Low frequency of AIP mutations in patients with young-onset sporadic pituitary macroadenomas.

PURPOSE: Mutations in the aryl hydrocarbon receptor interacting protein (AIP) gene cause familial isolated pituitary adenomas (FIPA). AIP mutations have also been found in patients with apparently sporadic pituitary adenomas, particularly in young patients with large adenomas. The aim of this study was to determine the frequency of AIP germline mutations in patients with young-onset sporadic pituitary macroadenomas. METHODS: The AIP gene was sequenced in 218 Portuguese patients with sporadic pituitary macroadenomas diagnosed before the age of 40 years. RESULTS: Heterozygous rare sequence variants in AIP were identified in 18 (8.3%) patients. However, only four (1.8%) patients had pathogenic or likely pathogenic variants. These consisted of two already known mutations (p.Arg81* and p.Leu115Trpfs*41) and two novel mutations (p.Glu246*, p.Ser53Thrfs*36). All four patients had GH-secreting adenomas diagnosed between the ages of 14 and 25 years. The frequency of AIP pathogenic or likely pathogenic variants in patients under the age of 30 and 18 years was 3.4% and 5.0%, respectively. CONCLUSION: The frequency of AIP mutations in this cohort was lower than in other studies. Previous reports may have overestimated the contribution of AIP mutations due to the inclusion of genetic variants of uncertain significance. The identification of novel AIP mutations expands the known spectrum of genetic causes of pituitary adenomas and may help understand the role of AIP mutations in the molecular mechanisms underlying pituitary tumorigenesis.

Space colonization by branching trachea explains the morphospace of a simple respiratory organ.

Branching morphogenesis helps increase the efficiency of gas and liquid transport in many animal organs. Studies in several model organisms have highlighted the molecular and cellular complexity behind branching morphogenesis. To understand this complexity, computational models have been developed with the goal of identifying the "major rules" that globally explain the branching patterns. These models also guide further experimental exploration of the biological processes that execute and maintain these rules. In this paper we introduce the tracheal gills of mayfly (Ephemeroptera) larvae as a model system to study the generation of branched respiratory patterns. First, we describe the gills of the mayfly Cloeon dipterum, and quantitatively characterize the geometry of its branching trachea. We next extend this characterization to those of related species to generate the morphospace of branching patterns. Then, we show how an algorithm based on the "space colonization" concept (SCA) can generate this branching morphospace via growth towards a hypothetical attractor molecule (M). SCA differs from other branch-generating algorithms in that the geometry generated depends to a great extent on its perception of the "external" space available for branching, uses few rules and, importantly, can be easily translated into a realistic "biological patterning algorithm". We identified a gene in the C. dipterum genome (Cd-bnl) that is orthologous to the fibroblast growth factor branchless (bnl), which stimulates growth and branching of embryonic trachea in Drosophila. In C. dipterum, this gene is expressed in the gill margins and areas of finer tracheolar branching from thicker trachea. Thus, Cd-bnl may perform the function of M in our model. Finally, we discuss this general mechanism in the context of other branching pattern-generating algorithms.

Zika in Rio de Janeiro: Assessment of basic reproduction number and comparison with dengue outbreaks.

Zika virus infection was declared a public health emergency of international concern in February 2016 in response to the outbreak in Brazil and its suspected link with congenital anomalies. In this study, we use notification data and disease natural history parameters to estimate the basic reproduction number (R 0) of Zika in Rio de Janeiro, Brazil. We also obtain estimates of R 0 of dengue from time series of dengue cases in the outbreaks registered in 2002 and 2012 in the city, when DENV-3 and DENV-4 serotypes, respectively, had just emerged. Our estimates of the basic reproduction number for Zika in Rio de Janeiro based on surveillance notifications (R 0 = 2·33, 95% CI: 1·97-2·97) were higher than those obtained for dengue in the city (year 2002: R 0 = 1·70 [1·50-2·02]; year 2012: R 0 = 1·25 [1·18-1·36]). Given the role of Aedes aegypti as vector of both the Zika and dengue viruses, we also derive R 0 of Zika as a function of both dengue reproduction number and entomological and epidemiological parameters for dengue and Zika. Using the dengue outbreaks from previous years allowed us to estimate the potential R 0 of Zika. Our estimates were closely in agreement with our first Zika's R 0 estimation from notification data. Hence, these results validate deriving the potential risk of Zika transmission in areas with recurring dengue outbreaks. Whether transmission routes other than vector-based can sustain a Zika epidemic still deserves attention, but our results suggest that the Zika outbreak in Rio de Janeiro emerged due to population susceptibility and ubiquitous presence of Ae. aegypti.

