Protective Effects of Deswelling on Stromal Collagen Denaturation After a Corneal Femtosecond Laser Cut

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Porcine Cornea Storage Ex Vivo Model as an Alternative to Human Donor Tissues for Investigations of Endothelial Layer Preservation

Purpose: Due to the growing shortage of human corneas for research, we developed a porcine cornea storage model with qualitative features comparable to human tissues.Methods: We established a decontamination procedure for porcine eye bulbs to ensure corneal storage at 31 degrees C to 35 degrees C for up to 28 days without contamination. We compared human and porcine corneas under hypothermic (2-8 degrees C) or culture (31-35 degrees C) conditions for central corneal thickness (CCT), corneal transparency, endothelialmorphology, endothelial cell density (ECD), and a novelmethod to quantifywhole endothelialmortality. We also examined portions of lamellar tissues consisting of Descemet's membrane and endothelial cells under the microscope after Alizarin red staining.Results: Our decontamination procedure reduced corneal contamination from 94% (control corneas without decontamination) to 18% after 28 days of storage at 31 degrees C to 35 degrees C. ECD, CCT, transparency, and morphology were significantly higher in porcine corneas than in human corneas at day 0. Nevertheless, the qualitative parameters of porcine and human corneas showed comparable trends under both investigated storage conditions for up to 14 days.Conclusions: The presented corneal storage model provides a reliable alternative to human tissues for preliminary corneal investigations.Translational Relevance: The porcine cornea storagemodel can be used to investigate the efficacy and safety of new media, substances, or storage conditions. Furthermore, themethod developed to assess the percentage of endothelialmortality is tissue conservative and can be used in eye banks to monitor endothelial mortality during storage of tissues intended for transplantation

Safety of silicone oils as intraocular medical device: An in vitro cytotoxicity study

This study aimed to assess the cytotoxic e!ect of low molecular weight components (LMWC) and conventional silicone oils (SOs) 1000 cSt with di!erent degree of puri"cation (raw, intermediate, and puri"ed) using in vitro cytotoxicity tests. Direct contact cytotoxicity tests were performed in BALB 3T3 and human retinal pigment epithelial cells (ARPE-19) using quantitative and qualitative evaluation according to the ISO 10993-5 (2009) standards. Conventional SOs 1000 cSt in form of raw, intermediate (intermediate product obtained during distillation process), and puri"ed SO ("nal product after distillation) and a concentrate of LMWC (including siloxane chains with molecular weight up to 1557 g/mol) were directly applied to 100% of cell layer area for 24 h. Cell viability was quanti"ed using 3-(4,5-dimethylthiazole-2-yl)-2,5–28 diphenyltetrazolium bromide (MTT) and neutral red uptake assays in ARPE-19 and BALB3T3, respectively. All tested samples, including the concentrate of LMWC, resulted to be not cytotoxic according to ISO 10993-5 in both qualitative and quantitative evaluations. However, the cellular viability was signi"cantly higher in the intermediate and puri"ed SO compared with the raw SO in ARPE-19 cells. No reduction in cell viability was detected by LMWC. The absence of cytotoxicity was observed for all tested samples in both BALB3T3 and ARPE-19 after 24 h of application. A direct cytotoxic e!ect is not likely to be involved in the potential complications related to SO and LMWC. Long-term potential adverse e!ects of SO could be related to the raw material and to di!erent concentrations of LMWC

Residual Antibiotics in Decontaminated Human Cardiovascular Tissues Intended for Transplantation and Risk of Falsely Negative Microbiological Analyses

<div><p>We investigated the presence of antibiotics in cryopreserved cardiovascular tissues and cryopreservation media, after tissue decontamination with antibiotic cocktails, and the impact of antibiotic residues on standard tissue bank microbiological analyses. Sixteen cardiovascular tissues were decontaminated with bank-prepared cocktails and cryopreserved by two different tissue banks according to their standard operating procedures. Before and after decontamination, samples underwent microbiological analysis by standard tissue bank methods. Cryopreserved samples were tested again with and without the removal of antibiotic residues using a RESEP tube, after thawing. Presence of antibiotics in tissue homogenates and processing liquids was determined by a modified agar diffusion test. All cryopreserved tissue homogenates and cryopreservation media induced important inhibition zones on both <i>Staphylococcus aureus</i>- and <i>Pseudomonas aeruginosa</i>-seeded plates, immediately after thawing and at the end of the sterility test. The RESEP tube treatment markedly reduced or totally eliminated the antimicrobial activity of tested tissues and media. Based on standard tissue bank analysis, 50% of tissues were found positive for bacteria and/or fungi, before decontamination and 2 out of 16 tested samples (13%) still contained microorganisms after decontamination. After thawing, none of the 16 cryopreserved samples resulted positive with direct inoculum method. When the same samples were tested after removal of antibiotic residues, 8 out of 16 (50%) were contaminated. Antibiotic residues present in tissue allografts and processing liquids after decontamination may mask microbial contamination during microbiological analysis performed with standard tissue bank methods, thus resulting in false negatives.</p></div

Results of microbiological analysis.

