Post mortem development of meat quality as related to changes in cytoskeletal proteins of chicken muscles.

1. A procedure was developed to separate high and medium molecular weight myofibrillar proteins from chicken muscular tissue with a high resolution by flat bed sodium-dodecyl-sulphate polyacrylamide gel electrophoresis (SDS-PAGE) and subsequent detection by either a general protein stain or Western blotting. These procedures were used to analyse the degradation process of cytoskeletal proteins in chicken breast and leg muscles during meat ageing. 2. This study demonstrates the degradation of all the examined cytoskeletal proteins: titin, nebulin and desmin as well as vinculin, a protein component of the costamere structure. All the examined proteins were found to be degraded during ageing of chicken breast and leg muscles. 3. Degradation of titin, nebulin and desmin started at 3 h post mortem in breast muscle. Intact titin and nebulin disappeared within 1 d. Intact desmin and vinculin were not detectable after 3 d post mortem. In leg muscle, the degradation process of all the examined proteins evolved much more slowly than in breast chicken muscles. 4. The changes observed in shear force, myofibrillar fragmentation and cooking loss were related to changes in cytoskeletal proteins and used to identify marker proteins or degradation products for the purpose of monitoring the development of meat ageing. The ageing process was faster in breast muscle than in leg muscle. 5. Significant correlations were found between degradation processes of titin, nebulin, and desmin and shear force, as well as myofibril fragmentation index of breast and leg muscles.

"Daisy-like" crystals: A rare and unknown type of urinary crystal.

BACKGROUND: Crystals are well known structures of urinary sediment, most of which are identified by the combined knowledge of crystal morphology, birefringence features at polarized light, and urine pH. In this paper, we report on a cohort of subjects whose urine contained a very rare type of crystal, which we first described in 2004 and which, based on its peculiar morphology, we define as "daisy-like crystal" (DLcr). METHODS: Reports on DLcr were spontaneously sent to our laboratory over a 10.5-year period by different laboratory professionals and by one veterinary clinician who, in their everyday work, had come across DLcr. After the examination of DLcr images submitted, a number of other information were requested and partly obtained. RESULTS: DLcr were found in 9 human beings in 7 different laboratories, located in 4 countries (Italy, Belgium, Croatia, France). DLcr were found mostly in female (8/9), at all ages (3.5 to 93years), mostly in alkaline urine (pH6.0 to 7.5), at variable specific gravity values (1.010 to 1.030), either as isolated particles (2/8) or in association with other crystals (5/8) and/or leucocytes or bacteria (3/8). In addition, DLcr were found in the urine of a 1-year-old dog, examined in a veterinary clinic of Czech Republic. In 3 cases, DLcr were identified by manual microscopy, while in 7 cases by automated urine sediment analyzers. CONCLUSIONS: This paper confirms the possible presence in the urine of DLcr. However, further cases are needed to clarify their frequency, clinical meaning, and composition.

Selective targeting of liposomes to macrophages using a ligand with high affinity for the macrophage scavenger receptor class A.

Macrophages play an important role in inflammatory processes and are crucially involved in the onset and progression of atherosclerosis and tumorigenesis. Therefore, macrophages are regarded as an excellent target for therapeutic intervention. Since the scavenger receptor class A (SRA) is highly expressed on macrophages, we developed in the present study an SRA-specific particulate drug carrier by providing phosphatidylcholine liposomes with a targeting ligand for SRA. To enable firm association with liposomes, the high-affinity SRA ligand decadeoxyguanine was covalently attached via a linker to lithocholic oleate (LCO-dA(2)dG(10)). Incorporation of LCO-dA(10)dG(2) into liposomes resulted in an increased electronegative surface charge and a dramatically enhanced serum clearance (t(1/2) < 2 min versus > 5 h). The LCO-dA(2)dG(10)-induced liposome clearance was fully dependent on SRA, as the clearance could be efficiently inhibited by the SRA competitor polyinosinic acid. LCO-dA(2)dG(10) enhanced the affinity of liposomes for SRA in vivo selectively, since introduction of overall or clustered negative charges by other modifications (e.g. oxidation, inclusion of phosphatidylserine, or exposure of glutamic acid residues) did not affect their serum clearance substantially, albeit that these modifications resulted in an at least equally high negative surface charge. LCO-dA(2)dG(10) also increased the association of liposomes with RAW264.7 cells, resulting in an enhanced intracellular delivery and bioactivity of encapsulated dexamethasone-phosphate. Therefore, the SRA-specificity of LCO-dA(2)dG(10)-liposomes may be applied for the specific delivery of drugs to macrophages, which may be of therapeutic benefit in general inflammatory disorders, atherosclerosis, and tumorigenesis.

