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The usefulness of lutein/trypan blue vital dye for the staining of corneal endothelium: a pilot study on DMEK pretreated tissues
PURPOSE: The study aims to evaluate the usefulness of lutein/trypan blue vital dye for the staining of corneal tissues and endothelium–Descemet membrane (EDM) for Descemet membrane endothelial keratoplasty (DMEK). METHODS: Sixteen human corneal tissues (Eye Bank, Rome, Italy) were used. Corneal endo...
Autores principales: | , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Springer Berlin Heidelberg
2022
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9707145/ https://www.ncbi.nlm.nih.gov/pubmed/36445446 http://dx.doi.org/10.1007/s00417-022-05909-x |
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author | Gisoldi, Rossella Colabelli Lodato, Gemma Balzamino, Bijorn Omar Esposito, Graziana Micera, Alessandra Pocobelli, Augusto |
author_facet | Gisoldi, Rossella Colabelli Lodato, Gemma Balzamino, Bijorn Omar Esposito, Graziana Micera, Alessandra Pocobelli, Augusto |
author_sort | Gisoldi, Rossella Colabelli |
collection | PubMed |
description | PURPOSE: The study aims to evaluate the usefulness of lutein/trypan blue vital dye for the staining of corneal tissues and endothelium–Descemet membrane (EDM) for Descemet membrane endothelial keratoplasty (DMEK). METHODS: Sixteen human corneal tissues (Eye Bank, Rome, Italy) were used. Corneal endothelium was tested at 25 s (T0), 1 min (T1), 2 min (T2), and 4 min (T4) from dye addition. Staining intensity and cell counting were compared. Stripped EDM was analyzed for selected apoptotic (AP, caspases, BCL2, BAX) and differentiation (VEGF-A, TGF-β1RI, SMAD3/7, SMA) targets and changes in target expression. Protein extracts were analyzed through SDS-PAGE/IB. RESULTS: Although trypan blue staining produced the same color intensity of lutein/trypan blue dye in half the time, lutein/trypan blue reached a good and adequate color intensity at T4, which persisted even on excised and washed EDM grafts. Lutein/trypan blue-stained EDM showed a reduced number of blue-stained cells and AP immunoreactivity was significantly reduced in the same samples. An increased BCL2 transcript and a reduced BAX transcript were detected in lutein/trypan blue-stained EDM. No significant changes were observed for the main effector caspases (3/9) upon both treatments and the target genes representative of endothelial cell trans-differentiation (TGF-β1RI, SMAD3/7, SMA). A trend in vascular endothelial growth factor (VEGF-A) regulation was observed in lutein/trypan blue-treated EDM grafts. CONCLUSION: Obtained results suggest that lutein/trypan blue dye deserves attention in the DMEK field and support the potential routine use of this dye as a valid alternative to trypan blue for all procedures devoted to the assessment of endothelial cell viability and visualization of EDM graft before DMEK grafting. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s00417-022-05909-x. |
format | Online Article Text |
id | pubmed-9707145 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2022 |
publisher | Springer Berlin Heidelberg |
record_format | MEDLINE/PubMed |
spelling | pubmed-97071452022-11-29 The usefulness of lutein/trypan blue vital dye for the staining of corneal endothelium: a pilot study on DMEK pretreated tissues Gisoldi, Rossella Colabelli Lodato, Gemma Balzamino, Bijorn Omar Esposito, Graziana Micera, Alessandra Pocobelli, Augusto Graefes Arch Clin Exp Ophthalmol Cornea PURPOSE: The study aims to evaluate the usefulness of lutein/trypan blue vital dye for the staining of corneal tissues and endothelium–Descemet membrane (EDM) for Descemet membrane endothelial keratoplasty (DMEK). METHODS: Sixteen human corneal tissues (Eye Bank, Rome, Italy) were used. Corneal endothelium was tested at 25 s (T0), 1 min (T1), 2 min (T2), and 4 min (T4) from dye addition. Staining intensity and cell counting were compared. Stripped EDM was analyzed for selected apoptotic (AP, caspases, BCL2, BAX) and differentiation (VEGF-A, TGF-β1RI, SMAD3/7, SMA) targets and changes in target expression. Protein extracts were analyzed through SDS-PAGE/IB. RESULTS: Although trypan blue staining produced the same color intensity of lutein/trypan blue dye in half the time, lutein/trypan blue reached a good and adequate color intensity at T4, which persisted even on excised and washed EDM grafts. Lutein/trypan blue-stained EDM showed a reduced number of blue-stained cells and AP immunoreactivity was significantly reduced in the same samples. An increased BCL2 transcript and a reduced BAX transcript were detected in lutein/trypan blue-stained EDM. No significant changes were observed for the main effector caspases (3/9) upon both treatments and the target genes representative of endothelial cell trans-differentiation (TGF-β1RI, SMAD3/7, SMA). A trend in vascular endothelial growth factor (VEGF-A) regulation was observed in lutein/trypan blue-treated EDM grafts. CONCLUSION: Obtained results suggest that lutein/trypan blue dye deserves attention in the DMEK field and support the potential routine use of this dye as a valid alternative to trypan blue for all procedures devoted to the assessment of endothelial cell viability and visualization of EDM graft before DMEK grafting. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s00417-022-05909-x. Springer Berlin Heidelberg 2022-11-29 2023 /pmc/articles/PMC9707145/ /pubmed/36445446 http://dx.doi.org/10.1007/s00417-022-05909-x Text en © The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature 2022, Springer Nature or its licensor (e.g. a society or other partner) holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law. This article is made available via the PMC Open Access Subset for unrestricted research re-use and secondary analysis in any form or by any means with acknowledgement of the original source. These permissions are granted for the duration of the World Health Organization (WHO) declaration of COVID-19 as a global pandemic. |
spellingShingle | Cornea Gisoldi, Rossella Colabelli Lodato, Gemma Balzamino, Bijorn Omar Esposito, Graziana Micera, Alessandra Pocobelli, Augusto The usefulness of lutein/trypan blue vital dye for the staining of corneal endothelium: a pilot study on DMEK pretreated tissues |
title | The usefulness of lutein/trypan blue vital dye for the staining of corneal endothelium: a pilot study on DMEK pretreated tissues |
title_full | The usefulness of lutein/trypan blue vital dye for the staining of corneal endothelium: a pilot study on DMEK pretreated tissues |
title_fullStr | The usefulness of lutein/trypan blue vital dye for the staining of corneal endothelium: a pilot study on DMEK pretreated tissues |
title_full_unstemmed | The usefulness of lutein/trypan blue vital dye for the staining of corneal endothelium: a pilot study on DMEK pretreated tissues |
title_short | The usefulness of lutein/trypan blue vital dye for the staining of corneal endothelium: a pilot study on DMEK pretreated tissues |
title_sort | usefulness of lutein/trypan blue vital dye for the staining of corneal endothelium: a pilot study on dmek pretreated tissues |
topic | Cornea |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9707145/ https://www.ncbi.nlm.nih.gov/pubmed/36445446 http://dx.doi.org/10.1007/s00417-022-05909-x |
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