Infliximab partially impairs the anti-Mycobacterium tuberculosis immune responses of severe psoriasis patients with positive tuberculin skin-test.

BACKGROUND: Infliximab and etarnecept are now widely used for treating severe psoriasis. However, these drugs, especially infliximab, increased the risk of tuberculosis reactivation. Surprisingly, epidemiological data suggest that the tuberculosis rate in patients taking infliximab in São Paulo State, Brazil, is similar to that of some developed, non-endemic countries. OBJECTIVE: The aim of this study was to better understand the effect of infliximab on Mycobacterium tuberculosis (Mtb) immune responses of psoriasis patients in an endemic setting (Brazil). METHODS: We evaluated the tuberculosis-specific immune responses of severe psoriasis patients and healthy individuals, both tuberculin skin test (TST) positive, in the presence/absence of infliximab. Patients had untreated severe psoriasis, no co-morbidities affecting the immune responses and a TST >10 mm. Healthy TST(+) (>10 mm) individuals were evaluated in parallel. PBMC cultures from both groups were stimulated with different Mycobacterium tuberculosis (Mtb) antigens (ESAT-6, 85B and Mtb lysate) and phytohemagglutinin, with or without infliximab (5 µg/mL). Parameters evaluated were TNF-α, IFN-γ and IL-10 secretion by ELISA, overnight IFN-γ ELISpot and lymphocyte proliferative response (LPR). RESULTS: Infliximab almost abolished TNF-α detection in PBMC supernatants of both groups. It also significantly reduced the LPR to phytohemagglutinin and the Mtb antigens as well as the IFN-γ levels secreted into day 5 supernatants in both groups. There was no concomitant exaggerated IL-10 secretion that could account for the decreases in these responses. ELISpot showed that, contrasting with the central-memory responses above, infliximab did not affect effector-memory INF-γ-releasing T-cell numbers. CONCLUSIONS: Infliximab affected some, but not all aspects of the in vitro antituberculosis immune responses tested. The preserved effector-memory responses, putatively related to exposure to environmental mycobacteria, may help to explain the lower than expected susceptibility to tuberculosis reactivation in our setting.

Glycaemic variability using continuous glucose monitoring and endothelial function in the metabolic syndrome and in Type 2 diabetes.

AIMS: Subjects who are at increased risk of developing diabetes may have increased glycaemic variability associated with endothelial dysfunction and possibly subclinical atherosclerosis, which may lead to increased cardiovascular risk observed at the time of diabetes diagnosis. To investigate this hypothesis, we measured endothelial function, carotid intima-media thickness and glycaemic variability using 48-h continuous subcutaneous glucose monitoring in 3 groups of overweight or obese subjects--those without the metabolic syndrome, and those with the metabolic syndrome with or without newly diagnosed Type 2 diabetes. METHODS: Consecutive subjects, aged 30-65 years with a body mass index >or= 25 kg/m(2) were recruited. Patients were classified as with or without the metabolic syndrome,or as metabolic syndrome with newly diagnosed Type 2 DM. Glycaemic variability was calculated in terms of the coefficient of variation. Endothelial function was measured using brachial artery flow-mediated dilation. RESULTS: We identified 75 subjects. Mean flow mediated dilation decreased (P < 0.001) and carotid intima-media thickness increased (P < 0.05) across groups. Flow mediated dilation predictors included mean 48-h continuous subcutaneous glucose monitoring values (beta = -0.022; P < 0.005) and the coefficient of variation (beta = -0.10; P = 0.01). Carotid intima-media thickness predictors included age (beta = 0.009; P < 0.001) and flow mediated dilation (beta = -0.014; P = 0.076). Patients re-stratified according to cut-offs for mean 48-h glycaemia and variability demonstrated that subjects with high mean glycaemia but low coefficient of variability had similar flow mediated dilation and carotid intima-media thickness to subjects with low mean glycaemia but high coefficient of variation. CONCLUSIONS: This study suggests that glycaemic variability influences endothelial function even in non-diabetic subjects. Such variability may explain the increased cardiovascular risk observed in patients prior to developing overt Type 2 diabetes.

