Inhibition of antigen-induced cutaneous responses of ponies with insect hypersensitivity by the histamine-1 receptor antagonist chlorpheniramine

A whole-body extract of Culicoides impunctatus induced a biphasic increase in oedema formation in ponies with insect hypersensitivity, with maxima after one and eight hours. The Culicoides antigen did not induce similar responses in ponies with no previous history of the disease. In insect-hypersensitive ponies the local administration of chlorpheniramine (12 micrograms) completely inhibited oedema formation in response to histamine (0.04 microgram) and to Culicoides antigen (0.5 microgram) at one hour, and the response to Culicoides antigen at eight hours was inhibited by 63 per cent. Chlorpheniramine also partially inhibited the accumulation of eosinophils and neutrophils induced by Culicoides antigen after two hours.     

Effect of a 5-lipoxygenase inhibitor and leukotriene antagonist (PF 5901) on antigen-induced airway responses in neonatally immunized rabbits

1. The effect of a single intratracheal dose (10 mg) of PF 5901 (2-[3(1-hydroxyhexyl) phenoxymethyl] quinoline hydrochloride, a specific inhibitor of the 5-lipoxygenase pathway of arachidonic acid metabolism and a leukotriene D4 antagonist) on airway changes induced in response to Alternaria tenuis aerosol challenge was assessed in adult rabbits neonatally immunized. Leukotriene generation was determined in vivo by measuring leukotriene B4 (LTB4) levels in bronchoalveolar lavage (BAL) fluid and ex vivo by measuring calcium ionophore-stimulated production of LTB4 in whole blood. 2. While PF 5901 (10 mg) had no significant effect on the acute bronchoconstriction induced by antigen, this dose was sufficient to inhibit significantly the increase in airway responsiveness to inhaled histamine 24 h following antigen challenge (P < 0.05). 3. Total leucocyte infiltration into the airways induced by antigen, as assessed by bronchoalveolar lavage, was significantly inhibited by pretreatment with PF 5901 (10 mg). However, the pulmonary infiltration of neutrophils and eosinophils induced by antigen was unaltered by prior treatment with PF 5901 (10 mg). 4. PF 5901 (10 mg) had no effect on ex vivo LTB4 synthesis in whole blood. However, the antigen-induced increase in LTB4 levels in BAL 24 h following challenge was significantly inhibited (P < 0.05). 5. We suggest from the results of the present study that the antigen-induced airway hyperresponsiveness to inhaled histamine in immunized rabbits is mediated, at least in part, by products of the 5-lipoxygenase metabolic pathway, and is not dependent on the extent of eosinophil or neutrophil influx into the airway lumen.     

Time-course of antigen-induced airway inflammation in the guinea-pig and its relationship to airway hyperresponsiveness

The causative relationship between airway inflammation and hyperreactivity is unclear, since inflammatory changes have been examined at one or, at most, a few time-points after antigen challenge in both human asthma and animal models. We have made a detailed investigation of inflammatory and functional changes in the airways up to 8 days after antigen challenge in guinea-pigs. In particular, we examined the hypothesis that eosinophil-derived mediators contribute to tissue damage and the development of airway hyperresponsiveness. Following antigen challenge, the influx of inflammatory cells and mediator release in airway tissue and bronchoalveolar lavage fluid were correlated temporally with histopathological changes in airway tissue and airway responsiveness. Eosinophil influx was demonstrable at 4 h. Eosinophilia peaked after 24 h and persisted for at least 8 days. Parallel increases in the concentrations of major basic protein and eosinophil cationic protein in bronchoalveolar lavage fluid indicated that the eosinophils were activated. Eosinophilia was accompanied by subepithelial oedema and epithelial damage co-localized with major basic protein immunoreactivity. A transient neutrophilia (< 48 h duration) and an increase in neutrophil elastase in bronchoalveolar lavage fluid peaked at 14 h. The proportion of airway macrophages with an activated morphology increased at 8 h and remained markedly elevated until 72 h. Airways were hyperresponsive to histamine at 4 h and for at least 8 days. The antigen-induced airway inflammation resemble in time-course and histopathology that seen in antigen-challenged asthmatics, and indicate that the eosinophil and its cytotoxic proteins may be major mediators of airway mucosal damage and airway hyperresponsiveness.     

