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An-Fei Li, Tsuyoshi Sato, Robert Haimovici, Tamami Okamoto, Sayon Roy; High Glucose Alters Connexin 43 Expression and Gap Junction Intercellular Communication Activity in Retinal Pericytes. Invest. Ophthalmol. Vis. Sci. 2003;44(12):5376-5382. doi: 10.1167/iovs.03-0360.
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purpose. To investigate the role of the gap junction protein, connexin-43 (Cx43) in the maintenance of retinal vascular homeostasis in diabetic retinopathy.
methods. In human retinal pericytes (HRPs) and bovine retinal pericytes (BRPs) grown for 7 days in normal (5 mM) or high (30 mM)-glucose medium, the Cx43 protein level was determined by Western blot analysis. Parallel experiments were performed in HRPs to determine the Cx43 mRNA level by RT-PCR, the distribution and localization of Cx43 protein by immunostaining, and gap junction intercellular communication (GJIC) activity by a scrape-loading dye transfer technique. Distribution and localization of Cx43 protein was also determined in pericyte–endothelial cell cocultures.
results. Western blot analysis of the Cx43 protein level in HRPs and BRPs indicated reduced Cx43 expression in the high-glucose condition (69.1% ± 17% of control, P = 0.004; 62.3% ± 19% of control, P = 0.001, respectively). The Cx43 mRNA level in HRPs grown in high-glucose medium also showed significant reduction (71.4% ± 16.8% of control, P = 0.02). The relative number of Cx43 plaques indicative of Cx43 localization at specific sites of contact between adjacent cells showed significant reduction in the high-glucose condition (61% ± 10% of control, P = 0.002); similarly, a significant reduction in the number of plaques was observed in cocultures grown in high-glucose medium compared with those in normal medium (59.4% ± 29% of control, P = 0.001). Cells with reduced Cx43 expression showed significantly reduced transfer of lucifer yellow (61% ± 13% of control, P = 0.001; r = 0.9).
conclusions. High-glucose–induced downregulation of Cx43 expression and inhibition of GJIC in retinal pericytes may play a role in the disruption of vascular homeostasis in diabetic retinopathy.
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