Fluorescent in situ hybridization reveals multiple expression domains for SpBrn1/2/4 and identifies a unique ectodermal cell type that co-expresses the ParaHox gene SpLox.

Here we report on the expression of two transcription factors previously reported to be involved in endodermal patterning, SpLox and SpBrn1/2/4. We describe three distinct domains of expression of the pou-domain gene SpBrn1/2/4. Endodermal expression of this gene is restricted to the foregut. SpBrn1/2/4 is also expressed in two distinct ectodermal domains: throughout the stomodeal ectoderm, and within cells scattered throughout the ciliated band. The ParaHox gene SpLox is demonstrated to also be expressed in ectodermal bilateral cell pairs in prism and early pluteus stage larvae. Double fluorescent in situ hybridization reveals that these cell pairs co-express both SpLox and SpBrn1/2/4, thus marking a novel cell type within the ciliary band of the sea urchin larvae.

Disseminated blue naevus and malignant blue naevus associated with excessive aromatase syndrome.

We report a case of a 17-year-old boy who had a giant congenital blue naevus with multiple satellite pigmented lesions. Later the patient developed melanoma arising in the pre-existing lesion. He also had gynaecomastia and was diagnosed as having aromatase excess syndrome. To our knowledge, the association of these two rare conditions has not been previously reported. Further studies should be performed to investigate this unusual combination, which may have a genetic, endocrine or local cutaneous link leading to its occurrence.

Bivalent IAP antagonists, but not monovalent IAP antagonists, inhibit TNF-mediated NF-κB signaling by degrading TRAF2-associated cIAP1 in cancer cells.

The inhibitor of apoptosis (IAP) proteins have pivotal roles in cell proliferation and differentiation, and antagonizing IAPs in certain cancer cell lines results in induction of cell death. A variety of IAP antagonist compounds targeting the baculovirus IAP protein repeat 3 (BIR3) domain of cIAP1have advanced into clinical trials. Here we sought to compare and contrast the biochemical activities of selected monovalent and bivalent IAP antagonists with the intent of identifying functional differences between these two classes of IAP antagonist drug candidates. The anti-cellular IAP1 (cIAP1) and pro-apoptotic activities of monovalent IAP antagonists were increased by using a single covalent bond to combine the monovalent moieties at the P4 position. In addition, regardless of drug concentration, treatment with monovalent compounds resulted in consistently higher levels of residual cIAP1 compared with that seen following bivalent compound treatment. We found that the remaining residual cIAP1 following monovalent compound treatment was predominantly tumor necrosis factor (TNF) receptor-associated factor 2 (TRAF2)-associated cIAP1. As a consequence, bivalent compounds were more effective at inhibiting TNF-induced activation of p65/NF-κB compared with monovalent compounds. Moreover, extension of the linker chain at the P4 position of bivalent compounds resulted in a decreased ability to degrade TRAF2-associated cIAP1 in a manner similar to monovalent compounds. This result implied that specific bivalent IAP antagonists but not monovalent compounds were capable of inducing formation of a cIAP1 E3 ubiquitin ligase complex with the capacity to effectively degrade TRAF2-associated cIAP1. These results further suggested that only certain bivalent IAP antagonists are preferred for the targeting of TNF-dependent signaling for the treatment of cancer or infectious diseases.

Expression of a hyperthermophilic aspartate aminotransferase in Escherichia coli.

The gene for an archaebacterial hyperthermophilic enzyme, aspartate aminotransferase from Sulfolobus solfataricus (AspATSs), was expressed in Escherichia coli and the enzyme purified to homogeneity. A suitable expression vector and host strain were selected and culture conditions were optimized so that 6-7 mg of pure enzyme per litre of culture were obtained repeatedly. The recombinant enzyme and the authentic AspATSs are indistinguishable: in fact, they have the same molecular weight, estimated by means of SDS-PAGE and gel filtration, the same Km values for 2-oxo-glutarate and cysteine sulphinate and the same UV-visible spectra. Moreover, recombinant AspATSs is thermophilic and thermostable just as the enzyme extracted from Sulfolobus solfataricus. The protocol described may be used to produce thermostable arachaebacterial enzymes in mesophilic hosts.

