The adaptor protein PID1 regulates receptor-dependent endocytosis of postprandial triglyceride-rich lipoproteins.

ObjectiveInsulin resistance is associated with impaired receptor dependent hepatic uptake of triglyceride-rich lipoproteins (TRL), promoting hypertriglyceridemia and atherosclerosis. Next to low-density lipoprotein (LDL) receptor (LDLR) and syndecan-1, the LDLR-related protein 1 (LRP1) stimulated by insulin action contributes to the rapid clearance of TRL in the postprandial state. Here, we investigated the hypothesis that the adaptor protein phosphotyrosine interacting domain-containing protein 1 (PID1) regulates LRP1 function, thereby controlling hepatic endocytosis of postprandial lipoproteins.MethodsLocalization and interaction of PID1 and LRP1 in cultured hepatocytes was studied by confocal microscopy of fluorescent tagged proteins, by indirect immunohistochemistry of endogenous proteins, by GST-based pull down and by immunoprecipitation experiments. The in vivo relevance of PID1 was assessed using whole body as well as liver-specific Pid1-deficient mice on a wild type or Ldlr-deficient (Ldlr-/-) background. Intravital microscopy was used to study LRP1 translocation in the liver. Lipoprotein metabolism was investigated by lipoprotein profiling, gene and protein expression as well as organ-specific uptake of radiolabelled TRL.ResultsPID1 co-localized in perinuclear endosomes and was found associated with LRP1 under fasting conditions. We identified the distal NPxY motif of the intracellular C-terminal domain (ICD) of LRP1 as the site critical for the interaction with PID1. Insulin-mediated NPxY-phosphorylation caused the dissociation of PID1 from the ICD, causing LRP1 translocation to the plasma membrane. PID1 deletion resulted in higher LRP1 abundance at the cell surface, higher LDLR protein levels and, paradoxically, reduced total LRP1. The latter can be explained by higher receptor shedding, which we observed in cultured Pid1-deficient hepatocytes. Consistently, PID1 deficiency alone led to increased LDLR-dependent endocytosis of postprandial lipoproteins and lower plasma triglycerides. In contrast, hepatic PID1 deletion on an Ldlr-/- background reduced lipoprotein uptake into liver and caused plasma TRL accumulation.ConclusionsBy acting as an insulin-dependent retention adaptor, PID1 serves as a regulator of LRP1 function controlling the disposal of postprandial lipoproteins. PID1 inhibition provides a novel approach to lower plasma levels of pro-atherogenic TRL remnants by stimulating endocytic function of both LRP1 and LDLR in the liver

Intravitreal injection of low-dose gentamicin for the treatment of recurrent or persistent uveitis in horses: Preliminary results.

Background: Despite appropriate medical therapy, many horses with equine recurrent uveitis continue to suffer from recurrent bouts of inflammation. Surgical interve tion via the pars plana vitrectomy or suprachoroidal cyclosporine implant placement may control and/or prevent recurrences, however, these procedures may be contraindicated, unavailable, or declined by an owner. Thus, an effective adjunctive treatment option may help to improve the clinical outcomes in those situations. There are several anecdotal reports on the use of intravitreal gentamicin injections, but to date, no data evaluating the complication rate and/or treatment effect following this treatment have been published. Thus, the aim of this prospective study was to describe the intravitreal gentamicin injection technique, describe the associated peri-injection (within 24 h) and post- injection (30 to 780 days) complications, and to report the effects of the injection on the clinical signs of uveitis. Additionally, evaluation of the systemic and ocular Leptospira-status, and its effect on the treatment outcome was performed. A total of 86 horses of various ages, breeds, and gender presenting with recurrent or persistent uveitis were treated via intravitreal injection of 4 mg of undiluted gentamicin (0.04 ml, Genta 100, 100 mg/ml in 35 horses) or preservative-free gentamicin (0.05 ml, 80 mg/ml in 52 horses) under sedation and local anesthesia. All 86 horses were observed for immediate peri-injection and post-injection complications. Response to therapy was evaluated in 59 of the 86 horses (follow-up: 30 to 780 days). Results: Peri-injection complications consisted of subconjunctival (26/86; 30.2%) or intracameral hemorrhage (4/86; 4.7%); both of which completely resolved within 5 days. Post-injection complications consisted of cataract formation/maturation (5/59 horses, 8.5%) and diffuse retinal degeneration (3/59 eyes 5.1%). The majority of horses 52/59 (88.1%) with a minimum follow-up period of 30 days were controlled (absence of recurrent or persistent inflammation) at their last recheck examination. Recurrent inflammation was documented in 5/59 (8.5%) horses and persistent inflammation was diagnosed in 2/59 (3.4%) horses. Conclusions: Intravitreal injection of low-dose gentamicin shows promise at controlling different types and stages of uveitis. The ability of intravitreal injections of low-dose gentamicin (4 mg) to control persistent and recurrent inflammation warrants further investigation

Intravitreal injection of low-dose gentamicin for the treatment of recurrent or persistent uveitis in horses: Preliminary results.

