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Inhibition of reactive oxygen species by Lovastatin downregulates vascular endothelial growth factor expression and ameliorates blood-retinal barrier breakdown in db/db mice: role of NADPH oxidase 4.

Li J, Wang JJ, Yu Q, Chen K, Mahadev K, Zhang SX - Diabetes (2010)

Bottom Line: Moreover, inhibition of Nox4 attenuated hypoxia-induced upregulation of HIF-1alpha and high-glucose-elicited phosphorylation of STAT3.Finally, depletion of Nox4 by adenovirus-delivered Nox4 small interfering RNA significantly decreased retinal NADPH oxidase activity and VEGF expression and reduced retinal vascular premeability in db/db mice.Activation of Nox4 plays an important role in high-glucose- and hypoxia-mediated VEGF expression and diabetes-induced BRB breakdown.

View Article: PubMed Central - PubMed

Affiliation: Harold Hamm Oklahoma Diabetes Center and Section of Endocrinology and Diabetes, University of Oklahoma Health Sciences Center, Oklahoma City, Oklahoma, USA.

ABSTRACT

Objective: Oxidative stress is a key pathogenic factor in diabetic retinopathy. We previously showed that lovastatin mitigates blood-retinal barrier (BRB) breakdown in db/db mice. The purpose of this study is to determine the mechanisms underlying the salutary effects of lovastatin in diabetic retinopathy.

Research design and methods: Expression of NADPH oxidase (Nox) 4, vascular endothelial growth factor (VEGF), and hypoxia-inducible factor (HIF)-1alpha; production of reactive oxygen species (ROS); and retinal vascular permeability were measured in cultured retinal capillary endothelial cells (RCECs) and in db/db mice treated with lovastatin.

Results: Expressions of Nox4 and VEGF were significantly increased in retinas of db/db mice and reduced by lovastatin treatment. In cultured RCECs, hypoxia and high glucose upregulated mRNA and protein expression of Nox4, ROS generation, and VEGF level. These changes were abrogated by pretreatment with lovastatin or NADPH oxidase inhibitor diphenyleneiodonium chloride. Overexpression of Nox4 increased basal level of ROS generation, HIF-1alpha, and VEGF expression in RCECs. In contrast, blockade of Nox4 activity using adenovirus-expressing dominant-negative Nox4 abolished hypoxia- and high-glucose-induced ROS production and VEGF expression. Moreover, inhibition of Nox4 attenuated hypoxia-induced upregulation of HIF-1alpha and high-glucose-elicited phosphorylation of STAT3. Finally, depletion of Nox4 by adenovirus-delivered Nox4 small interfering RNA significantly decreased retinal NADPH oxidase activity and VEGF expression and reduced retinal vascular premeability in db/db mice.

Conclusions: Activation of Nox4 plays an important role in high-glucose- and hypoxia-mediated VEGF expression and diabetes-induced BRB breakdown. Inhibition of Nox4, at least in part, contributes to the protective effects of lovastatin in diabetic retinopathy.

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Effect of lovastatin on VEGF and Nox4 expression in retinas of db/db mice. Db/db mice and nondiabetic littermate controls were gastric gavaged with or without lovastatin (10 mg/kg/day) for 6 weeks. VEGF (A) and Nox4 (B) expression in retinas were determined by Western blotting analysis and semiquantified by densitometry (means ± SE, n = 6). *P < 0.05 or **P < 0.01 vs. nondiabetic control; †P < 0.05 vs. db/db mice.
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Figure 1: Effect of lovastatin on VEGF and Nox4 expression in retinas of db/db mice. Db/db mice and nondiabetic littermate controls were gastric gavaged with or without lovastatin (10 mg/kg/day) for 6 weeks. VEGF (A) and Nox4 (B) expression in retinas were determined by Western blotting analysis and semiquantified by densitometry (means ± SE, n = 6). *P < 0.05 or **P < 0.01 vs. nondiabetic control; †P < 0.05 vs. db/db mice.

Mentions: Overexpression of VEGF in the retina induced by diabetes plays a critical role in BRB breakdown and vascular leakage in diabetic retinopathy (32). To address if the protection of BRB by lovastatin is associated with the inhibition of VEGF signaling, we determined the effects of lovastatin on retinal VEGF expression in db/db mice. As shown in Fig. 1A, VEGF expression was significantly increased in retinas of db/db mice, which was normalized by lovastatin treatment to nearly the normal level. These results suggest that inhibition of VEGF by lovastatin is associated with its salutary effect on diabetes-induced BRB breakdown. To determine whether Nox4 is implicated in diabetic retinopathy, we measured Nox4 level in the retina of db/db mice. We found that Nox4 expression in the retina was significantly increased by twofold in db/db mice when compared with age- and sex-matched nondiabetic littermates (P < 0.01) (Fig. 1B). Lovastatin treatment almost completely reversed the increase in Nox4 expression in the db/db retina (P < 0.05) (Fig. 1B). These results suggest a potential role of Nox4 in VEGF upregulation and BRB breakdown in diabetic retinopathy.


