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Liver macrophages fulfill various homeostatic functions and represent an essential line of defense against pathogenic insults. However, it remains unclear whether a history of infectious disease in the liver instructs long-term alterations to the liver macrophage compartment., info:eu-repo/semantics/published

AimLow-grade inflammation is the hallmark of non-alcoholic fatty liver diseases (NAFLD) and non-alcoholic steatohepatitis (NASH). The leakage of microbiota-derived products can contribute to liver inflammation during NAFLD/NASH development. Here, we assessed the roles of gut microbial DNA-containing extracellular vesicles (mEVs) in regulating liver cellular abnormalities in the course of NAFLD/NASH.MethodsWe performed studies with Vsig4-/- , C3-/- , cGAS-/- , and their wild-type littermate mice. Vsig4+ macrophage population and bacterial DNA abundance were examined in both mouse and human liver by either flow cytometric or immunohistochemistry analysis. Gut mEVs were adoptively transferred into Vsig4-/- , C3-/- , cGAS-/- , or littermate WT mice, and hepatocyte inflammation and HSC fibrogenic activation were measured in these mice.ResultsNon-alcoholic fatty liver diseases and non-alcoholic steatohepatitis development was concomitant with a diminished liver Vsig4+ macrophage population and a marked bacterial DNA enrichment in both hepatocytes and HSCs. In the absence of Vsig4+ macrophages, gut mEVs translocation led to microbial DNA accumulation in hepatocytes and HSCs, resulting elevated hepatocyte inflammation and HSC fibrogenic activation. In contrast, in lean WT mice, Vsig4+ macrophages remove gut mEVs from bloodstream through a C3-dependent opsonization mechanism and prevent the infiltration of gut mEVs into hepatic cells. Additionally, Vsig4-/- mice more quickly developed significant liver steatosis and fibrosis than WT mice after Western diet feeding. In vitro treatment with NASH mEVs triggered hepatocyte inflammation and HSC fibrogenic activation. Microbial DNAs are key cargo for the effects of gut mEVs by activating cGAS/STING.ConclusionAccumulation of microbial DNAs fuels the development of NAFLD/NASH-associated liver abnormalities.

BackgroundObesity, non-alcoholic fatty liver disease (NAFLD) and its more advanced form non-alcoholic steatohepatitis (NASH) are important causes of morbidity and mortality worldwide. Bile acid dysregulation is a pivotal part in their pathogenesis. The aim of this study was to evaluate the bile acid sequestrant colesevelam in a microbiome-humanized mouse model of diet-induced obesity and steatohepatitis.MethodsGerm-free C57BL/6 mice were associated with stool from patients with NASH and subjected to 20 weeks of Western diet feeding with and without colesevelam.ResultsColesevelam reduced Western diet-induced body and liver weight gain in microbiome-humanized mice compared with controls. It ameliorated Western diet-induced hepatic inflammation, steatosis, fibrosis and insulin resistance. Colesevelam increased de novo bile acid synthesis and decreased hepatic cholesterol content in microbiome-humanized mice fed a Western diet. It further induced the gene expression of the antimicrobials Reg3g and Reg3b in the distal small intestine and decreased plasma levels of LPS.ConclusionsColesevelam ameliorates Western diet-induced steatohepatitis and obesity in microbiome-humanized mice.

AimLow-grade inflammation is the hallmark of non-alcoholic fatty liver diseases (NAFLD) and non-alcoholic steatohepatitis (NASH). The leakage of microbiota-derived products can contribute to liver inflammation during NAFLD/NASH development. Here, we assessed the roles of gut microbial DNA-containing extracellular vesicles (mEVs) in regulating liver cellular abnormalities in the course of NAFLD/NASH.MethodsWe performed studies with Vsig4-/- , C3-/- , cGAS-/- , and their wild-type littermate mice. Vsig4+ macrophage population and bacterial DNA abundance were examined in both mouse and human liver by either flow cytometric or immunohistochemistry analysis. Gut mEVs were adoptively transferred into Vsig4-/- , C3-/- , cGAS-/- , or littermate WT mice, and hepatocyte inflammation and HSC fibrogenic activation were measured in these mice.ResultsNon-alcoholic fatty liver diseases and non-alcoholic steatohepatitis development was concomitant with a diminished liver Vsig4+ macrophage population and a marked bacterial DNA enrichment in both hepatocytes and HSCs. In the absence of Vsig4+ macrophages, gut mEVs translocation led to microbial DNA accumulation in hepatocytes and HSCs, resulting elevated hepatocyte inflammation and HSC fibrogenic activation. In contrast, in lean WT mice, Vsig4+ macrophages remove gut mEVs from bloodstream through a C3-dependent opsonization mechanism and prevent the infiltration of gut mEVs into hepatic cells. Additionally, Vsig4-/- mice more quickly developed significant liver steatosis and fibrosis than WT mice after Western diet feeding. In vitro treatment with NASH mEVs triggered hepatocyte inflammation and HSC fibrogenic activation. Microbial DNAs are key cargo for the effects of gut mEVs by activating cGAS/STING.ConclusionAccumulation of microbial DNAs fuels the development of NAFLD/NASH-associated liver abnormalities.