Baricitinib: JAK inhibition for rheumatoid arthritis.

Rheumatoid arthritis (RA), a chronic autoimmune inflammatory disease characterized by inflammation and joint destruction, is associated with pain, progressive disability, systemic comorbidities and early death. Conventional disease-modifying antirheumatic drugs (DMARDs) and biological DMARDs (bDMARDs) have been able to achieve remission or a very low disease activity status for RA. Nevertheless, since many patients do not reach a sufficient response with DMARDs or present with unacceptable side effects, new therapies are needed. Baricitinib (LY-3009104, INCB-028050), a new potent and selective tyrosine-protein kinase JAK1/JAK2 inhibitor, has shown clinical efficacy in patients with RA refractory to aggressive standard-of-care treatment (with both conventional DMARDs and bDMARDs) when administered orally at 4 mg q.d. in pivotal phase III clinical trials. In these studies, radiographic joint damage assessments showed significant improvements with baricitinib, and the drug was well tolerated with no unexpected safety findings. A phase III study aimed at assessing long-term (4 years) safety and efficacy of baricitinib is ongoing. Registration processes are ongoing in Europe, the U.S. and Japan.

Cardiac troponin-I on diagnosis predicts early death and refractoriness in acquired thrombotic thrombocytopenic purpura. Experience of the French Thrombotic Microangiopathies Reference Center.

BACKGROUND: Cardiac involvement is a major cause of mortality in patients with thrombotic thrombocytopenic purpura (TTP). However, diagnosis remains underestimated and delayed, owing to subclinical injuries. Cardiac troponin-I measurement (cTnI) on admission could improve the early diagnosis of cardiac involvement and have prognostic value. OBJECTIVES: To assess the predictive value of cTnI in patients with TTP for death or refractoriness. PATIENTS/METHODS: The study involved a prospective cohort of adult TTP patients with acquired severe ADAMTS-13 deficiency (< 10%) and included in the registry of the French Reference Center for Thrombotic Microangiopathies. Centralized cTnI measurements were performed on frozen serum on admission. RESULTS: Between January 2003 and December 2011, 133 patients with TTP (mean age, 48 ± 17 years) had available cTnI measurements on admission. Thirty-two patients (24%) had clinical and/or electrocardiogram features. Nineteen (14.3%) had cardiac symptoms, mainly congestive heart failure and myocardial infarction. Electrocardiogram changes, mainly repolarization disorders, were present in 13 cases. An increased cTnI level (> 0.1 µg L(-1) ) was present in 78 patients (59%), of whom 46 (59%) had no clinical cardiac involvement. The main outcomes were death (25%) and refractoriness (17%). Age (P = 0.02) and cTnI level (P = 0.002) showed the greatest impact on survival. A cTnI level of > 0.25 µg L(-1) was the only independent factor in predicting death (odds ratio [OR] 2.87; 95% confidence interval [CI] 1.13-7.22; P = 0.024) and/or refractoriness (OR 3.03; 95% CI 1.27-7.3; P = 0.01). CONCLUSIONS: A CTnI level of > 0.25 µg L(-1) at presentation in patients with TTP appears to be an independent factor associated with a three-fold increase in the risk of death or refractoriness. Therefore, cTnI level should be considered as a prognostic indicator in patients diagnosed with TTP.