Inhibition of antigen-induced muscle contractions by inhibitors of thromboxane pathway in rat small intestine

The cell cycle is driven by the sequential activation of a family of cyclin-dependent kinases (CDK) in association with cyclins. In mammalian cells the timing of activation of cyclin A-associated kinase activity coincides with the onset of DNA synthesis in S-phase. Using in vitro replication of SV40 origin-containing DNA as a model system, we have analyzed the proteins associated with DNA during initiation of DNA replication in S-phase cell extracts. This analysis reveals that, in addition to replication initiation proteins, cyclin A and cdk2 are also specifically associated with DNA. The association of cyclin A and cdk2 with DNA during initiation is cell cycle regulated and occurs specifically in the presence of SV40 origin-containing plasmid and SV40 T antigen (the viral replication initiator protein). The interactions among proteins involved in initiation play an important role in DNA replication. We therefore investigated the ability of cyclin A and cdk2 to associate with replication initiation proteins. Under replication initiation conditions, cyclin A and cdk2 from S-phase extracts specifically associate with SV40 T antigen. Further, the interaction of cyclin A-cdk2 with SV40 T antigen is mediated via cyclin A, and purified recombinant cyclin A associates directly with SV40 T antigen. Taken together, our results suggest that cyclin A and cdk2 are components of the SV40 replication initiation complex, and that protein-protein interactions between cyclin A-cdk2 and T antigen may facilitate the association of cyclin A-cdk2 with the complex.     

Inhibition by dexamethasone of antigen-induced c-Jun N-terminal kinase activation in rat basophilic leukemia cells

Antigen stimulation of IgE-sensitized rat basophilic leukemia RBL-2H3 cells induced activation of c-Jun N-terminal kinase (JNK) within a few minutes with maximum activity attained 40 min later. The increase in JNK activity was accompanied with an increase in phosphorylation of c-Jun in the cells. The Ag-induced JNK activation was inhibited by the phosphatidylinositol 3-kinase inhibitors wortmannin (10-100 nM) and LY 294002 (100 microM) but not by the protein kinase C inhibitors calphostin C (1 and 3 microM) and Ro 31-8425 (1 and 3 microM). Pretreatment with dexamethasone (10 and 100 nM) for 18 h inhibited the Ag-induced increase in JNK activity in a concentration-dependent manner. At least 6 h of preincubation with dexamethasone was necessary to inhibit the Ag-induced JNK activation. The phosphorylation of c-Jun induced by the Ag stimulation was reduced by pretreatment with dexamethasone without reduction of the content of c-Jun protein. The Ag-induced activation of the JNK kinase kinase mitogen-activated protein kinase-extracellular signal-regulated kinase kinase-1 was also inhibited by pretreatment with dexamethasone at 10 and 100 nM. These findings indicate that dexamethasone reduces JNK protein level and inhibits the Ag-induced activation of JNK resulting in the inhibition of c-Jun phosphorylation.     

Inhibition of smooth muscle cell proliferation and neointimal growth by low-anticoagulant heparin

Low-anticoagulant heparin (LA-heparin) obtained by affinity chromatography on antithrombin III Sepharose inhibits the proliferation of cultured arterial smooth muscle cells in an in vitro bioassay system as effectively as standard heparin. A growth inhibition of smooth muscle cells of about 60% is achieved when LA-heparin or heparin is added to the culture medium to a concentration of 50 micrograms/ml. In normolipemic rats LA-heparin suppresses the formation of neointimal thickenings and stenosis after balloon catheter-induced deendothelialization of the carotid artery. In terms of mass a dose of 5 mg/kg body weight/d given subcutaneously twice daily one week before and 2 weeks after balloon injury the cross sectional area of the neointima is reduced to 36% as compared with the nontreated control group (100%). This 64% reduction is statistically highly significant (p < 0.001). After treatment with 0.5 mg LA-heparin/kg/d the reduction of the neointima was 11% (p < 0.05). At a dose of 5 mg/kg body weight single or repeated administrations of LA-heparin caused only a small and transient increase in activated partial thromboplastin time values. The results show that subcutaneous administration of LA-heparin very effectively prevents smooth muscle cell proliferation and balloon catheter-induced neointimal growth. The well tolerated systemic application of this chemically non-modified LA-heparin might justify clinical trials for prevention of restenosis after percutaneous transluminal coronary angioplasty or other invasive cardiovascular interventions without complications of bleeding.     

Inhibitory effect of NPC-17731 on BK-induced and antigen-induced airway reactions in guinea-pigs

PURPOSE: To identify which aspects of social relations among 70-year-old men and women are predictive of mortality 11 years later. METHODS: The baseline study in 1984 included 734 70-year-old men and women in Glostrup (county of Copenhagen). The variables comprised the structure and the function of the social network, education, income and functional ability. Eleven years later, in November 1995, information about deaths was obtained from the Central National Register. RESULTS: The study showed an independent association between social relations and mortality. Men who did not help others with repairs and who lived alone and women with no social support to other tasks had increased risk of dying during the follow-up period. CONCLUSIONS: This study supports (1) that there is an association between social relations and mortality, (2) that two aspects of the function of social relations matters: (a) to receive support for small or larger tasks needed, (b) to help others with different tasks, and (3) that social relations may serve different functions for men and women.     

