Anti-inflammatory effects of Radix Aucklandiae herbal preparation ameliorate intestinal mucositis induced by 5-fluorouracil in mice

Abstract

Ethnopharmacological relevance

5-Fluorouracil (5-FU) is a chemotherapy agent that is widely used in clinical oncologic practice. However, intestinal mucositis is the most frequently occurring side effect of cancer therapy with 5-FU. Based on a literature survey, Radix Aucklandiae herbal preparation potentially ameliorates intestinal mucositis in 5-FU-treated mice.

Aim of the study

The aim of this study was to investigate the inflammation and gastrointestinal regulation of intestinal mucositis induced by 5-FU, including the intestinal morphology, as well as the reduction in food intake, body weight loss, and diarrhea.

Materials and methods

Intestinal mucositis was induced in mice by 5-FU (30 mg/kg, i.p., for 5 consecutive days). The dose-dependent Radix Aucklandiae herbal preparation (0.3, 1, and 3 g/kg/day, p.o.), loperamide (3 mg/kg/day, p.o.) or celecoxib (40 mg/kg/day, p.o.) was concurrently administered until the 7th day. Physical status observation, diarrhea assessment, serum proinflammatory cytokine levels, intestinal villus height and crypt depth, and total goblet cells from tissues were assessed.

Results

The dosage regimen of 5-FU administration caused severe intestinal mucositis in mice, including damage to the intestinal morphology, accompanied by a reduction in food intake, body weight loss, and diarrhea. The high-dose Radix Aucklandiae herbal preparation significantly relieves 5-FU-induced intestinal mucositis by enhancing proliferative activity in epithelial crypts; improving anepithymia, body weight loss, and diarrhea; and displaying protective effects on goblet cells in intestinal mucosal epithelia. Activation of NF-κB in the intestinal mucositis model was also suppressed by the Radix Aucklandiae herbal preparation, suggesting that it is a potent inhibitor of NF-κB and proinflammatory cytokines, such as IL-1β, IL-6, TNF-α, and COX-2.

Conclusions

Our data support the conclusion that the Radix Aucklandiae herbal preparation could effectively ameliorate 5-FU-induced gastrointestinal toxicity and be applied clinically for the prevention of intestinal mucositis during chemotherapy.

Introduction

5-Fluorouracil (5-FU) is widely known as one of the most commonly used chemotherapy drugs in clinical oncologic practice, especially for gastrointestinal cancer (Yamada et al., 2014). 5-FU has mainly used as a thymidylate synthase (TS) inhibitor. Interruption of this enzyme prevents the synthesis of pyrimidine thymidine nucleosides, which are required for DNA replication (Longley et al., 2003). Repeated 5-FU treatment causes severe intestinal mucositis, including morphological damage, altered gut motility and pH value, and intestinal crypt damage, which is accompanied by a reduction in food intake (Sakai et al., 2016), body weight loss and diarrhea (Smith et al., 2008; Gosselink et al., 2004). Inflammation, severe ulceration and even hemorrhage may spread all over the gastrointestinal tract, especially in the small intestine (Sonis et al., 2004). However, this compound has serious side effects at high doses. The most common toxic reaction to cancer chemotherapy is intestinal mucositis, and the individual is malnourished tolerance to cancer therapy is reduced (Song et al., 2013). Some studies discovered that 50%–80% of patients undergoing 5-FU chemotherapy develop clinical intestinal mucositis and gastrointestinal symptoms such as vomiting, abdominal bloating, and diarrhea (Benson III et al., 2004; Hamouda et al., 2017). The phenomenon of diarrhea represents a dose limiting toxic event with mucositis and can be a major cause of treatment discontinuation and decreased drug efficacy (Boussios et al., 2012).

Intestinal mucositis is a serious problem in oncology because it not only causes very serious pathologies along the alimentary tract but also impairs the quality of life of oncologic patients and may even be life-threatening, requiring interruption of chemotherapy and termination of treatment (Kuiken et al., 2015; Zur, 2011). In addition, chemotherapy cycles complicated with mucositis may prolong the time and cost of hospitalization (Dekkers, 2009). Loperamide is the most frequently used medication for the treatment of diarrhea with chemotherapy. Nevertheless, previous studies have demonstrated the association of loperamide and other antimotility agents with the pathogenesis of toxic megacolon in patients with gastroenteritis, so the potential risk of loperamide for 5-FU-related diarrhea should be considered. (McGregor et al., 2007). Celecoxib is the selective cyclooxygenase-2 (COX-2) inhibitor, which thought to provide the majority of the beneficial anti-inflammatory effects. Recently discoveries have shown that COX-2 positive advanced gastric cancer patients have benefited are more clinical by celecoxib combined with chemotherapy (Guo et al., 2019). Therefore, loperamide and celecoxib are suitable as the anti-diarrheal and anti-inflammatory positive control group in this study. To date, except for well-known antidiarrheal agents, antibiotics and mucosal protective agents, no sufficient strategies have been developed to manage 5-FU-related intestinal mucositis (Chang et al., 2020).

