IFN-γ promoter polymorphisms do not affect QuantiFERON® TB Gold In-Tube test results in a Canadian population.

BACKGROUND: Several studies have shown polymorphisms within the interferon-gamma (IFN-γ) promoter influence cytokine expression. The interferon-gamma release assay (IGRA) relies on the ability to produce IFN-γ in response to tuberculosis (TB) specific antigens. This study determined the relationship between the IFN-γ +874 A/T promoter polymorphism and the performance of the QuantiFERON®-TB Gold In-Tube (QFT-GIT) test in an ethnically diverse Canadian population. METHODS: A total of 190 participants were categorised into three groups based on history of and exposure to TB: active TB (n = 55), TB exposed (n = 55) and presumably TB unexposed controls (n = 80). All participants underwent QFT-GIT testing, and DNA was extracted from whole blood and probed for polymorphism at position +874 (T/A) of intron 1 of IFN-γ. Statistical relationships between the QFT-GIT results, polymorphisms and demographic data were evaluated. RESULTS: IFN-γ +874 genotype frequencies among the entire study population (n = 190) were A/A (45.8%), T/A (39.5%), and T/T (14.7%). Among the three study groups, there was no correlation between QFT-GIT results and the IFN-γ +874 A/T genotype, and no correlation of genotype with IFN-γ production in response to either Mycobacterium tuberculosis antigens or mitogenic stimulation. CONCLUSION: Our results indicate that the IFN-γ +874 promoter polymorphism does not influence QFT-GIT performance in this study population.

Controlled hydrostatic pressure stress downregulates the expression of ribosomal genes in preimplantation embryos: a possible protection mechanism?

The efficiency of various assisted reproductive techniques can be improved by preconditioning the gametes and embryos with sublethal hydrostatic pressure treatment. However, the underlying molecular mechanism responsible for this protective effect remains unknown and requires further investigation. Here, we studied the effect of optimised hydrostatic pressure treatment on the global gene expression of mouse oocytes after embryonic genome activation. Based on a gene expression microarray analysis, a significant effect of treatment was observed in 4-cell embryos derived from treated oocytes, revealing a transcriptional footprint of hydrostatic pressure-affected genes. Functional analysis identified numerous genes involved in protein synthesis that were downregulated in 4-cell embryos in response to hydrostatic pressure treatment, suggesting that regulation of translation has a major role in optimised hydrostatic pressure-induced stress tolerance. We present a comprehensive microarray analysis and further delineate a potential mechanism responsible for the protective effect of hydrostatic pressure treatment.

Pycnogenol efficiency on glycaemia, motor nerve conduction velocity and markers of oxidative stress in mild type diabetes in rats.

The aim of this study was to describe the effects of Pycnogenol at various doses on preprandial and postprandial glucose levels, the levels of thiobarbituric acid reactive substances (TBARs) and N-acetyl-beta-d-glucosaminidase (NAGA) and on motor nerve conduction velocity (MNCV) in streptozotocin (STZ)-induced diabetic rats. Pycnogenol treatment (10, 20, 50 mg/kg body weight (b.w.)/day) lasted for 8 weeks after induction of diabetes. Pycnogenol significantly decreased elevated levels of preprandial glycaemia in treated animals at all doses. At doses of 10 mg/kg b.w./day and 20 mg/kg b.w./day it significantly decreased elevated levels of postprandial glycaemia compared with diabetic non-treated animals. Pycnogenol failed to induce a significant decrease of postprandial glycaemia at a dose of 50 mg/kg b.w./day. Pycnogenol improved significantly the impaired MNCV at doses of 10 and 20 mg/kg b.w./day compared with non-treated animals. The levels of TBARs were elevated in diabetic rats. The levels of NAGA increased gradually despite the treatment. Pycnogenol failed to affect the increased levels of TBARs and NAGA. Pycnogenollowered the elevated levels of glycaemia and reduced the decline in motor nerve conduction velocity in STZ-induced diabetic rats. The effect of Pycnogenol on postprandial glycaemic levels and MNCV was not dose-dependent.

Preservation of fertility in reproductive-age women with the diagnosis of cancer.

