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. 2024 Sep 2;34(3):743–755. doi: 10.1007/s10068-024-01696-9

Long-term Pu-erh tea alleviates inflammatory bowel disease via the regulation of intestinal microbiota and maintaining the intestinal mucosal barrier

Bi-Yun Feng 1,5, Pei-Lu Chen 1,5, Ling Yan 1,5, Wei-Feng Huang 3,✉, Cheng-Fu Li 4, Li-Tao Yi 2,6,7,✉, Guang-Hui Xu 1,5,8,✉
PMCID: PMC11822139  PMID: 39958166

Abstract

Inflammatory bowel disease (IBD) is a chronic gastrointestinal condition with increasing global prevalence. Current therapies are limited, leading to exploration of novel treatments like Pu-erh tea, a fermented tea recognized for its health benefits. This study shows that long-term consumption of Pu-erh tea significantly reduces IBD symptoms in DSS-induced mice by moderating inflammation and enhancing oxidative responses in the colon. Pu-erh tea notably increases the abundance of specific gut microbiota, particularly enhancing Firmicutes, Bacteroidota, and Proteobacteria phyla, and raising levels of Lactobacillus and Muribaculaceae genera. Key species such as Lactobacillus johnsonii, Lactobacillus reuteri, and Lactobacillus murinus also showed increased abundance. Additionally, Pu-erh tea helps restore the integrity of the intestinal barrier. These findings highlight the potential of Pu-erh tea as a complementary dietary strategy for IBD, potentially improving disease management and patient outcomes through its effects on the intestinal microbiota and mucosal barrier.

Supplementary Information

The online version contains supplementary material available at 10.1007/s10068-024-01696-9.

Keywords: Pu-erh tea, Inflammatory bowel disease (IBD), Microbiota, Intestinal mucosal barrier, Inflammation

Introduction

Inflammatory bowel disease (IBD), including ulcerative colitis and Crohn's disease, is a chronic inflammatory disease of the intestinal tract. IBD affects millions of individuals worldwide, with a rising incidence in both developed and developing countries. The etiology of IBD remains complex and multifactorial, involving interactions between genetic, environmental, and immunological factors (Kofla-Dlubacz et al., 2022). The current therapeutic approaches for IBD primarily focus on symptom management and immunosuppression, aiming to reduce inflammation and maintain remission (Kotla and Rochev, 2023). However, these treatments often have limited efficacy, potential side effects, and fail to address the underlying causes of the disease. For example, long-term use of immunosuppressive agents and biologics can lead to serious side effects such as increased susceptibility to infections, liver toxicity, and an elevated risk of certain cancers. Hence, there is a growing need for novel therapeutic strategies that target the fundamental mechanisms driving IBD pathogenesis while minimizing adverse effects.

Recent researches have revealed a critical role of the intestinal microbiota in the development and progression of IBD (Santana et al., 2022). The gut microbiota consists of a vast and diverse community of microorganisms residing in the gastrointestinal tract, which play a vital role in maintaining gut homeostasis and regulating immune responses. Alterations in the composition and function of the intestinal microbiota, termed dysbiosis, have been observed in individuals with IBD (Andoh and Nishida, 2023; Pu et al., 2022), indicating a potential causative role in disease pathogenesis.

