The prion protein (PrPC) is highly expressed within the nervous system. Similar to other GPI-anchored proteins, PrPC is found in lipid rafts, membrane domains enriched in cholesterol and sphingolipids. PrPC raft association, together with raft lipid composition, appears essential for the conversion of PrPC into the scrapie isoform PrPSc, and the development of prion disease. Controversial findings were reported on the nature of PrPC-containing rafts, as well as on the distribution of PrPC between rafts and non-raft membranes. We investigated PrPC/ganglioside relationships and their influence on PrPC localization in a neuronal cellular model, cerebellar granule cells. Our findings argue that in these cells at least two PrPC conformations coexist: in lipid rafts PrPC is present in the native folding (α-helical), stabilized by chemico-physical condition, while it is mainly present in other membrane compartments in a PrPSc-like conformation. We verified, by means of antibody reactivity and circular dichroism spectroscopy, that changes in lipid raft-ganglioside content alters PrPC conformation and interaction with lipid bilayers, without modifying PrPC distribution or cleavage. Our data provide new insights into the cellular mechanism of prion conversion and suggest that GM1-prion protein interaction at the cell surface could play a significant role in the mechanism predisposing to pathology.
It has been shown recently that PrP (prion protein) and the calcium channel auxiliary α2δ subunits interact in neurons and expression systems [Senatore, Colleoni, Verderio, Restelli, Morini, Condliffe, Bertani, Mantovani, Canovi, Micotti, Forloni, Dolphin, Matteoli, Gobbi and Chiesa (2012) Neuron 74, 300–313]. In the present study we examined whether there was an effect of PrP on calcium currents. We have shown that when PrP is co-expressed with calcium channels formed from CaV2.1/β and α2δ-1 or α2δ-2, there is a consistent decrease in calcium current density. This reduction was absent when a PrP construct was used lacking its GPI (glycosylphosphatidylinositol) anchor. We have reported previously that α2δ subunits are able to form GPI-anchored proteins [Davies, Kadurin, Alvarez-Laviada, Douglas, Nieto-Rostro, Bauer, Pratt and Dolphin (2010) Proc. Natl. Acad. Sci. U.S.A. 107, 1654–1659] and show further evidence in the present paper. We have characterized recently a C-terminally truncated α2δ-1 construct, α2δ-1ΔC, and found that, despite loss of its membrane anchor, it still shows a partial ability to increase calcium currents [Kadurin, Alvarez-Laviada, Ng, Walker-Gray, D’Arco, Fadel, Pratt and Dolphin (2012) J. Biol. Chem. 1287, 33554–33566]. We now find that PrP does not inhibit CaV2.1/β currents formed with α2δ-1ΔC, rather than α2δ-1. It is possible that PrP and α2δ-1 compete for GPI-anchor intermediates or trafficking pathways, or that interaction between PrP and α2δ-1 requires association in cholesterol-rich membrane microdomains. Our additional finding that CaV2.1/β1b/α2δ-1 currents were inhibited by GPI–GFP, but not cytosolic GFP, indicates that competition for limited GPI-anchor intermediates or trafficking pathways may be involved in PrP suppression of α2δ subunit function.
PrP, but not GPI-anchorless PrP, suppresses CaV2.1/β/α2δ-1 or α2δ-2 calcium channel currents. However, PrP does not inhibit CaV2.1/β currents formed with anchorless α2δ-1, rather than full-length α2δ-1. The results of the present study suggest that α2δ subunits and PrP may compete for GPI-anchor pathways.
α2δ; auxiliary subunit; calcium channel; GPI anchor; prion protein; CaV, voltage-gated Ca2+; DRM, detergent-resistant membrane; GPI, glycosylphosphatidylinositol; HA, haemagglutinin; KO, knockout; MBS, Mes-buffered saline; PI-PLC, phosphatidylinositol-specific phospholipase C; PNGase F, peptide N-glycosidase F; PrP, prion protein; PrPC, normal cellular PrP; PrPSc, PrP scrapie; WT, wild-type
One of the main challenges for neurodegenerative disorders that are principally incurable is the development of new therapeutic strategies, which raises important medical, scientific and societal issues. Creutzfeldt-Jakob diseases are rare neurodegenerative fatal disorders which today remain incurable. The objective of this study was to evaluate the efficacy of the down-regulation of the prion protein (PrP) expression using siRNA delivered by, a water-in-oil microemulsion, as a therapeutic candidate in a preclinical study. After 12 days rectal mucosa administration of Aonys/PrP-siRNA in mice, we observed a decrease of about 28% of the brain PrPC level. The effect of Aonys/PrP-siRNA was then evaluated on prion infected mice. Several mice presented a delay in the incubation and survival time compared to the control groups and a significant impact was observed on astrocyte reaction and neuronal survival in the PrP-siRNA treated groups. These results suggest that a new therapeutic scheme based an innovative delivery system of PrP-siRNA can be envisioned in prion disorders.
