June 2022
Volume 63, Issue 7
Open Access
ARVO Annual Meeting Abstract  |   June 2022
CCN2 regulates the extracellular matrix and facilitates retinal ganglion cell maturation in the developing retina
Author Affiliations & Notes
  • Charles Karrasch
    Cell Biology, SUNY Downstate Health Sciences University The School of Graduate Studies, Brooklyn, New York, United States
    Ophthalmology, SUNY Downstate Health Sciences University The School of Graduate Studies, Brooklyn, New York, United States
  • Sohyun Moon
    Cell Biology, SUNY Downstate Health Sciences University, Brooklyn, New York, United States
    Ophthalmology, SUNY Downstate Health Sciences University, Brooklyn, New York, United States
  • Golam Mohiuddin
    Cell Biology, SUNY Downstate Health Sciences University The School of Graduate Studies, Brooklyn, New York, United States
    Ophthalmology, SUNY Downstate Health Sciences University The School of Graduate Studies, Brooklyn, New York, United States
  • Oleg Evgrafov
    Cell Biology, SUNY Downstate Health Sciences University, Brooklyn, New York, United States
  • Brahim Chaqour
    Cell Biology, SUNY Downstate Health Sciences University, Brooklyn, New York, United States
    Ophthalmology, SUNY Downstate Health Sciences University, Brooklyn, New York, United States
  • Footnotes
    Commercial Relationships   Charles Karrasch None; Sohyun Moon None; Golam Mohiuddin None; Oleg Evgrafov None; Brahim Chaqour None
  • Footnotes
    Support  None
Investigative Ophthalmology & Visual Science June 2022, Vol.63, 4279. doi:
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      Charles Karrasch, Sohyun Moon, Golam Mohiuddin, Oleg Evgrafov, Brahim Chaqour; CCN2 regulates the extracellular matrix and facilitates retinal ganglion cell maturation in the developing retina. Invest. Ophthalmol. Vis. Sci. 2022;63(7):4279.

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

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Abstract

Purpose : Extracellular matrix (ECM) remodeling is a key factor in retinal development. Recent work suggests the ECM protein Cellular Communication Network Factor 2 (CCN2) regulates retinal progenitor cell survival, proliferation, and differentiation. We employed transcriptional profiling of murine embryonic retinas to test the hypothesis that CCN2 regulates the ECM in a cell type-dependent manner and mediates cell fate specification.

Methods : Germline CCN2 deletion was achieved using a Ccn2fl/fl; CMV-Cre mouse model. Embryonic day (E)18 CCN2-/- and CCN2+/+ littermate control retinas from both sexes were used for bulk RNA sequencing (RNA-seq) (n=3), RT-qPCR (n≥3), and single-cell (sc)RNA-seq (n=2). RNA-seq and scRNA-seq data were interrogated by Gene Set Enrichment Analysis (GSEA).

Results : In CCN2-/- retinas, GSEA of RNA-seq data revealed significant repression of the Core Matrisome gene set (p=0.017, q=0.20), representing the core components of the ECM proteome. New retinas were probed via RT-qPCR for a selection of Core Matrisome genes, which confirmed broad ECM repression. Subsequent scRNA-seq and GSEA were performed with new retinas. Each retinal cell type possessed a unique ECM expression signature, and Ccn2 was specifically expressed by retinal progenitor cells (RPCs). CCN2 deficiency repressed the structural ECM genes Col1a2 (-2.58-fold, p<0.01) and Col5a2 (-1.88-fold, p<0.001) specifically in RPCs. Damaged cells were approximately 3-fold more prevalent in CCN2-/- retinas compared to controls (17.6% vs. 5.76%), suggesting decreased viability following CCN2 deletion. Among retinal ganglion cells (RGCs), CCN2-/- retinas contained a greater proportion of immature RGCs compared to controls (71.2% vs 51.8%). Accordingly, RGCs from CCN2-/- retinas repressed a fetal RGC gene set (p<0.001, q<0.001) and upregulated a fetal RPC gene set (p<0.001, q<0.001). Finally, GSEA of scRNA-seq data revealed cell type-specific regulation of the Core Matrisome, which in CCN2-/- retinas was repressed in RPCs (p=0.017) yet upregulated in RGCs (p=0.021) and horizontal cells (p=0.016).

Conclusions : In the developing retina, CCN2 mediates pan-retinal ECM gene expression, yet differentially regulates the ECM in a cell type-dependent manner. CCN2 is critical for RPC-specific expression of structural collagen genes, retinal cell viability, and RGC specification from RPCs.

This abstract was presented at the 2022 ARVO Annual Meeting, held in Denver, CO, May 1-4, 2022, and virtually.

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