167 research outputs found
A Liposome-Based Mycobacterial Vaccine Induces Potent Adult and Neonatal Multifunctional T Cells through the Exquisite Targeting of Dendritic Cells
BACKGROUND: In the search for more potent and safer tuberculosis vaccines, CAF01 was identified as a remarkable formulation. Based on cationic liposomes and including a synthetic mycobacterial glycolipid as TLR-independent immunomodulator, it induces strong and protective T helper-1 and T helper-17 adult murine responses to Ag85B-ESAT-6, a major mycobacterial fusion protein. Here, we assessed whether these properties extend to early life and how CAF01 mediates its adjuvant properties in vivo. METHODS/FINDINGS: Following adult or neonatal murine immunization, Ag85B-ESAT-6/CAF01 similarly reduced the post-challenge bacterial growth of M. bovis BCG, whereas no protection was observed using Alum as control. This protection was mediated by the induction of similarly strong Th1 and Th17 responses in both age groups. Multifunctional Th1 cells were already elicited after a single vaccine dose and persisted at high levels for at least 6 months even after neonatal priming. Unexpectedly, this potent adjuvanticity was not mediated by a massive targeting/activation of dendritic cells: in contrast, very few DCs in the draining lymph nodes were bearing the labeled antigen/adjuvant. The increased expression of the CD40 and CD86 activation markers was restricted to the minute portion of adjuvant-bearing DCs. However, vaccine-associated activated DCs were recovered several days after immunization. CONCLUSION: The potent adult and neonatal adjuvanticity of CAF01 is associated in vivo with an exquisite but prolonged DC uptake and activation, fulfilling the preclinical requirements for novel tuberculosis vaccines to be used in early life
Regulation of Neuronal APL-1 Expression by Cholesterol Starvation
Background: Alzheimer’s disease (AD) is a neurodegenerative disorder characterized by the deposition of b-amyloid plaques composed primarily of the amyloid-b peptide, a cleavage product of amyloid precursor protein (APP). While mutations in APP lead to the development of Familial Alzheimer’s Disease (FAD), sporadic AD has only one clear genetic modifier: the e4 allele of the apolipoprotein E (ApoE) gene. Cholesterol starvation in Caenorhabditis elegans leads to molting and arrest phenotypes similar to loss-of-function mutants of the APP ortholog, apl-1 (amyloid precursor-like protein 1), and lrp-1 (lipoprotein receptor-related protein 1), suggesting a potential interaction between apl-1 and cholesterol metabolism. Methodology/Principal Findings: Previously, we found that RNAi knock-down of apl-1 leads to aldicarb hypersensitivity, indicating a defect in synaptic function. Here we find the same defect is recapitulated during lrp-1 knock-down and by cholesterol starvation. A cholesterol-free diet or loss of lrp-1 directly affects APL-1 levels as both lead to loss of APL-1::GFP fluorescence in neurons. However, loss of cholesterol does not affect global transcription or protein levels as seen by qPCR and Western blot. Conclusions: Our results show that cholesterol and lrp-1 are involved in the regulation of synaptic transmission, similar to apl-1. Both are able to modulate APL-1 protein levels in neurons, however cholesterol changes do not affect global apl-1 transcription or APL-1 protein indicating the changes are specific to neurons. Thus, regulation of synaptic transmission an
Caenorhabditis elegans BAH-1 Is a DUF23 Protein Expressed in Seam Cells and Required for Microbial Biofilm Binding to the Cuticle
The cuticle of Caenorhabditis elegans, a complex, multi-layered extracellular matrix, is a major interface between the animal and its environment. Biofilms produced by the bacterial genus Yersinia attach to the cuticle of the worm, providing an assay for surface characteristics. A C. elegans gene required for biofilm attachment, bah-1, encodes a protein containing the domain of unknown function DUF23. The DUF23 domain is found in 61 predicted proteins in C. elegans, which can be divided into three distinct phylogenetic clades. bah-1 is expressed in seam cells, which are among the hypodermal cells that synthesize the cuticle, and is regulated by a TGF-β signaling pathway
