Published online Dec 14, 2018. doi: 10.3748/wjg.v24.i46.5234
Peer-review started: September 4, 2018
First decision: October 4, 2018
Revised: October 31, 2018
Accepted: November 7, 2018
Article in press: November 8, 2018
Published online: December 14, 2018
To determine whether cell division cycle (Cdc)42 is regulated by microRNA (miR)-15a in the development of pediatric inflammatory bowel disease (IBD).
We cultured 293T cells, used plasmids and performed dual-luciferase assay to determine whether Cdc42 is a miR-15a target gene. We cultured Caco-2 cells, and stimulated them with tumor necrosis factor (TNF)-α. We then employed lentiviruses to alter the expression of miR-15a and Cdc42. We performed quantitative reverse transcription polymerase chain reaction (qRT-PCR) and immunofluorescence to determine whether Cdc42 is regulated by miR-15a in Caco-2 cells. Finally, we collected ileocecal tissue by endoscopy from patients and performed qRT-PCR to examine the expression of miR-15a and Cdc42 in pediatric IBD patients.
Target Scan and dual-luciferase assay revealed that Cdc42 was a miR-15a target gene. MiR-15a expression increased (P = 0.0038) and Cdc42 expression decreased (P = 0.0013) in cells stimulated with TNF-α, and the expression of the epithelial junction proteins zona occludens (ZO)-1 (P < 0.05) and E-cadherin (P < 0.001) decreased. Cdc42 levels decreased in miR-15a-mimic cells (P < 0.001) and increased in miR-15a inhibitor cells (P < 0.05). ZO-1 and E-cadherin decreased in miR-15a-mimic cells (P < 0.001) but not in the miR-15a inhibitor + TNF-α cells. In Lv-Cdc42 + TNF-α cells, ZO-1 and E-cadherin expression increased compared to the Lv-Cdc42-NC + TNF-α (P < 0.05) or miR-15a-mimic cells (P < 0.05). Fifty-four pediatric IBD patients were included in this study, 21 in the control group, 19 in the Crohn’s disease (CD) active (AC) group, seven in the CD remission (RE) group, and seven in the ulcerative colitis (UC) group. MiR-15a increased and Cdc42 decreased in the CD AC group compared to the control group (P < 0.05). miR-15a decreased and Cdc42 increased in the CD RE group compared to the CD AC group (P < 0.05). miR-15a was positively correlated with the Pediatric Crohn’s disease Activity Index (PCDAI) (P = 0.006), while Cdc42 was negatively correlated with PCDAI (P = 0.0008). Finally, miR-15a expression negatively correlated with Cdc42 in pediatric IBD patients (P = 0.0045).
MiR-15a negatively regulates epithelial junctions through Cdc42 in Caco-2 cells and pediatric IBD patients.
Core tip: This study aimed to determine whether cell division cycle (Cdc)42 was regulated by microRNAs in pediatric inflammatory bowel disease (IBD) development. We studied microRNA (miR)-15a - Cdc42 signaling in pediatric IBD patients and found that miR-15a negatively regulated Cdc42 in pediatric IBD patients, and both were strongly correlated with the severity of IBD. Additionally, this study provides evidence that Cdc42 is a target gene of miR-15a, and this will provide a critical clue for understanding IBD development.