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Super Resolution and Endoscopic Two Photon Microscopy - Imaging of Cell Migration in Inflammation, Metastasis and Regeneration

English title Super Resolution and Endoscopic Two Photon Microscopy - Imaging of Cell Migration in Inflammation, Metastasis and Regeneration
Applicant Niess Jan Hendrik
Number 170809
Funding scheme R'EQUIP
Research institution Abt. für Gastroenterologie und Hepatologie Universitätsspital Basel
Institution of higher education University of Basel - BS
Main discipline Immunology, Immunopathology
Start/End 01.01.2017 - 31.12.2018
Approved amount 450'000.00
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All Disciplines (2)

Discipline
Immunology, Immunopathology
Experimental Cancer Research

Keywords (4)

two photon microscope; metastasis; cell migration; immunology

Lay Summary (German)

Lead
Zellmigration ist eine wichtiger Prozess bei Entstehung von Entzündung und Metastasen. Untersuchungen zur Zellmigration in lebenden Organismen ist schwierig, da Zellen klein sind, sich schnell bewegen und die zu untersuchenden Zellen tief im Gewebe liegen können. Zweiphotonenmikroskopie erlaubt dynamische Prozesse in lebenden Organismen zu untersuchen.
Lay summary

Zellmigration bedeutet die Bewegung der kleinsten lebenden Einheit eines Lebewesens, die ein bedeutender biologischer Prozess ist, der bei Entstehung von Entzündung, Absiedelung eines Tumors und Erholung von Krankheiten (Regeneration) eine wichtige Rolle hat. Die Untersuchung von Zellmigration in lebenden Organsimen ist eine Herausforderung für bildgebende Verfahren, da Zellen klein sind, Zellmigration ein dynamischer Prozess ist und die zu untersuchenden Zellen sich oft tief im Gewebe befinden. Die Zweiphotonenmikroskopie ist die führende Technologie, um Zellmigration in lebenden Organismen zu untersuchen, da diese fluoreszierenden Farbstoffe auf zellulärem Niveau tief in Geweben darzustellen vermag. In diesem Projekt beantragen wir ein neues Zweiphotonenmikroskop, welches eine hohe Auflösung und Untersuchungen in tiefliegenden Geweben besitzt. Die Integration des Mikroskops in die lokale Infrastruktur erlaubt, wiederholende Untersuchungen am gleichen Organismus durchzuführen. 

Direct link to Lay Summary Last update: 12.12.2016

Responsible applicant and co-applicants

Collaboration

Group / person Country
Types of collaboration
Christian Riedel, University of Ulm Germany (Europe)
- in-depth/constructive exchanges on approaches, methods or results
Valentin Djonov, Institute of Anatomy, Bern University Switzerland (Europe)
- in-depth/constructive exchanges on approaches, methods or results
Markus Affolter, Biozentrum, Basel University Switzerland (Europe)
- in-depth/constructive exchanges on approaches, methods or results
Walter Paul Weber, University Hospital Basel Switzerland (Europe)
- in-depth/constructive exchanges on approaches, methods or results

Associated projects

Number Title Start Funding scheme
163202 The molecular switch between normal and aberrant angiogenesis by VEGF 01.10.2015 Project funding (Div. I-III)
163938 Discovery of fundamental mechanisms underlying the development of cancer metastasis 01.04.2016 SNSF Professorships
182357 Decoding the molecular regulation of intussusceptive angiogenesis for therapeutic targeting 01.10.2018 Project funding (Div. I-III)
166428 Molecular mechanisms of breast tumorigenesis, metastasis, and resistance to therapy in tumors with a mutated PI3K pathway 01.12.2016 Project funding (Div. I-III)
158312 VEGF and Sema3A signaling for vascularized bone grafts 01.02.2015 Marie Heim-Voegtlin grants
146290 Investigation into the role of IL-19 producing macrophages in intestinal inflammation 01.12.2013 Project funding (Div. I-III)
175548 Intestinal macrophages and dendritic cells in oral tolerance and colitis 01.09.2018 Project funding (Div. I-III)
162575 Dissecting mechanisms of T cell dysfunction in patients with lung and ovarian cancer 01.05.2016 Project funding (Div. I-III)

Abstract

Cell migration plays an essential role in inflammation, metastasis, and regeneration. Studying cell migration in vivo challenges the limits of imaging because cells are small, migration is dynamic, and cells of interest are often located deep in the body or tissue. Two-photon microscopy has emerged as the leading technology to study cell migration in vivo because it can quickly image fluorescent reporters at cellular resolution and hundreds of microns deep in living mice. However, there are challenges to two-photon imaging that limit 1) image resolution 2) depth of imaging, 3) duration of experiment and 4) user friendliness. Here we propose a novel two-photon microscope that overcomes these limitations with the following features:1) Increased resolution via ‘Super-resolution Optical Fluctuation Imaging’ (SOFI). SOFI increases resolution four times beyond the physical limitations imposed by optics. This is of particular use for two-photon microscopy where the resolution is often compromised in order to image deeper. There are no commercially available two-photon microscopes offering the level of resolution proposed here, and access to such resolution before market availability is a competitive advantage.2) Extended depth of imaging via endoscopy. The depth of two-photon imaging is limited to the first few hundred microns of mouse tissues. Therefore invasive surgeries are often required to expose and image deeper organs. As a consequence, mice often have to be sacrificed after imaging. Equipping the two-photon microscope with an endoscope enables deep imaging with minimal invasion and thus allows repeated imaging of the same mouse during long term cell migration studies.3) Prolonged duration of experiment via mouse housing. Many in vivo studies require mouse sacrifice after two-photon imaging because hygiene laws prevent return of mice to housing. Mouse housing in the same room as the microscope enables repeated imaging of the same mouse, which is essential for long term cell migration studies in vivo. Furthermore, such mouse housing enables import of extramural mice for two-photon experiments, and thus facilitates collaboration and widespread use. Funding, design and designation of this room is already approved and organized by the DBM, independent of the R’Equip.4) Improved user friendliness via integration with Nikon. Custom designed and cutting-edge technology can often be user-unfriendly and therefore limit use. Historically, this has been the case for non-commercial two-photon microscopes which are unwieldy, e.g. requiring manual laser alignment. Integration of a custom designed super resolution and endoscopic two-photon with a commercial Nikon microscope creates an instrument that is both cutting-edge and highly usable. Nikon overcomes many of the previous usability challenges by automation, including laser alignment. Furthermore the Nikon two-photon microscope has several market-leading and patented features such as resonant scanning and online denoising, which improve live imaging by reducing toxicity of laser scanning. Considering these features together, such a high-tech and open in vivo two-photon microscope is not otherwise available in the Basel area. The project proposals here demonstrate that fluorescent mouse lines and in vivo hypotheses already exist in waiting, and only lack the means to image in vivo. In the absence of a two-photon intravital microscope, most research has been limited to ex vivo and in vitro studies, which ultimately beg the question of what happens in vivo. And in the cases where two-photon in vivo studies have already been employed through external collaboration, researchers have found limitations in resolution, depth of imaging, duration of experiment, and usability. Therefore these projects clearly reflect the urgent need for this novel in vivo imaging instrument and suggest what potentially high impact and clinically relevant data would be generated if the grant were awarded.
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