Dietary interventions and nutritional strategies have emerged as promising avenues for IBD management. Among these, Pu-erh tea is a Chinese fermented tea with a long history of consumption, has gained attention due to its potential health benefits. Pu-erh tea is derived from the leaves of the Camellia sinensis plant. It undergoes a unique microbial fermentation process, resulting in distinct biochemical profiles (Wang et al., 2022). Pu-erh tea has been associated with various health effects, including antioxidant and anti-inflammatory properties (Liu et al., 2021). Its consumption has been linked to improvements in mental disorders, metabolic disorders, cardiovascular health, and gastrointestinal function. For example, Pu-erh tea supplement altered the gut microbiota and modulated short-chain fatty contents in depression (Zhao et al., 2023). Pu-erh tea decreased body weight gain, reversed adipose inflammation, reduced metabolic endotoxemia, and changed the relative abundance of Firmicutes and Bacteroidetes in obese mice (Lu et al., 2019). Moreover, recent studies showed that Pu-erh tea alleviated acute dextran sulfate sodium (DSS)-treated mice (Huang et al., 2020, 2021). For example, Pu-erh tea has been demonstrated to inhibit intestinal oxidative stress-mediated inflammation pathways, upregulate intestinal tight junction proteins, promote M2 macrophage polarization, and improve the intestinal immune barrier by reshaping microbiota. These findings suggest that Pu-erh tea has practical effects on alleviating and preventing intestinal inflammation when administered for a short duration (one week). However, the chronic nature of IBD necessitates long-term therapeutic strategies. Our study aims to investigate the sustained benefits of long-term Pu-erh tea consumption, particularly focusing on its prolonged anti-inflammatory, antioxidative effects, and its role in maintaining gut microbiota and mucosal barrier integrity over an extended period. Therefore, to investigate the effects of long-term daily Pu-erh tea supplement on the intestinal microbiota, the microbial diversity, microbial difference, and functional prediction in the fecal of IBD patients were investigated. Besides, to investigate the impact of Pu-erh tea on the integrity of the intestinal mucosal barrier, tight junction proteins were also detected.

Materials and methods

Animals

C57BL/6 J mice (male, 8 weeks old, 23–25 g body weight) were obtained from Slac Laboratory Animal Center of Shanghai (Shanghai, China). Mice were acclimatized in the laboratory for 1 week at a relative humidity maintaining 50–75% and a temperature maintaining 22–25 °C. Mice were given a standard diet and free access to water. All procedures were approved by the ethnic committees of Huaqiao University (A2022043) and performed in accordance with the National guidelines of the China Council on Animal Care.

Extraction of Pu-erh tea

Ripened Pu-erh tea was immersed to water (w/v = 1:4), boiled for one hour and then filtered through filter paper and 0.22 um filter membrane respectively, and the filtrate was concentrated and lyophilized to obtain Pu-erh tea extract for further investigation. The contents of tea polyphenols in Pu-erh tea extract were determined by the Folin-phenol method, and the contents of tea polysaccharides were determined by the phenol–sulfuric acid method. The content of tea polyphenols in Pu-erh tea extract is 11.36%. The content of tea polysaccharides in Pu-erh tea extract is 5.65%.

Reagents

Dextran sodium sulfate (DSS, D808272) MW 40000 was purchased from Macklin (Shanghai, China). Sulfasalazine (BP22550), IL-1β ELISA kit (BP80277), IL-6 ELISA kit (BP80293), TNF-α ELISA kit (BP80317) and IL-10 ELISA kit (BP80278), and ZO-1 antibody (BP65421) were purchased from Purdue Bioscience (New York, USA). Claudin-1 (ab242370) and occludin (ab216327) antibodies were purchased from Abcam (Cambridge, USA). Superoxide dismutase (SOD, BC0175) and malondialdehyde (MDA, BC0025) kits were acquired from Solarbio (Beijing, China).

Drug administration

The mice were randomly grouped into six groups including normal vehicle group, DSS vehicle group, positive control group (sulfasalazine at 50 mg/kg), Pu-erh tea (150 mg/kg), Pu-erh tea (300 mg/kg) and Pu-erh tea (600 mg/kg). The mice in the DSS vehicle and the treatment groups were given drinking water for 5 days and then 2.5% DSS for 5 days for four cycles, while the normal vehicle group was given drinking water throughout. Pu-erh tea was dissolved in saline, and the positive drug sulfasalazine was dissolved in saline and orally administrated every day. The normal vehicle group and the DSS vehicle group were orally administrated with the same dose of saline every day for consistency.

Disease activity index (DAI)

DAI scores were recorded during the modeling period, including the degree of weight loss, fecal characteristics and blood in the stool. The scores are shown in Table S1.

