Curcuma longa extract exerts a myorelaxant effect on the ileum and colon in a mouse experimental colitis model, independent of the anti-inflammatory effect

Abstract

Background: Curcuma has long been used as an anti-inflammatory agent in inflammatory bowel disease. Since gastrointestinal motility is impaired in inflammatory states, the aim of this work was to evaluate if Curcuma Longa had any effect on intestinal motility.

Methods: The biological activity of Curcuma extract was evaluated against Carbachol induced contraction in isolated mice intestine. Acute and chronic colitis were induced in Balb/c mice by Dextran Sulphate Sodium administration (5% and 2.5% respectively) and either Curcuma extract (200 mg/kg/day) or placebo was thereafter administered for 7 and 21 days respectively. Spontaneous contractions and the response to Carbachol and Atropine of ileum and colon were studied after colitis induction and Curcuma administration.

Results: Curcuma extract reduced the spontaneous contractions in the ileum and colon; the maximal response to Carbachol was inhibited in a non-competitive and reversible manner. Similar results were obtained in ileum and colon from Curcuma fed mice. DSS administration decreased the motility, mainly in the colon and Curcuma almost restored both the spontaneous contractions and the response to Carbachol after 14 days assumption, compared to standard diet, but a prolonged assumption of Curcuma decreased the spontaneous and Carbachol-induced contractions.

Conclusions: Curcuma extract has a direct and indirect myorelaxant effect on mouse ileum and colon, independent of the anti-inflammatory effect. The indirect effect is reversible and non-competitive with the cholinergic agent. These results suggest the use of curcuma extract as a spasmolytic agent.

Figure 1. Photomicrographs of hematoxylin and eosin staining of Colon sections from DSS (5%w/v over 7 days) treated mice, before(a) and after 7 days Curcuma extract administration (b) and from DSS (2.5%,w/v, three cycles over 52 days) (c) and after 21 days Curcuma extract administration (d).

(a) Intestinal mucosal with edema and marked/moderate acute inflammation composed by neutrophils, lymphocites and plasmacells. The intestinal glands show moderate regenerative hyperplasia.(H&E 10×). (b) Moderate mucosal inflammation composed by lymphocytes and some neutrophils; the intestinal glands show mild regenerative hyperplasia. (H&E 5×). (c) Chronic colitis. The mucosa shows a chronic infiltrate composed by lymphocytes and plasmacells. The glands show mild regenerative hyperplasia. (H&E 5×). (d) Intestinal mucosal with mild inflammation composed by lymphocytes, plasmacells and rare neutrophils. The submucosa shows many foam macrophages (H&E 20×).

Figure 2. A schematic representation of the experimental design.

Figure 3. Inhibition of spontaneous motility of ileum (upper panel) and colon (lower panel) by curcuma extract.

The segments were suspended in organ baths containing gassed warm Krebs solution under a load of 1 g maintained at 37°C. Tension changes in longitudinal muscle length were recorded. In control conditions (a), after curcuma extract additioned medium (b) and after washing (c).For details see Methods section.

Figure 4. Effect of curcuma extract on carbachol-induced contraction in isolated mouse ileum (A) and distal colon (B).

a) Cumulative concentration-response curves were obtained before and after exposure to curcuma extract for 30 min. Each point is the mean ± SEM (n  = 3–5). b) Time course of curcuma extract effect on Carbachol-induced contraction in isolated mouse ileum (A) or distal colon (B) (100%). Cumulative concentration-response curves were obtained before and after exposure to curcuma extract (0.05 mg/mL) for 5, 30 and 45 min. Each point is the mean ± SEM (n  = 3–5). c) Time course of curcuma extract (0.05 mg/mL) on carbachol-induced contraction in isolated mouse ileum (A) and distal colon (B). Cumulative concentration-response curves were obtained before and after exposure to curcuma extract (0.05 mg/mL) and following 5, 30 and 60 min washing. Data are the mean ± SEM (n  = 3–5). Where error bars are not shown, these are covered with the point itself.

Figure 5. Potency of CCh (pEC₅₀) (dark blu) and atropine (pA₂) (light blu) on ileum (a) and on distal colon (red) and (pink) respectively.

(1): control mice. (2): mice with acute DSS induced colitis; (3): acute colitis mice fed the usual diet over 7 days after stopping DSS; (4): acute colitis mice fed curcuma over 7 days after stopping DSS. The dashed line shows the control values for CCh pEC₅₀. The dotted line shows the control values for Atropine pA₂. When error bar are not shown these are covered by the point itself. The contraction induced by Carbachol and the antagonistic effects of Atropine were used as reference. In healthy animals Carbachol is by 3.55 times more potent on the ileum than on the colon. Atropine presents similar antagonistic activity on both tissues: these data have been used as a comparison with the effect of Carbachol and Atropine on tissues from acute colitis animals. Carbachol pEC₅₀ decreased by 3.80 times in the ileum and 1.58 times in the colon and Atropine pA ₂ was reduced by 1.86 and 1.12 times respectively in the ileum and the colon, compared with the corresponding intestinal segments of control mice. Curcuma administration improved the response to Carbachol in the colon, but not in the ileum, also if it did not completely restore the normal values.

Figure 6. Potency of CCh (pD₂) and Atropine (pA₂) on the ileum (a, b, c) and on the colon (d, e, f) from mice with chronic colitis.

1: control mice, 2: chronic DSS induced colitis, 3: DSS chronic colitis mice assuming standard diet over 7 (a) 14 (b) and 21 (c) days after stopping DSS; 4: DSS chronic colitis mice assuming curcuma extract over 7 (a), 14 (b) and 21 (c) days after stopping DSS. The dashed line shows the control values for CCh pEC₅₀. The dotted line shows the control values for Atropine pA₂. When error bar are not shown these are covered with the point itself Carbachol. The contraction induced by Carbachol and the antagonistic effects of Atropine were used as reference. In healthy animals Carbachol is by 3.55 times more potent on the ileum than on the colon. Atropine presents similar antagonistic activity on both tissues: these data have been used as a comparison with the effect of Carbachol and Atropine on tissues from chronic colitis animals. After induction of chronic colitis, Charbacol pEC₅₀ decreased by 8.71 times in the ileum and 2.34 times in the colon and Atropine pA ₂ was increased by 1.35 and 3.09 times in the ileum and the colon respectively, compared with the corresponding intestinal segments of control mice. Curcuma administration improved the response to Carbachol in the ileum by 9.12 times and by 2.45 times in the colon after 14 days administration; Atropine pA₂ was decreased by 1.78 and 1.66 respectively in the two intestinal segments.