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Research Article
Gary Peh
Singapore Eye Research Institute
Corresponding Author
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Following corneal transplantation, there is an initial decline in corneal endothelial cells (CECs) following graft preparation and surgery. Monitoring post-transplantation is only possible months after surgery by specular microscopy, which has a limited field of view. We have developed a labelling approach using 1,1’-dioctadecyl-3,3,3’,3’-tetramethylindotricarbocyanine iodide (DIR) dye solution, that enabled tracking of labelled CECs in vivo for at least one month. Initial in vitro optimization of dye concentration, cellular toxicity and real-time cell migration was assessed using propagated primary CECs. Subsequent in vivo evaluation of cellular labelling was assessed within a rabbit wound healing model. Finally, real-time visualization of human cadaver donor tissue incubated in DIR transplanted into rabbits was achieved using the Heidelberg Spectralis. Results revealed detectable fluorescence increased with concentration to a plateau of 100µg/ml, with no toxicity of CECs at any concentration evaluated. DIR-labelled CECs were detectable in vivo upto 1 month, and transplanted labelled donor graft could be visualized and were trackable in vivo. Acute endothelial rejection in 1 rabbit was evidenced by detectable DIR positive cells within the anterior chamber. DIR imaging allowed for detailed imaging of the transplanted corneal endothelium, and enabled non-invasive observation of the corneal endothelial morphology following transplantation.
Keywords
corneal transplantation, in vivo imaging, pre-clinical animal model, 1,1’-dioctadecyl-3,3,3’,3’-tetramethylindotricarbocyanine iodide (DIR)
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This preprint is under consideration at Scientific Reports. A preprint is a preliminary version of a manuscript that has not completed peer review at a journal. Research Square does not conduct peer review prior to posting preprints. The posting of a preprint on this server should not be interpreted as an endorsement of its validity or suitability for dissemination as established information or for guiding clinical practice.
Learn more about In Review
Research Article
Gary Peh
Singapore Eye Research Institute
Corresponding Author
Badges
Prescreen
This manuscript passed our Prescreen assessment
Complete author information
Appropriate declaration statements
Potential risks to human health
Prescreen
Peer Review Timeline
CURRENT STATUS: Under Review
Version 1
Posted 10 Mar, 2021
No community comments so far
Review #1 received
Received 28 Mar, 2021
Review #2 received
Received 28 Mar, 2021
Reviewer #5 agreed
On 05 Mar, 2021
Reviewer #1 agreed
On 05 Mar, 2021
Reviewer #7 agreed
On 05 Mar, 2021
Reviewer #4 agreed
On 05 Mar, 2021
Reviewer #9 agreed
On 05 Mar, 2021
Reviewer #8 agreed
On 05 Mar, 2021
Reviewer #10 agreed
On 05 Mar, 2021
Reviewer #3 agreed
On 05 Mar, 2021
Reviewer #2 agreed
On 05 Mar, 2021
Reviewers invited
Invitations sent on 05 Mar, 2021
Editor assigned
On 02 Mar, 2021
Editor invited
On 02 Mar, 2021
Submission checks complete
On 02 Mar, 2021
First submitted
On 16 Feb, 2021
License:
This work is licensed under a CC BY 4.0 License. Read Full License
Following corneal transplantation, there is an initial decline in corneal endothelial cells (CECs) following graft preparation and surgery. Monitoring post-transplantation is only possible months after surgery by specular microscopy, which has a limited field of view. We have developed a labelling approach using 1,1’-dioctadecyl-3,3,3’,3’-tetramethylindotricarbocyanine iodide (DIR) dye solution, that enabled tracking of labelled CECs in vivo for at least one month. Initial in vitro optimization of dye concentration, cellular toxicity and real-time cell migration was assessed using propagated primary CECs. Subsequent in vivo evaluation of cellular labelling was assessed within a rabbit wound healing model. Finally, real-time visualization of human cadaver donor tissue incubated in DIR transplanted into rabbits was achieved using the Heidelberg Spectralis. Results revealed detectable fluorescence increased with concentration to a plateau of 100µg/ml, with no toxicity of CECs at any concentration evaluated. DIR-labelled CECs were detectable in vivo upto 1 month, and transplanted labelled donor graft could be visualized and were trackable in vivo. Acute endothelial rejection in 1 rabbit was evidenced by detectable DIR positive cells within the anterior chamber. DIR imaging allowed for detailed imaging of the transplanted corneal endothelium, and enabled non-invasive observation of the corneal endothelial morphology following transplantation.
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