Histopathological evaluation

The collected colonic tissues were fixed in 4% paraformaldehyde, dehydrated, embedded in paraffin, sectioned at 5 μm thickness and stained with hematoxylin and eosin (H&E), and observed for histopathological changes under a light microscope. According to the results of HE staining of colonic tissues, the colon was scored histologically with the following criteria: (1) Intestinal epithelial cells: normal morphology, 0; loss of cupped cells, 1; extensive loss of cupped cells, 2; loss of crypt, 3; extensive loss of crypt, 4. (2) Inflammatory infiltration: no inflammatory infiltration, 0; infiltration of mucosal layer, 1; infiltration of the mucosal muscle layer, 2 points; extensive infiltration of the mucosal muscle layer, accompanied by swelling of the mucosal muscle layer, 3; infiltration of the submucosal layer, 4.

Inflammatory cytokines and oxidative stress assay

The colon tissue was homogenized in PBS. After centrifugation of the mixture at 12,000 g for 15 min at 4 °C, the supernatant was collected and used for inflammatory cytokine assays. Levels of IL-1β, IL-6, TNF-α and IL-10 were measured according to the instructions of the ELISA kits. The oxidative stress levels (SOD and MDA) were determined according to the instructions of the assay kits.

Gut microbiota analysis

16S rRNA sequencing of fecal bacteria was performed using the Illumina HiSeq platform. The hypervariable region V3–V4 of the bacterial 16S rRNA gene were amplified with primer pairs 338F (5′-ACTCCTACGGGAGGCAGCAG-3′) and 806R (5′-GGACTACHVGGGTWTCTAAT-3′). Taxonomic assignment was performed using the Naive bayes consensus taxonomy classifier implemented in Qiime2 and the SILVA 16S rRNA database (v138). Non-repeat sequences were clustered according to the optional taxonomic unit (OTU) 97% similarity analysis. Based on the OTU data, alpha diversity index and beta diversity were calculated for each sample bacterial community. A weighted UniFrac distance algorithm combined with principal coordinate analysis (PCoA) and hierarchical clustering analysis was used to compare the differences in the structure of each sample's bacterial community. Based on the sequencing results, PICRUSt software was used to predict the function of the intestinal flora.

Immunofluorescence

Colon tissue was taken into 4% paraformaldehyde and subsequently transferred to 30% sucrose solution for dehydration, after which it was used for frozen sections. For staining, tissue slices were washed in TBST solution for 5 min × 2, followed by antigen repair, and subsequently closed for 1 h at room temperature by adding a closure solution. After that, the primary antibody (dilution at 1:100) was incubated overnight at 4 °C and the secondary antibody was incubated at room temperature for two hours on the next day. The tissue slides were attached to the slides after adding DAPI dropwise in the dark room and dried at room temperature. The expression of colonic intestinal mucosal barrier protein was observed by a confocal microscope (Leica SP8) using equivalent acquisition settings to maintain data uniformity, across the relevant excitation and emission spectra.

Statistical analyses

Experimental data were expressed as mean ± SEM. Differences between groups were determined by one-way ANOVA followed by Tukey's multiple comparison. Graphs were produced using GraphPad Prism software. P < 0.05 was used as the statistical difference.

Results and discussion

Long term Pu-erh tea consumption alleviated chronic DSS-induced IBD symptoms in mice

Debilitating symptoms and impaired quality of life are typically observed in IBD patients (Roda et al., 2020). The etiology of IBD remains complex and multifactorial, involving dysregulated immune responses, genetic predisposition, and environmental factors. Traditional treatment strategies for IBD often involve immunosuppressive agents with various side effects (McLean and Cross, 2014). Hence, there is a pressing need to explore natural compounds that possess anti-inflammatory properties and can mitigate IBD-like symptoms. In detail, a significant increase in DAI scores (indexed by weight change, stool properties and occult blood) was observed in DSS mice starting from day 8 (third day after DSS administration) compared to normal controls (Fig. 1). High doses of Pu-erh tea and positive drug sulfasalazine significantly reduced the severity of DAI scores from day 17, and low dose of Pu-erh tea from day 26 and middle dose of Pu-erh tea from day 25 intermittently reduced DAI scores (Table S2). Meanwhile, DSS mice showed significantly shorter colon compared with normal mice, and the change was significantly reversed after the consumption of middle and high doses of Pu-erh tea.

