September 2016
Volume 57, Issue 12
Open Access
ARVO Annual Meeting Abstract  |   September 2016
Chymase/Angiotensin I axis: A pathway for Angiotensin II generation in diabetic human vitreous
Author Affiliations & Notes
  • Shaina M Rubino
    Ophthalmology, Wake Forest University, Winston Salem, North Carolina, United States
  • Ana-Maria Nae
    Wake Forest School of Medicine, Winston Salem, North Carolina, United States
  • Vishak John
    Ophthalmology, Wake Forest University, Winston Salem, North Carolina, United States
  • Sarfaraz Ahmad
    General Surgery, Wake Forest University, Winston Salem, North Carolina, United States
  • Mohammad S Ola
    Biochemistry, King Saud University, Riyadh, Saudi Arabia
  • Carlos M Ferrario
    General Surgery, Wake Forest University, Winston Salem, North Carolina, United States
  • Craig Greven
    Ophthalmology, Wake Forest University, Winston Salem, North Carolina, United States
  • Footnotes
    Commercial Relationships   Shaina Rubino, None; Ana-Maria Nae, None; Vishak John, None; Sarfaraz Ahmad, None; Mohammad Ola, None; Carlos Ferrario, None; Craig Greven, None
  • Footnotes
    Support  NIH Short-term Training Grant, 5T35 DK007400-36
Investigative Ophthalmology & Visual Science September 2016, Vol.57, 5431. doi:
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      Shaina M Rubino, Ana-Maria Nae, Vishak John, Sarfaraz Ahmad, Mohammad S Ola, Carlos M Ferrario, Craig Greven; Chymase/Angiotensin I axis: A pathway for Angiotensin II generation in diabetic human vitreous. Invest. Ophthalmol. Vis. Sci. 2016;57(12):5431.

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

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Abstract

Purpose : Expanding knowledge of the biochemical pathway responsible for diabetic retinopathy offers an opportunity for novel therapies with the recognition of each new component. While several studies show that the vitreous fluid level of Angiotensin II (Ang II) is significantly higher in diabetic patients with proliferative diabetic retinopathy (PDR) than in non-diabetic patients or diabetic patients without retinopathy, it is not known whether the enzyme chymase is present in vitreous fluids of diabetic patients and is responsible for Ang II formation. We tested the hypothesis that chymase-mediated Ang II formation from angiotensinogen precursors is upregulated in vitreous fluid of poorly controlled human diabetic subjects through the assay of chymase activity in samples of human vitreous fluid.

Methods : Vitreous samples were collected from non-diabetic patients (n=11) and patients with proliferative diabetic retinopathy (n=11) at the time of vitreoretinal surgery and then rapidly frozen at -20°C until assay. Chymase activity was measured in collected samples preincubated with or without a chymase inhibitor (chymostatin, 50 µM). Radiolabelled 125I-Ang I was then added into the reaction medium and incubated at 37oC. The reaction was stopped by adding 1% phosphoric acid and centrifuged. 125I-Ang II product generation from 125I-Ang I by chymase was measured. Chymase activity was normalized by protein concentration and results were expressed in terms of fmol Ang II formation/min/mg protein. Samples were categorized by several factors including patient gender, diabetic status, and ocular comorbidities.

Results : Chymase activity was significantly higher in vitreous fluid of diabetic patients (0.70 ± 0.16 fmol/min/mg protein, N=11, P<0.05) as compared to non-diabetic patients (0.35 ± 0.06 fmol/min/mg protein, N=11) as well as in those with proliferative diabetic retinopathy, retinal detachment, or vitreous hemorrhage (0.65 ± 0.13 fmol/min/mg protein, N=14, P<0.05) compared to those without those comorbidities (0.28 ± 0.07 fmol/min/mg protein, N=8).

Conclusions : Our studies show that the chymase-mediated pathway in vitreous fluid may be responsible for Ang II formation and contribute to the development of retinopathy in diabetic subjects. Chymase inhibitors may have potential therapeutic implications in the management of diabetic retinopathy.

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

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