBF regulation. The basal release of NO plays a role in resting JBF regulation, but NO does not affect the AT1 receptor-mediated vasoconstriction of joint blood vessels. [source] Influence of high-dose methotrexate therapy (HD-MTX) on glomerular and tubular kidney functionPEDIATRIC BLOOD & CANCER, Issue 6 2003Lutz Hempel MD Abstract Background The present investigation was intended to further clarify the mechanisms involved in renal dysfunction following high-dose methotrexate (HD-MTX) treatment. Patients and Methods Fifty eight predominately pediatric patients [39 male, 19 female; mean age 12.3 years (range 2.2,34.1)] suffering from acute lymphoblastic leukemia (ALL, n,=,28), Non Hodgkins lymphoma (NHL, n,=,13), osteosarcoma (n,=,8), malignant brain tumor (n,=,6), or an ALL relapse (n,=,3), were prospectively examined. In the course of 220 infusions of HD-MTX, glomerular and tubular renal function was determined by measuring proteinuria and glomerular filtration rate (GFR), as well as renal excretion of alpha-1-microglobulin (AMG) and N -acetyl-,- D -glucosaminidase (NAG). It was investigated whether there were differences in MTX toxicity in dependence on the administered dose (1, 5, or 12 g/m2 BSA), on the combination with other cytostatic agents (ifosfamide or cyclophosphamide), on the metabolism of MTX into 7-OH-MTX, and on pre-treatment with MTX. Results The administration of HD-MTX has no direct tubulotoxic effect. The disturbance in glomerular function was dose dependently and indicated by an increase in proteinuria as well as by a decrease in GFR; all changes were completely reversible and did not correlate to the metabolism of MTX to 7-OH-MTX. Increasing the number of MTX therapeutic cycles did not increase the nephrotoxicity of MTX. Conclusion MTX is not directly tubulotoxic. Its side effects on glomeruli are usually without clinical relevance. Med Pediatr Oncol 2003;40:348,354. © 2003 Wiley-Liss, Inc. [source] Antiinflammatory activities of flavonoids and a triterpene caffeate isolated from Bauhinia variegataPHYTOTHERAPY RESEARCH, Issue 7 2008Yerra Koteswara Rao Abstract In the continuing search for novel antiinflammatory agents, six flavonoids, namely kaempferol (1), ombuin (2), kaempferol 7,4,-dimethyl ether 3- O - , - d -glucopyranoside (3), kaempferol 3- O - , - d -glucopyranoside (4), isorhamnetin 3- O - , - d -glucopyranoside (5) and hesperidin (6), together with one triterpene caffeate, 3, - trans -(3,4-dihydroxycinnamoyloxy)olean-12-en-28-oic acid (7) were isolated from the non-woody aerial parts of Bauhinia variegata. Compounds 1,7 were evaluated as inhibitors of some macrophage functions involved in the inflammatory process. These seven compounds significantly and dose dependently inhibited lipopolysaccharide (LPS) and interferon (IFN)- , induced nitric oxide (NO), and cytokines [tumor necrosis factor (TNF)- , and interleukin (IL)-12]. The concentration causing a 50% inhibition (IC50) of NO, TNF- , and IL-12 production by compounds 1, 2 and 7 was approximately 30, 50 and 10 µM, respectively, while at 50, 200 and 40 µM compounds 3, 4, and 5, 6 showed 15,30% inhibition, respectively. On the other hand, compounds 3 and 7 showed no inhibitory effect, while compounds 1, 4,6 reduced by around 10,30% the synthesis of NO by macrophages, when inducible NO synthase was already expressed with LPS/IFN- , for 24 h. These experimental findings lend pharmacological support to the suggested folkloric uses of the plant B. variegata in the management of inflammatory conditions. Copyright © 2008 John Wiley & Sons, Ltd. [source] Inhibition of interleukin-6 receptor directly blocks osteoclast formation in vitro and in vivoARTHRITIS & RHEUMATISM, Issue 9 2009Roland Axmann Objective To investigate the efficacy of a murine anti,interleukin-6 receptor (anti,IL-6R) antibody in directly blocking tumor necrosis factor (TNF), and RANKL-mediated osteoclastogenesis in vitro and in vivo. Methods The efficacy of a murine antibody against IL-6R in blocking osteoclast differentiation of mononuclear cells stimulated with RANKL was tested. In addition, arthritic human