Fig. 1.

Fig. 1

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Daily Pu-erh tea consumption attenuates DSS-induced IBD symptoms and histopathological alterations in mice. (A )Representative Anus images. (B) Dynamic DAI in four cycles. (C) Representative colon images. (D) Colon length. (E) Representative images of H&E-stained colon tissue. (F) Histopathological scores of colons. Scale bar (black) was shown as 25 μm. N = 5–8 per group. *p < 0.05, **p < 0.01

Compared with the normal group, DSS induced severe colonic pathological damage, including intestinal epithelial cell damage and inflammatory cell infiltration (Fig. 1E, F), and the histological score of the colon was significantly higher. Consumption of low, middle and high doses of Pu-erh tea, as well as positive drug sulfasalazine reduced intestinal epithelial cell injury and inflammatory cell infiltration, and the histological scores of the colon were all decreased.

Traditional treatment strategies for IBD often involve the use of immunosuppressive agents such as corticosteroids, thiopurines, and biologics. While these medications can be effective in reducing inflammation and inducing remission, they are associated with a range of potential side effects. For instance, long-term corticosteroid use can lead to complications such as osteoporosis, diabetes, and hypertension. Thiopurines, like azathioprine and mercaptopurine, carry risks of liver toxicity and myelosuppression, while biologics, such as infliximab and adalimumab, can increase the risk of serious infections and malignancies. These side effects can significantly impact the quality of life of IBD patients and highlight the need for safer therapeutic options. Pu-erh tea, with its natural anti-inflammatory and antioxidative properties, presents a promising alternative with a potentially better safety profile. Our study demonstrates that long-term consumption of Pu-erh tea significantly reduces IBD symptoms in DSS-induced mice. Unlike conventional treatments, Pu-erh tea does not carry the same risks of severe side effects, making it a safer option for chronic use.

Long term Pu-erh tea consumption alleviated chronic IBD related inflammatory response and antioxidant capacity

Our study investigates the therapeutic potential of Pu-erh tea for IBD by examining its effects on inflammation, oxidative stress, gut microbiota, and the intestinal mucosal barrier. These four aspects are closely linked in the pathogenesis and progression of IBD.

Firstly, chronic inflammation is a hallmark of IBD, leading to the release of pro-inflammatory cytokines that perpetuate tissue damage. This inflammatory response is often accompanied by increased oxidative stress (Gambini and Stromsnes, 2022). In the present experiment, serum and colonic tissue levels of pro-inflammatory cytokines were examined (Fig. 2A–F). The study showed that the levels of pro-inflammatory cytokines were significantly increased in the DSS group compared to the normal group. On the contrary, the levels of serum and colonic IL-1β, IL-6 and TNF-α were significantly reduced by the consumption of Pu-erh tea. These findings suggest that observed improvement in diarrhea symptoms may be attributed to the anti-inflammatory properties of Pu-erh tea as Pu-erh inhibited pro-inflammatory cytokines in both serum and colon.

Fig. 2.

Fig. 2

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Daily Pu-erh tea consumption decreases inflammatory cytokine and oxidative levels in DSS-induced IBD mice. (A) Serum IL-1β. (B) Serum IL-6. (C) Serum TNF-α. (D) Colonic IL-1β. (E) Colonic IL-6. (F) Colonic TNF-α. G Colonic SOD. (H) Colonic MDA. N = 8 per group. *p < 0.05, **p < 0.01

Severe oxidative stress is one of the mechanisms involved in the pathogenesis of IBD. In the present study, the activity of SOD in DSS-induced IBD colonic tissue was significantly decreased, while the MDA level was significantly increased as compared with the normal group (Fig. 2G, H). Pu-erh tea could tend to increase SOD activity (a tendency trend but not significance) and significantly reduce MDA levels. These findings imply that Pu-erh tea can improve the antioxidant level in the colon of DSS-induced IBD mice.

