June 2021
Volume 62, Issue 8
Open Access
ARVO Annual Meeting Abstract  |   June 2021
Proteomic analyses of extracellular matrix and basal deposits in iPSC-RPE models of macular dystrophies
Author Affiliations & Notes
  • Sonal Dalvi
    Ophthalmology, University of Rochester Medical Center, Rochester, New York, United States
  • Chad A. Galloway
    Pathology and Laboratory Medicine, University of Rochester Medical Center, Rochester, New York, United States
    Biomedical Genetics, University of Rochester, Rochester, New York, United States
  • Michael Roll
    Ophthalmology, University of Rochester Medical Center, Rochester, New York, United States
  • Priscila Passerotti Vaciski Barbosa
    Ophthalmology, University of Rochester Medical Center, Rochester, New York, United States
  • Geeng-Fu Jang
    Ophthalmic Research, Cleveland Clinic Cole Eye Institute, Cleveland, Ohio, United States
  • Jack S Crabb
    Ophthalmic Research, Cleveland Clinic Cole Eye Institute, Cleveland, Ohio, United States
    Cleveland Clinic Lerner Research Institute, Cleveland, Ohio, United States
  • John W Crabb
    Ophthalmic Research, Cleveland Clinic Cole Eye Institute, Cleveland, Ohio, United States
    Cleveland Clinic Lerner Research Institute, Cleveland, Ohio, United States
  • Ruchira Singh
    Ophthalmology, University of Rochester Medical Center, Rochester, New York, United States
    Biomedical Genetics, University of Rochester, Rochester, New York, United States
  • Footnotes
    Commercial Relationships   Sonal Dalvi, None; Chad A. Galloway, None; Michael Roll, None; Priscila Barbosa, None; Geeng-Fu Jang, None; Jack Crabb, None; John Crabb, None; Ruchira Singh, None
  • Footnotes
    Support  Retina Research Foundation and Research to Prevent Blindness, RPB’s Career Development Award, Unrestricted Challenge Grant to Department of Ophthalmology at the University of Rochester, R01EY028167, R01EY030183
Investigative Ophthalmology & Visual Science June 2021, Vol.62, 1577. doi:
  • Views
  • Share
  • Tools
    • Alerts
      ×
      This feature is available to authenticated users only.
      Sign In or Create an Account ×
    • Get Citation

      Sonal Dalvi, Chad A. Galloway, Michael Roll, Priscila Passerotti Vaciski Barbosa, Geeng-Fu Jang, Jack S Crabb, John W Crabb, Ruchira Singh; Proteomic analyses of extracellular matrix and basal deposits in iPSC-RPE models of macular dystrophies. Invest. Ophthalmol. Vis. Sci. 2021;62(8):1577.

      Download citation file:


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

      ×
  • Supplements
Abstract

Purpose : Induced pluripotent stem cell-retinal pigment epithelium (iPSC-RPE) in vitro models of maculopathies such as Sorsby’s fundus dystrophy (SFD) and Doyne honeycomb retinal dystrophy (DHRD) display drusen-like basal deposits that appear consistent with the human diseases. The purpose of this study was to evaluate the proteomic relatedness of the basal deposits in the SFD and DHRD iPSC-RPE cultures with the human drusen proteome.

Methods : Extracellular matrix (ECM) and/or basal deposits were isolated from parallel patient-derived SFD and DHRD iPSC-RPE cultures and control (unaffected family member and/or gene-corrected line) iPSC-RPE cultures that were grown in culture for 40-100 days. ECM/basal deposit proteins were extracted in SDS, and digested with trypsin. Peptides were fractionated by reversed-phase high performance liquid chromatography (RPHPLC) at pH 10 then analyzed by Liquid chromatography–mass spectrometry (LC MS/MS) and proteins identified using the human UniProt database. Bioinformatic analyses was performed using PANTHER and Ingenuity Pathway analysis software.

Results : Proteome analyses identified 1689 and 2237 number of total proteins with 2 or more unique peptides in the ECM of SFD and DHRD cultures respectively. Of note, 61 proteins present in ECM/basal deposit underlying SFD iPSC-RPE cultures and 56 proteins present in ECM/basal deposit underlying in DHRD iPSC-RPE cultures were common with the 123 proteins previously reported in human AMD drusen (2002 PNAS, (23) 14682-14687). Furthermore, 49 of the previously identified drusen proteins in the AMD eyes were seen in both SFD and DHRD iPSC-RPE cultures. Notably, cytoskeletal, calcium-binding, and inflammation associated proteins constituted the majority of proteins detected in the ECM/basal deposits from SFD and DHRD cultures.

Conclusions : LC MS/MS analyses demonstrate overlap between the proteome of drusen from AMD donor eyes and basal deposits in SFD and DHRD iPSC-RPE cultures. The results further support the utility of these in vitro models for mechanistic studies of macular degeneration.

This is a 2021 ARVO Annual Meeting abstract.

×
×

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.

×