Protective effect of Sesbania grandiflora on acetic acid induced ulcerative colitis in mice by inhibition of TNF-α and IL-6

Abstract

Ethnopharmacological relevance

The plant Sesbania grandiflora (Linn) belonging to the family Fabaceae is commonly known as sesbania, agathi, and katurai. The plant is accredited for alleviating a spectrum of ailments including inflammation, colitis, diarrhea, dysentery, leprosy, gout, rheumatism, jaundice, bronchitis, convulsion and anxiety. It is also used as antitumour, anthelmintic, and laxatives in Ayurveda and Siddha system of Indian traditional medicine.

Aim

To reveal protective effect of Sesbania grandiflora in acetic acid induced ulcerative colitis in mice.

Materials and method

Polyphenol, flavonoid and flavanone contents of different extracts of S. grandiflora leaves were quantified and correlated with their antioxidant capacity in-vitro (DPPH assay) for identification of potential fraction. In further studies hydroalocholic extract (HASG, 100 and 200 mg/kg) was evaluated for protective effect towards acetic acid induced ulcerative colitis (UC) animals administered with 150 µl of 5% acetic acid once, intrarectally. The colonic mucosal injury was assessed by estimating disease activity index (DAI), which took into account weight loss, stool consistency and occult/gross bleeding. Macroscopic changes like colon length, spleen weights, ulcer area and ulcer index were determined. Haematological parameters like WBC count, RBC count, Hb (g/dL), HCT (%), PLT count and FFA level were determined. Biochemical analysis was carried out for asserting the levels of tissue myeloperoxidase (MPO) accumulation, SOD concentrations, reduced GSH and lipid peroxidation in UC induced and treated animals. The cardinal inflammatory biomarkers like nitric oxide (NO), tumor necrosis factor-α (TNF-α) and interleukin (IL-6) were determined. Histopathological investigation was carried out and scores were calculated.

Results

HASG showed presence of highly polymerized polyphenols and flavonoids amongst other extracts of S. grandiflora, which is correlated to its rich antioxidant potential (IC₅₀ =19.21). HPLC fingerprinting quantifies the presence of quercetin in concentration of 81.7 µg/mg of HASG. HASG (200 mg/kg) and Prednisolone (2 mg/kg) significantly reduced DAI and macroscopic scores. The haematological changes in experimental animals were restored upon treatment with HASG and Prednisolone. HASG showed potent antioxidant activity (In-vivo) by restoring the levels of SOD, GSH, MPO, MDA and NO. HASG was found to inhibit FFA levels, which may indicate inhibition of TLR4 receptor mediated inflammation. The levels of serological biomarkers like TNF-α and IL-6 were found to be suppressed. Histopathological investigation reveals decrease signs of ulceration, necrosis, cellular infiltration, hyperaemia in HASG treated animals. The results of HASG (200 mg/kg) were found to be comparable with Prednisolone (2 mg/kg) significantly.

Conclusion

The protective action of HASG against acetic acid induced UC is attributed to the antioxidant like action (In-vitro and In-vivo) of highly polymerized polyphenols and flavonoids especially quercetin. Also HASG was found to reduce the levels of TNF-α and IL-6, thereby suppressing their inflammatory response in UC.

Introduction

Ulcerative colitis (UC) is a debilitating, recurrent chronic inflammatory bowel disorder (IBD) that afflicts large population throughout the world. UC results into formation of inflammatory lesions in mucosal and sub-mucosal layers of only colon with clinical manifestation like abdominal pain, purulent stool and colonic edema. The consistent pain and bowel urgencies impair quality of personal and social life. It also reduces working ability of patient (El-Meligy et al., 2013). The disease is more common in western populations especially in urban rather than rural areas however the annual incidence rate shows high prevalence of IBD day by day in tropical countries like India and Southeast Asia (Ray et al., 2016).

Allopathic therapeutic strategies for UC include medicines like salicylate, corticosteroids, antimicrobials and immunosuppressive agents. However, these agents produce serious side effects like diarrhea, abdominal cramps with pain and discomfort on chronic use (Xu et al., 2004). Herbal medicines had been proved to be effective for chronic inflammatory ailments like UC with lesser side effects, however mystery over their molecular mechanism are still unresolved and needs further exploration (El-Meligy et al., 2013). The secondary plant metabolites like flavonoids and polyphenols have been shown to exert pleiotropic effects both in-vivo and in-vitro for treatment of ailments like UC (Guruvayoorappan et al., 2013; Shigeshiro et al., 2013). The mechanism of actions suggested are free radical scavenging capacity (Biasi et al., 2011, Guazelli et al., 2013) and inhibition of NF-kB signalling in intestinal mucosa (Romier et al., 2009). In addition, these agents were found to reduce lipid peroxidations and increase the levels of antioxidant defence components like SOD and GSH (Dodda et al., 2014). They are also reported to decrease the level of COX2 and iNOS enzymes (Abdallah et al., 2011). Flavonoids and polyphenols were found to be effective by interacting with toll like receptors (TLRs) (Moura et al., 2015, Turner et al., 2011 and Jungbauer et al., 2012) and seems to maintain tight junction integrity of intestinal mucosa during colitis to prevent neutrophil and bacterial toxins infiltration (Piechota-Polanczyk et al., 2014).

The plant Sesbania grandiflora (Family: Leguminosae) is a traditionally revered plant for internal consumption and medicinal values from the era of Maharshi Agasthya (learned monk from ancient India) past several centuries. This plant goes as a principal ingredient in the preparation of ayurvedic medicinal preparation like Grahani kapata rasa (maintains the balance of Vata and Kapha and used in digestive diseases). The leaves and bark of the plants are used in treatment of colitis, ulcers of mouth and alimentary canal, stomach disorders like diarrhea and dysentery of infants and also in stomatitis (Kritikar, 1993, Duke and Wain, 1981). Sesbania grandiflora has been reported for its hepatoprotective activities due to its antioxidant potential (Kasture et al., 2002; Pari and Uma, 2003). Sesbania grandiflora had gain the attention of author for exploration of its protective effect in IBD due to presence of high amount flavonoids like quercetin, myricetin and kaempferol (Krasaekoopt et al., 2005; Mustafa et al., 2010). Sesbania extract shows quenching capabilities against ROS which is also a major cause of mucosal destruction in IBD (Ramesh et al., 2008, Doddola et al., 2008, Sreelatha et al., 2011, Anto et al., 2010, Kumaravel et al., 2011). Polyphenol from S. grandiflora has demonstrated inhibitory effect on some pathogenic bacteria and stimulated growth of probiotic organisms like Lactobacillus acidophilus (China et al., 2012). Considering the traditional knowledge, ethnomedicinal uses and available reported information on plant the investigation was preceded. In the present study we attempted to evaluate the potency of hydroalcoholic extract of S. grandiflora in mouse model of UC. Mice were injected intra-rectally with acetic acid to develop UC. These animals were treated with HASG and results were compared with UC group receiving prednisolone, a standard clinical therapy for UC. Protective mechanism of HASG in UC was investigated in the context of different stages of disease progression (Fig. 1).