Cinnamon extract reduces symptoms, inflammatory mediators and mast cell markers in murine IL-10^−/− colitis

Abstract

Inflammatory bowel disease (IBD) shows an increasing prevalence and harm in western countries. Conventional therapies are associated with bad compliance and adverse side effects. Natural substances like cinnamon extract (CE) could be an additional therapy. We found recently that CE acts anti-inflammatory on mast cells — discussed of being relevant in IBD. Here, we analysed the effects of CE on murine IL-10^−/− colitis as model for IBD. Mice were treated 12 weeks with or without CE in drinking water. Clinical scores and disease activity index were assessed. Colonic tissue samples were analysed for infiltration, tissue damage, bowel wall thickness, expression of pro-inflammatory mediators, mast cell proteases, tight junction proteins, and NF-κB signaling. Following treatment with CE, symptoms of murine colitis as well as increased infiltration of immune cells, tissue damage and bowel wall thickness in colon tissue of IL-10^−/− mice were diminished significantly. MIP-2, TNF, IFNγ, CCL2, CCL3, CCL4 and IL-1β as well as MC-CPA, MCP-1 and MCP-4 were strongly upregulated in IL-10^−/− mice compared to WT, but noteworthy not in CE group. Expression of tight junction proteins was not influenced by CE. Phosphorylation of IκB was slightly down-regulated in CE treated IL-10^−/− mice compared to IL-10^−/− controls. In summary, CE decreases inflammatory symptoms and expression of inflammatory markers in murine IL-10^−/− colitis. CE has no influence on tight junction proteins, but seems acting via reducing pro-inflammatory mediators and recruitment of neutrophil granulocytes probably by inhibiting NF-κB signaling.

Introduction

Inflammatory bowel disease (IBD) is a chronic immune-mediated inflammatory condition of the intestinal tract connected with strong symptoms like diarrhoea [1]. The prevalence of IBD even increased in the last decades in western society where the highest IBD prevalence could be detected. Clinically, the disease is characterized by two phenotypes — Crohn’s disease (CD) and ulcerative colitis (UC) — affecting the colon (CD and UC) and/or distal small intestine (CD) superficially (UC) or in a transmural manner (CD) [2]. Unfortunately, so far medications like aminosalicylates, antibiotics, immunosuppressants, anti-TNF agents, corticosteroids and non-steroidal anti-inflammatory drugs are often associated with adverse side effects like nausea, headache, diarrhoea or even more severe reactions [2], [3]. Therefore, concerning the current compliance rate studies report about 30–60% of patients being non-adherent [4], [5]. Because of its multifunctional background, the causalities of IBD are not yet fully revealed, but a combination of a genetic predisposition and environmental factors is discussed [1], [2]. An uncontrolled immune response to components of the normal intestinal microbiota together with defects in intestinal barrier function may play a role in pathogenesis [1]. Because of missing therapeutic strategies, new additional substances with better compliance and lower side effects among afflicted persons — like natural plant substances with immunomodulatory activities — would be of great interest.

Cinnamon extract (CE) from cinnamon bark of Cinnamomum ceylanicum is known for its anti-oxidative, anti-diabetic, anti-neoplastic, anti-allergic and anti-inflammatory effects in vivo and in vitro [6], [7], [8], [9], [10]. We recently found that CE inhibits activation of mast cells isolated from human intestinal tissue [11]. CE decreases release of stored and de novo synthesized mast cell mediators like β-hexosaminidase and cysteinyl leukotrienes, but also expression of diverse pro-inflammatory cytokines [11]. CE also shows anti-inflammatory activities in LPS-activated macrophages by inhibiting production of NO, cyclooxygenase-2 and prostaglandin E2 as well as MyD88, iNOS and TNF expression via suppressing NF-κB pathway [12]. First hints exist suggesting a protective role of cinnamon in acute murine colitis [13], [14], but its influence on chronic colitis better resembling IBD has not been analysed so far. In acute TNBS-induced colitis, it was shown that CE possesses an inhibitory effect on symptoms of colitis as well as on infiltration and tissue damage [14]. Moreover, in this colitis model cinnamon oil decreased bacterial translocation induced by intestinal mucosal barrier disruption [13] suggesting a role of cinnamon in protecting intestinal barrier function.

A number of murine models of mucosal inflammation resembling IBD exist [15]. Genetic murine IL-10^−/− model — developed by Kühn et al. in 1993 [16] — shows similar histology, physiology and biochemistry as human IBD — especially as Crohn’s disease — with a chronic transmural enterocolitis [17]. The main part of the intestine influenced by inflammation is the colon — when kept in specific pathogen free conditions inflammation is even limited to colon [18], [19]. For IL-10^−/− Balbc mice, the expected rate of spontaneous colitis until an age of 12 weeks is about 69 % [18]. IL-10 principally produced by activated macrophages and Th2 cells promotes humoral, Th2 cytokine-driven immune responses and inhibits Th1 immune responses [19]. Colitis in IL-10^−/− is mediated by unregulated actions of Th1-type CD4⁺ T cells dependent on a dysregulated IL-12 production by macrophages [19]. Th1-type CD4⁺ T cells produce large amounts of IFNγ and TNF leading to colitis [19].

Mast cell degranulation seems to be relevant for IBD pathogenesis [20]. It is supposed that mast cells can influence the epithelial barrier function by acting on expression and distribution of tight junction proteins via proteases [21]. Moreover, mast cell mediators like chemokines can influence inflammation by acting on recruitment and activation of immune cells like neutrophil granulocytes [22], [23]. After mast cell stimulation, release of a wide range of stored mediators like histamine and mast cell specific proteases like mast cell carboxypeptidase A (MC-CPA), chymases MCP-1, -2, -4, -5 and tryptase MCP-6 is induced, as well as de novo synthesis of diverse pro-inflammatory cytokines and chemokines like TNF, Gro-β, IL-1β [21], [24], [25], [26]. TNF can stimulate ion secretion from intestinal epithelium [27]. In humans, mast cell tryptase can stimulate release of CXCL8 from epithelial cells [28] and human chymase is known to alter epithelial permeability in vitro [29].

Here, we examined if CE shows beneficial effects on murine IL-10^−/− colitis concerning its symptoms, inflammatory and mast cell markers as well as epithelial barrier dysfunction and therefore could be considered as possible anti-inflammatory substance in inflammatory intestinal diseases, e.g. for the use as nutraceutical.