Bone marrow chimeric mice reveal a dual role for CD36 in Plasmodium berghei ANKA infection

BACKGROUND: Adhesion of Plasmodium-infected red blood cells (iRBC) to different host cells, ranging from endothelial to red blood cells, is associated to malaria pathology. In vitro studies have shown the relevance of CD36 for adhesion phenotypes of Plasmodium falciparum iRBC such as sequestration, platelet mediated clumping and non-opsonic uptake of iRBC. Different adhesion phenotypes involve different host cells and are associated with different pathological outcomes of disease. Studies with different human populations with CD36 polymorphisms failed to attribute a clear role to CD36 expression in human malaria. Up to the present, no in vivo model has been available to study the relevance of different CD36 adhesion phenotypes to the pathological course of Plasmodium infection. METHODS: Using CD36-deficient mice and their control littermates, CD36 bone marrow chimeric mice, expressing CD36 exclusively in haematopoietic cells or in non-haematopoietic cells, were generated. Irradiated CD36(-/- )and wild type mice were also reconstituted with syngeneic cells to control for the effects of irradiation. The reconstituted mice were infected with Plasmodium berghei ANKA and analysed for the development of blood parasitaemia and neurological symptoms. RESULTS: All mice reconstituted with syngeneic bone marrow cells as well as chimeric mice expressing CD36 exclusively in non-haematopoietic cells died from experimental cerebral malaria between day 6 and 12 after infection. A significant proportion of chimeric mice expressing CD36 only in haematopoietic cells did not die from cerebral malaria. CONCLUSION: The analysis of bone marrow chimeric mice reveals a dual role of CD36 in P. berghei ANKA infection. Expression of CD36 in haematopoietic cells, most likely macrophages and dendritic cells, has a beneficial effect that is masked in normal mice by adverse effects of CD36 expression in non-haematopoietic cells, most likely endothelial cells

Infection by and protective immune responses against Plasmodium berghei ANKA are not affected in macrophage scavenger receptors A deficient mice

BACKGROUND: Scavenger receptors (SRs) recognize endogenous molecules modified by pathological processes as well as components of diverse microorganisms. Mice deficient for both SR-AI and II are more susceptible to infections by a variety of bacterial and viral pathogens. RESULTS: Here we show that SR-A deficient mice and wild type mice are equally susceptible to malaria infection both during liver and blood stages. Moreover, like wild type mice, SR-A deficient mice are able to mount a protective immune response against radiation attenuated sporozoites. CONCLUSION: Our results do not reveal a function of SR-A I and II receptors in the Plasmodium berghei ANKA infection, both in the development of CM and parasitemia control. Moreover, these receptors appear not to be required for the establishment of a protective immune response against the malaria liver stages

A 3D printable adapter for solid-state fluorescence measurements: the case of an immobilized enzymatic bioreceptor for organophosphate pesticides detection

The widespread use of pesticides in the last decades and their accumulation into the environment gave rise to major environmental and human health concerns. To address this topic, the scientific community pointed out the need to develop methodologies to detect and measure the presence of pesticides in different matrices. Biosensors have been recently explored as fast, easy, and sensitive methods for direct organophosphate pesticides monitoring. Thus, the present work aimed at designing and testing a 3D printed adapter useful on different equipment, and a membrane support to immobilize the esterase-2 from Alicyclobacillus acidocaldarius (EST2) bioreceptor. The latter is labelled with the IAEDANS, a bright fluorescent probe. EST2 was selected since it shows a high specificity toward paraoxon. Our results showed good stability and replicability, with an increasing linear fluorescent intensity recorded from 15 to 150 pmol of labelled EST2. Linearity of data was also observed when using the immobilized labelled EST2 to detect increasing amounts of paraoxon, with a limit of detection (LOD) of 0.09 pmol. This LOD value reveals the high sensitivity of our membrane support when mounted on the 3D adapter, comparable to modern methods using robotic workstations. Notably, the use of an independent support significantly simplified the manipulation of the membrane during experimental procedures and enabled it to match the specificities of different systems. In sum, this work emphasizes the advantages of using 3D printed accessories adapted to respond to the newest research needs. Graphical abstract: [Figure not available: see fulltext.