Intestinal mucositis due to 5-FU is believed to generate an oxidative process, initially generating DNA and non-DNA damage, and then primary mucosal damage will cause epithelial apoptosis and the secretion of inflammatory cytokines. Nuclear factor kappa-B (NF-kB) triggers the production of proinflammatory cytokines such as tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), interleukin-1β (IL-1β) and COX-2, thus resulting in intestinal mucosal damage and dysfunction (Al-Asmari et al., 2015; Chen et al., 2016). The basic layer of the small intestine consists of the epithelial layer that is folded into finger-like protrusions (villi), the lamina propria that forms the villi, and the muscular mucosa (Madara, 2010). Goblet cells play a very important role in the epithelial layer of the small intestine. Mucins synthesized and secreted by intestinal goblet cells constitute the mucus layer on the intestinal epithelial surface, which is the first line of innate host defense mechanism mainly as a mechanical barrier in the intestine (Johansson and Hansson, 2013). However, in addition to a significant decrease in total goblet cell numbers, 5-FU has been shown to increase levels of the proinflammatory cytokines TNF-α, IL-6, and IFN-γ and impede intestinal barrier function (Yeung et al., 2015).

Currently, many cancer patients use alternative therapies to reduce the side effects associated with cancer therapies (Tachjian et al., 2010). Herbal medicines and remedies are the most commonly used complementary and alternative medicine (CAM) therapies. There is a growing body of evidence that certain complementary interventions can help control some of the symptoms and side effects of cancer treatments (Deng and Cassileth, 2005). Cancer patients often combine herbal supplements with conventional treatments in order to improve quality of life, alleviate the side effects of chemotherapy, strengthen the immune system, and slow down the progression of cancer (Sparreboom et al., 2004; Tascilar et al., 2006). In the clinic, some herbs have been shown to ameliorate chemotherapy-induced toxicity and have potentially beneficial effects on cancer progression (Yang et al., 2010).

The Radix Aucklandiae herbal preparation (RAHP) was exactly the same as the Xiang-Sha-Liu-Jun-Zi-Tang (XSLJZT) traditional Chinese formula. According to the National Health Insurance Research Database (NHIRD) in Taiwan, the herbal medicine formula XSLJZT is the most commonly prescribed formulation combined with chemotherapy (Wang et al., 2014; Chao et al., 2014). In a clinical trial, 32 patients with irritable bowel syndrome (IBS) were treated with XSLJZT for 28 days. After observation, oral administration of XSLJZT lowered the mean diarrhea score in patients with IBS demonstrating the significant therapeutic performances of XSLJZT (Shih et al., 2019). In our previous study reported that Identification of RAHP consists of eight Chinese herb substances by HPLC–MS/MS including Radix Aucklandiae (Aucklandia lappa Decne., Asteraceae); Amomi Fructus (Amomum villosum Lour., Zingiberaceae); Pinelliae praeparatum Rhizoma [Pinellia ternate (Thunb.) Breit., Araceae]; Citri reticulatae Pericarpium (Citrus reticulate Blanco, Rutaceae); Ginseng Radix (Panax ginseng C. A. Mey., Araliaceae); Atractylodis macrocephalae Rhizoma (Atractylodes macrocephala Koidz., Asteraceae); Poria [Poria cocos (Schw.) Wolf., Polyporaceae]; Glycyrrhizae Radix (Glycyrrhiza uralensis Fisch., Leguminosae) at the ratio of 2:2:2.5:2:2.5:5:5:2 (Liu et al., 2017a, Wang et al., 2016). RAHP was originally described in the Chinese Qing Dynasty classic book “TCM Prescriptions by Ancient and Modern Well Known Physicians”, which advances the function of promoting Qi circulation and strengthens the composition of the spleen and stomach (Xiao et al., 2012). RAHP is a phenomenally effective prescription for improving the clinical condition of damp-cold stagnation affecting the middle jiao (middle burner) from spleen and stomach deficiency (Li and Yu, 2008). RAHP significantly promotes gastrointestinal peristalsis and gastric emptying, improves electrogastrogram, reduces gastric sensitivity, and regulates gastrointestinal hormones (Kim et al., 2017). Some studies have suggested that RAHP appears to be suitable for the treatment of functional dyspepsia caused by multiple factors and as a potential multi-target therapeutic effect (Xiao et al., 2012). Several RCTs of RAHP on colitis have been conducted to confirm the anti-inflammatory effect and intestinal flora improvement (Heng et al., 2020). Costunolide and dehydrocostus are two of the major active components of Radix Aucklandiae in RAHP, and these ingredients have been confirmed to reduce 5-FU-induced intestinal mucositis by suppressing oxidative stress and regulating inflammatory homeostasis, enhancing proliferative activity in the epithelium and crypts (Chen et al., 2016).

However, up-to-date therapeutic strategies to manage chemotherapy-induced intestinal mucositis are not available. Our hypothesis is that the Radix aucklandiae herbal preparation may potentially ameliorate intestinal mucositis induced by 5-FU in mice. To investigate the mechanism of this hypothesis, the mice were randomly divided into control and herbal medicine treated groups. In addition, inflammatory cytokine analysis, measurement of NF-κB p65 and COX-2 production in middle jejunum and colon tissue, histologic examination of intestines and analysis of goblet cells were conducted to assess 5-FU-induced intestinal mucositis in mice.