Involvement of nitrogen oxide in the cerebral vasoconstriction during respiration with high pressure oxygen

High pressure oxygen evokes a cerebral vasoconstriction and diminishes cerebral blood flow with the aid of mechanisms which are not yet sufficiently studied. We were checking a hypothesis that the hyperbaric oxygen (HBO2) inactivates cerebral nitrogen oxide (NO), interrupts its basal relaxing effect, and evokes a vasoconstriction. In our experiments, HBO2 decreased cerebral blood flow depending on the pressure. Inhibiting the NO-synthase weakened basal vasorelaxation in breathing with atmosphere air and eliminated the vasoconstriction in exposure to the HBO2. Inactivation of O2 prevented the HBO2-induced vasoconstriction. The data obtained reveal that diminishing of cerebral blood flow in HBO is related to the NO inactivation and weakening of its basal vasorelaxing effect. Possible mechanisms of the NO inactivation may involve its reaction with oxygen and superoxide anion which lead to diminishing of the tissue NO concentration and weakening of its vasorelaxing effect.     

Contributions of nitric oxide synthase isoforms to pulmonary oxygen toxicity, local vs. mediated effects

Reactive species of oxygen and nitrogen have been collectively implicated in pulmonary oxygen toxicity, but the contributions of specific molecules are unknown. Therefore, we assessed the roles of several reactive species, particularly nitric oxide, in pulmonary injury by exposing wild-type mice and seven groups of genetically altered mice to >98% O2 at 1, 3, or 4 atmospheres absolute. Genetically altered animals included knockouts lacking either neuronal nitric oxide synthase (nNOS(-/-)), endothelial nitric oxide synthase (eNOS(-/-)), inducible nitric oxide synthase (iNOS(-/-)), extracellular superoxide dismutase (SOD3(-/-)), or glutathione peroxidase 1 (GPx1(-/-)), as well as two transgenic variants (S1179A and S1179D) having altered eNOS activities. We confirmed our earlier finding that normobaric hyperoxia (NBO2) and hyperbaric hyperoxia (HBO2) result in at least two distinct but overlapping patterns of pulmonary injury. Our new findings are that the role of nitric oxide in the pulmonary pathophysiology of hyperoxia depends both on the specific NOS isozyme that is its source and on the level of hyperoxia. Thus, iNOS predominates in the etiology of lung injury in NBO2, and SOD3 provides an important defense. But in HBO2, nNOS is a major contributor to pulmonary injury, whereas eNOS is protective. In addition, we demonstrated that nitric oxide derived from nNOS is involved in a neurogenic mechanism of HBO2-induced lung injury that is linked to central nervous system oxygen toxicity through adrenergic/cholinergic pathways.     

Nitric oxide synthase 3 deficiency limits adverse ventricular remodeling after pressure overload in insulin resistance

Insulin resistance (IR) and systemic hypertension are independently associated with heart failure. We reported previously that nitric oxide synthase 3 (NOS3) has a beneficial effect on left ventricular (LV) remodeling and function after pressure-overload in mice. The aim of our study was to investigate the interaction of IR and NOS3 in pressure-overload-induced LV remodeling and dysfunction. Wild-type (WT) and NOS3-deficient (NOS3(-/-)) mice were fed either a standard diet (SD) or a high-fat diet (HFD) to induce IR. After 9 days of diet, mice underwent transverse aortic constriction (TAC). LV structure and function were assessed serially using echocardiography. Cardiomyocytes were isolated, and levels of oxidative stress were evaluated using 2',7'-dichlorodihydrofluorescein diacetate. Cardiac mitochondria were isolated, and mitochondrial respiration and ATP production were measured. TAC induced LV remodeling and dysfunction in all mice. The TAC-induced decrease in LV function was greater in SD-fed NOS3(-/-) mice than in SD-fed WT mice. In contrast, HFD-fed NOS3(-/-) developed less LV remodeling and dysfunction and had better survival than did HFD-fed WT mice. Seven days after TAC, oxidative stress levels were lower in cardiomyocytes from HFD-fed NOS3(-/-) than in those from HFD-fed WT. N(ω)-nitro-L-arginine methyl ester and mitochondrial inhibitors (rotenone and 2-thenoyltrifluoroacetone) decreased oxidative stress levels in cardiomyocytes from HFD-fed WT mice. Mitochondrial respiration was altered in NOS3(-/-) mice but did not worsen after HFD and TAC. In contrast with its protective role in SD, NOS3 increases LV adverse remodeling after pressure overload in HFD-fed, insulin resistant mice. Interactions between NOS3 and mitochondria may be responsible for increased oxidative stress levels in HFD-fed WT mice hearts.     

