Confocal imaging of basal cell cancers in vivo and in thick skin excisions ex vivo

2002 ◽  
Author(s):  
Milind Rajadhyaksha ◽  
Peter J. Dwyer ◽  
Gregg M. Menaker ◽  
Thomas J. Flotte ◽  
James M. Zavislan ◽  
...  
Keyword(s):  
Basal Cell ◽  
Ex Vivo ◽  
Confocal Imaging ◽  
Thick Skin

Terahertz pulsed imaging of basal cell carcinoma ex vivo and in vivo

2004 ◽  
Vol 151 (2) ◽  
pp. 424-432 ◽  
Author(s):  
V.P. Wallace ◽  
A.J. Fitzgerald ◽  
S. Shankar ◽  
N. Flanagan ◽  
R. Pye ◽  
...  
Keyword(s):  
Basal Cell Carcinoma ◽  
Cell Carcinoma ◽  
Basal Cell ◽  
Ex Vivo ◽  
Terahertz Pulsed Imaging

scholarly journals Modeling Immune Checkpoint Inhibitor Efficacy in Syngeneic Mouse Tumors in an Ex Vivo Immuno-Oncology Dynamic Environment

2020 ◽  
Vol 21 (18) ◽  
pp. 6478
Author(s):  
Daniel T. Doty ◽  
Julia Schueler ◽  
Vienna L. Mott ◽  
Cassie M. Bryan ◽  
Nathan F. Moore ◽  
...  
Keyword(s):  
Real Time ◽  
Mouse Models ◽  
Immune Checkpoint ◽  
Checkpoint Inhibitors ◽  
Ex Vivo ◽  
Confocal Imaging ◽  
Checkpoint Blockade ◽  
Syngeneic Mouse

The immune checkpoint blockade represents a revolution in cancer therapy, with the potential to increase survival for many patients for whom current treatments are not effective. However, response rates to current immune checkpoint inhibitors vary widely between patients and different types of cancer, and the mechanisms underlying these varied responses are poorly understood. Insights into the antitumor activities of checkpoint inhibitors are often obtained using syngeneic mouse models, which provide an in vivo preclinical basis for predicting efficacy in human clinical trials. Efforts to establish in vitro syngeneic mouse equivalents, which could increase throughput and permit real-time evaluation of lymphocyte infiltration and tumor killing, have been hampered by difficulties in recapitulating the tumor microenvironment in laboratory systems. Here, we describe a multiplex in vitro system that overcomes many of the deficiencies seen in current static histocultures, which we applied to the evaluation of checkpoint blockade in tumors derived from syngeneic mouse models. Our system enables both precision-controlled perfusion across biopsied tumor fragments and the introduction of checkpoint-inhibited tumor-infiltrating lymphocytes in a single experiment. Through real-time high-resolution confocal imaging and analytics, we demonstrated excellent correlations between in vivo syngeneic mouse and in vitro tumor biopsy responses to checkpoint inhibitors, suggesting the use of this platform for higher throughput evaluation of checkpoint efficacy as a tool for drug development.

scholarly journals P397 Response to biologics in IBD patients assessed by Computerized image analysis of Probe Based Confocal Laser Endomicroscopy with molecular labeling

2021 ◽  
Vol 15 (Supplement_1) ◽  
pp. S406-S407
Author(s):  
M Iacucci ◽  
E Grisan ◽  
N Labarile ◽  
O Nardone ◽  
S C Smith ◽  
...  
Keyword(s):  
Predictive Value ◽  
Ex Vivo ◽  
Binding Activity ◽  
Biological Agent ◽  
Confocal Imaging ◽  
Confocal Laser Endomicroscopy ◽  
Confocal Laser ◽  
Tertiary Referral Centre ◽  
Fluorescent Intensity

