Limosilactobacillus johnsoni and Limosilactobacillus mucosae and Their Extracellular Vesicles Alleviate Gut Inflammatory Injury by Mediating Macrophage Polarization in a Lipopolysaccharide-Challenged Piglet Model

Abstract

Background: Limosilactobacillus johnsoni (L. j) and Limosilactobacillus mucosae (L. m) can alleviate the inflammatory response.

Objectives: This study aimed to elucidate the underlying mechanisms by which L. j- and L. m-derived extracellular vesicles (EVs) mitigate lipopolysaccharide (LPS)-induced intestinal injury.

Methods: Piglets were assigned to 4 groups: oral phosphate-buffered saline inoculation for 2 wk prior to intraperitoneal injection of physiological saline or LPS, and oral L. j/L. m inoculation for 2 wk prior to intraperitoneal injection of LPS. The intestinal integrity, macrophage markers, cytokine levels, and microbiota were determined. The cytokine levels and macrophage phenotype were detected after L. j/L. m and their EVs were coincubated with macrophages. The levels of cytokines, tight junction proteins, and apoptosis were measured after intestinal epithelial cells were cocultured with macrophages.

Results: LPS challenge decreased jejunal villus length; expression levels of zonula occludens-1 (ZO-1), occludin, arginase-1 (Arg1), and interleukin (IL)-10; and number of CD163⁺ cells and increased the expression levels of inducible nitric oxide synthase (iNOS), IL-1β, IL-6, and tumor necrosis factor (TNF)-α compared with that in the control. L. j and L. m pretreatment rescued the aforementioned indicators compared with LPS challenge. Pretreatment of L. j and L. m and their EVs reversed the levels of IL-1β, IL-6, TNF-α, and IL-10 and the gene expression of iNOS and Arg1 in the LPS group in macrophages. Pretreatment with L. j and L. m-derived EVs increased ZO-1 and occludin mRNA expression and reduced IL-1β, caspase-3, and bax gene expression in intestinal epithelial cells of the coculture system. Enzyme-treated EVs were less effective than native EVs.

Conclusions: This study suggests that EVs secreted by L. j and L. m control inflammation by modulating macrophage polarization, thereby improving intestinal barrier function.

Keywords: Limosilactobacillus johnsoni; Limosilactobacillus mucosae; intestinal epithelial cells; macrophage; piglet model.