Statins are among the most prescribed drugs worldwide and their recently discovered anti-inflammatory effect seems to have an important role in inhibiting proinflammatory cytokine production, chemokines expression and counteracting the harmful effects of sepsis on the coagulation system. We decided to perform a meta-analysis of all randomized controlled trials ever published on statin therapy in septic patients to evaluate their effect on survival and length of hospital stay.
Data sources and study selection
Articles were assessed by four trained investigators, with divergences resolved by consensus. BioMedCentral, PubMed, Embase and the Cochrane Central Register of clinical trials were searched for pertinent studies. Inclusion criteria were random allocation to treatment and comparison of statins versus any comparator in septic patients.
Data extraction and synthesis
Data from 650 patients in 5 randomized controlled studies were analyzed. No difference in mortality between patients receiving statins versus control (44/322 [14%] in the statins group vs 50/328 [15%] in the control arm, RR = 0.90 [95% CI 0.65 to 1.26], p = 0.6) was observed. No differences in hospital stay (p = 0.7) were found.
Published data show that statin therapy has no effect on mortality in the overall population of adult septic patients. Scientific evidence on statins role in septic patients is still limited and larger randomized trials should be performed on this topic.
Synaptic dysfunction is an important cause of neurological symptoms in prion diseases, a class of clinically heterogeneous neurodegenerative disorders caused by misfolding of the cellular prion protein (PrPC). Experimental data suggest that accumulation of misfolded PrPC in the endoplasmic reticulum (ER) may be crucial in synaptic failure, possibly because of the activation of the translational repression pathway of the unfolded protein response. Here, we report that this pathway is not operative in mouse models of genetic prion disease, consistent with our previous observation that ER stress is not involved. Building on our recent finding that ER retention of mutant PrPC impairs the secretory trafficking of calcium channels essential for synaptic function, we propose a model of pathogenicity in which intracellular retention of misfolded PrPC results in loss of function or gain of toxicity of PrPC-interacting proteins. This neurotoxic modality may also explain the phenotypic heterogeneity of prion diseases.
Growing evidence suggests that a physiological activity of the cellular prion protein (PrPC) plays a crucial role in several neurodegenerative disorders, including prion and Alzheimer’s diseases. However, how the functional activity of PrPC is subverted to deliver neurotoxic signals remains uncertain. Transgenic mice expressing PrP with a deletion of residues 105–125 in the central region (referred to as ΔCR PrP) provide important insights into this problem. Tg(ΔCR) mice exhibit neonatal lethality and massive degeneration of cerebellar granule neurons, a phenotype that is dose-dependently suppressed by the presence of wild-type PrP. When expressed in cultured cells, ΔCR PrP induces large, ionic currents that can be detected by patch-clamping techniques. Here, we have tested the hypothesis that abnormal ion channel activity underlies the neuronal death seen in Tg(ΔCR) mice. We find that ΔCR PrP induces abnormal ionic currents in neurons in culture and in cerebellar slices, and that this activity sensitizes the neurons to glutamate-induced, calcium-mediated death. In combination with ultrastructural and biochemical analyses, these results demonstrate a role for glutamate-induced excitotoxicity in PrP-mediated neurodegeneration. A similar mechanism may operate in other neurodegenerative disorders due to toxic, β-rich oligomers that bind to PrPC.
