Fecal Microbiota Transplantation Attenuates COPD by Regulating the MMP-9/TIMP-1 Pathway

WeiXu1

YangYang2

BingbingLi5

fuqiangLi1

BaofangZhang3

YijuCheng4

HongpiaoLi6

MinglianCheng3✉Emailchengml@21cn.com

Department of gastroenterologyThe affiliated Hospital of Guizhou Medical UniversityGuiyang

Department of nephrologyThe First People’s Hospital Of GuiyangGuiyang

Department of Infectious DiseasesThe affiliated Hospital of Guizhou Medical UniversityGuiyang

The fourth People’s Hospital Of GuiyangGuiyang

5Department of GastroenterologyThe First Affiliated Hospital of Jinan University, Jinan UniversityGuangzhouGuangdongChina

Department of GastroenterologyXinfeng County People’s HospitalGuangdongChina

Wei Xu¹,Yang Yang²,Bingbing Li⁵,fuqiang Li¹,Baofang Zhang³,Yiju Cheng⁴,Hongpiao Li⁶,Minglian Cheng³

¹Department of gastroenterology.The affiliated Hospital of Guizhou Medical University,Guiyang

²Department of nephrology, The First People's Hospital Of Guiyang,Guiyang

³Department of Infectious Diseases,The affiliated Hospital of Guizhou Medical University,Guiyang

⁴The fourth People's Hospital Of Guiyang,Guiyang

⁵Department of Gastroenterology, The First Affiliated Hospital of Jinan University, Jinan University, Guangzhou, Guangdong, China

⁶Department of Gastroenterology, Xinfeng County People's Hospital, Guangdong, China

Correspondence:Minglian Cheng,chengml@21cn.com

Wei Xu,Yang Yang,Bingbing Li

These authors have contributed equally to this work

Abstract

Background

Chronic obstructive pulmonary disease (COPD) is a leading cause of morbidity and mortality worldwide, with limited therapeutic options beyond symptom control. Airway remodeling driven by imbalance of the MMP-9/TIMP-1 axis is central to COPD progression. While fecal microbiota transplantation (FMT) has shown promise in immune regulation, its therapeutic potential and mechanism in COPD remain undefined. We aimed to determine whether FMT alleviates COPD by modulating the MMP-9/TIMP-1 signaling pathway.

Methods

Using an in vivo mouse model was established using cigarette smoke exposure and intratracheal lipopolysaccharide. Following model induction, rats received daily intragastric FMT for four weeks. Lung pathology, MMP-9/TIMP-1 expression, inflammatory cytokines, immune cell subsets, and gut microbiota diversity were analyzed.

Results

COPD model rats exhibited severe airway injury, upregulated MMP-9/TIMP-1 signaling, heightened inflammation, and reduced microbial diversity. FMT treatment significantly attenuated airway remodeling, restored MMP-9/TIMP-1 balance, reduced inflammatory cytokine levels, rebalanced Th17/Treg cells, and improved gut microbiota richness and diversity.

Conclusion

Our findings demonstrate for the first time that FMT can ameliorate COPD by suppressing MMP-9/TIMP-1–mediated airway remodeling through modulation of the gut–lung axis. This work highlights FMT as a novel therapeutic strategy with mechanistic insight into its role in COPD treatment.

Keywords:

Chronic obstructive pulmonary disease

fecal microbiota transplantation

MMP-9/TIMP-1

1 Introduction

Chronic obstructive pulmonary disease (COPD) is one of the most common respiratory disorders and is often caused by long-term inhalation of cigarette smoke [1]. It results in irreversible airflow limitation and ventilatory dysfunction due to chronic and persistent airway inflammation, small airway disease, emphysema, and loss of lung elasticity [2]. COPD is a major cause of global mortality and disability. According to estimates from the Global Burden of Disease Study 2019, it ranked sixth among 369 diseases in terms of disability-adjusted life years (DALYs), up from eleventh in 1990 [3], making it the third leading cause of death worldwide. It has become a significant public health challenge and has greatly increased the global disease burden. The pathogenesis of COPD is complex, and effective treatment strategies have yet to be fully established. Currently, inhaled corticosteroids and bronchodilators are the first-line therapies for COPD [1]. Although these drugs can alleviate symptoms, they have limited effects on disease progression and mortality reduction, and are associated with serious adverse effects. Therefore, there is an urgent need to develop more effective therapeutic approaches for COPD [4].