Expanding the genetic spectrum of ANOS1 mutations in patients with congenital hypogonadotropic hypogonadism.

STUDY QUESTION: What is the prevalence and functional consequence of ANOS1 (KAL1) mutations in a group of men with congenital hypogonadotropic hypogonadism (CHH)? SUMMARY ANSWER: Three of forty-two (7.1%) patients presented ANOS1 mutations, including a novel splice site mutation leading to exon skipping and a novel contiguous gene deletion associated with ichthyosis. WHAT IS KNOWN ALREADY: CHH is characterized by lack of pubertal development and infertility, due to deficient production, secretion or action of GnRH, and can be associated with anosmia/hyposmia (Kallmann syndrome, KS) or with a normal sense of smell (normosmic CHH). Mutations in the anosmin-1 (ANOS1) gene are responsible for the X-linked recessive form of KS. STUDY DESIGN, SIZE, DURATION: This cross-sectional study included 42 unrelated men with CHH (20 with KS and 22 with normosmic CHH). PARTICIPANTS/MATERIALS, SETTING, METHODS: Patients were screened for mutations in the ANOS1 gene by DNA sequencing. Identified mutations were further investigated by RT-PCR analysis and multiplex ligation-dependent probe amplification (MLPA) analysis. MAIN RESULTS AND THE ROLE OF CHANCE: Hemizygous mutations were identified in three (7.1%) KS cases: a novel splice acceptor site mutation (c.542-1G>C), leading to skipping of exon 5 in the ANOS1 transcript in a patient with self-reported normosmia (but hyposmic upon testing); a recurrent nonsense mutation (c.571C>T, p.Arg191*); and a novel 4.8 Mb deletion involving ANOS1 and eight other genes (VCX3B, VCX2, PNPLA4, VCX, STS, HDHD1, VCX3A and NLGN4X) in KS associated with ichthyosis. LIMITATIONS, REASONS FOR CAUTION: Objective olfactory testing was not performed in all cases of self-reported normosmia and this may have underestimated the olfactory deficits. WIDER IMPLICATIONS OF THE FINDINGS: This study further expands the spectrum of known genetic defects associated with CHH and suggests that patients with self-reported normal olfactory function should not be excluded from ANOS1 genetic testing. STUDY FUNDING/COMPETING INTEREST(S): This study was funded by the Portuguese Foundation for Science and Technology. The authors have no conflicts of interest. TRIAL REGISTRATION NUMBER: N/A.

Effects of periodic perturbations on the oscillatory behavior in the NO+H2 reaction on Pt(100).

The mean field model proposed by Makeev and Nieuwenhuys [J. Chem. Phys. 108, 3740 (1998)] simulates the oscillatory behavior experimentally observed in the NO+H2 reaction on the surface Pt(100). This model reproduces quite well the kinetic oscillations and the transition to chaos via the Feigenbaum route, that is to say, through bifurcations involving period doubling. From this model, we analyze the response of the natural oscillations of period-1 (P1, one maximum) to periodic perturbations superposed to the partial pressure of one of the reactants. The perturbed model reproduces the periodic states found in the autonomous model, the route to chaos through bifurcations with period doubling, and the appearance of chaos via the route of intermittency, which shows alternation of periodic oscillations with intervals of disordered oscillations in the same time evolution. Experimentally it has been observed that the reaction shows a great sensitivity to reactant partial pressures and temperature. In experimental conditions slightly different to those considered in Makeev and Nieuwenhuys (MN) model, oscillations with period-3 (P3, three maxima) have been observed. At T=457 K and certain pressures, these P3 oscillations do not appear in MN model, although they appear at T=456 K . The same effect (P3 oscillations) is obtained at T=457 K in our perturbed model, due to the modulation of pH2. In a second step we show how the modulation of the perturbing frequency influences on the oscillations P1 of the perturbed system. The results show that the periodic behavior loses its regularity at low values of the normalized amplitude and of the modulated frequency of the perturbation. Other aspect observed in the perturbed model is that the amount of products varies in relation to nonperturbed model. When the oscillations are periodic or they follow the Feigenbaum route to chaos, the production average decreases or slightly increases, whereas it always increases if there are intermittencies, the most significant percentage increase being for NH3 (nearly 10%).

