Effects of ketotifen on human lymphocytes in vitro and in vivo

The effects of the antiasthmatic drug ketotifen (CAS 34580-13-7) on human mononuclear leukocytes were studied in vitro and in vivo. In vitro ketotifen concentration-dependently inhibited mitogen-stimulated lymphocyte proliferation. High ketotifen concentrations also inhibited T-lymphocyte mitogen- and adenosine triphosphate stimulated increases in intracellular Ca2+ in lymphocytes and the U937 human monocyte precursor cell line, respectively; this involved inhibition of both Ca2+ influx and intracellular mobilization. In in vivo experiments, treatment of healthy volunteers with 1 mg ketotifen b.i.d. for 7 d did not alter the number or subset composition of circulating lymphocytes. Moreover, the mitogen-stimulated in vitro proliferation of lymphocytes obtained before and after ketotifen treatment in vivo was similar. It is concluded that high ketotifen concentrations can inhibit the activation of resting lymphocytes in vitro but standard ketotifen treatment does not notably affect the number of function of circulating lymphocytes in vivo.     

The prognostic significance of deletion of the long arm of chromosome 20 in myeloid disorders

OBJECTIVE: The effect of cigarette smoking on gallbladder (GB) emptying and refilling after a fatty meal was examined in 10 healthy volunteers (four women and six men, mean age 27.6 yr). METHODS: On three different days, the subjects underwent in randomized order: a control test without smoking (C), or they smoked two cigarettes during the early (0-20 min; S0-20), or late (20-40 min; S20-40) phase of the meal-induced GB emptying. GB volumes were measured ultrasonographically before the meal and at 10, 20, 30, 40, 60, 90, 120, 150, and 180 min postprandially. Two-way ANOVA was applied for statistical assessment of the results. RESULTS: The fasted GB volumes amounted to 15.7 +/- 1.8 cm3 (C), 15.0 +/- 1.7 cm3 (S0-20), and 18.4 +/- 2.3 cm3 (S20-40), F2;18 = 1.524, NS. Maximum GB emptying was observed until 60 min after the meal, with a nadir of the GB volume amounting to 7.3 +/- 1.3 cm3 (C), 6.6 +/- 1.2 cm3 (S0-20), and 7.1 +/- 1.1 cm3 (S20-40). No significant difference was found between the stimuli tested when absolute GB volumes were considered: F2;180 = 2.725, NS. Analysis of the GB emptying-refilling curves normalized for the fasted GB volume revealed that a significant inhibitory effect was produced by smoking two cigarettes during the late phase of GB emptying on the subsequent GB refilling: F2;162 = 11.066, p < 0.001 for the whole curve, and F2;72 = 7.126, p < 0.005 for the refilling phase. A significant contrast was found next between S20-40 and the control day (p < 0.001 whole curve; p < 0.005 refilling phase only), as well as between S20-40 and S0-20 (p < 0.001 whole curve; p < 0.025 refilling phase only). CONCLUSION: We conclude that smoking two cigarettes does not disturb the fatty meal-induced GB contraction in healthy humans. Subsequent GB refilling is delayed if smoking takes place during the late phase of the postprandial GB contraction.     

The effect of different protein diets on longevity and various biochemical parameters of aged rats]

In this work 23 month old rats were fed for 200 days with different protein diets (NT-diet: 19% protein, 72% of animal origin and LP-diet: 8.8% protein exclusively of vegetable origin). Some metabolic parameters and lifespan (on the base of a 50% death-rate) were determined. The relations of the liver free amino acids glycine + alanine and tyrosine + phenylalanine + branched chain amino acids and the ratio of phenylalanine/tyrosine were determined to be higher in the LP-group. Phenylalanine in liver and urea concentrations in liver and serum were lower in the LP-group. Furthermore the dopamine or serotonin levels were significantly lower in lateral and medial or lateral regions of the hypothalamus respectively in LP-diet fed rats. The norepinephrine content was not modified by the diets. The median lifespan of 23 month old rats was higher by 24% following LP-treatment. These results suggest that the protein component (amino acids) of different diets may modify metabolic parameters and lifespan of animals by mechanisms in which the central regulation may be involved.     

Plasma viral RNA load predicts disease progression in accelerated feline immunodeficiency virus infection

Viral RNA load has been shown to indicate disease stage and predict the rapidity of disease progression in human immunodeficiency virus type 1 (HIV-1)-infected individuals. We had previously demonstrated that feline immunodeficiency virus (FIV) RNA levels in plasma correlate with disease stage in infected cats. Here we expand upon those observations by demonstrating that plasma virus load is 1 to 2 logs higher in cats with rapidly progressive FIV disease than in long-term survivors. Differences in plasma FIV RNA levels are evident by 1 to 2 weeks after infection and are consistent throughout infection. We also evaluated humoral immune responses in FIV-infected cats for correlation with survival times. Total anti-FIV antibody titers did not differ between cats with rapidly progressive FIV disease and long-term survivors. These findings indicate that virus replication plays an important role in FIV disease progression, as it does in HIV-1 disease progression. The parallels in virus loads and disease progressions between HIV-1 and FIV support the idea that the accelerated disease model is well suited for the study of therapeutic agents directed at reducing lentiviral replication.     