Inhibition of reactive oxygen species by Lovastatin downregulates vascular endothelial growth factor expression and ameliorates blood-retinal barrier breakdown in db/db mice: role of NADPH oxidase 4.

Li J, Wang JJ, Yu Q, Chen K, Mahadev K, Zhang SX - Diabetes (2010)

Effect of lovastatin on VEGF and Nox4 expression in retinas of db/db mice. Db/db mice and nondiabetic littermate controls were gastric gavaged with or without lovastatin (10 mg/kg/day) for 6 weeks. VEGF (A) and Nox4 (B) expression in retinas were determined by Western blotting analysis and semiquantified by densitometry (means ± SE, n = 6). *P < 0.05 or **P < 0.01 vs. nondiabetic control; †P < 0.05 vs. db/db mice.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2874715&req=5

Figure 1: Effect of lovastatin on VEGF and Nox4 expression in retinas of db/db mice. Db/db mice and nondiabetic littermate controls were gastric gavaged with or without lovastatin (10 mg/kg/day) for 6 weeks. VEGF (A) and Nox4 (B) expression in retinas were determined by Western blotting analysis and semiquantified by densitometry (means ± SE, n = 6). *P < 0.05 or **P < 0.01 vs. nondiabetic control; †P < 0.05 vs. db/db mice.
Mentions: Overexpression of VEGF in the retina induced by diabetes plays a critical role in BRB breakdown and vascular leakage in diabetic retinopathy (32). To address if the protection of BRB by lovastatin is associated with the inhibition of VEGF signaling, we determined the effects of lovastatin on retinal VEGF expression in db/db mice. As shown in Fig. 1A, VEGF expression was significantly increased in retinas of db/db mice, which was normalized by lovastatin treatment to nearly the normal level. These results suggest that inhibition of VEGF by lovastatin is associated with its salutary effect on diabetes-induced BRB breakdown. To determine whether Nox4 is implicated in diabetic retinopathy, we measured Nox4 level in the retina of db/db mice. We found that Nox4 expression in the retina was significantly increased by twofold in db/db mice when compared with age- and sex-matched nondiabetic littermates (P < 0.01) (Fig. 1B). Lovastatin treatment almost completely reversed the increase in Nox4 expression in the db/db retina (P < 0.05) (Fig. 1B). These results suggest a potential role of Nox4 in VEGF upregulation and BRB breakdown in diabetic retinopathy.

Bottom Line: Moreover, inhibition of Nox4 attenuated hypoxia-induced upregulation of HIF-1alpha and high-glucose-elicited phosphorylation of STAT3.Finally, depletion of Nox4 by adenovirus-delivered Nox4 small interfering RNA significantly decreased retinal NADPH oxidase activity and VEGF expression and reduced retinal vascular premeability in db/db mice.Activation of Nox4 plays an important role in high-glucose- and hypoxia-mediated VEGF expression and diabetes-induced BRB breakdown.

View Article: PubMed Central - PubMed

Affiliation: Harold Hamm Oklahoma Diabetes Center and Section of Endocrinology and Diabetes, University of Oklahoma Health Sciences Center, Oklahoma City, Oklahoma, USA.

ABSTRACT

Objective: Oxidative stress is a key pathogenic factor in diabetic retinopathy. We previously showed that lovastatin mitigates blood-retinal barrier (BRB) breakdown in db/db mice. The purpose of this study is to determine the mechanisms underlying the salutary effects of lovastatin in diabetic retinopathy.

Research design and methods: Expression of NADPH oxidase (Nox) 4, vascular endothelial growth factor (VEGF), and hypoxia-inducible factor (HIF)-1alpha; production of reactive oxygen species (ROS); and retinal vascular permeability were measured in cultured retinal capillary endothelial cells (RCECs) and in db/db mice treated with lovastatin.

Results: Expressions of Nox4 and VEGF were significantly increased in retinas of db/db mice and reduced by lovastatin treatment. In cultured RCECs, hypoxia and high glucose upregulated mRNA and protein expression of Nox4, ROS generation, and VEGF level. These changes were abrogated by pretreatment with lovastatin or NADPH oxidase inhibitor diphenyleneiodonium chloride. Overexpression of Nox4 increased basal level of ROS generation, HIF-1alpha, and VEGF expression in RCECs. In contrast, blockade of Nox4 activity using adenovirus-expressing dominant-negative Nox4 abolished hypoxia- and high-glucose-induced ROS production and VEGF expression. Moreover, inhibition of Nox4 attenuated hypoxia-induced upregulation of HIF-1alpha and high-glucose-elicited phosphorylation of STAT3. Finally, depletion of Nox4 by adenovirus-delivered Nox4 small interfering RNA significantly decreased retinal NADPH oxidase activity and VEGF expression and reduced retinal vascular premeability in db/db mice.

Conclusions: Activation of Nox4 plays an important role in high-glucose- and hypoxia-mediated VEGF expression and diabetes-induced BRB breakdown. Inhibition of Nox4, at least in part, contributes to the protective effects of lovastatin in diabetic retinopathy.

Show MeSH
Related in: MedlinePlus