Large-scale isolation of human hepatocytes for therapeutic application.

During the last decade, hepatocyte transplantation has been suggested as a safe and potentially effective clinical option for the treatment of acute or decompensating chronic liver failure as well as for hereditary liver disease. Currently, one of the major limiting factors for clinical application is the insufficient access to suitable liver cell preparations. In cooperation with the German and Catalane organ procurement organizations, a routine procedure for the isolation of hepatocytes from donor organs rejected for transplantation (n = 117) has been established. The process is performed according to the current EC Guidelines for Good Manufacturing Practice (cGMP) and all corresponding national laws and regulations concerning donor organ and tissue procurement. In about 50% of the cases (n = 58) the three-step perfusion procedure has been completed with an average total cell yield of 5.9 x 10(9) cells per organ, the cell preparations displaying a mean viability of 64%. The mean specific yield was 3.6 x 10(6) total and 2.6 x 10(6) viable cells per gram liver tissue, respectively. Specific cell yields from three infantile donor livers were considerably higher. No correlation between isolation efficiency and cold ischemia time or donor age was found within the adult organ donors. In contrast, organs with a severe steatosis generally did not result in successful cell isolation. Results of sterility and endotoxin determination are also presented. In summary, a standardized and cGMP conform method of hepatocyte isolation from nontransplantable liver organs was established, which reproducibly yields large amounts of hepatocytes suitable for therapeutic application.

[Lamellar keratoplasty - new concepts]. / Lamellierende Keratoplastik - neue Konzepte.

New concepts of lamellar keratoplasty techniques mainly deal with the refinement of the so-called deep lamellar endothelial keratoplasty (DLEK). The aim of the deep lamellar technique is to expose bare Descemet's membrane of the recipient and to suture in a full thickness graft. Graft rejections are not known with DLEK. We describe the preparation techniques of Anwar, Melles, and Krumeich and discuss the advantages and disadvantages. The femtosecond laser enables the surgeon to cut the cornea non-mechanically with a cutting accuracy of +/- 10 microns. Fuchs endothelial dystrophy can be treated by posterior lamellar keratoplasty (POLK). In this technique only a sheet of Descemet's and endothelium are replaced. In severe alkali burn with limbal stem cell deficiency a lamellar corneo-scleral disc can be grafted in order to restore the limbus. Routine central penetrating keratoplasty should follow at least half a year later.