A Cell Permeable Peptide Targeting the Intracellular Loop 2 of Endothelin B Receptor Reduces Pulmonary Hypertension in a Hypoxic Rat Model

Cell permeable peptides (CPP) aid cellular uptake of targeted cargo across the hydrophobic plasma membrane. CPP-mediated cargo delivery of receptor signaling motifs provides an opportunity to regulate specific receptor initiated signaling cascades. Both endothelin-1 receptors, ETA and ETB, have been targets of antagonist therapies for individuals with pulmonary arterial hypertension (PAH). These therapies have had success but have been accompanied by adverse reactions. Also, unlike the CPP which target specific signaling cascades, the antagonists target the entire function of the receptor. Using the CPP strategy of biased antagonism of the ETB receptor’s intracellular loop 2 (ICB2), we demonstrate blunting of hypoxic pulmonary hypertension (HPH) in the rat, including indices of pulmonary arterial pressure, right ventricular hypertrophy and pulmonary vascular remodeling. Further, ex vivo analysis of the pulmonary artery treated with the IC2B peptide upon injection manifests marked reductions in Akt and ERK activation. Both kinases have been intimately related to cell proliferation and vascular contraction, the hallmarks of PAH. These observations in sum illustrate an involvement of the ETB receptor in HPH and furthermore provide a basis for a novel, CPP-based, strategy in the treatment of PAH, ultimately able to target not only ET-1, but also other factors involved in the development of PAH

The Mitochondrial Genome of Baylisascaris procyonis

BACKGROUND: Baylisascaris procyonis (Nematoda: Ascaridida), an intestinal nematode of raccoons, is emerging as an important helminthic zoonosis due to serious or fatal larval migrans in animals and humans. Despite its significant veterinary and public health impact, the epidemiology, molecular ecology and population genetics of this parasite remain largely unexplored. Mitochondrial (mt) genomes can provide a foundation for investigations in these areas and assist in the diagnosis and control of B. procyonis. In this study, the first complete mt genome sequence of B. procyonis was determined using a polymerase chain reaction (PCR)-based primer-walking strategy. METHODOLOGY/PRINCIPAL FINDINGS: The circular mt genome (14781 bp) of B. procyonis contained 12 protein-coding, 22 transfer RNA and 2 ribosomal RNA genes congruent with other chromadorean nematodes. Interestingly, the B. procyonis mtDNA featured an extremely long AT-rich region (1375 bp) and a high number of intergenic spacers (17), making it unique compared with other secernentean nematodes characterized to date. Additionally, the entire genome displayed notable levels of AT skew and GC skew. Based on pairwise comparisons and sliding window analysis of mt genes among the available 11 Ascaridida mtDNAs, new primer pairs were designed to amplify specific short fragments of the genes cytb (548 bp fragment) and rrnL (200 bp fragment) in the B. procyonis mtDNA, and tested as possible alternatives to existing mt molecular beacons for Ascaridida. Finally, phylogenetic analysis of mtDNAs provided novel estimates of the interrelationships of Baylisasaris and Ascaridida. CONCLUSIONS/SIGNIFICANCE: The complete mt genome sequence of B. procyonis sequenced here should contribute to molecular diagnostic methods, epidemiological investigations and ecological studies of B. procyonis and other related ascaridoids. The information will be important in refining the phylogenetic relationships within the order Ascaridida and enriching the resource of markers for systematic, population genetic and evolutionary biological studies of parasitic nematodes of socio-economic importance

Cerebral microbleeds and stroke risk after ischaemic stroke or transient ischaemic attack: a pooled analysis of individual patient data from cohort studies

