Probiotic supplementation reduces IL-6 and improves clinical outcomes in patients with mild-to-moderate COPD: a randomized placebo-controlled trial

Abstract

Background

Chronic obstructive pulmonary disease (COPD) is characterized by persistent inflammation and progressive airflow limitation. Emerging evidence highlights the gut–lung axis as a potential therapeutic target, with probiotics proposed to modulate Th17-related inflammatory pathways.

Methods

In this randomized, double-blind, placebo-controlled trial, 50 patients with mild-to-moderate COPD were enrolled; 44 completed the 8-week intervention (23 probiotics, 21 placebo). Participants received either a multistrain probiotic formulation or placebo. Outcomes included spirometry, COPD Assessment Test (CAT), modified Medical Research Council (mMRC) dyspnea scale, and serum IL-6, IL-17, and TGF-β levels.

Results

Probiotic supplementation significantly improved FEV1 and FVC within the intervention group, although between-group spirometric differences were not significant. IL-6 levels declined significantly following probiotic therapy, with a significantly greater reduction compared to placebo, whereas IL-17 and TGF-β remained unchanged. CAT scores improved significantly in the probiotic group, exceeding the minimal clinically important difference and demonstrating a significant between-group effect. No significant change was observed in mMRC scores.

Conclusion

Eight weeks of probiotic supplementation was associated with reduced systemic IL-6 levels and clinically meaningful improvement in patient-reported outcomes in mild-to-moderate COPD. These findings support a potential adjunctive role for probiotics and warrant larger mechanistic trials.

Trial registration

Registered on 26 December 2024 in the Iranian Registry of Clinical Trials (IRCT), registration number IRCT20241211064025N1.

Supplementary Information

The online version contains supplementary material available at 10.1186/s12890-026-04423-8.

Introduction

Chronic obstructive pulmonary disease (COPD) is a progressive respiratory disorder characterized by persistent airflow limitation and chronic inflammation of the airways and lungs. It is a leading cause of morbidity and mortality worldwide, creating a significant healthcare and socioeconomic burden. The disease arises from the interplay of environmental exposures—most notably cigarette smoke and air pollution—and host factors, resulting in structural changes such as airway narrowing, mucus overproduction, and destruction of alveolar walls [1]. The immunopathology of COPD involves both innate and adaptive immune mechanisms. While macrophages and neutrophils play a central role, increasing attention has turned to T helper 17 (Th17) cells—a subset of CD4⁺ T lymphocytes producing interleukin-17 (IL-17) [2]. IL-17 is a potent mediator of neutrophil recruitment and activation, driving the secretion of additional pro-inflammatory cytokines and chemokines [3, 4]. Th17 differentiation is supported by transforming growth factor-beta (TGF-β) and IL-6, with IL-23 maintaining cell stability [5]. In COPD, elevated IL-17 levels are associated with neutrophilic airway inflammation, mucus hypersecretion, and tissue damage, making the Th17 cytokine axis (TGF-β, IL-6, and IL-17) a potential therapeutic target. Elevated Th17-related cytokines have also been associated with increased exacerbation frequency and relative corticosteroid resistance, particularly in neutrophil-dominant COPD phenotypes [6]. Therefore, targeting the Th17 pathway may have clinical implications beyond inflammatory control, potentially influencing disease progression and therapeutic responsiveness.

Conventional COPD management—including bronchodilators, inhaled corticosteroids, and lifestyle modification—focuses on symptom control and reducing exacerbations. These treatments have limited impact on the persistent immune imbalance, particularly in neutrophil-dominant inflammation, where corticosteroid responsiveness is often poor [7]. In particular, IL-17-driven neutrophilic inflammation is often less responsive to corticosteroid therapy, highlighting the need for adjunctive strategies targeting the Th17–neutrophil axis [8]. As a result, there is interest in complementary interventions capable of modulating immune responses and attenuating chronic inflammation.

Probiotics, defined as live microorganisms that confer health benefits when consumed in adequate amounts, have traditionally been studied in gastrointestinal health. In recent years, their potential to influence systemic immunity through the gut–lung axis has gained attention [9]. This bidirectional communication between intestinal microbiota and the respiratory system enables gut-derived immune and metabolic signals to affect pulmonary health.

Probiotics exert immunomodulatory effects through multiple mechanisms, including reinforcement of intestinal barrier integrity, reduction of microbial translocation, production of anti-inflammatory metabolites such as short-chain fatty acids, and competitive inhibition of pathogenic microorganisms. The multi-strain formulation based on prior evidence demonstrating that Lactobacillus and Bifidobacterium species can suppress NF-κB activation, reduce IL-6 production, and modulate Th17 polarization in inflammatory conditions [5].

