Activation of the PTEN-PI3K-mTORC1 pathway consolidates metabolic programs that sustain cancer cell growth and proliferation. Here we show that mechanistic target of rapamycin complex 1 (mTORC1) ...regulates polyamine dynamics, a metabolic route that is essential for oncogenicity. By using integrative metabolomics in a mouse model and human biopsies of prostate cancer, we identify alterations in tumours affecting the production of decarboxylated S-adenosylmethionine (dcSAM) and polyamine synthesis. Mechanistically, this metabolic rewiring stems from mTORC1-dependent regulation of S-adenosylmethionine decarboxylase 1 (AMD1) stability. This novel molecular regulation is validated in mouse and human cancer specimens. AMD1 is upregulated in human prostate cancer with activated mTORC1. Conversely, samples from a clinical trial with the mTORC1 inhibitor everolimus exhibit a predominant decrease in AMD1 immunoreactivity that is associated with a decrease in proliferation, in line with the requirement of dcSAM production for oncogenicity. These findings provide fundamental information about the complex regulatory landscape controlled by mTORC1 to integrate and translate growth signals into an oncogenic metabolic program.
Chronic cholestasis is associated with increased bacterial infections and sepsis resulting in higher mortality in humans. In the current study, we investigated the relevance of gp130-dependent ...pathways after bile duct ligation (BDL). BDL was performed in conditional gp130 knockout (loxP/Cre system) mice and respective controls. Liver injury, regulation of the acute phase response, and the impact on survival and bacterial infections were determined. Acute BDL resulted in increased IL-6 levels, Stat3 activation, and an increase in acute-phase proteins (serum-amyloid-A SAA), which was blocked in gp130-deleted animals. In addition, the antimicrobial gene hepcidin was regulated in a gp130-dependent manner after BDL. During chronic cholestasis Stat3 activation was dramatically reduced, while high SAA levels were maintained via gp130-dependent signaling. Inhibition of gp130-dependent pathways resulted in higher mortality and liver damage, which was associated with higher infiltration of immune-activated cells and increased germ number in the liver. In conclusion, during acute and chronic cholestasis, the gp130 system is essential for controlling the acute-phase response. Lack of gp130 expression results in pronounced bacterial growth in bile and liver after BDL, which is associated with higher mortality. Activation of gp130-dependent pathways after BDL is essential and appears to be a therapeutic target during cholestasis.
TGF-β2 (TGF-β, transforming growth factor beta), the less-investigated sibling of TGF-β1, is deregulated in rodent and human liver diseases. Former data from bile duct ligated and MDR2 knockout (KO) ...mouse models for human cholestatic liver disease suggested an involvement of TGF-β2 in biliary-derived liver diseases.
As we also found upregulated
in liver tissue of patients with primary sclerosing cholangitis (PSC) and primary biliary cholangitis (PBC), we now fathomed the positive prospects of targeting TGF-β2 in early stage biliary liver disease using the MDR2-KO mice. Specifically, the influence of
silencing on the fibrotic and inflammatory niche was analysed on molecular, cellular and tissue levels.
-induced expression of fibrotic genes in cholangiocytes and hepatic stellate cellswas detected.
expression in MDR2-KO mice was blunted using
-directed antisense oligonucleotides (AON). Upon AON treatment, reduced collagen deposition, hydroxyproline content and αSMA expression as well as induced
expression reflected a significant reduction of fibrogenesis without adverse effects on healthy livers. Expression analyses of fibrotic and inflammatory genes revealed AON-specific regulatory effects on
,
,
,
and
expression. Further, AON treatment of MDR2-KO mice increased tissue infiltration by F4/80-positive cells including eosinophils, whereas the number of CD45-positive inflammatory cells decreased. In line,
and CD45 expression correlated positively in PSC/PBC patients and localised in similar areas of the diseased liver tissue.
Taken together, our data suggest a new mechanistic explanation for amelioration of fibrogenesis by TGF-β2 silencing and provide a direct rationale for TGF-β2-directed drug development.