TNF,,transgenic mice were treated with anti,IL-6R antibody, and osteoclast formation and bone erosion were assessed in arthritic paws. Results Blockade of IL-6R dose dependently reduced osteoclast differentiation and bone resorption in monocyte cultures stimulated with RANKL or RANKL plus TNF. In human TNF,,transgenic mice, IL-6R blockade did not inhibit joint inflammation, but it strongly reduced osteoclast formation in inflamed joints as well as bone erosion in vivo. Neither the cell influx into joints nor the synovial expression of IL-6 and RANKL changed with IL-6R blockade, while the synovial expression of IL-1 was significantly reduced. In contrast, TNF-mediated systemic bone loss was not inhibited by IL-6R blockade. Conclusion These data suggest that blockade of IL-6R directly affects osteoclast formation in vitro and in vivo, suggesting a direct and specific effect of anti,IL-6R therapy on osteoclasts independently of its antiinflammatory effects. This effect adds significantly to the structure-sparing potential of pharmacologic blockade of IL-6R in arthritis. [source] Mechanism of tyrosinase inhibition by deoxyArbutin and its second-generation derivativesBRITISH JOURNAL OF DERMATOLOGY, Issue 6 2008S. Chawla Summary Background, Disorders, such as age spots, melasma and hyperpigmentation at sites of actinic damage, emanate from the augmentation of an increased amount of epidermal melanin. Objectives, The ineptness of current therapies in treating these conditions, as well as high cytotoxicity, mutagenicity, poor skin penetration and low stability of skin-depigmenting formulations led us to investigate new compounds that meet the medical requirements for depigmentation agents. We have shown previously that the tyrosinase inhibitor deoxyArbutin (dA) is a more effective and less toxic skin lightener than hydroquinone (HQ). Methods, The efficacy and reversibility of dA and its derivatives on inhibiting tyrosine hydroxylase and DOPAoxidase was assessed using standard assays. Results, dA and its second-generation derivatives inhibit tyrosine hydroxylase and DOPAoxidase activities of tyrosinase dose dependently thereby inhibiting melanin synthesis in intact melanocytes, when used at concentrations that retain 95% cell viability in culture. This depigmenting effect was completely reversible when the compounds were removed. Tyrosinase inhibition was also observed in vitro when tested using human and purified mushroom tyrosinase, establishing that they are direct enzyme inhibitors. Lineweaver,Burk reciprocal plot analysis using mushroom tyrosinase illustrated that dA and its derivatives are more robust competitive inhibitors than HQ, when tyrosine is used as substrate. Conclusions, Thus, dA and its second-generation derivatives, which inhibit melanogenesis at safe concentrations by specifically acting on the tyrosinase enzyme at a post-translational level, are promising agents to ameliorate hyperpigmented lesions or lighten skin. [source] Novel function of DUSP14/MKP6 (dual specific phosphatase 14) as a nonspecific regulatory molecule for delayed-type hypersensitivityBRITISH JOURNAL OF DERMATOLOGY, Issue 5 2007Y. Nakano Summary Background, Nonspecific unresponsive states of delayed-type hypersensitivity (DTH) to unrelated antigens are induced in mice by a single administration of hapten. In these studies, we found a unique regulatory mechanism of contact hypersensitivity (CHS) mediated by nonspecific suppressor factor (NSF) induced by the intravenous injection of hapten-conjugated syngeneic spleen cells. NSF is a , 45-kDa protein released from the macrophage-like suppressor cells and binds selectively to dendritic cells (DCs). Moreover, NSF-treated DCs release a second , 20-kDa NSF (NSFint). Objectives, To try and identify NSF and characterize its function. Methods, The suppressor activity was evaluated by inhibition of the passive transfer of CHS by the effector cells sensitized with hapten and the antigen-presenting cell (APC) activity of hapten-primed draining lymph node cells (DLNCs) to induce CHS. NSF-containing supernatants obtained from the culture