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Loss of Sprouty2 in the colonic epithelium limits inflammatory cytokine production and enhances Muc2 and Lgr5 expression during colitis
Author(s) -
Schumacher Michael A,
Hsieh Jonathan J,
Almohazey Dana,
Warburton David,
Washington M. Kay,
Frey Mark R
Publication year - 2017
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.31.1_supplement.1049.12
Subject(s) - cytokine , colitis , mapk/erk pathway , tumor necrosis factor alpha , intestinal epithelium , myeloid , inflammatory bowel disease , immunology , epithelium , biology , cancer research , medicine , signal transduction , microbiology and biotechnology , pathology , disease
OBJECTIVE Sprouty2 (Spry2) is an intracellular modulator of MAPK signaling. Its expression is suppressed by tumor necrosis factor (TNF) in airway epithelium, but its regulation and activity in the intestinal tract are unknown. Aberrant MAPK signaling is associated with increased inflammatory cytokine production in diseases including inflammatory bowel disease, raising the possibility that Spry2 may play a role in regulating cytokine expression in the colon. MAPK is also an important regulator of secretory cell differentiation in the intestine. Here we tested the hypothesis that Spry2 regulation of MAPK signaling during colitis controls inflammatory cytokine production and goblet cell function, and thus impacts intestinal response to injury. METHODS We generated VillinCre;Spry2FLOXXED mice that lack Spry2 in the intestinal epithelium (Spry2KOIE) and LysMCre;Spry2FLOXXED mice that lack Spry2 in cells of myeloid lineage. Mice were given 3% dextran sodium sulfate (DSS) in drinking water for 4 days followed by 3 days without DSS and colitis severity was assessed. Cytokine levels were determined by qPCR. Epithelial crypt cultures (colonoids) were generated to assess regulation of ISC and cell lineages. The role of MAPK signaling was tested with MEK inhibitor U0126. RESULTS DSS colitis caused a 59% reduction of colonic Spry2 (p<0.05). Spry2KOIE mice subjected to DSS had decreased levels of TNF, CXCL2, IL1β, and IFNγ versus wildtype littermates, and were protected from histological damage (injury score reduced by 30%, p=0.037), weight loss, and colon shortening. In contrast, mice with Spry2 deletion in myeloid cells developed acute colitis similar to controls, suggesting an epithelial specific effect of Spry2 in colitis. Spry2KOIE mice showed higher colonic Lgr5 (ISC marker) and Muc2 (goblet cell marker) levels. In vitro, Spry2KOIE‐derived colonoids also showed a 3‐fold increase (p=0.005) in Muc2 expression and elevated Lgr5 expression. Because goblet cell differentiation is thought to be dependent upon MAPK signaling, we analyzed colonoids treated with the MAPK inhibitor, U0126, but surprisingly did not see reversal of the elevated Muc2 in Spry2KOIE colonoids. CONCLUSION Our work demonstrates that Spry2 is robustly expressed in the colonic epithelium, and expression is regulated by injury and inflammation. This may represent a protective mechanism against tissue injury by limiting inflammatory cytokine production, thus supporting Lgr5+ ISCs and inducing expression of the protective mucin, Muc2. Elevated Muc2 in Spry2KOIE colonoids was independent of MAPK activation, suggesting loss of Spry2 may elicit colonic protection through an alternative route of inhibiting inflammatory cytokine production. Support or Funding Information NIH R01DK095004 (MRF)