Curcumin, the major component of food flavour turmeric, reduces mucosal injury in trinitrobenzene sulphonic acid-induced colitis

Abstract

1 Inflammmatory bowel disease (IBD) is characterized by oxidative and nitrosative stress, leucocyte infiltration and upregulation of proinflammatory cytokines. In this study, we have investigated the protective effects of curcumin, an anti-inflammatory and antioxidant food derivative, on 2,4,6- trinitrobenzene sulphonic acid-induced colitis in mice, a model for IBD. 2 Intestinal lesions (judged by macroscopic and histological score) were associated with neutrophil infiltration (measured as increase in myeloperoxidase activity in the mucosa), increased serine protease activity (may be involved in the degradation of colonic tissue) and high levels of malondialdehyde (an indicator of lipid peroxidation). 3 Dose-response studies revealed that pretreatment of mice with curcumin (50 mg kg(-1) daily i.g. for 10 days) significantly ameliorated the appearance of diarrhoea and the disruption of colonic architecture. Higher doses (100 and 300 mg kg(-1)) had comparable effects. 4 In curcumin-pretreated mice, there was a significant reduction in the degree of both neutrophil infiltration (measured as decrease in myeloperoxidase activity) and lipid peroxidation (measured as decrease in malondialdehyde activity) in the inflamed colon as well as decreased serine protease activity. 5 Curcumin also reduced the levels of nitric oxide (NO) and O(2)(-) associated with the favourable expression of Th1 and Th2 cytokines and inducible NO synthase. Consistent with these observations, nuclear factor-kappaB activation in colonic mucosa was suppressed in the curcumin-treated mice. 6 These findings suggest that curcumin or diferuloylmethane, a major component of the food flavour turmeric, exerts beneficial effects in experimental colitis and may, therefore, be useful in the treatment of IBD.

Figure 1

Effect of curcumin pretreatment on the macroscopic damage score of colonic tissues after induction of colitis. Mice were treated with 0.1 ml of TNBS (60 mg ml⁻¹) intracolonically and assessed at various times (2–8 days) after treatment. Colonic damage was scored on a scale of 0 (normal) to 5 (severe) by two independent observers. Values are means±s.d. of 10 rats for each group. Results for the curcumin treatment group were compared against those for the TNBS-treated group with a two-sided Wilcoxon's rank-sum test. ^*P<0.001 vs TNBS.

Figure 2

Effect of curcumin on colon injury. (a) Mucosal from control mice did not show any histological modifications, (b) TNBS-induced mucosal injury (at 4 days) associated with transmural necrosis and oedema and submucosal infiltration of inflammatory cells and (c) pretreatment with curcumin prevented the disturbances in morphology associated with TNBS treatment. Original magnification: × 500.

Figure 3

Effect of curcumin pretreatment on (a) spleen, (b) colon and (c) body weight. A significant increase in weight was observed at 4 days after TNBS administration in spleen and colon. Curcumin pretreatment (50, 100 and 300 mg kg⁻¹) significantly prevented the loss in body weight (c) as well as reduced the organ weight (a and b). Values are means±s.d. of 10 rats for each group. ^*P<0.01 vs TNBS.

Figure 4

Effect of curcumin on NO and O₂⁻ production. (a) NO production in colonic tissue and (b) O₂⁻ production in neutrophils. TNBS treatment caused a significant increase of both NO and O₂⁻ generation, which were prevented by curcumin pretreatment. Values are means±s.d. of 10 rats for each group. ^*P<0.01 vs TNBS.

Figure 5

Effect of curcumin on neutrophil infiltration and lipid peroxidation. (a) Myeloperoxidase activity and (b) malondialdehyde levels in the colon. Myeloperoxidase activity and malondialdehyde levels were significantly increased in TNBS-treated mice in comparison to control. Curcumin pretreatment showed a significant reduction of both myeloperoxidase activity and malondialdehyde levels. Values are means±s.d. of 10 rats for each group. ^*P<0.01 vs TNBS.

Figure 6

Protease activity in TNBS-treated animals. (a) TNBS treatment showed a several-fold increase in protease activity (lane 2) over the untreated control animals (lane 1). Curcumin-pretreated mice showed a marked reduction in protease activity (lane 3) compared to TNBS-treated mice. Numbers on the right indicate molecular mass in kDa. (b) Band intensities were quantified by densitometry.

Figure 7

Suppression of Th1 phenotype in TNBS-treated mice subjected to curcumin pretreatment (50 mg kg⁻¹) as analysed by RT–PCR. (a) Expression of IFN-γ, IL-12, IL-4, iNOS and β-actin mRNA by colon tissue samples of control and treated mice. RT–PCR products were visualized by ethidium bromide staining. RNA samples were obtained from six mice in each group. Results are representative of three separate samples. β-actin expression levels were used as controls for RNA content and integrity. (b) Band intensities were quantified by densitometry.

Figure 8

Effect of curcumin pretreatment on NF-κB activation. NF-κB activity was upregulated in nuclear extracts from TNBS-induced colonic mucosa and suppressed by curcumin pretreatment. Nuclear extracts of the colonic tissue from control mice (lane 1), untreated mice with TNBS-induced colitis in absence (lane 2) and presence of 50 molar excess of unlabelled probe (lane 5) and curcumin (300 and 50 mg kg⁻¹) treated mice with TNBS-induced colitis (lane 3 and 4) were analysed.