The active site of Sulfolobus solfataricus aspartate aminotransferase.

Aspartate aminotransferase from the archaebacterium Sulfolobus solfataricus binds pyridoxal 5' phosphate, via an aldimine bond, with Lys-241. This residue has been identified by reducing the enzyme in the pyridoxal form with sodium cyanoboro[3H]hydride and sequencing the specifically labeled peptic peptides. The amino acid sequence centered around the coenzyme binding site is highly conserved between thermophilic aspartate aminotransferases and differs from that found in mesophilic isoenzymes. An alignment of aspartate aminotransferase from Sulfolobus solfataricus with mesophilic isoenzymes, attempted in spite of the low degree of similarity, was confirmed by the correspondence between pyridoxal 5' phosphate binding residues. Using this alignment it was possible to insert the archaebacterial aspartate aminotransferase into a subclass, subclass I, of pyridoxal 5' phosphate binding enzymes comprising mesophilic aspartate aminotransferases, tyrosine aminotransferases and histidinol phosphate aminotransferases. These enzymes share 12 invariant amino acids most of which interact with the coenzyme or with the substrates. Some enzymes of subclass I and in particular aspartate aminotransferase from Sulfolobus solfataricus, lack a positively charged residue, corresponding to Arg-292, which in pig cytosolic aspartate aminotransferase interacts with the distal carboxylate of the substrates (and determines the specificity towards dicarboxylic acids). It was confirmed that aspartate aminotransferase from Sulfolobus solfataricus does not possess any arginine residue exposed to chemical modifications responsible for the binding of omega-carboxylate of the substrates. Furthermore, it has been found that aspartate aminotransferase from Sulfolobus solfataricus is fairly active when alanine is used as substrate and that this activity is not affected by the presence of formate. The KM value of the thermophilic aspartate aminotransferase towards alanine is at least one order of magnitude lower than that of the mesophilic analogue enzymes.

Opsin evolution in the Ambulacraria.

Opsins--G-protein coupled receptors involved in photoreception--have been extensively studied in the animal kingdom. The present work provides new insights into opsin-based photoreception and photoreceptor cell evolution with a first analysis of opsin sequence data for a major deuterostome clade, the Ambulacraria. Systematic data analysis, including for the first time hemichordate opsin sequences and an expanded echinoderm dataset, led to a robust opsin phylogeny for this cornerstone superphylum. Multiple genomic and transcriptomic resources were surveyed to cover each class of Hemichordata and Echinodermata. In total, 119 ambulacrarian opsin sequences were found, 22 new sequences in hemichordates and 97 in echinoderms (including 67 new sequences). We framed the ambulacrarian opsin repertoire within eumetazoan diversity by including selected reference opsins from non-ambulacrarians. Our findings corroborate the presence of all major ancestral bilaterian opsin groups in Ambulacraria. Furthermore, we identified two opsin groups specific to echinoderms. In conclusion, a molecular phylogenetic framework for investigating light-perception and photobiological behaviors in marine deuterostomes has been obtained.

Results of 1-year growth hormone (GH)-releasing hormone-(1-44) treatment on growth, somatomedin-C, and 24-hour GH secretion in six children with partial GH deficiency.