Abstract Background The increase in therapeutic choices in inflammatory bowel diseases (IBD) imposed the identification of personalized therapeutic strategy. Confocal laser endomicroscopy (CLE) is a new endoscopic tool developed to obtain virtual in vivo histology. This study aimed to identify CLE in vivo and ex vivo features predictive of response for patients starting biologics. Methods We performed a prospective observational study at a single tertiary referral centre. 29 IBD patients (14 ulcerative colitis-UC and 15 Crohn’s Disease-CD) underwent colonoscopy with CLE before and 12-14 weeks after starting anti -TNF or 16 weeks after anti α4β7 integrin. CLE parameters analyzed were: crypt distribution along the mucosal surface, crypt area (CA), eccentricity, diameter, inter-cryptic distance (ICD), vessel tortuosity (VT), fluorescein leakage through the colonic mucosa (FLCM). Ex-vivo binding activity of fluorescein labelled biologics on corresponding biopsies was also assessed. Mosaicism reconstruction of CLE images were analyzed using a dedicated software algorithm (CellvizioViewer, Mauna Kea Technologies, ParisFrance). A dedicated Graphical User Interface was designed to enable a semiautomated analysis. Results After treatment, VT resulted significantly changed in overall IBD population (p<0.05), FLCM were reduced in UC patients (p<0.05), whilst CA, eccentricity and ICD decreased in CD patients (p< 0.05). The univariate regression analysis identified FLCM as the most accurate parameter for predicting responsiveness (AUROC) 83%, accuracy 83%, positive predictive value-PPV 94% and negative predictive value-NPV 57%). FLCM and ICD were the best discriminants in responders Vs non responders in UC patients (AUROC 85%, accuracy 85%, PPV 100% and NPV 71%); whilst VT, CA and ICD in CD patients (AUROC 95%, 86% and 83%; accuracy 90%, 90% and 88%; PPV 100%, 100% and 86%; and NPV 75%, 75% and 100%, respectively). Ex vivo confocal imaging revealed that UC patients, especially those responders, had higher basal fluorescent intensity signals with a significant reduction after biological treatment (p< 0.05), whereas in CD patients no significant change was found. An increased mucosal binding to the fluorescent labelled biological agent was associated to a higher likelihood of response to the treatment; this effect was higher in UC patients (AUROC 81%, accuracy 77%, PPV 100%, NPV 63%) compared to CD patients (AUROC 64%, accuracy 79%, PPV 80%, NPV 50%). Conclusion FLCM and ICD were the best discriminants of response in UC, while VT, CA and ICD in CD. A higher mucosal binding to a biological agent before treatment was observed in responders UC patients but not in CD patients.

Basal cell carcinoma: the utility of in vivo and ex vivo confocal microscopy

2018 ◽  
Vol 32 (12) ◽  
pp. 2090-2096 ◽  
Author(s):  
C. Longo ◽  
S. Borsari ◽  
R. Pampena ◽  
E. Benati ◽  
C. Bombonato ◽  
...  
Keyword(s):  
Confocal Microscopy ◽  
Basal Cell Carcinoma ◽  
Cell Carcinoma ◽  
Basal Cell ◽  
Ex Vivo

scholarly journals TRPML1 channels initiate Ca2+ sparks in vascular smooth muscle cells

2020 ◽  
Vol 13 (637) ◽  
pp. eaba1015 ◽  
Author(s):  
Pratish Thakore ◽  
Harry A. T. Pritchard ◽  
Caoimhin S. Griffin ◽  
Evan Yamasaki ◽  
Bernard T. Drumm ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Vascular Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Vascular Smooth Muscle Cells ◽  
Transient Receptor Potential ◽  
Ex Vivo ◽  
Ryanodine Receptors ◽  
Muscle Cells ◽  
Confocal Imaging