Inhibition of meizothrombin and meizothrombin(desF1) by heparin cofactor II

Meizothrombin and meizothrombin(desF1) are intermediates formed during the conversion of prothrombin to thrombin by factor Xa, factor Va, phospholipids, and Ca2+ (prothrombinase). These intermediates are active toward synthetic peptide substrates but have limited ability to interact with platelets or macromolecular substrates such as fibrinogen. Meizothrombin and meizothrombin(desF1) activate protein C, however, and may exert primarily an anticoagulant effect. In this study, we investigated the inhibition of meizothrombin and meizothrombin(desF1) by two glycosaminoglycan-dependent protease inhibitors, heparin cofactor II (HCII) and antithrombin (AT). Purified recombinant meizothrombin and meizothrombin(desF1) were inhibited by HCII in the presence of dermatan sulfate with maximal second-order rate constants of 8 x 10(6) M-1.min-1 and 1.8 x 10(7) M-1.min-1, respectively, but were inhibited less than one-tenth as fast by AT in the presence of heparin. Similarly, the products of the prothrombinase reaction were inhibited in situ more effectively by HCII than by AT. When HCII and dermatan sulfate were present continuously during the prothrombinase reaction, meizothrombin was trapped as a sodium dodecyl sulfate-stable complex with HCII and no amidolytic activity could be detected with a thrombin substrate. Our findings indicate that HCII is an effective inhibitor of meizothrombin and meizothrombin(desF1) and, therefore, might regulate the anticoagulant activity of these proteases.     

Prevention of Th2-like cell responses by coadministration of IL-12 and IL-18 is associated with inhibition of antigen-induced airway hyperresponsiveness, eosinophilia, and serum IgE levels

Allergic asthma is thought to be regulated by Th2 cells, and inhibiting this response is a promising mode of intervention. Many studies have focused on differentiation of Th cells to the Th1 or Th2 subset in vitro. IL-4 is essential for Th2 development, while IL-12 induces Th1 development, which can be enhanced by IL-18. In the present study, we investigated whether IL-12 and IL-18 were able to interfere in Th2 development and the associated airway symptoms in a mouse model of allergic asthma. Mice were sensitized with OVA using a protocol that induces IgE production. Repeated challenges by OVA inhalation induced elevated serum levels of IgE, airway hyperresponsiveness, and a predominantly eosinophilic infiltrate in the bronchoalveolar lavage concomitant with the appearance of Ag-specific Th2-like cells in lung tissue and lung-draining lymph nodes. Whereas treatments with neither IL-12 nor IL-18 during the challenge period were effective, combined treatment of IL-12 and IL-18 inhibited Ag-specific Th2-like cell development. This inhibition was associated with an absence of IgE up-regulation, airway hyperresponsiveness, and cellular infiltration in the lavage. These data show that, in vivo, the synergistic action of IL-12 and IL-18 is necessary to prevent Th2-like cell differentiation, and consequently inhibits the development of airway symptoms in a mouse model of allergic asthma.     

Effects of uroguanylin and guanylin against antigen-induced bronchoconstriction and airway microvascular leakage in sensitized guinea-pigs

Uroguanylin and guanylin are isolated mainly from the gastrointestinal tract and are activators of guanylyl cyclase C receptor (GC-C), which mediates the production of intracellular cyclic guanosine 3',5'-monophosphate (cyclic GMP). The bronchodilator effects of agents that raise cyclic GMP levels, such as atrial natriuretic peptide, have been reported, and uroguanylin mRNA has recently been detected in extra-gastrointestinal tissues, including the lung, suggesting their role in pulmonary activity. In the first step of this study, we examined the relaxant effects of uroguanylin and guanylin on isolated tracheal smooth muscle of guinea-pigs, and measured tissue cyclic GMP levels by means of enzymeimmunoassay. Uroguanylin produced concentration-dependent relaxant effects on resting tone and significant elevated cyclic GMP levels. Guanylin produced the same, but less potent, effects. In this study, we first investigated the effects of uroguanylin and guanylin on antigen-induced bronchoconstriction and airway microvascular leakage in actively sensitized guinea-pigs. Anesthetized male guinea-pigs, ventilated via a tracheal cannula, were placed in a plethysmograph to measure pulmonary mechanics for 10 min after challenging with 1 mg/kg of ovalbumin. Evans blue dye was then extravasated into their airway tissues to measure microvascular leakage. Intravenous pretreatment with uroguanylin significantly inhibited ovalbumin-induced bronchoconstriction and microvascular leakage in a dose-dependent manner. These inhibitory effects were mimicked by 8-bromoguanosine 3', 5'-cyclic monophosphate. This study is the first to show that uroguanylin not only had a potent bronchodilatory effect but also inhibited microvascular leakage. These results encouraged us to continue the above experimental and clinical studies in bronchial asthma.     

