Dataset Information

The usefulness of lutein/trypan blue vital dye for the staining of corneal endothelium: a pilot study on DMEK pretreated tissues.

ABSTRACT:

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.

SUBMITTER: Gisoldi RC

PROVIDER: S-EPMC9707145 | biostudies-literature | 2022 Nov

REPOSITORIES: biostudies-literature

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Publications

The usefulness of lutein/trypan blue vital dye for the staining of corneal endothelium: a pilot study on DMEK pretreated tissues.

Gisoldi Rossella Colabelli RC Lodato Gemma G Balzamino Bijorn Omar BO Esposito Graziana G Micera Alessandra A Pocobelli Augusto A

Graefe's archive for clinical and experimental ophthalmology = Albrecht von Graefes Archiv fur klinische und experimentelle Ophthalmologie 20221129 5

<h4>Purpose</h4>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).<h4>Methods</h4>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 apopt ...[more]

PMID: 36445446

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Project description:BackgroundWe evaluated the visual outcomes and complications of "endothelium-out" and "endothelium-in" Descemet membrane endothelial keratoplasty (DMEK) graft insertion techniques.Materials and methodsElectronic searches were conducted in CENTRAL, Cochrane databases, PubMed, EMBASE, ClinicalTrials.gov. Study designs included clinical trials, comparative observational studies, and large case series (≥25 eyes). PRISMA guidelines were used for abstracting data and synthesis. Random-effects models were employed for meta-analyses.Results21,323 eyes (95 studies) were included. Eighty-six studies reported on "endothelium-out" techniques; eight studies reported on "endothelium-in" techniques. One study compared "endothelium-out" to "endothelium-in" techniques. Eighteen "endothelium-out" studies reported that 42.5-85% of eyes achieved best-corrected visual acuity (BCVA) ≥20/25 at 6 months; pooled proportion of eyes achieving BCVA ≥20/25 at 6 months was 58.7% (95% CI 49.4-67.7%,15 studies). Three "endothelium-in" studies reported that 44.7-87.5% of eyes achieved BCVA of ≥20/25 at 6 months; pooled proportion of eyes achieving BCVA ≥20/25 at 6 months was 62.4% (95% CI 33.9-86.9%). Pooled mean endothelial cell loss was lower in the "endothelium-in" studies (28.1 ± 1.3%, 7 studies) compared to "endothelium-out" studies (36.3 ± 6.9%,10 studies) at 6 months (p = 0.018). Graft re-bubbling rates were higher in the "endothelium-out" studies (26.2%, 95% CI 21.9-30.9%, 74 studies) compared to "endothelium-in" studies (16.5%, 95% CI 8.5-26.4%, 6 studies), although statistical significance was not reached (p = 0.440). Primary graft failure rates were comparable between the two groups (p = 0.552). Quality of evidence was considered low and significant heterogeneity existed amongst the studies.ConclusionReported rates of endothelial cell loss were lower in "endothelium-in" DMEK studies at 6 months compared to "endothelium-out" studies. Outcomes of "endothelium-in" techniques were otherwise comparable to those reported in "endothelium-out" studies. Given the technical challenges encountered in "endothelium-out" procedures, surgeons may consider "endothelium-in" techniques designed for easier intra-operative DMEK graft unfolding. "Endothelium-in" studies evaluating outcomes at longer time points are required before conclusive comparisons between the two techniques can be drawn.

Project description:Background and purposeCorneal fluorescein staining is one of the most important diagnostic tests in dry eye disease (DED). Nevertheless, the result of this examination is depending on the grader. So far, there is no method for an automated quantification of corneal staining commercially available. Aim of this study was to develop a software-assisted grading algorithm and to compare it with a group of human graders with variable clinical experience in patients with DED.MethodsFifty images of eyes stained with 2 µl of 2% fluorescein presenting different severity of superficial punctate keratopathy in patients with DED were taken under standardized conditions. An algorithm for detecting and counting superficial punctate keratitis was developed using ImageJ with a training dataset of 20 randomly picked images. Then, the test dataset of 30 images was analyzed (1) by the ImageJ algorithm and (2) by 22 graders, all ophthalmologists with different levels of experience. All graders evaluated the images using the Oxford grading scheme for corneal staining at baseline and after 6-8 weeks. Intrarater agreement was also evaluated by adding a mirrored version of all original images into the set of images during the 2nd grading.ResultsThe count of particles detected by the algorithm correlated significantly (n = 30; p < 0.01) with the estimated true Oxford grade (Sr = 0,91). Overall human graders showed only moderate intrarater agreement (K = 0,426), while software-assisted grading was always the same (K = 1,0). Little difference was found between specialists and non-specialists in terms of intrarater agreement (K = 0,436 specialists; K = 0,417 non-specialists). The highest interrater agreement was seen with 75,6% in the most experienced grader, a cornea specialist with 29 years of experience, and the lowest was seen in a resident with 25,6% who had only 2 years of experience.ConclusionThe variance in human grading of corneal staining - if only small - is likely to have only little impact on clinical management and thus seems to be acceptable. While human graders give results sufficient for clinical application, software-assisted grading of corneal staining ensures higher consistency and thus is preferrable for re-evaluating patients, e.g., in clinical trials.

Dataset Information

The usefulness of lutein/trypan blue vital dye for the staining of corneal endothelium: a pilot study on DMEK pretreated tissues.

Purpose

Methods

Results

Conclusion

Publications

The usefulness of lutein/trypan blue vital dye for the staining of corneal endothelium: a pilot study on DMEK pretreated tissues.

Similar Datasets

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