During prion infection, the normal, protease-sensitive conformation of prion protein (PrPC) is converted via seeded polymerization to an abnormal, infectious conformation with greatly increased protease-resistance (PrPSc). In vitro, protein misfolding cyclic amplification (PMCA) uses PrPSc in prion-infected brain homogenates as an initiating seed to convert PrPC and trigger the self-propagation of PrPSc over many cycles of amplification. While PMCA reactions produce high levels of protease-resistant PrP, the infectious titer is often lower than that of brain-derived PrPSc. More recently, PMCA techniques using bacterially derived recombinant PrP (rPrP) in the presence of lipid and RNA but in the absence of any starting PrPSc seed have been used to generate infectious prions that cause disease in wild-type mice with relatively short incubation times. These data suggest that lipid and/or RNA act as cofactors to facilitate the de novo formation of high levels of prion infectivity. Using rPrP purified by two different techniques, we generated a self-propagating protease-resistant rPrP molecule that, regardless of the amount of RNA and lipid used, had a molecular mass, protease resistance and insolubility similar to that of PrPSc. However, we were unable to detect prion infectivity in any of our reactions using either cell-culture or animal bioassays. These results demonstrate that the ability to self-propagate into a protease-resistant insoluble conformer is not unique to infectious PrP molecules. They suggest that the presence of RNA and lipid cofactors may facilitate the spontaneous refolding of PrP into an infectious form while also allowing the de novo formation of self-propagating, but non-infectious, rPrP-res.
Mice lacking the prion protein (PrPC) gene (Prnp), Ngsk Prnp0/0 mice, show late-onset cerebellar Purkinje cell (PC) degeneration because of ectopic overexpression of PrPC-like protein (PrPLP/Dpl). Because PrPC is highly expressed in cerebellar neurons (including PCs and granule cells), it may be involved in cerebellar synaptic function and cerebellar cognitive function. However, no studies have been conducted to investigate the possible involvement of PrPC and/or PrPLP/Dpl in cerebellum-dependent discrete motor learning. Therefore, the present cross-sectional study was designed to examine cerebellum-dependent delay eyeblink conditioning in Ngsk Prnp0/0 mice in adulthood (16, 40, and 60 weeks of age). The aims of the present study were two-fold: (1) to examine the role of PrPC and/or PrPLP/Dpl in cerebellum-dependent motor learning and (2) to confirm the age-related deterioration of eyeblink conditioning in Ngsk Prnp0/0 mice as an animal model of progressive cerebellar degeneration. Ngsk Prnp0/0 mice aged 16 weeks exhibited intact acquisition of conditioned eyeblink responses (CRs), although the CR timing was altered. The same result was observed in another line of PrPc-deficient mice, ZrchI PrnP0/0 mice. However, at 40 weeks of age, CR incidence impairment was observed in Ngsk Prnp0/0 mice. Furthermore, Ngsk Prnp0/0 mice aged 60 weeks showed more significantly impaired CR acquisition than Ngsk Prnp0/0 mice aged 40 weeks, indicating the temporal correlation between cerebellar PC degeneration and motor learning deficits. Our findings indicate the importance of the cerebellar cortex in delay eyeblink conditioning and suggest an important physiological role of prion protein in cerebellar motor learning.
Transmissible spongiform encephalopathies (TSEs) or prion diseases are associated with accumulations of disease specific PrP (PrPd) in the central nervous system (CNS) and often the lymphoreticular system (LRS). Accumulations have additionally been recorded in other tissues including the peripheral nervous system and adrenal gland. Here we investigate the effect of sheep scrapie on the morphology and the accumulation of PrPd in the adrenal medulla of scrapie affected sheep using light and electron microscopy. Using immunogold electron microscopy, non-fibrillar forms of PrPd were shown to accumulate mainly in association with chromaffin cells, occasional nerve endings and macrophages. PrPd accumulation was associated with distinctive membrane changes of chromaffin cells including increased electron density, abnormal linearity and invaginations. Internalisation of PrPd from the chromaffin cell plasma membrane occurred in association with granule recycling following hormone exocytosis. PrPd accumulation and internalisation from membranes is similarly associated with perturbations of membrane structure and trafficking in CNS neurons and tingible body macrophages of the LRS. These data suggest that a major toxic effect of PrPd is at the level of plasma membranes. However, the precise nature of PrPd-membrane toxicity is tissue and cell specific suggesting that the normal protein may act as a multi-functional scaffolding molecule. We further suggest that the co-localisation of PrPd with exocytic granules of the hormone trafficking system may provide an additional source of infectivity in blood.
