All-Optical Wide-Field Selective Imaging of Fluorescent Nanodiamonds in Cells, In Vivo and Ex Vivo

BackgroundNoninvasive intraoperative optical biopsy that provides real-time imaging of histoarchitectural (cell resolution) features of brain tumors, especially at the margin of invasive tumors, would be of great value. To assess clinical-grade confocal laser endomicroscopy (CLE) and to prepare for its use intraoperatively in vivo, we performed an assessment of CLE ex vivo imaging in brain lesions.MethodsTissue samples from patients who underwent intracranial surgeries with fluorescein sodium (FNa)–based wide-field fluorescence guidance were acquired for immediate intraoperative ex vivo optical biopsies with CLE. Hematoxylin-eosin–stained frozen section analysis of the same specimens served as the gold standard for blinded neuropathology comparison. FNa 2 to 5 mg/kg was administered upon induction of anesthesia, and FNa 5 mg/kg was injected for CLE contrast improvement. Histologic features were identified, and the diagnostic accuracy of CLE was assessed.ResultsOf 77 eligible patients, 47 patients with 122 biopsies were enrolled, including 32 patients with gliomas and 15 patients with other intracranial lesions. The positive predictive value of CLE optical biopsies was 97% for all specimens and 98% for gliomas. The specificity of CLE was 90% for all specimens and 94% for gliomas. The second FNa injection in seven patients, a mean of 2.6 h after the first injection, improved image quality and increased the percentage of accurately diagnosed images from 67% to 93%. Diagnostic CLE features of lesional glioma biopsies and normal brain were identified. Seventeen histologic features were identified.ConclusionsResults demonstrated high specificity and positive predictive value of ex vivo intraoperative CLE optical biopsies and justify an in vivo intraoperative trial. This new portable, noninvasive intraoperative imaging technique provides diagnostic features to discriminate lesional tissue with high specificity and is feasible for incorporation into the fluorescence-guided surgery workflow, particularly for patients with invasive brain tumors.

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Spatial light modulator based active wide-field illumination for ex vivo and in vivo quantitative NIR FRET imaging

Biomedical Optics Express ◽

10.1364/boe.5.000944 ◽

2014 ◽

Vol 5 (3) ◽

pp. 944 ◽

Cited By ~ 27

Author(s):

Lingling Zhao ◽

Ken Abe ◽

Shilpi Rajoria ◽

Qi Pian ◽

Margarida Barroso ◽

...

Keyword(s):

Spatial Light Modulator ◽

Ex Vivo ◽

Wide Field ◽

Light Modulator

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Active Wide-field Illumination For In Vitro, Ex Vivo and In Vivo NIR FRET Imaging

Biomedical Optics 2014 ◽

10.1364/biomed.2014.bw3b.5 ◽

2014 ◽

Cited By ~ 1

Author(s):

Lingling Zhao ◽

Shilpi Rajoria ◽

Margarida Barroso ◽

Xavier Intes

Keyword(s):

Ex Vivo ◽

Wide Field

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High-resolution wide-field human brain tumor margin detection and in vivo murine neuroimaging

10.1101/252080 ◽

2018 ◽

Cited By ~ 2

Author(s):

Derek Yecies ◽

Orly Liba ◽

Elliott SoRelle ◽

Rebecca Dutta ◽

Edwin Yuan ◽

...

