In 2018, an outbreak of classical swine fever (CSF) was identified in Japan for the first time in 26 years. To prevent the spread of CSF to wild boars, bait vaccination with a commercial vaccine ...derived from the live attenuated C-strain for wild boars has been introduced in Japan since March 2019. Although conventional and real-time RT-PCR methods are suitable for screening and confirmation of suspected cases of the disease, neither of the methods used in Japan can discriminate between CSF virus field strains and live attenuated vaccines. Currently, Sanger sequencing of PCR products is used to distinguish them. In recent years, nanopore sequencing, which is based on a novel sequencing technology, has been adopted for laboratory diagnosis. To save the time and effort required for Sanger sequencing of PCR products, we aimed to determine whether nanopore sequencing would be applicable to CSF virus strain genotyping. PCR products from twenty-nine wild boar samples were evaluated and nanopore sequencing succeeded in discriminating between vaccine and epidemic strains in 28 of 29 samples in less than 90 minutes of total examination time per test. The comparative studies of the sequencing accuracy between Sanger and nanopore sequencing indicated a concordance of ＞96% suggesting that nanopore sequencing could be a promising method for CSF virus strain genotyping despite the short examination time.
Atypical bovine spongiform encephalopathy (BSE), first identified in 2004, poses a threat due to the potential to spread the disease to cattle and other animals, including humans. Here, we estimated ...prion titers in various tissues of cattle infected with atypical BSE using a real-time quaking-induced conversion assay that detects amyloid seeding activity of a disease-specific prion protein, PrPSc, a major component of prions. PrPSc was detected both in and outside of nerve tissues, and some of the peripheral nerve tissues contained relatively high prion titers. Low titers of prions were also observed in masseter, jejunum, and adrenal glands. Quantitative data on prion infectivity in tissues of atypical BSE-affected cattle is useful to assess the risk of atypical BSE.
To determine oral transmissibility of the L-type bovine spongiform encephalopathy (BSE) prion, we orally inoculated 16 calves with brain homogenates of the agent. Only 1 animal, given a high dose, ...showed signs and died at 88 months. These results suggest low risk for oral transmission of the L-BSE agent among cattle.
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Dostopno za:
DOBA, IZUM, KILJ, NUK, ODKLJ, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Recently, autophagy has been associated with the TLR signaling pathway to eliminate intracellular pathogens in the innate immune system. However, it is unknown if other pathways regulate autophagy ...during the immunologic response. Given the critical role of the purinergic P2X7 receptor (P2X7R) pathway during various immunologic functions (i.e., caspase activation and IL-1beta secretion), the principal objective here was to determine whether the P2X7R pathway may regulate autophagy in immune cells. We observed in both MG6 mouse microglial cells and primary microglia that activation of P2X7R by ATP increases the expression of microtubule-associated protein 1 light chain 3 (LC3)-II, the autophagosomal membrane-associated form of LC3, in an extracellular Ca(2+)-dependent manner. Consistent with this, immunohistochemistry showed extensive formation of LC3-immunopositive dots, and electron microscopy demonstrated accumulation of autophagosomes and autophagolysosomes in ATP-treated cells. Importantly, the up-regulation of LC3-II by P2X7R activation was not affected by autophagy inhibitors, such as 3-methyladenine and PI3K inhibitors. Furthermore, while lysosomal functions were impaired by ATP treatment, autophagolysosomal components were released into the extracellular space. Similarly, a phagocytosis assay using Escherichia coli BioParticles showed that phagosome maturation was impaired in ATP-treated cells and a robust release of LC3-immunopositive phagolysosomes was induced along with a radial extension of microtubule bundles. Taken together, the data suggest a novel mechanism whereby the P2X7R signaling pathway may negatively regulate autophagic flux through the impairment of lysosomal functions, leading to stimulation of a release of autophagolysosomes/phagolysosomes into the extracellular space.
The disease-associated prion protein (PrPSc) has the ability to seed the conformational conversion of normal prion proteins into the amyloid fibril form. This prion seeding activity can be measured ...using an in vitro amplification assay termed real-time quaking-induced conversion (RT-QuIC). There is a strong correlation between RT-QuIC positivity and prion infection; however, the relationship between seeding activity and infectivity remains elusive. In this study, we used endpoint dilution RT-QuIC on the brain homogenates from wild-type mice with mouse-adopted bovine spongiform encephalopathy (mBSE) at defined intervals during the incubation period and evaluated the temporal relationship among prion seeding dose, levels of proteinase-resistant PrPSc (PrPres), and infectious titer. We found that the infectious titer reached a plateau by 100 days postinfection, whereas seeding dose and PrPres levels were continuously elevated. Our calculation showed that the doubling time (
) for seeding dose from 40 to 100 days postinoculation was closer to the
for PrPres levels than to the
for prion titer. Although an uncoupling of seeding doses and PrPres levels was observed at end-stage disease in this model, our findings suggest that there is substantial but not complete overlap between PrPSc with seeding activity and PrPres rather than infectious PrPSc.
