July 2019
Volume 60, Issue 9
Open Access
ARVO Annual Meeting Abstract  |   July 2019
Tryptase release after degranulation of Mast Cells in a Model for Geographic Atrophy
Author Affiliations & Notes
  • Rajkumar Baldeosingh
    Johns Hopkins University, Baltimore, Maryland, United States
  • Shuntaro Ogura
    Johns Hopkins University, Baltimore, Maryland, United States
  • Siva Pramodh Kambhampati
    Johns Hopkins University, Baltimore, Maryland, United States
  • Malia Michelle Edwards
    Johns Hopkins University, Baltimore, Maryland, United States
  • Gerard A Lutty
    Johns Hopkins University, Baltimore, Maryland, United States
  • Footnotes
    Commercial Relationships   Rajkumar Baldeosingh, None; Shuntaro Ogura, None; Siva Kambhampati, None; Malia Edwards, None; Gerard Lutty, None
  • Footnotes
    Support  NIH Grant RO-1-EY-016151 (GL), EY-01765 (Wilmer), Bayer AG
Investigative Ophthalmology & Visual Science July 2019, Vol.60, 1231. doi:
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      Rajkumar Baldeosingh, Shuntaro Ogura, Siva Pramodh Kambhampati, Malia Michelle Edwards, Gerard A Lutty; Tryptase release after degranulation of Mast Cells in a Model for Geographic Atrophy. Invest. Ophthalmol. Vis. Sci. 2019;60(9):1231.

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

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Purpose : In subjects with geographic atrophy (GA), degranulated choroidal Mast Cells (MCs) are elevated in number (Bhutto et. al, BJO, 2016) and MC tryptase is present in Bruch’s membrane (BM) (McLeod et. al, IOVS, 2017). We have previously reported on a rat model that demonstrates the functional and pathological significance of MC degranulation on retinal pigment epithelial cells (RPE) and choroid and determined that GA-like changes occur 6 weeks after stimulating MC degranulation. The purpose of this study was to determine the time course of MC degranulation and detect tryptase release and deposit in this rat model.

Methods : A hydrogel with 48/80 (a snake venom-like compound) or a blank hydrogel was injected subconjunctivally into Sprague-Dawley rats. Rats were sacrificed at days 3, 7, 14, 28 and 36, post-surgery. Tissue was then fixed, cryopreserved and sectioned or flat-mounted. MCs in tissue were stained with nonspecific esterase (NSE) and Alcian blue (AB). Cryosections were stained with Hematoxylin & Eosin (H&E) or immunolabeled with antibodies against tryptase or c-Kit, and/or stained GS-Lectin and DAPI. Sections were examined and imaged on a Zeiss 710 Confocal Microscope.

Results : Significant choroidal MC degranulation was observed at all time points post implantation of subconjuntival 48/80 hydrogel but not in the blank hydrogel eyes (p<0.005), with the highest average MC degranulation occurring at days 3, 7 and 14. In addition, at days 3, 7 and 14, tryptase deposits were observed in choroidal stroma and BM in 48/80 eyes but no tryptase deposits were observed in blank hydrogel eyes. H&E staining showed no significant thinning or damage to the choroid and retina in 48/80 eyes at these early time points.

Conclusions : Inducing MC degranulation in a slow release fashion resulted in loss of RPE at 6 weeks post implantation of 48/80 hydrogel in our model. This study, however, showed that MC degranulation peaks at 3, 7 and 14 days in our model. The results also demonstrate that tryptase released during degranulation of choroidal mast cells binds to BM in advance of RPE atrophy, which is similar to our observations in human GA subjects. Tryptase activates MMPs, which can degrade BM membrane components, thus leading to profound effects on the function, health and transport of RPE in our rat model and in patients with GA.

This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.


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