Background: Despite appropriate medical therapy, many horses with equine recurrent uveitis continue to suffer from recurrent bouts of inflammation. Surgical interve tion via the pars plana vitrectomy or suprachoroidal cyclosporine implant placement may control and/or prevent recurrences, however, these procedures may be contraindicated, unavailable, or declined by an owner. Thus, an effective adjunctive treatment option may help to improve the clinical outcomes in those situations. There are several anecdotal reports on the use of intravitreal gentamicin injections, but to date, no data evaluating the complication rate and/or treatment effect following this treatment have been published. Thus, the aim of this prospective study was to describe the intravitreal gentamicin injection technique, describe the associated peri-injection (within 24 h) and post- injection (30 to 780 days) complications, and to report the effects of the injection on the clinical signs of uveitis. Additionally, evaluation of the systemic and ocular Leptospira-status, and its effect on the treatment outcome was performed. A total of 86 horses of various ages, breeds, and gender presenting with recurrent or persistent uveitis were treated via intravitreal injection of 4 mg of undiluted gentamicin (0.04 ml, Genta 100, 100 mg/ml in 35 horses) or preservative-free gentamicin (0.05 ml, 80 mg/ml in 52 horses) under sedation and local anesthesia. All 86 horses were observed for immediate peri-injection and post-injection complications. Response to therapy was evaluated in 59 of the 86 horses (follow-up: 30 to 780 days). Results: Peri-injection complications consisted of subconjunctival (26/86; 30.2%) or intracameral hemorrhage (4/86; 4.7%); both of which completely resolved within 5 days. Post-injection complications consisted of cataract formation/maturation (5/59 horses, 8.5%) and diffuse retinal degeneration (3/59 eyes 5.1%). The majority of horses 52/59 (88.1%) with a minimum follow-up period of 30 days were controlled (absence of recurrent or persistent inflammation) at their last recheck examination. Recurrent inflammation was documented in 5/59 (8.5%) horses and persistent inflammation was diagnosed in 2/59 (3.4%) horses. Conclusions: Intravitreal injection of low-dose gentamicin shows promise at controlling different types and stages of uveitis. The ability of intravitreal injections of low-dose gentamicin (4 mg) to control persistent and recurrent inflammation warrants further investigation

Interleukin-7 Links T Lymphocyte and Intestinal Epithelial Cell Homeostasis

Interleukin-7 (IL-7) is a major survival factor for mature T cells. Therefore, the degree of IL-7 availability determines the size of the peripheral T cell pool and regulates T cell homeostasis. Here we provide evidence that IL-7 also regulates the homeostasis of intestinal epithelial cells (IEC), colon function and the composition of the commensal microflora. In the colon of T cell-deficient, lymphopenic mice, IL-7-producing IEC accumulate. IEC hyperplasia can be blocked by IL-7-consuming T cells or the inactivation of the IL-7/IL-7R signaling pathway. However, the blockade of the IL-7/IL-7R signaling pathway renders T cell-deficient mice more sensitive to chemically-induced IEC damage and subsequent colitis. In summary, our data demonstrate that IL-7 promotes IEC hyperplasia under lymphopenic conditions. Under non-lymphopenic conditions, however, T cells consume IL-7 thereby limiting IEC expansion and survival. Hence, the degree of IL-7 availability regulates both, T cell and IEC homeostasis

Sulfur isotopes in rivers : insights into global weathering budgets, pyrite oxidation, and the modern sulfur cycle

