Inflammatory bowel disease (IBD) is a chronic inflammatory disease of unknown cause, and effective treatment options have not been discovered [1]. Recent data increasingly indicate that signaling molecules and pathways are responsible for the onset and progression of the disease. There is evidence that the increased regulation of inflammatory mediators, such as cytokines, chemokines, and adhesion molecules, plays an essential role in both human and experimental colitis models [2,3]. Nuclear factor-kappa B (NF-κB) is responsible for the gene expression regulation of inflammatory mediators such as tumor necrosis factor-α (TNF-α), interleukin 6 (IL-6), and nitric oxide synthase (iNOS) [4,5]. NF-κB is a key molecule involved in intestinal inflammation [6]. Studies that investigate the effect of anti-inflammatory drugs on the insula in IBD are difficult. For this reason, studies use animal models to investigate approaches to reduce inflammation and to activate various pathways.

Hericium erinaceus (HE), known as lion’s mane, is a medicinal mushroom and has been historical used in traditional Chinese medicine. HE mycelium and fruit bodies show great promise as substances that improve health such as brain and nerve health. The animal and in vitro experiments that investigated substances extracted from HE showed good results. In particular, the mycelium extract has been used clinically as an anti-inflammatory agent and in cytotoxic and cardiovascular antineoplastic research as an antibacterial and neuroprotective agent [7,8]. HE represents an important opportunity to treat Alzheimer’s and Parkinson’s diseases because HE is used as an anti-inflammatory medicine and promotes nerve growth factor gene expression.

The aim of this study was to evaluate the protective effect of HE treatment in a TNBS-induced colitis rat model.

Twenty-four Wistar albino rats (225 to 275 g) were included in this study. Rats were kept in wire-mesh bottomed cages under a 12-hour light/12-hour dark cycle at a constant room temperature (22ºC ± 2ºC). A standard chow diet and water were provided. The study was approved by the Istanbul University ethics committee (number 2013-88).

HE is an edible and medicinal mushroom and has also been used as a traditional Chinese medicine. Various studies have shown that the content of the HE is useful for medical purposes because of its high protein, low fat and high nutritional value [12,13]. HE contains erinaceus hericenone, CH [14,15], erinacines AK [16,17], orcinol derivatives (mycelium) [18], sialic acid-binding lectin [19], ergosterol and beta-sitosterol [20]. Since hericenone and erinacine strongly stimulate nerve growth factor synthesis, which allows for neuron repair and regeneration, HE increases nerve growth factor synthesis, and its compound activity slows neuronal cell death [17]. For this reason, HE can be used as a nutritional therapy in the treatment of dementia, Alzheimer’s disease and Parkinson’s disease. The orcinol derivatives obtained from the mycelium of HE show antibacterial and antifungal activity against certain pathogenic microorganisms (such as Bacillus subtilis and Saccharomyces cerevisiae) [21]. Another component of HE, sialic acid-binding lectin, can be used to identify, quantify, localize, purify, and characterize many biomolecules (such as glycoconjugates). Sialic acid plays an important role at the protein and cell levels in the biological recognition mechanisms, and lectins can be used as specific probes for sialic acid species, which act as molecular markers in pathological and physiological development [22]. Ergosterol is a major fungal sterol, and beta-sitosterol reduces the absorption of cholesterol in the intestinal lumen. In the study by Mbambo et al. [23], these stressors were shown to have antifungal activity. HE also has a prebiotic effect; it contained the polysaccharide β-glucan and the monosaccharides mannose (approximately 55.70%), arabinose (approximately 0.7%), rhamnose (approximately 3.75%), xylose (approximately 1.60%), galactose (approximately 5.12%), and glucose (approximately 33.10%) [24]. Since HE contains physiologically important compounds, there are some studies on gastritis and gastrointestinal infections. Extracts of HE mycelium have been indicated to inhibit gastric mucosal damage and improve gastric ulcers [25], with the inclusion of HE in the diet improved colonic health and IBD, and HE has anti-inflammatory activity [26,27].

The results of our study suggest that treatment with HE has a positive impact on colitis in TNBS-induced rats by reducing oxidative damage in blood and tissue. This is indicated by the significantly decreased NF-κB, MDA, NO, IL6, and TNF-α levels in blood and tissue. Additionally, the current study shows macroscopic and microscopic colitis scores that indicate less mucosal damage and improved mucosal structure.

IBD is a chronic, recurrent disease characterized by idiopathic inflammation of the gastrointestinal tract whose pathogenesis is not known precisely. IBD mainly consists of two clinical forms, Crohn’s disease (CD) and ulcerative colitis (UC). UC is characterized by recurrent weakness of the mucosal layer of the colon [28]. It is most commonly seen in the rectum, spreading to other parts of the colon with continuity. CD is a disease of the digestive tract that may be anywhere from oral to annular, with focal, asymmetric, or transmural involvement [29]. Some clinical studies have shown that genetic factors increase the risk of developing IBD [30]. The effects of environmental factors on the formation of IBD and processes are not fully understood. Clinical and epidemiological studies indicate that this disease can develop in genetically susceptible individuals after exposure to different antigens or environmental factors. Exacerbations and healing may be associated with these factors. According to our results, HE can be a protective agent for IBD.

