September 2016
Volume 57, Issue 12
Open Access
ARVO Annual Meeting Abstract  |   September 2016
Ultrastructural evidence of defective regeneration of the epithelial basement membrane as a major factor in the generation and persistence of corneal myofibroblasts after corneal injury in rabbits
Author Affiliations & Notes
  • Gustavo K. Marino
    Ophthalmology, Cleveland Clinic, Cleveland, Ohio, United States
  • Abirami Santhanam
    Ophthalmology, Cleveland Clinic, Cleveland, Ohio, United States
  • K P Connie Tam
    Ophthalmology, Cleveland Clinic, Cleveland, Ohio, United States
  • Steven E Wilson
    Ophthalmology, Cleveland Clinic, Cleveland, Ohio, United States
  • Footnotes
    Commercial Relationships   Gustavo Marino, None; Abirami Santhanam, None; K P Connie Tam, None; Steven Wilson, None
  • Footnotes
    Support  Supported by EY10056, EY015638 and Research to Prevent Blindness, New York, NY
Investigative Ophthalmology & Visual Science September 2016, Vol.57, 1282. doi:
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      Gustavo K. Marino, Abirami Santhanam, K P Connie Tam, Steven E Wilson; Ultrastructural evidence of defective regeneration of the epithelial basement membrane as a major factor in the generation and persistence of corneal myofibroblasts after corneal injury in rabbits. Invest. Ophthalmol. Vis. Sci. 2016;57(12):1282.

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      © 2017 Association for Research in Vision and Ophthalmology.

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Abstract

Purpose : Determine the correlation between the ultrastructure of the epithelial basement membrane (EBM) and the presence/absence of corneal myofibroblasts in scarred corneas after excimer laser surgery, incisional trauma, and bacterial infection

Methods : Rabbits were divided into 3 groups according to the mechanism of injury: (1) -9D PRK (VISX Star S4 IR laser); (2) partial-thickness (~300μm depth) corneal incisions; (3) P. aeruginosa corneal ulcer (~6x6mm). Fellow eyes were used as unwounded controls. Immunohistochemical analysis was used to detect the myofibroblast marker α-SMA and transmission electron microscopy at 30,000X was performed to determine whether the lamina lucida (LL) and lamina densa (LD) was present

Results : Group 1 had high numbers of α-SMA+ cells in the anterior stroma at 1 month after surgery and the EBM was irregular and lacked typical LL/LD morphology. At 3 months, there were no α-SMA+ cells and the EBM was fully regenerated and indistinguishable from control corneas. Group 2 had opacity at the incisions at 1, 2, and 3 months after surgery. None of these corneas had α-SMA+ cells and the EBM overlying the incisions was fully regenerated at all time points. Group 3 had large numbers of α-SMA+ cells from the anterior to posterior stroma at 1 month after infection and had no evidence of LL/LD overlying the corneal opacity (Figure)

Conclusions : Defective EBM correlated with the presence of corneal myofibroblasts in scarred corneas after high correction PRK or pseudomonas aeruginosa keratitis, which supports the hypothesis that generation and persistence of corneal myofibroblasts is regulated by the EBM after corneal injuries.

This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.

 

Immunohistochemistry for alpha-smooth muscle actin in central rabbit corneas: (A) Unwounded control; (C) -9D PRK at 1 month after surgery; (E) Partial-thickness corneal incision at 1 month after surgery; (G) Cornea at 1 month after a treated pseudomonas aeruginosa corneal ulcer (400X mag); and Transmission electron microscopy of the anterior central cornea of rabbits: (B) Unwounded control; (D) -9D PRK at 1 month after surgery; (F) Partial-thickness corneal incision at 1 month after surgery; (H) Cornea at 1 month after a treated pseudomonas aeruginosa corneal ulcer (30,000X mag). Arrowheads=a-SMA+ cells; arrows=lamina densa; “X”=absence of lamina lucida/lamina densa.

Immunohistochemistry for alpha-smooth muscle actin in central rabbit corneas: (A) Unwounded control; (C) -9D PRK at 1 month after surgery; (E) Partial-thickness corneal incision at 1 month after surgery; (G) Cornea at 1 month after a treated pseudomonas aeruginosa corneal ulcer (400X mag); and Transmission electron microscopy of the anterior central cornea of rabbits: (B) Unwounded control; (D) -9D PRK at 1 month after surgery; (F) Partial-thickness corneal incision at 1 month after surgery; (H) Cornea at 1 month after a treated pseudomonas aeruginosa corneal ulcer (30,000X mag). Arrowheads=a-SMA+ cells; arrows=lamina densa; “X”=absence of lamina lucida/lamina densa.

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