1Department of Ophthalmology, University of Michigan, 2MidWest Eye Banks
Woodward, M. A., Titus, M., Mavin, K., Shtein, R. M. Corneal Donor Tissue Preparation for Endothelial Keratoplasty. J. Vis. Exp. (64), e3847, doi:10.3791/3847 (2012).
Over the past ten years, corneal transplantation surgical techniques have undergone revolutionary changes1,2. Since its inception, traditional full thickness corneal transplantation has been the treatment to restore sight in those limited by corneal disease. Some disadvantages to this approach include a high degree of post-operative astigmatism, lack of predictable refractive outcome, and disturbance to the ocular surface. The development of Descemet's stripping endothelial keratoplasty (DSEK), transplanting only the posterior corneal stroma, Descemet's membrane, and endothelium, has dramatically changed treatment of corneal endothelial disease. DSEK is performed through a smaller incision; this technique avoids 'open sky' surgery with its risk of hemorrhage or expulsion, decreases the incidence of postoperative wound dehiscence, reduces unpredictable refractive outcomes, and may decrease the rate of transplant rejection3-6.
Initially, cornea donor posterior lamellar dissection for DSEK was performed manually1 resulting in variable graft thickness and damage to the delicate corneal endothelial tissue during tissue processing. Automated lamellar dissection (Descemet's stripping automated endothelial keratoplasty, DSAEK) was developed to address these issues. Automated dissection utilizes the same technology as LASIK corneal flap creation with a mechanical microkeratome blade that helps to create uniform and thin tissue grafts for DSAEK surgery with minimal corneal endothelial cell loss in tissue processing.
Eye banks have been providing full thickness corneas for surgical transplantation for many years. In 2006, eye banks began to develop methodologies for supplying precut corneal tissue for endothelial keratoplasty. With the input of corneal surgeons, eye banks have developed thorough protocols to safely and effectively prepare posterior lamellar tissue for DSAEK surgery. This can be performed preoperatively at the eye bank. Research shows no significant difference in terms of the quality of the tissue7 or patient outcomes8,9 using eye bank precut tissue versus surgeon-prepared tissue for DSAEK surgery. For most corneal surgeons, the availability of precut DSAEK corneal tissue saves time and money10, and reduces the stress of performing the donor corneal dissection in the operating room. In part because of the ability of the eye banks to provide high quality posterior lamellar corneal in a timely manner, DSAEK has become the standard of care for surgical management of corneal endothelial disease.
The procedure that we are describing is the preparation of the posterior lamellar cornea at the eye bank for transplantation in DSAEK surgery (Figure 1).
1. Setup: Aseptic Technique11
3. Representative Results
Proper mechanical microkeratome dissection of the donor cornea results in smooth, uniform lamellar corneal donor tissue. This is shown by an optical coherence tomography (OCT) image of the cornea in cross section (Figure 9). The final posterior corneal tissue should be of adequate thickness as measured by pachymetry. Over the past 6 months at the Michigan Eye Bank, the mean pre-processing corneal thickness was 558 microns, and post-processing thickness of the posterior corneal tissue was 158 microns. Eighty-seven percent of microkeratome-dissected donor tissue measured between 100-200 microns, 10.9% measured >200 microns, and 2.1% measured <100 microns.
The posterior corneal tissue must retain high endothelial cell density (ECD) to provide improved endothelial function to the transplant recipient. Specular microscopy is used to measure ECD. In the past 6 months, all of the corneal tissue deemed eligible for endothelial keratoplasty from the Michigan Eye Bank had an ECD of >2200 cells/mm2. Ninety-nine percent of the precut DSAEK tissue had a post-processing ECD >2400 cells/mm2, and 19% had an ECD > 3000 cells/mm2. The mean change in pre- to post-processing ECD was negligible at 1.2% (p=0.0003, paired two-tailed t-test). Complications that can occur during the procedure include loss of pressure, asymmetric dissection, and significant tissue indentation (resulting in corneal endothelial cell loss). Improper dissection can also result in tissue perforation.
Figure 1. Overall Scheme of Corneal Tissue Preparation for Endothelial Keratoplasty Figures 1a to 1e are cross-section representations of the donor cornea preparation. Figures 1f to 1i are cross-section representations of the transplant recipient's cornea during DSAEK surgery.
Figure 2. Corneal Tissue in Media. Corneal tissue is in preservation media after harvesting from donor.
Figure 3. Corneal Tissue on Artificial Anterior Chamber.
Figure 4. Corneal Tissue Thickness Check. The corneal thickness is checked using ultrasound pachymetry probe on surface of cornea.
Figure 5. Corneal Tissue Dissection.
Figure 6. Anterior Cap of Cornea after Dissection. The anterior cornea cap is free after microkeratome dissection is complete.
Figure 7. Dissected Cornea in Viewing Chamber. After dissection is completed, the cornea is returned to the viewing chamber for transportation.
Figure 8. Cornea in Packaging for Transportation. The cornea in the viewing chamber is then securely packaged for transportation to the surgeon.
Figure 9. OCT Appearance of Cornea. The OCT of the final dissected cornea (in cross-section) shows that the cornea is split into two halves. The interface is more intensely reflective than the surrounding tissue as indicated by the arrow.
