ORIGINAL ARTICLE

Anti-inflammatory effects of ethanolic extract from Abeliophyllum distichum (Miseon Tree) leaves in mice with dextran sulfate sodium-induced ulcerative colitis

Hye-Jung Moon¹, Youn-Soo Cha^1,2 and Kyung-Ah Kim^3*

¹Department of Food Science and Human Nutrition, Jeonbuk National University, Jeonju, Republic of Korea; ²K-Food Research Center, Jeonju, Republic of Korea; ³Department of Food and Nutrition, Chungnam National University, Daejeon, Republic of Korea

Abstract

Abeliophyllum distichum (Miseon tree), a native Korean plant, is known for the high phenolic content in its leaves. The ethanolic leaf extracts of A. distichum have shown antioxidant, anti-obesity, and anti-inflammatory properties. However, studies on its potential to improve colitis are limited. This study aimed to determine whether the ethanolic extract of A. distichum leaves (ADE) could alleviate dextran sulfate sodium (DSS)-induced colitis in mice. The mice were divided into three groups, and the experimental group was given 300 mg/kg ADE for 4 weeks. One week before the end of the experiment, 3% DSS was added to the drinking water to induce colitis. The clinical symptoms of colitis and damage to colon tissue, including the increase in Enterobacteriaceae abundance and a decrease in Bifidobacterium in the colon, were evaluated during DSS treatment. DSS overactivated the nuclear factor (NF)-κB signaling pathway, resulting in excessive production of pro-inflammatory cytokines. In contrast, ADE alleviated the DSS-induced colitis symptoms, protected against colonic tissue damage, and restored the balance of Enterobacteriaceae and Bifidobacterium levels in the colon. Moreover, ADE effectively inhibited the DSS-induced overactivation of the NF-κB signaling pathway within the colon and mitigated abnormal inflammatory responses. These findings suggest that ADE protects against colitis by modulating the growth of some intestinal strains and the NF-κB pathway in the colon, supporting its potential as a natural agent.

Keywords: Abeliophyllum distichum; dextran sulfate sodium; colitis; NF-kappa B

 

 

A chronic inflammation of the mucosal layer extending from the rectum to the colon is known as ulcerative colitis (UC) (1). Clinical characteristics of UC include diarrhea, abdominal pain, bloody diarrhea, rectal urgency, and weight loss. UC affects various organs, including the skin, joints, biliary tract, and eyes, causing extraintestinal symptoms (2). The occurrence of UC began with the industrialization and westernization of society. In the 20th century, the prevalence of UC was high in Western nations, including North America, Europe, and Oceania. However, in the 21st century, it has spread to Asia, Africa, and South America, posing a threat to global health (3). However, the exact mechanism of UC pathogenesis remains unclear, and it can result from alterations in the balance of intestinal bacteria, damage to the mucosal barrier, and disruptions in immune regulation by environmental variables and genetic susceptibility (4, 5).

Current therapeutic agents for UC include aminosalicylates, corticosteroids, immunomodulators, biologics, and small-molecule drugs to suppress excessive inflammatory responses. However, these drugs are associated with adverse reactions and high treatment costs (6–8). In addition, UC carries a cumulative relapse risk of 70–80% after 10 years, and despite drug treatment, the remission rate is only 20–30%, making it challenging to treat (9–11). These factors have led to an ongoing effort to develop newer, more effective treatments for UC that are less prone to showing adverse reactions (6, 8). For example, based on Ingenuity Pathway Analysis conducted on patients with UC, interleukin (IL)-6, IL-1β, and tumor necrosis factor (TNF)-α, and nuclear factor (NF)-κB were identified as potential key regulators of inflammation (12). In particular, the targeting of the NF-κB signaling pathway is a key approach for treating UC, and numerous studies have evaluated natural products that target this pathway (6, 13, 14).

