Highly sensitive imaging of renal microcirculation in vivo using ultrahigh sensitive optical microangiography

AbstractThe electrophysiological activities in the human body generate electric and magnetic fields that can be measured noninvasively by electrodes on the skin, or even, not requiring any contact, by magnetometers. This includes the measurement of electrical activity of brain, heart, muscles and nerves that can be measured in vivo and allows to analyze functional processes with high temporal resolution. To measure these extremely small magnetic biosignals, traditionally highly sensitive superconducting quantum-interference devices have been used, together with advanced magnetic shields. Recently, they have been complemented in usability by a new class of sensors, optically pumped magnetometers (OPMs). These quantum sensors offer a high sensitivity without requiring cryogenic temperatures, allowing the design of small and flexible sensors for clinical applications. In this letter, we describe the advantages of these upcoming OPMs in two exemplary applications that were recently carried out at Physikalisch-Technische Bundesanstalt (PTB): (1) magnetocardiography (MCG) recorded during exercise and (2) auditory-evoked fields registered by magnetoencephalography.

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Real-Time In Vivo Bioluminescent Imaging for Evaluating the Efficacy of Antibiotics in a Rat Staphylococcus aureus Endocarditis Model

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.49.1.380-387.2005 ◽

2005 ◽

Vol 49 (1) ◽

pp. 380-387 ◽

Cited By ~ 68

Author(s):

Yan Q. Xiong ◽

Julie Willard ◽

Jagath L. Kadurugamuwa ◽

Jun Yu ◽

Kevin P. Francis ◽

...

Keyword(s):

Staphylococcus Aureus ◽

Real Time ◽

Imaging System ◽

Bioluminescent Imaging ◽

Vancomycin Therapy ◽

Treatment Groups ◽

Highly Sensitive ◽

Relevant Animal Model ◽

And Control

ABSTRACT Therapeutic options for invasive Staphylococcus aureus infections have become limited due to rising antimicrobial resistance, making relevant animal model testing of new candidate agents more crucial than ever. In the present studies, a rat model of aortic infective endocarditis (IE) caused by a bioluminescently engineered, biofilm-positive S. aureus strain was used to evaluate real-time antibiotic efficacy directly. This strain was vancomycin and cefazolin susceptible but gentamicin resistant. Bioluminescence was detected and quantified daily in antibiotic-treated and control animals with IE, using a highly sensitive in vivo imaging system (IVIS). Persistent and increasing cardiac bioluminescent signals (BLS) were observed in untreated animals. Three days of vancomycin therapy caused significant reductions in both cardiac BLS (>10-fold versus control) and S. aureus densities in cardiac vegetations (P < 0.005 versus control). However, 3 days after discontinuation of vancomycin therapy, a greater than threefold increase in cardiac BLS was observed, indicating relapsing IE (which was confirmed by quantitative culture). Cefazolin resulted in modest decreases in cardiac BLS and bacterial densities. These microbiologic and cardiac BLS differences during therapy correlated with a longer time-above-MIC for vancomycin (>12 h) than for cefazolin (∼4 h). Gentamicin caused neither a reduction in cardiac S. aureus densities nor a reduction in BLS. There were significant correlations between cardiac BLS and S. aureus densities in vegetations in all treatment groups. These data suggest that bioluminescent imaging provides a substantial advance in the real-time monitoring of the efficacy of therapy of invasive S. aureus infections in live animals.

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Simulation and Analysis of Highly Sensitive MEMS Cantilever Designs for “in vivo Label Free” Biosensing

Procedia Technology ◽

10.1016/j.protcy.2014.08.012 ◽

2014 ◽

Vol 14 ◽

pp. 85-92 ◽

Cited By ~ 7

Author(s):

Deep Kishore Parsediya ◽

Jawar Singh ◽

Pavan Kumar Kankar

Keyword(s):

Label Free ◽

Highly Sensitive

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DNAJB6b is Downregulated in Synucleinopathies

Journal of Parkinson s Disease ◽

10.3233/jpd-202512 ◽

2021 ◽

pp. 1-13

Author(s):

Jonas Folke ◽

Sertan Arkan ◽

Isak Martinsson ◽

Susana Aznar ◽

Gunnar Gouras ◽

...

