June 2023
Volume 64, Issue 8
Open Access
ARVO Annual Meeting Abstract  |   June 2023
Safety and synergistic anti-fibrotic effects of Lisinopril (ACE inhibitor) in combination with Fasudil (ROCK inhibitor) for the canine cornea in vitro
Author Affiliations & Notes
  • Brayden Routh
    Veterinary Medicine and Surgery, University of Missouri, Columbia, Missouri, United States
  • Ratnakar Tripathi
    Veterinary Medicine and Surgery, University of Missouri, Columbia, Missouri, United States
    Harry S. Truman VA Hospital, Columbia, Missouri, United States
  • Elizabeth A Giuliano
    Veterinary Medicine and Surgery, University of Missouri, Columbia, Missouri, United States
  • Nishant R. Sinha
    Veterinary Medicine and Surgery, University of Missouri, Columbia, Missouri, United States
    Harry S. Truman VA Hospital, Columbia, Missouri, United States
  • Praveen Balne
    Veterinary Medicine and Surgery, University of Missouri, Columbia, Missouri, United States
    Harry S. Truman VA Hospital, Columbia, Missouri, United States
  • Laila A Suleiman
    Veterinary Medicine and Surgery, University of Missouri, Columbia, Missouri, United States
  • Suneel Gupta
    Veterinary Medicine and Surgery, University of Missouri, Columbia, Missouri, United States
    Harry S. Truman VA Hospital, Columbia, Missouri, United States
  • Nathan Hesemann
    Harry S. Truman VA Hospital, Columbia, Missouri, United States
  • Prashant R. Sinha
    Veterinary Medicine and Surgery, University of Missouri, Columbia, Missouri, United States
    Harry S. Truman VA Hospital, Columbia, Missouri, United States
  • Rajiv R Mohan
    Veterinary Medicine and Surgery, University of Missouri, Columbia, Missouri, United States
    Harry S. Truman VA Hospital, Columbia, Missouri, United States
  • Footnotes
    Commercial Relationships   Brayden Routh None; Ratnakar Tripathi None; Elizabeth Giuliano None; Nishant Sinha None; Praveen Balne None; Laila Suleiman None; Suneel Gupta None; Nathan Hesemann None; Prashant Sinha None; Rajiv Mohan None
  • Footnotes
    Support  None
Investigative Ophthalmology & Visual Science June 2023, Vol.64, 4587. doi:
  • Views
  • Share
  • Tools
    • Alerts
      ×
      This feature is available to authenticated users only.
      Sign In or Create an Account ×
    • Get Citation

      Brayden Routh, Ratnakar Tripathi, Elizabeth A Giuliano, Nishant R. Sinha, Praveen Balne, Laila A Suleiman, Suneel Gupta, Nathan Hesemann, Prashant R. Sinha, Rajiv R Mohan; Safety and synergistic anti-fibrotic effects of Lisinopril (ACE inhibitor) in combination with Fasudil (ROCK inhibitor) for the canine cornea in vitro. Invest. Ophthalmol. Vis. Sci. 2023;64(8):4587.

      Download citation file:


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

      ×
  • Supplements
Abstract

Purpose : Angiotensin-converting enzyme (ACE) and rho-kinase (ROCK) inhibitors are shown to treat fibrosis in many non-ocular tissues. This study evaluated safety and anti-fibrotic effects of the FDA-approved drugs, Lisinopril (ACE inhibitor) and Fasudil (ROCK inhibitor), alone and in combination for the canine cornea using an in vitro model.

Methods : Primary canine corneal fibroblasts (CCFs) generated from donor corneas of healthy dogs (n=20) were used. CCFs were grown in the +/- of TGFβ1 (5ng/ml), a fibrosis-causing agent, for 72h in serum-free conditions at 37°C in a humidified CO2 incubator to produce myofibroblasts. Cultures were grown +/- ACE and ROCK inhibitors and +/- TGFβ. The cultures were then subjected to MTT, trypan blue, Click-iT EdU Proliferation, phase-contrast microscopy, immunofluorescence (IF), and qRT-PCRs for safety and efficacy evaluations.

Results : A series of dose-dependent and time-dependent assays were performed with Lisinopril (1-50μM) and Fasudil (1-10nM). The 1-25μM doses of Lisinopril and 1-3nM doses of Fasudil caused an insignificant (0-5%) decrease in cellular viability and proliferation of CCFs. Conversely, 30-50μM doses of Lisinopril and 4-10nM of Fasudil decreased cell viability (31-67%) and cell proliferation (39-77%). Phase-contrast microscopy supported these findings. Collective analyses of 4 techniques found the 25μM dose of Lisinopril and the 3nM dose of Fasudil were safe, non-toxic, and optimal for therapeutic evaluations. CCFs grown in the presence of TGFβ1 (5ng/ml) in serum-free conditions without Lisinopril and/or Fasudil led to >93% (p<0.001) formation of myofibroblasts in cultures. Treatments of Lisinopril (25μM) or Fasudil (3nM), alone or in-combination, to CCFs grown in the presence of TGFβ1 showed remarkable inhibition of myofibroblast formation via phase-contrast microscopy. The myofibroblast inhibition was up to 30% higher (p<0.01 or p<0.1) when these two drugs were added in-combination. The αSMA, a biomarker for corneal fibrosis, IF studies supported brightfield microscopy observation. The results of qRT-PCR and IF studies of fibrosis markers are pending.

Conclusions : An ophthalmic formulation consisting of Lisinopril and Fasudil may offer an effective method of treating canine corneal fibrosis safely. Additional studies are warranted.

This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.

×
×

This PDF is available to Subscribers Only

Sign in or purchase a subscription to access this content. ×

You must be signed into an individual account to use this feature.

×