An Experimental Research On The Effects Of Intermediate And Long-term Cryopreservation Methods On Corneal Endothelium Viability From American

Background：According to the statistics of the second china national samplesurvey in2008,china has12.33million people who are visual disability，and about4million people caused by corneal disease among them，corneal disease is the secondleading cause of legal blindness.Most of them could see again by corneal transplantoperation. But the number of corneal transplant operations in our country every yearwas only4000to5000, Most of corneas were donated by our donors, The method ofcorneal preservation is mainly moist chamber storage(MCS),Corneal preservationtime is not more than24hours.Small Parts of corneas were donated by Americans，Method is the preservation of intermediate term medium(M-K or Optisolliquid),Corneal preservation time is4to14day.Few Method is the long-termpreservation(cryopreservation or in glycerol).Corneal transplant operations used thecorneas donated by Americans for our patients in the Northern hospital，and restoredsight to the corneal disease patients. So the quality of corneas by long-timepreservation and long-distance transportation are very important， Including allcorneas，the activity of corneal endothelial cells is most important.They will be keyto if corneal transplant operation will be successful， and if the occurrence ofcomplications will be happened. So we will use different methods study of the activityof corneal endothelial cells, and through combine the advantages of the fourmethods，we will find the best method of corneal metaphase preservation. Next，wewill study77corneas acceptted time from march2012to march2013been donatedby Americans.Objective: To evaluate corneal endothelium viability and clinical applicabilityby intermediate and long-term cryopreservation methods and by long-time and distance trasport.In order to abtain zhe atandard methods of corneal preservationand trasport.Methods: To study zhe77corneals received by department of ophthalmologyof Northern Hospital since march2012to march2013.Through the examinations ofcorneal s‘s Trypan blue and alizarin red staining、Pathological sections、 Scanningelectron microscope and Transmission electron microscope，to analysis cornealendothelium viability and ultrastructural.Results:1. Corneal endothelial cell density(ECD): There are not significantdifferences between Group I(corneal preservation time:4-7day) to Group II (cornealpreservation time:8-14day),P>0.05.and there are significant differences betweenGroup III (corneal preservation time:above14d) to both Group I and II,P<0.05.Thereis not significant differences between Group M-K solution to Group Optisolsolution.(Picture3)2.Endothelial survival rate(ESR): ESR is significant decreased in GroupIII,include1was saved by M-K solution,and2were saved by Optisol solution.Thenucleus are stained blue by Trypan blue(Picture1,2).The corneals cant be used inclinical operation of corneal transplant.includ penetrating keratoplasty (PKP) andcorneal endothelial cell transplantation.3.Pathology: There are five clear layers in Group I and Group II ofpathological examination, Endothelial cells are single layer and behind thedescemet’s membrane. The nucleus are stained violet,and the cell walls arecomplete.But the endothelial cells are missed in Group III(pic4-9).4.Ultrastructural:4.1Scanning electron microscope:The endothelial cells are hexagonal、flatarrangement and almost zhe same size in Group I and Group II(pic10),and there aremany microvillis in them.cell-cell junctions are very closing. But it is contrary inGroup III(pic11).4.2Transmission electron microscope: The nucleus and the cell membranesare intact in Group I and Group II, the nucleus is long oval、 flat arrangement.thereare many microvillis beside it.By contrast,the nucleus are losed or karyolysis,andthe mitochondrions are swelling,and the endoplasmic reticulums are ecstatic(pic13).Conclusion: 1.Most of the corneal endothelium viability and density conform to the clinicalstandard of corneal transplant operation by intermediate and long-termcryopreservation methods and by long-time and distance trasport from AmericaArizona’s eye bank.The corneals could be used in corneal transplant.and the time ofthe corneal saved and transported should be since4day to14day.2. This is almost undifferentiated in the matter of prervation of corneasbetween the M-K solution and to Optisol solution. corneal endothelium viabilityand density are good effect of preservation since4day to14day.