Abeliophyllum distichum (known in Korea as Miseon tree) is a deciduous shrub native to the Korean Peninsula and is protected by the Korean government (15). However, with recent advancements in the mass propagation of the A. distichum, various studies have been conducted on its fruits, flowers, stems, and leaves (16–19). The leaves of A. distichum contain various phenolic compounds, such as chlorogenic acid, caffeic acid, rutin, ferulic acid, and verbascoside (also known as acteoside) as well as polyphenolic compounds such as gallic acid, taxifolin, narirutin, and rutin (18, 20). The 70% ethanol extract of A. distichum leaves (ADE) demonstrated stronger inhibition of androgen receptor signaling than did the extracts from distilled water and 95% hexane, indicating its potential to prevent prostatic hyperplasia (21). ADE has also been shown to ameliorate obesity by lowering the expression of genes linked to adipogenesis in adipose tissue in vivo (22). Moreover, ADE have anti-inflammatory effects via suppressing lipopolysaccharide (LPS)-induced TNF-α production by inhibiting mitogen-activated protein kinase (MAPK)/NF-κB signaling in mouse macrophages (20). However, despite its widely researched anti-inflammatory properties, few studies have investigated the ability of A. distichum to ameliorate UC.

Therefore, in the present study, we investigated the potential of ADE to alleviate inflammation in mice with UC induced by dextran sulfate sodium (DSS). Specifically, we focused on the effects of ADE on the NF-κB signaling pathway, a critical regulator of inflammation, to evaluate its potential as a natural agent for alleviating colitis.

Materials and methods

Sample preparation

The ADE samples used in this experiment were provided by Korea Prime Pharmacy Co., Ltd. (Gwangju, Korea) and were prepared as described in a previous study (23). In summary, ADE samples were prepared by extraction at 75°C for 4 h with a 20-fold addition of 70% ethanol to dried ADE, followed by filtration, concentration, and lyophilization.

Animals

Male C57BL/6J mice, 5-week-old, were obtained from Central Lab Animal, Inc. (Seoul, Republic of Korea). Animals were acclimated for 6 days under consistent conditions, including a temperature of 22 ± 2°C, humidity of 50 ± 5%, and 12-h light/12-h dark cycles. After acclimation, 21 mice were divided into three experimental groups (n = 7 per group): Nor group with untreated normal control mice; DSS group with DSS-treated control mice; and ADE group containing mice treated with DSS and ADE. All animal experiments were approved by the Animal Ethics Committee of Chungnam National University (IACUC approval number: 202112-CNU-214).

Sample treatment and colitis induction in mice

Samples were administered by oral gavage to mice for 29 days. In the ADE group, ADE was orally administered at 300 mg/kg body weight daily. Phosphate-buffered saline (PBS) was supplied in the same amounts to the Nor and DSS groups. On day 21, UC was induced by replacing the drinking water of the DSS and ADE groups with a 3% DSS solution (w/v; MW: 36,000–50,000 Da, MP Biochemical, Solon, OH, USA) and allowing them to drink for 6 days.

After the day following the conclusion of the DSS drinking period, the fecal samples were collected. All mice were euthanized, and blood and colon tissue were obtained. Colon tissues were measured in length and weight, some were fixed in 4% formalin, and the remaining colon tissues were stored at −80°C.

Evaluation of the severity of colitis

The severity of UC was assessed using the disease activity index (DAI), a system that scores body weight loss, stool consistency, and gross bleeding over the DSS administration. The DAI score ranged from 0 to 9, with weight loss scores of 0 (< 1%) to 3 (> 10%), stool formation scores of 0 (none) to 3 (watery diarrhea), and blood in stool scores of 0 (none) to 3 (gross bleeding).

Histological studies

Hematoxylin and eosin (H&E) staining was used for histopathological analysis of colon tissue. Colon tissue sections embedded in paraffin after fixation in 4% formalin were stained with H&E and observed under a light microscope (DM2500; Leica Microsystems, Wetzlar, Germany) installed in the Center for University Research Facilities (CURF) at Jeonbuk National University (Jeonju, Korea).

Fecal microbial population

The fecal samples were diluted in sterile PBS and plated onto the respective selective media: Bifidobacterium, blood liver agar (BL agar, MBcell, Seoul, Republic of Korea), at 37°C for 48 h; Enterobacteriaceae, desoxycholate agar (MBcell), at 37°C for 24 h. Results were expressed as Colony Forming Units per gram of feces.

