To exclude unspecific effects of the chitin particles, we included glass beads of comparable size or just PBS as controls.

Chitin did neither induce nor inhibit Th2 polarization under neutral or polarizing conditions, respectively (Fig. 1C). This result led us to conclude that reduced Th2-cell expansion in vivo is not due to a direct inhibitory effect of chitin on T cells. The reduced frequency of TCR-tg cells in lung and LN of OVA/chitin-treated mice (Fig. 1A and B) could be due to inefficient homing, impaired proliferation or reduced survival of transferred T cells. To address the possibility that chitin inhibits T-cell proliferation, we labeled total splenocytes from BALB/c mice with CFSE and stimulated them in vitro with anti-TCR/anti-CD28 in the presence or absence of chitin. Proliferation of CD4+ T cells was inhibited by 30–40% selleck in the presence of chitin (Fig. 2A and B). Next, we sought to determine whether the inhibitory function of chitin was mediated by binding of chitin to the mannose receptor which has previously been shown to

serve as chitin receptor 11. T cells were cultured in the presence of chitin and soluble mannan in order to block binding of chitin to the mannose receptor as described earlier 11. However, the inhibitory activity of chitin was not reduced in the presence of INCB018424 in vitro mannan and mannan alone did not inhibit T-cell proliferation (Fig. 2A and B). To further determine whether inhibition of T-cell proliferation can be induced by direct interaction of chitin with T cells, we stimulated purified and CFSE-labeled CD4+ T cells on anti-TCR/anti-CD28-coated plates in the presence or absence of chitin or glass. Chitin or glass had no influence on the efficiency of T-cell proliferation, suggesting that the inhibitory effect of chitin is mediated by an accessory cell

type (Fig. 2C). The accessory cell type that mediates chitin-induced inhibition of T-cell proliferation might be a macrophage population. Indeed, cocultures of macrophages and T cells revealed Atezolizumab order that chitin caused a strong inhibitory effect (Fig. 3A and B). We have previously shown that chitin upregulates expression of Arg1, an enzyme which is induced in macrophages by Stat6-mediated signaling from the IL-4 receptors and therefore served as marker to indicate differentiation into AAM 9, 23, 24. However, it has recently been shown that Arg1 can also be induced by TLR-mediated signaling in a Stat6-independent fashion 25. Therefore, other markers like the chitinase-like protein Ym1 or “found in inflammatory zone 1” (Fizz1)/“resistin-like molecule α” appear to be more specific for AAM 26.