June 2020
Volume 61, Issue 7
Free
ARVO Annual Meeting Abstract  |   June 2020
Modulation of autophagy-related genes in healthy, keratoconus and injured corneas
Author Affiliations & Notes
  • Lynn M. Martin
    Harry S. Truman Memorial Veterans’ Hospital, Columbia, Missouri, United States
    VHC and Biomedical Sciences, University of Missouri College of Veterinary Medicine, Columbia, Missouri, United States
  • Rayne Lim
    Harry S. Truman Memorial Veterans’ Hospital, Columbia, Missouri, United States
    VHC and Biomedical Sciences, University of Missouri College of Veterinary Medicine, Columbia, Missouri, United States
  • Nishant Rajiv Sinha
    Harry S. Truman Memorial Veterans’ Hospital, Columbia, Missouri, United States
    VHC and Biomedical Sciences, University of Missouri College of Veterinary Medicine, Columbia, Missouri, United States
  • Alexandria Hofmann
    VHC and Biomedical Sciences, University of Missouri College of Veterinary Medicine, Columbia, Missouri, United States
  • Ratnakar Tripathi
    Harry S. Truman Memorial Veterans’ Hospital, Columbia, Missouri, United States
    VHC and Biomedical Sciences, University of Missouri College of Veterinary Medicine, Columbia, Missouri, United States
  • Sydney Green
    VHC and Biomedical Sciences, University of Missouri College of Veterinary Medicine, Columbia, Missouri, United States
  • Sabeeh Kamil
    Harry S. Truman Memorial Veterans’ Hospital, Columbia, Missouri, United States
    VHC and Biomedical Sciences, University of Missouri College of Veterinary Medicine, Columbia, Missouri, United States
  • DeAna Grant
    VHC and Biomedical Sciences, University of Missouri College of Veterinary Medicine, Columbia, Missouri, United States
  • Philip J. Johnson
    VHC and Biomedical Sciences, University of Missouri College of Veterinary Medicine, Columbia, Missouri, United States
  • Mikayla Fraunfelder
    VHC and Biomedical Sciences, University of Missouri College of Veterinary Medicine, Columbia, Missouri, United States
  • Rajiv R Mohan
    Harry S. Truman Memorial Veterans’ Hospital, Columbia, Missouri, United States
    VHC and Biomedical Sciences, University of Missouri College of Veterinary Medicine, Columbia, Missouri, United States
  • Footnotes
    Commercial Relationships   Lynn Martin, None; Rayne Lim, None; Nishant Sinha, None; Alexandria Hofmann, None; Ratnakar Tripathi, None; Sydney Green, None; Sabeeh Kamil, None; DeAna Grant, None; Philip Johnson, None; Mikayla Fraunfelder, None; Rajiv Mohan, None
  • Footnotes
    Support  RO1EY17294 National Eye Institute, NIH, Bethesda, Maryland, USA, 1I01BX00035701 Veteran Health Affairs, Washington DC USA, the Ruth M. Kraeuchi Missouri Endowment Chair Ophthalmology Fund.
Investigative Ophthalmology & Visual Science June 2020, Vol.61, 2909. doi:
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      Lynn M. Martin, Rayne Lim, Nishant Rajiv Sinha, Alexandria Hofmann, Ratnakar Tripathi, Sydney Green, Sabeeh Kamil, DeAna Grant, Philip J. Johnson, Mikayla Fraunfelder, Rajiv R Mohan; Modulation of autophagy-related genes in healthy, keratoconus and injured corneas. Invest. Ophthalmol. Vis. Sci. 2020;61(7):2909.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Purpose : Autophagy plays a key role in cellular homeostatic maintenance by discarding and recycling cytoplasmic proteins and dysfunctional organelles. Recently, we identified the involvement of autophagy in corneal wound healing and fibrosis. This study investigated (a) the expression of autophagy-related genes (Atgs) in healthy and diseased human and rabbit corneas and (b) autophagy's response to select treatments and contribution to corneal wound healing using donor human corneas and primary human corneal stromal fibroblast (hCSFs) cells.

Methods : Donor healthy and diseased human corneas, normal and wounded rabbit corneas and primary cultures of hCSFs cells generated from healthy human corneas were utilized for experiments. Cultures were grown in MEM medium with 1% or 10% FBS in +/- TGFβ1 or +/- starvation (FBS), +/- chloroquine or +/- rapamycin treatment. Serial corneal tissue sections prepared from healthy and diseased tissues (human and rabbit) were used to study differential Atg expression. Immunofluorescence, 2D/3D confocal microscopy, western blotting, qPCR, MTT, TUNEL, LDH, wound-scratch, TEM and other commercial assays were used for wound healing studies.

Results : Expression of LC3, BECN1 and SQSTM1 Atgs was detected in the selected human and rabbit corneas in vitro and in vivo. Starvation conditions decreased SQSTM1 and BCN1 and increased LC3-II protein expression in vitro as compared to the control (untreated hCSFs). TEM identified autophagosomal structures within primary hCSFs. Analysis of mRNA for treatment conditions and TGFβ1 stimulation regarding the levels of Atgs in hCSFs compared to the control (untreated hCSFs) is underway. Results of in vivo wound healing experiments are pending.

Conclusions : Autophagy mechanisms are used by the cornea for cellular homeostasis and become altered during corneal wound repair. Ongoing additional studies will provide more specific functional roles of Atgs in corneal wound healing and disease modulation.

This is a 2020 ARVO Annual Meeting abstract.

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