The role of ketoconazole in the QTc interval prolonging effects of H1-antihistamines in a guinea-pig model of arrhythmogenicity.

We have carried out experiments to re-evaluate the influence of ketoconazole (400 mg kg-1,p.o.) on the effects of ebastine, terfenadine and loratadine on the QTc interval in conscious guinea-pigs. Following a previously described protocol of oral drug administration, but using telemetric recording of the ECG, we have found that the prolongation of the QTc interval attributed to ebastine and terfenadine is in fact entirely due to ketoconazole, and that neither terfenadine, ebastine nor loratadine produce any additional effects on subsequent administration.

Effects of H1 antihistamines on animal models of QTc prolongation.

OBJECTIVE: The clinical use of some nonsedating H1 antihistamines (histamine H1 receptor antagonists) has been associated with a rare but life-threatening type of arrhythmia, torsade de pointes, especially when these drugs are coadministered with cytochrome P450 (CYP) 3A4 enzyme inhibitors. On the basis of the latter observation and the fact that most of these H1 antihistamines undergo extensive first-pass metabolism to active metabolites apparently devoid of cardiovascular adverse effects, this arrhythmogenicity has been attributed to the parent drug. The objective of this study was to find an animal model with the ability to predict the proclivity of drugs to produce torsade de pointes. DESIGN: Two experimental approaches were used: (i) blockade of CYP3A4 metabolism by coadministration of ketoconazole to increase the plasma concentrations of the parent compound in the conscious guinea-pig, and (ii) administration of the compound directly into the coronary circulation of the anaesthetised dog in order to circumvent first-pass metabolism. RESULTS: The first approach demonstrated that terfenadine administered in the presence of ketoconazole prolongs the corrected QT (QTc) interval of the electrocardiogram, whereas ebastine does not. Similarly, when terfenadine was administered through the coronary circulation, a statistically significant increase in the QTc interval was also seen, whereas ebastine and carebastine were without effect. Thus, it is clear that ebastine was much better tolerated than terfenadine from a cardiovascular standpoint, since ebastine and its metabolite are devoid of effects on cardiac repolarisation, as measured by the QTc interval in these animal models.

Inhibition of lipopolysaccharide-induced bowel erythrocyte extravasation in rats, and of mesenteric hypoperfusion in dogs, by phosphodiesterase inhibitors.

Sepsis is intricately associated with mesenteric ischemia. The remote complications of mesenteric ischemia are essentially those of sepsis, whether as a cause or as a consequence. Experimental endotoxic shock induces bowel hypoperfusion, erythrocyte extravasation and intestinal necrosis. The effects of pentoxifylline, rolipram and denbufylline, three phosphodiesterase inhibitors, were studied on endotoxin-induced bowel erythrocyte extravasation and intestinal and renal hypoperfusion, in conscious rats and anaesthetized dogs, respectively. Two hours after lipopolysaccharide i.v. injection in rats, erythrocyte extravasation was evident throughout the intestinal musculature and mucosa, apparently without affecting lungs, heart, kidneys, liver or pancreas. Pretreatment with the non-selective phosphodiesterase inhibitor, pentoxifylline, or selective phosphodiesterase IV inhibitors such as denbufylline or rolipram reduced intestinal haemoconcentration. In the anaesthetized dog, pentoxifylline and denbufylline both inhibited the E. coli lipopolysaccharide-induced mesenteric blood flow fall, without affecting renal blood flow or cardiac index. In conclusion, phosphodiesterase inhibitors protected from intestinal damage and bowel hypoperfusion after lipopolysaccharide challenge. This action may thus play a role in the protective effects against endotoxin-induced lethal toxicity previously described for phosphodiesterase inhibitors.

Cardiovascular safety profile of almotriptan, a new indolic derivative for the treatment of migraine.