Familial Danish dementia (FDD) is an autosomal dominant neurodegenerative disease caused by a 10-nucleotide duplication-insertion in the BRI2 gene. FDD is clinically characterized by loss of vision, hearing impairment, cerebellar ataxia and dementia. The main neuropathologic findings in FDD are the deposition of Danish amyloid (ADan) and the presence of neurofibrillary tangles (NFTs). Here we investigated tau accumulation and truncation in double transgenic (Tg-FDD-Tau) mice generated by crossing transgenic mice expressing human Danish mutant BRI2 (Tg-FDD) with mice expressing human 4-repeat mutant Tau-P301S (Tg-Tau). Compared to Tg-Tau mice, we observed a significant enhancement of tau deposition in Tg-FDD-Tau mice. In addition, a significant increase in tau cleaved at aspartic acid (Asp) 421 was observed in Tg-FDD-Tau mice. Tg-FDD-Tau mice also showed a significant decrease in synaptophysin levels, occurring before widespread deposition of fibrillar ADan and tau can be observed. Thus, the presence of soluble ADan/mutant BRI2 can lead to significant changes in tau metabolism and synaptic dysfunction. Our data provide new in vivo insights into the pathogenesis of FDD and the pathogenic pathway(s) by which amyloidogenic peptides, regardless of their primary amino acid sequence, can cause neurodegeneration.
Machado-Joseph disease (MJD) or spinocerebellar ataxia type 3 (SCA3) is an autosomal dominantly-inherited neurodegenerative disorder caused by the over-repetition of a CAG codon in the MJD1 gene. This expansion translates into a polyglutamine tract that confers a toxic gain-of-function to the mutant protein – ataxin-3, leading to neurodegeneration in specific brain regions, with particular severity in the cerebellum. No treatment able to modify the disease progression is available. However, gene silencing by RNA interference has shown promising results. Therefore, in this study we investigated whether lentiviral-mediated allele-specific silencing of the mutant ataxin-3 gene, after disease onset, would rescue the motor behavior deficits and neuropathological features in a severely impaired transgenic mouse model of MJD. For this purpose, we injected lentiviral vectors encoding allele-specific silencing-sequences (shAtx3) into the cerebellum of diseased transgenic mice expressing the targeted C-variant of mutant ataxin-3 present in 70% of MJD patients. This variation permits to discriminate between the wild-type and mutant forms, maintaining the normal function of the wild-type allele and silencing only the mutant form. Quantitative analysis of rotarod performance, footprint and activity patterns revealed significant and robust alleviation of gait, balance (average 3-fold increase of rotarod test time), locomotor and exploratory activity impairments in shAtx3-injected mice, as compared to control ones injected with shGFP. An important improvement of neuropathology was also observed, regarding the number of intranuclear inclusions, calbindin and DARPP-32 immunoreactivity, fluorojade B and Golgi staining and molecular and granular layers thickness. These data demonstrate for the first time the efficacy of gene silencing in blocking the MJD-associated motor-behavior and neuropathological abnormalities after the onset of the disease, supporting the use of this strategy for therapy of MJD.
Prion diseases, including sheep scrapie, are neurodegenerative diseases with the fundamental pathogenesis involving conversion of normal cellular prion protein (PrPC) to disease-associated prion protein (PrPSc). Chemical inhibition of prion accumulation is widely investigated, often using rodent-adapted prion cell culture models. Using a PrPSc-specific ELISA we discovered a monocationic phenyl-furan-benzimidazole (DB772), which has previously demonstrated anti-pestiviral activity and represents a chemical category previously untested for anti-prion activity, that inhibited PrPSc accumulation and prion infectivity in primary sheep microglial cell cultures (PRNP 136VV/154RR/171QQ) and Rov9 cultures (VRQ-ovinized RK13 cells). We investigated potential mechanisms of this anti-prion activity by evaluating PrPC expression with quantitative RT-PCR and PrP ELISA, comparing the concentration-dependent anti-prion and anti-pestiviral effects of DB772, and determining the selectivity index. Results demonstrate at least an approximate two-log inhibition of PrPSc accumulation in the two cell systems and confirmed that the inhibition of PrPSc accumulation correlates with inhibition of prion infectivity. PRNP transcripts and total PrP protein concentrations within cell lysates were not decreased; thus, decreased PrPC expression is not the mechanism of PrPSc inhibition. PrPSc accumulation was multiple logs more resistant than pestivirus to DB772, suggesting that the anti-PrPSc activity was independent of anti-pestivirus activity. The anti-PrPSc selectivity index in cell culture was approximately 4.6 in microglia and 5.5 in Rov9 cells. The results describe a new chemical category that inhibits ovine PrPSc accumulation in primary sheep microglia and Rov9 cells, and can be used for future studies into the treatment and mechanism of prion diseases.
