Universität zu Köln

Steven, Philipp, Bock, Felix, Huettmann, Gereon and Cursiefen, Claus (2011). Intravital Two-Photon Microscopy of Immune Cell Dynamics in Corneal Lymphatic Vessels. PLoS One, 6 (10). SAN FRANCISCO: PUBLIC LIBRARY SCIENCE. ISSN 1932-6203

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Abstract

Background: The role of lymphatic vessels in tissue and organ transplantation as well as in tumor growth and metastasis has drawn great attention in recent years. Methodology/Principal Findings: We now developed a novel method using non-invasive two-photon microscopy to simultaneously visualize and track specifically stained lymphatic vessels and autofluorescent adjacent tissues such as collagen fibrils, blood vessels and immune cells in the mouse model of corneal neovascularization in vivo. The mouse cornea serves as an ideal tissue for this technique due to its easy accessibility and its inducible and modifiable state of pathological hem-and lymphvascularization. Neovascularization was induced by suture placement in corneas of Balb/C mice. Two weeks after treatment, lymphatic vessels were stained intravital by intrastromal injection of a fluorescently labeled LYVE-1 antibody and the corneas were evaluated in vivo by two-photon microscopy (TPM). Intravital TPM was performed at 710 nm and 826 nm excitation wavelengths to detect immunofluorescence and tissue autofluorescence using a custom made animal holder. Corneas were then harvested, fixed and analyzed by histology. Time lapse imaging demonstrated the first in vivo evidence of immune cell migration into lymphatic vessels and luminal transport of individual cells. Cells immigrated within 1-5.5 min into the vessel lumen. Mean velocities of intrastromal corneal immune cells were around 9 mm/min and therefore comparable to those of T-cells and macrophages in other mucosal surfaces. Conclusions: To our knowledge we here demonstrate for the first time the intravital real-time transmigration of immune cells into lymphatic vessels. Overall this study demonstrates the valuable use of intravital autofluorescence two-photon microscopy in the model of suture-induced corneal vascularizations to study interactions of immune and subsequently tumor cells with lymphatic vessels under close as possible physiological conditions.

Item Type:	Journal Article
Creators:	Creators Email ORCID ORCID Put Code Steven, Philipp UNSPECIFIED UNSPECIFIED UNSPECIFIED Bock, Felix UNSPECIFIED UNSPECIFIED UNSPECIFIED Huettmann, Gereon UNSPECIFIED UNSPECIFIED UNSPECIFIED Cursiefen, Claus UNSPECIFIED UNSPECIFIED UNSPECIFIED
URN:	urn:nbn:de:hbz:38-487109
DOI:	10.1371/journal.pone.0026253
Journal or Publication Title:	PLoS One
Volume:	6
Number:	10
Date:	2011
Publisher:	PUBLIC LIBRARY SCIENCE
Place of Publication:	SAN FRANCISCO
ISSN:	1932-6203
Language:	English
Faculty:	Unspecified
Divisions:	Unspecified
Subjects:	no entry
Uncontrolled Keywords:	Keywords Language IN-VIVO; LYMPHANGIOGENESIS; INFLAMMATION; ENDOTHELIUM; MACROPHAGES; EXPRESSION; INDUCTION; MIGRATION; TUMOR Multiple languages Multidisciplinary Sciences Multiple languages
URI:	http://kups.ub.uni-koeln.de/id/eprint/48710