BackgroundObesity, non-alcoholic fatty liver disease (NAFLD) and its more advanced form non-alcoholic steatohepatitis (NASH) are important causes of morbidity and mortality worldwide. Bile acid dysregulation is a pivotal part in their pathogenesis. The aim of this study was to evaluate the bile acid sequestrant colesevelam in a microbiome-humanized mouse model of diet-induced obesity and steatohepatitis.MethodsGerm-free C57BL/6 mice were associated with stool from patients with NASH and subjected to 20 weeks of Western diet feeding with and without colesevelam.ResultsColesevelam reduced Western diet-induced body and liver weight gain in microbiome-humanized mice compared with controls. It ameliorated Western diet-induced hepatic inflammation, steatosis, fibrosis and insulin resistance. Colesevelam increased de novo bile acid synthesis and decreased hepatic cholesterol content in microbiome-humanized mice fed a Western diet. It further induced the gene expression of the antimicrobials Reg3g and Reg3b in the distal small intestine and decreased plasma levels of LPS.ConclusionsColesevelam ameliorates Western diet-induced steatohepatitis and obesity in microbiome-humanized mice.

Background & aimsAlthough Ikaros (IKZF1) is a well-established transcriptional regulator in leukocyte lymphopoiesis and differentiation, its role in myeloid innate immune responses remains unclear. Sirtuin 1 (SIRT1) is a histone/protein deacetylase involved in cellular senescence, inflammation, and stress resistance. Whether SIRT1 signaling is essential in myeloid cell activation remains uncertain, while the molecular communication between Ikaros and SIRT1, two major transcriptional regulators, has not been studied.MethodsWe undertook molecular and functional studies to interrogate the significance of the myeloid Ikaros-SIRT1 axis in innate immune activation and whether it may serve as a homeostatic sentinel in human liver transplant recipients (hepatic biopsies) and murine models of sterile hepatic inflammation (liver warm ischemia-reperfusion injury in wild-type, myeloid-specific Sirt1-knockout, and CD11b-DTR mice) as well as primary bone marrow-derived macrophage (BMM) cultures (Ikaros silencing vs. overexpression).ResultsIn our clinical study, we identified increased post-reperfusion hepatic Ikaros levels, accompanied by augmented inflammasome signaling yet depressed SIRT1, as a mechanism of hepatocellular damage in liver transplant recipients. In our experimental studies, we identified infiltrating macrophages as the major source of Ikaros in IR-stressed mouse livers. Then, we demonstrated that Ikaros-regulated pyroptosis - induced by canonical inflammasome signaling in BMM cultures - was SIRT1 dependent. Consistent with the latter, myeloid-specific Ikaros signaling augmented hepatic pyroptosis to aggravate pro-inflammatory responses in vivo by negatively regulating SIRT1 in an AMPK-dependent manner. Finally, myeloid-specific SIRT1 was required to suppress pyroptosis, pro-inflammatory phenotype, and ultimately mitigate hepatocellular injury in ischemia-stressed murine livers.ConclusionThese findings identify the Ikaros-SIRT1 axis as a novel mechanistic