Tracheobronchial schwannomas are rare diseases. Common signs and symptoms of this tumor include cough, wheezing, and dyspnea. In contrast, pneumothorax is an exceptional presentation. This study ...reports the first case of bronchial schwannoma presenting with pneumothorax. A 79-year-old woman was diagnosed with pneumothorax by chest radiography. Chest computed tomography unexpectedly revealed a tumor occluding the right main bronchus. Following the pathological diagnosis of bronchial schwannoma, the patient underwent thoracoscopic tumor enucleation. The airway lumens are consequently secured postoperatively. We reviewed the literature and discussed the mechanisms and treatment options for bronchial benign tumor-associated pneumothorax. Pneumothorax should be aware of a rare presentation of non-malignant tracheobronchial tumors.
Highlights • Mechanisms underlying GAPDH secretion from mammalian cells are poorly understood. • Exogenous ATP triggered GAPDH release from LPS-primed microglial cells. • P2X7 receptor plays a key ...role in the regulation of GAPDH release. • Exogenous GAPDH modulated LPS-induced activation of p38 MAP kinase in microglial cells. • Released GAPDH might regulate the neuroinflammatory response in the brain.
H-type bovine spongiform encephalopathy (BSE) has been identified in aged cattle in Europe and North America. To determine the localization of disease-associated prion protein (PrPSc) in the ...peripheral nerve tissues of cattle affected with H-type BSE, we employed highly sensitive immunohistochemical and immunofluorescence techniques with the tyramide signal amplification (TSA) system. PrPSc deposition was detected in the inferior ganglia, sympathetic nerve trunk, vagus nerve, spinal nerves, cauda equina, and adrenal medulla, using this system. Notably, granular PrPSc deposits were present mainly in the Schwann cells and fibroblast-like cells and occasionally along certain nerve fibers at the surface of the axons. In the adrenal gland, PrPSc immunolabeling was observed within the sympathetic nerve fibers and nerve endings in the adrenal medulla. Although our results were limited to only 3 experimental cases, these results suggest that the TSA system, a highly sensitive immunohistochemical procedure, may help in elucidating the peripheral pathogenesis of H-type BSE.
Atypical bovine spongiform encephalopathy (BSE) has recently been identified in Europe, North America, and Japan. It is classified as H-type and L-type BSE according to the molecular mass of the ...disease-associated prion protein (PrPSᶜ). To investigate the topographical distribution and deposition patterns of immunolabeled PrPSᶜ, H-type BSE isolate was inoculated intracerebrally into cattle. H-type BSE was successfully transmitted to 3 calves, with incubation periods between 500 and 600 days. Moderate to severe spongiform changes were detected in the cerebral and cerebellar cortices, basal ganglia, thalamus, and brainstem. H-type BSE was characterized by the presence of PrP-immunopositive amyloid plaques in the white matter of the cerebrum, basal ganglia, and thalamus. Moreover, intraglial-type immunolabeled PrPSᶜ was prominent throughout the brain. Stellate-type immunolabeled PrPSᶜ was conspicuous in the gray matter of the cerebral cortex, basal ganglia, and thalamus, but not in the brainstem. In addition, PrPSᶜ accumulation was detected in the peripheral nervous tissues, such as trigeminal ganglia, dorsal root ganglia, optic nerve, retina, and neurohypophysis. Cattle are susceptible to H-type BSE with a shorter incubation period, showing distinct and distinguishable phenotypes of PrPSᶜ accumulation.
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Induction of heat shock protein 70 (HSP70) is known to be effective against various diseases. We are interested in HSP70 induction capability of an antitumor antibiotic bleomycin ...which produces oxidative stress by iron chelate formation and oxygen activation in a cell. The HSP70 induction activity of bleomycin and its six metal core analogs was examined, and a compound HPH-1Trt of 10 μM was found to induce this protein in a pheochromocytoma cell line and some T cell and monocytic cell lines. Its mechanism is increase of HSP70 mRNA, but higher concentration of this compound showed toxicity. Two new derivatives were then synthesized, and one of them named DHPH-1Trt was shown to have less toxicity and higher HSP70 induction activity. This study would lead to a clue for new HSP70 inducer clinically used in near future.
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