A sensitive S-band noise receiver developed for the Mariner Mars 1964 spacecraft program

Sensitive S-band noise receiver for Mariner Mars 1964 spacecraft progra

Effects of fluoroquinolones and tetracyclines on mitochondria of human retinal MIO-M1 cells

Our goal was to explore the detrimental impacts of ciprofloxacin (CPFX) and tetracycline (TETRA) on human retinal Müller (MIO-M1) cells in vitro. Cells were exposed to 30, 60 and 120 μg/ml of CPFX and TETRA. The cellular metabolism was measured with the MTT assay. The JC-1 and CM-H2DCFDA assays were used to evaluate the levels of mitochondrial membrane potential (MMP) and ROS (reactive oxygen species), respectively. Mitochondrial DNA (mtDNA) copy number, along with gene expression levels associated with apoptotic (BAX, BCL2-L13, BCL2, CASP-3 and CASP-9), inflammatory (IL-6, IL-1β, TGF-α, TGF-β1 and TGF-β2) and antioxidant pathways (SOD2, SOD3, GPX3 and NOX4) were analyzed via Quantitative Real-Time PCR (qRT-PCR). Bioenergetic profiles were measured using the Seahorse® XF Flux Analyzer. Cells exposed 24 h to 120 μg/ml TETRA demonstrated higher cellular metabolism compared to vehicle-treated cells. At each time points, (i) all TETRA concentrations reduced MMP levels and (ii) ROS levels were reduced by TETRA 120 μg/ml treatment. TETRA caused (i) higher expression of CASP-3, CASP-9, TGF-α, IL-1B, GPX3 and SOD3 but (ii) decreased levels of TGF-B2 and SOD2. ATP production and spare respiratory capacity declined with TETRA treatment. Cellular metabolism was reduced with CPFX 120 μg/ml in all cultures and 60 μg/ml after 72 h. The CPFX 120 μg/ml reduced MMP in all cultures and ROS levels (72 h). CPFX treatment (i) increased expression of CASP-3, CASP-9, and BCL2-L13, (ii) elevated the basal oxygen consumption rate, and (iii) lowered the mtDNA copy numbers and expression levels of TGF-B2, IL-6 and IL-1B compared to vehicle-control cells. We conclude that clinically relevant dosages of bactericidal and bacteriostatic antibiotics can have negative effects on the cellular metabolism and mitochondrial membrane potential of the retinal MIO-M1 cells in vitro. It is noteworthy to mention that apoptotic and inflammatory pathways in exposed cells were affected significantly This is the first study showing the negative impact of fluoroquinolones and tetracyclines on mitochondrial behavior of human retinal MIO-M1 cells

Valosin-Containing Protein (VCP) Disease and Familial Alzheimer’s Disease: Contrasts and Overlaps

Introduction Contrasts between two entities may be illuminating because of the emphasis on what each is not. Here we describe two proteinopathies producing brain neurodegeneration in mature adults, autosomal dominant valosin-containing protein (VCP) disease and Familial Alzheimer’s disease (FAD) caused by presenillin-1 (PSEN1) mutations, illustrating both contrasting patterns of clinical presentation and known neuropathologic and imaging features, and points of congruence

Pathogenic variants of Valosin-containing protein induce lysosomal damage and transcriptional activation of autophagy regulators in neuronal cells