Almotriptan is a new 5-HT(1B/1D) receptor agonist effective for treating acute migraine attacks with or without aura. As 3-5% of patients treated with sumatriptan experience chest symptoms thought to be of cardiac origin, we investigated the cardiovascular safety profile of almotriptan in comparison with that of sumatriptan in six animal models. Almotriptan did not modify blood pressure or heart rate in conscious telemetered normotensive Wistar rats (p.o.), in anaesthetised beagle dogs (i.v.), or in conscious beagle dogs (i.v.), and only produced transient increases when administered (s.c.) to telemetered cynomolgus monkeys. Almotriptan did not consistently affect the duration of the electrocardiogram (ECG) intervals in anaesthetised beagle dogs even when the drug was administered into the coronary artery, nor was ECG morphology altered in telemetered cynomolgus monkeys. In contrast, sumatriptan i.v. consistently increased mean blood pressure and heart rate in conscious beagle dogs. Finally, almotriptan did not modify coronary blood flow at a dose of up to 0.3 mg/kg i.v. in conscious beagle dogs. Thus, almotriptan has a favourable cardiovascular safety profile.

Functional profile of almotriptan in animal models predictive of antimigraine activity.

Almotriptan is a new 5-HT(1B/1D) receptor agonist whose clinical efficacy for the treatment of migraine attacks has been demonstrated in Phase III clinical trials. We now compare the functional profile of almotriptan (assessed using animal models) with that of sumatriptan. Almotriptan selectively increased carotid vascular resistance in anaesthetised cats after intravenous or intraduodenal administration (ED(100)=11 microg/kg, i.v.; ED(50)=339 microg/kg, i. d.) and in anaesthetised beagle dogs following intravenous administration (ED(50)=116 microg/kg). A study in anaesthetised cats also demonstrated that almotriptan acts by selectively increasing the resistance of the carotid arteriovenous anastomoses without adversely affecting brain irrigation. In addition, almotriptan inhibited meningeal extravasation produced by electrical stimulation of the trigeminal ganglion in anaesthetised guinea pigs in the dose range of 0.3-3 mg/kg, i.v. In conclusion, almotriptan is both a selective constrictor affecting intracranial blood vessels and an inhibitor of neurogenically evoked plasma protein extravasation of the dura mater.

Pharmacological characterization of almotriptan: an indolic 5-HT receptor agonist for the treatment of migraine.

Almotriptan (3-[2-(dimethylamino)ethyl]-5-(pyrrolidin-1-ylsulfonylmethyl )-1H-indo le) has been studied in several models predictive of activity and selectivity at 5-HT receptors. Almotriptan showed low nanomolar affinity for the 5-HT(1B) and 5-HT(1D) receptors in several species, including the human, while affinity for 5-HT receptors other than 5-HT(1B/1D) was clearly less. Affinity for 5-HT(7) and 5-HT(1A) receptors was approximately 40 and 60 times lower than that for 5-HT(1B/1D) receptors, respectively. Almotriptan did not exhibit significant affinity for several non-5-HT receptors studied up to 100 microM. Almotriptan inhibited forskolin-stimulated cyclic AMP accumulation in HeLa cells transfected with 5-HT(1B) or 5-HT(1D) human receptors. In this model, almotriptan had the same efficacy as serotonin and an affinity in the low nanomolar range. It induced vasoconstriction in several vessels in which it was compared with sumatriptan. In isolated dog saphenous veins, almotriptan elicited concentration-dependent contractions with an EC(50) of 394 nM. In both these systems, almotriptan behaved as a full agonist. Infusion of almotriptan into the porcine meningeal vasculature induced vasoconstriction. In contrast, in the pig renal and rabbit mesenteric arteries, it had a very low maximal efficacy even at 100 microM, with similar results obtained in the rabbit renal artery. The results suggest that almotriptan is a potent and selective 5-HT(1B/1D) receptor agonist, with selectivity for the cranial vasculature as compared with peripheral vessels.

Pseudo-analbuminaemia due to the presence of a slow albumin variant moving into the alpha 1 zone.