In recent years, the role of the microbiota in modulating host immune function has received increasing attention. Most COPD-related studies have focused on the respiratory microbiota [5], but the gut harbors the largest and most diverse microbial community in the human body. Emerging evidence indicates a close interaction between gut microbiota composition and pulmonary immunity [6]. Cigarette smoke, a major contributing factor to COPD, not only harms pulmonary health but also adversely affects the gastrointestinal system, promoting the development of intestinal disorders [7]. COPD patients have a higher incidence of inflammatory bowel disease (IBD) [8], and they often exhibit gut microbiota dysbiosis and increased intestinal permeability [9]. Similarly, IBD patients show altered gut microbiota composition and decreased pulmonary function [10]. The interaction between gut microbiota and lung function is referred to as the "gut-lung axis" [7].

Many respiratory diseases impair the intestinal barrier, and alterations in gut microbiota composition are associated with the onset and progression of COPD [11]. Dysbiosis can lead to an increase in opportunistic pathogens, bacterial translocation, and endotoxin release. Once the intestinal barrier is compromised, innate immune responses are activated, with Toll-like receptor 4 (TLR4) playing a key role in infection suppression [12]. This, in turn, activates systemic immunity, promotes bacterial translocation to the lungs, exacerbates immune-mediated lung injury [13], and increases the expression of inflammatory cytokines such as interleukin-1β (IL-1β), interleukin-6 (IL-6), and tumor necrosis factor-α (TNF-α) [14]. Inflammation and oxidative stress affect protease homeostasis, leading to enhanced extracellular matrix (ECM) degradation and resulting in progressive alveolar destruction and airway remodeling [15]. Matrix metalloproteinases (MMPs) are a family of proteolytic enzymes that promote inflammation and degrade ECM components, while tissue inhibitors of metalloproteinases (TIMPs) inhibit MMP activity. Studies have shown that a protease-antiprotease imbalance is associated with airway damage in COPD patients. Among these, MMP-9 levels and the MMP-9/TIMP-1 ratio are considered strong predictors of emphysema in COPD [16].

Fecal microbiota transplantation (FMT), which involves transplanting functional gut microbiota from healthy donors into patients to restore intestinal ecological balance, has become an effective therapy for Clostridioides difficile infection [17]. It has also demonstrated good efficacy and safety in clinical trials for inflammatory bowel disease and irritable bowel syndrome [18, 19]. Given its high efficacy and safety profile, FMT has started to be explored for COPD treatment. In one study, researchers used FMT to treat mice with cigarette smoke-induced COPD and found that it effectively alleviated airway and systemic symptoms, alveolar structural damage, and intestinal pathological changes [20]. However, the potential mechanisms by which gut microbiota exert effects via the gut-lung axis and target molecules in FMT remain unclear.

Therefore, we hypothesize that FMT may have therapeutic effects on COPD by regulating the MMP-9/TIMP-1 pathway. A stable rat model of COPD was established using cigarette smoke exposure combined with intratracheal instillation of lipopolysaccharide (LPS). FMT was used to alter the gut microbiota in these rats to counteract COPD. Advanced techniques such as 16S rDNA amplification and sequencing were employed to analyze the composition and diversity of gut microbiota, investigate the effects of FMT on gut microbial structure and immune responses, and reveal the involved regulatory pathways.

2 Methods

2.1 Animals, Model Induction, and Fecal Microbiota Transplantation

Male Sprague-Dawley (SD) rats, aged 6–8 weeks, specific pathogen-free (SPF) grade, were purchased from Beijing Huafukang Biological Technology Co., Ltd. (License No. SCXK [Jing] 2019-0008). Animals were housed under controlled conditions: temperature 20–26°C, humidity 40%–70%, with standard feed and water provided ad libitum. Rats were randomly divided into five groups: Blank Control Group (Control): normal rats (n = 8);Negative Control Group (Saline solution group): model rats receiving saline transplantation (n = 8);Positive Control Group (Model group): normal rats receiving microbiota from model rats (n = 6);Experimental Group A (Microflora therapy group): model rats receiving microbiota from normal rats (n = 6).All animals underwent a one-week acclimatization period. A stable-phase chronic obstructive pulmonary disease (COPD) model was established by intratracheal instillation of lipopolysaccharide (LPS) combined with cigarette smoke exposure. Specifically, rats in groups 2 and 4 were anesthetized and administered 1 ml/kg of LPS solution (1 mg/ml) via intratracheal instillation on days 1 and 15. After instillation, rats were held upright and rotated for 1 minute to ensure even distribution of the solution in the lungs. From days 2–14 and 16–28, these rats were exposed to passive cigarette smoke in a fumigation chamber for 30 minutes daily. Rats in groups 1 and 3 received no treatment. After modeling, two rats each from groups 1 and 2 were euthanized for lung histological validation using hematoxylin-eosin (HE) staining. Following successful modeling (i.e., after the final smoking session), treatment was initiated: group 1 received no treatment; group 2 was gavaged with saline; group 3 was gavaged with fecal microbiota suspension from COPD model rats; group 4 was gavaged with fecal microbiota from normal rats. The gavage dose was 10 ml/kg, administered once daily for four weeks.

