Objective: Non-alcoholic steatohepatitis (NASH) is characterized by a robust pro-inflammatory component at both hepatic and systemic levels together with a disease-specific gut microbiome signature. Protein tyrosine phosphatase 1B (PTP1B) plays distinct roles in non-immune and immune cells, in the latter inhibiting pro-inflammatory signaling cascades. In this study we have explored the role of PTP1B in the composition of gut microbiota and gut barrier dynamics in methionine and choline-deficient (MCD) diet-induced NASH in mice. Methods: Gut features and barrier permeability were characterized in wild-type (PTP1B WT) and PTP1B-deficient (PTP1B KO) mice fed a chow or methionine/choline-deficient (MCD) diet during 4 weeks. The impact of inflammation was studied in intestinal epithelial and enteroendocrine cells. Secretion of GLP-1 was evaluated in primary colonic cultures and plasma of mice. Results: We found that a shift in the gut microbiota shape, disruption of gut barrier function, higher levels of serum bile acids and decreased circulating glucagon-like peptide (GLP)-1 are features during NASH. Surprisingly, despite the pro-inflammatory phenotype of global PTP1B deficient mice, they were partly protected against the alterations in gut microbiota composition during NASH and presented better gut barrier integrity and less permeability under this pathological condition. These effects concurred with higher colonic mucosal inflammation, decreased serum bile acids, protection against the drop of circulating GLP-1 levels during NASH in comparison to their wild-type counterparts together with increased expression of GLP-2-sensitive genes in the gut. At the molecular level, stimulation of enteroendocrine STC-1 cells with a pro-inflammatory conditioned medium (CM) from lipopolysaccharide (LPS)- stimulated macrophages triggered pro-inflammatory signaling cascades that were further exacerbated by a PTP1B inhibitor. Likewise, the pro-inflammatory CM induced GLP-1 secretion in primary colonic cultures, an effect augmented by PTP1B inhibition. Conclusion: Altogether our results have unraveled a potential role of PTP1B in the gut-liver axis during NASH, likely mediated by increased sensitivity to GLPs, with potential therapeutic value.