Extraintestinal gamogony of Aggregata octopiana in the reared common octopus (Octopus vulgaris) (Cephalopoda: Octopodidae)

Aggregata octopiana (Apicomplexa, Aggregatidae) is the most prevalent coccidian in the wild common octopus (Octopus vulgaris), whose heteroxenous life cycle includes gamogony and sporogony undergoing in the octopus digestive tract. In the infected reared octopi, we observed an unusual extraintestinal distribution of the coccidian, with both gamogony and sporogony ongoing in dermal and gill tissue. Oocysts and macrogamonts were embedded in the dermal connective tissue of octopian arms, demarcated by a thin cyst wall or multilayered dark membrane. In gill connective and epithelial tissue all developmental stages were observed, eliciting hemocytic infiltration. Sometimes a complete substitution of the tissue by cysts and developmental stages occurred, resulting in necrosis of gill tissue.     

An asparaginyl endopeptidase mediates in vivo protein backbone cyclization

Proteases can catalyze both peptide bond cleavage and formation, yet the hydrolysis reaction dominates in nature. This presents an interesting challenge for the biosynthesis of backbone cyclized (circular) proteins, which are encoded as part of precursor proteins and require post-translational peptide bond formation to reach their mature form. The largest family of circular proteins are the plant-produced cyclotides; extremely stable proteins with applications as bioengineering scaffolds. Little is known about the mechanism by which they are cyclized in vivo but a highly conserved Asn (occasionally Asp) residue at the C terminus of the cyclotide domain suggests that an enzyme with specificity for Asn (asparaginyl endopeptidase; AEP) is involved in the process. Nicotiana benthamiana does not endogenously produce circular proteins but when cDNA encoding the precursor of the cyclotide kalata B1 was transiently expressed in the plants they produced the cyclotide, together with linear forms not commonly observed in cyclotide-containing plants. Observation of these species over time showed that in vivo asparaginyl bond hydrolysis is necessary for cyclization. When AEP activity was suppressed, either by decreasing AEP gene expression or using a specific inhibitor, the amount of cyclic cyclotide in the plants was reduced compared with controls and was accompanied by the accumulation of extended linear species. These results suggest that an AEP is responsible for catalyzing both peptide bond cleavage and ligation of cyclotides in a single processing event.     

Study of angiotensin-(1-7) vasoactive peptide and its beta-cyclodextrin inclusion complexes: complete sequence-specific NMR assignments and structural studies

We report the complete sequence-specific hydrogen NMR assignments of vasoactive peptide angiotensin-(1-7) (Ang-(1-7)). Assignments of the majority of the resonances were accomplished by COSY, TOCSY, and ROESY peak coordinates at 400MHz and 600MHz. Long-side-chain amino acid spin system identification was facilitated by long-range coherence transfer experiments (TOCSY). Problems with overlapped resonance signals were solved by analysis of heteronuclear 2D experiments (HSQC and HMBC). Nuclear Overhauser effects (NOE) results were used to probe peptide conformation. We show that the inclusion of the angiotensin-(1-7) tyrosine residue is favored in inclusion complexes with beta-cyclodextrin. QM/MM simulations at the DFTB/UFF level confirm the experimental NMR findings and provide detailed structural information on these compounds in aqueous solution.     

SIRT1 deacetylase protects against neurodegeneration in models for Alzheimer's disease and amyotrophic lateral sclerosis

A progressive loss of neurons with age underlies a variety of debilitating neurological disorders, including Alzheimer's disease (AD) and amyotrophic lateral sclerosis (ALS), yet few effective treatments are currently available. The SIR2 gene promotes longevity in a variety of organisms and may underlie the health benefits of caloric restriction, a diet that delays aging and neurodegeneration in mammals. Here, we report that a human homologue of SIR2, SIRT1, is upregulated in mouse models for AD, ALS and in primary neurons challenged with neurotoxic insults. In cell-based models for AD/tauopathies and ALS, SIRT1 and resveratrol, a SIRT1-activating molecule, both promote neuronal survival. In the inducible p25 transgenic mouse, a model of AD and tauopathies, resveratrol reduced neurodegeneration in the hippocampus, prevented learning impairment, and decreased the acetylation of the known SIRT1 substrates PGC-1alpha and p53. Furthermore, injection of SIRT1 lentivirus in the hippocampus of p25 transgenic mice conferred significant protection against neurodegeneration. Thus, SIRT1 constitutes a unique molecular link between aging and human neurodegenerative disorders and provides a promising avenue for therapeutic intervention.     

Abnormal systolic blood pressure during treadmill test and brachial artery flow-mediated vasodilatation impairment

