Effects of Probiotics and Prebiotics on Intestinal Microbiota in Acute Colitis Mice

0
0
0

Effects of Probiotics and Prebiotics on Intestinal Microbiota in Mice with Acute Colitis Based on 16S rRNA Gene Sequencing

Introduction

The gut microbiota constitutes the largest microbial ecosystem in the human body, forming a dynamic equilibrium with the host to maintain intestinal structure, immune homeostasis, and resistance against pathogens. Dysbiosis of this ecosystem is closely linked to inflammatory bowel diseases (IBD), including ulcerative colitis (UC). Acute colitis models induced by dextran sulfate sodium (DSS) mimic human UC symptoms, providing a platform to study therapeutic interventions. Probiotics (e.g., Bifidobacterium and Lactobacillus) and prebiotics (e.g., lactitol and inulin) have emerged as potential modulators of gut microbiota, offering alternatives to traditional anti-inflammatory drugs with fewer side effects. This study investigates the effects of probiotics, prebiotics, and their combinations on inflammation and microbial composition in DSS-induced acute colitis mice using 16S rRNA sequencing.

Methods

Experimental Design

Sixty male C57BL/6J mice (6–8 weeks old) were divided into six groups:

  1. Blank control: No DSS, no intervention.
  2. Model control: 2.5% DSS for 5 days, no intervention.
  3. Probiotics: DSS + daily gavage of Lactobacillus acidophilus, L. rhamnosus, and Bifidobacterium lactis (1 × 10⁹ CFU/mouse).
  4. Synbiotics: DSS + probiotics (5 × 10⁸ CFU) combined with inulin.
  5. Lactitol: DSS + lactitol (6.6 g/kg/day).
  6. Probiotics + Lactitol: DSS + half-dose probiotics (5 × 10⁸ CFU) and lactitol (3.3 g/kg/day).

After 5 days of DSS exposure, interventions were administered for 7 days. Mice were sacrificed, and colonic tissues were collected for histopathological scoring. Fecal and mucosal samples were analyzed using 16S rRNA sequencing.

Histopathological Evaluation

Colonic inflammation was scored based on three parameters:

  1. Severity of inflammation (0–4: none to severe).
  2. Extent of injury (0–3: mucosal to transmural).
  3. Crypt damage (0–4: intact to complete loss).
    Scores were multiplied by the percentage of affected tissue (1–4× for 0–25% to 76–100% involvement).

Microbiota Analysis

DNA from fecal and mucosal samples was sequenced using the MiSeq PE300 platform. Alpha diversity (Observed Species index), principal component analysis (PCA), and taxonomic profiling were performed. Metastats analysis identified significant genus-level differences (abundance >1%, P < 0.05).

Results

Histopathological Outcomes

Intervention groups showed significantly lower histopathological scores compared to the model control group (P < 0.05, Table 1). No statistical differences were observed among intervention groups, indicating comparable anti-inflammatory efficacy.

Table 1: Histopathological Scores Group Score (Mean ± SD)
Blank control 0
Model control 27.00 ± 7.94
Probiotics 5.40 ± 2.79*
Synbiotics 7.25 ± 2.87*
Lactitol 7.20 ± 2.86*
Probiotics + Lactitol 9.33 ± 5.69*

Compared to model control (P* < 0.05).

Body Weight Changes

All DSS-exposed groups exhibited weight loss during inflammation. By day 7, blank control mice weighed significantly more than other groups (P = 0.0139), but no differences were noted among intervention groups.

Fecal Microbiota Dynamics

Baseline (W0 Feces)

No significant differences in alpha diversity or PCA clustering were observed, confirming uniform baseline microbiota.

Mid-Intervention (D4 Feces)

Alpha diversity decreased in all DSS groups vs. blank controls (P < 0.05). Key findings:

  • Model control: Reduced Lactobacillus (0.0020 vs. 0.0122, P = 0.0188) and increased Bacteroides (0.3519 vs. 0.0208, P = 0.0002).
  • Probiotics + Lactitol: Elevated Akkermansia (0.0404 vs. 0.0087, P = 0.0178) and Faecalibacterium (0.2854 vs. 0.0589, P = 0.0215) compared to model controls.

Post-Intervention (W1 Feces)

Alpha diversity remained lower in DSS groups. Notable changes:

  • Bacteroides abundance stayed elevated in DSS groups (e.g., model control: 0.3679 vs. blank: 0.0099, P = 0.0016).
  • Lachnospiraceae_NK4A136_group increased in intervention groups (probiotics: 0.2010 vs. model: 0.0320, P = 0.0352).
  • Akkermansia was higher in probiotics + lactitol vs. synbiotics (0.0215 vs. 0.0013, P = 0.0315).

Mucosal Microbiota

Mucosal samples exhibited higher alpha diversity than feces. Key differences:

  • Mucispirillum was mucosa-enriched (0.0207 vs. 0.0001 in feces, P = 0.0034).
  • Akkermansia abundance in lactitol group exceeded model control (0.0138 vs. 0.0055, P = 0.0415), probiotics (0.0138 vs. 0.0022, P = 0.0041), and synbiotics (0.0138 vs. 0.0011, P = 0.0034).
  • Faecalibacterium decreased in model control (0.0009 vs. blank: 0.0265, P = 0.0131).

Discussion

Anti-Inflammatory Effects

Probiotics and prebiotics significantly reduced colitis severity, aligning with prior studies showing Lactobacillus and Bifidobacterium mitigate inflammation via immune modulation and barrier reinforcement. Synergistic effects of combined therapies were not superior to individual treatments, possibly due to dose adjustments or lactitol’s broader microbial effects.

Microbial Shifts in Colitis

DSS-induced colitis mirrored human UC dysbiosis:

  • Pathobiont Expansion: Bacteroides increased, consistent with its association mucosal damage.
  • Beneficial Genera Depletion: Lactobacillus, Roseburia, and Faecalibacterium (butyrate producers) declined, impairing anti-inflammatory signaling and epithelial repair.

Lactitol’s Role in Promoting Akkermansia

Lactitol uniquely enriched Akkermansia, a mucin-degrading bacterium linked to improved metabolic health and barrier function. Its genome encodes enzymes that may metabolize lactitol, fostering colonization. Akkermansia’s rise correlates with reduced endotoxemia and inflammation, suggesting prebiotics targeting specific taxa could enhance colitis management.

Mucosal vs. Fecal Microbiota

Mucosal communities differed markedly from luminal flora, highlighting niche-specific microbial adaptations. Mucispirillum, a mucus colonizer, was mucosa-predominant, underscoring the importance of sampling site in microbiota studies.

Limitations and Future Directions

  • Taxonomic Resolution: Genus-level analysis limited species-specific insights.
  • Temporal Dynamics: Mucosal microbiota changes during intervention were not tracked.
  • Mechanistic Gaps: Causality between microbial shifts and inflammation requires metagenomic/metabolomic validation.

Conclusion

Probiotics and lactitol ameliorated DSS-induced colitis by restoring beneficial genera (Bifidobacterium, Lactobacillus) and suppressing pathobionts (Bacteroides). Lactitol specifically promoted Akkermansia, a promising therapeutic target. These findings underscore the potential of microbiota-targeted therapies in IBD, warranting clinical trials to validate translatability.

doi.org/10.1097/CM9.0000000000000308

Was this helpful?

0 / 0