2.2 Hematoxylin-Eosin (HE) Staining

Cardiac tissue was removed and rinsed in running water for several hours, followed by dehydration through graded ethanol solutions (70%, 80%, 90%). The tissue was then immersed in a 1:1 solution of absolute ethanol and xylene for 15 minutes, then in xylene I and xylene II each for 15 minutes until transparent. Tissues were transferred to a 1:1 mixture of xylene and paraffin for 15 minutes, followed by paraffin I and II infiltration for 50–60 minutes each. Paraffin embedding and sectioning were performed. Sections were baked, dewaxed, and rehydrated. After immersion in distilled water, sections were stained with hematoxylin for 3 minutes, differentiated with ethanol-hydrochloric acid for 15 seconds, washed, blued for 15 seconds, rinsed with running water, stained with eosin for 3 minutes, dehydrated, cleared, mounted, and examined microscopically.

2.3 16S rDNA Microbial Diversity Sequencing

Fecal samples were collected from three rats per group before and after microbiota transplantation. DNA was extracted from fecal samples and subjected to high-throughput 16S rDNA sequencing to analyze intestinal microbial diversity and compare differences in microbiota composition and abundance across groups.

2.4 ELISA Detection

Serum samples were collected from rats, and levels of IL-1β, IL-6, and TNF-α were measured using enzyme-linked immunosorbent assay (ELISA), strictly following the kit instructions.

2.5 Flow Cytometry Analysis

Bronchoalveolar lavage fluid (BALF) was collected and centrifuged at 1500 rpm for 3 minutes, washed twice with PBS; Fivefold Binding Buffer was diluted to 1× with double-distilled water; Cells were resuspended in 300 µl pre-cooled 1× Binding Buffer; Each tube received 5 µl Annexin V-FITC and 10 µl PI; After gentle mixing, samples were incubated in the dark at room temperature for 10 minutes;200 µl of pre-cooled 1× Binding Buffer was added; Samples were analyzed using flow cytometry.

2.6 Quantitative Real-Time PCR (qPCR)

Lung tissues were homogenized in Trizol for mRNA extraction. RNA concentration and purity were assessed, followed by reverse transcription into cDNA using a commercial kit. qPCR was conducted with β-actin as an internal control to determine the relative expression levels of TLR4, NF-κB, MMP-9, and TIMP-1 in lung tissues (primer sequences detailed

in Supplementary Table 1).

2.7 Immunohistochemistry (IHC)

Paraffin Section Preparation: Baking: sections were baked at 65°C for 2 hours; Dewaxing: placed in xylene for 10 minutes, replaced with fresh xylene for another 10 minutes; Rehydration: sequential immersion in 100%, 100%, 95%, 80% ethanol and purified water for 5 minutes each. Antigen Retrieval: sections were placed in citrate buffer in a retrieval box and heated in a pressure cooker. After steam release, heating was stopped and allowed to cool naturally, followed by PBS rinsing. Endogenous Peroxidase Blocking: sections were incubated with freshly prepared 3% hydrogen peroxide for 10 minutes in a humidified chamber at room temperature and then rinsed with PBS. Non-specific Blocking: PBS washing (3×, 5 minutes each), blotted dry, then incubated with 5% BSA at 37°C for 30 minutes.

2.8 Immunoreaction:

Primary antibody incubation: excess blocking solution was removed, and sections were incubated with diluted anti-RIPK3 antibody (1:150) overnight at 4°C in a humidified chamber; Secondary antibody incubation: sections were brought to room temperature for 45 minutes, washed, and incubated with HRP-conjugated goat anti-rabbit IgG (H + L) (1:150) at 37°C for 30 minutes, followed by PBS washing. Color Development and Counterstaining: DAB staining for 5–10 minutes under microscopic observation, rinsed with PBS or tap water for 1 minute; Hematoxylin counterstaining for 3 minutes, followed by differentiation, bluing, rinsing, dehydration, clearing, mounting, and microscopy.

2.9 Pulmonary Function Testing

Pulmonary function was assessed using the Buxco Pulmonary Function Analysis System (Buxco Electronics, DSI Buxco-PFT). Rats were anesthetized with 3% pentobarbital sodium (1 ml/kg), intubated, and placed in a forced pulmonary maneuver system. With a controlled ventilation rate of approximately 60 breaths per minute, respiratory flow, oral pressure, esophageal pressure, and heart rate were monitored. Parameters measured included inspiratory capacity (IC), lung resistance (RL), forced expiratory flow at 75% of forced vital capacity (FEF75), forced expiratory volume at 100 milliseconds (FEV100), dynamic lung compliance (Cdyn), and forced vital capacity (FVC). Each parameter was averaged from three measurements per rat.