They can also alter T cell polarization, promoting regulatory T cells and reducing excessive Th1 or Th17 responses depending on the immune environment. Studies in asthma [10], inflammatory bowel disease [11], and rheumatoid arthritis [12] have shown that certain probiotic strains reduce IL-6 and IL-17 while enhancing anti-inflammatory mediators such as TGF-β in a regulatory context.

Although research on probiotics in COPD is still emerging, preliminary findings are promising. Experimental models of chronic airway inflammation have shown reduced neutrophil recruitment and downregulated IL-17 following probiotic administration. Clinical studies suggest potential benefits such as improved antioxidant capacity, fewer respiratory infections, and modulation of systemic inflammatory markers [13, 14]. These effects may be particularly valuable in mild to moderate COPD, where early immune modulation could slow disease progression and improve patient outcomes.

Despite this potential, few randomized clinical trials have explored the impact of probiotics on Th17 manners in COPD or examined how these immune changes relate to clinical measures such as lung function. Addressing this gap could help clarify whether probiotic supplementation is a viable adjunct to conventional COPD therapy. Therefore, this randomized clinical trial aims to assess the effects of probiotic supplementation in patients with mild to moderate COPD, focusing on changes in serum levels of Th17-related cytokines (TGF-β, IL-6, and IL-17), respiratory function test results, and patient-reported outcomes measured by the COPD Assessment Test (CAT) and the modified Medical Research Council (mMRC) dyspnea scale.

Materials and methods

Study design and ethical approval

This study was conducted as a randomized, double-blind, placebo-controlled clinical trial. The study protocol was reviewed and approved by the Ethics Committee of Semnan University of Medical Sciences (IR.SEMUMS.REC.1403.202) and registered at the Iranian Registry of Clinical Trials (https://irct.behdasht.gov.ir: IRCT20241211064025N1). The trial was conducted in accordance with the Declaration of Helsinki and reported following the Consolidated Standards of Reporting Trials (CONSORT) guidelines.

Participants

Fifty eligible individuals were screened for participation between January 2025 and June 2025. Recruitment was completed in June 2025, and all follow-up assessments were finalized by August 2025. Inclusion criteria were as follows: patients aged 40–75 years with a confirmed diagnosis of mild to moderate COPD, according to the Global Initiative for Chronic Obstructive Lung Disease (GOLD) criteria 2024 [15], were considered eligible. All participants had a smoking history of ≥ 10 pack-years and demonstrated a post-bronchodilator FEV1/FVC ratio < 70%, with FEV1 between 40 and 78% of the predicted value.

Exclusion criteria included a history of other chronic pulmonary diseases (such as asthma, pulmonary fibrosis, or tuberculosis), respiratory tract infection within 4 weeks prior to enrollment, malignancy, coronary artery disease, autoimmune disorders, primary or secondary immunodeficiency, long-term systemic corticosteroid use, and receipt of antibiotics, probiotic supplements, or high-dose vitamin supplementation within 3 months before screening. Inhaled corticosteroids were permitted provided that the dose had been stable for at least 3 months prior to enrollment, and their distribution was comparable between groups at baseline. Participants who developed SARS-CoV-2 infection during the study period were excluded from the final analysis. Written informed consent was obtained from all participants prior to enrollment.

Sample size consideration

Given the limited prior data regarding the effects of probiotic supplementation on inflammatory cytokines in COPD, this study was designed as an exploratory pilot trial. Based on preliminary assumptions targeting a moderate effect size (Cohen’s d = 0.7) for inflammatory outcomes, a minimum of 20 participants per group was estimated to provide approximately 80% statistical power at a two-sided alpha level of 0.05. The final enrolled sample (n = 44) was therefore considered sufficient for evaluation of the primary endpoint, while secondary outcomes should be interpreted cautiously.

Randomization and blinding

Participants were randomly assigned in a 1:1 ratio to receive either probiotic supplementation or placebo. Randomization was performed using computer-generated random numbers. Allocation concealment was ensured through sequentially numbered, opaque, sealed envelopes prepared by an independent researcher who was not involved in participant recruitment, intervention administration, or outcome assessment. Both participants and investigators remained blinded to treatment allocation throughout the study period. Blinding was maintained until completion of data analysis.

