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A.T. Dias, M. Martins, A.A. S. L. Filho, A.N. Odashiro, A.C. S. Lourenco, J.P. Gomes, R. Belfort, Jr., M.N. Burnier, Jr.; Histopathological Findings and Immunohistochemical Expression of Vimentin, Cytokeratin 8 and Cytokeratin 18 in Freeze–Dried and Fresh–Frozen Preserved Human Amniotic Membranes . Invest. Ophthalmol. Vis. Sci. 2006;47(13):5049.
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© ARVO (1962-2015); The Authors (2016-present)
To evaluate and compare histopathologic results and immunohistochemical expression of Vimentin, Cytokeratin 8 and Cytokeratin 18 obtained from freeze–dried (FD) and fresh–frozen (FF) preserved human amniotic membranes (HAM).
A total of 64 samples of HAM, obtained from 4 immediate post–partum women, were evaluated. For each amniotic membrane, 8 samples were FD and 8 were FF. After 1, 7, 30 and 90 days, 2 samples from each group (freeze–dried and fresh–frozen) were analyzed by Hematoxilin & Eosin (HE) and Periodic Acid–Schiff (PAS) staining. Immunohistochemical expression of Vimentin in the conective tissue, Cytokeratin 8 and Cytokeratin 18 in the epithelium were also performed. The samples were classified semi–quantitatively according to degree of epithelial vacuolization (0–3), autolysis (0–3), integrity of basement membrane (0–2), and the expression of Vimentin, Cytokeratin 8 and Cytokeratin 18 (0–3). The mean scores of each feature were compared between each method and each placenta using independent samples t test and Anova test. A p–value of less than 0.05 was considered to be statistically significant.
The average score and standard deviation for the semi–quantitative characteristics were calculated: Epithelial vacuolization: 1.25±0.84 (FD – n=32) and 2.38±0.79 (FF – n=32)
Autolysis: 0.41±0.50 (FD – n=32) and 1.53±0.88 (FF– n=32)
Integrity of basement membrane: 1.87±0.34 (FD – n=31) and 1.28±0.46 (FF – n=32)
Vimentin: 2.55±0.57 (FD – n=31) and 1.31±0.74 (FF – n=32)
Cytokeratin 8: 1.90±0.60 (FD – n=31) and 0.97±0.65 (FF – n=32)
Cytokeratin 18: 2.50±0.51 (FD – n=30) and 1.34±0.48 (FF – n=32)
Analysis of each of the antibodies’ expressions yielded statistically significant differences (p<0.001).
Freeze–drying and fresh–freezing are effective methods for HAM preservation. Upon histopathological analysis and immunohistochemical evaluation of the specimens, FD proved to have a better preservation of its structure than FF.
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