Dr.PranavMore
Cornea Fellow,
Cornea Services,
Aravind Eye Hospital,
Madurai.
Apart from the suitability of the donor tissue, the survival of the tissue after enucleation from a donor till the time of transplantation greatly depends on the preservation methods used. Unlike tissues such as bone or heart valves that may be extensively processed and altered from their natural state, corneas must be transplanted as a viable living tissue. Thus the aim of any current corneal storage technique is simply to maintain this living viable state while holding the cornea for the period between donation and transplantation. To enable us to achieve this aim various techniques have evolved over the past 30 years and are still evolving. Storage techniques are being used not only to store corneal tissue but also for preservation of scleral tissue.
Depending on he duration of storage corneal tissue preservation can be broadly classified into
- Short Term method
- Intermediate method
- Long Term method
All storage media to be stored at 40C and should be brought to room temperatures about 1 hr prior to transplantation.
|
Method |
Max.Duration of Storage |
Disadvantages |
Short Term |
Moist chamber at 4 0C |
48 hrs |
Surgery has to be performed in an emergency.
Significant endothelial cell loss.
|
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M-K Medium(most preferred) |
Upto 4 days |
Limited interval of preservation.
Some reports on endophthalmitis present.
|
Intermediate Term |
K-sol |
10 days |
Withdrawn from market due to bacterial contamination during manufacture. |
|
Corneal Storage Medium (CSM) |
7 days |
No longer available. |
|
Dexsol(Dextran supplemented CSM) |
2 weeks |
|
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Optisol G-S |
2 weeks |
Expensive
Not more effective than Dexsol.
|
Long Term |
Organ culture(340C) |
4-5weeks |
Requires 2 wks quarantine to detect contamination. |
|
Cryopreservation(-700C) |
Upto 1 yr |
Expensive and complex.
Careful monitoring for freezing and thawing required.
|
|
Short Term Method
- Moist Chamber Storage
Filatov first described the use of moist chamber storage of whole eyes. This is achieved by placing an enucleated eye in a sealed chamber together with gauze, usually moistened by a saline or antibiotic solution, and then placed at 4C. Care must be taken not to immerse the eye in solution as this will be absorbed by the cornea and cause stromal oedema. Although the moist chamber technique was used successfully and with confidence for over forty years (and is still employed in some countries) its main drawback is the limitation to around 24 hours of storage. This is because the endothelium of a cornea stored on the globe is subjected to the toxic build up of metabolic waste and necrotic tissue in the stagnant aqueous humor. To overcome this problem the idea evolved of removing the cornea and placing it in a biologically defined environment.
Intermediate Term Method(Hypothermic Corneal Storage)
- M-K medium
McCarey and Kaufman modified an existing tissue culture medium, TC 199, by adding dextran as an osmotic agent to compensate for the inactivity of the corneas normal water removal mechanism at 4C. This extended reliable storage time to 2-3 days and the media, M-K medium, quickly became the storage system of choice for eye banks following publication of several series of successful transplants. Over the years the M-K formulation has been improved by the addition of the more stable HEPES buffer and the replacement of the penicillin streptomycin antibiotics to gentamicin which has a greater spectrum against gram-negative bacteria.
- K-Sol medium
Kaufman et al in the mid-eighties added 2.5% chondroitin sulfate to the basic M-K formulation and the resultant solution, K-Sol, successfully extended corneoscleral storage times to 7-10 days. However one of the problems with media containing chondroitin sulfate is that the layers of the cornea can absorb some of its smaller molecular weight moieties, and the resultant osmotic flow of water causes the cornea to swell.
- Dexsol medium
Optisol
Optisol, the latest commercially available storage medium, further improves on Dexsol by maintaining better endothelial cell morphology and thinness of the corneas While the original publications suggested that Optisol was suitable for up to 14 days storage, in practice the maximum storage time most Eye Banks feel happy with is 7-10 days with storage between 2C - 6C. The manufacturers have also published that 48 hours room temperature storage (approximately 21C) still provides adequate corneal preservation. Optisol is now usually available as Optisol-GS that contains both gentamicin and streptomycin to give broad antimicrobial coverage.
Long Term Method
- Organ Culture -
This long term storage method using storage temperatures between 31C and 37C was developed by Doughman and Sperling and has been further developed in Europe This technique provides preservation of up to 35 days. This extended storage time overcomes some of the inherent organizational problems if matching high-risk recipients with tissue typed donor corneas is undertaken. In addition, the culture system provides a functional assessment of the cornea allowing the use of good corneas from marginal donor groups (such as older age groups, septicemia donors, extended postmortem times). These advantages have seen organ culture adopted as the preferred method of corneal storage across Europe, the UK and in New Zealand.
