Differential effects of curcumin on vasoactive factors in the diabetic rat heart

Abstract

Background: Increased oxidative stress has been associated with the pathogenesis of chronic diabetic complications, including cardiomyopathy. Recent studies indicate that curcumin, a potent antioxidant, may be beneficial in preventing diabetes-induced oxidative stress and subsequent secondary complications. We have investigated the effects of curcumin on the nitric oxide (NO) pathway in cardiac tissues and cultured cells.

Methods: Streptozotocin-induced diabetic rats were treated with curcumin for a period of one month. Heart tissues were then analyzed for endothelial NO synthase (eNOS) and inducible NO synthase (iNOS) mRNA expression. Oxidative protein and DNA damage were assessed by immunohistochemical analysis of nitrotyrosine and 8-hydroxy-2'-deoxyguanosine (8-OHdG). Heart tissues were further subjected to endothelin-1 (ET-1) mRNA expression. In order to further characterize the effects of curcumin, we assayed microvascular endothelial cells (MVECs). Cultured MVECs, exposed either to glucose or glucose and varying concentrations of curcumin, were assessed for alterations of NOS expression and activation of nuclear factor-kappaB (NF-kappaB) and activating protein-1 (AP-1). Oxidative stress and ET-1 expression levels were also assayed.

Results: Our results indicate that one month of diabetes causes an upregulation of both eNOS and iNOS mRNA levels, and nitrotyrosine and 8-OHdG immunoreactivity in the heart. Treatment of diabetic rats with curcumin reduced eNOS and iNOS levels in association with reduced oxidative DNA and protein damage. Interestingly, curcumin further increased vasoconstrictor ET-1 in the heart. Exposure of MVECs to high glucose increased both eNOS and iNOS levels and oxidative stress. Curcumin prevented NOS alteration and oxidative stress in a dose-dependent manner which was mediated by nuclear factor-kappaB and activating protein-1. Exposure to curcumin also increased ET-1 levels in the MVECs.

Conclusion: Our studies indicate the differential effects of curcumin in vasoactive factor expression in the heart and indicate the importance of tissue microenvironment in the treatment of diabetic complications.

Figure 1

Effect of curcumin on diabetes-induced vasoactive factor expression, showing real time RT-PCR analysis of (A) NOS, and (B) ET-1 transcript levels in the heart tissues [CO, control; DM, diabetics; DM-C, diabetics treated with 150 mg/kg/d curcumin; *p < 0.05 compared to CO; †p < 0.05 compared to DM; n = 4/group].

Figure 2

Diabetes-induced oxidative stress as assessed by (A) nitrotyrosine, and (B) 8-OHdG staining. Curcumin decreased the level of oxidative DNA and protein damage [Original magnification X400; *p < 0.05 compared to CO; †p < 0.05 compared to DM].

Figure 3

Effect of high glucose levels (A) and curcumin (B) on NOS mRNA levels and transcription factor activity in ECs. ECs exposed to 25 mM glucose were treated with varying curcumin concentrations (0.1, 10, and 100 μM in B) and assayed for eNOS and iNOS expression (A and B) and transcription factor activity (C and D) [CO, 5 mM glucose; HG, 25 mM glucose; HG-C, HG + curcumin (μM); *p < 0.05 compared to CO; †p < 0.05 compared to HG; ‡p < 0.05 compared to 0.1 μM].

Figure 4

ET-1 (A) and HO (B) expression in ECs cultured with 25 mM glucose and 100 μM curcumin as assessed by real time RT-PCR [HG-C, HG + 100 μM curcumin; *p < 0.05 compared to CO; †p < 0.05 compared to HG].