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F. Bock, K. Gottschalk, B. O. Bachmann, C. Cursiefen; Intravital Detection of Lymphatic Vessels in Vascularized Corneas Using Confocal Microscopy (HRT II With Cornea Module) in the Mouse Model. Invest. Ophthalmol. Vis. Sci. 2010;51(13):1569.
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© ARVO (1962-2015); The Authors (2016-present)
We recently demonstrated the specific visualization of lymphatic vessels in vascularized murine corneas in-vivo using the two photon-microscopy. Since lymphatic vessels in the cornea are not visible using normal slit-lamp magnification and since the mouse offers various models to analyse hem- and lymphangiogenesis, we established the detection of corneal lymphatic vessels in vivo by confocal microscopy in the mouse model.
Corneal neovascularization was induced in 6-8 weeks old BALB/c mice corneas by intrastromal placement of 11-0 nylon sutures. Double-immunohistochemistry using CD31 as panendothelial marker and LYVE-1 as specific lymphatic endothelial marker on whole mounts of these corneas was used to confirm combined corneal hemangiogenesis and lymphangiogenesis. The HRT II with cornea module (Heidelberg Engineering, Rostocker Modul) was used to analyze these corneas for the presence of blood and lymphatic vessels. To clearly label lymphatic vessels in vivo, india ink was injected into vacularized corneas prior to confocal imaging. Afterwards, these eyes were analyzed using hematoxylin&eosin histology.
Immunohistochemistry confirmed that placement of nylon sutures caused combined hem- and lymphangiogenesis into the mouse cornea. HRT imaging revealed numerous dark vascular structures with plenty of fast moving bright erythrocytes (blood vessels). In addition, black and empty appearing vascular structures were found close to, but separate from blood vessels. The diameter of these vessels was larger than that of the blood vessels. 24 hours after india ink injection, bright particles moving very slowly were found in these previously empty appearing (lymphatic) vessels. H&E histology localized india ink particles selectively in stromal lymphatic (i.e. non-erythrocyte-filled) vessels.
Using the mouse as a model to detect lymphatic vessels intravital by confocal microscopy allows the analysis of the lymphatic status in different well established murine models.
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