The Probiotic Mixture VSL#3 Attenuates Colitis, Decreases Macrophage Infiltration, and Reduces Serum Cytokine Levels in an Acute Colitis Model
Authors: Isidro Raymond A. , López Abdón , Vázquez Abner E. , Cruz Myrella L. , Hernández Siomara , Cabán Josué , Isidro Angel A. , Appleyard Caroline B. 
Published: November 2023

Background

Our previous work has shown that the probiotic mixture VSL#3 alters the morphology and secretory profile of proinflammatory (M1), anti‐inflammatory (M2), and unpolarized macrophages in vitro. The effect of VSL#3 on macrophages in vivo is unclear. This knowledge may prove useful for developing and improving treatments for colitis in patients with Inflammatory Bowel Disease.

Aim

Examine the effect of VSL#3 administration on disease parameters, macrophage phenotype and infiltration, and serum cytokine levels in an acute colitis model.

Methods

Male rats were randomized to one of the following groups: a normal group (n=5), a colitis group (n=9), or a colitis+VSL#3 group (n=12). Rats in the colitis+VSL#3 group received 10 billion CFU of the probiotic in drinking water daily from a week prior to colitis induction until the time of sacrifice, whereas the control rats received water during this period. Rats underwent colitis induction by intracolonic administration of 0.5 mL of trinitrobenzene sulfonic acid (60 mg/mL) and were sacrificed 72 hours later. The colon of each rat was resected and evaluated for macroscopic and microscopic damage. Double immunofluorescence staining of colonic sections for CD68 and inducible nitric oxide synthase (iNOS) was used to assess total macrophage (CD68+) and M1 macrophage (CD68+iNOS+) numbers. Serum cytokine levels at the time of sacrifice were measured with a multiplex bead array.

Results

All of the colitis rats significantly lost weight when compared to normal rats (p<0.001), which gained weight. The probiotic did not have a significant effect on weight loss. All of the colitis rats had significant macroscopic (p<0.001) and microscopic damage (p<0.01) in the colon compared to normal rats as expected. Rats in the colitis+VSL#3 group had lower macroscopic damage scores overall (p<0.01), and lower microscopic damage scores in the proximal colon (p<0.01), when compared to rats in the colitis group. This improvement was not found in the mid or distal colon. M1 macrophages were mainly limited to areas of mucosa and submucosa that stained for iNOS. Within these areas, rats with colitis contained significantly higher percentages of M1 macrophages than normal rats (p<0.05), treatment with VSL#3 did not decrease the percentage of M1 macrophages in rats with colitis, and total macrophage numbers did not differ significantly between the three groups. Overall, colitis induction led to a significant increase in submucosal macrophage infiltration (p<0.01), and this infiltration was significantly reduced by treatment with VSL#3 (p<0.01). Mucosal macrophage infiltration was significantly higher in the colitis group when compared to normal controls (p<0.001) and the colitis+VSL#3 group (p<0.01). Colitis induction significantly increased serum levels of GRO/KC, MCP‐1, and VEGF. Serum levels of EGF, IL‐5, IL‐12p70, IL‐13, IL‐17A, IL‐18, MIP‐1α, and TNF‐α were significantly increased in the colitis group, but not the colitis+VSL#3 group, when compared to normal controls. Treatment with VSL#3 significantly reduced serum levels of EGF, eotaxin, IL‐2, and VEGF.

Conclusion

In this model of acute colitis, the probiotic VSL#3 reduces colitis severity, colonic macrophage infiltration, and serum cytokine and chemokine levels. Our results suggest that VSL#3 treatment does not dampen the proinflammatory phenotype of M1 macrophages, consistent with our previously reported in vitro results for human monocyte‐derived M1 macrophages.

Support or Funding Information

R25GM082406 & William Townsend Porter Predoctoral Fellowship from the American Physiological Society