Measurement of pro-inflammatory cytokines

Enzyme-linked immunosorbent assay (ELISA) kits were employed to analyze pro-inflammatory cytokines in both colon tissue and serum. The manufacturer’s guidelines were followed in order to quantify the levels of TNF-α (Invitrogen, Vienna, Austria), IL-1β, and IL-6 (R&D Systems, Minneapolis, MN, USA).

Messenger RNA (mRNA) analysis by Quantitative real-time polymerase chain reaction (PCR) (qRT-PCR)

TRIZol reagent (Takara Bio, Inc.) was utilized to extract total RNA from the colon. The extracted total RNA was then synthesized into cDNA using PrimeScript RT Master Mix (Takara Bio, Inc.). The gene expression was quantified employing a 7500 Real-Time PCR System (Applied Biosystems, Foster City, CA, USA). The relative expression of the target gene was normalized to glyceraldehyde 3-phosphate dehydrogenase (GAPDH) expression, a housekeeping gene.

Western blotting

The colon tissue was homogenized and centrifuged to obtain a supernatant. The supernatant was used for western blotting analysis to confirm the expression of the target protein. Primary antibodies were used at a dilution of 1:1,000 for NF-κB phosphorylated-p65 (p-p65), NF-κB p65, induced nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), and β-actin (Cell Signaling Technology, Danvers, MA, USA). Secondary antibodies of horseradish peroxidase-labeled anti-rabbit or anti-mouse were used at a 1:3,000 dilution (Cell Signaling Technology). The protein bands on the prepared membranes were visualized using a ChemiDoc system (ATTO LuminoGraph II, ATO, Tokyo, Japan). Signals were quantified using ImageJ software (National Institute of Health, Bethesda, Maryland, USA).

Statistical analysis

All results were expressed as mean ± standard deviation (SD). Statistical analysis was performed utilizing SPSS 18.0 software (SPSS Inc., Chicago, IL, USA). Differences among groups were evaluated for significance using a one-way analysis of variance (ANOVA) followed by Duncan’s post hoc tests at a significance level of P < 0.05.

Results

Alleviatory effects of ADE on the clinical symptoms of colitis

The DSS and ADE groups exhibited significant reductions in body weight in comparison with that of the Nor group by the end of the experiment (Fig. 1A, B). Additionally, the DAIs in both the DSS and ADE groups were higher than those in the Nor group from 1 day after DSS administration until the end of the DSS administration period (Fig. 1C). However, after cessation of DSS administration, the ADE group demonstrated faster recovery from colitis symptoms than did the DSS group, as indicated by a reduction in the DAI for 2 days.

Fig. 1. Effect of the 70% ethanol extract of Abeliophyllum distichum leaves (ADE) on colitis symptoms. (A) Body weight changes during the entire experimental period. (B) Body weight changes during DSS administration. (C) DAI during DSS administration. Data are presented as mean ± standard deviation (n = 7). The letters next to the values indicate significant differences at the P < 0.05 level (a > b > c; P < 0.05). Nor, normal group; DSS, dextran sulfate sodium group; ADE, a group treated with ADE at 300 mg/kg body weight along with DSS administration.

Effects of ADE on the length-to-weight ratio of the colon and the histopathological changes in colitis

The effect of ADE on colon length was confirmed when colitis was induced (Fig. 2A–C). Colon length in the DSS group was significantly shortened to approximately 25% of that in the Nor group. While the colon length in the ADE group showed no significant difference from that in the DSS group, it showed a tendency to increase by approximately 13% (Fig. 2A, B). The colon weight-to-length ratio in the DSS group was 21.66 mg/cm, a significant increase of 51.15% from that of the Nor group (14.33 mg/cm). However, the corresponding value in the ADE group was 19.61 mg/cm, which was 9.46% lower than that in the DSS group (Fig. 2C).

Fig. 2. Effects of the 70% ethanol extract of Abeliophyllum distichum leaves (ADE) on DSS-induced colon tissue damage. (A) Representative images of colonic length for each group. (B) Statistical analysis of colonic length. (C) Statistical analysis of colon weight-to-length ratio. (D) Representative hematoxylin and eosin images. Data are presented as mean ± standard deviation (n = 7). Lowercase letters above the bars indicate significant differences at the P < 0.05 level (a > b > c). Nor, normal group; DSS, dextran sulfate sodium group; ADE, a group treated with ADE at 300 mg/kg body weight along with DSS administration.

