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Macrophage heme oxygenase-1-SIRT1-p53 axis regulates sterile inflammation in liver ischemia-reperfusion injury.

ABSTRACT: BACKGROUND & AIMS:Hepatic ischemia-reperfusion injury (IRI), characterized by exogenous antigen-independent local inflammation and hepatocellular death, represents a risk factor for acute and chronic rejection in liver transplantation. We aimed to investigate the molecular communication involved in the mechanism of liver IRI. METHODS:We analyzed human liver transplants, primary murine macrophage cell cultures and IR-stressed livers in myeloid-specific heme oxygenase-1 (HO-1) gene mutant mice, for anti-inflammatory and cytoprotective functions of macrophage-specific HO-1/SIRT1 (sirtuin 1)/p53 (tumor suppressor protein) signaling. RESULTS:Decreased HO-1 expression in human post-reperfusion liver transplant biopsies correlated with a deterioration in hepatocellular function (serum ALT; p<0.05) and inferior patient survival (p<0.05). In the low HO-1 liver transplant biopsy group, SIRT1/Arf (alternative reading frame)/p53/MDM2 (murine double minute 2) expression levels decreased (p<0.05) while cleaved caspase 3 and frequency of TUNEL+cells simultaneously increased (p<0.05). Immunofluorescence showed macrophages were the principal source of HO-1 in human and mouse IR-stressed livers. In vitro macrophage cultures revealed that HO-1 induction positively regulated SIRT1 signaling, whereas SIRT1-induced Arf inhibited ubiquitinating activity of MDM2 against p53, which in turn attenuated macrophage activation. In a murine model of hepatic warm IRI, myeloid-specific HO-1 deletion lacked SIRT1/p53, exacerbated liver inflammation and IR-hepatocellular death, whereas adjunctive SIRT1 activation restored p53 signaling and rescued livers from IR-damage. CONCLUSION:This bench-to-bedside study identifies a new class of macrophages activated via the HO-1-SIRT1-p53 signaling axis in the mechanism of hepatic sterile inflammation. This mechanism could be a target for novel therapeutic strategies in liver transplant recipients. LAY SUMMARY:Post-transplant low macrophage HO-1 expression in human liver transplants correlates with reduced hepatocellular function and survival. HO-1 regulates macrophage activation via the SIRT1-p53 signaling network and regulates hepatocellular death in liver ischemia-reperfusion injury. Thus targeting this pathway in liver transplant recipients could be of therapeutic benefit.

SUBMITTER: Nakamura K

PROVIDER: S-EPMC5884687 | biostudies-literature | 2017 Dec

REPOSITORIES: biostudies-literature

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Macrophage heme oxygenase-1-SIRT1-p53 axis regulates sterile inflammation in liver ischemia-reperfusion injury.

Nakamura Kojiro K Zhang Min M Kageyama Shoichi S Ke Bibo B Fujii Takehiro T Sosa Rebecca A RA Reed Elaine F EF Datta Nakul N Zarrinpar Ali A Busuttil Ronald W RW Araujo Jesus A JA Kupiec-Weglinski Jerzy W JW

Journal of hepatology 20170823 6

<h4>Background & aims</h4>Hepatic ischemia-reperfusion injury (IRI), characterized by exogenous antigen-independent local inflammation and hepatocellular death, represents a risk factor for acute and chronic rejection in liver transplantation. We aimed to investigate the molecular communication involved in the mechanism of liver IRI.<h4>Methods</h4>We analyzed human liver transplants, primary murine macrophage cell cultures and IR-stressed livers in myeloid-specific heme oxygenase-1 (HO-1) gene ...[more]