Keyword(s):

Neurodegenerative Diseases ◽

Substantia Nigra Pars Compacta ◽

Endogenous Protein ◽

Highly Sensitive ◽

Brain Material ◽

Health And Disease ◽

Isoform B

Background: α-synuclein (α-syn) aggregation contributes to the progression of multiple neurodegenerative diseases. We recently found that the isoform b of the co-chaperone DNAJB6 is a strong suppressor of a-syn aggregation in vivo and in vitro. However, nothing is known about the role of the endogenous isoform b of DNAJB6 (DNAJB6b) in health and disease, due to lack of specific antibodies. Objective: Here we generated a novel anti-DNAJB6b antibody to analyze the localization and expression this isoform in cells, in tissue and in clinical material. Methods: To address this we used immunocytochemistry, immunohistochemistry, as well as a novel quantitative DNAJB6 specific ELISA method. Results: The endogenous protein is mainly expressed in the cytoplasm and in neurites in vitro, where it is found more in dendrites than in axons. We further verified in vivo that DNAJB6b is expressed in the dopaminergic neurons of the substantia nigra pars compacta (SNpc), which is a neuronal subpopulation highly sensitive to α-syn aggregation, that degenerate to a large extend in patients with Parkinson’s disease (PD) and multiple system atrophy (MSA). When we analyzed the expression levels of DNAJB6b in brain material from PD and MSA patients, we found a downregulation of DNAJB6b by use of ELISA based quantification. Interestingly, this was also true when analyzing tissue from patients with progressive supranuclear palsy, a taupathic atypical parkinsonian disorder. However, the total level of DNAJB6 was upregulated in these three diseases, which may indicate an upregulation of the other major isoform of DNAJB6, DNAJB6a. Conclusion: This study shows that DNAJB6b is downregulated in several different neurodegenerative diseases, which makes it an interesting target to further investigate in relation to amyloid protein aggregation and disease progression.

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Analysis of free and protein-bound nitrotyrosine in human plasma by a gas chromatography/mass spectrometry method that avoids nitration artifacts

Biochemical Journal ◽

10.1042/bj3450453 ◽

2000 ◽

Vol 345 (3) ◽

pp. 453-458 ◽

Cited By ~ 49

Author(s):

Matthew T. FROST ◽

Barry HALLIWELL ◽

Kevin P. MOORE

Keyword(s):

Reactive Nitrogen Species ◽

Plasma Proteins ◽

Biological Fluids ◽

Plasma Concentrations ◽

Normal Subjects ◽

Gas Chromatography Mass Spectrometry ◽

Highly Sensitive ◽

Acidic Conditions ◽

First Time

Measurement of nitrotyrosine in biological fluids and tissues is increasingly being used to monitor the production of reactive nitrogen species in vivo. The detection of nitrotyrosine in vivo has been reported with the use of a variety of methods including immunoassay, HPLC and GLC/MS. The validity of HPLC and immunoassays have been questioned with regard to their selectivity and sensitivity limits. In principle, the measurement of nitrotyrosine by GLC/MS permits a highly specific, highly sensitive and fully quantitative assay. The nitration of tyrosine under acidic conditions in the presence of nitrite is well documented. Derivatization for the full quantification of nitrotyrosine by using GLC/MS can lead to the artifactual nitration of tyrosine if performed under acidic conditions in the presence of nitrite. We describe a novel alkaline method for the hydrolysis and derivatization of nitrotyrosine and tyrosine, and demonstrate its applicability to the measurement of plasma concentrations of both free and protein-bound nitrotyrosine and tyrosine. A detection limit of 1 pg for nitrotyrosine and 100 pg for tyrosine has been achieved. Our method allows, for the first time, the analysis of free and protein-bound nitrotyrosine and tyrosine in biological samples. The plasma concentrations (means±S.E.M.) of free tyrosine and nitrotyrosine in eight normal subjects were 12±0.6 μg/ml and 14±0.7 ng/ml respectively. Plasma proteins contained tyrosine and nitrotyrosine at 60.7±1.7 μg/mg and 2.7±0.4 ng/mg respectively.

