Protective potential of curcumin in L-NAME-induced hypertensive rat model: AT1R, mitochondrial DNA synergy

Abstract

Background & objectives: Hypertension can be induced by inhibiting nitric oxide synthesis with L-NAME, which also has a role in oxidative stress. Curcumin has strong antioxidant property. Our aim was to examine the possible preventive role of curcumin on renal dysfunction secondary to hypertension.

Material & methods: Twenty-four adult male Albino rats were divided in four groups: normal (N); curcumin (C; received curcumin 100 mg/kg/day by oral gavage for 10 weeks); hypertensive (H; received L-NAME 40 mg/kg/day in their drinking water for 4 weeks); and hypertensive-curcumin (HC; received L-NAME and curcumin). Arterial blood pressure was evaluated non-invasively for 4 weeks. Rats were then sacrificed for assessment of oxidative stress (catalase, lipid peroxidase, reduced glutathione and superoxide dismutase), renal function and structure, and biomarkers of apoptosis (Bcl-2 and caspase-3). AT1R expression and renal mtDNA integrity were also assessed.

Results: Curcumin attenuated the effects of L-NAME on blood pressure and renal function. The renal histopathological changes observed in the L-NAME group were improved by curcumin administration. The expression of Bcl2 and caspase-3 was improved associated with downregulation of AT1R in curcumin treated groups. The antioxidant markers and mtDNA fragmentation show marked increase in hypertensive group which significantly decreased after curcumin treatment.

Conclusion: Curcumin improved blood pressure elevation renal dysfunction. These improvements mediated through anti-oxidant capabilities and downregulation of AT1R favoring reduced apoptosis and preserved mitochondrial DNA.

Keywords: AT1R; L-NAME; curcumin; mtDNA; oxidative stress; renal function.

Figure 1

Mean Arterial blood pressure (MABP) in all studied groups. All data are expressed as mean ± SEM and analyzed using one-way ANOVA. *Significant in comparison to normal group. $Significant in comparison to Curcumin group. #Significant in comparison to Hypertensive group.

Figure 2

Kidney function tests (serum urea and Creatinine level) and Plasma nitrate/nitrite levels in all studied groups. All data are expressed as mean ± SEM and analyzed using one-way ANOVA. *Significant in comparison to normal group. $Significant in comparison to Curcumin group. #Significant in comparison to Hypertensive group.

Figure 3

Photomicrographs of renal cortex of all study groups: (A) normal group, (B) curcumin group, (C) hypertensive group and (D) cur + hypertensive group. A &B show renal corpuscles with normal glomerular (G) blood capillaries covered by nuclei of podocytes and mesangial cells and surrounded with urinary space. Proximal convoluted (PT) and the distal convoluted tubules (DT) show normal appearance. It also shows normal blood arterioles (arrows). (C1) Shows collapsed glomeruli (G) or complete loss (LG). The PT and DT show shrinkage of their lining cells with pyknosis of their nuclei. In (C1-I) there is extravasation of RBCs in the renal interstitium (arrows), lymphocytic infiltration (green arrow) and dilated congested capillaries (*). Thickened wall of arterioles (black arrow), are shown in (C1-II). (C2) Shows hyaline material deposits in the renal interstitium (arrows), hydropic degeneration of the cells lining the PCTs (PT) and edematous arteriolar wall (green arrow). (D) Shows few corpuscles have lost glomeruli (★). It shows normal arteriolar wall (green arrow) but surrounded with dilated capillaries (*). No lymphocytic infiltration, RBCs extravasation or hyalinosis is present. H&E ×400.

Figure 4

(A) Photomicrograph for the renal cortex of all study groups: (A-D) PAS ×400. (E-H) Masson’s trichrome ×400. (A, E) normal group, (B, F) curcumin group, (C, G) hypertensive group and (D, H) cur + hypertensive group. In (A, B) there are normal regular basal lamina of glomerular capillaries (G), urinary spaces (arrow heads), continuous with that of PCT whose cells have apical brush border (black arrows). It also shows normal basal lamina of the DCT whose cells has no brush border (green arrows). (C) Shows increase in the PAS stained material. The Bowman capsules show increased thickness of their basal laminae (arrow) and so the glomeruli (G). Thickening in the capillary walls (DC) with hyaline material depositions in the interstitium (white arrows). (D) Shows normal thickness of basal laminae around the renal tubules (PT&DT), renal corpuscles and glomerular capillaries (G). No hyaline deposits are present. (E, F) Have little amount of green material in the interstitium, around tubules and corpuscles (arrow heads), surrounding the arterioles (arrow) and in the mesangium (G). (G) Shows increase in the greenish collagen content around the arterioles, renal tubules and corpuscles (arrows) and in the glomerular mesangium (G). (H) Shows normal distribution and amount of greenish collagen. (B) Mean PAS and Masson stain intensity percentage was quantified by Fiji image analyzer software. All data are expressed as mean ± SEM and analyzed using one-way ANOVA. *Significant in comparison to normal group. $Significant in comparison to Curcumin group. #Significant in comparison to Hypertensive group.

Figure 5

Phptomicrograph of the renal cortex of all study groups: (A) normal group, (B) curcumin group, (C) hypertensive group, (D) cur + hypertensive group. Brown cytoplasmic reaction in the cells linining the tubules. The glomeruli (G) show negative reaction in both stains in all groups. BCL2 & Caspase 3 immunostaining ×200. (E) Bcl-2 and caspase-3 immunohistochemical intensity was quantified by Fiji image analyzer software. All data are expressed as mean ± SEM and analyzed using one-way ANOVA. *Significant in comparison to normal group. $Significant in comparison to Curcumin group. #Significant in comparison to Hypertensive group.

Figure 6

AT1R expression in blood vessels of all studied groups. All data are expressed as mean ± SEM and analyzed using one-way ANOVA. *Significant in comparison to normal group. $Significant in comparison to Curcumin group. #Significant in comparison to Hypertensive group.

Figure 7

Oxidative stress level in renal tissue of all studied groups. All data are expressed as mean ± SEM and analyzed using one-way ANOVA and Bonferroni post-hoc test. *Significant in comparison to normal group. $Significant in comparison to Curcumin group. #Significant in comparison to Hypertensive group.

Figure 8

Mitochondrial DNA (mtDNA) in the experimental groups, Lane M, 1 kb DNA ladder; lane N: intact mtDNA isolated from normal group; lane C: mtDNA samples isolated from curcumin group; lane H: mtDNA samples isolated from hypertensive group and lanes HC: mtDNA sample isolated from treated group.