Intervention

Participants in the intervention arm received one probiotic capsule daily after lunch for eight consecutive weeks. The control group received a visually identical placebo capsule containing 500 mg of starch. The intervention duration of eight weeks was selected based on prior clinical evidence indicating that modulation of systemic inflammatory markers and clinical symptoms following probiotic supplementation typically occurs within 6–8 weeks of continuous administration [9]. Both probiotic and placebo capsules were manufactured and packaged by Zist-Takhmir Co. (Tehran, Iran), licensed by the Iranian Food and Drug Administration. Each probiotic capsule contained a multi-strain formulation of freeze-dried live bacteria, with a total viable count of approximately 3.4 × 10¹⁰ colony-forming units (CFU) per capsule. The formulation included the following bacterial strains: Lactobacillus casei, Lactobacillus acidophilus, Lactobacillus rhamnosus, Lactobacillus bulgaricus, Bifidobacterium breve, Bifidobacterium longum, and Streptococcus thermophilus. The approximate contribution of each strain per capsule was as follows: L. casei (3 × 10⁹ CFU), L. acidophilus (3 × 10⁹ CFU), L. rhamnosus (7 × 10⁹ CFU), L. bulgaricus (5 × 10⁸ CFU), B. breve (2 × 10¹⁰ CFU), B. longum (1 × 10⁹ CFU), and S. thermophilus (3 × 10⁸ CFU). These values represent the estimated viable bacterial count per capsule at the time of manufacture. In addition, each probiotic capsule contained 38.5 mg of fructo-oligosaccharide (FOS) as a prebiotic component. The placebo capsules did not contain any live microorganisms or prebiotic compounds and were identical in appearance, weight, and packaging to ensure blinding. Participants were instructed to store the capsules in a refrigerator at 2–7 °C throughout the study period to maintain bacterial viability, as recommended by the manufacturer. The probiotic formulation was designed to ensure stability and viability under these storage conditions until the expiration date. The selected strains were chosen based on previous evidence supporting their anti-inflammatory and immunomodulatory properties in respiratory and systemic inflammatory conditions [5, 9, 10]. To monitor adherence, participants were contacted weekly to encourage compliance and to document any adverse events. At follow-up visits, unused capsules were collected, and adherence was calculated based on capsule counts. Compliance above 80% was considered acceptable. Self-reported adherence was also recorded.

Outcomes

The primary outcomes were changes in pulmonary function and clinical symptom scores after 8 weeks of intervention. Pulmonary function tests (PFTs), including FEV1, FVC, and FEV1/FVC ratio, were measured using a calibrated spirometer in accordance with American Thoracic Society/European Respiratory Society (ATS/ERS) guidelines [16]. Symptom burden and health status were assessed using the COPD Assessment Test (CAT) [17] and the modified Medical Research Council (mMRC) dyspnea scale [18]. Cough frequency and dyspnea severity categories were derived from individual CAT components and mMRC grading, respectively. Secondary outcomes included changes in plasma concentrations of cytokines, including IL-6, IL-17, and TGF-β. Blood samples were collected at baseline and at the end of the intervention period.

Cytokine measurement by ELISA

Peripheral venous blood (5 mL) was collected and centrifuged at 1500 × g for 10 min to separate serum. Samples were aliquoted and stored at − 80 °C until analysis. Serum concentrations of IL-6, IL-17, and TGF-β were measured using commercially available ELISA kits (KPG, Iran) according to the manufacturer’s instructions. Briefly, serum samples or recombinant standards were added to pre-coated microplates. After incubation for 2 h, detection antibodies were applied, followed by avidin-HRP conjugate. Tetramethylbenzidine (TMB) substrate was added, and the reaction was stopped using 1 N sulfuric acid. Optical density was measured at 450 nm using a microplate reader. Cytokine concentrations were determined using standard curves generated from known recombinant cytokine standards.

Statistical analysis

Data are presented as mean ± standard error of the mean (SEM). Normality of distribution was evaluated using the Kolmogorov–Smirnov test. Comparative analyses were conducted on a per-protocol basis, including only participants who completed the intervention period and had available outcome data at both baseline and week 8. For normally distributed variables, paired t-tests were used to assess within-group changes, and independent t-tests were applied for between-group comparisons. For non-normally distributed variables, Wilcoxon signed-rank tests and Mann–Whitney U tests were used as appropriate. Correlation analyses between pulmonary function parameters and clinical symptom scores were performed using Pearson correlation coefficients. Sensitivity analyses were conducted to evaluate the influence of potential outliers on correlation results. All statistical tests were two-sided. Given the exploratory nature of the study and the limited sample size, no formal correction for multiple comparisons (e.g., Bonferroni adjustment) was applied. Therefore, the findings should be interpreted as hypothesis-generating. Due to the exploratory design of the study and the small number of dropouts, analyses were conducted on a per-protocol basis. A p-value < 0.05 was considered statistically significant. Statistical analyses were performed using SPSS software (version 25.0; IBM Corp., Armonk, NY, USA) and GraphPad Prism (version 8.0, GraphPad Software, USA).

