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M. Bobbie, R. Raithel, A. Simon, S. Roy; Effect of Connexin 43 Downregulation on the Development of Retinal Vascular Lesions in Diabetic Retinopathy. Invest. Ophthalmol. Vis. Sci. 2009;50(13):5903.
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To determine whether the development of acellular capillaries (AC) and pericyte loss (PL) is influenced by connexin 43 (Cx43) downregulation in Cx43 heterozygous knockout (Cx43+/- KO) mice and Cx43+/- KO mice with diabetes.
Retinas isolated from four groups of mice (normal, diabetic, Cx43+/- KO, and diabetic plus Cx43+/- KO) were subjected to trypsin digestion technique to isolate the capillary networks. The capillary networks were stained with hematoxylin and periodic acid-Schiff, and digital images of random fields of the network were captured. The images were analyzed for AC and PL. TUNEL assays were performed to identify retinal vascular cells undergoing apoptosis. Western blot analysis was performed with protein isolated from the contralateral retina to determine retinal Cx43 protein levels in the four groups of mice.
Western blot analysis showed reduced retinal Cx43 protein levels in diabetic, Cx43+/- KO and diabetic plus Cx43+/- KO mice (66±15% of control; 51±11% of control; 52±7% of control, respectively) compared to those of normal mice. Retinas from diabetic, Cx43+/- KO, and diabetic plus Cx43+/- KO mice showed significant increase in the number of AC (212±17% of control, p=0.034; 206±8% of control, p=0.016; and 235±33% of control, p=0.008, respectively) compared to those of normal mice. Similarly the number of PL was increased in diabetic, Cx43+/- KO, and diabetic plus Cx43+/- KO compared to normal mice (233±17% of control, p=0.003; 200±54% of control, p=0.024; 250±33% of control, p=0.001, respectively). An increase in the number of TUNEL positive cells was observed in retinal capillaries of diabetic, Cx43+/- KO, and diabetic plus Cx43+/- KO mice compared to that of normal mice.
These findings suggest that decreased Cx43 expression in the retina may contribute to the development of AC and PL, similar to those observed in characteristic lesions of diabetic retinopathy.
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