2.10 Statistical Analysis

Statistical analyses were performed using SPSS 20.0. Quantitative data were expressed as mean ± standard deviation. Normality and homogeneity of variance were tested. Unpaired t-tests were used for intergroup comparisons. One-way ANOVA or repeated measures ANOVA was employed for multi-group comparisons. A significance threshold of α = 0.05 was set, with P < 0.05 considered statistically significant.

3 Results

3.1 FMT Alleviates Inflammatory Response in COPD

To investigate alterations in the microbiota composition and the effects of fecal microbiota transplantation (FMT), a stable COPD rat model was established using SD rats through cigarette smoke exposure combined with intratracheal instillation of LPS. FMT effectively alleviated airway inflammation associated with COPD. Some rats underwent passive microbial transplantation for intervention. Cigarette smoke and LPS exposure induced pulmonary inflammation. Compared with the control group, the Saline solution group exhibited a lower percentage of Treg cells and a higher percentage of Th17 cells. The gut microbiota from COPD rats induced pulmonary inflammation in healthy mice. Similarly, in the Model group, the Treg cell percentage was reduced and Th17 increased compared to the control group. However, transplanting the gut microbiota from healthy rats into COPD rats significantly alleviated lung inflammation: the Microflora therapy group showed a lower proportion of Th17 cells and a higher proportion of Treg cells in lung tissue compared with the Model group (Figure. 1A-D).Additionally, both COPD rats and healthy rats receiving microbiota from COPD rats demonstrated enhanced systemic inflammation, with dysbiosis elevating levels of IL-1β, IL-6, and TNFα. In contrast, FMT significantly reduced the systemic inflammatory response induced by cigarette smoke and LPS ( Figure.1E-G). These findings indicate that FMT can restore and improve inflammation in COPD.

Fig. 1

FMT alleviates local and systemic inflammation induced by COPD. (A–D) Flow cytometry analysis of Treg and Th17 cell populations in lung tissues. (E–F) ELISA results showing serum levels of inflammatory cytokines IL-1β, IL-6, and TNFα in rats. *=p < 0.05; **p = < 0.01; ***p = < 0.001; ****p = < 0.0001.

3.2 FMT Alleviates Pulmonary Pathological Damage and Improves Lung Function in COPD

Compared with control rats, HE staining revealed that the gut microbiota plays a significant role in the pathogenesis of COPD. Normal rats receiving microbiota from COPD donors exhibited marked signs of lung injury: the model group showed deformed bronchial lumens, pronounced epithelial cell shedding, thinning of the alveolar walls, and substantial inflammatory cell infiltration around the airways. These pathological features were markedly reduced in the Microflora Therapy Group that received healthy microbiota. In this group, the bronchial structure remained relatively intact, with some epithelial shedding and mild inflammatory cell infiltration observed (Fig. 2A).Pulmonary function assessment is widely regarded as the gold standard for diagnosing chronic obstructive pulmonary disease (COPD). In this study, compared with the normal control group, both the Saline Solution Group and the Model Group exhibited significant reductions in inspiratory capacity (IC), forced vital capacity (FVC), and dynamic compliance (Cdyn), suggesting that dysbiosis leads to impaired pulmonary ventilation and decreased lung elasticity. In contrast, rats in the Microflora Therapy Group that received fecal microbiota transplantation (FMT) showed significant recovery in IC, FVC, and Cdyn levels (Fig. 2B–G), indicating a degree of lung function improvement. These findings suggest that FMT may have potential therapeutic value in ameliorating COPD-related pulmonary dysfunction.

Fig. 2

FMT ameliorates pathological damage and improves lung function in COPD. (A) Representative images of lung histopathology under H&E staining at 100× and 400× magnification. (B–G) Pulmonary function test results showing changes in IC, FVC, RL, Cdyn, FEF75, and FEV100 in rats. *=p < 0.05; **p = < 0.01; ***p = < 0.001; ****p = < 0.0001.

3.3 FMT Enhances Gut Microbiota Diversity in COPD Rats

Microbial composition analysis showed that Bacteroidota and Firmicutes were the dominant phyla across all groups. Before transplantation, both the Control and Saline solution groups had similar levels of Bacteroidota and Firmicutes. After transplantation, the ratio in the Control group shifted, with Bacteroidota becoming more dominant than Firmicutes. In contrast, the Saline solution and Model groups showed higher Firmicutes abundance than Bacteroidota and increased overall microbiota diversity. The Model group also exhibited significantly increased abundance of Spirochaetota and Desulfobacterota, which was notably reduced after FMT ( Fig. 3A-B).