Long term Pu-erh tea consumption reshaped the composition of gut microbiota in DSS-induced mice

The gut microbiota plays an important role in maintaining intestinal homeostasis and modulating the immune response. Dysbiosis, or an imbalance in gut microbiota, has been implicated in the initiation and perpetuation of IBD (Zhang et al., 2021). For example, an increase in pathogenic bacteria and a decrease in beneficial bacteria can disrupt gut homeostasis and promote inflammation. Changes in the gut microbiota can influence both inflammation and oxidative stress. Beneficial bacteria, such as Lactobacillus, produce short-chain fatty acids (SCFAs) that have anti-inflammatory properties and can enhance the intestinal barrier function (Markowiak-Kopec and Slizewska, 2020). Conversely, pathogenic bacteria can produce metabolites that exacerbate oxidative stress and inflammation. Therefore, the alteration of gut microbiota induced by DSS and Pu-erh tea was evaluated. As shown in Fig. 3A, the Venn analysis indicated there was no difference among the samples at Phylum level. At Genus level, there were 9 exclusive Genus in DSS group, however, there was no exclusive Genus in Normal group (Fig. 3B). At Species level, there were 15 exclusive Species and 1 exclusive Specie in DSS and Normal group, respectively (Fig. 3C). According to principal coordinate analysis (PCoA) analysis, the individual samples were clearly separable between normal and DSS groups at Phylum, Genus or Species level (Fig. 3D–F), indicating the distinct microbiome clusters between normal and DSS groups. According to community barplot analysis and Circos analysis, Firmicutes, Bacteroidota and Proteobacteria were the three Phylum with the highest abundance among the all six groups. Lactobacillus, Muribaculaceae and Bacteroides were the three Genus with the highest abundance among the all six groups. Lactobacillus johnsonii, Lactobacillus intestinalis and Lactobacillus reuteri were the three Species with the highest abundance among the all six groups (Fig. 3G–L).

Fig. 3.

Fig. 3

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Daily Pu-erh tea consumption reshapes gut microbiota compositions. (A) Venn analysis at Phylum level. (B) Venn analysis at Genus level. (C) Venn analysis at Species level. (D) PCoA analysis at Phylum level. (E) PCoA analysis at Genus level. (F) PCoA analysis at Species level. (G) Community barplot analysis at Phylum level. (H) Community barplot analysis at Genus level. (I) Community barplot analysis at Species level. (J) Analysis of dominant Phylum analysis. (K) Analysis of dominant Genus analysis. (L) Analysis of dominant Species analysis. N = 6 per group

Further analysis showed that differences among the groups at Phylum, Genus and Species levels (Fig. 4). At the phylum level, DSS significantly decreased the relative abundance of Firmicutes, Campilobacterota and Actinobacteriota, but increased the relative abundance of Proteobacteria (Fig. 4A). The relative abundance of Firmicutes was restored after daily Pu-erh tea consumption. At the Genus level, DSS significantly decreased the relative abundance of Lactobacillus, but increased the relative abundance of Muribaculum (Fig. 4E). The relative abundance of Lactobacillus was also restored after daily Pu-erh tea consumption (Fig. 4F). At the Species level, DSS significantly decreased the relative abundance of Lactobacillus johnsonii, Lactobacillus murinus, Lactobacillus reuteri and Lactobacillus intestinalis (Fig. 4I). Daily Pu-erh tea consumption could significantly increase the relative abundance of Lactobacillus johnsonii (Fig. 4J).

Fig. 4.