Although open surgical repair of thoracoabdominal aortic aneurysm (TAAA) has evolved significantly over the last few decades, technical challenges and current morbidity and mortality of the inclusion technique are still significant, particularly in patients with extensive aneurysms or prior aortic surgery and in poor surgical candidates. Hybrid TAAA repair is an appealing technique and may represent a “bridge” solution as we wait for larger series and reproducible results from the evolving experience with total endovascular TAAA repair with fenestrated and branched endografts.
Thoracoabdominal aortic aneurysm (TAAA); extensive aneurysms; total endovascular TAAA repair
Thoracoabdominal aortic aneurysm (TAAA) hybrid repair consists of aortic visceral branch rerouting followed by TAAA endograft exclusion. This technique has been shown to represent a technically feasible strategy in selected patients.
We analyzed 52 high-risk patients who underwent hybrid TAAA repair between 2001 and 2012 in our centre with a variety of visceral rerouting configurations and of commercially available thoracic endografts. Thirty-seven simultaneous (71.2%) and 15 staged procedures (21.8%) were performed with a four-vessel revascularization in 18 cases (34.6%), a three-vessel revascularization in 11 cases (21.2%) and a two-vessel revascularization in 23 cases (44.2%).
No intraoperative deaths were observed. We recorded a perioperative mortality rate of 13.5% (n=7), including deaths from multiorgan failure (n=2), myocardial infarction (n=2), coagulopathy (n=1), pancreatitis (n=1) and bowel infarction (n=1). Perioperative morbidity rate was 28.8% (n=15), including 2 cases of transient paraparesis and 1 case of permanent paraplegia. Renal failure (n=5), pancreatitis (n=3), respiratory failure (n=3) and dysphagia (n=1) were also observed. At median follow-up of 23.9 months procedure-related mortality rate was 9.6%: two patients died from visceral graft occlusion and three from aortic rupture. There were three endoleaks and one endograft migration, none of which resulted in death. Five patients (9.6%) died as a consequence of unrelated events.
Typical complications of conventional TAAA open surgery have not been eliminated by hybrid repair, and significant mortality and morbidity rates have been recorded. Fate of visceral bypasses and incidence of endoleak and other endograft-related complications needs to be carefully assessed. Hybrid TAAA repair should currently be limited to high-risk surgical patients with unfit anatomy for endovascular repair.
Thoracoabdominal; aorta; aneurysm; hybrid; endovascular
Open surgical repair of thoracoabdominal aortic aneurysms has evolved significantly over the last decades thanks to technical improvements, especially in the area of organ protection. However, despite adjunctive strategies, morbidity and mortality rates are still not negligible. Repair of the thoracoabdominal aorta represents a formidable challenge for surgeons, anesthesiologists and patients alike. While operative repair is generally carried out in specialized institutions, knowledge of the state-of-the-art diagnostic, anesthesiologic, surgical and endovascular aspects will certainly be of great value to all physicians involved in the care of these patients at any level. This “How to” video will explain all of these diagnostic, anesthesiologic and surgical aspects in our daily practice.