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PGRMC1-dependent lipophagy promotes ferroptosis in paclitaxel-tolerant persister cancer cells

Journal of Experimental & Clinical Cancer Research ◽

10.1186/s13046-021-02168-2 ◽

2021 ◽

Vol 40 (1) ◽

Author(s):

Ji Hyeon You ◽

Jaewang Lee ◽

Jong-Lyel Roh

Keyword(s):

Cancer Cells ◽

Lipid Droplets ◽

Tumor Xenograft ◽

Acid Oxidation ◽

Mouse Tumor ◽

Autophagy Induction ◽

Highly Sensitive ◽

Xenograft Models

Abstract Background Progesterone receptor membrane component 1 (PGRMC1) is a heme-binding protein inducing dimerization with cytochrome P450, which mediates chemoresistance. Increased PGRMC1 expression is found in multiple types of resistant cancers, but the role of PGRMC1 in the ferroptosis of cancer cells remains unrevealed. Therefore, we examined the role of PGRMC1 in promoting ferroptosis in paclitaxel-tolerant persister cancer cells (PCC). Methods The effects of ferroptosis inducers and PGRMC1 gene silencing/overexpression were tested on head and neck cancer (HNC) cell lines and mouse tumor xenograft models. The results were analyzed about cell viability, death, lipid ROS and iron production, mRNA/protein expression and interaction, and lipid assays. Results PCC had more free fatty acids, lipid droplets, and fatty acid oxidation (FAO) than their parental cells. PCC was highly sensitive to inhibitors of system xc− cystine/glutamate antiporter (xCT), such as erastin, sulfasalazine, and cyst(e)ine deprivation, but less sensitive to (1S,3R)-RSL3. PGRMC1 silencing in PCC reduced ferroptosis sensitivity by xCT inhibitors, and PGRMC1 overexpression in parental cells increased ferroptosis by xCT inhibitors. Lipid droplets were degraded along with autophagy induction and autophagosome formation by erastin treatment in PCC. Lipophagy was accompanied by increased tubulin detyrosination, which was increased by SIRT1 activation but decreased by SIRT1 inhibition. FAO and lipophagy were also promoted by the interaction between lipid droplets and mitochondria. Conclusion PGRMC1 expression increased FAO and ferroptosis sensitivity from in vivo mice experiments. Our data suggest that PGRMC1 promotes ferroptosis by xCT inhibition in PCC.

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Click-chemistry enabled directed evolution of glycosynthases for bespoke glycans synthesis

10.1101/2020.03.23.001982 ◽

2020 ◽

Cited By ~ 1

Author(s):

Ayushi Agrawal ◽

Chandra Kanth Bandi ◽

Tucker Burgin ◽

Youngwoo Woo ◽

Heather B. Mayes ◽

...

Keyword(s):

Single Cell ◽

Click Chemistry ◽

Directed Evolution ◽

Cell Sorting ◽

Screening Methods ◽

Carbohydrate Active Enzymes ◽

E Coli ◽

Highly Sensitive ◽

Increased Activity

AbstractEngineering of carbohydrate-active enzymes like glycosynthases for chemoenzymatic synthesis of bespoke oligosaccharides has been limited by the lack of suitable directed evolution based protein engineering methods. Currently there are no ultrahigh-throughput screening methods available for rapid and highly sensitive single cell-based screening of evolved glycosynthase enzymes employing azido sugars as substrates. Here, we report a fluorescence-based approach employing click-chemistry for the selective detection of glycosyl azides (versus free inorganic azides) that facilitated ultrahigh-throughput in-vivo single cell-based assay of glycosynthase activity. This discovery has led to the development of a directed evolution methodology for screening and sorting glycosynthase mutants for synthesis of desired fucosylated oligosaccharides. Our screening technique facilitated rapid fluorescence activated cell sorting of a large library of glycosynthase variants (>106 mutants) expressed in E. coli to identify several novel mutants with increased activity for β-fucosyl-azide activated donor sugars towards desired acceptor sugars, demonstrating the broader applicability of this methodology.

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