A serum has been studied, in which direct observation of cellulose acetate electrophoresis may induce misdiagnosis of analbuminaemia. Immunogically, and by saline fractionation, the serum contained albumin, although in a decreased quantity. Gel filtration shows the majority of this albumin is to be in the 4.5 S20 peak and to have a low electrophoretic mobility, overlapping with that of alpha 1 globulins. Possible explanations of the presence of this slow-moving albumin are discussed.

DNA radiolysis in DNA-protein complexes: a stochastic simulation of attack by hydroxyl radicals.

PURPOSE: To propose an improved version of RADACK, a stochastic simulation of radiolytic attack on DNA, that takes into account the reactivity of each amino acid of a specifically bound protein with hydroxyl radicals. To apply it to the natural lactose operator-repressor complex taking advantage of recently reported structures. To compare the obtained probabilities of DNA strand break induction with those calculated with the previous versions and with an experimental pattern of strand break probabilities. MATERIALS AND METHODS: Models of complexes close to the natural ones, derived from crystallography- and NMR-based structures recently available in the PDB databank, were used. The specific chemical reactivity of each amino acid was introduced in the new version of RADACK (the reactivity model). The probabilities of strand break induction by the irradiation of the complex were calculated with this new version as well as with previous ones. RESULTS: The patterns of probabilities of strand break induction calculated with the improved version of RADACK were partially different from those obtained with previous versions. The patterns obtained for both, using putative models of natural complexes, were consistent with the experimental results, but some discrepancies were suggestive of slight differences between these structures and the real natural system. The crystallographic structure agreed best with the experimental results. CONCLUSIONS: A new version of RADACK was validated that took into account the reactivity of atoms in both DNA and protein. The putative modelled structures of a natural lactose operator-repressor complex were discussed.

Determination of a buspirone metabolite in plasma samples.

A high-performance liquid chromatographic (HPLC) method is described for the assay of the active metabolite [1-(2-pyrimidinyl)piperazinel of buspirone, an anxiolytic agent, in rat plasma. The method is based on the use of ion-pair HPLC coupled to a liquid-solid extraction scheme. Samples of rat plasma (2 ml) with internal standard (1-phenylpiperazine), adjusted to pH 10.5 with borate buffer, were loaded on to a preactivated C-18 cartridge. The metabolite and the internal standard were eluted with 5 ml of methanol and injected on to a reversed-phase 10-microm Spherisorb ODS-2 column. The column was eluted with a mobile phase of 0.005 M sodium lauryl sulphate in citrate buffer (pH 3.6)-acetonitrile (65:35, v/v) at 2 ml min(-1). Detection was carried out at 248 nm. The recovery of the metabolite was 55%. The method was applied to the determination of the metabolite in rat plasma after oral dosing (25 mg kg(-1)) of the parent compound.

Determination of flunarizine in plasma by a new high-performance liquid chromatography method. Application to a bioavailability study in the rat.

A reversed-phase high performance liquid chromatographic (HPLC) method is described for the study of the pharmacokinetics of flunarizine. The method involves selective liquid-solid extraction of flunarizine and meclizine (as internal standard) from samples of rat plasma. The optimization of the extraction and HPLC separation parameters are discussed. Recoveries were satisfactory and the relative standard deviation for replicate assays was below 10%. The sensitivity of the method would allow the detection of flunarizine in plasma at 13 ng ml(-1). Kinetic parameters for a bioequivalency study between flunarizine and a liquid formulation (oral drops) have been evaluated; the relative bioavailability was 88%.

Size distribution and sources of aerosol in Launceston, Australia, during winter 1997.

