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M. E. Hajee, S. Hasan, E. M. Shrier, M. Hanna, D. Lazzaro, B. Chaqour; Differential Expression of the CCN Proteins in Retinal Blood Vessels During Development. Invest. Ophthalmol. Vis. Sci. 2009;50(13):2947.
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The retinal vasculature develops in the third-trimester human fetus and during the first two postnatal weeks in mice. The deeper and intermediate networks of the retinal vasculature grow outwardly in a concentric manner following gradients of growth factors/signaling molecules. The CCN proteins, Cyr61 and CTGF, are structurally-related, multifaceted matricellular proteins which regulate both developmental and pathological angiogenesis by means of cell migration, proliferation and survival. The present study was designed to examine the expression profile of the CCN proteins during development of the murine retinal vasculature.
Expression of the Cyr61 and CTGF genes was examined in mouse retina from postnatal day (P) 1 to P21 by real-time PCR using specifically designed primers. CCN copy number was determined from the fractional cycle at which a threshold amount of amplicon DNA is reached (threshold cycle or Ct) and normalized to 18S rRNA. Localization of the CCN proteins in retinal tissue was determined by immunohistochemistry in whole mounts retinas and in radial sections.
Three distinct peaks with the highest expression levels of the Cyr61 gene were identified at P4, P10 and P14 while, in the adult retina, Cyr61 transcripts were virtually undetectable. The expression of the CTGF gene decreased progressively between P1 and P8 and remained constant thereafter. Immunoreactivity to the Cyr61 protein at P 4 appeared in superficial retinal vessels, which grow radially from the optic nerve and in the deeper retinal vasculature which forms during the second week (P10) through branching of the superficial layer. The protein localized primarily in both small and moderately sized vessels. There was little evidence of association of Cyr61 with the microvasculature.
Cyr61 expression fluctuated the most during retinal vessel development. Its levels were essentially augmented when the number of sprouting vessels increased considerably and were decreased as the vascular network become denser in the retina, which underscores a potentially important role of this protein in steering the growth of vascular sprouts, by angiogenesis.
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