Purpose: To present light and electron microscopic as well as immunohistochemical findings after corneal cross-linking (CXL).
Methods: Six keratoconus corneas after CXL, 12 keratoconus corneas without CXL, and 7 normal corneas were examined by light microscopy, indirect immunohistochemistry using antibodies against proapoptotic BAX, antiapoptotic survivin, and BCL-2, and anti–smooth muscle actin and, in part, by transmission electron microscopy. Direct immunofluorescence with 4′6-diamidino-2-phenylindole was performed to analyze keratocytes/area in the anterior, middle, posterior, peripheral, and central corneal stroma.
Results: The period between CXL and keratoplasty ranged from 5 to 30 months. All keratoconus corneas showed the typical histological changes. Increased proapoptotic BAX expression and/or antiapoptotic survivin expression were noticed in keratocytes and endothelium in 2 keratoconus specimens after CXL. Smooth muscle actin was only observed in subepithelial scar tissue of 2 keratoconus corneas without CXL. Keratoconus corneas after CXL revealed a significant reduction in keratocyte counts in the entire cornea (P = 0.003) compared with keratoconus corneas without CXL and normal corneas. This difference was because of a loss of keratocytes in the anterior (P = 0.014) and middle (P = 0.024) corneal stroma. Keratocytes in CXL corneas were reduced in the center (P = 0.028) and the periphery (P = 0.047).
Conclusions: CXL in human keratoconus can cause considerable morphologic corneal changes up to 30 months postoperatively. Especially noteworthy is a long-lasting, maybe permanent, keratocyte loss in the anterior and middle corneal stroma involving the central and peripheral cornea. As long-term corneal damage after CXL is of genuine concern, particular care should be taken to perform this procedure only in accordance with investigational protocols.
*Department of Ophthalmology, Ludwig-Maximilians-University, Munich, Germany
†University Eye Hospital, Basel, Switzerland
‡University Eye Hospital, Bonn, Germany
§University Eye Hospital, Homburg-Saar, Germany
¶Eye Hospital, Luzern, Switzerland
‖University Eye Hospital, Freiburg, Germany.
Reprints: Elisabeth M. Messmer, Department of Ophthalmology, Ludwig-Maximilians-University, Mathildenstr 8, 80336 Munich, Germany (e-mail: firstname.lastname@example.org).
The authors state that they have no funding or conflicts of interest to disclose.
Received September 9, 2011
Accepted January 27, 2012