Outbreak of acute fasciolosis in sheep farms in a Mediterranean area arising as a possible consequence of climate change.

The objective of the present study was to investigate whether climate change in recent years have influenced the onset of acute outbreaks of Fasciola hepatica in ovine farms in southern Italy. In May-June 2014, a severe outbreak of F. hepatica occurred in three sheep farms in the Campania region. Clinical, coprological and necroscopic examinations were performed. Morbidity and mortality due to F. hepatica were 3-67% and 3-50%, respectively. Coprological examinations showed high values of F. hepatica eggs per gram (EPG) of faeces (860-1,240). Similarly, high adult parasitic burdens were found in animals that had sucombed (124-426 flukes). The study area was georeferenced and climatic data (temperature, humidity, days of rain and total amount of rainfall) were recorded at four georeferenced meterological stations in the study area. Montly data were processed and analyzed for the period 2000-2013 to evaluate the change of the climatic parameters during these years. The results show that there was a significant increase (P<0.001) of temperature, increased rainfall and increase in the number of rainy days compared to previous years. In addition to the outbreak reported here, we discuss the potential effects of climate change on the epidemiology of F. hepatica and the implications for sheep farming in the Mediterranean area

Protease-activated receptor 2 sensitizes the capsaicin receptor transient receptor potential vanilloid receptor 1 to induce hyperalgesia

Inflammatory proteases (mast cell tryptase and trypsins) cleave protease-activated receptor 2 (PAR2) on spinal afferent neurons and cause persistent inflammation and hyperalgesia by unknown mechanisms. We determined whether transient receptor potential vanilloid receptor 1 (TRPV1), a cation channel activated by capsaicin, protons, and noxious heat, mediates PAR2-induced hyperalgesia. PAR2 was coexpressed with TRPV1 in small- to medium-diameter neurons of the dorsal root ganglia (DRG), as determined by immunofluorescence. PAR2 agonists increased intracellular [Ca2+] ([Ca2+]i) in these neurons in culture, and PAR2-responsive neurons also responded to the TRPV1 agonist capsaicin, confirming coexpression of PAR2 and TRPV1. PAR2 agonists potentiated capsaicin-induced increases in [Ca2+]i in TRPV1-transfected human embryonic kidney (HEK) cells and DRG neurons and potentiated capsaicin-induced currents in DRG neurons. Inhibitors of phospholipase C and protein kinase C (PKC) suppressed PAR2-induced sensitization of TRPV1-mediated changes in [Ca2+]i and TRPV1 currents. Activation of PAR2 or PKC induced phosphorylation of TRPV1 in HEK cells, suggesting a direct regulation of the channel. Intraplantar injection of a PAR2 agonist caused persistent thermal hyperalgesia that was prevented by antagonism or deletion of TRPV1. Coinjection of nonhyperalgesic doses of PAR2 agonist and capsaicin induced hyperalgesia that was inhibited by deletion of TRPV1 or antagonism of PKC. PAR2 activation also potentiated capsaicin-induced release of substance P and calcitonin gene-related peptide from superfused segments of the dorsal horn of the spinal cord, where they mediate hyperalgesia. We have identified a novel mechanism by which proteases that activate PAR2 sensitize TRPV1 through PKC. Antagonism of PAR2, TRPV1, or PKC may abrogate protease-induced thermal hyperalgesia

Antigene MYCN Silencing by BGA002 Inhibits SCLC Progression Blocking mTOR Pathway and Overcomes Multidrug Resistance

: Small-cell lung cancer (SCLC) is the most aggressive lung cancer type, and is associated with smoking, low survival rate due to high vascularization, metastasis and drug resistance. Alterations in MYC family members are biomarkers of poor prognosis for a large number of SCLC. In particular, MYCN alterations define SCLC cases with immunotherapy failure. MYCN has a highly restricted pattern of expression in normal cells and is an ideal target for cancer therapy but is undruggable by traditional approaches. We propose an innovative approach to MYCN inhibition by an MYCN-specific antigene-PNA oligonucleotide (BGA002)-as a new precision medicine for MYCN-related SCLC. We found that BGA002 profoundly and specifically inhibited MYCN expression in SCLC cells, leading to cell-growth inhibition and apoptosis, while also overcoming multidrug resistance. These effects are driven by mTOR pathway block in concomitance with autophagy reactivation, thus avoiding the side effects of targeting mTOR in healthy cells. Moreover, we identified an MYCN-related SCLC gene signature comprehending CNTFR, DLX5 and TNFAIP3, that was reverted by BGA002. Finally, systemic treatment with BGA002 significantly increased survival in MYCN-amplified SCLC mouse models, including in a multidrug-resistant model in which tumor vascularization was also eliminated. These findings warrant the clinical testing of BGA002 in MYCN-related SCLC