Fig. 3

FMT improves gut microbiota diversity in COPD rats. (A) Analysis of microbial richness. (B) Heatmap showing the composition of microbiota at various taxonomic levels.

3.4 MMP-9/TIMP-1 Pathway Regulates Airway Epithelial Barrier Function

The airway epithelial barrier is the first line of defense against inhaled smoke and pathogens. Barrier dysfunction in COPD contributes to impaired defense, persistent inflammation, and infection exacerbation. The MMP-9/TIMP-1 pathway is associated with airway epithelial barrier integrity. Immunohistochemical results showed increased MMP-9 and TIMP-1 expression and decreased E-cadherin and occludin expression in the Saline solution and Model groups compared to the Control group. In the Microflora therapy group, FMT significantly downregulated MMP-9 and TIMP-1 while upregulating E-cadherin and occludin (Fig. 4A-B), indicating that FMT improves epithelial barrier function via regulation of the MMP-9/TIMP-1 pathway.

Fig. 4

FMT improves gut microbiota diversity in COPD rats. (A) Analysis of microbial richness. (B) Heatmap showing the composition of microbiota at various taxonomic levels.

3.5 FMT Ameliorates COPD via MMP-9/TIMP-1 Pathway Regulation

Gut dysbiosis leads to overgrowth of potential pathogens, bacterial translocation, and endotoxin release, which breach the gut barrier and activate pulmonary immune responses via the TLR4 signaling pathway. To investigate how gut microbiota influence the TLR4/NF-κB signaling axis in COPD, we assessed relative expression levels of TLR4, NF-κB, MMP-9, and TIMP-1 in lung tissue. Compared to the Control group, these markers were elevated in both the Saline solution and Model groups, with significant differences observed in the Model group. Notably, the Model group showed significantly higher TLR4 and MMP-9 expression compared with the Saline solution group, suggesting that microbial dysbiosis activates the TLR4/NF-κB pathway and upregulates MMP-9/TIMP-1 expression. However, these expression levels were significantly reduced in the Microflora therapy group (Fig. 5A-D), indicating that FMT may alleviate COPD through modulation of the MMP-9/TIMP-1 axis.

Fig. 5

FMT treats COPD by regulating the MMP-9/TIMP-1 pathway.(A–D) qPCR analysis of lung tissue to assess the expression levels of genes related to the MMP-9/TIMP-1 pathway, including TLR4, NF-κB, MMP-9, and TIMP-1.*=p < 0.05; **p = < 0.01; ***p = < 0.001; ****p = < 0.0001.

4 Discussion

The gastrointestinal system is the largest organ in the human body [21], and it harbors a vast and complex gut microbiota. With advances in high-throughput sequencing and metagenomics, it is now possible to obtain dynamic insights into changes in the human gut microbiota, shedding light on the intricate interactions between gut microbes and disease. Current research indicates that the onset and progression of chronic obstructive pulmonary disease (COPD) are closely related to gut health and alterations in the intestinal microenvironment [22]. Fecal microbiota transplantation (FMT) can modulate the gut microbiota of COPD patients, thereby suppressing local and systemic inflammation, mitigating lung tissue damage, and improving pulmonary function. Our study demonstrates that FMT exerts therapeutic effects on COPD by regulating the MMP-9/TIMP-1 signaling pathway.

Gut microbiota dysbiosis may affect the respiratory system by compromising the integrity of the intestinal mucosa. Mucins secreted by intestinal epithelial cells restrict the translocation of harmful bacteria from the intestinal lumen to the epithelium. Once the intestinal barrier is disrupted, this restriction is lifted, increasing the risk of bacterial translocation to distant organs and potentially resulting in severe systemic responses [23]. In our COPD rat model induced by cigarette smoke exposure combined with LPS instillation, gut microbiota dysbiosis was characterized by increased abundances of Firmicutes, Spirochaetota, and Desulfobacterota, alongside a decreased abundance of Bacteroidota. A clinical study involving fecal samples from 73 healthy individuals and 99 COPD patients revealed a relatively lower proportion of Bacteroidota and a higher proportion of Firmicutes in COPD patients compared with healthy controls [24]. Moreover, a dominance of Spirochaetota was observed in both COPD patients and animal models, indicating microbial dysbiosis [24,25]. These findings suggest that Firmicutes, Spirochaetota, and Desulfobacterota may represent pathogenic microbiota in COPD.