The effect of ADE on the histopathology of the colon tissue after induction of colitis was also observed (Fig. 2D). The DSS group exhibited the histopathological features of colitis, including crypt loss, destruction of goblet cells, and infiltration of inflammatory cells into the mucosal and submucosal layers. Furthermore, thickening of the submucosa and muscle layer was observed, along with epithelial cell destruction. However, the ADE group demonstrated relief from DSS-induced colon tissue damage, including inflammatory cell infiltration into the mucosa and submucosa, as well as crypt loss.

Effects of ADE on the commensal bacterial population in colitis

Changes in the commensal bacteria in the colon have been reported to cause colitis (24, 25). Changes in Enterobacteriaceae and Bifidobacterium were confirmed in the feces when ADE was administered during colitis induction with DSS (Fig. 3A, B). In comparison with that of the Nor group, the DSS group showed a significant increment of approximately 24-fold in the abundance of Enterobacteriaceae and a 4-fold reduction in the abundance of Bifidobacterium. However, the ADE group showed a reduction in the abundance of Enterobacteriaceae and an increase in the abundance of Bifidobacterium to levels similar to those in the Nor group.

Fig. 3. Effects of the 70% ethanol extract of Abeliophyllum distichum leaves (ADE) on the populations of Enterobacteriaceae and Bifidobacterium in the feces of mice with DSS-induced colitis. (A) Enterobacteriaceae and (B) Bifidobacterium counts in feces. Data are presented as mean ± standard deviation (n = 6). Lowercase letters above the bars indicate significant differences at the P < 0.05 level (a > b > c). Nor, normal group; DSS, Dextran sulfate sodium group; ADE, a group treated with ADE at 300 mg/kg body weight along with DSS administration.

Effects of ADE on the pro-inflammatory cytokines in colitis

Alterations in pro-inflammatory cytokines were observed in the colon and serum of ADE-treated mice (Table 1). The levels of TNF-α and IL-6 in the colon tissue, as well as TNF-α, IL-6, and IL-1β in the serum, were significantly higher in the DSS group than in the Nor group. Conversely, the ADE group demonstrated a significant reduction in the levels of all pro-inflammatory cytokines that were elevated in the DSS group, thereby inhibiting the progression of inflammation.

Table 1. Effects of the 70% ethanol extract of Abeliophyllum distichum leaves (ADE) on pro-inflammatory cytokines in the colon and serum in DSS-induced colitis mice

Parameters		Nor	DSS	ADE
Serum (pg/mL)	TNF-α	1.82 ± 0.53c	7.89 ± 1.25a	5.54 ± 1.80b
	IL-1β	0.05 ± 0.03b	0.08 ± 0.04a	0.03 ± 0.01b
	IL-6	0.45 ± 0.05b	2.65 ± 1.20a	1.15 ± 0.33b
Colon (pg/μg protein)	TNF-α	28.76 ± 25.71b	64.88 ± 22.50a	32.68 ± 15.34b
	IL-1β	2.85 ± 0.71c	37.32 ± 14.19a	25.28 ± 7.52b
	IL-6	6.92 ± 0.57a	7.18 ± 1.06a	6.57 ± 1.37a
Data are presented as mean ± standard deviation (n = 7). The letters next to the values indicate significant differences at the P < 0.05 level (a > b > c; P < 0.05). ADE, a group treated with ADE at 300 mg/kg body weight along with DSS administration.

Effects of ADE on mRNA and protein expression related to the NF-κB signaling pathway in colitis

The NF-κB signaling pathway has been suggested to be a major signaling pathway related to colitis (26). The effects of ADE on mRNA and protein expression related to the NF-κB signaling pathway were examined (Fig. 4A). The DSS group showed significantly upregulated mRNA expression of the pro-inflammatory cytokines TNF-α, IL-1β, and IL-6 in comparison with the Nor group. Additionally, the assessment of inflammation-related factors showed significantly elevated mRNA expression of NF-κB, iNOS, COX-2, and monocyte chemoattractant protein (MCP)-1 in the DSS group. In contrast, the ADE group showed significantly lower inflammation-related mRNA expression levels than the DSS group.