Thoracoabdominal aortic aneurysm; diagnosis; open surgical rapair
Coronary atherosclerosis, the main condition predisposing to acute myocardial infarction, has an inflammatory component caused by stimuli that are yet unknown. We molecularly investigated the nature of the immune response within human coronary lesion in four coronary plaques obtained by endoluminal atherectomy from four patients. We constructed phage-display libraries containing the IgG1/kappa antibody fragments produced by B-lymphocytes present in each plaque. By immunoaffinity, we selected from these libraries a monoclonal antibody, arbitrarily named Fab7816, able to react both with coronary and carotid atherosclerotic tissue samples. We also demonstrated by confocal microscopy that this monoclonal antibody recognized human transgelin type 1, a cytoskeleton protein involved in atherogenesis, and that it co-localized with fibrocyte-like cells transgelin+, CD68+, CD45+ in human sections of coronary and carotid plaques. In vitro fibrocytes obtained by differentiating CD14+ cells isolated from peripheral blood mononuclear cells also interacted with Fab7816, thus supporting the hypothesis of a specific recognition of fibrocytes into the atherosclerotic lesions. Interestingly, the same antibody, cross-reacted with the outer membrane proteins of Proteus mirabilis and Klebsiella pneumoniae (and possibly with homologous proteins of other enterobacteriaceae present in the microbiota). From all the other three libraries, we were able to clone, by immunoaffinity selection, human monoclonal antibodies cross-reacting with bacterial outer membrane proteins and with transgelin. These findings demonstrated that in human atherosclerotic plaques a local cross-reactive immune response takes place.
Thoracic outlet syndrome is an often misdiagnosed syndrome which consists of a neurovascular compression at the upper thoracic outlet. The clinical presentation can be variable, ranging from mild symptoms to venous thrombosis and muscle atrophy. Many aetiologies, both congenital and acquired, related either to bony or soft tissue anomalies, have been associated with this syndrome. As a consequence, the diagnosis is often challenging and sometimes it can be obtained only with surgical exploration. Additionally, no specific clinical test is considered diagnostic of thoracic outlet syndrome. However, the recent advances in imaging techniques together with a careful clinical evaluation give the surgeon the chance to recognize the constricting anatomy before surgery in many cases. No standard surgical procedure has been identified; however, in literature the largest series have been treated with transaxillary first rib resection. Here we report our experience in the surgical treatment of this syndrome with a minimum follow-up of three years. Our approach consists of performing a supraclavicular decompression without routine first rib resection. This allows for identifying and removing the constricting anatomy in most cases, with satisfactory results in 96.9% of patients and a low complication rate.
How mutant prion protein (PrP) leads to neurological dysfunction in genetic prion diseases is unknown. Tg(PG14) mice synthesize a misfolded mutant PrP which is partially retained in the neuronal endoplasmic reticulum (ER). As these mice age, they develop ataxia and massive degeneration of cerebellar granule neurons (CGNs). Here, we report that motor behavioral deficits in Tg(PG14) mice emerge before neurodegeneration and are associated with defective glutamate exocytosis from granule neurons due to impaired calcium dynamics. We found that mutant PrP interacts with the voltage-gated calcium channel α2δ-1 subunit, which promotes the anterograde trafficking of the channel. Owing to ER retention of mutant PrP, α2δ-1 accumulates intracellularly, impairing delivery of the channel complex to the cell surface. Thus, mutant PrP disrupts cerebellar glutamatergic neurotransmission by reducing the number of functional channels in CGNs. These results link intracellular PrP retention to synaptic dysfunction, indicating new modalities of neurotoxicity and potential therapeutic strategies.
► Mutant PrP dysrupts glutamatergic transmission in cerebellar granule neurons (CGNs) ► Mutant PrP impairs depolarization-evoked calcium dynamics in CGNs ► Mutant PrP binds to the α2δ-1 subunit of voltage-gated calcium channels (VGCCs) ► Mutant PrP misfolding and intracellular retention impairs membrane delivery of VGCCs
How mutant prion protein causes neurological dysfunction in genetic prion diseases is not fully known. Senatore et al. report intracellular accumulation of misfolded prion impairs voltage-gated calcium channel transport to synapses, altering glutamatergic neurotransmission and cerebellum-dependent motor function.
