Department of Physiology and Neurobiology

University of Connecticut

Storrs, CT

BASIC RESEARCH PROJECT

Modulation of retinal vascularization by endothelial cell genetic reprogramming

Research Interests

Abnormal formation of new blood vessels is the fundamental cause of catastrophic vision loss during ocular neovascularization, stressing the urgent need for new treatments. Current therapies aim to destroy aberrant vascularization by neutralizing Vascular Endothelial Growth Factor-A (VEGF). Yet, these approaches do not address the causes that underlie vascular pathology, such as hypoxia and inflammation. We therefore aim to identify VEGF-independent signaling pathways, that could promote vascular repair and regeneration of the diseased eye. One pathway that holds promise is Transforming Growth Factor (TGF) β and its receptor (R) 1 (TGFBR1), which are specifically required during developmental vascular growth in the neuroretina. Dr. Zarkada hypothesizes that activation of TGFβ/TGFBR1 signaling could force sprouting endothelial cells to acquire the abilities to form a leak-proof and properly patterned vasculature in the neuroretina. She will therefore generate a new genetic mouse model to test this hypothesis during developmental retinal vascularization.

Plans for 2026

Based on previous studies, Dr. Zarkada will use her lab’s newly generated mouse model to elucidate how ectopic expression of TGFBR1 would affect developmental vascularization in the postnatal mouse neuroretina and in a pre-clinical model of retinopathy of prematurity (oxygen-induced retinopathy; OIR). In addition, her team will assess possible neuroprotective roles of TGFBR1 over expression in OIR, as well as the impact of such over expression on the preservation of visual acuity in retinopathy.

Specific Aims: Aim 1  the team will perform in vivo validation of endothelial-specific overexpression of TGFBR1 and analyze the effects of increased TGFβ/TGFBR1 signaling during developmental retinal angiogenesis in honozygous GOF mice. In Aim 2, the team will subject TGFBR1 gain of function mice to a preclinical model or retinopathy of prematurity (oxygen-induced retinopathy; OIR), and assess the therapeutic potential of TGFβ/TGFBR1 activation in retinopathy by evaluating pathological neovascularization, neuronal degeneration and visual function.

Progress in 2025

In 2025, Dr. Zarkada’s lab generated and characterized an endothelial-specific TGFBR1 gain-of-function (GOF) transgenic mouse model. the model is based on a novel technology, known as FLEx-switch, which mediates permanent site-specific recombination of targeted sequences.  The team’s analysis of mice containing one copy of TGFBR1-HA reveals mild and variable expression retinal endothelial cells, which does not severely affect vascular retina development. the lab demonstrated that TGFBR1 is not involved in the vascularization of organs outside the mouse central nervous system, including the retina and brain. In addition, Dr. Zarkada published a novel protocol to analyze deep layer retinal vascularization in Methods in Molecular Biology (August 2025)

Progress in 2024

During the previous year of this pilot award, Dr. Zarkada generated a TGFBR1 gain-of-function transgenic mouse model. The model is based on a novel technology, known as FLEx-switch, which mediates permanent site-specific recombination of targeted sequences. Validation of our construct revealed zero spontaneous expression (leakage) at baseline conditions, and increased activity of TGFBR1 after cell transfection in vitro. First generation offspring has been born in September 2024, and Dr. Zarkada has  confirmed germline transmission and CRE-induced recombination. In addition, the lab has optimized its approach to analyze deep layer retinal vascularization, with a novel protocol that utilizes cross sections of freshly prepared traditional retinal flat mounts. This protocol has been accepted for publication as a methods paper in Methods in Molecular Biology.