Keyword(s):

Brain Tumor ◽

White Matter ◽

Ex Vivo ◽

Field Of View ◽

Low Grade ◽

Wide Field ◽

Tumor Margin ◽

Wide Field Of View ◽

Margin Detection

AbstractCurrent in vivo neuroimaging techniques provide limited field of view or spatial resolution and often require exogenous contrast. These limitations prohibit detailed structural imaging across wide fields of view and hinder intraoperative tumor margin detection. Here we present a novel neuroimaging technique, speckle-modulating optical coherence tomography (SM-OCT), which allows us to image the brains of live mice and ex vivo human samples with unprecedented resolution and wide field of view using only endogenous contrast. The increased effective resolution provided by speckle elimination reveals white matter fascicles and cortical layer architecture in the brains of live mice. To our knowledge, the data reported herein represents the highest resolution imaging of murine white matter structure achieved in vivo across a wide field of view of several millimeters. When applied to an orthotopic murine glioblastoma xenograft model, SM-OCT readily identifies brain tumor margins with near single-cell resolution. SM-OCT of ex vivo human temporal lobe tissue reveals fine structures including cortical layers and myelinated axons. Finally, when applied to an ex vivo sample of a low-grade glioma resection margin, SM-OCT is able to resolve the brain tumor margin. Based on these findings, SM-OCT represents a novel approach for intraoperative tumor margin detection and in vivo neuroimaging.

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The mouse lung early cellular innate immune response is not sufficient to control fungal infection with Cryptococcus neoformans

10.1101/679274 ◽

2019 ◽

Author(s):

Jacob Rudman ◽

Helen Maria Marriott ◽

Leo M. Carlin ◽

Simon Andrew Johnston

Keyword(s):

Immune Response ◽

Cryptococcus Neoformans ◽

Post Infection ◽

Opportunistic Infections ◽

Ex Vivo ◽

Mouse Lung ◽

Cell Mediated Immunity ◽

Wide Field ◽

Cryptococcal Infection

AbstractCryptococcus neoformanscauses life-threatening infection in the immunocompromised. This and other opportunistic pathogens are an increasing threat as immunosuppression increases globally. To counter antibiotic resistance, there is precedent for developing immune enhancing therapy. However, our understanding of how immunocompetent patients resolve these infections is poor as opportunistic infections typically resolve subclinically. Because this has led to a lack of clinical data, we rely on animal models. Currentin vivoinfection models either lack mammalian immunity or are not compatible with long term high content imaging required to model the complexities of human host-pathogen interactions. Therefore, we have developed anex vivomurine precision cut lung slice (PCLS) model to understand innate immunity in cryptococcosis. C57BL/6 mice were sacrificed 0 or 24 hours post infection withKN99αcryptococci. Lungs were inflated with 37°C agarose, 300μm thick PCLS were prepared on a vibratome and imaged by confocal or wide-field fluorescence microscopy. Using PCLS and immunofluorescence, we demonstrate cryptococcal replication and clearance rates are balanced over the first 24 hours of infection. Cell-mediated immunity is alveolar macrophage centric, although alveolar macrophages demonstrate limited phagocytosis of cryptococci and enable intracellular cryptococcal replication.Cryptococcus neoformansresponded to the lung environment by forming enlarged cells, although these were not large enough to be titan cells. To further understand cryptococcal proliferationin vivo, we also infected animals withplb1mutantCryptococcus neoformansthat has been shown to exhibit proliferation defectsin vivo. We found no difference in fungal burden withplb1infected animals 24 hours post infection, but observed significantly larger fungal cells and no incidences of phagocytosis. Thus, the PCLS model can be used to assess the lung immune response early in cryptococcal infection, demonstrating that resident lung macrophages cannot control cryptococcal infection and offer an intracellular niche forCryptococcus neoformansgrowth.

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Effect of Some Metabolic Inhibitors on Vinblastine-Induced Ribosomal-Granular Material Complexes

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100066796 ◽

1971 ◽

Vol 29 ◽

pp. 548-549

Author(s):

Awtar Krishan ◽

Dora Hsu

Keyword(s):

