Effects of Dietary Copper Deficiency on Colonic Barrier Integrity, Inflammatory Markers, and Gut Microbiota Composition in Mice

Introduction: This study sought to explore the impact of dietary Cu deficiency on colonic health, including assessments of histopathology, barrier function, inflammatory response, and gut microbiota composition. Methods: Weaned mice were fed a copper-deficient diet for four weeks, followed by one week of intraperitoneal copper sulfate administration as a proof-of-concept rescue intervention. Colonic pathology was assessed by H&E staining, goblet cell changes by AB-PAS staining, and intestinal barrier integrity by immunofluorescence. Inflammatory cytokine levels were measured by ELISA, while protein and mRNA expression of inflammatory markers were detected by Western blot and qRT-PCR. Gut microbiota composition, diversity, and signature genus abundance were analyzed by 16S sequencing. Results: Compared to the control group, CuD mice exhibited histopathological damage in the colon, including mucosal thinning and inflammatory cell infiltration. The number of goblet cells and the expression of mucin MUC2 were significantly reduced, and the expression of tight junction proteins (ZO-1, Occludin) was downregulated, indicating impairment of both the physical and chemical intestinal barriers. Concurrently, Cu deficiency markedly elevated systemic and colonic levels of pro-inflammatory cytokines (TNF-α, IL-1β, and IL-6), and enhanced NF-κB phosphorylation. To explore potential microbial contributions to these colonic alterations, we subsequently analyzed the gut microbiota composition by 16S rRNA sequencing. This analysis revealed that Cu deficiency significantly reduced the α-diversity and species richness of the gut microbiota. This dysbiosis was characterized by a decreased abundance of beneficial bacteria (e.g., Bacteroidota, Muribaculaceae) and an increased abundance of Desulfobacterota, a pro-inflammatory taxon, as well as Akkermansia, a mucin-degrading bacterium with context-dependent effects on gut health. Intraperitoneal administration of copper sulfate (CuD + CuSO4) partially reversed the histopathological and inflammatory changes; its effect on the gut microbiota was not assessed. Conclusions: Dietary Cu deficiency is associated with colonic injury, and these alterations were accompanied by intestinal barrier disruption, an activated inflammatory response, and gut microbiota dysbiosis. These findings provide experimental evidence highlighting the importance of copper nutrition in maintaining colonic homeostasis, though further mechanistic studies are needed to establish causal relationships.