TRPML1 (transient receptor potential mucolipin 1) is a Ca2+-permeable, nonselective cation channel localized to the membranes of endosomes and lysosomes and is not present or functional on the plasma membrane. Ca2+ released from endosomes and lysosomes into the cytosol through TRPML1 channels is vital for trafficking, acidification, and other basic functions of these organelles. Here, we investigated the function of TRPML1 channels in fully differentiated contractile vascular smooth muscle cells (SMCs). In live-cell confocal imaging studies, we found that most endosomes and lysosomes in freshly isolated SMCs from cerebral arteries were essentially immobile. Using nanoscale super-resolution microscopy, we found that TRPML1 channels present in late endosomes and lysosomes formed stable complexes with type 2 ryanodine receptors (RyR2) on the sarcoplasmic reticulum (SR). Spontaneous Ca2+ signals resulting from the release of SR Ca2+ through RyR2s (“Ca2+ sparks”) and corresponding Ca2+-activated K+ channel activity are critically important for balancing vasoconstriction. We found that these signals were essentially absent in SMCs from TRPML1-knockout (Mcoln1−/−) mice. Using ex vivo pressure myography, we found that loss of this critical signaling cascade exaggerated the vasoconstrictor responses of cerebral and mesenteric resistance arteries. In vivo radiotelemetry studies showed that Mcoln1−/− mice were spontaneously hypertensive. We conclude that TRPML1 is crucial for the initiation of Ca2+ sparks in SMCs and the regulation of vascular contractility and blood pressure.

scholarly journals Confocal imaging of carbon dioxide laser-ablated basal cell carcinomas: An ex-vivo study on the uptake of contrast agent and ablation parameters

2015 ◽  
Vol 48 (2) ◽  
pp. 133-139 ◽  
Author(s):  
Heidy Sierra ◽  
Shadi Damanpour ◽  
Brian Hibler ◽  
Kishwer Nehal ◽  
Anthony Rossi ◽  
...  
Keyword(s):  
Carbon Dioxide ◽  
Contrast Agent ◽  
Basal Cell ◽  
Carbon Dioxide Laser ◽  
Ex Vivo ◽  
Confocal Imaging ◽  
Basal Cell Carcinomas ◽  
Ex Vivo Study

scholarly journals A crosscorrelation methodology for in vivo pharmacokinetic study by the trans-scale fluorescent system

2022 ◽  
Author(s):  
Yueqing Gu ◽  
Siwen Li ◽  
Qiao Lin ◽  
Yi Ma ◽  
Lu Qian ◽  
...  
Keyword(s):  
Pharmacokinetic Study ◽  
Time Course ◽  
Drug Disposition ◽  
Ex Vivo ◽  
Temporal Correlation ◽  
Confocal Imaging ◽  
Innovative Methodology ◽  
Micro Level ◽  
Spatio Temporal

Abstract Conventional single-organ-isolation-based pharmacokinetics study is short of time-course information and exists considerable inaccuracy due to the inter-individual differences and characteristic imparities between in vivo and ex vivo tissues/cells. The in vivo time-course and multi-organs study of model drugs in living subjects could afford precise spatio-temporal correlation. Herein, a revolutionized trans-dimensional fluorescence system was home built, with the macro-level detection part for simultaneous pharmacokinetic study in different organs, and one confocal imaging needle for micro-level visualizing cellular uptake of drugs with super-high resolution (0.472 μm). Correlating these simultaneous acquired trans-scale data, an innovative physiologically-based pharmacokinetics (PBPK) model was firstly created for predicting drug disposition in other species. Its accuracy and reliability was firmly supported by the high consistent predicted-data with the real-measured data in mice and in human, respectively. This study provides an innovative methodology and revolutionized instrument for in vivo real-time advancing assessment of druggability.

