48 research outputs found
TAIL1: an isthmin-like gene, containing type 1 thrombospondin-repeat and AMOP domain, mapped to ARVD1 critical region.
No full textIsthmins represent a novel family of vertebrate secreted proteins containing one copy of the thrombospondin type 1 repeat (TSR), which in mammals is shared by several proteins with diverse biological functions, including cell adhesion, angiogenesis, and patterning of developing nervous system. We have determined the genomic organization of human TAIL1 (thrombospondin and AMOP containing isthmin-like 1), a novel isthmin-like gene encoding a protein that contains a TSR and a C-terminal AMOP domain (adhesion-associated domain in MUC4 and other proteins), characteristic of extracellular proteins involved in adhesion processes. TAIL1 gene encompasses more than 24.4 kb. Analysis of the DNA sequence surrounding the putative transcriptional start region revealed a TATA-less promoter located in a CpG island. Several consensus binding sites for the transcription factors Sp1 and MZF-1 were identified in this promoter region. In humans, TAIL1 gene is located on chromosome 14q24.3 within ARVD1 (arrhythmogenic right ventricular dysplasia/cardiomyopathy, type 1) critical region; preliminary evidence suggests that it is expressed in several tissues, showing multiple alternative splicing
Serum paraoxonase 1 (PON1) activity in bottlenose dolphins (Tursiops truncates): influence of different substrates and genetic polymorphisms
No full textBackground: Paraoxonase-1 (PON1) is an HDL-bound esterase and lipolactonase involved in organophosphate detoxification, protection against oxidative damage. PON1 acts as negative acute phase reactant (APR). Based on the different PON1 functions, several substrates have been identified to evaluate PON1 activities, which are influenced by single-nucleotide polymorphisms (SNPs) such as Leu55Met and Gln192Arg. No data on PON1 activity and SNPs are available for dolphins.
Objective: to evaluate the serum PON1 activity using different substrates and SNPs in bottlenose dolphins.
Methods: Serum samples from under human-care bottlenose dolphins were used: in 42 healthy animals paraoxonase activity was evaluated using paraoxon as substrate. Arylesterase and lactonase activity were evaluated in samples from 6 healthy and 6 diseased animals using 4-nitrophenyl acetate (4-nPA), phenyl-acetate, and dihydrocoumarin. In 15 animals, PON1 gene was screened for Leu55Met and Gln192Arg SNPs.
Results: paraoxonase activity showed mean values of 6.6±4.5 U/mL (median: 7.2 U/mL). Arylesterase activity using 4-nPA was 186.2±20.4 U/L (median: 187.2 U/L) in healthy dolphins and 177.5±14.3 U/L (median: 177.5 U/L) in diseased dolphins, while using phenyl-acetate, PON1 activity had high variability with no reaction or negative results. The lactonase activity was not detectable in any dolphins. All the animals showed homozygosis for Leu55 and Arg192.
Conclusions: paraoxonase activity in bottlenose dolphins is lower compared to domestic animals, thus its use as a negative APR is hampered. Arylesterase activity was higher, but without significant differences between healthy and diseased animals. The SNPs seems not to be associated with an elevated paroxonase activity, as reported for humans
Genomic and structural investigation on Dolphin Morbillivirus (DMV) in Mediterranean fin whales (Balaenoptera physalus)
No full textDolphin Morbillivirus (DMV) has been deemed as one of the most relevant threats for fin whales (Balaenoptera physalus) being responsible for a mortality outbreak in the Mediterranean Sea in the last years. Knowledge of the complete viral genome is essential to understand any structural changes that could modify virus pathogenesis and viral tissue tropism.
We report the complete DMV sequence of N, P/V/C, M, F and H genes identified from a fin whale and the comparison of primary to quaternary structure of proteins between this fin whale strain and some of those isolated during the 1990-‘92 and the 2006-‘08 epidemics. Some relevant substitutions were detected, particularly Asn52Ser located on F protein and Ile21Thr on N protein. Comparing mutations found in the fin whale DMV with those occurring in viral strains of other cetacean species, some of them were proven to be the result of diversifying selection, thus allowing to speculate on their role in host adaptation and on the way they could affect the interaction between the viral attachment and fusion with the target host cells
Molecular analysis of dolphin morbillivirus: A new sensitive detection method based on nested RT-PCR
No full textCetacean Morbillivirus (CeMV) has been identified as the most pathogenic virus for cetaceans. Over the past three decades, this RNA virus has caused several outbreaks of lethal disease in odontocetes and mysticetes worldwide. Isolation and identification of CeMV RNA is very challenging in whales because of the poor preservation status frequently shown by tissues from stranded animals. Nested reverse transcription polymerase chain reaction (nested RT-PCR) is used instead of conventional RT-PCR when it is necessary to increase the sensitivity and the specificity of the reaction. This study describes a new nested RT-PCR technique useful to amplify small amounts of the cDNA copy of Cetacean morbillivirus (CeMV) when it is present in scant quantity in whales' biological specimens. This technique was used to analyze different tissues (lung, brain, spleen and other lymphoid tissues) from one under human care seal and seven cetaceans stranded along the Italian coastline between October 2011 and September 2015. A well-characterized, 200 base pair (bp) fragment of the dolphin Morbillivirus (DMV) haemagglutinin (H) gene, obtained by nested RT-PCR, was sequenced and used to confirm DMV positivity in all the eight marine mammals under study. In conclusion, this nested RT-PCR protocol can represent a sensitive detection method to identify CeMV-positive, poorly preserved tissue samples. Furthermore, this is also a rather inexpensive molecular technique, relatively easy to apply
Involvement of the plasma membrane Ca2+-ATPase in the short-term response of Arabidopsis thaliana cultured cells to oligogalacturonides
