Fig 2: LEV-mediated regulation of the SIRT5/p53 axis affects proliferation and glycolysis in intestinal epithelial cells, influencing colon tumor formation. Note: (A) Schematic diagram illustrating the construction of a mouse model of colitis-associated tumors; (B) co-immunoprecipitation (co-IP) experiment detecting the acetylation level of p53 in mouse colonic polyp tissues from each group; (C) statistical analysis of the number and burden of colonic polyps in each group of mice; (D) immunohistochemical staining detecting the positive expression of Ki67 protein in mouse colonic polyp tissues from each group (scale bar = 100 μm); (E) immunohistochemical staining detecting the positive expression of cell cycle-related proteins Cyclin D1 and p27 in mouse colonic polyp tissues from each group (scale bar = 100 μm); (F) glucose uptake in mouse colonic polyp tissues from each group; (G) lactate production in mouse colonic mucosal tissues from each group; (H) Western blot detecting the expression of glycolytic rate-limiting enzymes GLUT1 and HKII in mouse colonic mucosal tissues from each group. * represents a difference compared to the WT group (P < 0.05), # represents a difference compared to the LEVs + WT group (P < 0.05), 6 mice per group

Fig 3: The Effect of LEVs-Mediated SIRT5/p53 Axis on Proliferation and Glycolysis in Caco-2 Colorectal Cancer Cells. Note: (A) RT-qPCR detection of p53 knockdown efficiency; (B) Co-IP experiment to detect succinylation levels of p53 in different groups of Caco-2 cells; (C) EdU staining to detect proliferation of different groups of Caco-2 cells (scale bar = 25 μm); (D) Clonogenic assay to detect colony formation of different groups of Caco-2 cells; (E) Flow cytometry analysis to detect cell cycle changes in different groups of Caco-2 cells; (F) Western blot detection of expression changes of cell cycle-related proteins in different groups of Caco-2 cells; (G) Glucose uptake in different groups of Caco-2 cells; (H) Lactate generation in different groups of Caco-2 cells; (I) Western blot detection of expression of glycolysis rate-limiting enzymes in different groups of Caco-2 cells. * indicates P < 0.05 compared to sh-NC or Control group, ** indicates P < 0.01 compared to sh-NC group, # indicates P < 0.05 compared to LEVs + sh-NC group. Cell experiments were repeated three times

Fig 4: Molecular mechanism diagram illustrating how exosome-like vesicles derived from Lactobacillus regulate glycolysis metabolic reprogramming and abnormal proliferation of intestinal epithelial cells through SIRT5-mediated acetylation modification of p53, affecting colon tumor formation

Fig 5: The Effect of LEVs-Mediated SIRT5 Expression Regulation on Proliferation and Glycolysis in Caco-2 Colorectal Cancer Cells. Note: (A-B) RT-qPCR and Western blot detection of SIRT5 mRNA and protein expression in Caco-2 cells overexpressing SIRT5; (C-D) RT-qPCR and Western blot detection of SIRT5 mRNA and protein expression in different groups of Caco-2 cells; (E) EdU staining to detect proliferation of different groups of Caco-2 cells (scale bar = 25 μm); (F) Clonogenic assay to detect colony formation of different groups of Caco-2 cells; (G) Flow cytometry analysis to detect cell cycle changes in different groups of Caco-2 cells; (H) Western blot detection of expression changes of cell cycle-related proteins in different groups of Caco-2 cells; (I) Glucose uptake in different groups of Caco-2 cells; (J) Lactate generation in different groups of Caco-2 cells; (K) Western blot detection of expression of glycolysis rate-limiting enzymes in different groups of Caco-2 cells. * indicates P < 0.05 compared to oe-NC or PBS group, # indicates P < 0.05 compared to LEVs + oe-NC group. Cell experiments were repeated three times