The MYCN inhibitor BGA002 restores the retinoic acid response leading to differentiation or apoptosis by the mTOR block in MYCN-amplified neuroblastoma

Background: Neuroblastoma is a deadly childhood cancer, and MYCN-amplified neuroblastoma (MNA-NB) patients have the worst prognoses and are therapy-resistant. While retinoic acid (RA) is beneficial for some neuroblastoma patients, the cause of RA resistance is unknown. Thus, there remains a need for new therapies to treat neuroblastoma. Here we explored the possibility of combining a MYCN-specific antigene oligonucleotide BGA002 and RA as therapeutic approach to restore sensitivity to RA in NB. Methods: By molecular and cellular biology techniques, we assessed the combined effect of the two compounds in NB cell lines and in a xenograft mouse model MNA-NB. Results: We found that MYCN-specific inhibition by BGA002 in combination with RA (BGA002-RA) act synergistically and overcame resistance in NB cell lines. BGA002-RA also reactivated neuron differentiation (or led to apoptosis) and inhibited invasiveness capacity in MNA-NB. Moreover, we found that neuroblastoma had the highest level of mRNA expression of mTOR pathway genes, and that BGA002 led to mTOR pathway inhibition followed by autophagy reactivation in MNA-NB cells, which was strengthened by BGA002-RA. BGA002-RA in vivo treatment also eliminated tumor vascularization in a MNA-NB mouse model and significantly increased survival. Conclusion: Taken together, MYCN modulation mediates the therapeutic efficacy of RA and the development of RA resistance in MNA-NB. Furthermore, by targeting MYCN, a cancer-specific mTOR pathway inhibition occurs only in MNA-NB, thus avoiding the side effects of targeting mTOR in normal cells. These findings warrant clinical testing of BGA002-RA as a strategy for overcoming RA resistance in MNA-NB

A novel MYCN-specific antigene oligonucleotide deregulates mitochondria and inhibits tumor growth in MYCN-amplified neuroblastoma

Approximately half of high-risk neuroblastoma is characterized by MYCN amplification. N-Myc promotes tumor progression by inducing cell growth and inhibiting differentiation. MYCN has also been shown to play an active role in mitochondrial metabolism, but this relationship is not well understood. Although N-Myc is a known driver of the disease, it remains a target for which no therapeutic drug exists. Here, we evaluated a novel MYCN-specific antigene PNA oligonucleotide (BGA002) in MYCN-amplified (MNA) or MYCN-expressing neuroblastoma and investigated the mechanism of its antitumor activity. MYCN mRNA and cell viability were reduced in a broad set of neuroblastoma cell lines following BGA002 treatment. Furthermore, BGA002 decreased N-Myc protein levels and apoptosis in MNA neuroblastoma. Analysis of gene expression data from patients with neuroblastoma revealed that MYCN was associated with increased reactive oxygen species (ROS), downregulated mitophagy, and poor prognosis. Inhibition of MYCN caused profound mitochondrial damage in MNA neuroblastoma cells through downregulation of the mitochondrial molecular chaperone TRAP1, which subsequently increased ROS. Correspondingly, inhibition of MYCN reactivated mitophagy. Systemic administration of BGA002 downregulated N-Myc and TRAP1, with a concomitant decrease in MNA neuroblastoma xenograft tumor weight. In conclusion, this study highlights the role of N-Myc in blocking mitophagy in neuroblastoma and in conferring protection to ROS in mitochondria through upregulation of TRAP1. BGA002 is a potently improved MYCN-specific antigene oligonucleotide that reverts N-Myc\u2013dysregulated mitochondrial pathways, leading to loss of the protective effect of N-Myc against mitochondrial ROS. Significance: A second generation antigene peptide oligonucleotide targeting MYCN induces mitochondrial damage and inhibits growth of MYCN-amplified neuroblastoma cells

Identifying the ion channels responsible for signaling gastro-intestinal based pain

We are normally unaware of the complex signalling events which continuously occur within our internal organs. Most of us only become cognisant when sensations of hunger, fullness, urgency or gas arise. However, for patients with organic and functional bowel disorders pain is an unpleasant and often debilitating reminder. Furthermore, chronic pain still represents a large unmet need for clinical treatment. Consequently, chronic pain has a considerable economic impact on health care systems and the afflicted individuals. In order to address this need we must understand how symptoms are generated within the gut, the molecular pathways responsible for generating these signals and how this process changes in disease states.Stuart M. Brierley, Patrick A. Hughes, Andrea M. Harrington, Grigori Y. Rychkov and L. Ashley Blacksha