The inner-most layer of the cornea, the endothelium, maintains the optical clarity of the cornea by maintaining corneal dehydration. Numerous disease states specifically affect the health and viability of the corneal endothelium including dystrophies, infections, inflammatory processes, and degenerations. Until recently, corneal transplantation to replace diseased corneal endothelium required full thickness transplantation of the cornea. In the past decade, targeted replacement of the posterior cornea by partial thickness corneal transplantation, such as DSAEK surgery, has been able to achieve the same goal with a shorter recovery time. Regardless of technique, transplanted tissues must retain a healthy corneal endothelium to maintain optical clarity in the transplanted cornea; as a result, all tissue processing techniques have additional steps to protect the corneal endothelium.
Automated microkeratome dissection is currently the standard for corneal tissue preparation for endothelial keratoplasty. Use of the microkeratome was shown to decrease donor tissue perforation and speed visual recovery as compared to manual dissection 5. When DSAEK was initially introduced, some surgeons purchased microkeratome equipment and performed donor lamellar dissection in the operating room at the start of surgery. If tissue preparation failed, the surgery would have to be cancelled causing inconvenience and expense to the patient and the surgeon. In 2006 eye banks began performing microkeratome dissection "off site" under controlled conditions in the eye bank laboratory a day prior to surgery 12. If tissue preparation fails in this setting, there is time to perform another dissection with fresh tissue.
Microkeratome dissection of donor posterior lamellar corneal tissue requires gentle and precise technique to minimize complications. The cornea must be placed gently on the artificial anterior chamber (AAC) to avoid any damage to the delicate endothelial cells. Once the cornea is mounted and aligned on the AAC, the locking ring must be positioned and adjusted for a tight seal to maintain high pressure within the chamber and decrease the chances of tissue collapse during the microkeratome dissection. To ensure uniform dissection, the microkeratome head should be rotated in a constant, smooth manner. After dissection, the tissue must again be handled carefully to minimize endothelial cell damage. Further, it is critical to confirm that the lamellar cut is centered and complete in order to ensure that the posterior lamella will be suitable for use in surgery 13.
Corneal donor perforation is a dreaded complication of this procedure; it becomes less frequent with experience. If repeated perforations occur during microkeratome passes, it is important to check multiple steps in the procedure. The cornea needs to be properly mounted and aligned on the AAC. A tight seal and high pressure should be obtained prior to microkeratome pass and maintained throughout the procedure. On occasion, residual conjunctiva exists on the sclera rim and needs to be dissected off prior to mounting. The microkeratome blade should be sharp and without notches. The correct microkeratome head thickness should be selected based on the estimated corneal thickness. The microkeratome head must be oscillating properly prior to the pass. With further problems with this technique, consult the microkeratome provider to verify that all equipment is working properly.
Even when performed perfectly, there are some limitations to this technique. Microkeratome heads are available to cut corneal tissue at various thicknesses; however, the relationship of microkeratome head to final tissue thickness is imprecise with the same head providing a wide range of final tissue thicknesses 7. In addition, a significant learning curve exists and complications in preparation are more frequent for new users. Performing the lamellar dissection in the eye bank setting allows trained technicians who perform this procedure many times per day to optimize their technique, thereby minimizing complications due to inexperience.
Modifications of the technique for microkeratome-assisted corneal dissection for endothelial keratoplasty exist. Some prefer to use viscoelastic material or corneal storage media rather than balanced salt solution beneath the corneal button on the artificial anterior chamber with the intent of providing increased protection to the corneal endothelium. There are also variations in the method for determining the location in which the microkeratome incision should be initiated. Different eye banks and different corneal surgeons may have particular preferences as to how the cornea is marked for identification of the anterior corneal cap or posterior lamellar orientation. It should be noted that manufacturers make similar equipment to that used in this protocol. The particular manufacturer's instructions should be followed with whichever equipment is used.
Dependable eye bank preparation of dissected corneal tissue for DSAEK surgery has been beneficial for the widespread acceptance of this relatively new surgical technique. Once surgeons became confident in eye banks performing the "first step" in corneal transplantation - namely tissue preparation, many practical limitations to DSAEK were alleviated. Eye banks have been able to reliably keep up with the demand for processed tissue with safe, effective, and timely supply of donor tissue. High quality DSAEK tissue can now be ordered in advance and operating room time and tissue preparation complications have decreased 9, 14-15.
This protocol describes the preparation of DSAEK grafts from the perspective of eye bank processing of corneal donor tissue; however, this technique is directly applicable to surgeons interested in preparing their own lamellar corneal donor tissue as well as those interested in performing laboratory-based or clinical studies on lamellar corneal transplantation.
Future directions for corneal lamellar preparation include use of the femtosecond laser for tissue processing, creation of "ultra-thin" DSAEK corneal tissue, or preparation of corneal endothelial scrolls for Descemet's membrane endothelial keratoplasty (DMEK) surgery. Endothelial keratoplasty is a surgical field in evolution and the symbiotic relationship between the eye bank and corneal surgeon will continue to change as surgical techniques are optimized.
Drs. Woodward and Shtein have nothing to disclose. Mr. Mavin and Mr. Titus are employees of Midwest Eye Banks.
Supported in part by NIH/NEI EY017885 (RMS).
|Artificial Anterior Chamber base||Moria||19161-562|
|Artificial Anterior Chamber cap||Moria||19172-220|
|Microkeratome Head 300||Moria||O304|
|Microkeratome Head 350||Moria||N791|