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Time-Lapse In Vivo Imaging of Corneal Angiogenesis: The Role of Inflammatory Cells in Capillary Sprouting
Linköping University, Department of Clinical and Experimental Medicine, Ophthalmology. Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Sinnescentrum, Department of Ophthalmology UHL/MH.
Linköping University, Department of Clinical and Experimental Medicine, Ophthalmology. Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Sinnescentrum, Department of Ophthalmology UHL/MH.
Östergötlands Läns Landsting, Centre for Diagnostics, Department of Clinical Pathology and Clinical Genetics.
Linköping University, Department of Clinical and Experimental Medicine, Ophthalmology. Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Sinnescentrum, Department of Ophthalmology UHL/MH.
2011 (English)In: Investigative Ophthalmology and Visual Science, ISSN 0146-0404, Vol. 52, no 6, 3060-3068 p.Article in journal (Refereed) Published
Abstract [en]

PURPOSE. To elucidate the temporal sequence of events leading to new capillary sprouting in inflammatory corneal angiogenesis.

METHODS. Angiogenesis was induced by corneal suture placement in Wistar rats. The inflamed region was examined by time-lapse in vivo confocal microscopy for up to 7 days. At 6 and 12 hours and 1, 2, 4, and 7 days, corneas were excised for flat mount immunofluorescence with primary antibodies for CD31, CD34, CD45, CD11b, CD11c, Ki-M2R, NG2, and alpha-SMA. From days 0 to 4, the in vivo extravasation and expansion characteristics of single limbal vessels were quantified.

RESULTS. Starting hours after induction and peaking at day 1, CD45(+)CD11b(+) myeloid cells extravasated from limbal vessels and formed endothelium-free tunnels within the stroma en route to the inflammatory stimulus. Limbal vessel diameter tripled on days 2 to 3 as vascular buds emerged and transformed into perfused capillary sprouts less than 1 day later. A subset of spindle-shaped CD11b(+) myeloid-lineage cells, but not dendritic cells or mature macrophages, appeared to directly facilitate further capillary sprout growth. These cells incorporated into vascular endothelium near the sprout tip, co-expressing endothelial marker CD31. Sprouts had perfusion characteristics distinct from feeder vessels and many sprout tips were open-ended.

CONCLUSIONS. Time-lapse in vivo corneal confocal microscopy can be used to track a temporal sequence of events in corneal angiogenesis. The technique has revealed potential roles for myeloid cells in promoting vessel sprouting in an inflammatory corneal setting.

Place, publisher, year, edition, pages
Research in Vision and Opthalmology , 2011. Vol. 52, no 6, 3060-3068 p.
National Category
Medical and Health Sciences
Identifiers
URN: urn:nbn:se:liu:diva-69178DOI: 10.1167/iovs.10-6101ISI: 000291100800026OAI: oai:DiVA.org:liu-69178DiVA: diva2:424306
Note
Original Publication: Beatrice Bourghardt Peebo, Per Fagerholm, Catharina Traneus-Rockert and Neil Lagali, Time-Lapse In Vivo Imaging of Corneal Angiogenesis: The Role of Inflammatory Cells in Capillary Sprouting, 2011, INVESTIGATIVE OPHTHALMOLOGY and VISUAL SCIENCE, (52), 6, 3060-3068. http://dx.doi.org/10.1167/iovs.10-6101 Copyright: Research in Vision and Opthalmology http://www.arvo.org/Available from: 2011-06-17 Created: 2011-06-17 Last updated: 2012-05-09Bibliographically approved
In thesis
1. Angiogenesis from a new perspective
Open this publication in new window or tab >>Angiogenesis from a new perspective
2012 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Angiogenesis is the emergence of new blood and lymph vessels from existing ones. In the pathologic form it contributes to the onset and progression of numerous different human disorders such as cancer, inflammation, atherosclerosis and blinding eye diseases. There exist a number of models to study angiogenesis, both in vitro and in vivo, but there is no single perfect model so far. Consequently there is a need to develop new angiogenesis assays for evaluating blood and lymph vessel behaviour in different physiologic settings.

The aim of this thesis was to gain insight into in vivo angiogenesis introducing a new technique in an inflammatory corneal model. The method involved in vivo examination of the cornea and subsequent comparison of in vivo findings with ex vivo immunohistochemical analysis of the same tissue samples. An existing suture model for inflammatory angiogenesis in the cornea was modified for in vivo observations with a clinically-approved corneal confocal microscope.

In this thesis, corneal lymph vessels were characterized for the first time in vivo and findings from the experimental bench could be applied in a clinical setting, where presumed lymphatics were observed in a corneal transplant patient with rejection. Furthermore, the technique was extended to investigate time-lapse processes in sprouting and regressing capillaries, and led to a number of new observations. CD11b+ myeloid cells constitute the first bulk of infiltrating inflammatory cells and contribute to inflammatory sprouting and regression in numerous ways including pre-patterning of the corneal stroma and guiding of capillary sprouts. Newly formed hemangiogenic sprouts are perfused with a slow-moving fluid and have a lumen. In blood vessel regression, capillary remodeling occurred by abandonment of sprout tips in close association with macrophages and vascular loops formed by presumed intussusceptive angiogenesis. In addition, a network of pericyte- and endothelium-free basement membrane tubes was formed after desertion or degradation of vascular endothelium in former corneal capillaries.

In conclusion, we introduce a new in vivo technique for investigating angiogenesis in a corneal model were in vivo findings can be interpreted with ex vivo definitions of specific cell types by immunohistochemistry. Findings from pre-clinical experiments have been possible to apply in a clinical setting when examining patients with corneal pathology.

Place, publisher, year, edition, pages
Linköping: Linköping University Electronic Press, 2012. 91 p.
Series
Linköping University Medical Dissertations, ISSN 0345-0082 ; 1284
National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:liu:diva-73137 (URN)978-91-7519-999-3 (ISBN)
Public defence
2012-01-20, Nils-Holger salen, Hälsouniversitetet, Campus US, Linköpings universitet, Linköping, 13:00 (Swedish)
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Supervisors
Available from: 2011-12-19 Created: 2011-12-19 Last updated: 2014-06-05Bibliographically approved

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