Characterization of four basic models of indirect pharmacodynamic responses

Four basic models of indirect pharmacodynamic responses were characterized in terms of changing dose, Imax or Smax, and IC50 or SC50 to examine the effects of these fundamental drug properties on response profiles. Standard pharmacokinetic parameters were used for generating plasma concentration, and response-time profiles using computer simulations. Comparisons to theoretical expectations were made. In all four models, the maximum response (Rmax) (inhibition or stimulation) and the time of its occurrence (TRmax) were dependent on the model, dose, Imax or Smax, and IC50 or SC50 values. An increase in dose or a decrease in IC50 or SC50 by the same factor produced, as theoretically expected, identical and superimposable pharmacodynamic response patterns in each of the models. Some parameters (TRmax, ABEC) were nearly proportional to log dose, while others (Rmax, CRmax) were nonlinear. Assessment of expected response signature patterns as demonstrated in this report may be helpful in experimental designs and in assigning appropriate models to pharmacodynamic data.     

Inhibition by heparin of thrombin-catalyzed activation of the factor VIII-von Willebrand factor complex

The activation of factor VIII (fVIII) by thrombin is associated with heavy chain cleavages at Arg372 and Arg740 and light chain cleavage at Arg1689. In a defined, plasma-free assay of fVIII activation and at physiological ionic strength and pH, heparin inhibited the rate of activation of either human or porcine fVIII by thrombin in either the presence or absence of von Willebrand factor (vWf). The inhibitory effect of heparin was associated with inhibition of all three thrombin-catalyzed bond cleavages. At plasma concentrations of fVIII (approximately 1 nM) and vWf (approximately 35 nM), the rate of fVIII activation was inhibited by 50% at approximately 0.1 unit/ml heparin, which is below the normal range of heparin concentrations in plasma during therapeutic anticoagulation (0.2-0.7 unit/ml). We propose that, in addition to catalyzing the inhibition of thrombin and other intrinsic pathway coagulation proteases by antithrombin, heparin functions as an anticoagulant by direct inhibition of the activation of the fVIII-vWf complex by thrombin.     

Inhibition by imipramine of ATP-evoked responses in rat pheochromocytoma cells

Determination of allantoin in bovine milk based on high-performance liquid chromatography (HPLC) is described. Following dilution and filtration, milk samples were analysed directly. Separation and quantification of allantoin was achieved using a Spherisorb 5 NH2 column (250 x 4.6 mm ID), acetonitrile-water (90:10, v/v) mobile phase at a flow-rate of 2.0 ml min(-1), temperature 20 degrees C and monitoring the effluent at 214 nm. Total analysis time was 10 min. Recovery of allantoin added to milk was 97 (+/-3.7, n = 30)%. Lowest detectable concentration was 1 micromol l(-1). Within-day and between-day variability were less than 3%. Advantages of improved retention and separation of allantoin, and less complicated sample preparation exist over current methods.     

Fifteen years of cutaneous and mucocutaneous leishmaniasis in Bolivia: a retrospective study

The photosensitizing properties of p-nitroacetophenone (PNAP), a well-known radiosensitizer, have been studied in near UV region. The mechanism of PNAP photosensitization has been investigated by testing the efficiency of singlet oxygen production using photooxidation of 2,2,6,6-tetramethylpiperidine (TEMP) and photodegradation of guanosine. In both the cases, the enhancement effect of deuterated solvents has been observed. Results of these experiments suggest the significant role of type II mechanisms in PNAP photosensitization.     

