Effect of the Probiotic Mixture VSL#3 on Macrophage and Proliferating Cell Numbers in the Liver of Rats Undergoing Acute Colitis
Authors: Gonzalez Torres Mayra I. , Isidro Raymond A. , López Abdon , Vázquez Abner E. , Cruz Myrella L. , Isidro Angel A. , Appleyard Caroline B. 
Published: November 2023

IntroductionPatients with Inflammatory Bowel Disease are vulnerable to developing extra‐intestinal disorders, usually in the liver and biliary system. Macrophages have been shown to accumulate in the liver in several animal models of colitis. The probiotic mixture VSL#3, which consists of 8 bacterial strains, has been suggested to exert its beneficial effects in part by decreasing proliferation. As with most tissue‐resident macrophages, Kupffer cells (hepatic macrophages) are thought to have proliferative capacity. Our aim was to examine the effect of the probiotic mixture VSL#3 on the liver of rats undergoing acute colitis.MethodsMale Sprague Dawley rats were randomly assigned to normal (n=5), colitis (n=9) or colitis+VSL#3 (n=12) groups. For a week prior to colitis induction and until the time of sacrifice, colitis+VSL#3 rats were treated daily with 10 billion CFU of the probiotic in drinking water. Control rats received water during this period. Colitis was induced by intracolonic administration of trinitrobenzene sulfonic acid (0.5 mL, 60 mg/mL). Rats were euthanized three days after colitis induction. At the time of sacrifice liver samples from each rat were collected. Hematoxylin and eosin staining of liver tissue was used to assess histology. Macrophage numbers and cell proliferation was assessed using double immunofluorescence staining of formalin‐fixed paraffin‐embedded liver tissue for CD68 and Ki67. Total macrophages (CD68+), proliferating cells (Ki67+) and proliferating macrophages (CD68+Ki67+) were quantified in the hepatic parenchyma surrounding the central vein and the portal areas of each sample.ResultsTissue from rats undergoing colitis demonstrated inflammatory infiltrates in the portal areas, sinusoids, and hepatic parenchyma. Colitis rats predominantly contained mild portal infiltrates and patchy but diffuse sinusoidal infiltrates, with occasional neutrophilic infiltration in the parenchyma. Colitis+VSL#3 rats mostly displayed inflammatory infiltration of the larger portal areas with mixed neutrophilic and mononuclear infiltration in the sinusoids. The total number of macrophages (CD68+) in proximity to the central veins and the portal areas was significantly higher in colitis (p<0.01) and colitis+VSL#3 (p<0.05) rats when compared to normals. The majority of these macrophages were negative for Ki67. The percentage of macrophages that were Ki67+ was not substantially different between the three groups. Normal, colitis, and colitis+VSL#3 rats contained similar numbers of total Ki67+ cells and of Ki67+CD68+ cells. A significantly (p<0.05) higher percentage of proliferating cells (Ki67+) were macrophages (CD68+Ki67+) in the areas surrounding the central vein, but not in the portal areas, of the colitis and colitis+VSL#3 groups when compared to the normal group.ConclusionIn agreement with previous studies, colitis led to macrophage accumulation in the liver. However, the probiotic VSL#3 did not significantly affect this accumulation in our acute colitis model. Neither colitis alone nor colitis in conjunction with probiotic treatment significantly altered the prevalence of proliferating cells in the liver. Further studies are needed to better understand the effect of intestinal inflammation on liver physiology.Support or Funding InformationR25GM096955, R25GM082406 & APS Porter Fellowship