Immune factors play an effective role in the pathogenesis of IBD. Some cytokines are proinflammatory (IL-1, IL-2, IL-6, and TNF-α), and some are anti-inflammatory (IL-8, IL-10, IL-11, and TNF-γ). HE treatment improved the inflammatory response of rats because treatment of TNBS-induced rats with HE resulted in increased levels of some anti-inflammatory cytokines compared to the untreated group [24]. Our study also showed that feeding TNBS-treated rats HE had a good inflammatory effect because the TNF-α and IL-6 levels of HE-fed rats were lower than those of the control group. It was also demonstrated that the administration of HE decreased inflammation, MDA levels and MPO activity in rats with TNBS-induced colitis.

Several compounds that initiate the immune response in the digestive tract have been identified, and experimental models, in which these compounds are generated, have been used to understand the pathogenesis of the disease and to develop drug treatments. TNBS is one of these compounds, and the disease model created by this substance is widely used in experimental studies since it produces effects similar to human UC [31,32]. In this study, cytokines such as TNF-α, MPO, NO, MDA, and IL-6 were investigated to determine the cellular activities of the TNBS-induced colitis model because cytokines are key signaling molecules of the intestinal immune system that play an important role in IBD. In vivo and in vitro studies indicate that the levels of proinflammatory cytokines such as TNF-α and IL-6 are elevated in IBD and that the severity of inflammation is directly proportional to the levels of these cytokines [33]. Moreover, the macroscopic colitis score of this study showed that colonic wall thickness, ulcers, severe adhesions between the colon and other organ hyperemia were observed on the 7th day in TNBS-induced rats. The comparison of the microscopic colitis scores of the TNBS-induced group and HE treatment group showed that the HE treatment group had less mucosal injury, improved mucosal structure, and epithelial integrity. In one study, the HE-treated group showed more pronounced improvement than did the TNBS-induced group. Mucosal erosion and ulcers were observed in the TNBS-induced group, and less injury was observed in the HE group than in the TNBS group. Our results clearly demonstrated that the HE extracts could promote the protection of the gastric mucosa and give positive results in the experimental models of IBD.

One previous study showed that HE extracts contributed to the growth of beneficial gut bacteria and ameliorated host immunity in an experimental IBD model. When TNBS-induced rats were treated with HE extracts, the expression levels of inflammatory factors were changed. MPO levels declined, and TNF-α levels in the peripheral blood and colon of the normal group were lower than those in TNBS-treated rats. In our study, according to the biochemical results, there was a decrease in MDA, NO, IL6, and TNF-α levels in the HE treatment group compared with the TNBS-induced colitis group. Treatment with HE enhanced all the inflammatory responses that we investigated.

Excess TNF-α expression damages epithelial barriers and induces apoptosis in epithelial cells [34]. In accordance with the literature, TNF-a expression was more intense in the areas of inflammation and ulceration in the colon mucosa of the colitis group. Zhou et al. [35] showed that TNF-a expression was increased in a TNBS-induced colitis model. This study showed that HE treatment decreased TNF-α and IL-6 levels because the TNF-α and IL-6 levels in the TNBS-induced colitis model were significantly higher than those in the HE group.

One of the previous observations showed that MPO levels in the HE group were lower than those in the colitis group, and HE extracts decreased colon inflammation in rats with IBD [22]. When MPO activity was examined, the values obtained from the colitis group were considerably higher than those of the control group (p < 0,05), and inflammation was observed. The MPO levels measured at the tissue level were significantly lower in the treatment groups than those in the colitis group (p < 0.05). Furthermore, the MDA level of the colonic tissues was found to be increased in the TNBS-treated group compared to the sham group. Treatment with HE caused a significant decrease in the MDA level in TNBS-induced colitis rats.

NF-κB proteins are transcription factors that control apoptosis, cell cycle progression, cell growth, and differentiation. The NF-κB pathway is involved in colonic inflammation, and increased NF-κB protein expression has been observed in the mucosa of patients with unspecific colitis, UC, and CD [36]. Proinflammatory cytokines induce NF-κB activation in different cell types, and this could be initiated by oxidative stress, which has a significant role in the pathogenesis and progression of IBD. Our biochemical findings also indicate that NF-κB expression decreased with HE administration, whereas NF-κB overexpression was observed in TNBS-induced colitis. The inhibition of NF-κB activation could be a possible treatment strategy for IBD.

As a limitation of this study, we analyzed MDA, NO, TNF-α, MPO, and NF-κB values to measure oxidative damage, which is similar to some studies [5,35,37]. The antioxidant effects of HE, the total antioxidant concentration and the protein expression levels of heme oxygenase-1 (HO-1) and NAD(P)H:quinone oxidoreductase (NQO1). were not analyzed in this study so that there was not much data complexity during the experimental phase and to ensure that the costs of the experiment did not increase. In future studies by our group, these values will be examined.

In conclusion, oral HE treatment showed good efficacy in the experimental colitis model. Biochemical indexes and microscopic and macroscopic colitis scores were analyzed. TNF-α, MPO, NO, MDA, and IL6 cytokines, which affect TBNS-induced colitis models, were examined, and TNF-α, MPO, NO, MDA, and IL6 levels were lower in the HE treatment group than in the colitis groups. Less mucosal injury was detected in the HE treatment group than in the colitis group. In light of these results, HE has clinical potential for the improvement of IBD because HE has an anti-inflammatory properties. The main disadvantage of oral therapy with HE is the unknown dose required for the anti-inflammation effect. For this reason, this issue will be investigated in future studies.