Fig. 4. Effects of the 70% ethanol extract of Abeliophyllum distichum leaves (ADE) on the expression of genes and proteins related to the NF-κB signaling pathway in the colon of DSS-induced colitis mice. (A) Gene expression and (B) protein expression related to the NF-κB signaling pathway in colon tissue. Data are presented as mean ± standard deviation (n = 7). Lowercase letters above the bars indicate significant differences at the P < 0.05 level (a > b > c). Nor, normal group; DSS, dextran sulfate sodium group; ADE, a group treated with ADE at 300 mg/kg body weight along with DSS administration.

Moreover, in the DSS group, NF-κB p-p65 was excessively activated, and the expression levels of the downstream inflammation-related enzymes iNOS and COX-2 were also increased (Fig. 4B, C). Conversely, the ADE group showed inhibition of the activity of NF-κB p-p65 that was increased by DSS, thereby reducing the expression of iNOS and COX-2. Therefore, ADE alleviates excessive inflammatory reactions by inhibiting the overactive NF-κB signaling pathway during colitis.

Discussion

UC treatment aims to achieve immediate symptom relief and promote mucosal healing, ultimately enhancing the patient’s overall quality of life (27). We investigated whether ADE, a natural product endemic to the Korean Peninsula that has shown anti-inflammatory effects on macrophages (20), has the potential to prevent colitis in mice. Although ADE did not inhibit the body weight loss in DSS-induced colitis mice, it significantly reduced the DAI. ADE has been previously shown to prevent obesity (22), suggesting that it may not influence body weight recovery. However, the anti-inflammatory effects of ADE are thought to ameliorate symptoms such as rectal bleeding and diarrhea. Thus, the anti-inflammatory effects of ADE may have reduced damage to colon tissue, thereby ensuring recovery from symptoms such as rectal bleeding and diarrhea.

Additionally, UC is characterized by pathological features such as goblet cell depletion and neutrophil inflow, and morphological changes such as shortened colon length (28) and ADE improved these symptoms. Verbascoside and chlorogenic acid, major components of ADE, have also been shown to improve colon length, mucosal damage, and infiltration of inflammatory cells in mice with colitis (18, 23, 29, 30). Thus, the polyphenols in the ADE may serve as the potential components for preventing colitis.

Inflammatory bowel diseases (IBDs) such as UC are characterized by an increase in Enterobacteriaceae and a decrease in Bifidobacterium (31, 32). We found that ADE contributed to the restoration of this balance. This result is similar to those obtained with dietary resveratrol administration in DSS-induced colitis mice and could be attributed to the increased abundance of beneficial bacteria, such as Bifidobacterium and Lactobacillus that protect against tissue damage from potentially harmful bacteria, such as Enterobacteriaceae (33). However, further research is required to understand the impact of ADE on the overall gut microbiota and the improvement of colitis.

A healthy intestine maintains intestinal immune homeostasis through appropriate interactions between the resident microbial community and the host immune system. In contrast, UC occurs when the balance of the colonic mucosal immune system is disrupted due to overactivation of the transcription factor NF-κB, which is caused by various factors, such as bacterial LPSs, pro-inflammatory cytokines, and viruses (26, 34). NF-κB consists of five subunits, of which p65 can directly activate the transcription of the target gene (26, 34). Activated NF-κB promotes the gene transcription of pro-inflammatory cytokines (e.g. IL-1β, IL-6, and TNF-α), as well as inflammation-related factors (e.g. iNOS and COX-2) (14). In addition, excessive production of cytokines and chemokines (e.g. MCP-1) resulting from NF-κB activation mediates the infiltration of inflammatory cells such as macrophages and neutrophils into damaged colon tissues, exacerbating colitis (35–37).

In this study, ADE inhibited the activation of NF-κB p-p65, which was typically heightened in the DSS group. This inhibition affected the expression of downstream genes and proteins, consequently contributing to the alleviation of colitis by reducing the concentration of inflammatory cytokines in both the colon and serum. Curcumin has been considered as a promising phytonutrient for treating IBD, with no observed side effects in animal studies and clinical trials (38). Similarly, ADE has demonstrated potential as a natural agent to prevent colitis. Based on these findings, further studies and subsequent clinical trials to identify the optimal concentration and bioactive compounds responsible for the colitis-protective effect of ADE may contribute to its development as a natural therapeutic agent.