Damage to the intestinal barrier induced by COPD can lead to the translocation of bacteria and their metabolites, such as endotoxins and lipopolysaccharides (LPS), which may promote disease progression [26]. Previous studies support the notion that imbalances between Bacteroidota and Firmicutes may act as potential triggers for inflammation [27]. In our study, the relative abundance of Bacteroidota was reduced and Firmicutes increased in the model group following FMT from diseased donors, consistent with findings in diabetes and obesity research [27, 28]. The abundances of Spirochaetota and Desulfobacterota were significantly elevated in COPD and mirrored similar outcomes observed in mice subjected to cigarette smoke-induced microbiota disruption [29]. Dysbiosis-induced bacterial translocation may activate pulmonary immune responses, where sustained inflammation leads to the release of multiple inflammatory mediators, promoting activation of various inflammatory cells, damaging lung tissue, and inducing oxidative stress [30]. Notably, the gut microbiota from COPD patients has been shown to exacerbate mucus hypersecretion and accelerate lung function decline in mice [31].

In our study, the model group exhibited severe local and systemic inflammation, bronchial lumen deformation, prominent epithelial shedding, thinning of alveolar walls, and significant infiltration of inflammatory cells. Elevated levels of IL-1β, IL-6, and TNF-α were observed, alongside decreased values of inspiratory capacity (IC), forced vital capacity (FVC), and dynamic compliance (Cdyn), consistent with previous findings.

The pathogenesis of COPD involves dysregulated protease activity, degradation of the extracellular matrix (ECM), progressive destruction of alveoli, and airway remodeling, ultimately resulting in functional impairment and irreversible loss of lung function [32]. Among these processes, airway remodeling is driven by chronic airway inflammation and repeated cycles of tissue injury and repair [33]. MMP-9 and TIMP-1 play critical roles in this process and are commonly used as key indicators for assessing ECM deposition and the extent of airway remodeling. Serum levels of matrix metalloproteinase-9 (MMP-9) and tissue inhibitor of metalloproteinase-1 (TIMP-1) can be used to diagnose and evaluate COPD severity [34]. MMP-9 regulates cell adhesion and acts on extracellular components or other proteins, thereby participating in angiogenesis, tissue remodeling, embryogenesis, and wound healing [35]. TIMP-1 is a specific inhibitor of MMP-9 and is synthesized and secreted by various inflammatory and structural cells in the lungs. It may inhibit MMP-mediated endothelial cell migration, promote collagen synthesis and fibroblast proliferation, suppress ECM degradation, and facilitate ECM deposition. TLR-4 regulates the expression of MMP-9 via activation of the NF-κB signaling pathway [36]. In our study, immunohistochemistry and RT-PCR data demonstrated significantly increased expression of MMP-9 and TIMP-1 in the COPD model. In contrast, their expression was markedly reduced following FMT, suggesting that the therapeutic effects of FMT may be associated with downregulation of MMP-9 and TIMP-1 expression in lung tissue.

There are several limitations to our study that should be addressed in future research. First, the relationship between gut microbiota and COPD remains largely correlative; further causal studies, such as bacterial depletion experiments, are needed to validate these findings. Second, although our animal model provides important insights into the mechanisms of FMT in COPD, its translational relevance to humans remains uncertain. Future studies should investigate whether FMT can improve COPD in patients by modulating the MMP/TIMP-1 pathway.

5 Conclusions

In conclusion, this study reveals the potential therapeutic effects of fecal microbiota transplantation (FMT) in improving chronic obstructive pulmonary disease (COPD), possibly through regulation of the MMP-9 and TIMP-1 signaling pathways. The expression changes of MMP-9 and TIMP-1 may play a key role in the bidirectional regulation between the lungs and gut microbiota, offering a novel theoretical basis and research direction for FMT in COPD treatment. However, current studies are limited by the lack of large-scale clinical validation and mechanistic depth. Therefore, further animal and clinical studies are needed to elucidate the specific mechanisms, safety, and efficacy of FMT interventions in COPD, thereby providing a stronger theoretical foundation and practical support for microbiota-based COPD therapy.

Declarations

Data Availability

The datasets used and/or analysed during the current study are available from the corresponding author on reasonable request.

Acknowledgements

Not applicable.

Clinical trial number.

Not applicable.

Funding

The author(s) declare that financial support was received for the research and/or publication of this article. The project was funded by the Science and Technology Fund of the Guizhou Provincial Health Commission (gzwkj2021-125) and the Health Research Project of Shaoguan (No. Y25066).