of spleen cells from mice that had been injected intravenously with oxazolone-conjugated syngeneic spleen cells 7 days before were prepared and purified with a Green A dye-affinity column, DEAE column and Sephacryl S-200 column. Then, samples of molecular mass of , 45 kDa were separated by native-PAGE (polyacrylamide gel electrophoresis) and nonreducing sodium dodecyl sulphate (SDS)-PAGE. After confirming the suppressor activity of proteins of , 45 kDa separated by native-PAGE, samples were separated by nonreducing SDS-PAGE, transferred onto polyvinylidene difluoride membranes and analysed using matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry. Results, Proteins of , 45 kDa eluted from a Sephacryl S-200 column and the slice of native-PAGE gel exhibited the strong suppressor activity. Analyses using MALDI-TOF mass spectrometry and MASCOT algorithm of the protein bands around 45 kDa separated by nonreducing SDS-PAGE identified NSF as a 22·5-kDa protein, dual specific phosphatase 14/MAP-kinase phophatase-6 (DUSP14/MKP6), which functions as a negative regulator of the MAP-kinase signalling. Western blot analyses revealed that recombinant DUSP14 (rDUSP14) exists as the mixture of 22·5-kDa monomer and 45-kDa dimer under nonreducing conditions, and monomers under reducing conditions. Treatment with rDUSP14 at 4 °C for 2 h suppressed the ability of effector cells to transfer CHS dose dependently and the APC function of DLNCs to induce CHS. Epicutaneous application of rDUSP14 immediately after challenge inhibited the subsequent CHS expression. rDUSP14 was bound specifically by major histocompatibility complex class II (Ia)-positive spleen cells (presumably DCs). The suppressor activity of NSF was neutralized by anti-DUSP14 monoclonal antibody. Expression of DUSP14 mRNA in the spleen was upregulated parallel to the unresponsive state induced by hapten-conjugated cells. NSF, NSFint and rDUSP14 exhibited the phosphatase activity towards p -nitrophenyl phosphate in vitro as alkaline phosphatase. Conclusions, These studies indicate for the first time that NSF is a dimer of DUSP14 secreted by macrophage-like suppressor cells by stimulation with hapten-conjugated cells and exerts a regulatory function on CHS through DCs as a secreted phosphatase. [source] A peptide (P2) derived from the variable heavy chain of an anti-P-selectin monoclonal antibody (LYP20) inhibits leucocyte adhesion to thrombin-activated platelets and endothelial cellsBRITISH JOURNAL OF HAEMATOLOGY, Issue 4 2003Joseph F. Murphy Summary. P-selectin, a member of the selectin family of adhesion molecules, is present in endothelial Weibel,Palade bodies and platelet ,-granules, and is rapidly expressed on their surface upon activation, resulting in leucocyte adhesion. LYP20 is a functional monoclonal antibody previously generated in our laboratory that binds with high affinity and specificity directed against P-selectin. This binding is largely imparted by the specific sequence of amino acids present on the hypervariable portions of the IgG chains. We now show that a peptide derived from the heavy chain of mAb LYP20 dose dependently inhibits the adhesion of poly morphonuclear cells to resting and thrombin-activated endothelial cells (EC) and platelets. The scrambled form of this peptide, identical in amino acid composition to the authentic peptide but with altered sequence, was not inhibitory at corresponding concentrations. Binding studies revealed that this peptide also dose dependently bound to both resting and thrombin-activated EC and platelets. Our results may prove useful for the development of new therapeutic inhibitors to modulate leucocyte interactions in inflammatory disorders. [source] Neuropeptide S is a stimulatory anxiolytic agent: a behavioural study in miceBRITISH JOURNAL OF PHARMACOLOGY, Issue 2 2008A Rizzi Background and purpose: Neuropeptide S (NPS) was recently identified as the endogenous ligand of an orphan receptor, now referred to as the NPS receptor. In vivo, NPS produces a unique behavioural profile by increasing wakefulness and exerting anxiolytic-like effects. In the