Results

Baseline characteristics of participants

A total of 50 patients with mild to moderate COPD were enrolled and randomly assigned to either the probiotic or placebo group (Fig. 1). Six participants did not complete the trial: two in the probiotic group did not receive the intervention, and four in the placebo group were excluded (one did not receive the intervention and three were lost to follow-up due to withdrawal of consent or inability to attend follow-up visits). Consequently, 44 participants (23 in the probiotic group and 21 in the placebo group) were included in the final per-protocol analysis. Among the analyzed participants, 19 were classified as GOLD stage I (mild) and 25 as GOLD stage II (moderate). Due to the limited sample size, subgroup analyses based on disease severity were not performed. The two groups were well matched in demographic and clinical characteristics at baseline (Table 1), with no statistically significant differences observed.

Fig. 1.

Flow diagram of patient enrolment, allocation, follow-up, and analysis in the randomized clinical trial. A total of 50 patients were randomized into probiotic (n = 25) and placebo (n = 25) groups. In the probiotic group, 23 patients received the intervention and were included in the analysis (2 did not receive intervention). In the placebo group, 24 patients received the placebo and 21 were analyzed (1 did not receive intervention and 3 were lost to follow-up)

Table 1.

Baseline demographic and clinical characteristics of patients with mild-to-moderate COPD in the probiotic and placebo groups

Variables (%)	Probiotic group (n = 23)	Placebo group (n = 21)	p-value
Age (mean ± SD)	62.5 ± 13.8	63.1 ± 13.9	> 0.05*
Sex			> 0.05
Male	15 (65.2%)	15 (71.4%)
Female	8 (34.8%)	7 (33.3%)†
Cough frequency			> 0.05
Mild	2 (8.7%)	1 (4.8%)
Moderate	15 (65.2%)	16 (76.2%)
Severe	6 (26.1%)	4 (19.0%)
Phlegm production			> 0.05
Mild	9 (39.1%)	10 (47.6%)
Moderate	9 (39.1%)	9 (42.9%)
Severe	5 (21.7%)	2 (9.5%)
Chest tightness severity			> 0.05
Mild	3 (13.0%)	3 (14.3%)
Moderate	10 (43.5%)	15 (71.4%)
Severe	10 (43.5%)	3 (14.3%)
Dyspnea severity			> 0.05
Mild	8 (34.8%)	2 (9.5%)
Moderate	12 (52.2%)	15 (71.4%)
Severe	3 (13.0%)	4 (19.0%)
Limitations in daily activities			> 0.05
Mild	11 (47.8%)	8 (38.1%)
Moderate	9 (39.1%)	10 (47.6%)
Severe	3 (13.0%)	3 (14.3%)
Sleep quality			> 0.05
Mild	12 (52.2%)	15 (71.4%)
Moderate	5 (21.7%)	4 (19.0%)
Severe	6 (26.1%)	2 (9.5%)
Energy levels			> 0.05
Mild	10 (43.5%)	11 (52.4%)
Moderate	11 (47.8%)	10 (47.6%)
Severe	2 (8.7%)	0 (0.0%)

*P-value for Age calculated using Independent Samples T-test. All other P-values calculated using Chi-Square test

Effects of probiotic supplementation on pulmonary function

Spirometry was performed at baseline and after eight weeks of intervention to assess pulmonary function. In the probiotic group, FEV1 increased significantly from 1.77 ± 0.73 L to 1.94 ± 0.92 L, representing a mean increase of 0.17 L (paired t-test, p = 0.008). FVC also increased from 2.40 ± 0.74 L to 2.74 ± 1.14 L (mean change: +0.34 L; p = 0.02). However, the FEV1/FVC ratio did not change significantly (p > 0.05).

In the placebo group, no significant within-group changes were observed in FEV1, FVC, or FEV1/FVC ratio (all p > 0.05). Between-group comparisons showed no significant differences at baseline or at follow-up for any spirometric parameter (all p > 0.05). Importantly, between-group comparisons of the mean changes (ΔFEV1 and ΔFVC) from baseline also did not reach statistical significance (Table 2).

Table 2.