<p>Positive (contaminated) samples determined by microbiological analysis performed by standard tissue bank methods and the RESEP tube method on tissue and liquid samples of human cardiovascular allografts processed by two cardiovascular tissue banks and tested before and after decontamination and after thawing.</p><p><i>* liquid samples were inoculated in BacT/ALERT blood culture vials.</i></p><p>Results of microbiological analysis.</p

Presence of antibiotic residues in decontaminated and cryopreserved tissue homogenates and cryopreservation media.

<p>Agar diffusion assays performed using <i>Staphylococcus aureus-</i>seeded plates (A) and <i>Pseudomonas aeruginosa</i>-seeded plates (B), immediately after thawing (initial) and after 14 days of sterility testing (final) on tissues and media undergoing decontamination procedures with Cocktail 1 and Cocktail 2, prepared by tissue banks, and BASE.128 as a control. The plot legend on <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0112679#pone-0112679-g001" target="_blank">Figure 1A</a> also applies to <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0112679#pone-0112679-g001" target="_blank">Figure 1B</a>.</p

Rose Bengal Photodynamic Antimicrobial Therapy: A review of the intermediate term clinical and surgical outcomes

To evaluate the intermediate term clinical outcomes of Rose Bengal Photodynamic Antimicrobial Therapy (RB-PDAT) for infectious keratitis. Secondarily, to evaluate the surgical outcomes of individuals that underwent optical keratoplasty after RB-PDAT. Retrospective cohort study. Retrospective chart review of 31 eyes from 30 consecutive individuals with infectious keratitis refractory to standard medical therapy who underwent RB-PDAT at the Bascom Palmer Eye Institute between January 2016 and July 2020. Data collected included demographics, risk factors for infectious keratitis, microbiological diagnosis, Best Spectacle-Corrected Visual Acuity (BCVA), clinical outcomes after RB-PDAT and complication rates post-keratoplasty. RB-PDAT was performed as described in previous studies. Graft survival was evaluated using Kaplan Meier curves with log-ranks in individuals that underwent keratoplasty after RB-PDAT. Mean age of the study population was 53±18.0 years. 70% were female; 53.3% self-identified as non-Hispanic White; 43.3% as Hispanic. Mean follow-up time was 28.0±14.4 months. Risk factors included contact lens use (80.6%), history of infectious keratitis (19.3%), and ocular surface disease (16.1%). Cultures were positive for Acanthamoeba (51.6%), Fusarium (12.9%), and Pseudomonas (6.5%). 22.5% of individuals with Acanthamoeba infection were treated with concomitant Miltefosine. Clinical resolution was achieved in 77.4% of individuals on average 2.72±1.85 months after RB-PDAT with 22.5% requiring therapeutic penetrating keratoplasties and 54.8% subsequently requiring optical penetrating keratoplasties. At 2 years, the overall probability of graft survival was 78.7% and the graft failure rate was 21.3%. RB-PDAT is a potential adjunct therapy for infectious keratitis that may reduce the need for a therapeutic penetrating keratoplasty. Cases that undergo keratoplasty after RB-PDAT may have a higher probability of graft survival at one year postoperatively

Clinical Features and Treatment Outcomes of Carbapenem-Resistant Pseudomonas aeruginosa Keratitis

Evaluation of the microbiological diagnostic profile of multidrug-resistant Pseudomonas aeruginosa keratitis and potential management with rose bengal-photodynamic antimicrobial therapy (RB-PDAT) is important. To document the disease progression of carbapenemase-resistant P aeruginosa keratitis after an artificial tear contamination outbreak. This retrospective observation case series included 9 patients 40 years or older who presented at Bascom Palmer Eye Institute and had positive test results for multidrug-resistant P aeruginosa keratitis between January 1, 2022, and October 31, 2023. Evaluation of type III secretion phenotype, carbapenemase-resistance genes blaGES and blaVIM susceptibility to antibiotics, and in vitro and in vivo outcomes of RB-PDAT against multidrug-resistant P aeruginosa keratitis. Among the 9 patients included in the analysis (5 women and 4 men; mean [SD] age, 73.4 [14.0] years), all samples tested positive for exoU and carbapenemase-resistant blaVIM and blaGES genes. Additionally, isolates were resistant to carbapenems as indicated by minimum inhibitory concentration testing. In vitro efficacy of RB-PDAT indicated its potential application for treating recalcitrant cases. These cases highlight the rapid progression and challenging management of multidrug-resistant P aeruginosa. Two patients were treated with RB-PDAT as an adjuvant to antibiotic therapy and had improved visual outcomes. This case series highlights the concerning progression in resistance and virulence of P aeruginosa and emphasizes the need to explore alternative therapies like RB-PDAT that have broad coverage and no known antibiotic resistance. The findings support further investigation into the potential effects of RB-PDAT for other multidrug-resistant microbes