The CTLA4 +49 A/G polymorphism is not associated with susceptibility to type 1 diabetes mellitus in the Portuguese population.

CTLA4 genetic polymorphisms have been associated with type 1 diabetes. We genotyped 207 patients and 249 controls for the most frequently investigated polymorphism of the CTLA4 gene (+49A/G (rs231775)). No significant differences were observed, suggesting that this polymorphism is not strongly associated with type 1 diabetes in the Portuguese population.

Combined GSTM1 and GSTT1 null genotypes are associated with a lower risk of papillary thyroid cancer.

Individual susceptibility to cancer is influenced by polymorphisms of genes encoding drug-metabolizing enzymes such as the glutathione S-transferases (GST). The null polymorphisms of the GSTM1 and GSTT1 genes have been associated to a modified risk of several cancers but studies of thyroid cancer have produced conflicting results. The aim of this study was to investigate the relationship between these polymorphisms and the risk of papillary thyroid cancer (PTC). A total of 188 patients with PTC and 247 controls were genotyped using a PCR-based assay. Odds ratios (OR) and 95% confidence intervals (CI) for each homozygous null genotype were determined. The frequency of each of the GSTM1 and GSTT1 null genotypes did not differ significantly between patients and controls (OR=0.83, 95%CI: 0.56-1.21; p=0.328; and OR=0.66, 95%CI: 0.39-1.12; p=0.123, respectively), but the frequency of individuals that had the combined GSTM1 null/GSTT1 null genotypes was significantly lower in the patient group (OR=0.50, 95%CI: 0.26-0.97; p=0.040). The GSTM1 null genotype was associated with a lower risk of advanced cancer stages (III/IV) (OR=0.50, 95%CI: 0.26-0.96; p=0.036) and the GSTT1 null genotype was associated with a lower risk of the follicular variant of PTC (OR=0.31, 95%CI: 0.10-0.97; p=0.044). These results suggest that GSTM1 and GSTT1 null genotypes are weak, yet possible, modifiers of the risk of PTC. This protective effect may be due to a role of the GSTM1 and GSTT1 encoded enzymes in the metabolic activation of putative thyroid carcinogens or in other pathways involved in thyroid carcinogenesis.

Stabilization from chaotic to periodical states in a model of cellular automaton for oxidation of CO.

The chaotic behavior of a chemical reaction can be controlled perturbing periodically some of the parameters externally governing the reaction. Based on the resonance phenomenon, the method of external forcing can convert chaotic behavior into a periodical one through the application of a sinusoidal modulation. In this paper we analyze the effect of a periodical perturbation on room temperature in a model of cellular automaton that studies catalytic oxidation of CO. This model considers the reaction of carbon monoxide and oxygen adsorbed on a surface allowing the variation of the surface temperature and analyzing the time oscillations in the reaction. The results of simulations of this model show quasiperiodical and chaotic behaviors. Then the strategy of control through periodical forcing is able to remove the chaotic dynamics by means of the stabilization of periodical solutions, there being enough of a perturbing harmonic function with only one frequency to transform a chaotic state of the system into a periodical state with periodicity 1.

Periodical forcing for the control of chaos in a chemical reaction.