Granular Material ◽

Rna Synthesis ◽

Culture Medium ◽

Actinomycin D ◽

Metabolic Inhibitors ◽

Protein And Rna Synthesis ◽

Apparent Reduction ◽

Plant Alkaloids ◽

In Cells

Cells exposed to antitumor plant alkaloids, vinblastine and vincristine sulfate have large proteinacious crystals and complexes of ribosomes, helical polyribosomes and electron-dense granular material (ribosomal complexes) in their cytoplasm, Binding of H3-colchicine by the in vivo crystals shows that they contain microtubular proteins. Association of ribosomal complexes with the crystals suggests that these structures may be interrelated.In the present study cultured human leukemic lymphoblasts (CCRF-CEM), were incubated with protein and RNA-synthesis inhibitors, p. fluorophenylalanine, puromycin, cycloheximide or actinomycin-D before the addition of crystal-inducing doses of vinblastine to the culture medium. None of these compounds could completely prevent the formation of the ribosomal complexes or the crystals. However, in cells pre-incubated with puromycin, cycloheximide, or actinomycin-D, a reduction in the number and size of the ribosomal complexes was seen. Large helical polyribosomes were absent in the ribosomal complexes of cells treated with puromycin, while in cells exposed to cycloheximide, there was an apparent reduction in the number of ribosomes associated with the ribosomal complexes (Fig. 2).

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The Role of Polyphenols in the Modulation of Plasma Non-Enzymatic Antioxidant Capacity (NEAC)

International Journal for Vitamin and Nutrition Research ◽

10.1024/0300-9831/a000116 ◽

2012 ◽

Vol 82 (3) ◽

pp. 228-232 ◽

Cited By ~ 7

Author(s):

Mauro Serafini ◽

Giuseppa Morabito

Keyword(s):

Antioxidant Capacity ◽

Dietary Intervention ◽

Ex Vivo ◽

The Body ◽

Dietary Polyphenols ◽

Enzymatic Antioxidant ◽

Endogenous Antioxidant

Dietary polyphenols have been shown to scavenge free radicals, modulating cellular redox transcription factors in different in vitro and ex vivo models. Dietary intervention studies have shown that consumption of plant foods modulates plasma Non-Enzymatic Antioxidant Capacity (NEAC), a biomarker of the endogenous antioxidant network, in human subjects. However, the identification of the molecules responsible for this effect are yet to be obtained and evidences of an antioxidant in vivo action of polyphenols are conflicting. There is a clear discrepancy between polyphenols (PP) concentration in body fluids and the extent of increase of plasma NEAC. The low degree of absorption and the extensive metabolism of PP within the body have raised questions about their contribution to the endogenous antioxidant network. This work will discuss the role of polyphenols from galenic preparation, food extracts, and selected dietary sources as modulators of plasma NEAC in humans.

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Kutane antioxidative Wirkung von Johanniskrautextrakt (Hypericum perforatum): In-vitro-, Ex-vivo- und In-vivo-Untersuchungen

Zeitschrift für Phytotherapie ◽

10.1055/s-0032-1313268 ◽

2012 ◽

Vol 33 (S 01) ◽

Author(s):

MC Meinke ◽

S Schanzer ◽

S Arndt ◽

A Kleemann ◽

J Lademann

Keyword(s):

Hypericum Perforatum ◽

Ex Vivo ◽

Antioxidative Wirkung

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Antithrombotic Effects and Bleeding Time Prolongation with Synthetic Platelet GPIIb/llla Inhibitors in Animal Models of Platelet-Mediated Thrombosis

Thrombosis and Haemostasis ◽

10.1055/s-0038-1642390 ◽

1994 ◽

Vol 71 (01) ◽

pp. 095-102 ◽

Cited By ~ 43

Author(s):

Désiré Collen ◽

Hua Rong Lu ◽

Jean-Marie Stassen ◽

Ingrid Vreys ◽

Tsunehiro Yasuda ◽

...