This research was funded by a Foster and Coco Stanback postdoctoral fellowship and a Marie Curie Career Integration Grant (CIG14-631752) to AB. JFA acknowledges the support of NSF-OCE grant 1340174 and NSF-EAR grant 1349858. WF acknowledges the support of a grant from the David and Lucile Packard Foundation.The biogeochemical sulfur cycle is intimately linked to the cycles of carbon, iron, and oxygen, and plays an important role in global climate via weathering reactions and aerosols. However, many aspects of the modern budget of the global sulfur cycle are not fully understood. We present new δ34S measurements on sulfate from more than 160 river samples from different geographical and climatic regions—more than 46% of the world's freshwater flux to the ocean is accounted for in this estimate of the global riverine sulfur isotope budget. These measurements include major rivers and their tributaries, as well as time series, and are combined with previously published data to estimate the modern flux-weighted global riverine δ34S as 4.4 ± 4.5‰ (V-CDT), and 4.8 ± 4.9‰ when the most polluted rivers are excluded. The sulfur isotope data, when combined with major anion and cation concentrations, allow us to tease apart the relative contributions of different processes to the modern riverine sulfur budget, resulting in new estimates of the flux of riverine sulfate due to the oxidative weathering of pyrites (1.3 ± 0.2 Tmol S/y) and the weathering of sedimentary sulfate minerals (1.5 ± 0.2 Tmol S/y). These data indicate that previous estimates of the global oxidative weathering of pyrite have been too low by a factor of two. As pyrite oxidation coupled to carbonate weathering can act as a source of CO2 to the atmosphere, this global pyrite weathering budget implies that the global CO2 weathering sink is overestimated. Furthermore, the large range of sulfur isotope ratios in modern rivers indicates that secular changes in the lithologies exposed to weathering through time could play a major role in driving past variations in the δ34S value of seawater.PostprintPostprintPostprintPeer reviewe

Different modes of state transitions determine pattern in the Phosphatidylinositide-Actin system

<p>Abstract</p> <p>Background</p> <p>In a motile polarized cell the actin system is differentiated to allow protrusion at the front and retraction at the tail. This differentiation is linked to the phosphoinositide pattern in the plasma membrane. In the highly motile <it>Dictyostelium </it>cells studied here, the front is dominated by PI3-kinases producing PI(3,4,5)tris-phosphate (PIP3), the tail by the PI3-phosphatase PTEN that hydrolyses PIP3 to PI(4,5)bis-phosphate. To study de-novo cell polarization, we first depolymerized actin and subsequently recorded the spontaneous reorganization of actin patterns in relation to PTEN.</p> <p>Results</p> <p>In a transient stage of recovery from depolymerization, symmetric actin patterns alternate periodically with asymmetric ones. The switches to asymmetry coincide with the unilateral membrane-binding of PTEN. The modes of state transitions in the actin and PTEN systems differ. Transitions in the actin system propagate as waves that are initiated at single sites by the amplification of spontaneous fluctuations. In PTEN-null cells, these waves still propagate with normal speed but loose their regular periodicity. Membrane-binding of PTEN is induced at the border of a coherent PTEN-rich area in the form of expanding and regressing gradients.</p> <p>Conclusions</p> <p>The state transitions in actin organization and the reversible transition from cytoplasmic to membrane-bound PTEN are synchronized but their patterns differ. The transitions in actin organization are independent of PTEN, but when PTEN is present, they are coupled to periodic changes in the membrane-binding of this PIP3-degrading phosphatase. The PTEN oscillations are related to motility patterns of chemotaxing cells.</p

Peroxisomal alterations in Alzheimer’s disease

In Alzheimer’s disease (AD), lipid alterations are present early during disease progression. As some of these alterations point towards a peroxisomal dysfunction, we investigated peroxisomes in human postmortem brains obtained from the cohort-based, longitudinal Vienna-Transdanube Aging (VITA) study. Based on the neuropathological Braak staging for AD on one hemisphere, the patients were grouped into three cohorts of increasing severity (stages I–II, III–IV, and V–VI, respectively). Lipid analyses of cortical regions from the other hemisphere revealed accumulation of C22:0 and very long-chain fatty acids (VLCFA, C24:0 and C26:0), all substrates for peroxisomal β-oxidation, in cases with stages V–VI pathology compared with those modestly affected (stages I–II). Conversely, the level of plasmalogens, which need intact peroxisomes for their biosynthesis, was decreased in severely affected tissues, in agreement with a peroxisomal dysfunction. In addition, the peroxisomal volume density was increased in the soma of neurons in gyrus frontalis at advanced AD stages. Confocal laser microscopy demonstrated a loss of peroxisomes in neuronal processes with abnormally phosphorylated tau protein, implicating impaired trafficking as the cause of altered peroxisomal distribution. Besides the original Braak staging, the study design allowed a direct correlation between the biochemical findings and the amount of neurofibrillary tangles (NFT) and neuritic plaques, quantified in adjacent tissue sections. Interestingly, the decrease in plasmalogens and the increase in VLCFA and peroxisomal volume density in neuronal somata all showed a stronger association with NFT than with neuritic plaques. These results indicate substantial peroxisome-related alterations in AD, which may contribute to the progression of AD pathology

