P087 Increased wnt ligands expression in M2c macrophages is associated with fibrosis in Stat6 knockout mice

Ortiz-Masia D.^*1,2, Salvador P.³, Macias-Ceja D.C.⁴, Gisbert-Ferrándiz L.³, Hernandez C.⁴, Calatayud S.³, Esplugues J.V.^3,4, Barrachina D.³

¹Universidad de Valencia-CIBERehd, Pharmacology, Valencia, Spain ²Universidad de Valencia, Medicine, Valencia, Spain ³Universidad de Valencia, Pharmacology, Valencia, Spain ⁴FISABIO Hospital Peset, Pharmacology, Valencia, Spain

Background

STAT6 plays a crucial role in M2a macrophage polarization in vitro and these cells mediate mucosal healing in an acute model of TNBS-colitis through the expression of Wnt ligands [1]. We have recently reported that STAT6 deficiency favours fibrosis in a murine model of TNBS colitis [2] and we aim to characterize here the functional relevance of the macrophage phenotype in fibrosis development.

Methods

WT or STAT6(−/−) mice were given TNBs (0.5, 0.5, 0.75, 0.75, 1, and 1 mg, intrarectally) or saline weekly and they were sacrificed 3, 5 or 7 weeks after the first TNBs administration. The percentage of CD206, CD16, and CD86 positive cells was analyzed by flow cytometry in F4/80+ macrophages isolated from the intestinal mucosa. The mRNA expression of Wnt ligands was evaluated in F4/80+ CD16+ macrophages isolated from the mucosa, 7 weeks after the first TNBs administration and results are expressed as fold induction vs vehicle-treated mice. Data are expressed as mean ± SEM with n≥8 in all groups (*p<0.05).

Results

TNBs increased the percentage of CD206 positive macrophages in the mucosa of TNBS-WT animals while it failed to do that in TNBS-STAT6(−/−) mice. The percentage of CD16 positive macrophages increased in a time-dependent manner only in the mucosa of STAT6(−/−)-TNBS-treated mice. The number of CD86+ cells was similar in TNBS-WT and TNBS-STAT6(−/−) mice.

Figure 1. Macrophages population.

In CD16+ macrophages isolated from TNBS-STAT6(−/−) mice the mRNA expression of canonical and non-canonical Wnt ligands was significantly increased compared with cells isolated from TNBS-WT mice.

Table 1. Wnt ligands

	Wnt ligand	2B	5A	6	7B	10A	10B
7-week	WT	1.6±0.2	2.1± 0.5	1.2±0.1	2.7±0.6	3.3± 0.8	1.9±0.2
7-week	STAT6(−/−)	1.9±0.4	6.4±0.6*	2.7±0.4*	15.3±2.2*	2.9±1.0	15.3±2.5*

Conclusion

An increased percentage of M2c macrophages which exhibited an increased expression of Wnt ligands is detected in the mucosa of TNBS-STAT6 knockout mice and it could mediate the increased fibrosis detected in these animals.

References:

[1] Cosín-Roger J, Ortiz-Masiá D, Calatayud S, Hernández C, Esplugues JV, Barrachina MD., (2016), The activation of Wnt signaling by a STAT6-dependent macrophage phenotype promotes mucosal repair in murine IBD., Mucosal Immunol. 2016 Jul;9(4):986–98. doi: 10.1038/mi.2015.123.

[2] P. Salvador, (2016), STAT6 deficiency alters macrophage polarization and promotes fibrosis in a murine model of chronic inflammation. ECCO2016

Posted in: Poster presentations: Basic science (2017)