Keyword(s):

Platelet Aggregation ◽

Bleeding Time ◽

Bolus Injection ◽

Cell Injury ◽

Ex Vivo ◽

Thrombus Formation ◽

Femoral Vein ◽

Thrombotic Occlusion ◽

Antithrombotic Effects

SummaryCyclic Arg-Gly-Asp (RGD) containing synthetic peptides such as L-cysteine, N-(mercaptoacetyl)-D-tyrosyl-L-arginylglycyl-L-a-aspartyl-cyclic (1→5)-sulfide, 5-oxide (G4120) and acetyl-L-cysteinyl-L-asparaginyl-L-prolyl-L-arginyl-glycyl-L-α-aspartyl-[0-methyltyrosyl]-L-arginyl-L-cysteinamide, cyclic 1→9-sulfide (TP9201) bind with high affinity to the platelet GPIIb/IIIa receptor.The relationship between antithrombotic effect, ex vivo platelet aggregation and bleeding time prolongation with both agents was studied in hamsters with a standardized femoral vein endothelial cell injury predisposing to platelet-rich mural thrombosis, and in dogs with a carotid arterial eversion graft inserted in the femoral artery. Intravenous administration of G4120 in hamsters inhibited in vivo thrombus formation with a 50% inhibitory bolus dose (ID50) of approximately 20 μg/kg, ex vivo ADP-induccd platelet aggregation with ID50 of 10 μg/kg, and bolus injection of 1 mg/kg prolonged the bleeding time from 38 ± 9 to 1,100 ± 330 s. Administration of TP9201 in hamsters inhibited in vivo thrombus formation with ID50 of 30 μg/kg, ex vivo platelet aggregation with an ID50 of 50 μg/kg and bolus injection of 1 mg/kg did not prolong the template bleeding time. In the dog eversion graft model, infusion of 100 μg/kg of G4120 over 60 min did not fully inhibit platelet-mediated thrombotic occlusion but was associated with inhibition of ADP-induccd ex vivo platelet aggregation and with prolongation of the template bleeding time from 1.3 ± 0.4 to 12 ± 2 min. Infusion of 300 μg/kg of TP9201 over 60 min completely prevented thrombotic occlusion, inhibited ex vivo platelet aggregation, but was not associated with prolongation of the template bleeding time.TP9201, unlike G4120, inhibits in vivo platelet-mediated thrombus formation without associated prolongation of the template bleeding time.

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Significance of Platelet β-Adrenoceptors for Platelet Responses In Vivo and In Vitro

Thrombosis and Haemostasis ◽

10.1055/s-0038-1646345 ◽

1992 ◽

Vol 68 (06) ◽

pp. 687-693 ◽

Cited By ~ 24

Author(s):

P T Larsson ◽

N H Wallén ◽

A Martinsson ◽

N Egberg ◽

P Hjemdahl

Keyword(s):

Ex Vivo ◽

High Dose ◽

Platelet Aggregability ◽

Adrenoceptor Agonist ◽

Dose Infusion ◽

Von Willebrand ◽

Willebrand Factor ◽

Factor Antigen

SummaryThe significance of platelet β-adrenoceptors for platelet responses to adrenergic stimuli in vivo and in vitro was studied in healthy volunteers. Low dose infusion of the β-adrenoceptor agonist isoprenaline decreased platelet aggregability in vivo as measured by ex vivo filtragometry. Infusion of adrenaline, a mixed α- and β-adrenoceptor agonist, increased platelet aggregability in vivo markedly, as measured by ex vivo filtragometry and plasma β-thromboglobulin levels. Adrenaline levels were 3–4 nM in venous plasma during infusion. Both adrenaline and high dose isoprenaline elevated plasma von Willebrand factor antigen levels β-Blockade by propranolol did not alter our measures of platelet aggregability at rest or during adrenaline infusions, but inhibited adrenaline-induced increases in vWf:ag. In a model using filtragometry to assess platelet aggregability in whole blood in vitro, propranolol enhanced the proaggregatory actions of 5 nM, but not of 10 nM adrenaline. The present data suggest that β-adrenoceptor stimulation can inhibit platelet function in vivo but that effects of adrenaline at high physiological concentrations are dominated by an α-adrenoceptor mediated proaggregatory action.

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