- Cryopreservation
Capella et al introduced cryopreservation in 1960 in which corneal tissue was immersed in cryoprotective solutions and stored at -700C for almost 1 year. However it is technically complicated requires well trained technician and expensive equipment. To add to this the donor must be less than 50 years of age and tissue must be harvested in less than 6 hours post mortem.
At the end, the most commonly used preservative media are MK and Optisol. MK medium is easily available in India as it is produced by the support of Rotary at Hyderabad and supplied to only recognized eye banks and costs approximately US$5/- per vial. Corneas can be preserved in this medium from 72 hours to 96 hours. The other medium is Optisol-GS. This has to be imported and costs approximately US$42/- per vial. Corneas can be stored in this medium for 10 to 14 days.
Serology
Reports of transmission of prion diseases like Creutzfeldt-Jakob5,6,7, rabies8 and Hep.B in the past few decades, has made it mandatory for a detailed and careful documentation of the cause of the death as well as the review of the medical records of the potential donor.Medical records should be checked especially for past history of receiving human pituitary-derived growth hormone, death with undiagnosed neurological disease, rabies, blood malignancies etc..
Apart from this the Eye Bank Association of America, has made it mandatory to perform the following serological tests on every donor -----
- HIV-1/HIV-2 (Human Immunodeficiency Virus
Human Immunodeficiency Virus has been detected in tears, conjunctiva and corneas from AIDS patient, although transmission via corneal transplantation has not been documented. A negative screening test must be documented prior to release of tissue for transplantation.
- Hepatitis B surface Antigen9 (HBs Ag):
Hepatitis B virus was always suspected to be transmissible through cornea and 3 such cases were reported at the American Academy of Ophthalmology, Las Vegas, 1988. HBs Ag has been found in corneal donors and the virus is thought to be transmittable by corneal transplantation. The risk of HBV transmission was shown to be much greater than that for HIV transmission. As both HIV & HBV are life threatening disease, the careful screening of donor blood is mandatory despite the lack of demonstrable transmission through corneal transplantation. Inability to test for these diseases in the donor presents serious risks, since the risk of infection by AIDS & Hepatitis through corneal transplantation is increasing.
- Hepatitis C antibodies (Anti HCV):
Screening for HCV antibody was deemed important when seroconversion (positive HCV antibody) was demonstrated in renal transplant receipients.
At least 5 cc of blood must be available for performing serology. The sample should preferably be collected in plain silicone coated vacutainer which is inexpensive. Many eye banks use the ELISA kits as the common screening method for HIV, HBsAg and Anti-HCV. The current kits are upto 97.1% specific and upto 99.6% sensitive in diagnosing the viral diseases in the donor.
The Future
In the future it may be possible for Eye Banks to enhance corneal epithelial and endothelial viability through the manipulation of growth factors. Similar manipulation could conceivably provide an opportunity to decontaminate the tissue of infectious agents including bacteria, viruses and prions. Such tissue engineering techniques may also be able to modify the immune rejection and wound healing responses of donor corneas, or indeed provide the ability for in vitro growth of a cornea.
References
- Filatov VP. Transplantation of cornea from preserved cadaver eyes. Lancet; 1:1395, 1937.
- McCarey BE, Kaufman HE. Improved corneal storage. Invest Ophthalmol 13: 165-73, 1974
- Aquavella JV, Van Horn DL, Haggerty CJ. Corneal preservation using M-K Medium. Am J Ophthalmol 80:791-799,1975.
- Lass JH, Bourne WM,Musch DC, et al : A randomized, prospective double-masked clinical trial of Optisol vs Dexsol corneal storage media. Arch.Ophthal 110: 1404, 1992
- Heckmann JG, Lang CJG, Petruch F et.al. Transmission of Creutzfeldt-Jakob disease via a corneal transplant. J Neurol Neurosurg Psychiatry 63: 388-90, 1997.
- Uchiyama K, Ischida C, Yago S, Kuramaya H, Kitmoto T. An autopsy case of Creutzfeldt-Jakob disease associated with corneal transplantation. Dementia 8: 466-73, 1994.
- Hogan RN, Heck E, Cavanagh HD. Risk of prion disease transmission from ocular donor tissue transplantation. Cornea 18: 2-11, 1999.
- Houff SA, Burton RC, Wilson RW et.al. Human-to-human transmission of rabies virus by corneal transplantation. N Eng J Med 300:603-604, 1979.
- Hoft RH, Pflugfelder SC, Forster RK, Ullman S, Polack FM, Schiff ER. Clinical evidence for hepatitis B transmission resulting from corneal transplantation. Cornea 16: 132-137, 1997.
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