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Alex A. Bigger-Allen, mark Graham, Joseph F Arboleda-Velasquez, Vincent Primo, Patricia A D'Amore; Adult Notch3 Deficient Mice Exhibit Extensive Mural Cell Loss in the Retinal Vasculature. Invest. Ophthalmol. Vis. Sci. 2015;56(7 ):3384. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
Vascular development is mediated in part by interactions between endothelial cells (ECS) and mural cells. This heterotypic signaling plays a critical role in the formation, maturation and stability of the blood vessels. Prior work using cell culture and animal models has implicated the Notch signaling pathway in vascular development and stability. We have previously demonstrated a critical role for Notch signaling in mural cell survival in vitro. In the present work, we examined the role of Notch signaling in the adult microvasculature in vivo.
Retinas were dissected from 200-day-old mice including wild type (WT), n = 4 and from Notch 3 knockout mice (Notch 3 -/-), n = 4. Flat mounts were stained by immunofluorescence using antisera against a-smooth muscle actin (αSMA) conjugated to Cy3 fluorophore as well as lectin conjugated to Alexa 488 fluorophore. Mural cell coverage was quantified using Image J by dividing the total mural cell area by the total vascularized area.
The retinal vasculature of adult Notch 3 knockout mice had 48% ± 14.22, p < 0.001 less aSMA positive area compared to the C57BL/6J control. Arterial vessels from the base of the optic nerve to as far as secondary and tertiary branches of the arteries exhibited significant lengths devoid of smooth muscle cells investment (Fig 1). Areas of vessels that were lacking smooth muscle cells displayed vessel dilation, which is reminiscent of the microaneurysms that characterize background diabetic retinopathy.
The absence of mural cells in 6- month-old retinal vessels of Notch 3 knockout mice suggests that Notch signaling is necessary for the stability and/or survival of mural cells in the adult. Further, these data support the hypothesis that the loss of mural cells contributes to the retinal degeneration observed in diabetic retinopathy.
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