Six children with short stature and partial GH deficiency in response to two pharmacological tests received GHRH for 12 months (10 micrograms/kg X day, sc) each evening. Twenty-four-hour GH secretion was studied before and after 3 and 12 months of treatment, and GHRH tests (2 micrograms/kg, iv) were done before and after 6 months of treatment. Plasma somatomedin-C was measured before and after 1.5, 3, 6, 9, and 12 months of treatment. Statural growth was measured at 3-month intervals. Mean growth velocity increased from 4.2 to 8.6 cm/yr, with a good result in five children and no response in the other. The growth response was substantial during the first 3 months. It was maintained during the following 6 months, and then decreased during the last 3 months. The peak plasma GH level in response to GHRH increased from 34.5 +/- 14.2 (+/-SD) ng/mL before treatment to 47.8 +/- 3.4 ng/mL after 6 months of treatment. Twenty-four-hour GH secretion increased in all parameters at 3 months (maximum peak, area under the curve, integrated concentration, and number of peaks) and at 12 months (with the exception of the maximum peak). Nycthemeral secretory profiles became normal, with reappearance of secretory pulses in two children, slight increases in three children, and no change in one child. Plasma somatomedin-C levels rose from 0.8 +/- 0.3 U/mL before treatment to 2.0 +/- 1.0 U/mL at 3 months, then decreased to 1.3 +/- 0.6 U/mL at 12 months. These results indicate that GHRH administered by sc injection for a 1-yr period stimulated growth and GH secretion. However, a decrease in activity was noted during the last 3 months of treatment. Tests for anti-GHRH antibodies were positive in the only child who did not respond to treatment.

Selective inhibition of sucrose and ethanol intake by SR 141716, an antagonist of central cannabinoid (CB1) receptors.

SR 141716, a selective central CB1 cannabinoid receptor antagonist, markedly and selectively reduces sucrose feeding and drinking as well as neuropeptide Y-induced sucrose drinking in rats. SR 141716 also decreases ethanol consumption in C57BL/6 mice. In contrast, blockade of CB1 receptors only marginally affects regular chow intake or water drinking. The active doses of SR 141716 (0.3-3 mg/kg) are in the range known to antagonize the characteristic effects induced by cannabinoid receptor agonists. These results suggest for the first time that endogenous cannabinoid systems may modulate the appetitive value of sucrose and ethanol, perhaps by affecting the activity of brain reward systems.

Neurobehavioural effects of SR 27897, a selective cholecystokinin type A (CCK-A) receptor antagonist.

The activity of SR 27897, a potent and selective CCK-A vs CCK-B receptor antagonist (Ki = 0.2 nM on guinea-pig pancreas vs 2000 nM on rat brain) was studied on behavioural, electrophysiological and biochemical effects induced by peripheral or central injection of CCK-8S. For comparative purposes, devazepide, a reference CCK-A receptor antagonist, was investigated in these same models. CCK-induced hypophagia and CCK-induced hypolocomotion in rats, two behavioural changes associated with the stimulation of peripheral CCK-A receptors, were dose-dependently antagonized by SR 27897 (ED50 = 0.003 and 0.002 mg/kg i.p., respectively) and devazepide (ED50 = 0.02 and 0.1 mg/kg i.p., respectively). CCK-induced decrease of cerebellar cGMP levels in mice was also reduced by SR 27897 (ED50 = 0.013 mg/kg) and by devazepide (0.084 mg/kg). The CCK-induced turning behaviour after intrastriatal injection in mice, and the potentiation of the rate suppressant activity of apomorphine on rat DA neurons, were blocked by higher doses of SR 27897 and devazepide, consistent with the probable central origin of these effects. The respective ED50s were 0.2 mg/kg i.p. for SR 27897 and 4.9 mg/kg i.p. for devazepide in the former model, while the respective minimal effective doses were 1.25 and 5 mg/kg i.p. in the latter test. In most tests the i.p./p.o. ratio for SR 27897 was near unity, suggesting a high oral bioavailability of the compound. Taken together, these findings support the notion that SR 27897 behaves as a potent CCK-A antagonist able to cross the blood brain barrier.

Effect of SR 59026A, a new 5-HT(1A) receptor agonist, on sexual activity in male rats.