Patterns of DNA Barcode Variation in Canadian Marine Molluscs

Background

Molluscs are the most diverse marine phylum and this high diversity has resulted in considerable taxonomic problems. Because the number of species in Canadian oceans remains uncertain, there is a need to incorporate molecular methods into species identifications. A 648 base pair segment of the cytochrome c oxidase subunit I gene has proven useful for the identification and discovery of species in many animal lineages. While the utility of DNA barcoding in molluscs has been demonstrated in other studies, this is the first effort to construct a DNA barcode registry for marine molluscs across such a large geographic area.

Methodology/Principal Findings

This study examines patterns of DNA barcode variation in 227 species of Canadian marine molluscs. Intraspecific sequence divergences ranged from 0–26.4% and a barcode gap existed for most taxa. Eleven cases of relatively deep (>2%) intraspecific divergence were detected, suggesting the possible presence of overlooked species. Structural variation was detected in COI with indels found in 37 species, mostly bivalves. Some indels were present in divergent lineages, primarily in the region of the first external loop, suggesting certain areas are hotspots for change. Lastly, mean GC content varied substantially among orders (24.5%–46.5%), and showed a significant positive correlation with nearest neighbour distances.

Conclusions/Significance

DNA barcoding is an effective tool for the identification of Canadian marine molluscs and for revealing possible cases of overlooked species. Some species with deep intraspecific divergence showed a biogeographic partition between lineages on the Atlantic, Arctic and Pacific coasts, suggesting the role of Pleistocene glaciations in the subdivision of their populations. Indels were prevalent in the barcode region of the COI gene in bivalves and gastropods. This study highlights the efficacy of DNA barcoding for providing insights into sequence variation across a broad taxonomic group on a large geographic scale.     

Expression of miRNAs in ovine fetal gonads: potential role in gonadal differentiation

Background  

Gonadal differentiation in the mammalian fetus involves a complex dose-dependent genetic network. Initiation and progression of fetal ovarian and testicular pathways are accompanied by dynamic expression patterns of thousands of genes. We postulate these expression patterns are regulated by small non-coding RNAs called microRNAs (miRNAs). The aim of this study was to identify the expression of miRNAs in mammalian fetal gonads using sheep as a model.     

Molecular evolution of mitochondrial 12S RNA and cytochrome b sequences in the pantherine lineage of Felidae

DNA sequence comparisons of two mitochondrial DNA genes were used to infer phylogenetic relationships among 17 Felidae species, notably 15 in the previously described pantherine lineage. The polymerase chain reaction (PCR) was used to generate sequences of 358 base pairs of the mitochondrial 12S RNA gene and 289 base pairs of the cytochrome b protein coding gene. DNA sequences were compared within and between 17 felid and five nonfelid carnivore species. Evolutionary trees were constructed using phenetic, cladistic, and maximum likelihood algorithms. The combined results suggested several phylogenetic relationships including (1) the recognition of a recently evolved monophyletic genus Panthera consisting of Panthera leo, P. pardus, P. onca, P. uncia, P. tigris, and Neofelis nebulosa; (2) the recent common ancestry of Acinonyx jubatus, the African cheetah, and Puma concolor, the American puma; and (3) two golden cat species, Profelis temmincki and Profelis aurata, are not sister species, and the latter is strongly associated with Caracal caracal. These data add to the growing database of vertebrate mtDNA sequences and, given the relatively recent divergence among the felids represented here (1-10 Myr), allow 12S and cytochrome b sequence evolution to be addressed over a time scale different from those addressed in most work on vertebrate mtDNA.      

Plutella australiana (Lepidoptera,Plutellidae), an overlooked diamondback moth revealed?by?DNA?barcodes

The genus Plutella was thought to be represented in Australia by a single introduced species, Plutella xylostella (Linnaeus), the diamondback moth. Its status as a major pest of cruciferous crops, and the difficulty in developing control strategies has motivated broad-ranging studies on its biology. Prior genetic work has generally supported the conclusion that populations of this migratory species are connected by substantial gene flow. However, the present study reveals the presence of two genetically divergent lineages of this taxonin Australia. One shows close genetic and morphological similarity with the nearly cosmopolitan Plutella xylostella. The second lineage possesses a similar external morphology, but marked sequence divergence in the barcode region of the cytochrome c oxidase I gene, coupled with clear differences in genitalia. As a consequence, members of this lineage are described as a new species, Plutella australiana Landry & Hebert, which is broadly distributed in the eastern half of Australia.     