Conclusions

ADE reversed the altered gut microbiota composition observed in colitis by reducing the abundance of Enterobacteriaceae while increasing that of Bifidobacterium. Moreover, ADE exhibits anti-inflammatory properties by mitigating pro-inflammatory cytokines through the suppression of the highly activated NF-κB signaling pathway, which is elevated during UC. Consequently, ADE has the potential as natural materials for the management of colitis.

Acknowledgements

The authors thank Korea Prime Pharmacy Co., Ltd. for providing the powdered ethanolic extract of A. distichum.

Authors’ contributions

KAK conceptualized the study, obtained funding, and carried out project administration. HJM, YSC, and KAK designed the experimental methods. HJM and KAK performed the experiments and analyzed the results. HJM wrote the original draft, and HJM and KAK reviewed and revised the manuscript. YSC and KAK provided resources and supervision.

References

1.	Gajendran M, Loganathan P, Jimenez G, Catinella AP, Ng N, Umapathy C, et al. A comprehensive review and update on ulcerative colitis. Dis Mon 2019; 65(12): 100851. doi: 10.1016/j.disamonth.2019.02.004
2.	Yu YR, Rodriguez JR. Clinical presentation of Crohn’s, ulcerative colitis, and indeterminate colitis: symptoms, extraintestinal manifestations, and disease phenotypes. Semin in Pediat Surg 2017; 26(6): 349–55. doi: 10.1053/j.sempedsurg.2017.10.003
3.	Ng SC, Shi HY, Hamidi N, Underwood FE, Tang W, Benchimol EI, et al. Worldwide incidence and prevalence of inflammatory bowel disease in the 21st century: a systematic review of population-based studies. Lancet 2017; 390(10114): 2769–78. doi: 10.1016/s0140-6736(17)32448-0
4.	Lobionda S, Sittipo P, Kwon HY, Lee YK. The role of gut microbiota in intestinal inflammation with respect to diet and extrinsic stressors. Microorganisms 2019; 7(8): 271. doi: 10.3390/microorganisms7080271
5.	Sartor RB. Mechanisms of disease: pathogenesis of Crohn’s disease and ulcerative colitis. Nat Clin Pract Gastroenterol Hepatol 2006; 3(7): 390–407. doi: 10.1038/ncpgasthep0528
6.	Zhou Y, Wang D, Yan W. Treatment effects of natural products on inflammatory bowel disease in vivo and their mechanisms: based on animal experiments. Nutrients 2023; 15(4): 1031. doi: 10.3390/nu15041031
7.	Burisch J, Vardi H, Schwartz D, Friger M, Kiudelis G, Kupčinskas J, et al. Health-care costs of inflammatory bowel disease in a pan-European, community-based, inception cohort during 5 years of follow-up: a population-based study. Lancet Gastroenterol Hepatol 2020; 5(5): 454–64. doi: 10.1016/s2468-1253(20)30012-1
8.	Cai Z, Wang S, Li J. Treatment of inflammatory bowel disease: a comprehensive review. Front Med (Lausanne) 2021; 8: 765474. doi: 10.3389/fmed.2021.765474
9.	Alsoud D, Verstockt B, Fiocchi C, Vermeire S. Breaking the therapeutic ceiling in drug development in ulcerative colitis. Lancet Gastroenterol Hepatol 2021; 6(7): 589–95. doi: 10.1016/s2468-1253(21)00065-0
10.	Fumery M, Singh S, Dulai PS, Gower-Rousseau C, Peyrin-Biroulet L, Sandborn WJ. Natural history of adult ulcerative colitis in population-based cohorts: a systematic review. Clin Gastroenterol Hepatol 2018; 16(3): 343–56.e3. doi: 10.1016/j.cgh.2017.06.016
11.	Volk N, Siegel CA. Defining failure of medical therapy for inflammatory bowel disease. Inflamm Bowel Dis 2019; 25(1): 74–7. doi: 10.1093/ibd/izy238
