Abstract
PurposeTo evaluate myofibroblast differentiation as an etiology of haze at the graft-host interface in a cat model of Descemet’s Stripping Automated Endothelial Keratoplasty (DSAEK).MethodsDSAEK was performed on 10 eyes of 5 adult domestic short-hair cats. In vivo corneal imaging with slit lamp, confocal, and optical coherence tomography (OCT) were performed twice weekly. Cats were sacrificed and corneas harvested 4 hours, and 2, 4, 6, and 9 days post-DSAEK. Corneal sections were stained with the TUNEL method and immunohistochemistry was performed for α-smooth muscle actin (α-SMA) and fibronectin with DAPI counterstain.ResultsAt all in vivo imaging time-points, corneal OCT revealed an increase in backscatter of light and confocal imaging revealed an acellular zone at the graft-host interface. At all post-mortem time-points, immunohistochemistry revealed a complete absence of α-SMA staining at the graft-host interface. At 4 hours, extracellular fibronectin staining was identified along the graft-host interface and both fibronectin and TUNEL assay were positive within adjacent cells extending into the host stroma. By day 2, fibronectin and TUNEL staining diminished and a distinct acellular zone was present in the region of previously TUNEL-positive cells.ConclusionsOCT imaging consistently showed increased reflectivity at the graft-host interface in cat corneas in the days post-DSAEK. This was not associated with myofibroblast differentiation at the graft-host interface, but rather with apoptosis and the development of a subsequent acellular zone. The roles of extracellular matrix changes and keratocyte cell death and repopulation should be investigated further as potential contributors to the interface optical changes.
Highlights
[3] Scatterometry and adaptive optics analyses suggest that corneal haze is a likely contributor to this sub-optimal vision. [1,4] In vivo human studies have investigated the locations and roles of corneal haze in visual acuity and other measures of visual function. [5,6] The graft-host interface has been identified as an area of increased corneal haze post-Descemet’s Stripping Automated Endothelial Keratoplasty (DSAEK)
All research was conducted with the approval of our Institutional Animal Care and Use Committee (IACUC), the University of Rochester Committee on Animal Research (UCAR)
DSAEK in Cats – surgical Outcomes All surgeries were performed successfully resulting in clear corneas (Figure 2) with no primary graft failures or tissue dislocations
Summary
Despite its many advantages over full thickness penetrating keratoplasty (PK) [1,2], endothelial keratoplasty can yield suboptimal post-operative visual acuity. [3] Scatterometry and adaptive optics analyses suggest that corneal haze is a likely contributor to this sub-optimal vision. [1,4] In vivo human studies have investigated the locations and roles of corneal haze in visual acuity and other measures of visual function. [5,6] The graft-host interface has been identified as an area of increased corneal haze post-DSAEK. [7] Interface reflectance, as measured by in vivo confocal imaging, decreases with time post-DSAEK and has been correlated with improvements in post-operative visual acuity. [8] while interface haze appears to be implicated in visual acuity limitations post-DSAEK, the cause of this interface haze remains unknown.In the normal cornea, keratocytes are quiescent cells that contribute to corneal clarity by maintaining the structure of the collagen lamellae and the composition of the extracellular matrix. [5,6] The graft-host interface has been identified as an area of increased corneal haze post-DSAEK. [7] Interface reflectance, as measured by in vivo confocal imaging, decreases with time post-DSAEK and has been correlated with improvements in post-operative visual acuity. In response to wounding stimuli, keratocytes become activated and differentiate into a myofibroblast phenotype. Myofibroblasts are less transparent than non-activated keratocytes,[14,15,16] and they further decrease stromal transparency by altering the organization and composition of the extracellular matrix.[17,18,19] In models of wound healing such as incisional wounds and photoablation, haze has been largely attributed to myofibroblast activity. DSAEK graft-host interface is due to myofibroblasts differentiation and activity Myofibroblasts are less transparent than non-activated keratocytes,[14,15,16] and they further decrease stromal transparency by altering the organization and composition of the extracellular matrix.[17,18,19] In models of wound healing such as incisional wounds and photoablation, haze has been largely attributed to myofibroblast activity. [15] In this study, we utilized a cat model of DSAEK to test the hypothesis that corneal haze at the post-
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