PMID: 28842295

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Project description:Background: Ischemia reperfusion injury (IRI) plays a major role in solid organ transplantation. The length of warm ischemia time is critical for the extent of tissue damage in renal IRI. In this experimental study we hypothesized that local release of labile heme in renal tissue is triggered by the duration of warm ischemia (15 vs. 45 min IRI) and mediates complement activation, cytokine release, and inflammation. Methods: To induce IRI, renal pedicle clamping was performed in male C57BL/6 mice for short (15 min) or prolonged (45 min) time periods. Two and 24 h after experimental ischemia tissue injury labile heme levels in the kidney were determined with an apo-horseradish peroxidase assay. Moreover, renal injury, cytokines, and C5a and C3a receptor (C5aR, C3aR) expression were determined by histology, immunohistochemistry and qPCR, respectively. In addition, in vitro studies stimulating bone marrow-derived macrophages with LPS and the combination of LPS and heme were performed and cytokine expression was measured. Results: Inflammation and local tissue injury correlated with the duration of warm ischemia time. Labile heme concentrations in renal tissue were significantly higher after prolonged (45 min) as compared to short (15 min) IRI. Notably, expression of the inducible heme-degrading enzyme heme oxygenase-1 (HO-1) was up-regulated in kidneys after prolonged, but not after short IRI. C5aR, the pro-inflammatory cytokines IL-6 and TNF-α as well as pERK were up-regulated after prolonged, but not after short ischemia times. Consecutively, neutrophil infiltration and up-regulation of pro-fibrotic cytokines such as CTGF and PAI were more pronounced in prolonged IRI in comparison to short IRI. In vitro stimulation of macrophages with LPS revealed that IL-6 expression was enhanced in the presence of heme. Finally, administration of the heme scavenger human serum albumin (HSA) reduced the expression of pro-inflammatory cytokines, C3a receptor and improved tubular function indicated by enhanced alpha 1 microglobulin (A1M) absorption after IRI. Conclusions: Our data show that prolonged duration of warm ischemia time increased labile heme levels in the kidney, which correlates with IRI-dependent inflammation and up-regulation of anaphylatoxin receptor expression.

Project description:BackgroundIschemia-reperfusion injury (IRI) is an important cause of graft dysfunction post-liver transplantation, where donor liver with severe steatosis is more sensitive to IRI. Liver IRI involves ferroptosis and can be alleviated by heme oxygenase-1-modified bone marrow mesenchymal stem cells (HO-1/BMMSCs).AimsTo explore the role and mechanism of HO-1/BMMSCs in severe steatotic liver IRI.MethodsA severe steatotic liver IRI rat model and a hypoxia/reoxygenation (H/R) of severe steatosis hepatocyte model were established. Liver and hepatocyte damage was evaluated via liver histopathology and cell activity. Ferroptosis was evaluated through ferroptosis indexes. Nuclear factor erythroid 2-related factor 2 (Nrf2) was knocked down in severe steatotic hepatocytes. The role of Nrf2 and AMPK in HO-1/BMMSC inhibition of ferroptosis was examined using the AMP-activated protein kinase (AMPK) pathway inhibitor Compound C.ResultsThe HO-1/BMMSCs alleviated severe steatotic liver IRI and ferroptosis. HO-1/BMMSCs promoted ferritin heavy chain 1(FTH1), Nrf2, and phosphorylated (p)-AMPK expression in the H/R severe steatotic hepatocytes. Nrf2 knockdown decreased FTH1 expression levels but did not significantly affect p-AMPK expression levels. The protective effect of HO-1/BMMSCs against H/R injury in severe steatotic hepatocytes and the inhibitory effect on ferroptosis were reduced. Compound C decreased p-AMPK, Nrf2, and FTH1 expression levels, weakened the HO-1/BMMSC protective effect against severe steatotic liver IRI and H/R-injured severe steatotic hepatocytes, and reduced the inhibition of ferroptosis.ConclusionsFerroptosis was involved in HO-1/BMMSC reduction of severe steatotic liver IRI. HO-1/BMMSCs protected against severe steatotic liver IRI by inhibiting ferroptosis through the AMPK-Nrf2-FTH1 pathway. HO-1/BMMSCs activate AMPK, which activates Nrf2, promotes its nuclear transcription, then promotes the expression of its downstream protein FTH1, thereby inhibiting ferroptosis and attenuating severe steatotic liver IRI in rats. Glu: glutamic acid; Cys: cystine; GSH: glutathione; GPX4: glutathione peroxidase 4; HO-1/BMMSCs: HO-1-modified BMMSCs; Fer-1: ferrostatin-1; DFO: deferoxamine; FTH1: ferritin heavy chain1; p-AMPK: phosphorylated AMP-activated protein kinase; Nrf2: nuclear factor erythroid 2-related factor 2; IRI: ischemia-reperfusion injury; MCD: methionine-choline deficiency.