Within- and between-group comparisons of spirometric parameters before and after probiotic supplementation in patients with COPD

Variable	Probiotic	Placebo	Between-group p-value
FEV1 (L)
Baseline	1.77 ± 0.73	1.87 ± 0.51	0.46
Follow-up	1.94 ± 0.92	1.89 ± 0.49	0.78
Within-group p-value	0.008	0.18
Mean change (Δ)	+ 0.17 ± 0.40	+ 0.02 ± 0.09	0.10
Effect size (Cohen’s d for Δ)			0.51
FVC (L)
Baseline	2.40 ± 0.74	2.78 ± 0.57	0.08
Follow-up	2.74 ± 1.14	2.91 ± 0.62	0.57
Within-group p-value	0.02	0.09
Mean change (Δ)	+ 0.34 ± 0.63	+ 0.12 ± 0.29	0.14
Effect size (Cohen’s d for Δ)			0.44
FEV1/FVC (%)
Baseline	64.05 ± 8.63	63.24 ± 8.92	0.76
Follow-up	64.18 ± 7.32	64.32 ± 7.90	0.72
Within-group p-value	0.87	0.33
Mean change (Δ)	+ 0.14 ± 4.12	+ 1.38 ± 4.12	0.32
Effect size (Cohen’s d for Δ)			0.30

Values are presented as mean ± SD

Between-group comparisons were performed using independent t-test or Mann–Whitney test

Within-group comparisons were performed using paired t-test or Wilcoxon signed-rank test

FEV1 forced expiratory volume in one second, FVC forced vital capacity

Impact of probiotics on Th17-related cytokines

Serum cytokine profiles were assessed to evaluate the immunomodulatory effects of probiotics. Distribution analysis using the Kolmogorov–Smirnov test confirmed normality of cytokine data; accordingly, parametric statistical tests were applied. As illustrated in Fig. 2A, patients receiving probiotics demonstrated a significant reduction in IL-6 levels at follow-up compared with baseline (p = 0.007), whereas no significant change was observed in the placebo group. Accordingly, the mean change in IL-6 (ΔIL-6) was − 19.6 ± 23.4 pg/mL in the probiotic group compared with + 2.8 ± 21.1 pg/mL in the placebo group. Between-group comparison of ΔIL-6 values (Fig. 2A) revealed a significantly greater reduction in the probiotic group (mean difference: −22.4 pg/mL; p = 0.02), with a large effect size (Cohen’s d = 0.99).

Fig. 2.

Serum cytokine levels in COPD patients before and after 8 weeks of intervention with probiotics or placebo. A IL-6 levels significantly decreased in the probiotic group compared to baseline and compared to the placebo group post-intervention. B IL-17 levels showed no significant changes within or between groups. C TGF-β levels remained unchanged following intervention. Data are presented as mean ± SEM. Within-group comparisons were performed using paired t-test; between-group comparisons were conducted using independent t-test. ** p < 0.01

In contrast, IL-17 and TGF-β levels showed no significant differences at the end of the intervention (p > 0.05). In the placebo group, no significant differences in IL-6, IL-17, or TGF-β concentrations were noted between baseline and follow-up.

Effects of probiotic supplementation on CAT and mMRC dyspnea scores

In the probiotic group, the CAT score significantly decreased from 18.68 ± 7.16 at baseline to 12.73 ± 6.28 after intervention, representing a mean reduction of 5.95 points (paired t-test, p < 0.0001). This improvement exceeded the established minimal clinically important difference (MCID) of 2 points. In the placebo group, the CAT score changed from 17.33 ± 5.81 to 15.95 ± 5.78, which was not statistically significant (p > 0.05).

The mean reduction in CAT score was − 5.95 ± 4.8 in the probiotic group compared with − 1.38 ± 4.3 in the placebo group. The between-group comparison of ΔCAT scores demonstrated a statistically significant difference (p < 0.0001), corresponding to a large effect size (Cohen’s d = 1.31).

No statistically significant difference was observed in CAT scores between the groups at baseline or after follow-up (p > 0.05 for both comparisons).

The mMRC dyspnea score did not show significant within-group changes in either the probiotic group (2.22 ± 1.06 to 2.13 ± 1.08; p > 0.05) or the placebo group (2.23 ± 0.83 to 2.21 ± 0.85; p > 0.05). No significant between-group differences were observed at baseline or follow-up (p > 0.05) (Table 3).

Table 3.

Within- and between-group comparisons of CAT and mMRC scores before and after probiotic supplementation in patients with COPD

Variable	Probiotic	Placebo	Between-group p-value
CAT score
Baseline	18.68 ± 7.16	17.33 ± 5.81	0.67
Follow-up	12.73 ± 6.28	15.95 ± 5.78	0.053
Within-group p-value	< 0.0001	0.08
Mean change (Δ)	−5.95 ± 4.47	−1.11 ± 2.54	< 0.0001
Effect size (Cohen’s d for Δ)			1.31
mMRC score
Baseline	2.22 ± 1.06	2.23 ± 0.83	0.97
Follow-up	2.13 ± 1.08	2.21 ± 0.85	0.96
Within-group p-value	0.32	0.33
Mean change (Δ)	−0.095 ± 0.44	0.02 ± 0.01	0.23
Effect size (Cohen’s d for Δ)			0.31