The aim of the study was to assess the relationship between systolic blood pressure during maximal treadmill test (SBP9mtt)) and flow-mediated vasodilation (FMD). Abnormal rise of SBP(mtt) is the phenomenon more frequent in hypertensive persons but it could be found in normotensive subjects too. 199 subjects referred to treadmill test were enrolled in the study. Four groups were formed: hypertensives with abnormal SBP(mtt) (group A), hypertensives with normal SBP(mtt) (group B), normotensives with abnormal SBP(mtt) (group C) and normotensives with normal SBP(mtt) (group D). Rise of SBP(mtt) above 200 mmHg was considered abnormal reaction. Simple linear regression analysis showed significant inverse relationship between SBP(mtt) and FMD (F = 20.2036, p < 0.001, R2 = 0.0956). Mean FMD index was worst in hypertensive subjects with abnormal SBP(mtt) (group A), followed by normotensives with abnormal SBP(mtt) (group C), hypertensives with normal SBP(mtt) (group B) and the best was in normotensives with normal SBP(mtt) (3.56 +/- 5.17, 4.19 +/- 5.14, 6.81 +/- 8.43 and 10.92 +/- 7.48%, respectively). In multivariate regression analysis FMD showed significant association with abnormal SBP(mtt) (p < 0.001) along with brachial artery diameter (p < 0.001), male gender (p < 0.001), but not with hypertension (p = 0.073), BMI (p = 0.137) and total cholesterol (p = 0.23) (coefficients: -0.26, -0.40, -0.27, -0.13, -0.11 and -0.07, respectively). There was a significant inverse relationship between SBP(mtt) and FMD. An impairment of FMD exists in normotensive subjects with abnormal SBP(mtt). In hypertensives with abnormal SBP(mtt) an additional impairment of FMD exists when compared to hypertensives with normal SBP(mtt). Abnormal SBP(mtt) should be taken into account in global cardiovascular risk assessment.     

Virtual bronchoscopy and 3D spiral CT reconstructions in the management of patient with bronchial cancer--our experience with Syngo 3D postprocessing software

Multislice helical CT generated virtual bronchoscopy (VB) represents one of the most recent developments in three-dimensional computer aided visualisation techniques. VB allows non-invasive and relatively accurate 3D evaluation of tracheobronchal tree. We performed virtual bronchoscopy and in-space 3D volume analysis on CT-data set acquired from sixty-four-year old male with bronchial cancer in order to demonstrate advantages and disadvantages of these methods in diagnostics and preoperative management of metastatic bronchial cancer. Siemens Somatom Emotion 16 helical CT scanner was used for data acquisition. Data post-processing was done with 3D Syngo 2006G software package from Siemens medical systems. CT scanning of the thorax was performed in heavy smoker with an expansive T4N1M1 malignant process in a superior lobe of the right lung accompanied with large metastatic lesion attached on the right lateral chest wall. Metastatic lesions were also found in vertebral column. In-space 3D analysis followed with virtual bronchoscopy had revealed obstruction of apical branch of superior lobe segmental bronchus. External compression done by tumor to the superior segmental and right main bronchus was found. We concluded that multi-slice CT in connection with VB became a possible non-invasive alternative to bronchoscopy, if tissue samples are not required.     

ANG II type 1 receptor antagonist irbesartan inhibits coronary angiogenesis stimulated by chronic intermittent hypoxia in neonatal rats

Chronic hypoxia has been shown to stimulate myocardial microvascular growth and improve cardiac ischemic tolerance in young and adult rats. The aim of this study was to determine whether the ANG II type 1 receptor (AT(1)) pathway was involved in these processes. Newborn Wistar rats, exposed to chronic intermittent hypoxia (8 h/day) for 10 days, were simultaneously treated with AT(1) receptor blocker irbesartan and compared with untreated animals. The major finding is that chronic hypoxia increased the capillary supply of myocardial tissue, which was even more pronounced in hypertrophied right ventricle, whereas increased arteriolar supply was found only in the left ventricle. This angiogenic response was completely prevented by irbesartan. Moreover, chronic hypoxia improved the postischemic recovery of cardiac contractile function during reperfusion, and this protective effect was also completely abolished by irbesartan. Chronic hypoxia increased the myocardial density of AT(1) but not of ANG II type 2 receptor subtypes, whereas the effect of irbesartan was not significant. The expression of caveolin-1alpha markedly increased in response to chronic hypoxia, and irbesartan prevented this effect. Neither hypoxia nor irbesartan treatment altered the expression of nitric oxide synthase 3, heat shock protein 90, and VEGF. It is concluded that the AT(1) receptor pathway plays an important role in coronary angiogenesis and improved cardiac ischemic tolerance induced in neonatal rats by chronic hypoxia.     

Geldanamycin promotes nuclear localisation and clearance of PHOX2B misfolded proteins containing polyalanine expansions

Polyalanine expansions in the PHOX2B gene have been detected in the vast majority of patients affected with congenital central hypoventilation syndrome, a neurocristopathy characterized by absence of adequate control of breathing, especially during sleep, with decreased sensitivity to hypoxia and hypercapnia. The correlation between length of the alanine expanded tracts and severity of congenital central hypoventilation syndrome respiratory phenotype has been confirmed by length-dependent cytoplasmic PHOX2B retention with formation of aggregates. To deepen into the molecular mechanisms mediating the effects of PHOX2B polyalanine expansions, we have set up experiments aimed at assessing the fate of cells characterized by PHOX2B polyalanine aggregates. In particular, we have observed that activation of the heat shock response by the drug geldanamycin is efficient both in preventing formation and in inducing clearance of PHOX2B pre-formed polyalanine aggregates in COS-7 cells expressing PHOX2B-GFP fused proteins, and ultimately also in rescuing the PHOX2B ability to transactivate the Dopamine-beta-Hydroxilase promoter. In addition, we have demonstrated elimination of PHOX2B mutant proteins by the proteasome and autophagy, two cellular mechanisms already been involved in the clearance of proteins containing expanded polyglutamine and polyalanine tracts. Moreover, our data suggest that geldanamycin effects on PHOX2B aggregates may be also mediated by the proteasome pathway. Finally, analysis of cellular toxicity due to polyalanine aggregates has confirmed the occurrence of cell apoptosis consequent to expression of PHOX2B carrying the longest expanded alanine tract and shown that geldanamycin can delay cell progression toward the most advanced apoptotic stages.