Kinin B-1 and B-2 receptors in pig vessels: characterization of two monoreceptor systems

The coronary artery and renal vein of the adult pig are sensitive and reliable monoreceptor systems for studying kinin receptors. The pig coronary artery with intact endothelium is highly sensitive to bradykinin (BK, pEC50 8.6), while being insensitive to the B1 receptor agonist, LysdesArg9BK. The tissue responds to BK with concentration-dependent relaxation, which is prevented by B2 receptor antagonists, particularly DArg[Hyp3, Thi5, DTic7, Oic8]BK (HOE 140, pKB 9.3), (E)-3-(6-acetoamido-3-pyridyl)-N-(N-{2, 4-dichloro-3-[(2-methyl-8-quinolinyl)oxy-methyl]phenyl}-N- methylaminocarbonyl-methyl)acrylamide (FR 173657), a new non peptide compound (pKB 9.3), while B1 receptor antagonists (e.g. Lys[Leu8]desArg9BK) are inactive. The order of potency of kinin-related peptides in this vessel is: LysBK > or = BK > [Hyp3]BK > [Aib7]BK, a sequence typical of a B2 receptor system. Antagonists such as HOE 140 and FR 173657, at high concentrations reduce the maximum effect of BK and thus behave as noncompetitive antagonists. The kinin B1 receptor was studied in the pig renal vein without endothelium and incubated for several hours in order to allow for the de novo formation of this functional site. After 7-8 h in vitro incubation, the vessel shows high sensitivity to LysdesArg9BK (pEC50 8.3) and is insensitive to BK. The pig renal vein responds to B1 receptor agonists with concentration-dependent contraction which maintains a stable plateau and is prevented by selective B1 receptor antagonists such as Lys[Leu8]desArg9BK (pKB 6.7). The most active antagonist has been found to be desArg9HOE 140 (pA2 7.6) which acts as competitive antagonist in this preparation. Some B2 antagonists (e.g. HOE 140) show weak (pKB 6.1) anti-B1 receptor activity while the non-peptide compound FR 173657 is inactive on the B1 receptor and therefore acts as a potent and selective kinin B2 receptor antagonist in the pig. The data obtained in this study allow us to compare the porcine B2 and B1 receptors with those of other species including man, and underline some interesting features that are unique to the porcine functional sites

Cattle gastrointestinal nematode egg-spiked faecal samples: High recovery rates using the Mini-FLOTAC technique

Background: Faecal egg count (FEC) techniques are commonly used to detect gastrointestinal nematodes (GINs) in cattle and to determine anthelmintic efficacy/resistance through the faecal egg count reduction test (FECRT). Mini-FLOTAC is one of the techniques recommended for a standardised FEC/FECRT of helminth eggs in cattle. However, only one paper evaluated the recovery rate of GIN eggs by Mini-FLOTAC (compared to McMaster and modified-Wisconsin method) in cattle, using only a level of contamination of 200 eggs per gram (EPG) of faeces and using GIN eggs collected from goat faeces to spike faecal samples from cattle. To further study the recovery rate of GIN eggs from cattle faeces, this study was conducted in two laboratories, one in Belgium and one in Italy to evaluate the sensitivity, accuracy, precision and reproducibility of the Mini-FLOTAC and McMaster techniques (at two reading levels: grids and chambers) for the detection of GIN eggs in spiked bovine faecal samples. Methods: In both countries, spiked cattle faecal samples with five different levels of egg contamination (10, 50, 100, 200 and 500 EPG) of GINs were used. The study was performed in both laboratories by the same expert operator and using the same standard operating procedures (SOPs) for the Mini-FLOTAC and McMaster techniques. Sensitivity, accuracy and precision were calculated for each technique and for each level of contamination. Statistical analyses were performed to evaluate differences in performance between the two techniques. Results: Mini-FLOTAC had a higher sensitivity (100% at all EPG levels for Mini-FLOTAC vs 0-66.6% for McMaster chambers and grids at levels< 100 EPG), a higher accuracy (98.1% mean value for Mini-FLOTAC vs 83.2% for McMaster grids and 63.8% for McMaster chambers) and a lower coefficient of variation (10.0% for Mini-FLOTAC vs 47.5% for McMaster grids and 69.4% for McMaster chambers) than McMaster. There was no significant difference in the recovery of GIN eggs between the two studies performed in Belgium and in Italy. Conclusions: The high GIN egg recovery rate detected by Mini-FLOTAC and the similar results obtained in Belgium and in Italy indicated that the diagnostic performance of a FEC technique was not dependent on the laboratory environment.[Figure not available: see fulltext.