Over the past few decades the number of young, reproductive age cancer survivors has increased as a result of improved and less destructive cancer treatments. Certain types of cancers are predominantly diagnosed among reproductive age women and a small proportion of cancers originating in the reproductive tract are also detected in this age group. Treatment in the past used to be definitive and in most cases led to sterility. In recent years, improved medical treatments and more conservative surgical approaches have been introduced increasing the number of young survivors of cancer treatment. These less invasive treatments seem to be associated with similar survival rates and fertility can be preserved in most cases. This has led to studies evaluating the reproductive options of these women. Conservative surgical techniques, the use of chemotherapeutic agents with a reduced gonadotoxic side-effect profile, and the application of more focused radiation therapy are associated with maintenance of fertility. In addition, assisted reproductive technology (ART) has undergone tremendous improvements and now offers several alternatives to those who wish to maintain fertility before or even after cancer therapy. This review summarizes the fertility sparing medical and surgical as well as ART options that reproductive age women desiring to maintain fertility may utilize if they face cancer therapy.

Effect of VULM 1457, an ACAT inhibitor, on serum lipid levels and on real time red blood cell flow in diabetic and non-diabetic hamsters fed high cholesterol-lipid diet.

Acyl-coenzyme A: cholesterol O-acyltransferase (ACAT) catalyzes the formation of cholesterol/fatty acyl-coenzyme A esters. Accumulation of cholesterol esters leads to pathological changes connected with atherosclerosis. We have evaluated effects of a newly synthesized ACAT inhibitor, 1-(2,6-diisopropyl-phenyl)-3-[4-(4'-nitrophenylthio)phenyl] urea (VULM 1457), on serum lipid (cholesterol and triglycerides) levels and velocity of red blood cells (RBC) in non-diabetic and diabetic hamsters fed on high cholesterol-lipid (HCHL) diet during 3 months. The VULM 1457 effects on the paw microcirculation were assessed using capillary microscopy by measuring (RBC) velocity in vivo. Hamsters fed on HCHL diet became hypercholesterolemic with a dramatic increase in serum lipids accompanied with significantly decreased RBC velocity. Diabetic hamsters fed on HCHL diet had further increased serum lipids with reduction of RBC velocity. The VULM 1457 inhibitor lowered cholesterol levels in both non-diabetic and diabetic hamsters fed on HCHL diet. The greater VULM 1457 effect was shown in diabetic hamsters fed on HCHL diet where VULM 1457 expressed hypotriglycerides effects, too. An improved RBC velocity-pronounced effect was observed in diabetic hamsters fed on HCHL diet treated with VULM 1457. These results suggest that the ACAT inhibitor, VULM 1457, is a prospective hypolipidemic and anti-atherogenic drug which treats diabetes.

Reactive oxygen species induced smooth muscle responses in the intestine, vessels and airways and the effect of antioxidants.

Numerous experimental data confirm the importance of reactive oxygen species (ROS) in physiological activities of smooth muscles and in the pathogenesis of various diseases with altered function of smooth muscles. The present study shows that smooth muscles of the intestine, airways and vessels, as well as their epithelium, endothelium and innervations, might be important targets of the ROS action. We demonstrated differences among the actions of various ROS (endogenous, exogenous, produced enzymatically, non-enzymatically) as well as among their actions in different smooth muscle tissues. Our results indicate that ROS are involved in changes in muscle tone, membrane conductance, calcium homeostasis, calcium-dependent processes, as well as in eicosanoid and nitric oxide metabolism. The effects of antioxidative enzymes (superoxide dismutase, catalase), of several drugs of natural origin (e.g. Kampo Medicines) and synthetic agents (e.g. stobadine, nitrosopine, ACE inhibitors) suggest that smooth muscle tissues are useful models to study ROS action and drug intervention in ROS induced injuries.

Extracellular ATP increases [Ca2+]i in primarily cultured pig coronary smooth muscle cells via a P2Y purinoceptor subtype.