Aerosols Transmit Prions to Immunocompetent and Immunodeficient Mice

Prions, the agents causing transmissible spongiform encephalopathies, colonize the brain of hosts after oral, parenteral, intralingual, or even transdermal uptake. However, prions are not generally considered to be airborne. Here we report that inbred and crossbred wild-type mice, as well as tga20 transgenic mice overexpressing PrPC, efficiently develop scrapie upon exposure to aerosolized prions. NSE-PrP transgenic mice, which express PrPC selectively in neurons, were also susceptible to airborne prions. Aerogenic infection occurred also in mice lacking B- and T-lymphocytes, NK-cells, follicular dendritic cells or complement components. Brains of diseased mice contained PrPSc and transmitted scrapie when inoculated into further mice. We conclude that aerogenic exposure to prions is very efficacious and can lead to direct invasion of neural pathways without an obligatory replicative phase in lymphoid organs. This previously unappreciated risk for airborne prion transmission may warrant re-thinking on prion biosafety guidelines in research and diagnostic laboratories

Atlas of the clinical genetics of human dilated cardiomyopathy

[Abstract] Aim. Numerous genes are known to cause dilated cardiomyopathy (DCM). However, until now technological limitations have hindered elucidation of the contribution of all clinically relevant disease genes to DCM phenotypes in larger cohorts. We now utilized next-generation sequencing to overcome these limitations and screened all DCM disease genes in a large cohort. Methods and results. In this multi-centre, multi-national study, we have enrolled 639 patients with sporadic or familial DCM. To all samples, we applied a standardized protocol for ultra-high coverage next-generation sequencing of 84 genes, leading to 99.1% coverage of the target region with at least 50-fold and a mean read depth of 2415. In this well characterized cohort, we find the highest number of known cardiomyopathy mutations in plakophilin-2, myosin-binding protein C-3, and desmoplakin. When we include yet unknown but predicted disease variants, we find titin, plakophilin-2, myosin-binding protein-C 3, desmoplakin, ryanodine receptor 2, desmocollin-2, desmoglein-2, and SCN5A variants among the most commonly mutated genes. The overlap between DCM, hypertrophic cardiomyopathy (HCM), and channelopathy causing mutations is considerably high. Of note, we find that >38% of patients have compound or combined mutations and 12.8% have three or even more mutations. When comparing patients recruited in the eight participating European countries we find remarkably little differences in mutation frequencies and affected genes. Conclusion. This is to our knowledge, the first study that comprehensively investigated the genetics of DCM in a large-scale cohort and across a broad gene panel of the known DCM genes. Our results underline the high analytical quality and feasibility of Next-Generation Sequencing in clinical genetic diagnostics and provide a sound database of the genetic causes of DCM.Hôpitaux de Paris; PHRC AOM0414

The Complete Genome Sequence of Thermoproteus tenax: A Physiologically Versatile Member of the Crenarchaeota

Here, we report on the complete genome sequence of the hyperthermophilic Crenarchaeum Thermoproteus tenax (strain Kra 1, DSM 2078(T)) a type strain of the crenarchaeotal order Thermoproteales. Its circular 1.84-megabase genome harbors no extrachromosomal elements and 2,051 open reading frames are identified, covering 90.6% of the complete sequence, which represents a high coding density. Derived from the gene content, T. tenax is a representative member of the Crenarchaeota. The organism is strictly anaerobic and sulfur-dependent with optimal growth at 86 degrees C and pH 5.6. One particular feature is the great metabolic versatility, which is not accompanied by a distinct increase of genome size or information density as compared to other Crenarchaeota. T. tenax is able to grow chemolithoautotrophically (CO2/H-2) as well as chemoorganoheterotrophically in presence of various organic substrates. All pathways for synthesizing the 20 proteinogenic amino acids are present. In addition, two presumably complete gene sets for NADH:quinone oxidoreductase (complex I) were identified in the genome and there is evidence that either NADH or reduced ferredoxin might serve as electron donor. Beside the typical archaeal A(0)A(1)-ATP synthase, a membrane-bound pyrophosphatase is found, which might contribute to energy conservation. Surprisingly, all genes required for dissimilatory sulfate reduction are present, which is confirmed by growth experiments. Mentionable is furthermore, the presence of two proteins (ParA family ATPase, actin-like protein) that might be involved in cell division in Thermoproteales, where the ESCRT system is absent, and of genes involved in genetic competence (DprA, ComF) that is so far unique within Archaea