Acute intensive insulin therapy exacerbates diabetic blood-retinal barrier breakdown via hypoxia-inducible factor-1α and VEGF
Vassiliki Poulaki, … , George D. Yancopoulos, Anthony P. Adamis
Vassiliki Poulaki, … , George D. Yancopoulos, Anthony P. Adamis
Published March 15, 2002
Citation Information: J Clin Invest. 2002;109(6):805-815. https://doi.org/10.1172/JCI13776.
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Article Vascular biology

Acute intensive insulin therapy exacerbates diabetic blood-retinal barrier breakdown via hypoxia-inducible factor-1α and VEGF

Abstract

Acute intensive insulin therapy is an independent risk factor for diabetic retinopathy. Here we demonstrate that acute intensive insulin therapy markedly increases VEGF mRNA and protein levels in the retinae of diabetic rats. Retinal nuclear extracts from insulin-treated rats contain higher hypoxia-inducible factor-1α (HIF-1α) levels and demonstrate increased HIF-1α–dependent binding to hypoxia-responsive elements in the VEGF promoter. Blood-retinal barrier breakdown is markedly increased with acute intensive insulin therapy but can be reversed by treating animals with a fusion protein containing a soluble form of the VEGF receptor Flt; a control fusion protein has no such protective effect. The insulin-induced retinal HIF-1α and VEGF increases and the related blood-retinal barrier breakdown are suppressed by inhibitors of p38 mitogen-activated protein kinase (MAPK) and phosphatidylinositol (PI) 3-kinase, but not inhibitors of p42/p44 MAPK or protein kinase C. Taken together, these findings indicate that acute intensive insulin therapy produces a transient worsening of diabetic blood-retinal barrier breakdown via an HIF-1α–mediated increase in retinal VEGF expression. Insulin-induced VEGF expression requires p38 MAPK and PI 3-kinase, whereas hyperglycemia-induced VEGF expression is HIF-1α–independent and requires PKC and p42/p44 MAPK. To our knowledge, these data are the first to identify a specific mechanism for the transient worsening of diabetic retinopathy, specifically blood-retinal barrier breakdown, that follows the institution of intensive insulin therapy.

Authors

Vassiliki Poulaki, Wenying Qin, Antonia M. Joussen, Peter Hurlbut, Stanley J. Wiegand, John Rudge, George D. Yancopoulos, Anthony P. Adamis

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Figure 1

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Upper panel: Conditioned media VEGF protein levels in RPE cells in vitro...
Upper panel: Conditioned media VEGF protein levels in RPE cells in vitro following exposure to high glucose and insulin (ins). Insulin increased VEGF protein levels (from 0.32 to 0.89 pg/μg total protein, n = 4, P < 0.001). Inhibition (inhib) of the p38 MAPK and PI 3-kinase suppressed the insulin-induced VEGF levels. Inhibition of p42/p44 MAPK or PKC had no effect. Hyperglycemia (HG) increased VEGF levels in RPE-conditioned media in vitro. Inhibition of p42/p44 MAPK, PKC, and p38 MAPK suppressed the hyperglycemia-induced VEGF increases, whereas PI 3-kinase inhibition had no effect. Asterisks denote statistically significant results (**P < 0.001). Lower panel: HIF-1α protein levels in RPE cells in vitro following exposure to high glucose and insulin. Insulin increased HIF-1α levels in vitro (HIF-1α/β-actin ratio 0.85 to 2.7, n = 4, P < 0.001). Inhibition of p38 MAPK or PI 3-kinase suppressed the insulin-induced HIF-1α increases (1.97 and 1.51, respectively; n = 4, P < 0.001), whereas inhibition of either PKC or p42/p44 had no effect (2.54 and 2.41, respectively; n = 4, P > 0.05). Hyperglycemia alone had no effect on HIF-1α levels in vitro (HIF-1α/β-actin ratio increased from 0.87 to 1.01, 0.99, 0.99, 0.99, and 0.99 for the mannose control, high glucose + p38,PI 3-kinase, MAPK, and PKC inhibitor-treated cells respectively). Numbers represent the HIF-1α/β-actin ratio.