Contributions

WX: Project administration, Formal analysis, Writing – original draft, Methodology, Supervision, Data curation, Software, Conceptualization, Visualization, Resources, Funding acquisition, Investigation, Validation, Writing – review & editing. YY: Resources, Investigation, Data curation, Writing – review & editing. BL: Conceptualization, Data curation, Investigation, Methodology, Software, Writing – original draft, Writing – review & editing. FL: Writing – original draft, Investigation, Data curation. BZ: Writing – original draft, Data curation, Investigation. YC: Data curation, Investigation, Writing – original draft. HL: Formal analysis, Validation, Writing – review & editing. MC: Conceptualization, Formal analysis, Funding acquisition, Investigation, Methodology, Project administration, Resources, Software, Supervision, Writing – original draft, Writing – review & editing.

Corresponding author

Correspondence to Minglian Cheng

Ethics declarations

Ethics approval and consent to participate

The animal study was approved by Animal Ethics Committee of Guizhou Medical University.

The study was conducted in accordance with the local legislation and institutional requirements.

Consent for publication

Not applicable.

Competing interests

The authors declare no competing interests.

Additional information

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Author Contribution

References

CELLI B R AGUSTÍA, CRINER G J, et al. Global Initiative for Chronic Obstructive Lung Disease 2023 Report: GOLD Executive Summary[J/OL]. Am J Respir Crit Care Med. 2023;207(7):819–37.

CHRISTENSON S A, SMITH B M, BAFADHEL M, et al. Chronic obstructive pulmonary disease[J/OL]. Lancet. 2022;399(10342):2227–42.

VOS T, LIM S S, ABBAFATI C, et al. Global burden of 369 diseases and injuries in 204 countries and territories, 1990–2019: a systematic analysis for the Global Burden of Disease Study 2019[J/OL]. Lancet. 2020;396(10258):1204–22.

HALPIN D M G, CRINER G J, PAPI A, Prevention of Chronic Obstructive Lung Disease. Global Initiative for the Diagnosis, Management, and. The 2020 GOLD Science Committee Report on COVID-19 and Chronic Obstructive Pulmonary Disease[J/OL]. American Journal of Respiratory and Critical Care Medicine, 2021, 203(1): 24–36.

LI R, LI J, ZHOU X. Lung microbiome: new insights into the pathogenesis of respiratory diseases[J/OL]. Signal Transduct Target Therapy. 2024;9:19.

YOU D, WU Y, LU M, et al. A genome-wide cross-trait analysis characterizes the shared genetic architecture between lung and gastrointestinal diseases[J/OL]. Nat Commun. 2025;16:3032.

WANG L, CAI Y, GARSSEN J et al. The Bidirectional Gut–Lung Axis in Chronic Obstructive Pulmonary Disease[J/OL]. Am J Respir Crit Care Med, 207(9): 1145–60.

BRASSARD P, VUTCOVICI M, ERNST P, et al. Increased incidence of inflammatory bowel disease in Québec residents with airway diseases[J/OL]. Eur Respir J. 2015;45(4):962–8.

KEELY S, HANSBRO PM. Lung-Gut Cross Talk: A Potential Mechanism for Intestinal Dysfunction in Patients With COPD[J/OL]. Chest. 2014;145(2):199–200.

10.

RODRIGUEZ-ROISIN R, BARTOLOME S D, HUCHON G, et al. Inflammatory bowel diseases, chronic liver diseases and the lung[J/OL]. Eur Respir J. 2016;47(2):638–50.

11.

Shi H, Zhao T, Geng R, Sun L, Fan H. The associations between gut microbiota and chronic respiratory diseases: a Mendelian randomization study. Front Microbiol. 2023;14:1200937.

12.

BUDDEN K F, GELLATLY S L, WOOD D L A, et al. Emerging pathogenic links between microbiota and the gut–lung axis[J/OL]. Nat Rev Microbiol. 2017;15(1):55–63.

13.

TAN J, yang TANGY, chun HUANGJ. Gut Microbiota and Lung Injury[M/OL]//CHEN P. Gut Microbiota and Pathogenesis of Organ Injury. Singapore: Springer; 2020. pp. 55–72. https://doi.org/10.1007/978-981-15-2385-4_5. [2025-04-27].

14.

TANG J, XU L, ZENG Y, et al. Effect of gut microbiota on LPS-induced acute lung injury by regulating the TLR4/NF-kB signaling pathway[J/OL]. Int Immunopharmacol. 2021;91:107272.

15.

BARNES PJ. Cellular and Molecular Mechanisms of Chronic Obstructive Pulmonary Disease[J/OL]. Clin Chest Med. 2014;35(1):71–86.

16.

DIMIC-JANJIC S, HODA M A, MILENKOVIC B, et al. The usefulness of MMP-9, TIMP-1 and MMP-9/TIMP-1 ratio for diagnosis and assessment of COPD severity[J/OL]. Eur J Med Res. 2023;28:127.

