[Morphological assessment of different amniotic membrane epithelial denuding techniques].
OBJECTIVE
To evaluate the morphological features of the amniotic membrane denuded by different techniques.


METHODS
Human amniotic membrane was collected at the time of delivery, fixed in increasing concentrations of glycerol (0-50% in DMEM) and preserved at -80 degrees C until the time of use. The study consisted of 4 groups: intact epithelium (control) and denuded by trypsin (2 mg/mL at 1:250), dispase (1.2 U/mL in Mg2+ and Ca2+ free Hank's balanced salt solution) or ethylenediaminetetraacetic acid (EDTA), 0.02%. Specimens were submitted to electron (scanning and transmission) microscopy analysis.


RESULTS
Scanning electron microscopy disclosed intact epithelium in the control group and its absence in the amniotic membranes denuded by trypsin and dispase. In those denuded by ethylenediaminetetraacetic acid there were areas with and without epithelium. When assessed by transmission electron microscopy, the epithelium was intact and firmly adhered to the basement membrane by hemidesmossomes in controls and in parts of ethylenediaminetetraacetic acid group. There were only collagen fibers in the dispase- and trypsin-treated groups.


CONCLUSIONS
Trypsin and dispase treatment of the amniotic membrane may cause complete denuding of the epithelium and basement membrane whereas ethylenediaminetetraacetic acid may leave some intact epithelium-areas and partially destroy the basement membrane in others.
de Melo
Gustavo Barreto
GB
Universidade Federal de São Paulo, São Paulo, SP, Brasil. gustavobmelo@oftalmo.epm.br
Gomes
José Alvaro Pereira
JA
da Glória
Maria Aparecida
MA
Martins
Maria Cristina
MC
Haapalainen
Edna Freymüller
EF
D000650:Amnion / Q000187:drug effects / Q000648:ultrastructure*; D018929:Cell Culture Techniques; D004492:Edetic Acid / Q000494:pharmacology; D010450:Endopeptidases / Q000494:pharmacology; D004847:Epithelial Cells / Q000648:ultrastructure*; D006801:Humans; D008854:Microscopy, Electron; D014357:Trypsin / Q000494:pharmacology
D004492:Edetic Acid; D010450:Endopeptidases; D014357:Trypsin; C023139:dispase