Although disruption of β-catenin signaling did not affect the frequency of CD4+ DC versus CD8α+ DC populations in the liver (Supporting Fig. 4), it did increase (P < 0.005) PTEN activity (Fig. 6C) and IL-12p40 mRNA expression (Fig. 6D) in hepatic DCs, as compared with NS siRNA controls. We investigated the regulatory role of β-catenin on apoptosis pathways by western blots. By 6 hours of reperfusion after 90 minutes of ischemia, knockdown of β-catenin Dabrafenib manufacturer in Ad-HO-1 or Ad-IL-10-transfected livers down-regulated Bcl-2/Bcl-xL (0.1-0.3 AU and 0.3-0.6 AU, respectively), yet up-regulated cleaved caspase-3 (2.4-2.7 AU) (Fig. 7A). In contrast, the expression of Bcl-2/Bcl-xL strongly up-regulated

in NS siRNA-treated livers after Ad-HO-1 or Ad-IL-10 (2.0-2.2 AU and 2.1-2.3 AU, respectively),

whereas the expression of cleaved caspase-3 was inhibited in NS siRNA-treated controls (0.3-0.5 AU). These results were confirmed by increased caspase-3 activity in siβ-cat- but not NS siRNA-treated mice (Fig. 7B: 4.12 ± 0.42 and 4.01 ± 0.4 versus 1.19 ± 0.29 and 1.08 ± 0.32, respectively, P < 0.001). We further analyzed IR-induced hepatic oncotic necrosis/apoptosis selleck chemicals llc by TUNEL staining (Fig. 7C,D). Livers in mice treated with siβ-cat showed increased frequency of TUNEL+ cells (Fig. 7Cc/e: 28.6 ± 10.8 and 26.1 ± 11.1, respectively), compared with NS siRNA controls (Fig. 7Cd/f: 6.5 ± 3.6 and 5.5 ± 3.2, respectively, P < 0.0001). Both innate and adaptive immune responses are essential in the mechanism of liver IRI.1 By regulating the initial

response in damaged/necrotic cells by way of TLR4 signaling, DCs are key mediators of immune homeostasis,25 yet by amplifying innate responses they may also promote the development of adaptive immunity.5, Pregnenolone 6 Our results highlight the regulatory role of β-catenin to orchestrate local inflammation, PTEN/PI3K and TLR4 signaling in IR-stressed liver. Our in vitro data support the regulatory function of STAT3-induced β-catenin in DC activation and PTEN/TLR4 signaling. Previous studies have implicated STAT3-mediated antiinflammatory phenotype in LPS-stimulated DCs.26 We found that CoPP- or rIL-10-induced STAT3 triggered translocation of β-catenin from the cytoplasm to the nucleus, and transcription of its target genes in BMDCs. Activation of β-catenin inhibited IL-12p40, TNF-α, and IL-6 expression, as well as DC maturation by down-regulating costimulatory CD40, CD80, and CD86. In addition, our findings suggest that GSK-3β may play a role in β-catenin activation and DC maturation. Interestingly, STAT3 knockdown in LPS-stimulated BMDCs depressed β-catenin and Akt but enhanced PTEN expression, leading to increased DC expression of proinflammatory mediators and costimulatory molecules, suggesting STAT3 can mediate β-catenin activation to program DC functions.