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ID naloge: 10 Letnik: 1997 Predmet: oftalmologija
Primerjava dveh postopkov
shranjevanja roženic in vitro
Avtor: Mojca Grobelnik, Kristina Morela
Mentor: Prof. dr. Gorazd Kolar
Somentor: As. dr. Vladimir Pfeifer, Dr. Matjaž Jeras, mag. Farm
IZHODIŠCA. Rešitev pomanjkanja primernih roženic umrlih dajalcev za presaditev predstavljajo ocesne banke, kjer lahko tkivo shranjujemo na razlicne nacine. Najpogosteje uporabljajo shranjevanje v tkivni kulturi pri 30°C do 37°C in v industrijsko pripravljenih medijih pri 4°C. V raziskavi smo želeli ugotoviti, kako dva razlicna postopka 14-dnevnega shranjevanja vplivata na kakovost roženic umrlih dajalcev. Odlocili smo se za primerjavo Optisola-GS® pri 4°C in tkivne kulture pri 31°C. Predvideli smo, da sta za 14-dnevno shranjevanje enako primerna oba postopka.
METODE. V laboratorijih Centra za tipizacijo tkiv smo v prospektivni raziskavi ocenjevali makroskopski videz in lastnosti endotelija pred shranjevanjem in po njem 16-ih roženic v Optisolu-GS® in 18-ih roženic v tkivni kulturi. Roženice umrlih dajalcev smo odvzeli na Inštitutu za sodno medicino Medicinske fakultete v Ljubljani. Primerjali smo prozornost, celovitost endotelija, gostoto endotelijskih celic, celicni mozaik ter obliko in meje posameznih endotelijskih celic roženic. Med shranjevanjem smo preverjali tudi sterilnost medijev.
REZULTATI. Dokazali smo, da primerjana medija glede celovitosti endotelija in celicnega mozaika nista enako primema za 14-dnevno shranjevanje (p < 0,05). Pri primerjavi roženic shranjenih v tkivni kulturi in Optisolu-GS® smo ugotovili statisticno znacilno spremembo v celovitosti endotelija po 14 dneh (p < 0,05). V tkivni kulturi so se majhne vrzeli med shranjevanjem zapolnile, v Optisolu-GS® pa so se pojavile nove vrzeli na prej neprekinjenem endoteliju, ali pa so se že prej obstojece še povecale. Prozornost roženic, meje in oblike endotelijskih celic pa se v obeh skupinah niso statisticno znacilno razlikovale.
ZAKLJUCKI. Rezultati primerjave vpliva obeh postopkov 14-dnevnega shranjevanja na kakovost roženic umrlih dajalcev govorijo v prid shranjevanja v tkivni kulturi pri 31 °C. Predvsem je treba poudariti možnost obnavljanja endotelija roženic v tkivni kulturi na racun povecanja in preureditve preostalih endotelijskih celic. V raziskavi smo potrdili, da uporaba Optisola-GS® pri 4°C ni primema za 14-dnevno shranjevanje, saj se v tem casu celovitost endotelija znacilno poslabša. Shranjevanje v tkivni kulturi pa je varno le, ce imamo na razpolago primeren laboratorij za delo v sterilnih razmerah in strokovno usposobljeno osebje. Zakljucimo lahko z dejstvom, da je postopek shranjevanja roženic v tkivni kulturi pri 31 °C primeren vsaj za dobo 14 dni ali vec, medtem ko je uporaba Optisola-GS® pri 4°C za 14-dnevno shranjevanje roženic bistveno manj primerna.
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[Abstract / English version]
Comparrison of two storage medium
on donor corneas
Author: Mojca Grobelnik, Kristina Morela
Mentor: Prof. dr. Gorazd Kolar
Co-mentor: As. dr. Vladimir Pfeifer, Dr. Matjaž Jeras, mag. Farm
BACKGROUND. Eye banks offer a solution to the problem of the unavailability of suitable human donor corneas to be used as transplants. In such banks tissue can be stored in different ways: The most frequently used method is storing in a tissue culture at 30 to 37°C, and in commercially available media at 4°C. Our study was aimed to show how the two different methods of 14-day storing influence the quality of the human donor corneas. Optisol-GS® at 4°C and tissue culture at 31 °C were compared, as we presumed that both procedures are equally suitable for 14-day storage.
METHODS. In the prospective study carried out in the laboratories of the Tissue Typing Center we were assessing the macroscopic appearance and characteristics of the endothelium before and after storage; 16 corneas were stored in Optisol-GS® and 18 in tissue culture. The corneas were taken from dead donors at the Institute of Forensic Medicine, Faculty of Medicine in Ljubljana. Features such as corneal transparency, integrity of the endothelium, density of endothelial cells, endothelial cell mosaic, as well as the shape and margins of individual endothelial cells of the cornea were compared. The sterility of media was monitored throughout the storage period.
RESULTS. We have proved that the media under study are not equally suitable for 14-day storage with respect to two features, i.e. integrity of the endothelium and cell mosaic (p < 0.05). The comparison of corneas stored in tissue culture with those stored in Optisol-GS® has revealed a statistically significant difference in integrity of the endothelium after 14 days of storage (p < 0.05). While in tissue culture small defects in the endothelium were filled in, the previously intact endothelium stored in Optisol-GS® showed new defects, or any previously existing defects became bigger. There were no statistically significant differences between the two groups with respect to transparency of the cornea, and the margins and shapes of endothelial cells.
CONCLUSIONS. The results of our study comparing the influence of two 14-day storage methods on the quality of human donor corneas confirm the advantage of storage in the tissue culture at 31°C. There, particularly the possibility of endothelial repair in tissue culture, attributable to compensatory hypertrophy and rearrangement of the remaining endothelial cells, should be pointed out. The study has confirmed that the use of Optisol-GS® at 4°C is not suitable for 14-day storage, as it significantly worsens the integrity of the endothelium. On the other hand, in order to ensure safe storage in culture medium, we need a suitable laboratory for work in sterile conditions and adequately qualified staff. Based on the above findings, it can be concluded that the procedure of cornea storage in tissue culture at 31°C is adequate at least for a period of 14 days or longer, while the use of Optisol-GS® at 4°C for 14-day storage is considerably less suitable.
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