Dissecting cAMP binding domain A in the RIalpha subunit of cAMP-dependent protein kinase. Distinct subsites for recognition of cAMP and the catalytic subunit

The two gene-duplicated cAMP binding domains in the regulatory subunits of cAMP dependent protein kinase are each comprised of an A helix, an eight-stranded beta-barrel, and a B and C helix (1). The A domain is required for high affinity binding to C, while the B domain regulates access to the A domain. Using a combination of a yeast two-hybrid screen coupled with deletion analysis, cAMP binding domain A of RI was dissected into two structurally and functionally distinct subsites, one that binds cAMP and another that binds the C subunit. The minimum stable subdomain required for binding to C in the 1-3 micromolar range is composed of residues 94-169, while residues 236-244, mapped to the C helix of cAMP binding domain A, were defined as a second surface necessary for high affinity (5-10 nanomolar) binding to C. This portion of the C helix, due to its position directly between the two subsites, serves as a molecular switch for either a cAMP-bound conformation or a C-bound conformation and can thus modulate interactions of cAMP binding domain A with cAMP, with C, and with cAMP binding domain B.     

Pulmonary vasoconstrictor effects of prostacyclin in rats: potential role of thromboxane receptors

Endogenous prostacyclin (PGI2; epoprostenol) is a potent endothelium-derived pulmonary vasodilator. However, the effects of exogenous PGI2 on isolated arteries could be either relaxant or contractile, depending on the species and organ studied. The present study investigated the distal pathways involved in the PGI2-induced contraction in rat intrapulmonary artery (PA) and relaxation in lamb PA. When vessels were precontracted with 30 mM K+, PGI2 (1 microM) induced relaxation in lamb PA but caused contraction in rat PA. Use of 30 mM K+, phenylephrine, serotonin, angiotensin II, or hypoxia to precontract the vessels did not alter the contractile effect of PGI2 in rat PA. Nevertheless, PGI2 produced a mild relaxation in rat PA precontracted by U-46619, a thromboxane A2 (TxA2)-receptor agonist, whereas the TxA2-receptor blocker SQ-29548 (0.1-0.5 microM) abolished the contractile response in rat PA. These data suggest that PGI2-induced contraction is mediated by activation of TxA2 receptors. The PGI2-induced modest relaxation in rat PA, which was only observed when TxA2 receptors were blocked by SQ-29548, suggests that the PGI2-mediated vasorelaxant pathway is diminished in these vessels. Simultaneous application of forskolin, an adenylate cyclase activator, and rolipram, a phosphodiesterase inhibitor, caused similar relaxation in both rat and lamb PA. This suggests that the adenosine 3',5'-cyclic monophosphate-dependent relaxing pathway is intact in rat PA and is comparable to that in lamb PA. On the basis of these data, we conclude that the pathways responsible for the paradoxical effects of PGI2 on rat and lamb PA are located upstream of the adenosine 3',5'-cyclic monophosphate-dependent relaxing pathway and that a paucity of PGI2 receptors in rat PA may be responsible.     

Replication of O6-methylguanine-containing DNA by repair and replicative DNA polymerases

The biological consequences of O6-methylguanine (m6G) in DNA are well recognized. When template m6G is encountered by DNA polymerases, replication is hindered and trans-lesion replication results in the preferential incorporation of dTMP opposite template m6G. Thus, unrepaired m6G in DNA is both cytotoxic and mutagenic. Yet, cell lines tolerant to m6G in DNA have been isolated, which indicates that some cellular DNA polymerases may replicate m6G-containing DNA with reasonable efficiency. Previous reports suggested that mammalian pol beta could not replicate m6G-containing DNA, but we find that pol beta can catalyze trans-lesion replication; however, the lesion must reside in the optimal context for pol beta activity, single- or short nucleotide gapped substrates. Primed single-stranded DNA templates, with or without template m6G, were poor substrates for pol beta as reported in earlier studies. In contrast, trans-lesion replication by bacteriophage T4 DNA polymerase was observed for primed single-stranded DNA templates. Replication of m6G-containing DNA by T4 DNA polymerase required the gp45 accessory protein that clamps the polymerase to the DNA template. The rate-limiting step in replicating m6G-containing DNAs by both DNA polymerases tested was incorporation of dTMP across from the lesion.