present study, we further evaluated the effects of in vivo supraspinal NPS in mice. Experimental approach: Effects of NPS, injected intracerebroventricularly (i.c.v.), on locomotor activity (LA), righting reflex (RR) recovery and on anxiety states (measured with the elevated plus maze (EPM) and stress-induced hyperthermia (SIH) tests) were assessed in Swiss mice. Key results: NPS (0.01,1 nmol per mouse) caused a significant increase in LA in naive mice, in mice habituated to the test cages and in animals sedated with diazepam (5 mg kg,1). In the RR assay, NPS dose dependently reduced the proportion of animals losing the RR in response to diazepam (15 mg kg,1) and their sleeping time. In the EPM and SIH test, NPS dose dependently evoked anxiolytic-like effects by increasing the time spent by animals in the open arms and reducing the SIH response, respectively. Conclusions and implications: We provide further evidence that NPS acts as a novel modulator of arousal and anxiety-related behaviours by promoting a unique pattern of effects: stimulation associated with anxiolysis. Therefore, NPS receptor ligands may represent innovative drugs for the treatment of sleep and anxiety disorders. British Journal of Pharmacology (2008) 154, 471,479; doi:10.1038/bjp.2008.96; published online 31 March 2008 [source] Biological properties of a specific G,q/11 inhibitor, YM-254890, on platelet functions and thrombus formation under high-shear stressBRITISH JOURNAL OF PHARMACOLOGY, Issue 1 2006Toshio Uemura 1The effects of YM-254890, a specific G,q/11 inhibitor, on platelet functions, thrombus formation under high-shear rate condition and femoral artery thrombosis in cynomolgus monkeys were investigated. 2YM-254890 concentration dependently inhibited ADP-induced intracellular Ca2+ elevation, with an IC50 value of 0.92±0.28 ,M. 3P-selectin expression induced by ADP or thrombin receptor agonist peptide (TRAP) was strongly inhibited by YM-254890, with IC50 values of 0.51±0.02 and 0.16±0.08 ,M, respectively. 4YM-254890 had no effect on the binding of fibrinogen to purified GPIIb/IIIa, but strongly inhibited binding to TRAP-stimulated washed platelets. 5YM-254890 completely inhibited platelet shape change induced by ADP, but not that induced by collagen, TRAP, arachidonic acid, U46619 or A23187. 6YM-254890 attenuated ADP-, collagen-, TRAP-, arachidonic acid- and U46619-induced platelet aggregation with IC50 values of <1 ,M, whereas it had no effect on phorbol 12-myristate 13-acetate-, ristocetin-, thapsigargin- or A23187-induced platelet aggregation. 7High-shear stress-induced platelet aggregation and platelet-rich thrombus formation on a collagen surface under high-shear flow conditions were concentration dependently inhibited by YM-254890. 8The antithrombotic effect of YM-254890 was evaluated in a model of cyclic flow reductions in the femoral artery of cynomolgus monkeys. The intravenous bolus injection of YM-254890 dose dependently inhibited recurrent thrombosis without affecting systemic blood pressure or prolonging template bleeding time. 9YM-254890 is a useful tool for investigating G,q/11 -coupled receptor signaling and the physiological roles of G,q/11. British Journal of Pharmacology (2006) 148, 61,69. doi:10.1038/sj.bjp.0706711 [source] EFFECTS OF ADMINISTRATION OF ORAL MAGNESIUM ON PLASMA GLUCOSE AND PATHOLOGICAL CHANGES IN THE AORTA AND PANCREAS OF DIABETIC RATSCLINICAL AND EXPERIMENTAL PHARMACOLOGY AND PHYSIOLOGY, Issue 8 2005Nepton Soltani SUMMARY 1.,Magnesium deficiency has recently been proposed as a novel factor implicated in the pathogenesis of the complications of diabetes. The purpose of the present study was to determine the relationship between oral Mg supplementation and changes in plasma glucose, calcium, haemogolobin, Ca/Mg ratio, blood pressure and the histology of the pancreas and vascular system in streptozotocin-induced diabetic rats. 2.,Ten days after the induction of diabetes in male Wistar rats, half the diabetic animals were divided into six groups, receiving 0, 1, 3, 10, 30 or 50 g/L MgSO4 added into the drinking water for 8 weeks. Plasma glucose and Mg were measured at days 1, 2, 3, 5, 7, 14 and 21 to find the optimum dose of Mg and the time-course of its effect. In addition, histological observations were undertaken. Eight weeks later, all animals were decapitated, the pancreas and thoracic aorta were removed carefully and immersed immediately in 10% formaldehyde for histological study. 