Project description:Hypercapnic acidosis (HCA) has protective effects in animal models of acute lung injury, but the mechanism underlying the effect of HCA is unclear. Heme oxygenase-1 (HO-1) is an antioxidant enzyme that protects tissue from inflammation injury. We investigated whether HO-1 contributes to the protective effects of HCA in ischemia-reperfusion (IR)-induced lung injury. Typical acute lung injury in rats was successfully induced by 40 min of ischemia and 90 min of reperfusion in an isolated perfused lung model. The rat lungs were randomly assigned to the control group, IR group or IR + HCA group with or without zinc protoporphyrin IX (ZnPP), an HO-1 activity inhibitor. At the end of the experiment, bronchoalveolar lavage fluid (BALF) and lung tissues were collected to evaluate the degree of lung injury. In in vitro experiments, HO-1 siRNA transfected A549 cells were exposed to a normoxic or hypoxia-reoxygenation (H/R) environment in the presence or absence of HCA. IR caused significant increases in the pulmonary arterial pressure, lung weight to body weight and wet/dry ratios, lung weight gain, capillary filtration coefficient, lung injury scores, neutrophil infiltration, and concentrations of protein and TNF-? in the BALF. IR also induced degradation of inhibitor of nuclear factor (NF)-?B-?, increased I?B kinase (IKK)-? phosphorylation and nuclear translocation of NF-?B, and up-regulated HO-1 expression and activity. Furthermore, IR decreased Bcl-2 protein expression and increased the number of active caspase-3 stained cells. HCA treatment enhanced HO-1 expression and activity, and accordingly reduced IKK-NF-?B signaling, inhibited apoptosis, and significantly attenuated IR-induced changes. Treatment with ZnPP partially blocked the protective effect of HCA. In addition, HO-1 siRNA significantly reversed HCA-mediated inhibition of NF-?B signaling in A549 cells subjected to H/R. In conclusion, the protective effect of HCA in IR lung injury in rats was mediated in part by the anti-inflammatory and anti-apoptotic action of HO-1.

Project description:BACKGROUND:Stroke is the leading cause of long-term motor disability and cognitive impairment. Recently, neurogenesis has become an attractive strategy for the chronic recovery of stroke. It is important to understand the molecular mechanism that promotes neural stem cell (NSC) neurogenesis for future NSC-based therapies. Our previous study showed that Momordica charantia polysaccharides (MCPs) exerted neuroprotective effects on stroke via their anti-oxidant and anti-inflammation activities. However, it remains unknown whether MCPs promote NSC neurogenesis after cerebral ischemic/reperfusion injury (IRI). METHODS:We investigated MCPs' function in differentiation of neural stem cells (NSCs) in vivo and in vitro experiments. Based on a middle cerebral artery occlusion (MCAO) rat model, the effect of MCPs on neuronal differentiation after MCAO was analyzed. Primary NSCs and neural stem cell line C17.2 were cultured and subjected to glutamate stimulation to establish the cell model of IRI. We evaluated the effect of MCPs on NSC differentiation in IRI cell model by Western blot and immunofluorescence staining. The SIRT1 activity of NSCs post glutamate stimulation was also evaluated by CELL SIRT1 COLORIMETRY ASSAY KIT. In addition, molecular mechanism was clarified by employing the activator and inhibitor of SIRT1. RESULTS:MCPs had no effects on the differentiation of neural stem cells under physiological conditions while shifted NSC differentiation potential from the gliogenic to neurogenic lineage under pathological conditions. Activation of SIRT1 with MCPs was responsible for the neuronal differentiation of C17.2-NSCs. The neuronal differentiation effect of MCPs was attributed to upregulation SIRT1-mediated deacetylation of ?-catenin. MCP-induced deacetylation via SIRT1 promoted nuclear accumulation of ?-catenin in NSCs. CONCLUSION:Our findings indicate that the deacetylation of ?-catenin by SIRT1 represents a critical mechanism of action of MCPs in promoting NSC neuronal differentiation. It provides an improved understanding of molecular mechanism underlying neuroprotective effects of MCPs in IRI, indicating its potential role on treating ischemic stroke especially chronic recovery.

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Macrophage heme oxygenase-1-SIRT1-p53 axis regulates sterile inflammation in liver ischemia-reperfusion injury.

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Macrophage heme oxygenase-1-SIRT1-p53 axis regulates sterile inflammation in liver ischemia-reperfusion injury.

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