A FRET Approach to Detect Paraoxon among Organophosphate Pesticides Using a Fluorescent Biosensor

The development of faster, sensitive and real-time methods for detecting organophosphate (OP) pesticides is of utmost priority in the in situ monitoring of these widespread compounds. Research on enzyme-based biosensors is increasing, and a promising candidate as a bioreceptor is the thermostable enzyme esterase-2 from Alicyclobacillus acidocaldarius (EST2), with a lipase-like Ser–His–Asp catalytic triad with a high affinity for OPs. This study aimed to evaluate the applicability of Förster resonance energy transfer (FRET) as a sensitive and reliable method to quantify OPs at environmentally relevant concentrations. For this purpose, the previously developed IAEDANS-labelled EST2-S35C mutant was used, in which tryptophan and IAEDANS fluorophores are the donor and the acceptor, respectively. Fluorometric measurements showed linearity with increased EST2-S35C concentrations. No significant interference was observed in the FRET measurements due to changes in the pH of the medium or the addition of other organic components (glucose, ascorbic acid or yeast extract). Fluorescence quenching due to the presence of paraoxon was observed at concentrations as low as 2 nM, which are considered harmful for the ecosystem. These results pave the way for further experiments encompassing more complex matrices

Point-of-care diagnostics of covid-19: From current work to future perspectives

Coronaviruses have received global concern since 2003, when an outbreak caused by SARS‐CoV emerged in China. Later on, in 2012, the Middle‐East respiratory syndrome spread in Saudi Arabia, caused by MERS‐CoV. Currently, the global crisis is caused by the pandemic SARS‐ CoV‐2, which belongs to the same lineage of SARS‐CoV. In response to the urgent need of diagnostic tools, several lab‐based and biosensing techniques have been proposed so far. Five main areas have been individuated and discussed in terms of their strengths and weaknesses. The cell‐culture detection and the microneutralization tests are still considered highly reliable methods. The genetic screening, featuring the well‐established Real‐time polymerase chain reaction (RT‐PCR), represents the gold standard for virus detection in nasopharyngeal swabs. On the other side, immunoassays were developed, either by screening/antigen recognition of IgM/IgG or by detecting the whole virus, in blood and sera. Next, proteomic mass‐spectrometry (MS)‐based methodologies have also been proposed for the analysis of swab samples. Finally, virus-biosensing devices were efficiently designed. Both electrochemical immunosensors and eye‐based technologies have been described, showing detection times lower than 10 min after swab introduction. Alternative to swab‐based techniques, lateral flow point‐of‐care immunoassays are already commercially available for the analysis of blood samples. Such biosensing devices hold the advantage of being portable for on‐site testing in hospitals, airports, and hotspots, virtually without any sample treatment or complicated lab precautions

Altered Expression of Protamine-like and Their DNA Binding Induced by Cr(VI): A Possible Risk to Spermatogenesis?

Chromium (VI) is the most dangerous oxidation state among the stable forms of chromium. In this work, we evaluated the effect of exposing Mytilus galloprovincialis for 24 h to 1, 10, and 100 nM chromium (VI) on the properties of Protamine-like (PLs) and their gene levels in the gonads. Specifically, we analyzed, by AU-PAGE and SDS-PAGE, PLs extracted from unexposed and exposed mussels. In addition, via EMSA, we evaluated the ability of PLs to bind DNA and also verified their potential to protect DNA from oxidative damage. Finally, we assessed possible alterations in gonadal expression of mt10, hsp70, and genes encoding for PLs-II/PL-IV and PL-III. We found that for all experimental approaches the most relevant alterations occurred after exposure to 1 nM Cr(VI). In particular, a comigration of PL-II with PL-III was observed by SDS-PAGE; and a reduced ability of PLs to bind and protect DNA from oxidative damage was recorded. This dose of chromium (VI) exposure was also the one that produced the greatest alterations in the expression of both mt10 and PL-II/PL-IV encoding genes. All of these changes suggest that this dose of chromium (VI) exposure could affect the reproductive health of Mytilus galloprovincialis

PKR is not obligatory for high-fat diet-induced obesity and its associated metabolic and inflammatory complications

Protein kinase R (PKR) has previously been suggested to mediate many of the deleterious consequences of a high-fat diet (HFD). However, previous studies have observed substantial phenotypic variability when examining the metabolic consequences of PKR deletion. Accordingly, herein, we have re-examined the role of PKR in the development of obesity and its associated metabolic complications in vivo as well as its putative lipid-sensing role in vitro. Here we show that the deletion of PKR does not affect HFD-induced obesity, hepatic steatosis or glucose metabolism, and only modestly affects adipose tissue inflammation. Treatment with the saturated fatty acid palmitate in vitro induced comparable levels of inflammation in WT and PKR KO macrophages, demonstrating that PKR is not necessary for the sensing of pro-inflammatory lipids. These results challenge the proposed role for PKR in obesity, its associated metabolic complications and its role in lipid-induced inflammation

Interleukin-6 Attenuates Insulin-Mediated Increases in Endothelial Cell Signaling but Augments Skeletal Muscle Insulin Action via Differential Effects on Tumor Necrosis Factor-α Expression

