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Although mechanisms of immune activation against liver ischemia reperfusion (IR) injury (IRI) have been studied extensively, questions regarding liver-resident macrophages, that is, Kupffer cells (KCs), remain controversial. Recent progress in the biology of tissue-resident macrophages implicates homeostatic functions of KCs. This study aims to dissect responses and functions of KCs in liver IRI. In a murine liver partial warm ischemia model, we analyzed liver-resident versus infiltrating macrophages by FACS and immunofluorescence staining. Our data showed that liver immune activation by IR was associated with not only infiltrations/activations of peripheral macrophages, but also necrotic depletion of KCs. Inhibition of receptor-interacting protein 1 (RIP1) by necrostatin-1s protected KCs from ischemia-induced depletion, resulting in the reduction of macrophage infiltration, suppression of proinflammatory immune activation, and protection of livers from IRI. The depletion of KCs by clodronate liposomes abrogated the effect of necrostatin-1s. Additionally, liver reconstitutions with KCs postischemia exerted anti-inflammatory/cytoprotective effects against IRI. These results reveal a unique response of KCs against liver IR, that is, RIP1-dependent necrosis, which constitutes a novel mechanism of liver inflammatory immune activation in the pathogenesis of liver IRI. [ABSTRACT FROM AUTHOR]

Glycogen synthase kinase 3 (Gsk3) a and b are both constitutively active and inhibited upon stimulation by N-terminal serine phosphorylation. Although roles of active Gsk3 in liver ischemia reperfusion injury (IRI) have been well appreciated, whether Gsk3 N-terminal serine phosphorylation has any functional significance in the disease process remains unclear. In a murine liver partial warm ischemia model, we studied Gsk3 N-terminal serine mutant knock-in (KI) mice and showed that liver IRI was decreased in Gsk3aS21A but increased in Gsk3bS9A mutant KI mice. Bone marrow chimeric experiments revealed that the Gsk3a, but not b, mutation in liver parenchyma protected from IRI, and both mutations in bone marrow-derived cells exacerbated liver injuries. Mechanistically, mutant Gsk3a protected hepatocytes from inflammatory (TNF-a) cell death by the activation of HIV-1 TAT-interactive protein 60 (TIP60)-mediated autophagy pathway. The pharmacological inhibition of TIP60 or autophagy diminished the protection of the Gsk3a mutant hepatocytes from inflammatory cell death in vitro and the Gsk3a mutant KI mice from liver IRI in vivo. Thus, Gsk3 N-terminal serine phosphorylation inhibits liver innate immune activation but suppresses hepatocyte autophagy in response to inflammation. Gsk3 aS21, but not bS9, mutation is sufficient to sustain Gsk4 activities in hepatocytes and protect livers from IRI via TIP60 activation. [ABSTRACT FROM AUTHOR]

Previous investigations demonstrated that protein tyrosine phosphatase, receptor type, O (PTPRO) acts as a tumor suppressor in liver cancer; however, little is known about its role in liver inflammation. Thus, we investigated the role of PTPRO in fulminant hepatitis (FH) using a Con A-induced mouse model. Significantly more severe liver damage, but attenuated inflammation, was detected in PTPRO-knockout (KO) mice, and PTPRO deficiency could confer this phenotype to wild-type mice in bone marrow transplantation. Moreover, hepatocytes with PTPRO depletion were more sensitive to TNF-α-induced apoptosis, and secretion of cytokines was significantly decreased in both T and NK/NKT cells and led to marked impairment of NF-κB activation. Intriguingly, wild-type and PTPRO-KO cells responded equally to TNF-α in activation of IKK, but NF-κB activation was clearly decreased in PTPRO-KO cells. PTPRO associated with ErbB2, and loss of PTPRO potentiated activation of the ErbB2/Akt/GSK-3β/β-catenin cascade. Increased β-catenin formed a complex with NF-κB and attenuated its nuclear translocation and activation. Importantly, in humans, PTPRO was much decreased in FH, and this was associated with enhanced β-catenin accumulation but reduced IFN-γ secretion. Taken together, our study identified a novel PTPRO/ErbB2/Akt/GSK-3β/β-catenin/NF-κB axis in FH, which suggests that PTPRO may have therapeutic potential in this liver disease. [ABSTRACT FROM AUTHOR]

