NF-κB/twist mediated regulation of colonic inflammation by lupeol in abating dextran sodium sulfate induced colitis in mice

Highlights

• An increased incidence of colon cancer has been observed in patients with colitis.

• DSS induced colitis imitates human colitis, by altering mucosal/inflammatory system.

• Lupeol (Lup) has anti colon cancer property, with no proof as protective for colitis.

• Lup alters mucin expression linked with inflammation, favoring colonic regeneration.

• Lup mediated NF-κB regulation in repressing colitis signifies the anti-inflammatory property.

Abstract

Due to the idiopathic inflammation of the gastrointestinal tract and ulcerative colitis, there is an increased risk of colorectal cancer in inflammatory bowel disease (IBD) patients. Chemoprevention remains vital and much interest has been focused on targetable inflammatory proteins associated with IBD. Dextran sodium sulfate (DSS) induced colitis mimics human IBD in mouse, which is used as a model to determine the action of Lupeol (a triterpene), found in certain fruits and Indian herbs, in the healing of colitis. DSS causes chronic inflammation of the colonic epithelium by altering mucosal immune system i.e. mucins and inflammatory proteins namely TNF-α, IL 1 and 2 and NF-κB. Lupeol supplement to colitis mice, showed colonic regeneration by altering the MUC 1 & 2 gene expression. Lupeol also modulates TWIST gene expression, NF-κB, mediated inflammation and assists down regulation of TNF-α and IL genes. Thus Lupeol has potent colonic healing activity and also acts as an anti-inflammatory agent in controlling colitis.

Introduction

After Crohn and Rosenberg’s (1925) first description about Inflammatory Bowel Disease-Colorectal cancer (IBD-CRC), several epidemiological studies have confirmed that there is an increased risk of CRC in IBD patients (Eaden, Abrams, & Mayberry, 2001). Idiopathic inflammation of the gastrointestinal tract causes IBD and when the inflammation proximally starts from the anal verge and spreads to all or part of the colon it is called ulcerative colitis (UC). Several epidemiological studies suggest that patients with IBD have a 20-fold increased risk for CRC. 10–15% of all deaths in IBD patients were due to the occurrence of colon cancer, where the severity of inflammation plays a vital role (María Luque Cabal, 2011, Munkholm, 2003).

The incidence of CRC in India, is considerably lower than that in western countries (Parkin, Bray, Ferlay, & Pisani, 2001). Population based time trend studies show an increasing trend in the incidence of CRC in India (Yeole, 2008). Studies from Northern India reported a population prevalence of UC of approximately 42 per 100,000 persons with a crude incidence rate of 6 per 100,000 (Makharia et al., 2012). This emphasises a remarkable increase in the incidence of CRC in India, due to change in life style, diet, stress and microbial agents which are the well-recognized risk factors associated with IBD.

While the exact cause of IBD is not completely identified, it is known to involve an interaction between the inflammatory genes, the mucosal immune system, and environmental factors. The pathogenesis of IBD-CRC is exemplified by chronic inflammation of the colonic epithelium leading to a depletion of the goblet cells, disruption of crypt morphology, ulceration, and an increased risk of carcinoma. Epithelial integrity, mucus production, and the presence and equilibrium of bacteria work as intestinal barriers. The adherent mucus layer of the gastrointestinal tract plays a vital physiological part as a lubricant and as a protective barrier against luminal contents (Corfield et al., 2000). The varying constituents and rapid turnover of the mucus barrier are vital factors in response to infection, a form of innate/adaptive immune response. The mucus layer is produced by specialized secretory cells that are found throughout the intestinal tract. Mucus is mainly composed of secretory mucin and membrane bound mucin. Studies have focused on the role of the alterations of these proteins in intestine and their association with IBD.

Intact epithelia are major sources of inflammatory mediators (e.g. TNF-α and IL-8) that initiate and amplify host responses to infection (Gao et al., 2013). A significant balance between pro- and anti-inflammatory pathways needs to be maintained for a homeostatic environment at epithelia to minimize tissue damage. Nuclear factor-κB (NF-κB) is found to be a vital molecule in the pathophysiology of inflammatory cascades and important in the progression of IBD (Marrero et al., 2000).

Chemoprevention of IBD remains an important goal, and surveillance programs are critical for early detection in these patients. The periods of active disease and remission of the colitis might be reduced by conventional treatment but the patients become refractory with side effects. Cellular and molecular evidence obtained from our previous study (Kumar, Raja, Ilakkiya, & Devaraj, 2014) suggest that crude extracts of A. marmelos inhibit DSS induced colonic inflammation at concentrations that showed no apparent toxicity in normal animals. This biological activity of A. marmelos extract was mainly due to the presence of its active components. Hence, in this study, the biological effect of lupeol-a lead molecule of A. marmelos extract, on the DSS induced colitis was studied.

Lupeol (Lup-20(29)-en-3b-ol) (Lup), a triterpene found in edible plants such as olive, fig, mango, strawberry, red grapes, vegetables, and medicinal plants, possesses strong antioxidant, anti-inflammatory, anti-arthritic, anti-mutagenic and anti-malarial activity in both in vitro and in vivo systems (Saleem, 2009). In this study, dextran sodium sulfate (DSS) induced colitis model, which mimics human IBD in mouse was used. This widely accepted model of experimental colitis was used to test lupeol’s potency in the healing/remission of colitis and to delineate its mechanism of action. In addition to the wide pharmacological activities, it was demonstrated, that lupeol regulates mucin gene expression which was linked with the inflammation regulation of repressing colitis.