Fig. 4

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Daily Pu-erh tea consumption attenuates the alterations of gut microbiota caused by DSS. (A) Significant changes between Normal and DSS groups at Phylum level. (B) The effects of Pu-erh tea on Firmicutes. (C) The effects of Pu-erh tea on Bacteroidota. (D) The effects of Pu-erh tea on Proteobacteria. (E) Significant changes between Normal and DSS groups at Genus level. (F) The effects of Pu-erh tea on Lactobacillus. (G) The effects of Pu-erh tea on norank_f__Muribaculaceae. (H) The effects of Pu-erh tea on Bacteroides. (I) Significant changes between Normal and DSS groups at Species level. (J) The effects of Pu-erh tea on Lactobacillus johnsonii. (K) The effects of Pu-erh tea on Lactobacillus reuteri. (L) The effects of Pu-erh tea on Lactobacillus murinus. N = 6 per group. *p < 0.05, **p < 0.01

Association analysis was performed between gut microbiota and serum inflammatory factors, colonic inflammatory factors and colonic oxidative stress marker proteins, and correlations with TOP 30 association strength were selected (Fig. 5). According to the matrix heat map results, at the level of the phylum, Proteobacteria had significant correlations with serum inflammatory cytokines, colonic inflammatory cytokines and colonic oxidative stress marker proteins. Firmicutes had negative correlations with serum inflammatory cytokines, colonic inflammatory cytokines and colonic MDA levels. At the Genus level, Lactobacillus had a negative correlation with serum inflammatory cytokines and colonic inflammatory cytokines, but no correlation with colonic oxidative stress marker proteins. At the Species level, several different Lactobacillus species such as Lactobacillus johnsonii, Lactobacillus murinus, Lactobacillus reuteri and Lactobacillus intestinalis were associated with serum inflammatory cytokines, colonic inflammatory cytokines and colonic oxidative stress marker proteins were differentially correlated.

Fig. 5.

Fig. 5

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Spearman correlation heatmap after daily Pu-erh tea consumption. (A) Correlation between gut microbiota and serum cytokines at Phylum level. (B) Correlation between gut microbiota and serum cytokines at Genus level. (C) Correlation between gut microbiota and serum cytokines at Species level. (D) Correlation between gut microbiota and colon cytokines at Phylum level. (E) Correlation between gut microbiota and colon cytokines at Genus level. (F) Correlation between gut microbiota and colon cytokines at Species level. (G)Correlation between gut microbiota and colon SOD/MDA at Phylum level. (H) Correlation between gut microbiota and colon SOD/MDA at Genus level. I Correlation between gut microbiota and colon SOD/MDA at Species level. N = 6 per group. *p < 0.05, **p < 0.01

Finally, we constructed analytical models by machine learning and other methods to classify and predict data information, evaluated the applicability and accuracy of the model classification, and gave information related to the importance of each part of data information based on the model. The performance of our machine learning models was evaluated using receiver operating characteristic (ROC) curve analysis and the area under the curve (AUC) metric (Fig. S1). The ROC curve is a graphical representation that illustrates the diagnostic ability of a binary classifier system as its discrimination threshold is varied. Specifically, the ROC curve plots the true positive rate (sensitivity) against the false positive rate (specificity) at various threshold settings. Sensitivity measures the proportion of actual positives that are correctly identified by the model, whereas specificity measures the proportion of actual negatives that are correctly identified. Thus, ROC curve provides a visual representation of the discriminatory ability of the model. The AUC represents the probability that the model will correctly classify a randomly chosen positive instance over a randomly chosen negative instance. In our study, the ROC curve and AUC analysis were used to assess the accuracy of the models in classifying samples and predicting IBD severity. An AUC close to 1 indicates high model accuracy, whereas an AUC close to 0.5 suggests a model with no discriminative power. Our models achieved an AUC close to 0.9, indicating a high level of accuracy in predicting disease progression and treatment response based on the phylum level of microbiota. In addition, the machine learning models provided key insights into the relationships between gut microbiota composition, inflammatory markers, and IBD severity. For example, the abundance of Proteobacteria was strongly correlated with increased serum and colon cytokine levels, suggesting its potential as a microbiological indicator for IBD progression. Similarly, the increased abundance of Lactobacillus species in Pu-erh tea-treated groups was associated with reduced inflammation and improved gut barrier function. By leveraging machine learning, we identified specific microbial and inflammatory signatures that differentiate between treatment groups and predict IBD severity.