12.	Bergemalm D, Andersson E, Hultdin J, Eriksson C, Rush ST, Kalla R, et al. Systemic inflammation in preclinical ulcerative colitis. Gastroenterology 2021; 161(5): 1526–39.e9. doi: 10.1053/j.gastro.2021.07.026
13.	Kim G, Jang M, Hwang I, Cho J, Kim S. Radish sprout alleviates DSS-induced colitis via regulation of NF-κB signaling pathway and modifying gut microbiota. Biomed Pharmacother 2021; 144: 112365. doi: 10.1016/j.biopha.2021.112365
14.	Terra X, Valls J, Vitrac X, Mérrillon JM, Arola L, Ardèvol A, et al. Grape-seed procyanidins act as antiinflammatory agents in endotoxin-stimulated RAW 264.7 macrophages by inhibiting NFκB signaling pathway. J Agric Food Chem 2007; 55(11): 4357–65. doi: 10.1021/jf0633185
15.	Kang U, Chang C-S, Kim YS. Genetic structure and conservation considerations of rare endemic Abeliophyllum distichum Nakai (Oleaceae) in Korea. J Plant Res 2000; 113(2): 127–38. doi: 10.1007/PL00013923
16.	Lee NN, Kim J-A, Kim Y-W, Choi YE, Moon HK. Effect of explant’s position and culture method on shoot proliferation and micro-cuttings for a rare and endangered species, Abeliophyllum distichum Nakai. J Plant Biotechnol 2015; 42(3): 228–34. doi: 10.5010/JPB.2015.42.3.228
17.	Park GH, Park JH, Eo HJ, Song HM, Woo SH, Kim MK, et al. The induction of activating transcription factor 3 (ATF3) contributes to anti-cancer activity of Abeliophyllum distichum Nakai in human colorectal cancer cells. BMC Complement Altern Med 2014; 14: 487. doi: 10.1186/1472-6882-14-487
18.	Yoo TK, Kim JS, Hyun TK. Polyphenolic composition and anti-melanoma activity of white forsythia (Abeliophyllum distichum Nakai) organ extracts. Plants (Basel) 2020; 9(6): 757. doi: 10.3390/plants9060757
19.	Lee HD, Kim JH, Pang QQ, Jung PM, Cho EJ, Lee S. Antioxidant activity and acteoside analysis of Abeliophyllum distichum. Antioxidants (Basel) 2020; 9(11): 1148. doi: 10.3390/antiox9111148
20.	Kim HW, Yu AR, Kang M, Sung NY, Lee BS, Park SY, et al. Verbascoside-rich Abeliophyllum distichum Nakai leaf extracts prevent LPS-induced preterm birth through inhibiting the expression of proinflammatory cytokines from macrophages and the cell death of trophoblasts induced by TNF-α. Molecules 2020; 25(19): 4579. doi: 10.3390/molecules25194579
21.	Choi YJ, Fan M, Tang Y, Moon S, Lee SH, Lee B, et al. Ameliorative effect of Abeliophyllum distichum Nakai on benign prostatic hyperplasia in vitro and in vivo. Nutr Res Pract 2022; 16(4): 419–34. doi: 10.4162/nrp.2022.16.4.419
22.	Eom J, Thomas SS, Sung NY, Kim DS, Cha YS, Kim KA. Abeliophyllum distichum ameliorates high-fat diet-induced obesity in C57BL/6J mice by upregulating the AMPK pathway. Nutrients 2020; 12(11): 3320. doi: 10.3390/nu12113320
23.	Thomas SS, Eom J, Sung NY, Kim DS, Cha YS, Kim KA. Inhibitory effect of ethanolic extract of Abeliophyllum distichum leaf on 3T3-L1 adipocyte differentiation. Nutr Res Pract 2021; 15(5): 555–67. doi: 10.4162/nrp.2021.15.5.555
24.	Wasilewska E, Zlotkowska D, Wroblewska B. Yogurt starter cultures of Streptococcus thermophilus and Lactobacillus bulgaricus ameliorate symptoms and modulate the immune response in a mouse model of dextran sulfate sodium-induced colitis. J Dairy Sci 2019; 102(1): 37–53. doi: 10.3168/jds.2018-14520