17.

MINKOFF N Z, ASLAM S, MEDINA M, et al. Fecal microbiota transplantation for the treatment of recurrent Clostridioides difficile (Clostridium difficile)[J/OL]. Cochrane Database Syst Rev. 2023;2023(4):CD013871.

18.

EL-SALHY M, HATLEBAKK J G, GILJA O H, et al. Efficacy of faecal microbiota transplantation for patients with irritable bowel syndrome in a randomised, double-blind, placebo-controlled study[J/OL]. Gut. 2020;69(5):859–67.

19.

PARAMSOTHY S, KAMM M A, KAAKOUSH N O, et al. Multidonor intensive faecal microbiota transplantation for active ulcerative colitis: a randomised placebo-controlled trial[J/OL]. Lancet. 2017;389(10075):1218–28.

20.

BUDDEN K F, SHUKLA S D, BOWERMAN K L, et al. Faecal microbial transfer and complex carbohydrates mediate protection against COPD[J/OL]. Gut. 2024;73(5):751–69.

21.

YANG Y, ZHANG H, WANG Y, et al. Promising dawn in the management of pulmonary hypertension: The mystery veil of gut microbiota[J/OL]. iMeta. 2024;3(2):e159.

22.

SONG X, DOU X. CHANG J, The role and mechanism of gut-lung axis mediated bidirectional communication in the occurrence and development of chronic obstructive pulmonary disease[J/OL]. Gut Microbes, 16(1): 2414805.

23.

WYPYCH T P, WICKRAMASINGHE L C, MARSLAND B J. The influence of the microbiome on respiratory health[J/OL]. Nature Immunology, SONG J, YANG H et al. Gut microbiota was highly related to the immune status in chronic obstructive pulmonary disease patients[J/OL]. Aging, 2024, 16(4): 3241–3256.

25.

TOMODA K, KUBO K, ASAHARA T, et al. Cigarette smoke decreases organic acids levels and population of bifidobacterium in the caecum of rats[J/OL]. J Toxicol Sci. 2011;36(3):261–6.

26.

LIM E Y, SONG E J, SHIN HS. Gut Microbiome as a Possible Cause of Occurrence and Therapeutic Target in Chronic Obstructive Pulmonary Disease[J/OL]. 2023, 33(9): 1111–8.

27.

QIN J, LI Y, CAI Z, et al. A metagenome-wide association study of gut microbiota in type 2 diabetes[J/OL]. Nature. 2012;490(7418):55–60.

28.

TURNBAUGH PJ, HAMADY M, YATSUNENKO T, et al. A core gut microbiome in obese and lean twins[J/OL]. Nature. 2009;457(7228):480–4.

29.

LAIMAN V, CHUANG H C, LO Y C, et al. Cigarette smoke-induced dysbiosis: comparative analysis of lung and intestinal microbiomes in COPD mice and patients[J/OL]. Respir Res. 2024;25:204.

30.

ZHANG Z M, WANG Y C CHENL et al. Protective effects of the suppressed NF-κB/TLR4 signaling pathway on oxidative stress of lung tissue in rat with acute lung injury[J/OL]. [2025][2025-05-08].

31.

LI N, DAI Z, WANG Z, et al. Gut microbiota dysbiosis contributes to the development of chronic obstructive pulmonary disease[J/OL]. Respir Res. 2021;22(1):274.

32.

FISCHER B M, PAVLISKO E, VOYNOW J A. Pathogenic triad in COPD: oxidative stress,protease–antiprotease imbalance, and inflammation[J/OL]. International Journal of Chronic Obstructive Pulmonary Disease, 2011, 6: 413–421.

33.

HIROTA N, MARTIN JG. Mechanisms of Airway Remodeling[J/OL]. Chest. 2013;144(3):1026–32.

34.

DIMIC-JANJIC S, HODA M A, MILENKOVIC B, et al. The usefulness of MMP-9, TIMP-1 and MMP-9/TIMP-1 ratio for diagnosis and assessment of COPD severity[J/OL]. Eur J Med Res. 2023;28:127.

35.

KLEINER D E, STETLER-STEVENSON W G. Matrix metalloproteinases and metastasis[J/OL]. Cancer Chemother Pharmacol. 1999;43(1):S42–51.

36.

EHRENTRAUT H, MEYER R, SCHWEDERSKI M, et al. Systemically Administered Ligands of Toll-Like Receptor 2, -4, and – 9 Induce Distinct Inflammatory Responses in the Murine Lung[J/OL]. Mediat Inflamm. 2011;2011:746532.

Yes