Values are presented as mean ± SD

Between-group comparisons were performed using independent t-test or Mann–Whitney test

Within-group comparisons were performed using paired t-test or Wilcoxon signed-rank test

CAT COPD Assessment Test, mMRC modified Medical Research Council

Safety and tolerability

Probiotic supplementation was well tolerated. Based on capsule count and weekly follow-up contacts, adherence was considered acceptable (> 80%) in the majority of participants in both groups, with no significant difference between groups. No serious adverse events were reported during the 8-week intervention. Mild gastrointestinal complaints were infrequent and comparable between groups. No participants discontinued the study due to adverse effects.

Interrelationship between pulmonary function parameters and clinical symptom scores

Correlation analysis demonstrated significant inverse associations between CAT scores and pulmonary function parameters. Higher CAT scores, reflecting greater symptom burden, were significantly associated with poorer lung function. Specifically, CAT scores showed a moderate negative correlation with the FEV1/FVC ratio (r = -0.40, p = 0.007; Fig. 3A) and with post-bronchodilator FEV1 (r = -0.42, p = 0.007; Fig. 3B). A similar association was observed with FVC (r = -0.34, p = 0.03; Fig. 3C).

Fig. 3.

Correlations between pulmonary function parameters and CAT score in patients with mild-to-moderate COPD. A Negative correlation between CAT score and FEV1/FVC ratio (r = -0.40, p = 0.007). B Negative correlation between CAT score and FEV1 (r = -0.42, p = 0.007). C Negative correlation between CAT score and FVC (r = -0.34, p = 0.03). D Positive correlation between FEV1/FVC ratio and FEV1 (r = 0.63, p < 0.0001). E Positive correlation between FEV1 and FVC (r = 0.79, p < 0.0001). Pearson correlation coefficients (r) and corresponding p-values are shown in each panel

As expected, positive correlations were confirmed among core spirometric indices. The FEV1/FVC ratio was positively correlated with FEV1 (r = 0.63, p < 0.0001; Fig. 3D), and FEV1 was strongly correlated with FVC (r = 0.79, p < 0.0001; Fig. 3E), demonstrating internal consistency and reliability of the pulmonary function measurements.

Discussion

COPD remains a major global health challenge, marked by progressive airflow limitation, persistent respiratory symptoms, and recurrent exacerbations. The underlying pathology is strongly influenced by chronic inflammation, immune dysregulation, and structural damage to the lungs [19]. Over the last decade, growing attention has been directed toward the role of gut–lung axis interactions in respiratory diseases, with probiotics emerging as potential modulators of immune balance and systemic inflammation [20]. Although probiotics are hypothesized to exert effects through the gut–lung axis, this study did not directly assess microbiome composition. Therefore, mechanistic conclusions regarding gut–lung crosstalk cannot be drawn. In this randomized clinical trial, we examined whether an eight-week probiotic intervention could improve clinical outcomes and modify key immune markers in patients with mild to moderate COPD. Although this exact multi-strain formulation has not been previously evaluated in COPD, several of its individual components have demonstrated anti-inflammatory effects in prior experimental and clinical studies [5, 9, 10]. Our findings indicate modest improvements in selected pulmonary function parameters and health status, accompanied by a significant reduction in circulating IL-6 levels, suggesting attenuation of systemic inflammatory activity. The pathogenesis of COPD involves a complex network of inflammatory cells and mediators. Among these, Th17 cells and their associated cytokines, particularly IL-17, IL-6, and TGF-β, play crucial roles in sustaining neutrophilic airway inflammation and driving tissue remodelling [21]. Elevated IL-17 levels have been linked to worse airflow obstruction and more frequent exacerbations, while IL-6 participates in systemic inflammatory signaling and is involved in pathways associated with Th17 differentiation. TGF-β, although traditionally considered anti-inflammatory, may also contribute to airway remodeling in chronic disease settings [2].

Our results demonstrated a significant reduction in IL-6 levels following probiotic supplementation, while IL-17 and TGF-β remained largely unchanged. The reduction in IL-6 indicates a decrease in systemic inflammatory activity; however, the underlying biological mechanisms were not directly investigated in this study [22]. Similar reductions in IL-6 have been reported in studies evaluating probiotic supplementation in asthmatic patients, particularly with Lactobacillus-containing formulations, supporting the plausibility of our findings [5]. Given the established role of IL-6 in inflammatory signaling pathways, this reduction may be consistent with pathways involved in systemic inflammation; however, its relevance to Th17 biology cannot be determined from the present data. This is consistent with preclinical studies showing that probiotics can suppress pro-inflammatory cytokine production and restore immune homeostasis by influencing gut microbial composition and associated metabolites [23]. Importantly, it remains unclear whether the observed reduction in IL-6 is linked to downstream Th17 responses, as these were not evaluated in this study.

