Ex Vivo Corneal Organ Culture Model for Wound Healing Studies

Castro, Nileyma; Gillespie, Stephanie R.; Bernstein, Audrey M.

doi:10.3791/58562-v

Cited by 9 publications

(4 citation statements)

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“…The absence of these factors may not only affect the state of the host tissue but also the localization and environment of the bacteria, potentially altering bacterial gene expression in response to their surroundings. Although there is evidence supporting tissue viability and epithelial healing in cultured corneas ex vivo in the literature [89,90], the 3 day storage period may introduce variability and deviate from in vivo conditions due to lack of host factors and slower healing process. Moreover, the presence of opacity after incubation and before infection highlights the need to consider the dynamic changes that occur in corneal tissue over time post-excision.…”

Section: Discussionmentioning

confidence: 99%

Increased tolerance to commonly used antibiotics in a Pseudomonas aeruginosa ex vivo porcine keratitis model

Okurowska,

Monk,

Karunakaran

2024

Microbiology

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Introduction. Bacterial keratitis, particularly caused by Pseudomonas aeruginosa, is challenging to treat because of multi-drug tolerance, often associated with the formation of biofilms. Antibiotics in development are typically evaluated against planktonic bacteria in a culture medium, which may not accurately represent the complexity of infections in vivo. Hypothesis/Gap Statement. Developing a reliable, economic ex vivo keratitis model that replicates some complexity of tissue infections could facilitate a deeper understanding of antibiotic efficacy, thus aiding in the optimization of treatment strategies for bacterial keratitis. Methodology. Here we investigated the efficacy of three commonly used antibiotics (gentamicin, ciprofloxacin and meropenem) against Pseudomonas aeruginosa cytotoxic strain PA14 and invasive strain PA01 using an ex vivo porcine keratitis model. Results. Both strains of P. aeruginosa were susceptible to the MIC of the three tested antibiotics. However, significantly higher concentrations were necessary to inhibit bacterial growth in the minimum biofilm eradication concentration (MBEC) assay, with both strains tolerating concentrations greater than 512 mg l−1 of meropenem. When MIC and higher concentrations than MBEC (1024 mg l−1) of antibiotics were applied, ciprofloxacin exhibited the highest potency against both P. aeruginosa strains, followed by meropenem, while gentamicin showed the least potency. Despite this, none of the antibiotic concentrations used effectively cleared the infection, even after 18 h of continuous exposure. Conclusions. Further exploration of antibiotic concentrations and aligning dosing with clinical studies to validate the model is needed. Nonetheless, our ex vivo porcine keratitis model could be a valuable tool for assessing antibiotic efficacy.

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Section: Discussionmentioning

confidence: 99%

Increased tolerance to commonly used antibiotics in a Pseudomonas aeruginosa ex vivo porcine keratitis model

Okurowska,

Monk,

Karunakaran

2024

Microbiology

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“…Typically, tears supply growth factors to aid the healing process (Klenkler et al, 2007) and this is a limitation of my ex vivo cornea model. Studies on ex vivo porcine corneas showed that corneal stratified squamous epithelium could regenerate within 72 hours (Schumann et al, 2021) to six days (Castro et al, 2019). In ex vivo human corneas, complete reepithelialization was observed after seven days, resulting in five to six layers of the stratified squamous epithelium.…”

Section: Discussionmentioning

confidence: 99%

“…The different healing rates observed in ex vivo corneal models may be related to several factors. The media composition, size, and depth of the wound and rocking of the cornea during incubation affect the recovery rate (Castro et al, 2019; Deshpande et al, 2015). Our study experiments were terminated once wound closure in untreated corneas was achieved, regardless of the total epithelial thickness.…”

Section: Discussionmentioning

confidence: 99%

The effect of ciprofloxacin and gentamicin on wound healing inex vivosheep cornea model

Okurowska,

Suarez,

Karunakaran

2023

Preprint

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PurposeOur objective was to assess the efficacy of anex vivosheep corneal model as an alternative for live animal testing in screening drug cytotoxicity. In pursuit of this goal, we investigated the impact of two commonly used topical antibiotics, ciprofloxacin and gentamicin, on wound healing. Furthermore, we examined different antibiotic dosages and dosing regimens to understand their effects comprehensively.MethodsThe epithelium onex vivo sheep corneas was removed with a scalpel, and the area was treated with ciprofloxacin (0.1, 0.3, and 1 mg mL-1), gentamicin (0.25, 1, and 3 mg mL-1), or phosphate-buffered saline (control). The corneas were exposed to treatments continuously or twice daily for ten minutes. Wound closure was observed by fluorescein retention and histological staining.ResultsUntreated corneas healed within 41 hours. Continuous exposure to both ciprofloxacin and gentamicin significantly reduced the corneal healing ability in a time- and concentration-dependent manner. Overall, ciprofloxacin was found to be more toxic than gentamycin. However, this model showed that the corneal epithelium could heal effectively when both antibiotics were administered intermittently.ConclusionCiprofloxacin demonstrated greater inhibition of wound healing compared to gentamicin, aligning within vivostudies. The administration of drops several times daily mitigated the toxic effects of antibiotics. Theex vivosheep wound healing model holds promise as an alternative approach toin vivotoxicity testing, enabling the swift evaluation of novel antimicrobial treatments and eye drop additives.

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“…The porcine eye was selected as an ex vivo animal model to simulate Acanthamoeba keratitis in humans. With numerous anatomical similarities to human eyes [ 13 , 14 ], porcine eyes have been utilized as a model for studying wound healing, as demonstrated by Nileyma Castro et al [ 15 ]. Porcine eyeballs were obtained from a nearby slaughterhouse.…”

Section: Methodsmentioning

confidence: 99%

Development of an Ex Vivo Porcine Eye Model for Exploring the Pathogenicity of Acanthamoeba

Shi,

Sung,

Huang

et al. 2024

Microorganisms

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Acanthamoeba, a widely distributed free-living amoeba found in various environments, is an opportunistic pathogen responsible for causing Acanthamoeba keratitis, a condition that may lead to blindness. However, identifying the pathogenicity of Acanthamoeba is challenging due to its complex life cycle, ability to adapt to different environments, variable virulence factors, and intricate interactions with the host immune system. Additionally, the development of an effective model for studying Acanthamoeba pathogenicity is limited, hindering a comprehensive understanding of the mechanisms underlying its virulence and host interactions. The aim of this study was to develop an ex vivo model for Acanthamoeba infection using porcine eyeballs and to evaluate the pathogenicity of the Acanthamoeba isolates. Based on slit lamp and biopsy analysis, the developed ex vivo model is capable of successfully infecting Acanthamoeba within 3 days. Histopathological staining revealed that clinical isolates of Acanthamoeba exhibited greater corneal stroma destruction and invasion in this model than environmental isolates. Our results highlight the importance of an ex vivo porcine eye model in elucidating the pathogenesis of Acanthamoeba infection and its potential implications for understanding and managing Acanthamoeba-related ocular diseases.

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Ex Vivo Corneal Organ Culture Model for Wound Healing Studies

Cited by 9 publications

References 0 publications

Increased tolerance to commonly used antibiotics in a Pseudomonas aeruginosa ex vivo porcine keratitis model

Increased tolerance to commonly used antibiotics in a Pseudomonas aeruginosa ex vivo porcine keratitis model

The effect of ciprofloxacin and gentamicin on wound healing inex vivosheep cornea model

Development of an Ex Vivo Porcine Eye Model for Exploring the Pathogenicity of Acanthamoeba

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