BACKGROUND: Cerebral microbleeds are a neuroimaging biomarker of stroke risk. A crucial clinical question is whether cerebral microbleeds indicate patients with recent ischaemic stroke or transient ischaemic attack in whom the rate of future intracranial haemorrhage is likely to exceed that of recurrent ischaemic stroke when treated with antithrombotic drugs. We therefore aimed to establish whether a large burden of cerebral microbleeds or particular anatomical patterns of cerebral microbleeds can identify ischaemic stroke or transient ischaemic attack patients at higher absolute risk of intracranial haemorrhage than ischaemic stroke. METHODS: We did a pooled analysis of individual patient data from cohort studies in adults with recent ischaemic stroke or transient ischaemic attack. Cohorts were eligible for inclusion if they prospectively recruited adult participants with ischaemic stroke or transient ischaemic attack; included at least 50 participants; collected data on stroke events over at least 3 months follow-up; used an appropriate MRI sequence that is sensitive to magnetic susceptibility; and documented the number and anatomical distribution of cerebral microbleeds reliably using consensus criteria and validated scales. Our prespecified primary outcomes were a composite of any symptomatic intracranial haemorrhage or ischaemic stroke, symptomatic intracranial haemorrhage, and symptomatic ischaemic stroke. We registered this study with the PROSPERO international prospective register of systematic reviews, number CRD42016036602. FINDINGS: Between Jan 1, 1996, and Dec 1, 2018, we identified 344 studies. After exclusions for ineligibility or declined requests for inclusion, 20 322 patients from 38 cohorts (over 35 225 patient-years of follow-up; median 1·34 years [IQR 0·19-2·44]) were included in our analyses. The adjusted hazard ratio [aHR] comparing patients with cerebral microbleeds to those without was 1·35 (95% CI 1·20-1·50) for the composite outcome of intracranial haemorrhage and ischaemic stroke; 2·45 (1·82-3·29) for intracranial haemorrhage and 1·23 (1·08-1·40) for ischaemic stroke. The aHR increased with increasing cerebral microbleed burden for intracranial haemorrhage but this effect was less marked for ischaemic stroke (for five or more cerebral microbleeds, aHR 4·55 [95% CI 3·08-6·72] for intracranial haemorrhage vs 1·47 [1·19-1·80] for ischaemic stroke; for ten or more cerebral microbleeds, aHR 5·52 [3·36-9·05] vs 1·43 [1·07-1·91]; and for ≥20 cerebral microbleeds, aHR 8·61 [4·69-15·81] vs 1·86 [1·23-1·82]). However, irrespective of cerebral microbleed anatomical distribution or burden, the rate of ischaemic stroke exceeded that of intracranial haemorrhage (for ten or more cerebral microbleeds, 64 ischaemic strokes [95% CI 48-84] per 1000 patient-years vs 27 intracranial haemorrhages [17-41] per 1000 patient-years; and for ≥20 cerebral microbleeds, 73 ischaemic strokes [46-108] per 1000 patient-years vs 39 intracranial haemorrhages [21-67] per 1000 patient-years). INTERPRETATION: In patients with recent ischaemic stroke or transient ischaemic attack, cerebral microbleeds are associated with a greater relative hazard (aHR) for subsequent intracranial haemorrhage than for ischaemic stroke, but the absolute risk of ischaemic stroke is higher than that of intracranial haemorrhage, regardless of cerebral microbleed presence, antomical distribution, or burden

Toxocara canis (Ascaridida: Nematoda): Mitochondrial gene content, arrangement and composition compared with other Toxocara species

