Regional differences and post-mortem stability of enzymatic activities in the retinal pigment epithelium.
BACKGROUND
The retinal pigment epithelium (RPE) is essential for the metabolism of the neural retina. As a result of dysfunction of the RPE, retinal degeneration occurs. A potential treatment for certain forms of retinal degenerations is transplantation of RPE cells. To determine optimal conditions for treatment of donor eyes before transplantation, activities of key proteases (aminopeptidase M, dipeptidylpeptidase II and IV and gamma-glutamyltranspeptidase) as indicators of RPE cell quality (viability and functional state) were measured.


METHODS
Protease activities were quantified in bovine RPE cells from different regions of the eyecup, after different times of storage of the bulbi, cryopreservation of the RPE cells and in RPE cell cultures. The distribution of the activities was compared to the pigmentation of the RPE cells, the thickness of the choroid and photoreceptor density.


RESULTS
Most proteases showed regional maxima. Prolonged storage of the bulbi decreased gamma-glutamyltranspeptidase and aminopeptidase M activities. Cryopreservation of the RPE cells for up to 6 weeks caused no loss in the enzymatic activities. Culture of RPE cells caused pronounced decreases in the activities of gamma-glutamyltranspeptidase and dipeptidylpeptidase IV. Storage of the bulbi at 4 degrees C for more than 50 h causes marked loss of enzymatic activities in RPE cells.


CONCLUSIONS
The decrease in gamma-glutamyltranspeptidase activity may be especially important because the RPE is exposed to high concentrations of reactive oxygen species. Whole bulbi should be stored for less than 50 h, but isolated RPE cells may be stored at -80 degrees C for weeks. Propagation of RPE cells by culture increases cell number; this effect may be counteracted by a decrease in the function of these cells.
Fröhlich
Eleonore
E
Anatomisches Institut, Osterbergstrasse 3, 72074 Tübingen, Germany. eleonore.froehlich@anatu.uni-tuebingen.de
Klessen
Christian
C
D000818:Animals; D018826:CD13 Antigens / Q000378:metabolism*; D002196:Capillaries; D002417:Cattle; D002478:Cells, Cultured; D015925:Cryopreservation; D018819:Dipeptidyl Peptidase 4 / Q000378:metabolism*; D004152:Dipeptidyl-Peptidases and Tripeptidyl-Peptidases / Q000378:metabolism*; D004795:Enzyme Stability; D005123:Eye; D005470:Fluorometry; D010857:Pigment Epithelium of Eye / Q000098:blood supply / Q000201:enzymology*; D011180:Postmortem Changes; D013997:Time Factors; D014021:Tissue Preservation; D005723:gamma-Glutamyltransferase / Q000378:metabolism*
D005723:gamma-Glutamyltransferase; D018826:CD13 Antigens; D004152:Dipeptidyl-Peptidases and Tripeptidyl-Peptidases; C100151:dipeptidyl peptidase II; D018819:Dipeptidyl Peptidase 4