Aberrant methylation of PSD disturbs Rac1-mediated immune responses governing neutrophil chemotaxis and apoptosis in ulcerative colitis-associated carcinogenesis

We previously reported that the Pleckstrin and Sec7 domain-containing (PSD)gene is preferentially methylated in patients with ulcerative colitis (UC) whodeveloped colorectal cancer (CRC), and is implicated in UC-associated carcinogenesisthrough its inhibition of apoptosis. This study aimed to determine the potentialeffect of PSD methylation on its downstream molecule, Ras-related C3 botulinumtoxin substrate 1 (Rac1), which governs neutrophil chemotaxis and apoptosis signaling.PSD was knocked down in a normal human fibroblast cell line (HNDF) and a neutrophil-likecell line (HL-60). Both NHDF and HL-60 cells exhibited numerous filamentous-actin(F-actin) rich membrane extensions, resulting in the activation of Rac1; thisactivation was hampered by PSD silencing. Lipopolysaccharide, a reactive oxygenspecies (ROS) inducer, stimulated NHDF cells to release ROS and activated caspase‑3/7in the presence of neutrophils, which was inhibited by PSD knockdown. Migrationassays demonstrated that chemotaxis of HL-60 cells was affected by PSD silencingin NHDF cells. Tissue sections from 6 UC patients with CRC and 15 UC patientswithout CRC were examined. To verify Rac1-mediated chemotaxis in tissue sections,we evaluated the grade of neutrophil infiltration by histological assessment andassessed F-actin and PSD expression by immunohistochemistry. Neutrophil infiltration,F-actin and PSD expression were significantly decreased in specimens from UC patientswith PSD methylation compared with those without. Decreased levels of F-actinexpression were observed in colorectal mucosa, as well as in infiltrating cellswith PSD methylation. PSD expression was preferentially inhibited in colorectalmucosa by PSD methylation, whereas PSD expression was rarely observed in infiltratingcells, regardless of PSD methylation status. These data indicate that aberrantmethylation of PSD occurs in UC-associated colorectal mucosa, enabling circumventionof Rac1-mediated immune responses governing neutrophil chemotaxis and apoptosis,and thus plays a pivotal role in the mechanisms underlying UC-associated carcinogenesis.