Haematopoietic stem and progenitor cells (HSPC) are tightly regulated in a balance between self-renewal and differentiation and undergo rapid expansion in response to infection. In the context of ...acute bacterial infection, the hematopoietic system needs energy to drive the immune response necessary for host survival. In an adult human, bone marrow adipose tissue accounts for up to 70% of bone marrow and we have previously shown that free fatty acids are transferred from adipocytes to AML blasts providing the AML blasts more energy for proliferation and survival (Shafat, Blood 2017). Therefore, we hypothesis that during an acute bacterial infection, the HSPC population uptakes free fatty acids (FFA) to support and sustain the rapid response to bacterial infection. To determine if HSPC acquired FFA we infected C57BL/6 with Salmonella Typhimurium (S.typhimurium) or treated C57BL/6 with lipopolysaccharide (LPS) for 2 and 16 hours. Using both models we show that the HSPC populations (HSC (haematopoietic stem cell), MPP (multipotent progenitor) and LSK (LN-, CD117+, Sca1+)) all accumulate FFA as measured by staining with neutral lipid-specific BODIPY 493/503 dye. To determine if this was uptake or accumulation of fatty acid within the HSPC of animals infected with S.typhimurium we used BODIPY®-dodecanoic acid fluorescent fatty acid to trace FFA uptake. HSPC populations from animals treated with LPS had a higher uptake of BODIPY®-dodecanoic acid when compared to the control treated mice. To assess metabolic changes within HSC population in response to infection, cells were sorted (LN-, CD117+, Sca1+, CD48-, CD150+ and CD34+) and analysed for gene expression changes. In HSC, increases were observed in several metabolic genes including FABP4, CD36 and CPT1 in response to S.typhimurium as well as LPS at 16 hours post stimulation. Moreover, following cell sorting, the LSK (LN-, CD117+, Sca1+) population were assessed for metabolic activity for different substrates. The LSK population from LPS treated animals had an increased dependency on beta oxidation when compared to the control. Moreover, etoximir (a beta-oxidation inhibitor) inhibited LSK basal metabolism and reduced LSK expansion (as measured by colony forming cell assay and Ki-67 staining) in animals treated with LPS. Our data indicates that infection drives uptake of FFA in different HSPC populations. The FFA are metabolised by beta oxidation which provides the energy for the expansion of leukocytes.
Bowles:Abbvie: Research Funding; Janssen: Research Funding. Rushworth:Abbvie: Research Funding; Janssen: Research Funding.
Cholestasis comprises aetiologically heterogeneous conditions characterized by accumulation of bile acids in the liver that actively contribute to liver damage. Sirtuin 1 (SIRT1) regulates liver ...regeneration and bile acid metabolism by modulating farnesoid X receptor (FXR); we here investigate its role in cholestatic liver disease. We determined SIRT1 expression in livers from patients with cholestatic disease, in two experimental models of cholestasis, as well as in human and murine liver cells in response to bile acid loading. SIRT1‐overexpressing (SIRToe) and hepatocyte‐specific SIRT1‐KO (knockout) mice (SIRThep–/–) were subjected to bile duct ligation (BDL) and were fed with a 0.1% DDC (3,5‐diethoxycarbonyl‐1,4‐dihydrocollidine) diet to determine the biological relevance of SIRT1 during cholestasis. The effect of NorUDCA (24‐norursodeoxycholic acid) was tested in BDL/SIRToe mice. We found that SIRT1 was highly expressed in livers from cholestatic patients, mice after BDL, and Mdr2 knockout mice (Mdr2–/–) animals. The detrimental effects of SIRT1 during cholestasis were validated in vivo and in vitro. SIRToe mice showed exacerbated parenchymal injury whereas SIRThep–/– mice evidenced a moderate improvement after BDL and 0.1% DDC feeding. Likewise, hepatocytes isolated from SIRToe mice showed increased apoptosis in response to bile acids, whereas a significant reduction was observed in SIRThep–/– hepatocytes. Importantly, the decrease, but not complete inhibition, of SIRT1 exerted by norUDCA treatment correlated with pronounced improvement in liver parenchyma in BDL/SIRToe mice. Interestingly, both SIRT1 overexpression and hepatocyte‐specific SIRT1 depletion correlated with inhibition of FXR, whereas modulation of SIRT1 by NorUDCA associated with restored FXR signaling. Conclusion: SIRT1 expression is increased during human and murine cholestasis. Fine‐tuning expression of SIRT1 is essential to protect the liver from cholestatic liver damage.