25.	Kataoka K, Ogasa S, Kuwahara T, Bando Y, Hagiwara M, Arimochi H, et al. Inhibitory effects of fermented brown rice on induction of acute colitis by dextran sulfate sodium in rats. Dig Dis Sci 2008; 53(6): 1601–8. doi: 10.1007/s10620-007-0063-3
26.	Wullaert A. Role of NF-κB activation in intestinal immune homeostasis. Int J Med Microbiol 2010; 300(1): 49–56. doi: 10.1016/j.ijmm.2009.08.007
27.	Segal JP, LeBlanc JF, Hart AL. Ulcerative colitis: an update. Clin Med (Lond) 2021; 21(2): 135–9. doi: 10.7861/clinmed.2021-0080
28.	Eichele DD, Kharbanda KK. Dextran sodium sulfate colitis murine model: an indispensable tool for advancing our understanding of inflammatory bowel diseases pathogenesis. World J Gastroenterol 2017; 23(33): 6016–29. doi: 10.3748/wjg.v23.i33.6016
29.	Liu Y, Huang W, Zhu Y, Zhao T, Xiao F, Wang Y, Lu B. Acteoside, the main bioactive compound in Osmanthus fragrans flowers, palliates experimental colitis in mice by regulating the gut microbiota. J Agric Food Chem 2022; 70(4): 1148–62. doi: 10.1021/acs.jafc.1c07583
30.	Vukelić I, Detel D, Pučar LB, Potočnjak I, Buljević S, Domitrović R. Chlorogenic acid ameliorates experimental colitis in mice by suppressing signaling pathways involved in inflammatory response and apoptosis. Food Chem Toxicol 2018; 121: 140–50. doi: 10.1016/j.fct.2018.08.061
31.	Duranti S, Gaiani F, Mancabelli L, Milani C, Grandi A, Bolchi A, et al. Elucidating the gut microbiome of ulcerative colitis: bifidobacteria as novel microbial biomarkers. FEMS Microbiol Ecol 2016; 92(12): fiw191. doi: 10.1093/femsec/fiw191
32.	Khorsand B, Asadzadeh Aghdaei H, Nazemalhosseini-Mojarad E, Nadalian B, Nadalian B, Houri H. Overrepresentation of Enterobacteriaceae and Escherichia coli is the major gut microbiome signature in Crohn’s disease and ulcerative colitis; a comprehensive metagenomic analysis of IBDMDB datasets. Front Cell Infect Microbiol 2022; 12: 1015890. doi: 10.3389/fcimb.2022.1015890
33.	Larrosa M, Yañéz-Gascón MJ, Selma MV, González-Sarrías A, Toti S, Cerón JJ, et al. Effect of a low dose of dietary resveratrol on colon microbiota, inflammation and tissue damage in a DSS-induced colitis rat model. J Agric Food Chem 2009; 57(6): 2211–20. doi: 10.1021/jf803638d
34.	Atreya I, Atreya R, Neurath MF. NF-kappaB in inflammatory bowel disease. J Intern Med 2008; 263(6): 591–6. doi: 10.1111/j.1365-2796.2008.01953.x
35.	Khan WI, Motomura Y, Wang H, El-Sharkawy RT, Verdu EF, Verma-Gandhu M, et al. Critical role of MCP-1 in the pathogenesis of experimental colitis in the context of immune and enterochromaffin cells. Am J Physiol Gastrointest Liver Physiol 2006; 291(5): G803–11. doi: 10.1152/ajpgi.00069.2006
36.	Bibi S, de Sousa Moraes LF, Lebow N, Zhu MJ. Dietary green pea protects against DSS-induced colitis in mice challenged with high-fat diet. Nutrients 2017; 9(5): 509. doi: 10.3390/nu9050509
37.	Yao D, Dong M, Dai C, Wu S. Inflammation and inflammatory cytokine contribute to the initiation and development of ulcerative colitis and its associated cancer. Inflamm Bowel Dis 2019; 25(10): 1595–602. doi: 10.1093/ibd/izz149
38.	Fallahi F, Borran S, Ashrafizadeh M, Zarrabi A, Pourhanifeh MH, Khaksary Mahabady M, et al. Curcumin and inflammatory bowel diseases: from in vitro studies to clinical trials. Mol Immunol 2021; 130: 20–30. doi: 10.1016/j.molimm.2020.11.016