2A). In contrast, claudin-18 this website and 23 mRNAs were not increased upon IL-4-stimulation, while claudin-8 and 9 mRNAs were not detected at all in C57BL6 thio-PEM. Thus, besides E-cadherin, claudin-1, 2 and 11 are members of the junction protein family whose mRNAs are induced in IL-4-stimulated thioglycollate-elicited

peritoneal macrophages. For this reason, we confined our further analysis to this limited set of claudin genes. Of note, the IL-4-mediated induction of these three claudins is largely STAT-6 dependent, as demonstrated by their significantly reduced upregulation in STAT-6-deficient thio-PEM (Fig. 2B). Finally, to extrapolate these findings to other macrophages, BALB/c bone marrow-derived macrophages (BMDM) were treated with IL-4 and assessed for Cdh1, Cldn1, Cldn2 and Cldn11 gene expression. Although Cdh1 and Cldn2 are still inducible by IL-4 in BMDM, the level of induction is less pronounced as compared to thio-PEM (Fig. 2C). As opposed to this, Cldn11 mRNA is strongly induced in BMDM. Of note, the differences in IL-4-mediated Cldn1, 2 and 11 gene inductions between BALB/c thio-PEM and BALB/c BMDM are not because of significant differences in the basal

expression levels of these genes in the https://www.selleckchem.com/products/gsk1120212-jtp-74057.html respective macrophage populations (Table S1). Together, these data confirm the IL-4-induced, STAT6-dependent gene expression of Cldn1, Cldn2 and in particular Cdh1 and Cldn11 in AAMs, but the extent of gene induction depends on the macrophage type. E-cadherin regulation in AAMs was already studied in detail before [8] and is therefore not included in the remainder of this manuscript. To evaluate whether enhanced gene expression of the selected claudins resulted in increased protein levels, we performed a FACS staining on naive and IL-4-stimulated BALB/c thio-PEM. While E-cadherin expression was clearly detected at the cell surface of IL-4-treated,

but not naive thio-PEM as documented before [8], claudin-1, 2 and 11 were below the detection limit (data not shown). Furthermore, no claudin-1, 2 and 11 proteins were detected by Western blot in complete cell lysates of HA 1077 IL-4-stimulated BALB/c thio-PEM. Complete brain and kidney homogenates were used as controls and scored positive for the tested claudins, validating the experimental procedure for detecting these proteins (Fig. S1). The effect of IL-4 on Cldn1 gene expression in macrophages was rather limited. Besides IL-4, other cytokines such as IL-10 and TGF-β have been reported to induce an M2 macrophage activation state. Therefore, BALB/c and C57BL/6 thio-PEM and BALB/c BMDM were treated with IL-4, IL-10 and TGF-β, and Cldn1 induction was assessed.