Insight into the normal function of PrPC, and how it can be subverted to produce neurotoxic effects, is provided by PrP molecules carrying deletions encompassing the conserved central region. The most neurotoxic of these mutants, Δ105–125 (called ΔCR), produces a spontaneous neurodegenerative illness when expressed in transgenic mice, and this phenotype can be dose-dependently suppressed by co-expression of wild-type PrP. Whether the toxic activity of ΔCR PrP and the protective activity or wild-type PrP are cell-autonomous, or can be exerted on neighboring cells, is unknown. To investigate this question, we have utilized co-cultures of differentiated neural stem cells derived from mice expressing ΔCR or wild-type PrP. Cells from the two kinds of mice, which are marked by the presence or absence of GFP, are differentiated together to yield neurons, astrocytes, and oligodendrocytes. As a surrogate read-out of ΔCR PrP toxicity, we assayed sensitivity of the cells to the cationic antibiotic, Zeocin. In a previous study, we reported that cells expressing ΔCR PrP are hypersensitive to the toxic effects of several cationic antibiotics, an effect that is suppressed by co-expression of wild type PrP, similar to the rescue of the neurodegenerative phenotype observed in transgenic mice. Using this system, we find that while ΔCR-dependent toxicity is cell-autonomous, the rescuing activity of wild-type PrP can be exerted in trans from nearby cells. These results provide important insights into how ΔCR PrP subverts a normal physiological function of PrPC, and the cellular mechanisms underlying the rescuing process.
Endovascular treatment of complex thoracic pathologic conditions involving the aortic arch can often be appropriate and safe; however, minimally invasive procedures are not always feasible, especially in emergent cases. We report the case of a 78-year-old woman who emergently presented in hemorrhagic shock with a ruptured chronic dissecting aneurysm that involved the aortic arch. Eight years earlier, she had undergone aortic valve replacement and plication of the ascending aorta, which was complicated a day later by Stanford type B dissection, malperfusion, and ischemia that required an axillobifemoral bypass. At the current admission, we successfully treated her surgically through a left thoracotomy, using moderate hypothermic extracorporeal circulation and advanced organ-protection methods. We discuss the surgical indications and our operative strategy in relation to open surgical repair versus endovascular treatment in patients with complex conditions.
Aneurysm, dissecting/radiography/surgery; aortic aneurysm, thoracic/radiography/surgery; aortic diseases/surgery; aortic rupture/surgery; treatment outcome; vascular surgical procedures
The cellular pathways activated by mutant prion protein (PrP) in genetic prion diseases, ultimately leading to neuronal dysfunction and degeneration, are not known. Several mutant PrPs misfold in the early secretory pathway and reside longer in the endoplasmic reticulum (ER) possibly stimulating ER stress-related pathogenic mechanisms. To investigate whether mutant PrP induced maladaptive responses, we checked key elements of the unfolded protein response (UPR) in transgenic mice, primary neurons and transfected cells expressing two different mutant PrPs. Because ER stress favors the formation of untranslocated PrP that might aggregate in the cytosol and impair proteasome function, we also measured the activity of the ubiquitin proteasome system (UPS). Molecular, biochemical and immunohistochemical analyses found no increase in the expression of UPR-regulated genes, such as Grp78/Bip, CHOP/GADD153, or ER stress-dependent splicing of the mRNA encoding the X-box-binding protein 1. No alterations in UPS activity were detected in mutant mouse brains and primary neurons using the UbG76V-GFP reporter and a new fluorogenic peptide for monitoring proteasomal proteolytic activity in vivo. Finally, there was no loss of proteasome function in neurons in which endogenous PrP was forced to accumulate in the cytosol by inhibiting cotranslational translocation. These results indicate that neither ER stress, nor perturbation of proteasome activity plays a major pathogenic role in prion diseases.
Aneurysm, dissecting/surgery; aortic aneurysm, thoracic/surgery; blood vessel prosthesis implantation; endovascular procedures; stents
Aortic diseases/therapy; blood vessel prosthesis implantation; bronchial fistula/therapy; esophageal fistula/therapy; vascular fistula/therapy
Pseuodaneurysms are an extremely rare complication of osteochondromas. We describe a case of traumatic pseudoaneurysm of the brachial artery presenting as a soft tissue mass in a patient who was treated for an osteochondroma 3 years earlier. This case demonstrates that radiographic follow-up of large osteochondromas is mandatory and that, in patients with soft tissue masses and a history of osteochondroma, pseudoaneurysms should be included in the differential diagnosis.
Pseudoaneurysm; Osteochondroma; Soft tissue mass; Brachial artery