In primarily cultured pig coronary smooth muscle cells, extracellular adenosine triphosphate (ATP; 10(-9) to 10(-3) M) dose-dependently increases intracellular calcium ([Ca2+]i). The [Ca2+]i transients measured by fura-2 fluorescence consist of peak and plateau phases with [Ca2+]i values of 191.84 +/- 5.67 nM (n = 10) and 91.67 +/- 1.89 nM, respectively. In Ca(2+)-free solution, the peak phases persisted, but there was a loss of the plateau response, indicating an initial ATP-stimulated intracellular Ca2+ release and a subsequent transarcolemmal Ca2+ entry. Various agonists have been used to characterize the P2 purinoceptor subtype involved in the ATP-induced Ca2+ transients. The rank order of potency was uridine triphosphate (UTP) > ATP >> 2-meSATP > beta,gamma-meATP = alpha,beta-meATP = adenosine = 0. To examine the refilling of ATP-sensitive stores, four repetitive 60-s ATP responses were produced throughout with a 5-min recovery period in between. Now the ATP peaks gradually declined in Ca(2+)-free solution, indicating the emptying of the stores. If, however, Ca2+ entry was allowed in the "refilling period" (i.e., between the ATP pulses), the Ca2+ peaks could be maintained or restored, respectively. The data suggest that the ATP-dependent [Ca2+]i transients may be mediated via a UTP > ATP-activated P2Y purinoceptor subtype, mediating both an intracellular Ca2+ release and a transarcolemmal Ca2+ influx. The refilling of Ca2+ stores may occur through the unstimulated membrane after agonist stimulation. A putative pathway may be a "capacitative" Ca2+ entry induced on depletion of intracellular Ca2+ stores.

[Delivery following cryopreserved embryo transfer]. / Szülés mélyfagyasztva tárolt praeembriók beültetését követöen.

In this case report the authors describe the first delivery following cryopreserved preembryo transfer in Hungary. They concluded that this method may help to improve the cumulative pregnancy rate during in vitro fertilization cycles and alleviate the ethical problems associated with the destruction of viable preembryos.

Voltage dependence of the pharmacological Mg2+ block of the Ca2+ entry into vascular smooth muscle cells.

We studied the voltage dependence of the inhibitory effects of the Ca2+ antagonist Mg2+, compared to nifedipine, on the transsarcolemmal Ca2+ uptake into primary monolayers of smooth muscle cells obtained from pig coronary arteries. The cellular Ca incorporation was analyzed by liquid scintillation. Depolarization was produced by elevation of K+0. The data suggest that depolarization known to improve the inhibition by organic Ca2+ antagonists of L-type voltage-gated Ca2+ channels attenuates the Ca2+ antagonism of Mg2+ at vascular smooth muscle cells.

Effect of pentacaine and its derivatives on the contractile responses of smooth muscle in the guinea-pig stomach.

The effect of a carbanilic local anesthetic pentacaine [(+/-)-trans-2- (1-pyrrolidinyl)cyclohexyl ester of 3(n)-pentyloxyphenyl-carbanilic acid] and some of its derivatives [K-1905 [(+/-)-trans-2-diethylaminocyclopentyl ester of 3(n)-pentyloxyphenyl-carbanilic acid], K-2002 [(+/-)-trans-2-(1-pyrrolidinyl)cyclohexyl ester of 4(n)-pentyloxyphenyl-carbanilic acid], K-2006 [(+/-)-trans-2-(1-pyrrolidinyl)cyclopentyl ester of 4(n)-pentyloxyphenyl-carbanilic acid], and carbanilates P2 [(+/-)-trans-2-(1-pyrrolidinyl)cyclohexyl ester of 4-methoxy-carbonylphenyl-carbanilic acid], P3 [(+/-)-trans-2-(1-pyrrolidinyl)cyclohexyl ester of 3-methoxy-phenyl-carbanilic acid], and PeJ, the quaternized derivative of pentacaine), as well as that of oxethazaine was studied on longitudinal antral and circular fundic smooth muscle strips of the guinea-pig stomach. All the carbanilates studied relaxed the smooth muscle, attenuated the spontaneous smooth muscle contractions and shifted the acetylcholine, histamine, and BaCl2 cumulative concentration effect curves to the right, reducing their maximum. There was no direct relationship between their relaxing potency and the ability to reduce the action of different stimulants. For the effectiveness of the carbanilates studied, substitution in the lipophilic part of the molecule was more important than in the hydrophilic part and the meta position was more advantageous than the para position. Pentyloxy-derivatives (pentacaine, K-1905, K-2002 and K-2006) were more active than the methylcarbonyloxy (P2)- and methoxy (P3)-derivatives. Opening of the heterocyclic ring (K-1905) in the hydrophilic part of the molecule did not affect significantly the potency of the derivative studied, while quaternization (PeJ) significantly reduced the potency. It is suggested that the carbanilates studied may affect the smooth muscle responses via changes in the membrane fluidity and Ca2+ availability, and that these effects might be partly responsible also for their antiulcer activity.