Although we did not detect significant changes in IL-17 or TGF-β levels after intervention, the reduction in IL-6 may represent an early change in inflammatory signaling; however, no conclusions can be drawn regarding Th17 differentiation. It is possible that eight weeks was insufficient to alter downstream effectors like IL-17, or that serum levels do not reflect local concentrations in lung tissue or sputum. Still, suppressing IL-6 could interrupt the inflammatory loop that perpetuates airway damage in COPD. Some probiotic strains, such as Lactobacillus and Bifidobacterium species, have been shown to inhibit NF-κB signaling and reduce IL-6 production in intestinal and immune cells—effects that may extend systemically via the gut–lung axis [5, 24]. IL-6 signaling is closely associated with activation of the STAT3 pathway, a key regulator of Th17 differentiation and chronic inflammatory responses. Previous studies have suggested that probiotics may influence inflammatory pathways such as NF-κB and STAT3 signaling [25]. However, these mechanisms were not assessed in the present study, and therefore their involvement remains speculative.

In addition, probiotic-derived short-chain fatty acids (SCFAs), such as butyrate and propionate, have been shown to modulate dendritic cell function and suppress excessive Th17 polarization through epigenetic and metabolic reprogramming, providing a plausible gut–lung axis mechanism linking IL-6 reduction to downstream immune regulation [26].

Spirometry remains the cornerstone of COPD evaluation, providing objective measures of airflow limitation and disease severity [27]. In our study, patients receiving probiotics demonstrated significant within-group improvements in FEV1 and FVC, whereas no significant changes were observed in the placebo group. However, between-group differences did not reach statistical significance, indicating that these findings should be interpreted with caution. Although the FEV1/FVC ratio did not change significantly, its directional trend was consistent with improvements in absolute lung volumes, suggesting a possible early physiological effect that may require longer follow-up to become statistically detectable. Overall, while these trends are encouraging, they do not constitute definitive evidence of a clinically meaningful spirometric benefit [28].

Several mechanisms may explain these improvements. By reducing systemic inflammation, probiotics may alleviate airway wall edema and neutrophil-mediated tissue damage, leading to better airflow and volume expansion [29]. In addition, probiotics have been reported to enhance epithelial barrier integrity and modulate local immune responses within the lung, potentially contributing to the stabilization of pulmonary function [30].

The gut–lung axis has been proposed as a potential pathway linking intestinal microbiota to respiratory health; however, this study did not investigate this mechanism. The gut microbiota influences systemic immunity through microbial metabolites SCFAs, modulation of dendritic cells, and regulation of T-cell polarization [26]. Probiotics may enhance gut barrier integrity, reduce bacterial translocation, and promote anti-inflammatory immune responses. In animal models of COPD, certain probiotic strains have reduced lung inflammation, decreased neutrophil infiltration, and improved histology [31]. Human studies remain limited, but our results add to a growing body of evidence suggesting that gut-directed interventions can influence lung health.

Beyond physiological measurements, COPD management increasingly emphasizes patient-reported outcomes, which provide critical insights into symptom burden and quality of life [32]. Our trial revealed a striking reduction in CAT scores among probiotic recipients, indicating an improvement in overall health status. This improvement occurred in parallel with reductions in IL-6, which may be associated with concurrent reductions in systemic inflammation; however, causal relationships cannot be established.

Interestingly, no significant change was observed in the mMRC dyspnea scale. This may reflect the narrower focus of mMRC on exertional breathlessness, whereas CAT captures a broader spectrum of COPD-related symptoms, including cough, sputum production, chest tightness, and sleep disturbance [33]. The greater sensitivity of CAT to multidimensional symptom changes likely explains why improvements were more readily detected with this instrument. Collectively, these findings suggest that probiotic supplementation may preferentially improve overall symptom burden and health status rather than exerting an isolated effect on dyspnea severity alone. Notably, the strong negative correlation between CAT scores and spirometric indices further supports its clinical relevance, as it reflects both physiological impairment and patient-reported symptom experience.

A strength of the present study is the integrated evaluation of clinical and functional outcomes. We observed consistent associations between patient-reported symptom burden and objective pulmonary function parameters. Specifically, higher CAT scores were significantly associated with lower FEV1, FVC, and FEV1/FVC ratios, highlighting the close relationship between airflow limitation and perceived health status in COPD. These findings reinforce the clinical relevance of spirometric impairment as a determinant of symptom severity and quality of life.