Control of the chaotic behavior of a chemical system can be achieved perturbing periodically some control parameters of the system. This procedure based on external forcing, which is based on the phenomenon of resonance, can change a chaotic behavior into a periodical one by means of the application of a sinusoidal perturbation. In this paper, the influence of a periodical modulation added to the parameter controlling the oxygen adsorption rate in a cellular automaton (CA) model studying CO oxidation is analyzed. This CA model considers the oxidation reaction of CO on a catalytic surface, taking into account the catalyst temperature variation in order to analyze the reaction time oscillatory behavior. Simulations of the CA model exhibit chaotic and quasiperiodical behaviors, and it can be shown that the periodical forcing strategy can suppress the chaotic dynamics by means of the stabilization of periodical solutions.

A cellular automaton for the modeling of oscillations in a surface reaction.

The reaction of CO and O over a catalytic surface is studied with a cellular automata (CA) model. We extend the CA model proposed by Mai and von Niessen [Phys. Rev. A 44 R6165 (1991)] taking into account the variation of the temperature of the catalyst with the aim of analyzing the existence of oscillations in this reaction. The rate constants for different processes which govern the reaction are chosen in the Arrhenius form. Quasiperiodic, aperiodic, O-poisoned, and CO-poisoned regimes are observed depending on the temperature relaxation parameter. The results from the CA model presented are in agreement with several oscillatory behaviors which the catalyzed oxidation of CO exhibits.

[Molecular characterisation of a kindred with MEN2A and clinical implications]. / Caracterização molecular de uma família com nem2a e suas implicações clínicas.

INTRODUCTION: MEN2A is an autossomal dominant cancer syndrome characterised by the presence of medullary thyroid cancer, pheochromocytoma and primary hyperparathyroidism. Germline mutations of the RET protooncogene constitute the molecular defect and can be identified in affected individuals. Genetic screening of family members at risk allows early diagnosis and preventive measures before the appearance of the disease. We present a family with several members affected with MEN2A, their molecular characterisation and the clinical implications of genetic testing. POPULATION AND METHODS: We studied 18 members distributed among three generations of a family of which four members were clinically affected with MEN2A and cutaneous lichen amyloidosis. RET gene mutations were screened for in affected individuals and their offspring by PCR-RFLP techniques. RESULTS: Genetic testing revealed a point mutation at codon 634 (TGC>TGG), in the heterozygous state, in all affected individuals. The same mutation was also found in a five years old asymptomatic child which after total thyroidectomy showed to have multifocal medullary thyroid carcinoma. DISCUSSION: Genetic screening is the most suitable method for pre-symptomatic diagnosis of MEN2A allowing an efficient and early identification of individuals who will later develop the disease. These can be monitored more closely and be submitted to a prophylactic thyroidectomy before the appearance of medullary thyroid carcinoma. The ideal moment for this intervention is still under discussion although the results of this study suggest that it should be undertaken before the age of five.

Genetic polymorphism of CYP2D6, GSTM1 and NAT2 and susceptibility to haematological neoplasias.

Xenobiotic-metabolizing enzymes constitute an important line of defence against a variety of carcinogens. Many are polymorphic, constituting the basis for the wide inter-individual variation in metabolic capacity and possibly a source of variation in the susceptibility to chemical-induced carcinogenesis. The aim of this study was to determine the existence of any association between the main genetic polymorphisms of cytochrome P450 2D6 (CYP2D6), glutathione S-transferase M1 (GSTM1) and N-acetyltransferase 2 (NAT2) and an altered risk for haematological neoplasias. A total of 160 patients and 128 controls were genotyped by means of PCR-RFLP-based assays. Mutated alleles comprising CYP2D6*4, GSTM1*0, NAT2*5A, *5B, *5C, *6 and *7 were analysed along with the wild-type alleles. The results showed a higher frequency of CYP2D6 extensive metabolizers carrying two functional alleles in the leukaemia group, when compared with controls (76.6 versus 57.0%, P = 0.008). No differences were found in the case of Hodgkin and non-Hodgkin lymphomas. Analysis of the GSTM1 and NAT2 polymorphisms failed to show an association with any of the neoplasias, although a near significant increase in fast acetylators was also found in the leukaemia group (50.0 versus 35.9%, P = 0.06). The results suggest an association of extensive metabolism with an increased risk for leukaemia, possibly by an increase in the metabolic activation of chemical carcinogens or linkage to another cancer-causing gene. Opposite findings presented in other studies may reflect geographical differences in the type of environmental carcinogens to which different populations are exposed.