Pharmacokinetics of heparin and its pharmacodynamic effect on plasma lipoprotein lipase activity and coagulation in healthy horses

We evaluated the pharmacokinetics of IV administered sodium heparin and the pharmacodynamic effect of heparin on lipoprotein lipase (LPL) activity. Horses were allotted to 3 groups. Plasma samples were obtained from each horse before and at various times for 6 hours after heparin administration for determination of heparin concentration, LPL activity, and activated partial thromboplastin time (APTT). The disposition of heparin was dose dependent. The area under the plasma heparin concentration vs time curve (AUC) increased more than proportionally with dose, indicating that heparin elimination was nonlinear. Total clearance of heparin was similar after the 40 and 80 IU/kg of body weight dosages, averaging 0.45 and 0.36 IU/kg/min, respectively. However, after administration of the 120 IU/kg dose, clearance was significantly less than that after the 40 IU/kg dose. The half-life of heparin averaged 53, 70, and 136 minutes after 40, 80, and 120 IU/kg, respectively, with significant differences observed between the low and high doses. In contrast to heparin, the area under the plasma concentration vs time curve for LPL activity increased less than proportionally with dose. Maximal LPL activity observed was independent of dose, averaging 4.8 mumol of free fatty acids/ml/h. The APTT was significantly prolonged for 120 minutes after administration of the 40 IU/kg dose. Correlation coefficients for LPL activity vs either plasma heparin concentration or APTT were less than 0.7, indicating that neither laboratory measure can be used to accurately predict plasma LPL activity.     

Inhibition of human T-cell responses to house dust mite allergens by a T-cell receptor peptide

Recent analysis of the usage of T-cell receptor (TcR) beta chain variable region (V beta) gene elements by house dust mite (HDM)-reactive T cells from an atopic donor suggested that TcR-V beta 3 gene products may form a major component of the human T-cell repertoire reactive to this common allergen. In this study a peptide analog of the TcR-V beta 3 complementarity determining region 2 (CDR2) is shown to inhibit the polyclonal human T-cell response to HDM; this effect is specific because inhibition is dependent on the presence of V beta 3 + T cells. This experimental approach has been used to determine whether the pattern seen in T-cell clones derived from one atopic donor reflects TcR-V beta usage in the polyclonal response to allergen in the general population. Inhibition of more than 50% of the polyclonal response to allergen by V beta 3-CDR2 peptide was observed in 16 of 21 donors tested, suggesting that TcR-V beta 3 gene usage may form a major component of the human HDM repertoire and as such offer a suitable target for T cell-directed specific immunotherapy in HDM-allergic individuals. Depletion of CD8+ T cells abolishes peptide-mediated inhibition of CD4+ T-cell proliferation to HDM, suggesting that induction of a CD8+ regulatory T-cell subset by the CDR2 peptide may modulate HDM-specific allergic T-cell responses.     

Inhibition of bovine dentin demineralization by a glutardialdehyde pretreatment: an in vitro caries study

A substantial literature shows that late components of visual evoked potentials (VEPs) are delayed in at least some forms of dementia in the elderly. The late-component delay is selective in that earlier components are not affected. More recent work with better defined clinical groups suggests that the selective late-component delay may be characteristic of Alzheimer's disease (AD) rather than an inevitable feature of dementia in general. To overcome problems in traditional VEP component latency measurement the present study uses reference-free Global Field Power (GFP) analysis to more objectively define VEP components and finds that the GFP peak corresponding to the late P2 component of the flash VEP is delayed in a probable AD group but not in a demented unlikely AD group, relative to age-equivalent healthy controls. The late-component delay is again found to be selective in that the GFP peak corresponding to the earlier P1 component of the flash VEP does not differ between groups. These findings further strengthen the evidence for electrocortical changes in the visual system of AD patients.     

Inhibition of nicotinic responses by cotinine in bovine adrenal chromaffin cells

We studied the effects of cotinine, the major metabolite of nicotine, on nicotine-induced increase in [3H]phorbol dibutyrate binding, activation of protein kinase C and [3H]noradrenaline release in primary cultured bovine adrenal chromaffin cells. Cotinine (1 mM, 15 min.) and nicotine (10 microM, 5 min.) increased the [3H]phorbol binding by 100% and 150%, respectively. Both a short-term (10 min.) and a long-term (24 hr) pretreatment with cotinine inhibited the effect of nicotine. A 24 hr pretreatment with cotinine (1 mM) also reduced the nicotine-induced increase in membrane-bound protein kinase C activity. Cotinine pretreatment (10 min.) dose-dependently inhibited the release of [3H]noradrenaline induced by nicotine and dimethylphenylpiperazinium. Cotinine pretreatment did not reduce the [3H]noradrenaline release induced by high extracellular potassium (56 mM) or veratrine (10 mg l-1). The results indicate that cotinine inhibits activation of protein kinase C and noradrenaline release induced by nicotinic agonists in primary cultures of bovine adrenal chromaffin cells. The results suggest that pre-existing cotinine could modify responses to acute exposure to nicotine in neural systems.