The data indicated that Pu-er tea mainly restored the abundance of Firmicutes, decreased the abundance of Bacteroidota and Proteobacteria at Phylum level. The increase in Firmicutes abundance following Pu-erh tea consumption is of particular interest, as alterations in Firmicutes have been consistently reported in IBD patients. Firmicutes are known to play a crucial role in maintaining gut health by producing short-chain fatty acids and promoting epithelial barrier integrity. The restoration of Firmicutes abundance by Pu-erh tea may contribute to the attenuation of IBD symptoms and improvement of gut barrier function. The decrease in Bacteroidota abundance observed in our study is consistent with previous reports in IBD patients. Bacteroides species have been associated with pro-inflammatory responses and increased mucosal permeability (Ryan et al., 2020). The reduction in Bacteroidota abundance following Pu-erh tea consumption may cause the mitigation of inflammation and restoration of gut barrier integrity. Of particular interest is the decrease in Proteobacteria abundance after Pu-erh tea supplementation. Proteobacteria, including Enterobacteriaceae, have been linked to intestinal inflammation and increased intestinal permeability (Shin et al., 2015). The correlation between the abundance of Proteobacteria at the phylum level and various serum and colon parameters suggests that Proteobacteria may serve as a potential microbiological indicator for IBD. Monitoring changes in Proteobacteria abundance could provide valuable insights into disease activity and treatment response in IBD patients. Pu-er tea restored the abundance of Lactobacillus and Muribaculaceae at Genus level. Pu-er tea increased the abundance of Lactobacillus johnsonii, Lactobacillus reuteri and Lactobacillus murinus at Species level. Lactobacillus, a genus known for its probiotic properties, was significantly increased in abundance. Lactobacillus species, including L. johnsonii, L. reuteri, and L. murinus, have been associated with anti-inflammatory effects and modulation of the immune response. The negative correlations observed between the abundance of Lactobacillus and serum and colon cytokines suggest that Lactobacillus species may serve as microbiological indicators for inflammation following Pu-erh tea consumption. Monitoring the abundance of Lactobacillus could provide valuable information on the anti-inflammatory effects of Pu-erh tea in IBD.

Long term Pu-erh tea consumption repaired intestinal mucosal barrier in the colon of mice

The intestinal mucosal barrier plays a crucial role in modulating gut homeostasis and protecting the underlying tissues from harmful substances (Lie et al., 2018; Schwartz et al., 2022). The intestinal mucosal barrier is composed of single-layer epithelial cells, which are connected by tight junction proteins. They create a physical barrier preventing the entry of harmful substances such as bacteria, toxins, and undigested food particles into the underlying tissues (Chelakkot et al., 2018; Song et al., 2023). In addition to its physical barrier function, the intestinal mucosa also produces mucus, antimicrobial peptides, and immunoglobulins that contribute to the defense against pathogens and maintain immune homeostasis in the gut (Herath et al., 2020; Tokuhara et al., 2019). In IBD, the mucosal barrier is compromised, leading to increased permeability and allowing the translocation of luminal contents across the intestinal epithelium (Wells et al., 2017). This translocation triggers an exaggerated immune response, resulting in chronic inflammation and tissue damage. Multiple factors contribute to the disruption of the mucosal barrier in IBD, including genetic predisposition, dysregulated immune responses, altered gut microbiota composition, and environmental factors (Chelakkot et al., 2018; Yu, 2018). The restoration of the intestinal mucosal barrier function is an important therapeutic goal in IBD management (Villablanca et al., 2022). Inflammation and oxidative stress can directly damage the intestinal barrier by disrupting tight junction proteins (e.g., ZO-1, claudin-1, occludin). This barrier dysfunction allows for the infiltration of luminal antigens, which further stimulate the immune response and perpetuate inflammation. In the context of the study, the results showed that all the tight junction proteins including ZO-1, claudin-1 and occludin were significantly reduced in response to DSS treatment (Fig. 6). In contrast, Pu-erh tea was found to increase the intensity of ZO-1, claudin-1 and occluding in the colon, suggesting that Pu-erh tea may enhance the barrier function and restore the selective permeability of the intestinal epithelium. Studies have demonstrated that nutrients can enhance the expression and assembly of tight junction proteins, such as claudins and occludin, thereby strengthening the integrity of the barrier and reducing permeability (Suzuki, 2020; Zhang et al., 2023). By preserving the tight junction structure and function, Pu-erh tea may improve mucosal barrier in IBD. The findings from our study are consistent with previous research demonstrating the therapeutic potential of tea and its polyphenolic components in alleviating IBD symptoms (Chen et al., 2022; El-Desoky Mohamady et al., 2022; Zhang et al., 2022). The mechanistic basis for these effects involves the modulation of pro-inflammatory signaling pathways and the promotion of tissue repair mechanisms.