The effect of SR 59026A, a new selective 5-HT(1A) receptor agonist, was evaluated on sexual behaviour of male rats in different experimental conditions. SR 59026A (1-10mg/kg p.o.) stimulated the copulatory behaviour of sexually experienced rats, as evidenced by a decrease in the number of pre-ejaculatory mounts and intromissions and a shortening of the ejaculation latency. SR 59026A also facilitated the sexual behaviour of naive male rats characterized by a low level of sexual performance: over the same dose range, the percentage of naive males that copulated was significantly increased and the ejaculation latency reduced. In experiments designed to evaluate the onset of sexual satiation, SR 59026A (1 and 3mg/kg) increased significantly the number of ejaculations and delayed the time of sexual satiation. Finally, in agreement with studies on other 5-HT(1A) receptor agonists, SR 59026A did not modify the occurrence of spontaneous erections in isolated male rats. Therefore, the present study shows that SR 59026A improves the sexual performance of male rats in a number of different experimental models, and the compound may prove to be of interest for the treatment of certain states of human male sexual dysfunction.

Properties of 3H-MPTP binding sites in human blood platelets.

Our study demonstrates that 3H-1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (3H-MPTP) specifically binds to platelet membrane sites in humans. This specific, high affinity and saturable binding has properties similar to those of 3H-MPTP binding to rat and monkey brain, with a higher affinity. Deprenyl, a specific inhibitor of MAO type B enzyme, was the most potent drug in displacing 3H-MPTP from platelet binding sites. Platelets are considered a good model for central aminergic neurons and are very rich with MAO enzymatic activity, exclusively of type B. Our findings support previous evidence indicating a correspondence between 3H-MPTP binding sites and MAO-B enzyme. Moreover the presence of 3H-MPTP binding sites on human platelets suggests the use of this peripheral tissue as a simple model to study at least partially the mechanisms of neurotoxic action of MPTP.

Kappa binding sites in guinea-pig brain membranes: evidence for a dynorphin-resistant subtype.

Using [3H]ethylketocyclazocine after suppression of mu, delta and sigma binding by 100 nM DAGO + 100 nM DADLE + 10 microM phencyclidine we showed the existence of a subclass of kappa sites insensitive to dynorphins 1-9 and 1-17, and alpha and beta neoendorphins, but sensitive to the non-peptidic opiates. The dynorphin-sensitive site probably corresponds to the "standard" kappa site while the importance of the dynorphin-resistant site remains to be established.

Effects of diazepam on extinction induced aggression in pigs.

Pigs were trained to press a panel with their snout to get food in an operant conditioning chamber. Aggressive behaviour which developed between two pigs submitted together to extinction was used as a baseline to study the effects of 1-2 mg/kg diazepam. When access to the response panel and feeding area was permitted, diazepam enhanced resistance to extinction and did not modify aggression. When access to the response panel and feeding area was not permitted, diazepam increased the severity of aggression observed between the animals. In both instances, plasma corticosteroid levels were depressed in diazepam-treated pigs. These results suggest that benzodiazepines do not act on frustation or aggressiveness per se, but rather strengthen the prevailing behavioural attitudes in the animals' repertoire at the time of test.

Encephalocraniocutaneous lipomatosis: a new case report and review of the literature.

Encephalocraniocutaneous lipomatosis is a rare neurocutaneous syndrome characterized by lipomatous hamartomas ranging in size from a few millimeters to several centimeters and affecting the head. Ocular anomalies and a variable degree of mental retardation with or without convulsions are usually observed. This disorder should be distinguished from other mosaic neurocutaneous phenotypes such as Proteus syndrome, oculocerebrocutaneous syndrome, and nevus sebaceous syndrome. We report the clinicopathologic findings of a 4-year-old Brazilian girl affected by this syndrome and review the literature. To our best knowledge, this is the first documented case of encephalocraniocutaneous lipomatosis occurring sporadically in South America.

[The presence of nucleotide sequences of the hepatitis B virus in the serum of HBsAg-negative blood donors in Sardinia]. / Presenza di sequenze nucleotidiche del virus dell'epatite B nel siero dei donatori di sangue HBsAg negativi in Sardegna.