Frameless linac-based stereotactic radiosurgery (SRS) for brain metastases: analysis of patient repositioning using a mask fixation system and clinical outcomes
<p>Abstract</p> <p>Purpose</p> <p>To assess the accuracy of patient repositioning and clinical outcomes of frameless stereotactic radiosurgery (SRS) for brain metastases using a stereotactic mask fixation system.</p> <p>Patients and Methods</p> <p>One hundred two patients treated consecutively with frameless SRS as primary treatment at University of Rome Sapienza Sant'Andrea Hospital between October 2008 and April 2010 and followed prospectively were involved in the study. A commercial stereotactic mask fixation system (BrainLab) was used for patient immobilization. A computerized tomography (CT) scan obtained immediately before SRS was used to evaluate the accuracy of patient repositioning in the mask by comparing the isocenter position to the isocenter position established in the planning CT. Deviations of isocenter coordinates in each direction and 3D displacement were calculated. Overall survival, brain control, and local control were estimated using the Kaplan-Meier method calculated from the time of SRS.</p> <p>Results</p> <p>The mean measured isocenter displacements were 0.12 mm (SD 0.35 mm) in the lateral direction, 0.2 mm (SD 0.4 mm) in the anteroposterior, and 0.4 mm (SD 0.6 mm) in craniocaudal direction. The maximum displacement of 2.1 mm was seen in craniocaudal direction. The mean 3D displacement was 0.5 mm (SD 0.7 mm), being maximum 2.9 mm. The median survival was 15.5 months, and 1-year and 2-year survival rates were 58% and 24%, respectively. Nine patients recurred locally after SRS, with 1-year and 2-year local control rates of 91% and 82%, respectively. Stable extracranial disease (P = 0.001) and KPS > 70 (P = 0.01) were independent predictors of survival.</p> <p>Conclusions</p> <p>Frameless SRS is an effective treatment in the management of patients with brain metastases. The presented non-invasive mask-based fixation stereotactic system is associated with a high degree of patient repositioning accuracy; however, a careful evaluation is essential since occasional errors up to 3 mm may occur.</p
ITPase deficiency causes a Martsolf-like syndrome with a lethal infantile dilated cardiomyopathy
Typical Martsolf syndrome is characterized by congenital cataracts, postnatal microcephaly, developmental delay, hypotonia, short stature and biallelic hypomorphic mutations in either RAB3GAP1 or RAB3GAP2. Genetic analysis of 85 unrelated “mutation negative” probands with Martsolf or Martsolf-like syndromes identified two individuals with different homozygous null mutations in ITPA, the gene encoding inosine triphosphate pyrophosphatase (ITPase). Both probands were from multiplex families with a consistent, lethal and highly distinctive disorder; a Martsolf-like syndrome with infantile-onset dilated cardiomyopathy. Severe ITPase-deficiency has been previously reported with infantile epileptic encephalopathy (MIM 616647). ITPase acts to prevent incorporation of inosine bases (rI/dI) into RNA and DNA. In Itpa-null cells dI was undetectable in genomic DNA. dI could be identified at a low level in mtDNA without detectable mitochondrial genome instability, mtDNA depletion or biochemical dysfunction of the mitochondria. rI accumulation was detectable in proband-derived lymphoblastoid RNA. In Itpa-null mouse embryos rI was detectable in the brain and kidney with the highest level seen in the embryonic heart (rI at 1 in 385 bases). Transcriptome and proteome analysis in mutant cells revealed no major differences with controls. The rate of transcription and the total amount of cellular RNA also appeared normal. rI accumulation in RNA–and by implication rI production—correlates with the severity of organ dysfunction in ITPase deficiency but the basis of the cellulopathy remains cryptic. While we cannot exclude cumulative minor effects, there are no major anomalies in the production, processing, stability and/or translation of mRNA
