Notch1 mediates acute colitis by downregulating IL17 and IL22 cytokines and cross talking with NFkB pathway
Authors: Pujada Adani , Bui Tien Anh , Merlin Didier , Garg Pallavi 
Published: June 2021

Introduction

Ulcerative colitis (UC) is an inflammatory bowel disease (IBD) that causes chronic inflammation and ulcers affecting the innermost lining of colon and rectum. Almost 1 million people suffer through UC in the United States alone, with a disease history of 16 years. Notch1 signaling is an evolutionarily conserved intercellular signaling pathway. It regulates cell fate, cell proliferation, cell differentiation, and cell death. However, Notch1 can act as a mediator or suppressor depending on the cellular context and disease stage.

Aim

Aim of this study is to understand the mechanistic role of Notch1 in UC.

Methods

We used 7 weeks old (both male and female) transgenic mice (C57/B6) with conditional deletion of Notch1 under villin promoter (Ncre). Notch1flox/flox mice (Nflox) were used as the control group since they were crossed with Crevillin mice to generate Ncre mice. Enteroids were isolated from 6 weeks old Ncre and Nflox mice to study proliferation and differentiation. Colonic inflammation was induced by the inflammatory agent dextran sodium sulfate (2.5% in drinking water for 5 days). Colonic tissues were used for western blot (WB) analysis and histochemistry.

Results

WB analysis showed that Notch1 signaling was absent in colon, small intestine, spleen and cecum of Ncre mice. However Notch 1 signaling was active in the liver of both Ncre and Nflox mice. In vivo permeability assay using 4kD FITC‐dextran showed that barrier function of Ncre mice was compromised compared to Nflox mice. Alcian blue ‐ periodic acid Schiff stain showed increased goblet cells among Ncre mice compared to Nflox mice. Enteroids isolated from Ncre and Nflox mice indicated compromised differentiation and altered proliferation among Ncre enteroids compared to Nflox ones on Days 2, 4 and 7. WB analysis exhibited decreased expression of PepT1 (enterocyte marker) and PCNA (proliferation maker) among Ncre enteroids compared to Nflox ones. We observed that Ncre mice were resistant to DSS induced colitis compared to Nflox mice as reflected by the body weight loss. Hematoxylin and Eosin staining exhibited decreased neutrophil infiltration, less damage to crypt architecture among Ncre mice compared to Nflox mice in acute colitis. QPCR analysis showed increased mRNA levels of IL6, TNFa, IL17 and IL22 cytokines among Ncre mice compared to Nflox mice in acute colitis. WBs indicated that Ncre mice had decreased expressions of NFkB and CyclinD (downstream target of NFkB and a proliferation marker). However there was no significant difference in protein expressions Cox2 or iNOS (inflammatory signaling pathways) among Ncre and Nflox mice as indicated by the WBs.

Conclusion

Our study shows that Notch1 mediates acute colitis by cross talking with NFkB pathway and promoting the abnormal proliferation. We also observed that Notch1 activation inhibits the activation of anti‐inflammatory cytokines (IL17 and IL22). However silencing of Notch1 signaling promotes goblet cell hyperplasia and inflammatory cytokines such as IL6 and TNFa. Therefore, our study suggests that instead of Notch1 silencing, NFkB inhibitors might be a better option as therapeutics for the colitis patients in the clinics.

Support or Funding Information

NIDDK‐R01 Grant DK064711 to DM and 3057‐CCFAgrant to PG

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.