Towards a comprehensive barcode library for arctic life - Ephemeroptera, Plecoptera, and Trichoptera of Churchill, Manitoba, Canada

Background

In order to understand the role of herbivores in trophic webs, it is essential to know what they feed on. Diet analysis is, however, a challenge in many small herbivores with a secretive life style. In this paper, we compare novel (high-throughput pyrosequencing) DNA barcoding technology for plant mixture with traditional microhistological method. We analysed stomach contents of two ecologically important subarctic vole species, Microtus oeconomus and Myodes rufocanus, with the two methods. DNA barcoding was conducted using the P6-loop of the chloroplast trnL (UAA) intron.

Results

Although the identified plant taxa in the diets matched relatively well between the two methods, DNA barcoding gave by far taxonomically more detailed results. Quantitative comparison of results was difficult, mainly due to low taxonomic resolution of the microhistological method, which also in part explained discrepancies between the methods. Other discrepancies were likely due to biases mostly in the microhistological analysis.

Conclusion

We conclude that DNA barcoding opens up for new possibilities in the study of plant-herbivore interactions, giving a detailed and relatively unbiased picture of food utilization of herbivores.     

Interaction of neuronal NOS and catecholamines in regulation of expression of proteins of apoptosis by vasopressinergic hypothalamic neurons

The work studied vasopressinergic neurons of hypothalamic supraoptic and paravenricular nuclei of the wild type mice and the neuronal nitric oxide synthase (nNOS) gene knockouted mice at a decrease of the brain catecholamine (CA) level caused by administration of the blocker of activity of tyrosine hydroxylase alpha-methyl-paratyrosine (alpha-MPT) and at the CA level decrease on the background of functional activity of the vasopressinergic neurons caused by dehydration of animals. There were analyzed changes in the number of neurons in both magnocellular hypothalamic nuclei expressing proapoptotic proteins caspase-8 and caspase-9, p53, and antiapoptotic protein Bcl-2. The disturbance of the CA-ergic innervation was shown to be a strong damaging factor leading to apoptosis of neurons regardless of the presence of nNOS in the cells. However, at disturbance of the CA-ergic innervation due to the 5-day mouse dehydration, no death of neurons by apoptosis was revealed. Thus, it is possible that functional activation prevents the hypothalamic vasopressinergic neurons from death at a decrease of the CA level in brain. The main difference of the nNOS gene knockouts is the absence of activation of the Bcl-2 expression under all used actions. This confirms our suggestion about interaction of CA and NO in triggering of expression of the antiapoptotic protein Bcl-2.     

Synthetic osteogenic extracellular matrix formed by coated silicon dioxide nanosprings

Background

Angiogenesis is widely investigated in conjunction with cancer development, in particular because of the possibility of early stage detection and of new therapeutic strategies. However, such studies are negatively affected by the limitations of imaging techniques in the detection of microscopic blood vessels (diameter 3-5 ??m) grown under angiogenic stress. We report that synchrotron-based X-ray imaging techniques with very high spatial resolution can overcome this obstacle, provided that suitable contrast agents are used.

Results

We tested different contrast agents based on gold nanoparticles (AuNPs) for the detection of cancer-related angiogenesis by synchrotron microradiology, microtomography and high resolution X-ray microscopy. Among them only bare-AuNPs in conjunction with heparin injection provided sufficient contrast to allow in vivo detection of small capillary species (the smallest measured lumen diameters were 3-5 ??m). The detected vessel density was 3-7 times higher than with other nanoparticles. We also found that bare-AuNPs with heparin allows detecting symptoms of local extravascular nanoparticle diffusion in tumor areas where capillary leakage appeared.

Conclusions

Although high-Z AuNPs are natural candidates as radiology contrast agents, their success is not guaranteed, in particular when targeting very small blood vessels in tumor-related angiography. We found that AuNPs injected with heparin produced the contrast level needed to reveal--for the first time by X-ray imaging--tumor microvessels with 3-5 ??m diameter as well as extravascular diffusion due to basal membrane defenestration. These results open the interesting possibility of functional imaging of the tumor microvasculature, of its development and organization, as well as of the effects of anti-angiogenic drugs.