P16INK4a Deletion Ameliorates Damage of Intestinal Epithelial Barrier and Microbial Dysbiosis in a Stress-Induced Premature Senescence Model of Bmi-1 Deficiency

<jats:p>This study aimed to determine whether <jats:italic>Bmi-1</jats:italic> deficiency leads to intestinal epithelial barrier destruction and microbiota dysfunction, which members of the microbial community alter barrier function with age, and whether <jats:italic>p16</jats:italic><jats:sup><jats:italic>INK4a</jats:italic></jats:sup> deletion could reverse the damage of intestinal epithelial barrier and microbial dysbiosis. Intestines from <jats:italic>Bmi-1</jats:italic>–deficient (<jats:italic>Bmi-1<jats:sup>–/–</jats:sup></jats:italic>), <jats:italic>Bmi-1</jats:italic> and <jats:italic>p16</jats:italic><jats:sup><jats:italic>INK4a</jats:italic></jats:sup> double-knockout (<jats:italic>Bmi-1<jats:sup>–/–</jats:sup>p16<jats:sup><jats:italic>INK4a</jats:italic>–/–</jats:sup></jats:italic>), and wild-type mice were observed for aging and inflammation. Duolink Proximity Ligation Assay, immunoprecipitation, and construction of <jats:italic>p16</jats:italic><jats:sup><jats:italic>INK4a</jats:italic></jats:sup> overexpressed adenovirus and the overexpressed plasmids of full-length, mutant, or truncated fragments for occludin were used for analyzing the interaction between p16<jats:sup><jats:italic>INK4a</jats:italic></jats:sup> and occludin. High-throughput sequencing of V4 region amplicon of 16S ribosomal RNA was conducted using intestinal microbiota. We found <jats:italic>Bmi-1</jats:italic> deficiency destructed barrier structure, barrier function, and tight junction (TJ) in intestinal epithelium; decreased the TJ proteins; increased tumor necrosis factor α (TNF-α)–dependent barrier permeability; and up-regulated proinflammatory level of macrophages induced by intestinal microbial dysbiosis. The transplantation of fecal microbiota from wild-type mice ameliorated TJ in intestinal epithelium of <jats:italic>Bmi-1<jats:sup>–/–</jats:sup></jats:italic> and <jats:italic>Bmi-1<jats:sup>–/–</jats:sup>p16<jats:sup><jats:italic>INK4a</jats:italic>–/–</jats:sup></jats:italic> mice. Harmful bacteria including <jats:italic>Desulfovibrio</jats:italic>, <jats:italic>Helicobacter</jats:italic>, and <jats:italic>Oscillibacter</jats:italic> were at a higher level in <jats:italic>Bmi-1<jats:sup>–/–</jats:sup></jats:italic> mice. More harmful bacteria <jats:italic>Desulfovibrio</jats:italic> entered the epithelium and promoted macrophages-secreted TNF-α and caused TNF-α–dependent barrier permeability and aging. Accumulated p16<jats:sup><jats:italic>INK4a</jats:italic></jats:sup> combined with occludin at the 1st–160th residue in cytoplasm of intestinal epithelium cells from <jats:italic>Bmi-1<jats:sup>–/–</jats:sup></jats:italic> mice, which blocked formation of TJ and the repair of intestinal epithelium barrier. <jats:italic>P16</jats:italic><jats:sup><jats:italic>INK4a</jats:italic></jats:sup> deletion could maintain barrier function and microbiota balance in <jats:italic>Bmi-1<jats:sup>–/–</jats:sup></jats:italic> mice through strengthening formation of TJ and decreasing macrophages-secreted TNF-α induced by <jats:italic>Desulfovibrio</jats:italic> entering the intestinal epithelium. Thus, Bmi-1 maintained intestinal TJ, epithelial barrier function, and microbiota balance through preventing senescence characterized by p16<jats:sup><jats:italic>INK4a</jats:italic></jats:sup> accumulation. The clearance of p16<jats:sup><jats:italic>INK4a</jats:italic></jats:sup>-positive cells in aging intestinal epithelium would be a new method for maintaining barrier function and microbiota balance. The residues 1–160 of occludin could be a novel therapeutic target for identifying small molecular antagonistic peptides to prevent the combination of p16<jats:sup><jats:italic>INK4a</jats:italic></jats:sup> with occludin for protecting TJ.</jats:p>