3.,To evaluate the effects of Mg on plasma glucose, calcium, haemoglobin, Mg and blood pressure, another group of animals was divided into four experimental groups, as follows: (i) non-diabetic controls received tap water for 8 weeks; (ii) acute diabetics received tap water for 10 days; (iii) chronic diabetic controls received tap water for 8 weeks; and (iv) Mg-treated chronic diabetic rats received 10 g/L MgSO4 added into the drinking water 10 days after the induction of diabetes for 8 weeks. 4.,Magnesium dose dependently affects plasma glucose levels. The peak effect was reached during the first 24 h following oral administration. Administration of 10 g/L MgSO4 results in the return of normal structure in the diabetic pancreas and aorta. Moreover, this concentration of MgSO4 causes glucose, haemoglobin, calcium, the Ca/Mg ratio and blood pressure to reach normal levels. Although the Mg level increases slightly following the administration of 10 g/L MgSO4 to diabetic rats, it never reaches control levels. 5.,On the basis of the results of the present study, it may be concluded that chronic Mg administration may have beneficial effects on diabetes. [source] Angiotensin I-Converting Enzyme And Metabolism Of The Haematological Peptide N -Acetyl-Seryl-Aspartyl-Lysyl-ProlineCLINICAL AND EXPERIMENTAL PHARMACOLOGY AND PHYSIOLOGY, Issue 12 2001Michel Azizi SUMMARY 1. Angiotensin I-converting enzyme (ACE) has two homologous active N- and C-terminal domains and displays activity towards a broad range of substrates. The tetrapeptide N -acetyl-seryl-aspartyl-lysyl-proline (AcSDKP) has been shown to be hydrolysed in vitro by ACE and to be a preferential substrate for its N-terminal active site. This peptide reversibly prevents the recruitment of pluripotent haematopoietic stem cells and normal early progenitors into the S-phase. 2. Angiotensin I-converting enzyme inhibitors, given as a single dose to normal subjects or during long-term treatment in hypertensive patients, result in plasma AcSDKP levels five- to six-fold higher and urine concentrations 40-fold higher than those of control subjects and/or patients. Thus, AcSDKP is a natural peptide hydrolysed by the N-terminal domain of ACE in vivo. In addition, ACE may be implicated in the process of haematopoietic stem cell regulation by permanently degrading this natural circulating inhibitor of cell entry into the S-phase. 3. Besides hydrolysis by ACE, the second very effective mechanism by which AcSDKP is cleared from plasma is glomerular filtration. Because of its high sensitivity and specificity, the measurement of AcSDKP in plasma and urine provides a valuable tool in screening specific inhibitors of the N-terminal domain of ACE and in monitoring ACE inhibition during chronic treatment. 4. The long-term consequences of AcSDKP accumulation are not known. During chronic ACE inhibition in rats, AcSDKP levels slightly increase in organs with high ACE content (kidneys, lungs). To significantly increase its concentration in target haematopoietic organs (the extracellular fraction of bone marrow), AcSDKP has to be infused on top of a captopril-based treatment. 5. A selective inhibitor of the N-domain of ACE in vitro and in vivo has been identified recently. The phosphinic peptide RXP 407 does not interfere with blood pressure regulation, but does increase, dose dependently, plasma concentrations of AcSDKP in mice, in contrast with lisinopril, which affects the metabolism of both AcSDKP and angiotensin I. N-Terminal-selective ACE inhibitors may be used to selectively control AcSDKP metabolism in target haematopoietic organs. This new therapeutic strategy may be of value for protecting haematopoietic cells from the toxicity of cancer chemotherapy. [source] | |||||||||||||