The use of TGF-β-induced CD4+Foxp3+ T cells (induced regulatory T cells [iTregs]) is an important prevention and treatment strategy in autoimmune diseases and other disorders. However, the potential use of iTregs as a treatment modality for acute graftversus- host disease (aGVHD) has not been realized because they may be unstable and less suppressive in this disease. We restudied the ability of iTregs to prevent and treat aGVHD in two mouse models. Our results showed that, as long as an appropriate iTreggeneration protocol is used, these iTregs consistently displayed a potent ability to control aGVHD development and reduce mortality in the aGVFH) animal models. iTreg infusion markedly suppressed the engraftment of donor CD8+ cells and CD4+ cells, the expression of granzyme A and B, the cytotoxic effect of donor CD8+ cells, and the production of T cell cytokines in aGVHD. Therefore, we conclude that as long as the correct methods for generating iTregs are used, they can prevent and even treat aGVHD. [ABSTRACT FROM AUTHOR]

Although the role of phosphatase and tensin homolog deleted on chromosome 10 (PTEN) in regulating cell proliferation is well established, its function in immune responses remains to be fully appreciated. In the current study, we analyzed myeloid-specific PTEN function in regulating tissue inflammatory immune response in a murine liver partial warm ischemia model. Myeloid-specific PTEN knockout (KO) resulted in liver protection from ischemia reperfusion injury (IRI) by deviating the local innate immune response against ischemia reperfusion toward the regulatory type: expression of proinflammatory genes was selectively decreased and anti-inflammatory IL-10 was simultaneously increased in ischemia reperfusion livers of PTEN KO mice compared with those of wild-type (WT) mice. PI3K inhibitor and IL-10-neutralizing Abs, but not exogenous LPS, recreated liver IRI in these KO mice. At the cellular level, Kupffer cells and peritoneal macrophages isolated from KO mice expressed higher levels of M2 markers and produced lower TNF-α and higher IL-10 in response to TLR ligands than did their WT counterparts. They had enhanced Stat3- and Stat6-signaling pathway activation, but diminished Stat1-signaling pathway activation, in response to TLR4 stimulation. Inactivation of Kupffer cells by gadolinium chloride enhanced proinflammatory immune activation and increased IRI in livers of myeloid PTEN KO mice. Thus, myeloid PTEN deficiency protects livers from IRI by facilitating M2 macrophage differentiation. [ABSTRACT FROM AUTHOR]

Liver fibrosis is a severe, life-threatening clinical condition resulting from nonresolving hepatitis of different origins. IL-17A is critical in inflammation, but its relation to liver fibrosis remains elusive. We find increased IL-17A expression in fibrotic livers from HBV-infected patients undergoing partial hepatectomy because of cirrhosis-related early-stage hepatocellular carcinoma in comparison with control nonfibrotic livers from uninfected patients with hepatic hemangioma. In fibrotic livers, IL-17A immunoreactivity localizes to the inflammatory infiltrate. In experimental carbon tetrachloride-induced liver fibrosis of IL-17RA-deficient mice, we observe reduced neutrophil influx, proinflammatory cytokines, hepatocellular necrosis, inflammation, and fibrosis as compared with control C57BL/6 mice. IL-17A is produced by neutrophils and T lymphocytes expressing the Th17 lineage-specific transcription factor Retinoic acid receptor-related orphan receptor γt. Furthermore, hepatic stellate cells (HSCs) isolated from naive C57BL/6 mice respond to IL-17A with increased IL-6, α-smooth muscle actin, collagen, and TGF-β mRNA expression, suggesting an IL-17A-driven fibrotic process. Pharmacologic ERK1/2 or p38 inhibition significantly attenuated IL-17A-induced HSC activation and collagen expression. In conclusion, IL-17A+ Retinoic acid receptor-related orphan receptor γt+ neutrophils and T cells are recruited into the injured liver driving a chronic, fibrotic hepatitis. IL-17A-dependent HSC activation may be critical for liver fibrosis. Thus, blockade of IL-17A could potentially benefit patients with chronic hepatitis and liver fibrosis. [ABSTRACT FROM AUTHOR]