In vitro and in vivo effects of nitric oxide synthase inhibitors and nitric oxide inactivators on the South American opossum ileocolonic junction.

The potential role of nitrergic nerves in the regulation of the South American (SA) opossum ileocolonic junction (ICJ) function was investigated. In vitro, the effects of nitric oxide (NO) synthase inhibitors and NO inactivators on the non-adrenergic non-cholinergic (NANC) nerve-mediated relaxations of the circular muscle of the SA opossum ICJ were determined by employing isolated strips. Electrical field stimulation (0.2-8.0 Hz) caused frequency-dependent NANC relaxations. Nicotine and ATP also induced concentration dependent NANC relaxations that were abolished by tetrodotoxin (TTX). The relaxation response induced by NANC nerve activation was reduced in a dose dependent manner by NO synthase inhibitors while vasoactive intestinal peptide (VIP) and sodium nitroprusside (SNP) induced relaxations were uninfluenced by these drugs. In vivo, the NO synthase inhibitor, L-NAME, administered into the local artery caused a raise in intraluminal pressure of the ICJ in anaesthetized SA opossums in a L-arginine-preventable manner. Hydroquinone and pyrogallol, while being able to reduce, in a superoxide dimutase (SOD) reversible manner, the relaxations induced by exogenous NO failed to affect the NANC nerve-induced relaxations. Finally, neurones and nerve fibres in the myenteric plexus as well as varicose nerve fibres on the circular smooth layer were positive for NADPH-diaphorase activity. These findings indicate that nitrergic nerves inhibit ICJ circular smooth muscle in vitro and in vivo but cast doubts on the neuromediator being the NO radical.

N-acetyltransferase genotypes in the Portuguese population.

Fast and slow acetylator phenotypes differ in their respective frequencies among different populations. A polymerase chain reaction-restriction fragment length polymorphism based genotyping assay was used to determine the frequency of the most important NAT2 polymorphisms in a group of 128 Portuguese individuals. The results showed that slow acetylators represented 64.1% of the group, and the frequencies of NAT2*4, *5A, *5B, *5C, *6 and *7 alleles were 0.211, 0.031, 0.379, 0.023, 0.328 and 0.027, respectively. These values are similar to those presented in other studies in Caucasians. The data obtained may be useful in epidemiological studies of the influence of acetylator polymorphisms on carcinogenesis or other environmental caused diseases.

Nonadrenergic-noncholinergic relaxations of isolated circular muscle from South American opossum esophagogastric junction: is nitric oxide the inhibitory mediator?

Nonadrenergic-noncholinergic (NANC) inhibitory nerves are responsible for most of the nerve induced relaxations of gastrointestinal muscle. It has recently been proposed that NANC nerves may release nitric oxide (NO) or a related compound derived from L-arginine. We have recently shown that the South American (SA) opossum is another suitable model to elucidate the mechanism involved in these NANC relaxations. In the present study the effect of NO synthase inhibitors as well as NO inactivators on the NANC-nerve induced relaxations of the circular muscle of the esophagogastric junction (EGJ) of the SA opossum was investigated. It was observed that the NO synthase inhibitors, L-NOARG and L-NAME, caused a concentration-dependent reduction of NANC-nerve induced relaxations which was reversed by L- but not D-arginine. The NO-donors sodium nitroprusside and hydroxilamine as well as NO caused concentration-dependent relaxations of the EGJ circular muscle. In the myenteric plexus of this region, NADPH-diaphorase positive neurons and nerve fibers were observed while in the circular muscle layer only numerous positive fibers were found. The NO inactivators, hydroquinone, pyrogallol and carboxy-PTIO, reduced NO-induced relaxations but failed to affect NANC nerve- and sodium nitroprusside-induced relaxations. Taken together, these findings indicate that NANC nerve induced relaxation of the SA opossum EGJ circular muscle is dependent on neural NO synthase activity and suggest that the neurotransmitter being released is a superoxide resistant molecule, which is unlikely to be the NO radical, or that the activity of NO synthase is required for the release of the actual neurotransmitter rather than for synthesizing the neuromediator.