The non-
receptor protein tyrosine kinase 2β (Pyk2) phosphorylated tricellular tight junction (tTJ) molecules angulin-1/LSR and
tricellulin (TRIC) and the inhibitor PF-431396 (PF43) suppress angulin-1/LSR and TRIC recruitment to tTJs. The disruption of the intestinal epithelial barrier by high mobility group box 1 (
HMGB1) and the inflammatory
cytokines TNFα and IFNγ contributes to downregulation of angulin-1/LSR and TRIC in 2.5D culture of Caco-2 cells as a novel model of
inflammatory bowel disease (IBD). In the present study, to investigate the roles of Pyk2 phosphorylated angulin-1/LSR and TRIC in the intestinal epithelial barrier, 2D and 2.5D cultures of Caco-2 cells were treated with the Pyk2 inhibitor PF-43 with or without
HMGB1, inflammatory
cytokines TNFα and IFNγ. Treatment with PF-43 increased expression of angulin-1/LSR, phosphorylated AMPK and phosphorylated MAPK and decreased that of phosphorylated JNK, with upregulation of the epithelial barrier and cellular metabolism measured as basal oxygen consumption rate (OCR) and
ATP production in 2D culture. Treatment with PF-43 prevented the downregulation of the epithelial barrier by
HMGB1 and inflammatory
cytokines in 2D culture. Treatment with PF-43 prevented the epithelial hyperpermeability induced by
HMGB1 and inflammatory
cytokines in 2.5D culture. In 2.5D culture, treatment with PF-43 inhibited the decreases of angulin-1/LSR, TRIC, pJNK, pAMPK and pMAPK induced by
HMGB1 and the inflammatory
cytokines. Treatment with PF-43 inhibited in part the induced phosphorylation of the
serine of angulin-1/LSR and TRIC. Pyk2 inhibitor PF-43 may have potential for use in
therapy for IBD via its actions with regard to phosphorylated tTJs and cellular metabolism.