Details
| Original language | English |
|---|---|
| Pages (from-to) | 4247-4255 |
| Number of pages | 9 |
| Journal | Investigative Ophthalmology and Visual Science |
| Volume | 52 |
| Issue number | 7 |
| Publication status | Published - Jun 2011 |
| Externally published | Yes |
Abstract
Purpose. Photochemical cross-linking of corneal stromal collagen using riboflavin and ultraviolet irradiation is an evolving treatment for keratoconus. The purpose of the present study was to investigate the wound-healing process in rabbit corneas after cross-linking. Methods. Photochemical cross-linking was performed according to a standard protocol on the right eyes of eight male New Zealand White rabbits; the left eyes served as controls. Untreated controls and cross-linked rabbit corneas were imaged 3 days, 6 days, and 6 weeks after treatment using a customized setup for three-dimensional nonlinear microscopy and confocal laser-scanning microscopy of reflected femtosecond light (fs-CLSM). Results. The combination of fs-CLSM in reflective mode and two-photon-excited fluorescence permitted differentiation of the following zones in the lamina propria of treated corneas 3 and 6 days after cross-linking: (1) an anterior zone with postapoptotic keratocyte debris, visible only on fs-CLSM in reflective mode; (2) a posterior zone with activated keratocytes with strong autofluorescence; and (3) surviving or restored keratocytes with moderate autofluorescence beyond the intermediate zone. Repopulation with normal keratocytes was achieved by 6 weeks. Bi-directional, second-harmonic generation (SHG) imaging showed no global differences in the fiber orientation and lamellar structure of stromal collagen at any time point. A relatively strong additional two-photon excited fluorescence occurred in the treated corneas with a diffuse three-dimensional spatial distribution. Conclusions. This combination of imaging modalities has the potential to become a new clinical instrument capable of visualizing corneal changes at the cellular and extracellular level.
ASJC Scopus subject areas
- Medicine(all)
- Ophthalmology
- Neuroscience(all)
- Sensory Systems
- Neuroscience(all)
- Cellular and Molecular Neuroscience
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In: Investigative Ophthalmology and Visual Science, Vol. 52, No. 7, 06.2011, p. 4247-4255.
Research output: Contribution to journal › Article › Research › peer review
}
TY - JOUR
T1 - Combined nonlinear and femtosecond confocal laser-scanning microscopy of rabbit corneas after photochemical cross-linking
AU - Krüger, Alexander
AU - Hovakimyan, Marine
AU - Ojeda, Diego F.Ramírez
AU - Stachs, Oliver
AU - Wree, Andreas
AU - Guthoff, Rudolf F.
AU - Heisterkamp, Alexander
PY - 2011/6
Y1 - 2011/6
N2 - Purpose. Photochemical cross-linking of corneal stromal collagen using riboflavin and ultraviolet irradiation is an evolving treatment for keratoconus. The purpose of the present study was to investigate the wound-healing process in rabbit corneas after cross-linking. Methods. Photochemical cross-linking was performed according to a standard protocol on the right eyes of eight male New Zealand White rabbits; the left eyes served as controls. Untreated controls and cross-linked rabbit corneas were imaged 3 days, 6 days, and 6 weeks after treatment using a customized setup for three-dimensional nonlinear microscopy and confocal laser-scanning microscopy of reflected femtosecond light (fs-CLSM). Results. The combination of fs-CLSM in reflective mode and two-photon-excited fluorescence permitted differentiation of the following zones in the lamina propria of treated corneas 3 and 6 days after cross-linking: (1) an anterior zone with postapoptotic keratocyte debris, visible only on fs-CLSM in reflective mode; (2) a posterior zone with activated keratocytes with strong autofluorescence; and (3) surviving or restored keratocytes with moderate autofluorescence beyond the intermediate zone. Repopulation with normal keratocytes was achieved by 6 weeks. Bi-directional, second-harmonic generation (SHG) imaging showed no global differences in the fiber orientation and lamellar structure of stromal collagen at any time point. A relatively strong additional two-photon excited fluorescence occurred in the treated corneas with a diffuse three-dimensional spatial distribution. Conclusions. This combination of imaging modalities has the potential to become a new clinical instrument capable of visualizing corneal changes at the cellular and extracellular level.
AB - Purpose. Photochemical cross-linking of corneal stromal collagen using riboflavin and ultraviolet irradiation is an evolving treatment for keratoconus. The purpose of the present study was to investigate the wound-healing process in rabbit corneas after cross-linking. Methods. Photochemical cross-linking was performed according to a standard protocol on the right eyes of eight male New Zealand White rabbits; the left eyes served as controls. Untreated controls and cross-linked rabbit corneas were imaged 3 days, 6 days, and 6 weeks after treatment using a customized setup for three-dimensional nonlinear microscopy and confocal laser-scanning microscopy of reflected femtosecond light (fs-CLSM). Results. The combination of fs-CLSM in reflective mode and two-photon-excited fluorescence permitted differentiation of the following zones in the lamina propria of treated corneas 3 and 6 days after cross-linking: (1) an anterior zone with postapoptotic keratocyte debris, visible only on fs-CLSM in reflective mode; (2) a posterior zone with activated keratocytes with strong autofluorescence; and (3) surviving or restored keratocytes with moderate autofluorescence beyond the intermediate zone. Repopulation with normal keratocytes was achieved by 6 weeks. Bi-directional, second-harmonic generation (SHG) imaging showed no global differences in the fiber orientation and lamellar structure of stromal collagen at any time point. A relatively strong additional two-photon excited fluorescence occurred in the treated corneas with a diffuse three-dimensional spatial distribution. Conclusions. This combination of imaging modalities has the potential to become a new clinical instrument capable of visualizing corneal changes at the cellular and extracellular level.
UR - http://www.scopus.com/inward/record.url?scp=80052340069&partnerID=8YFLogxK
U2 - 10.1167/iovs.10-7112
DO - 10.1167/iovs.10-7112
M3 - Article
C2 - 21498616
AN - SCOPUS:80052340069
VL - 52
SP - 4247
EP - 4255
JO - Investigative Ophthalmology and Visual Science
JF - Investigative Ophthalmology and Visual Science
SN - 0146-0404
IS - 7
ER -