The Carotid and Middle cerebral artery Occlusion Surgery Study (CMOSS): a study protocol for a randomised controlled trial
Intracellular Trafficking and Synaptic Function of APL-1 in Caenorhabditis elegans
Background: Alzheimer’s disease (AD) is a neurodegenerative disorder primarily characterized by the deposition of b-amyloid plaques in the brain. Plaques are composed of the amyloid-b peptide derived from cleavage of the amyloid precursor protein (APP). Mutations in APP lead to the development of Familial Alzheimer’s Disease (FAD), however, the normal function of this protein has proven elusive. The organism Caenorhabditis elegans is an attractive model as the amyloid precursor-like protein (APL-1) is the single ortholog of APP, and loss of apl-1 leads to a severe molting defect and early larval lethality. Methodology/Principal Findings: We report here that lethality and molting can be rescued by full length APL-1, C-terminal mutations as well as a C-terminal truncation, suggesting that the extracellular region of the protein is essential for viability. RNAi knock-down of apl-1 followed by drug testing on the acetylcholinesterase inhibitor aldicarb showed that loss of apl-1 leads to aldicarb hypersensitivity, indicating a defect in synaptic function. The aldicarb hypersensitivity can be rescued by full length APL-1 in a dose dependent fashion. At the cellular level, kinesins UNC-104/KIF-1A and UNC-116/kinesin-1 are positive regulators of APL-1 expression in the neurons. Knock-down of the small GTPase rab-5 also leads to a dramatic decrease in the amount of apl-1 expression in neurons, suggesting that trafficking from the plasma membrane to the early endosome is important for apl-1 function. Loss of function of a different small GTPase, UNC-108, on the contrary, leads t
The NS1 Glycoprotein Can Generate Dramatic Antibody-Enhanced Dengue Viral Replication in Normal Out-Bred Mice Resulting in Lethal Multi-Organ Disease
Antibody-enhanced replication (AER) of dengue type-2 virus (DENV-2) strains and production of antibody-enhanced disease (AED) was tested in out-bred mice. Polyclonal antibodies (PAbs) generated against the nonstructural-1 (NS1) glycoprotein candidate vaccine of the New Guinea-C (NG-C) or NSx strains reacted strongly and weakly with these antigens, respectively. These PAbs contained the IgG2a subclass, which cross-reacted with the virion-associated envelope (E) glycoprotein of the DENV-2 NSx strain, suggesting that they could generate its AER via all mouse Fcγ-receptor classes. Indeed, when these mice were challenged with a low dose (<0.5 LD50) of the DENV-2 NSx strain, but not the NG-C strain, they all generated dramatic and lethal DENV-2 AER/AED. These AER/AED mice developed life-threatening acute respiratory distress syndrome (ARDS), displayed by diffuse alveolar damage (DAD) resulting from i) dramatic interstitial alveolar septa-thickening with mononuclear cells, ii) some hyperplasia of alveolar type-II pneumocytes, iii) copious intra-alveolar protein secretion, iv) some hyaline membrane-covered alveolar walls, and v) DENV-2 antigen-positive alveolar macrophages. These mice also developed meningo-encephalitis, with greater than 90,000-fold DENV-2 AER titers in microglial cells located throughout their brain parenchyma, some of which formed nodules around dead neurons. Their spleens contained infiltrated megakaryocytes with DENV-2 antigen-positive red-pulp macrophages, while their livers displayed extensive necrosis, apoptosis and macro- and micro-steatosis, with DENV-2 antigen-positive Kuppfer cells and hepatocytes. Their infections were confirmed by DENV-2 isolations from their lungs, spleens and livers. These findings accord with those reported in fatal human “severe dengue” cases. This DENV-2 AER/AED was blocked by high concentrations of only the NG-C NS1 glycoprotein. These results imply a potential hazard of DENV NS1 glycoprotein-based vaccines, particularly against DENV strains that contain multiple mutations or genetic recombination within or between their DENV E and NS1 glycoprotein-encoding genes. The model provides potential for assessing DENV strain pathogenicity and anti-DENV therapies in normal mice
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