Progression and regression by verapamil of vitamin D3-induced calcific medial degeneration in coronary arteries of rats.

Vitamin D3-induced mural calcification represents an animal model for investigating experimental calcium (Ca) overload and calcification of arterial walls. In this study, long-term progression of calcific degeneration in coronary arteries of rats after one intoxication with vitamin D3 was examined, as well as possible regression of preestablished mural Ca overload with the Ca antagonist verapamil. Sprague-Dawley rats were treated with one intramuscular (i.m.) overdose of vitamin D3 [300,000 IU/kg body weight (b.w.)]. Oral verapamil therapy (100 mg/kg/day b.w. for 24 weeks) was initiated 14 days after the vitamin D3 intoxication. Arteriosclerotic alterations were verified by microchemical analyses of tissue Ca and of cholesterol contents with atomic absorption spectroscopy (special graphite tube technique) and gas chromatography, respectively, and by standard histological techniques. Serum lipids were determined by sequential ultracentrifugation. Between week 3 and week 26 after the vitamin D3 injection, a progressive Ca incorporation from 448.8 +/- 110 to 1,310 +/- 166.3% of control values (i.e., coronary Ca content in 32-week-old untreated control rats = 100%) was observed, associated with calcific morphological lesions, and reactive intimal plaque formation. Verapamil prevented this progression and induced a regression of preestablished mural Ca overload. Therefore, the coronary Ca content after 24 weeks of verapamil treatment amounted to only 146.3 +/- 53.8% of controls. The data indicate that an initial calcific lesion of coronary arteries may serve as crystallization nucleus for advancing Ca overload and morphological alterations.(ABSTRACT TRUNCATED AT 250 WORDS)

A comparative study of guinea pig and rat tracheal reactivity.

Species differences in behavior and responses of guinea pig (GPT) and rat (RT) tracheal smooth muscle to electrical field stimulation (EFS) and chemicals were investigated under semi-isometric conditions. Indomethacin augmented and papaverine reduced the contractions elicited by EFS in both tissues. They relaxed the GPT and did not affect the tone decline in the RT. Experiments with different load, cartilage content, thread and rubber strips suggest the role of elastic elements in passive elongation of the RT. This was independent of muscarinic, adrenergic, histaminergic and serotoninergic receptors, nitric oxide, eicosanoids and metabolic processes. The second response on repeated administration of acetylcholine was augmented and that of serotonin attenuated in both GPT and RT. Upon repeated elevation of [K+]o the tissues responded in an opposite manner. Further data are provided on similarities (modulation of cholinergic transmission by prostaglandins, sensitization to acetylcholine and desensitization to serotonin) and differences (innervation, responses to repeated elevation of [K+]o, presence/absence of prostaglandin-regulated basal tone and spontaneous activity) in reactivity of GPT and RT.

[Smooth muscle preparations as models for studies of drug effects on organs]. / Hladkosvalové preparáty ako modelové orgány stúdia úcinku farmák.

The action of drugs on processes in smooth muscles, in their innervation or mucosa results in changes in contractility of the gut, airways, vessels and urogenital system. Noteworthy insight has been gained into the basic common characteristics ot smooth muscles as well as into special properties of individual smooth muscle types whose fundamental properties have become adapted to a particular situation. This insight along with knowledge on the subcellular and cellular organization of smooth muscle cells and of their innervation, on the role of the mucosa, and introduction of sophisticated electrophysiological, biochemical, isotopic and morphological methods makes smooth muscle suitable for investigation of elemental physiological and pathophysiological processes and of targets of drug action. The complexity of the smooth muscle tissue allows to study the mechanisms of drug action on the peripheral cholinergic, adrenergic, nonadrenergic-noncholinergic nerves and their neuromediators, on the epithelial and endothelial cells and the biologically active substances which they release, on the membrane and subcellular receptors, receptor coupled processes, ion channels, enzymes, Ca2+ availability, etc. Since most of these mechanisms operate also in other tissues, the obtained results may characterize drug action in other systems as well.

Involvement of different Ca2+ sources in changes of responsiveness of guinea-pig trachea to repeated administration of histamine and acetylcholine.