Fig. 6.

Fig. 6

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Daily Pu-erh tea consumption enhances the function of the intestinal barrier impairment in mice exposed to DSS. (A) Representative ZO-1 fluorescence images. (B) Quantitative analysis of ZO-1 fluorescence intensity. (C) Representative claudin-1 fluorescence pictures. (D) Quantitative analysis of the fluorescence intensity of claudin-1. (E) Representative occludin fluorescence images. (F) Quantitative analysis of occludin fluorescence intensity. N = 5 per group. *p < 0.05, **p < 0.01

Our findings demonstrate that long-term consumption of Pu-erh tea significantly reduces pro-inflammatory cytokines and enhances antioxidant defenses (e.g., SOD levels) in DSS-induced mice. By breaking the cycle of inflammation and oxidative stress, Pu-erh tea helps alleviate IBD symptoms. On the one hand, the abundance of anti-inflammatory bacteria (e.g., Lactobacillus) and reducing pathogenic bacteria (e.g., Proteobacteria) by Pu-erh tea restore gut homeostasis and reduce inflammation. On the other hand, Pu-erh tea enhances the expression of tight junction proteins, thereby improving the integrity of the intestinal mucosal barrier. This reduces intestinal permeability and prevents the translocation of harmful substances that can trigger inflammation.

In conclusion, our study highlights the novel and sustained benefits of long-term Pu-erh tea consumption in the management of IBD. Unlike the short-term effects reported in previous studies, our research reveals prolonged anti-inflammatory and antioxidative benefits, substantial modulation of gut microbiota, and significant restoration of intestinal barrier integrity. These extended benefits emphasize the potential of Pu-erh tea as a natural therapeutic option for chronic IBD management. Further research is warranted to explore the underlying mechanisms and to validate these findings in human clinical trials, aiming to establish Pu-erh tea as a complementary or alternative treatment with long-lasting effects.

Supplementary Information

Below is the link to the electronic supplementary material.

Supplementary file1 (DOCX 406 KB) (405.7KB, docx)

Acknowledgements

This work was supported by Xiamen Science and Technology Plan Project (No. 3502Z20226034), supported by Traditional Chinese Medicine Foundation of Xiamen (Project XWZY-2023-0605) and the Youth Science and Technology of Xiamen Marine and Fishery Development Special Fund (23YYZP013QCA11). We would like to thank Instrumental Analysis Center of Huaqiao University for the help of confocal testing.

Author contributions

G.X., W.H. and L.Y. conceived and designed the study. B.F., P.C., L.Y. and G.X. did animal experiments and biochemical experiments. J.Z. and C.L. analyzed the data. B.F., G.X., L.Y. and W.H. wrote and revised the paper. All authors read and approved the final manuscript.

Declarations

Conflict of interest

The authors declare that there is no conflict of interest.

Footnotes

Publisher's Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Contributor Information

Wei-Feng Huang, Email: hwf0625@xmu.edu.cn.

Li-Tao Yi, Email: litaoyi@hqu.edu.cn.

Guang-Hui Xu, Email: xghcxm@163.com.

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