Methionine adenosyltransferase 1A (MAT1A) and glycine N‐methyltransferase (GNMT) are the primary genes involved in hepatic S‐adenosylmethionine (SAMe) synthesis and degradation, respectively. Mat1a ...ablation in mice induces a decrease in hepatic SAMe, activation of lipogenesis, inhibition of triglyceride (TG) release, and steatosis. Gnmt‐deficient mice, despite showing a large increase in hepatic SAMe, also develop steatosis. We hypothesized that as an adaptive response to hepatic SAMe accumulation, phosphatidylcholine (PC) synthesis by way of the phosphatidylethanolamine (PE) N‐methyltransferase (PEMT) pathway is stimulated in Gnmt−/− mice. We also propose that the excess PC thus generated is catabolized, leading to TG synthesis and steatosis by way of diglyceride (DG) generation. We observed that Gnmt−/− mice present with normal hepatic lipogenesis and increased TG release. We also observed that the flux from PE to PC is stimulated in the liver of Gnmt−/− mice and that this results in a reduction in PE content and a marked increase in DG and TG. Conversely, reduction of hepatic SAMe following the administration of a methionine‐deficient diet reverted the flux from PE to PC of Gnmt−/− mice to that of wildtype animals and normalized DG and TG content preventing the development of steatosis. Gnmt−/− mice with an additional deletion of perilipin2, the predominant lipid droplet protein, maintain high SAMe levels, with a concurrent increased flux from PE to PC, but do not develop liver steatosis. Conclusion: These findings indicate that excess SAMe reroutes PE towards PC and TG synthesis and lipid sequestration. (Hepatology 2013;58:1296–1305)
Background & Aims Disruption of the nuclear factor-κB ( NF-κB ) essential modulator ( NEMO ) in hepatocytes of mice (NEMOΔhepa mice) results in spontaneous liver apoptosis and chronic liver disease ...involving inflammation, steatosis, fibrosis, and development of hepatocellular carcinoma. Activation of caspase-8 (Casp8) initiates death receptor-mediated apoptosis. We investigated the pathogenic role of this protease in NEMOΔhepa mice or after induction of acute liver injury. Methods We created mice with conditional deletion of Casp8 in hepatocytes (Casp8Δhepa ) and Casp8Δhepa NEMOΔhepa double knockout mice. Acute liver injury was induced by Fas-activating antibodies, lipopolysaccharides, or concanavalin A. Spontaneous hepatocarcinogenesis was monitored by magnetic resonance imaging. Results Hepatocyte-specific deletion of Casp8 protected mice from induction of apoptosis and liver injury by Fas or lipopolysaccharides but increased necrotic damage and reduced survival times of mice given concanavalin A. Casp8Δhepa NEMOΔhepa mice were protected against steatosis and hepatocarcinogenesis but had a separate, spontaneous phenotype that included massive liver necrosis, cholestasis, and biliary lesions. The common mechanism by which inactivation of Casp8 induces liver necrosis in both injury models involves the formation of protein complexes that included the adaptor protein Fas-associated protein with death domain and the kinases receptor-interacting protein (RIP) 1 and RIP3—these have been shown to be required for programmed necrosis. We demonstrated that hepatic RIP1 was proteolytically cleaved by Casp8, whereas Casp8 inhibition resulted in accumulation of RIP complexes and subsequent liver necrosis. Conclusions Inhibition of Casp8 protects mice from hepatocarcinogenesis following chronic liver injury mediated by apoptosis of hepatocytes but can activate RIP-mediated necrosis in an inflammatory environment.
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