Objective: In small-for-size liver transplantation, portal hypertension aggravates endotoxin from the gut which accelerates the activation of inducible nitric oxide synthase (iNOS). However, there is little knowledge as to the effects of iNOS inhibitors on small-for-size graft damage. Our study was designed to investigate the role of an iNOS inhibitor both with and without lipopolysaccharide (LPS) treatment in ischemia-reperfusion injury of small-for-size liver transplantation. Methods: Subjecting Sprague-Dawley rats to small-for-size grafts liver transplantation, we investigated the time course of changes in hepatic expression of iNOS and endothelial nitric oxide synthase (eNOS). Meantime, we also investigated the effects of iNOS inhibitor, both with and without LPS treatment, at 6 h after reperfusion. Results: While iNOS mRNA expression reached a peak at 3 h, the highest protein level occurred at 6 h after reperfusion. Aminoguanidine (AG) significantly inhibited mRNA and protein expressions of iNOS. but not that of eNOS. However. LPS accelerated activation of iNOS, but suppressed the expression of eNOS. Meanwhile, compared with the untreated group, those treated with AG or LPS experienced worsened liver function and tissue damage, promoting neutrophil infiltration in the liver tissue. The difference between the LPS group and the LPS + AG group was found to be significant. In addition, AG and LPS treatments up- regulated the protein expression of ICAM-1 and NF-κB p65. Conclusion: In a small-for-size model of rat liver transplantation, regardless of LPS treatment, the inhibitor of iNOS. AG. attenuated iNOS expression, but worsened liver function and tissue damage. The subsequent increased neutrophil infiltration in liver tissue may be associated with up-regulation of ICAM-1 and NF-κB expressions. [ABSTRACT FROM AUTHOR]

Objective: Graft size is one of the major risk factors in adult-to-adult living donor liver transplantation and rapid regeneration is an essential post-operative requirement. Tschemic preconditioning (TPC) has been shown to be an effective strategy in the reduction of hepatic ischemia-reperfusion injury and stimulation of liver regeneration. This study was designed to evaluate the effects of IPC on liver regeneration in small-for-size liver grafts. Methods: We employed a rat orthotopic liver transplantation model using small-for-size (30%) grafts, in the presence or absence (control) of IPC (10 mm of ischemia followed by 15 mm of reperfusion). Survival rate, graft injury, hepatocellular proliferation, cell cycle progression, Stat3 activation, as well as TNF-α and IL-6 expression were assessed. Results: IPC significantly enhanced the extent of graft injury and hindered hepatic regeneration in small-for-size liver grafts. The 7-day survival rate was also reduced by IPC, but failed to reach statistical significance. IPC did not affect TNF-α levels, but significantly decreased the elevation of TL-6 after reperfusion. These findings were correlated with down-regulation of cyclin E and cyclin Dl, and decreased numbers of PCNA-positive nuclei in IPC grafts. These results were inconsistent with Stat3 activation, as P-Stat3 exhibited a stronger and prolonged pattern of expression in the IPC group, compared to controls. Conclusions: lschemic preconditioning may impair liver regeneration in small-for-size liver grafts by decreasing lL-6 and blunting cell cycle progression, through a mechanism at least partially independent of Stat3. [ABSTRACT FROM AUTHOR]