The presence of HBV DNA was assessed in the serum samples from 878 HBsAg negative Sardinian blood donors. They were composed of 481 (55%) donors selected because of abnormal serum alanine aminotransferase (ALT) levels during routine testing of their blood donation, and of 397 donors (45%) selected on the basis of normal serum ALT activities. HBV DNA sequences were detected in 37 (7.7%) out of 481 subjects with abnormal ALT and in 2 (0.5%) out of 397 subjects with normal ALT. Anti-HBc was detected in 199 (41%) of the 481 subjects with abnormal ALT and in 81 (20%) out of 397 subjects with normal ALT. Among the 39 subjects positive for serum HBV DNA, 12 (31%) were positive for anti-HBc, while 27 (69%) were negative for all serological HBV markers. These data show in Sardinia, where HBV infection is endemic, there is a high frequency of HBsAg negative HBV DNA positive individuals in whom multiplication of HBV may occur without conventional serological HBV markers, suggesting the possible existence of HBV-like viruses which may be responsible for some of the presumed non-A non-B hepatitis.

[Xenograft of pancreatic islets: preliminary results of a new immunoisolation method]. / Xenotrapianto di insule pancreatiche: risultati preliminari di una nuova metodica d'immunoisolamento.

Results of clinical islet transplantation remain disappointing despite the advances in islet technology. Availability of human organs and control of rejection by adequate immunosuppressive therapy remain the unsolved problems. Transplantation of xenogeneic tissue enclosed in immuno-separating membranes without immunosuppressive drugs may be a solution. In the present study porcine pancreatic islets were isolated by semiautomated method and purified utilizing discontinuous Euroficoll gradients on IBM 2991 cell separator. The porcine pancreatic islets were encapsulated with a new one-step method utilizing a home-made droplet generator. Each microcapsule contained one or two islets and microcapsule diameter was approximately that of the islets. This condition allows an optimal diffusion of insulin, glucose, nutrients and oxygen. Consequently, perifusion experiments with encapsulated porcine islets revealed a typical biphasic pattern of insulin release as it was seen in unencapsulated controls. Human erythrocytes were encapsulated and incubated with serum containing hemolysins and complement. These experiments showed that the encapsulated erythrocytes were protected against the hemolytic activity of Ig G and complement fractions. In conclusion, this encapsulation procedure allows the production of a very thin barium alginate membrane around the islets with very little increase of the total volume of transplanted tissue.

Prognostic role of aldosterone in patients with acute coronary syndrome: short and medium term follow-up.

AIMS: Different studies have shown a correlation between aldosterone, atherosclerosis and ischemia in the past decade. Evidence exists for the relationship between high levels of aldosterone and augmented risk of cardiovascular diseases, such as hypertension, cardiac failure, coronary artery disease and stroke. The objective of this study was to determine the prognostic role of aldosterone in patients with myocardial infarction. METHODS: The study population included 96 consecutive patients admitted to our department for ST-elevated and non-ST-elevated myocardial infarction from June 2009 to March 2012. Plasma aldosterone levels were measured at admission to hospital in all patients. A 2-year prospective follow-up was performed, and fatal events and non-fatal events, such as reinfarction, congestive heart failure and arrhythmias, were recorded. RESULTS: Aldosterone levels at admission were associated with incidence of congestive heart failure (P = 0.02), ventricular arrhythmias (P = 0.01) and all complications (P = 0.003) after 1-month follow-up. Moreover, high aldosterone levels gave important information in the medium term (24 ±â€Š6 months). Specifically, aldosterone was a predictive variable of reinfarction (P < 0.0001), congestive heart failure (P < 0.0001) and adverse events (P = 0.0002). The logistic regression analysis confirmed these results and showed that aldosterone may be predictive of adverse events at medium-term follow-up (odds ratio 1.1, 95% confidence interval 1.03-1.15, P = 0.02). CONCLUSION: These data show a strong and significant correlation between plasma aldosterone levels at admission for myocardial infarction and fatal and nonfatal adverse events. Aldosterone appears to be a main marker of adverse clinical outcome, in accordance with the literature. These data suggest the need to identify whether antialdosteronic drug treatment, applied acutely in patients with aldosterone elevation, can influence favorably the prognosis of patients with myocardial infarction.