Connective tissue growth factor deletion induces retinal vascular defects and barrier dysfunction through yes-associated protein-dependent mechanisms

Abstract

Purpose : Connective tissue growth factor (CTGF) is an inducible extracellular matrix protein with a major role in the initiation and progression of fibroproliferative diseases including proliferative diabetic retinopathy and oxygen-induced retinopathy. However, global CTGF deficiency produces skeletal dysplasia and perinatal lethality in mice. Despite its prominent expression in vascular cells e.g., endothelial cells (ECs) and pericytes, the role and mechanisms whereby CTGF regulates retinal vessel formation and regeneration are unknown. Herein, we used mouse genetics and omics’ approaches to study the regulation and function of CTGF in the retinal vasculature.

Methods : Mice with global, endothelial- or pericyte-specific deletion of CTGF are obtained by crossing mice with floxed CTGF alleles with tamoxifen-activated UBC-CreER^T2, Cdh5(PAC)-CreER^T2 or Cspg4-CreER^T2 transgenic mice respectively. Retinal vascular alterations and permeability are characterized at postnatal day P7 using immunohistochemical and biochemical techniques. Transcriptomic changes between three replicates of wild-type and CTGF mutant mouse retinas are determined by RNA-seq on prepared RNA libraries paired-end sequenced on Illumina HiSeq 2000 and subsequently analyzed using bioinformatics tools. Post hoc unpaired t-test is used to compare two means/groups.

Results : Either global or endothelial-specific deletion of CTGF reduce vessel area, microvessel density, and branching points and produces a leaky vascular network partially shaped into rudimentary arterioles venules and capillaries. Conversely, the vascular phenotype of wild-type (WT) and pericyte-specific CTGF-deficient mice are similar. Transcriptomic changes between WT and EC-specific CTGF mutant mouse retinas include cytoskeletal, extracellular matrix, integrin and transcriptional co-activator genes such as the yes-associated protein, YAP, known to regulates cell growth and organ size. CTGF deletion recapitulates the retinal vascular phenotype associated with YAP deficiency. Re-expression of YAP, at least in part, rescues the vascular defects and improves barrier function in CTGF-deficient mice.

Conclusions : CTGF normalizes the retinal vasculature and maintains barrier function through the regulation of the expression of YAP and its “regulome”.

This is a 2020 ARVO Annual Meeting abstract.