No full textTreatment of Arabidopsis thaliana cells with oligoga-lacturonides (OG) initiates a transient production of reactive oxygen species (IROS), the concentration of which in the medium peaks after about 20 min of treatment. The analysis of OG effects on Ca2+ fluxes shows that OG influence both Ca2+ influx and Ca2+ efflux (measured as Ca-45(2+) fluxes) in a complex way. During the first 10-15 min, OG stimulate Ca2+ influx and decrease its efflux, while at successive times of treatment, OG cause an increase of Ca2+ efflux and a slight decrease of its influx. Treatment with sub-muM concentrations of eosin yellow (EY), which selectively inhibits the Ca2+-ATPase of plasma membrane (PM), completely prevents the OG-induced increase in Ca2+ efflux. EY also suppresses the transient feature of OG-induced ROS accumulation, keeping the level of ROS in the medium high. The biochemical analysis of PM purified from OG-treated cells indicates that treatment with OG for 15 to 45 min induces a significant decrease in Ca2+-ATPase activation by exogenous calmodulin (CaM), and markedly increases the amount of CaM associated with the PM. During the same time span, OG do not influence the expression of At-ACA8, the main isoform of PM Ca2+-ATPase in suspension-cultured A. thaliana cells, and of CaM genes. Overall, the reported results demonstrate that the PM Ca2+-ATPase is involved in the response of plant cells to OG and is essential in regulation of the oxidative burst
Is beta-catenin signalling involved in the molecular pathogenesis of arrhythmogenic right ventricular cardiomyopathy? Study on DSG2 transgenic mice
No full textPurpose. Arrhythmogenic Right Ventricular Cardiomyopathy (ARVC) is emerging as a relatively common cause of sudden death in teenagers and athletes. ARVC is considered a disease of cell adhesion because mutations in desmosomal genes have been involved in the pathogenesis of ARVC in a significant proportion of patients. The aim of the present study is to elucidate the molecular pathogenesis of ARVC by generating DSG2 transgenic animal models.
Methods. Human full−length wild−type and mutant cDNA sequences for DSG2 were cloned into a vector containing alpha−MyHC promoter. Transgenic lines showing a comparable expression level of transgene have been generated for G100R, N266S, K294E and Q558X mutations and for wild-type DSG2. Phenotypic characterisation was performed by electrophysiological and histological analyses. In order to investigate on the molecular pathogenesis, electron miscroscopic, immunohystochemical and Western blot examination was performed as well.
Results. Histological examination of 6-month-old mouse hearts evidentiated cardiomyocytes loss and fibro-fatty tissue replacement, resembling the phenotype of ARVC patients and confirming the pathogenic effects of DSG2 mutations in an animal model. Immunofluorescent staining showed correct localization of DSG2 G100R, N266S, and K294E into the desmosomal complexes, while in mice carrying the DSG2-Q558X mutation the protein was retained in the cytoplasm. Electron microscopic analyses showed a reduced number of desmosomes at the intercalated discs, myofibrillar degeneration and damaged mitochondria. Beta-catenin overexpression has been detected by Western blotting analysis and immunofluorescence in hearts of transgenic mice carrying the mutated proteins. DSG2 transgenic mice have been inbreeded with beta-catenin-activated transgene driving expression of nuclear beta-galactosidase reporter (BAT-gal) transgenic mice, in order to confirm beta-catenin overexpression. Analyses of BAT-gal expression in embrionic and neonatal transgenic hearts revealed a higher beta-catenin transcriptional activity compared to wild-type hearts.
Conclusions. DSG2 transgenic mice represents an invaluable model to investigate molecular pathogenesis of ARVC. Further studies will be needed to assess if beta-catenin overexpression could lead to beta-catenin signalling defects
Cardio-circulatory assistance with an artificial, peripheral, external ventricle: hemodynamics
No full textThe results of an experimental study on cardio circulatory assistance on 20 dogs with the use of a novel artificial ventricle applied across either the iliac or femoral artery are reported. The artificial ventricle implements a presystolic and systolic decrement, and a post systolic increment of the endoaortic pressure. The apparatus is designed to permit the direct control of pressure and the shifting of hematic volumes in constant relation with the volume ejected by the left ventricle
Genotype-phenotype correlation in arrhythmogenic right-ventricular cardiomyopathy linked to desmoplakin mutation (ARVD8)
No full textIF: 6.2
Dolphin Morbillivirus, a major threat for Mediterranean fin whales (Balaenoptera physalus)
No full textBetween 2011 and 2013, Dolphin Morbillivirus (DMV) was molecularly identified in four Mediterranean fin whales (Balaenoptera physalus), with the responsible strain showing a high nucleoprotein, phosphoprotein and hemoagglutinin gene sequence homology with the virus causing the 2006-‘07 Mediterranean epidemic. In conclusion, DMV represents a serious threat for fin whales
Detection of hereditary bisalbuminemia in bottlenose dolphins (Tursiops truncatus, Montagu 1821): comparison between capillary zone and agarose gel electrophoresis
No full textHereditary bisalbuminemia is a relatively rare anomaly characterized by the occurrence of two albumin fractions on serum protein separation by electrophoresis. In human medicine, it is usually revealed by chance, is not been clearly associated with a specific disease and the causative genetic alteration is a point mutation of human serum albumin gene inherited in an autosomal codominant pattern. This type of alteration is well recognizable by capillary zone electrophoresis (CZE), whilst agarose gel electrophoresis (AGE) not always produces a clear separation of albumin fractions. The aims of this study is to report the presence of this abnormality in two separate groups of related bottlenose dolphins and to compare the results obtained with capillary zone and agarose gel electrophoresis