OBJECTIVE: The cytokine interleukin-6 (IL-6) stimulates AMP-activated protein kinase (AMPK) and insulin signaling in skeletal muscle, both of which result in the activation of endothelial nitric oxide synthase (eNOS). We hypothesized that IL-6 promotes endothelial cell signaling and capillary recruitment in vivo, contributing to increased glucose uptake. RESEARCH DESIGN AND METHODS: The effect of IL-6 with and without insulin on AMPK, insulin, and eNOS signaling in and nitric oxide (NO) release from human aortic endothelial cells (HAECs) was examined. The physiological significance of these in vitro signaling events was assessed by measuring capillary recruitment in rats during control and euglycemic-hyperinsulinemic clamps with or without IL-6 infusion. RESULTS: IL-6 blunted increases in insulin signaling, eNOS phosphorylation (Ser1177), and NO production and reduced phosphorylation of AMPK in HAEC in vitro and capillary recruitment in vivo. In contrast, IL-6 increased Akt phosphorylation (Ser473) in hindlimb skeletal muscle and enhanced whole-body glucose disappearance and glucose uptake during the clamp. The differences in endothelial cell and skeletal muscle signaling were mediated by the cell-specific, additive effects of IL-6 and insulin because this treatment markedly increased tumor necrosis factor (TNF)-alpha protein expression in HAECs without any effect on TNF-alpha in skeletal muscle. When HAECs were incubated with a TNF-alpha-neutralizing antibody, the negative effects of IL-6 on eNOS signaling were abolished. CONCLUSIONS: In the presence of insulin, IL-6 contributes to aberrant endothelial cell signaling because of increased TNF-alpha expression

Redox stress proteins are involved in adaptation response of the hyperthermoacidophilic archaeon Sulfolobus solfataricus to nickel challenge

<p>Abstract</p> <p>Background</p> <p>Exposure to nickel (Ni) and its chemical derivatives has been associated with severe health effects in human. On the contrary, poor knowledge has been acquired on target physiological processes or molecular mechanisms of this metal in model organisms, including Bacteria and Archaea. In this study, we describe an analysis focused at identifying proteins involved in the recovery of the archaeon <it>Sulfolobus solfataricus </it>strain MT4 from Ni-induced stress.</p> <p>Results</p> <p>To this purpose, <it>Sulfolobus solfataricus </it>was grown in the presence of the highest nickel sulphate concentration still allowing cells to survive; crude extracts from treated and untreated cells were compared at the proteome level by using a bi-dimensional chromatography approach. We identified several proteins specifically repressed or induced as result of Ni treatment. Observed up-regulated proteins were largely endowed with the ability to trigger recovery from oxidative and osmotic stress in other biological systems. It is noteworthy that most of the proteins induced following Ni treatment perform similar functions and a few have eukaryal homologue counterparts.</p> <p>Conclusion</p> <p>These findings suggest a series of preferential gene expression pathways activated in adaptation response to metal challenge.</p

Activation of AMP-Activated Protein Kinase by Interleukin-6 in Rat Skeletal Muscle: Association With Changes in cAMP, Energy State, and Endogenous Fuel Mobilization

OBJECTIVE: Interleukin-6 (IL-6) directly activates AMP-activated protein kinase (AMPK) in vivo and in vitro; however, the mechanism by which it does so is unknown. RESEARCH DESIGN AND METHODS: We examined this question in skeletal muscle using an incubated rat extensor digitorum longus (EDL) muscle preparation as a tool. RESULTS: AMPK activation by IL-6 coincided temporally with a nearly threefold increase in the AMP:ATP ratio in the EDL. The effects of IL-6 on both AMPK activity and energy state were inhibited by coincubation with propranolol, suggesting involvement of β-adrenergic signaling. In keeping with this notion, IL-6 concurrently induced a transient increase in cAMP, and its ability to activate AMPK was blocked by the adenyl cyclase inhibitor 2′5′-dideoxyadenosine. In addition, like other β-adrenergic stimuli, IL-6 increased glycogen breakdown and lipolysis in the EDL. Similar effects of IL-6 on AMPK, energy state, and cAMP content were observed in C2C12 myotubes and gastrocnemius muscle in vivo, indicating that they were not unique to the incubated EDL. CONCLUSIONS: These studies demonstrate that IL-6 activates AMPK in skeletal muscle by increasing the concentration of cAMP and, secondarily, the AMP:ATP ratio. They also suggest that substantial increases in IL-6 concentrations, such as those that can result from its synthesis by muscles during exercise, may play a role in the mobilization of fuel stores within skeletal muscle as an added means of restoring energy balance.United States Public Health Service (RO1DK19514, RO1DK067509); Ruth L. Kirschstein NRSA Postdoctoral Training Grant (HL-07224); Fonds de la Recherche en Santé du Québe