The role of Ca(i) and Ca(o) in changes of responsiveness of guinea pig tracheal smooth muscle strips to repeated applications of histamine and acetylcholine was investigated. Homologous desensitization to histamine developed when the airways were exposed to concentrations higher than 10(-5) mol/l, while sensitization to acetylcholine was recorded even when its highest concentration did not exceed 10(-5) mol/l. The maximum of the concentration response curves (CRC) was reduced upon repeated histamine, and enhanced upon repeated acetylcholine administration. There was, however, no significant difference in EC50 values for repeated CRCs of the stimulants. In Ca2+ free, EGTA (10(-4) mol/l) containing solution the second contraction elicited by single (10(-3) mol/l) or cumulative (10(-9)-10(-3) mol/l) histamine application was significantly smaller, while that elicited by acetylcholine did not differ significantly from the first one. In Ca(2+)-free, caffeine (10(-2) mol/l) and EGTA containing solution the contractile responses to repeated additions of Ca2+ (2.7 mmol/l) in histamine and acetylcholine (10(-3) mol/l) treated tracheae was decreased and unchanged, respectively. Addition of nifedipine (10(-6) mol/l) to this solution fully prevented Ca2+ in inducing contraction in histamine treated tracheae, while Ca2+ still induced contraction in acetylcholine treated tracheae. TMB-8 (10(-5) mol/l) was ineffective in blocking the remaining acetylcholine induced contractions. The present data suggest that contractions of the guinea pig trachea elicited by histamine and acetylcholine are due to release of intracellular Ca2+ from a caffeine sensitive store and to influx of Ca2+ from the extracellular space via voltage operated channels (VOC). Moreover, acetylcholine activated Ca2+ entry into guinea pig tracheal smooth muscle cells via the nifedipine insensitive mechanism, probably receptor operated channels (ROC). It is concluded that desensitization to histamine in the guinea pig trachea is most probably due to alterations in intracellular Ca2+ mobilization and Ca2+ influx via VOC. In contrast, sensitization to acetylcholine involved primarily enhanced Ca2+ influx via VOC and Ca2+ induced Ca2+ release.

Experimental vasoprotection by calcium antagonists against calcium-mediated arteriosclerotic alterations.

According to chemical analyses, the development of conventional human coronary artery plaques from "fatty streaks" to "fibrous plaques" and "complicated lesions" is dominated by progressive mural calcium (Ca) incorporation. The atherogenic significance of Ca ions and arterial Ca overload was examined under the influence of nicotine, oxidatively modified low-density lipoproteins, spontaneous hypertension, and an elevated extracellular Ca concentration or vitamin D3. Experiments were carried out either in vitro on cultured medial cells of rats or in vivo on various types of experimental arteriosclerosis of rats. Suitable Ca antagonists (verapamil, diltiazem, nifedipine, or nitrendipine) prevented experimental Ca overload of arterial walls and transmembrane Ca uptake into cultured medial cells produced by risk factors. Thus they protected, in vivo and/or in vitro, against the atherogenic potential of Ca ions, i.e., migration, proliferation, matrix production and intracellular Ca overload of vascular smooth-muscle cells, as well as calcification of elastic fibers. The data indicate that various Ca-consuming processes demand a progressive uptake of Ca into arterial walls if Ca-dominated types of arteriosclerosis develop. Under experimental conditions, specific Ca antagonists inhibit Ca-mediated arteriosclerotic alterations by preventing progressive mural Ca incorporation.

Effect of stobadine and histamine H1 and H2 blockers on histamine-induced contraction of guinea pig airways in vitro.

Histamine, a nonselective histamine receptor agonist, activates simultaneously both H1 and H2 receptors in the guinea pig trachea and lung strip. The resulting contraction is due to the prevalence of H1 receptors, because they are blocked by the selective H1 antagonist diphenhydramine. The H2 receptor antagonists cimetidine and ranitidine increased the sensitivity of both tissues to histamine by affecting primarily the amplitude of contractions induced by high histamine concentrations. Since the lung strips were more sensitized by ranitidine and low concentrations of the other studied antagonists (diphenhydramine, dithiadene, stobadine) than the tracheal smooth muscle, it is inferred that the density of H2 receptors is higher in peripheral than central airways. From all the studied histamine receptor antagonists only dithiadene was able to unmask the relaxation induced by H2 receptor activation indicative of its highest H1 selectivity. In the light of the concentration-dependent antihistaminic effect of stobadine, i.e. potentiation in low and inhibition in high concentrations, stobadine is suggested to belong to antihistaminics with no histamine receptor subtype selectivity.