1. Post-Mortem Ultra-Structural Analysis of a Cornea Transplanted With DMEK Jack Parker Jr, MD; Eitan Livny, MD; Vasilis S. Liarakos, MD, PhD; Gerrit R. Melles, MD, PhD The Netherlands Institute for Innovative Ocular Surgery (NIIOS), Melles Cornea Clinic, and the Amnitrans Eye Bank, Rotterdam, The Netherlands. *No authors have any financial interests to disclose

2. Background (1/2) After Descemet Membrane Endothelial Keratoplasty (DMEK), the cornea of the transplanted eye looks essentially “normal,” virtually indistinguishable from a healthy, unoperated eye when viewed by slit-lamp biomicroscopy… 3 months after DMEK for Fuchs Endothelial Dystrophy

3. Background (2/2) …However, in operations where we’ve had to replace a “failed” DMEK graft, we’ve observed that performing the Descemetorhexis is significantly more challenging than in a virgin eye. Seemingly, the donor DMEK graft “sticks” more firmly the recipient posterior stroma than does normal endothelium. As a result, we began to wonder: 1)Ultra-structurally (under high power microscopes with special tissue stains) does eyes with DMEK grafts really look indistinguishable from a normal, unoperated eyes 2)What explains this extra observed “stickiness” that DMEK grafts appear to have? 3)Might an explanation for this “stickiness” shed new light on the cause (and prevention of) DMEK’s most common complication – graft detachment?

4. Experimental Design To investigate these matters, we harvested the whole globe of a deceased patient formerly operated with a DMEK for Fuchs Endothelial Dystrophy (FED). The Patient: 76 year old white female, who received her DMEK previously at the Netherlands Institute for Innovative Ocular Surgery (NIIOS) The tissue: The cornea was removed, sectioned, and sent for light and transmission electron microscopy

5. After DMEK, before Death (1/2) Six months after the patient’s DMEK, the transplanted cornea was thin and clear, resulting in a best corrected distance visual acuity of 20/25 (0.8). Anterior segment optical coherence tomography (A), slit-lamp (B, ), and Pentacam images (C)

6. After DMEK, before Death (1/2) Two years after surgery (at the last clinic visit before the death) the cornea remained thin and clear with a healthy appearing endothelial cell population, essentially identical to a normal, unoperated eye. Vision was stable at 20/28 (0.7) Slit-lamp (A) and specular microscopy (B) images

7. Results: Light Microscopy after Death Light microscopy image of the patient’s cornea 2 years after DMEK. No identifiable scar tissue or inflammatory reaction is seen at the donor-to-host interface. Note the bridging collagen fibers connecting the host’s stroma to the donor’s DM (arrows). [The separation of the donor DM from the host posterior stroma is an artifact induced by tissue processing for light microscopy]

8. Results: Light Microscopy after Death Light microscopy of the edge of the DMEK graft. No scar or hypertrophic tissue or an overlap of the graft and the central rim of the descemetorhexis could be identified, and the graft edge appears similar to more central areas

9. Results: Transmission Electron Microscopy (TEM) after Death Transmission electron microscopy of the patient’s cornea 2 years after DMEK (A) and of a healthy naïve eye (B). Junction between donor DM and host stroma. Note the close attachment of the interfacial matrix of the donor DM to the host stroma and the projection of stromal collagen fibrils into the interfacial matrix (arrows). Fibril projection depth is 6400 nm (dashed bar). Note that the ultrastructure of the DMEK graft-to-stroma junction is virtually indistinguishable from the normal DM-to-stromal junction in a naive cornea. ABL, anterior banded layer of the DM; IFM, interfacial matrix; ST, stroma host; the arrowhead points to longspacing collagen fibers (bar = 500 nm).

10. Discussion Ultra-structurally, the DMEK graft looked identical to a normal, unoperated corneal endothelium, without any additional (or absent) identifiable structures The additional observed “stickiness” observed when stripping a failed DMEK graft from the posterior corneal surface may stem from: 1.Deeper projection of collagen fibrils into the DM 2.Larger number of linking collagen fibrils, 3.Higher concentration of adhesion molecules as occasionally seen in naive corneas 4. Other adhesion structures such as peg-like interdigitations of the interfacial matrix into the host stroma that were not seen

11. Conclusion DMEK seems to accurately restore the cornea’s normal anatomy The absence of identifiable scar tissue at the graft interface may explain the unprecedented visual outcomes of the operation

12. Dr Melles is a consultant for D.O.R.C. International/ Dutch Ophthalmic USA. Thank you! Jack Parkerjack.parker@gmail.com Gerrit Melles info@niios.com NIIOS Wet Lab Courses Limited enrollment Practical lectures Hands on instruction Watch live operations, see the patients the next day