Imaging of oxygen gradients in giant umbrella cells: an ex vivo PLIM study

2015 ◽  
Vol 309 (7) ◽  
pp. C501-C509 ◽  
Author(s):  
A. V. Zhdanov ◽  
A. V. Golubeva ◽  
I. A. Okkelman ◽  
J. F. Cryan ◽  
D. B. Papkovsky
Keyword(s):  
Molecular Mechanisms ◽  
Ex Vivo ◽  
Hypoxia Inducible Factor ◽  
Confocal Imaging ◽  
Tissue Imaging ◽  
In Vivo Studies ◽  
Enzymatic Reactions ◽  
Bladder Epithelium ◽  
Umbrella Cells

O2 plays a pivotal role in aerobic metabolism and regulation of cell and tissue function. Local differences and fluctuations in tissue O2 levels are well documented; however, the physiological significance of O2 microgradients, particularly at the subcellular level, remains poorly understood. Using the cell-penetrating phosphorescent O2 probe Pt-Glc and confocal fluorescence microscopy, we visualized O2 distribution in individual giant (>100-μm) umbrella cells located superficially in the urinary bladder epithelium. We optimized conditions for in vivo phosphorescent staining of the inner surface of the mouse bladder and subsequent ex vivo analysis of excised live tissue. Imaging experiments revealed significant (≤85 μM) and heterogeneous deoxygenation within respiring umbrella cells, with radial O2 gradients of up to 40 μM across the cell, or ∼0.6 μM/μm. Deeply deoxygenated (5–15 μM O2) regions were seen to correspond to the areas enriched with polarized mitochondria. Pharmacological activation of mitochondrial respiration decreased oxygenation and O2 gradients in umbrella cells, while inhibition with antimycin A dissipated the gradients and caused gradual reoxygenation of the tissue to ambient levels. Detailed three-dimensional maps of O2 distribution potentially can be used for the modeling of intracellular O2-dependent enzymatic reactions and downstream processes, such as hypoxia-inducible factor signaling. Further ex vivo and in vivo studies on intracellular and tissue O2 gradients using confocal imaging can shed light on the molecular mechanisms regulating O2-dependent (patho)physiological processes in the bladder and other tissues.

scholarly journals Phenotypically distinct neutrophils patrol uninfected human and mouse lymph nodes

2019 ◽  
Vol 116 (38) ◽  
pp. 19083-19089 ◽  
Author(s):  
Laurence S. C. Lok ◽  
Thomas W. Dennison ◽  
Krishnaa M. Mahbubani ◽  
Kourosh Saeb-Parsy ◽  
Edwin R. Chilvers ◽  
...  
Keyword(s):  
T Cell ◽  
Lymph Nodes ◽  
T Cell Activation ◽  
Cell Activation ◽  
Ex Vivo ◽  
Confocal Imaging ◽  
High Endothelial Venules ◽  
Two Photon Microscopy ◽  
Routine Surveillance

Neutrophils play a key role in innate immunity. As the dominant circulating phagocyte, they are rapidly recruited from the bloodstream to sites of infection or injury to internalize and destroy microbes. More recently, neutrophils have been identified in uninfected organs, challenging the classical view of their function. Here we show that neutrophils were present in lymph nodes (LNs) in homeostasis. Using flow cytometry and confocal imaging, we identified neutrophils within LNs in naive, unchallenged mice, including LNs draining the skin, lungs, and gastrointestinal tract. Neutrophils were enriched within specific anatomical regions, in the interfollicular zone, a site of T cell activation. Intravital two-photon microscopy demonstrated that LN neutrophils were motile, trafficked into LNs from both blood and tissues via high endothelial venules and afferent lymphatics, respectively, and formed interactions with dendritic cells in LNs. Murine and human LN neutrophils had a distinct phenotype compared with circulating neutrophils, with higher major histocompatibility complex II (MHCII) expression, suggesting a potential role in CD4 T cell activation. Upon ex vivo stimulation with IgG immune complex (IC), neutrophils up-regulated expression of MHCII and costimulatory molecules and increased T cell activation. In vivo, neutrophils were capable of delivering circulating IC to LNs, suggesting a broader functional remit. Overall, our data challenge the perception that neutrophil patrol is limited to the circulation in homeostasis, adding LNs to their routine surveillance territory.

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