Aim: Mutations in the valosin-containing protein (VCP) gene cause various lethal proteinopathies that mainly include inclusion body myopathy with Paget's disease of bone and frontotemporal dementia (IBMPFD) and amyotrophic lateral sclerosis (ALS). Different pathological mechanisms have been proposed. Here, we define the impact of VCP mutants on lysosomes and how cellular homeostasis is restored by inducing autophagy in the presence of lysosomal damage. Methods: By electron microscopy, we studied lysosomal morphology in VCP animal and motoneuronal models. With the use of western blotting, real-time quantitative polymerase chain reaction (RT-qPCR), immunofluorescence and filter trap assay, we evaluated the effect of selected VCP mutants in neuronal cells on lysosome size and activity, lysosomal membrane permeabilization and their impact on autophagy. Results: We found that VCP mutants induce the formation of aberrant multilamellar organelles in VCP animal and cell models similar to those found in patients with VCP mutations or with lysosomal storage disorders. In neuronal cells, we found altered lysosomal activity characterised by membrane permeabilization with galectin-3 redistribution and activation of PPP3CB. This selectively activated the autophagy/lysosomal transcriptional regulator TFE3, but not TFEB, and enhanced both SQSTM1/p62 and lipidated MAP1LC3B levels inducing autophagy. Moreover, we found that wild type VCP, but not the mutants, counteracted lysosomal damage induced either by trehalose or by a mutant form of SOD1 (G93A), also blocking the formation of its insoluble intracellular aggregates. Thus, chronic activation of autophagy might fuel the formation of multilamellar bodies. Conclusion: Together, our findings provide insights into the pathogenesis of VCP-related diseases, by proposing a novel mechanism of multilamellar body formation induced by VCP mutants that involves lysosomal damage and induction of lysophagy

Mesenchymal stem cell transplantation for diffuse alveolar hemorrhage in SLE

Background. A 19-year-old girl was diagnosed with systemic lupus erythematosus, based on findings of arthritis, malar rash, positive antinuclear antibody test and high levels of antibodies to double-stranded DNA. Two months after diagnosis, the patient presented with a sudden drop in blood hemoglobin level. Several days later, she developed bloody sputum, rapidly progressive dyspnea and hypoxemia. High-resolution CT showed diffuse alveolar infiltrates in both lung fields.Investigations. Physical examination, complete blood count, erythrocyte sedimentation rate, urinalysis, 24-h urine protein excretion, fecal occult blood test, d-dimer test, acid hemolysis test, activated partial thromboplastin time and prothrombin time, direct and indirect Coombs tests, bone marrow smear, arterial blood gas, sputum smear and culture, and high-resolution CT scan of the chest.Diagnosis. Diffuse alveolar hemorrhage associated with systemic lupus erythematosus.Management. The patient did not respond to pulsed intravenous methylprednisolone (two courses of 500 mg per day for 3 days) and intravenous immunoglobulin (20 g per day for 5 days). The patient was referred to a specialist treatment center for allogenic transplantation using umbilical-cord-derived mesenchymal stem cells. She underwent transplantation with an infusion of 8 - 10 7 mesenchymal stem cells. After showing dramatic improvements in her clinical condition, oxygenation level, radiographic and hematological status, the patient was discharged from hospital approximately 5 weeks after undergoing transplantation. © 2010 Macmillan Publishers Limited.postprin

Numerical and functional defects of blood dendritic cells in early- and late-stage breast cancer

The generation of antitumour immunity depends on the nature of dendritic cell (DC)–tumour interactions. These have been studied mostly by using in vitro-derived DC which may not reflect the natural biology of DC in vivo. In breast cancer, only one report has compared blood DC at different stages and no longitudinal evaluation has been performed. Here we conducted three cross-sectional and one one-year longitudinal assessments of blood DC in patients with early (stage I/II, n=137) and advanced (stage IV, n=36) disease compared to healthy controls (n=66). Patients with advanced disease exhibit markedly reduced blood DC counts at diagnosis. Patients with early disease show minimally reduced counts at diagnosis but a prolonged period (1 year) of marked DC suppression after tumour resection. While differing in frequency, DC from both patients with early and advanced disease exhibit reduced expression of CD86 and HLA-DR and decreased immunostimulatory capacities. Finally, by comparing a range of clinically available maturation stimuli, we demonstrate that conditioning with soluble CD40L induces the highest level of maturation and improved T-cell priming. We conclude that although circulating DC are compromised by loco-regional and systemic breast cancer, they respond vigorously to ex vivo conditioning, thus enhancing their immunostimulatory capacity and potential for immunotherapy

Interferon-inducible gene 202b controls CD8+ T cell-mediated suppression in anti-DNA Ig peptide-treated (NZB × NZW) F1 lupus mice