Normal 0 false false false EN-US X-NONE X-NONE Introduction: The complete mitochondrial (mt) genomes of nematodes sequenced thus far are circular and small in length, containing 37 genes, including 12–13 protein-coding genes: namely, cytochrome oxidase subunits 1–3 (cox1–3), nicotinamide adenine dinucleotide dehydrogenase subunits 1–6 and 4L (nad1–6 and nad4L), ATP synthase subunit 6 and 8 (atp6 and 8), cytochrome b (cytb), small subunit ribosomal RNA gene (rrnS), large subunit ribosomal RNA. The aims of the present study are to determine the gene order, composition, codon usage pattern, and translation initiation/termination codons of the mt genome of Toxocara canis found in Sri Lanka and to compare those findings with other Toxocara species. Methodology: Toxocara canis worms used in the present study were obtained from Sri Lanka. The total genomic DNA was extracted using a part of a single worm using an Easy-DNATM Kit. PCR reactions were performed as described previously by Wickramasinghe et al., (2009). Nucleotide sequences were determined with an ABI PRISM 3100 Avant DNA sequencer using a Big Dye Terminator v3.1 Cycle Sequencing Kit. The open-reading frames and codon usage profiles of protein-coding genes were analyzed using the program DNAsis and GENETYXMAC (ver. 6.0). Results: The partial mt genome of T. canis is 9611 bp in length. This partial mt genome contains 9 protein-coding genes (cox1-3, nad2-5, atp6 and cytb), 14 transfer RNA (tRNA) genes and one ribosomal RNA gene. All genes are transcribed in the same direction as found in other members of the genus Toxocara. The nucleotide compositions of the obtained partial mtDNA sequences for T. canis is biased toward A and T, with T being the most favored nucleotide and C the least favored, in accordance with mt genomes of other nematodes. The content of A+T is 67.48% for T. canis (19.9% A, 47.5% T, 23.3% G and 9.3% C). The start codons we inferred in T. canis were ATT, GTG, ATA, GTT, TTG and ATG. Stop codons were TAG, TA and T. Six of the 8 protein genes used TAG as a translation termination codon and the other two genes (atp6 and nad2) used TA and T respectively. The length of 14 tRNAs is ranging from 52 bp to 68 bp and 8 more tRNAs remain to be sequenced and identified. The rrnL gene of T. canis was identified by sequence comparison with other Toxocara species. The rrnL is located between tRNA-His and nad3. The rrnL gene is 958 bp in length. A non-coding region is located between nad4 and cox1. The length of this region is 112 bp in T. canis. Conclusion: The present study determined the partial mt genome sequence of T. canis obtained from a dog in Sri Lanka. This study provides molecular markers for studying the systematics, population genetics and ecology of the nematodes of socioeconomic importance. Furthermore, we observed a significant difference in nucleotide length of the mitochondrial protein-coding genes, tRNAs, rrnL and non-coding regions of T. canis from three different geographical locations, Sri Lanka, China and Australia. These variations should be further investigated in future studies. Note: Nucleotide data reported in this paper is available in the GenBankTM, EMBL and DDBJ databases under the accession number JN593098.DOI: http://dx.doi.org/10.4038/sljid.v4i2.6946 Sri Lankan Journal of Infectious Diseases 2014; Vol.4(2):90-98 <!--[endif] --

Production of Verrucarol

Verrucarol was obtained from a simple procedure that involved the hydrolysis of a crude extract of a culture of Myrothecium verrucaria ATCC 24571

Morphological and molecular characterization of Paragonimus westermani in northeastern India

Evidence for the presence of lung flukes of the Paragonimus westermani in India remains scant. In particular, evidence based on morphology of adult worms is lacking. Metacercariae of the genus Paragonimus, recovered from crabs in two regions of northeastern India, were raised to adulthood in laboratory rats. Morphologically, these worms appear to be P. westermani. DNA sequences from the second internal transcribed spacer (ITS2) and a portion of the ribosomal large subunit gene (28S) of the nuclear ribosomal RNA gene repeat, as well as fragments of the mitochondrial cytochrome c oxidase subunit 1 (cox1) and NADH dehydrogenase subunit 1 (nad1) genes, all supported this identification. Molecular phylogenetic methods were used for studying the relatedness of these Indian flukes with counterparts from southeast and far-east Asia. Molecular data showed that Indian representatives of the P. westermani complex represent a distinct lineage. It is unclear whether the Indian form can cause disease in humans as some members of the complex do elsewhere