As part of a pilot study into the chemical and physical properties of Australian fine particles, a suite of aerosol samples was collected at Ti Tree Bend in Launceston, Tasmania, during June and July 1997. This period represents midwinter in the Southern Hemisphere, a period when aerosol sources in Launceston are dominated by smoke from domestic wood burning. This paper describes the results from this measurement campaign, with the aim of assessing the effect of wood smoke on the chemical and physical characteristics of ambient aerosol. A micro orifice uniform deposit impactor (MOUDI) was used to measure the size distributions of the aerosol from 0.05 to 20 microns aerodynamic diameter. Continuous measurements of fine particle mass were made using a PM2.5 tapered element oscillating microbalance (TEOM) and light scattering coefficients at 530 nm were measured with nephelometers. Mass size distributions tended to be bimodal, with the diameter of the dominant mode tending to smaller sizes with increases in total mass. Non-sea salt potassium and polycyclic aromatic hydrocarbons (PAHs) were used as chemical tracers for wood smoke. Wood smoke was found to increase absolute particle mass (enough to regularly exceed air quality standards), and to concentrate mass in a single mode below 1 micron aerodynamic diameter. The acid-base equilibrium of the aerosol was altered by the wood smoke source, with free acidity hydrogen ion, non-sea salt sulfate, and ammonium concentrations being higher and the concentration of all species, including nitrate (to differing extents), focused in the fine particle size ranges. The wood smoke source also heavily influenced the aerosol scattering efficiency, adding to a strong diurnal cycle in both mass concentration and light scattering.

[Study of lipids (cholesterol, triglycerides and phospholipids), plasma lipoproteins (HDL-cholesterol) and apoproteins (apo A and apo B in patients with acute myocardial infarction]. / Estudio de lípidos (colesterol, triglicéridos y fosfolípidos), lipoproteínas plasmáticas (HDL-colesterol) y apoproteínas (apo A y apo B) en pacientes con infarto agudo de miocardio.

Several lipid parameters were evaluated in 88 patients with acute myocardial infarction (AMI). A reduction in cholesterol and phospholipid level was observed, with minimal values between the days 10-20, followed by a subsequent increase. Triglycerides increased after 12-14 hours, with a maximal value after 20-30 days. Cholesterol bound to high-density lipoproteins (HDL-cholesterol) decreased after 24 hours, with a minimal value after 20-30 days. Apoprotein A decreased from the initial measurement until days 20-30. The initial lipid profile (indicating the risk of coronary artery disease) was constituted by high cholesterol (particularly in males below 60 years), high triglyceride and low HDL-cholesterol levels. The latter finding was more marked and characteristic in the overall group of evaluated patients. All values were compared with a control group of 24 healthy persons. Relevant data for prognosis included the following: males above 60 years had a greater reduction in cholesterol and apoprotein A during AMI; the males who died had lower apoprotein A levels than the survivors; females with higher haptoglobin levels also had higher apoprotein B levels. All these findings were statistically significant (p less than 0.01).

[Evanescent exostosis. A report on one case (author's transl)]. / Exostose évanescente. Un cas.

Almost complete recovery after seven years of an isolated large exostosis of the lower end of the femur, discovered in a young boy at the age of 6 years.

[Lipids and beta-lipoproteins in hepatic cirrhosis]. / Lípidos y betalipoproteínas en cirrosis hepática.

The authors carried out a study of serum lipids and beta-lipoproteins using chemical methods commonly available in clinical pathological laboratories. The study was based on 72 men and women who were chemically and clinically healthy and 41 other patients with biopsy confirmed hepatic cirrhosis, clinically decompensated. In cirrhotic patients there is a decrease in lipid and beta-lipoprotein values in serum. In plasma cirrhotics show a decrease in beta-lipoproteins. In addition the beta-lipoprotien molecule has a different make-up in cirrhotics which statistically is significantly altered from that seen in healthy patients. The graphic correlation between cholesterol, phospholipids and esterified fatty acids in serum as well as in beta-lipoproteins shows a different theoretical equation in the healty and cirrhotic patient. The beta-lipoproteins in cirrhotics are less atherogenic than the corresponding molecules in healthy individuals. The results here presented suggest that there is an important quantitative and qualitative alternation in lipid metabolism and in the biosynthesis of plasma lipoproteins in the uncontrolled cirrhotic.