Administration of an artificial peptide (pConsensus) based on anti-DNA IgG sequences that contain major histocompatibility complex class I and class II T-cell determinants, induces immune tolerance in NZB/NZW F1 female (BWF1) mice. To understand the molecular basis of CD8+ Ti-mediated suppression, we previously performed microarray analysis to identify genes that were differentially expressed following tolerance induction with pCons. CD8+ T cells from mice tolerized with pCons showed more than two-fold increase in Ifi202b mRNA, an interferon inducible gene, versus cells from untolerized mice. Ifi202b expression increased through weeks 1–4 after tolerization and then decreased, reapproaching baseline levels at 6 weeks. In vitro polyclonal activation of tolerized CD8+ T cells significantly increased Ifi202b mRNA expression. Importantly, silencing of Ifi202b abrogated the suppressive capacity of CD8+ Ti cells. This was associated with decreased expression of Foxp3, and decreased gene and protein expression of transforming growth factor (TGF)β and interleukin-2 (IL-2), but not of interferon (IFN)-γ, IL-10, or IL-17. Silencing of another IFN-induced gene upregulated in tolerized CD8+ T cells, IFNAR1, had no effect on the ability of CD8+ T cells to suppress autoantibody production. Our findings indicate a potential role for Ifi202b in the suppressive capacity of peptide-induced regulatory CD8+ Ti cells through effects on the expression of Foxp3 and the synthesis of TGFβ

IBMPFD disease-causing mutant VCP/p97 proteins are targets of autophagic-lysosomal degradation

The ubiquitin-proteasome system (UPS) degrades soluble proteins and small aggregates, whereas macroautophagy (autophagy herein) eliminates larger protein aggregates, tangles and even whole organelles in a lysosome-dependent manner. VCP/p97 was implicated in both pathways. VCP/p97 mutations cause a rare multisystem disease called IBMPFD (Inclusion Body Myopathy with Paget's Disease and Frontotemporal Dementia). Here, we studied the role IBMPFD-related mutants of VCP/p97 in autophagy. In contrast with the wild-type VCP/p97 protein or R155C or R191Q mutants, the P137L mutant was aggregate-prone. We showed that, unlike commonly studied R155C or R191Q mutants, the P137L mutant protein stimulated both autophagosome and autolysosome formation. Moreover, P137L mutant protein itself was a substrate of autophagy. Starvation- and mTOR inhibition-induced autophagy led to the degradation of the P137L mutant protein, while preserving the wild-type and functional VCP/p97. Strikingly, similar to the P137L mutant, other IBMPFD-related VCP/p97 mutants, namely R93C and G157R mutants induced autophagosome and autolysosome formation; and G157R mutant formed aggregates that could be cleared by autophagy. Therefore, cellular phenotypes caused by P137L mutant expression were not isolated observations, and some other IBMPFD disease-related VCP/p97 mutations could lead to similar outcomes. Our results indicate that cellular mechanisms leading to IBMPFD disease may be various, and underline the importance of studying different disease-associated mutations in order to better understand human pathologies and tailor mutation-specific treatment strategies

Pathogenic p62/SQSTM1 mutations impair energy metabolism through limitation of mitochondrial substrates

Abnormal mitochondrial function has been found in patients with frontotemporal dementia (FTD) and amyotrophic lateral sclerosis (ALS). Mutations in the p62 gene (also known as SQSTM1) which encodes the p62 protein have been reported in both disorders supporting the idea of an ALS/FTD continuum. In this work the role of p62 in energy metabolism was studied in fibroblasts from FTD patients carrying two independent pathogenic mutations in the p62 gene, and in a p62-knock-down (p62 KD) human dopaminergic neuroblastoma cell line (SH-SY5Y). We found that p62 deficiency is associated with inhibited complex I mitochondrial respiration due to lack of NADH for the electron transport chain. This deficiency was also associated with increased levels of NADPH reflecting a higher activation of pentose phosphate pathway as this is accompanied with higher cytosolic reduced glutathione (GSH) levels. Complex I inhibition resulted in lower mitochondrial membrane potential and higher cytosolic ROS production. Pharmacological activation of transcription factor Nrf2 increased mitochondrial NADH levels and restored mitochondrial membrane potential in p62-deficient cells. Our results suggest that the phenotype is caused by a loss-of-function effect, because similar alterations were found both in the mutant fibroblasts and the p62 KD model. These findings highlight the implication of energy metabolism in pathophysiological events associated with p62 deficiency