While IL-6 levels decreased following probiotic supplementation, no significant changes were observed in IL-17 or TGF-β concentrations. Therefore, the immunomodulatory effect observed in this study appears to be predominantly related to modulation of systemic inflammation rather than direct alteration of the Th17 cytokine axis [34, 35].

Our results provide preliminary evidence supporting probiotics as a safe, accessible, and potentially effective adjunctive therapy for COPD. By attenuating systemic inflammation and improving patient-reported health status, probiotics could complement pharmacological strategies aimed at bronchodilation and anti-inflammatory control [36]. Importantly, probiotics may offer a cost-effective intervention with minimal side effects, particularly relevant in resource-limited settings where access to advanced therapies remains restricted.

While the magnitude of lung function improvement was modest, the significant enhancement in quality of life underscores the potential clinical value of probiotics. COPD management is not solely about preventing decline in FEV1; improving day-to-day symptom burden and well-being is equally important. In this respect, probiotics may fill an important therapeutic gap by addressing systemic inflammation and its impact on patient experience [37].

However, several limitations must be acknowledged. This study was designed as an exploratory pilot trial with a modest sample size, which may have limited the statistical power to detect small but potentially meaningful between-group differences in spirometric outcomes. The intervention period of eight weeks may have been insufficient to capture long-term effects on cytokine dynamics and structural lung remodeling, and we did not assess gut microbiota composition, limiting mechanistic insight into gut–lung axis modulation. Moreover, serum cytokine concentrations may not fully reflect local pulmonary inflammation, highlighting the need for future studies incorporating induced sputum or bronchoalveolar lavage analyses. The use of per-protocol analysis without intention-to-treat analysis represents a limitation, as it may introduce bias and affect the generalizability of the findings. Finally, the study population included only patients with mild to moderate COPD, and findings may not be generalizable to those with advanced disease or frequent exacerbations.

Future studies should expand upon these findings by conducting larger, longer-duration randomized clinical trials with adequate statistical power, ideally incorporating microbiome sequencing and metabolomic profiling. Such investigations would allow deeper mechanistic understanding of host–microbiome interactions in COPD and help identify biomarkers predicting response to probiotic therapy. In addition, exploring strain-specific probiotic effects and optimal dosing regimens will be essential for translating this approach into clinical practice.

Despite its exploratory design, the present study integrates clinical, functional, and immunological assessments within a randomized, double-blind, placebo-controlled framework in patients with mild-to-moderate COPD. By concurrently evaluating spirometry, patient-reported symptom burden, and systemic inflammatory markers, this work provides translational data that may inform the design of larger confirmatory trials.

Conclusion

In conclusion, probiotic supplementation in patients with mild-to-moderate COPD was associated with a significant reduction in circulating IL-6 levels and a clinically meaningful improvement in CAT scores that exceeded the established minimal clinically important difference. Improvements in FEV1 and FVC were observed in the probiotic group over the intervention period, whereas no comparable changes were detected in the placebo group. Although between-group differences in spirometric indices did not reach statistical significance, the observed trends alongside significant IL-6 suppression and improved CAT scores suggest a biologically plausible and clinically relevant adjunctive effect. Overall, the findings indicate that probiotic supplementation may provide anti-inflammatory and symptomatic benefits in patients with COPD and support its consideration as a complementary therapeutic approach. Further large-scale and mechanistically oriented clinical trials are warranted to confirm these effects and clarify the underlying pathways involved.

Authors’ contributions

Rasoul Baharlou and Mohammad Memarian conceived and planned the experiments. Sharareh Ebrahimi and Samaneh Mohammadi carried out the experiments. Sharareh Ebrahimi wrote the manuscript. Sharareh Ebrahimi contributed to sample preparation. Rasoul Baharlou and Mohammad Memarian and Sharareh Ebrahimi contributed to the interpretation of the results. All authors provided critical feedback and helped shape the research, analysis and manuscript.

Funding

This work was supported by a grant from Semnan University of Medical Sciences.

Data availability

All data generated or analyzed during this study are included in this article. Further inquiries can be directed to the corresponding author. The study was registered on the Iran Clinical Trials website at: ( https://irct.behdasht.gov.ir: IRCT20241211064025N1).

Declarations

Ethics approval and consent to participate

The study complied with the guidelines for human studies and was conducted in an accordance with the World Medical Association Declaration of Helsinki. The study protocol was approved by the Ethical Board of the Semnan University of Medical Sciences (IR.SEMUMS.REC.1403.202). Written informed consent to participate in the study and publication of their clinical details was obtained from the patient.

Competing interests

The authors declare no competing interests.