Metabolic impairments and liver and adipose depots alterations were reported in subjects with Alzheimer's disease (AD), highlighting the role of the liver-adipose-tissue-brain axis in AD pathophysiology. The gut microbiota might play a modulating role. We investigated the alterations to the liver and white/brown adipose tissues (W/BAT) and their relationships with serum and gut metabolites and gut bacteria in a 3xTg mouse model during AD onset (adulthood) and progression (aging) and the impact of high-fat diet (HFD) and intranasal insulin (INI). Glucose metabolism (18FDG-PET), tissue radiodensity (CT), liver and W/BAT histology, BAT-thermogenic markers were analyzed. 16S-RNA sequencing and mass-spectrometry were performed in adult (8 months) and aged (14 months) 3xTg-AD mice with a high-fat or control diet. Generalized and HFD resistant deficiency of lipid accumulation in both liver and W/BAT, hypermetabolism in WAT (adulthood) and BAT (aging), abnormal cytokine-hormone profiles, and liver inflammation were observed in 3xTg mice; INI could antagonize all these alterations. Specific gut microbiota-metabolome profiles correlated with a significant disruption of the gut-microbiota-liver-adipose axis in AD mice. In conclusion, fat dystrophy in liver and adipose depots contributes to AD progression, and associates with altered profiles of the gut microbiota, which candidates as an appealing early target for preventive intervention.

[ES] Las enfermedades hepáticas crónicas (EPC) constituyen un problema mundial con una demanda creciente de atención médica. La actividad inflamatoria y la fibrosis derivada son características histopatológicas comunes y relevantes en la EPC. Tradicionalmente, la biopsia hepática se ha considerado el gold standard para su diagnóstico y la clasificación de su nivel gravedad. La evaluación histológica establecida se basa en sistemas de puntuación semicuantitativos basados ¿¿en criterios bien definidos, aunque interpretativos relacionados con la gravedad de la EPC subyacente. Esta evaluación categórica subjetiva se basa en cambios arquitectónicos y celulares, pero no en alteraciones patológicas continuas numéricas. Por lo tanto, los informes de biopsia estándar son propensos a una gran variabilidad, lo que limita una evaluación crítica de las características histológicas en el entorno de la investigación y reduce la reproducibilidad de los resultados. Por otra parte, la biopsia hepática tiene otras limitaciones bien conocidas, ya que la técnica es un procedimiento costoso e invasivo, no bien aceptado por los pacientes y que no se puede usar libremente para el seguimiento. Además, es propenso a errores de submuestreo y sufre de variabilidad interobservador en la puntuación patológica. Más recientemente, se ha propuesto el uso del análisis de imagen digital asistido por computadora (DIA) de las secciones histológicas obtenidas en diferentes etiologías de EPC para una cuantificación más precisa de algunas características histológicas como la esteatosis y la fibrosis., [EN] Chronic liver disease (CLD) is a global problem with a growing demand for medical care. Inflammatory activity and derived fibrosis are common and relevant histopathological features in CLD. Traditionally, liver biopsy has been considered the gold standard for its diagnosis and classification of its level of severity. The established histological evaluation is based on semi-quantitative scoring systems based on well-defined but interpretive criteria related to the severity of the underlying CLD. This subjective categorical evaluation is based on architectural and cellular changes, but not on numerical continuous pathological changes. Therefore, standard biopsy reports are prone to great variability, limiting critical evaluation of histological features in the research setting and reducing reproducibility of results. On the other hand, liver biopsy has other well-known limitations, since the technique is an expensive and invasive procedure, little accepted by patients and cannot be freely used for follow-up. Furthermore, it is prone to subsampling errors and suffers from interobserver variability in the pathological score. More recently, the use of computer-aided digital image analysis (DIA) of histological sections obtained in different EPC etiologies has been proposed for a more precise quantification of some histological characteristics such as steatosis and fibrosis.

[ES] Las enfermedades hepáticas crónicas (EPC) constituyen un problema mundial con una demanda creciente de atención médica. La actividad inflamatoria y la fibrosis derivada son características histopatológicas comunes y relevantes en la EPC. Tradicionalmente, la biopsia hepática se ha considerado el gold standard para su diagnóstico y la clasificación de su nivel gravedad. La evaluación histológica establecida se basa en sistemas de puntuación semicuantitativos basados ¿¿en criterios bien definidos, aunque interpretativos relacionados con la gravedad de la EPC subyacente. Esta evaluación categórica subjetiva se basa en cambios arquitectónicos y celulares, pero no en alteraciones patológicas continuas numéricas. Por lo tanto, los informes de biopsia estándar son propensos a una gran variabilidad, lo que limita una evaluación crítica de las características histológicas en el entorno de la investigación y reduce la reproducibilidad de los resultados. Por otra parte, la biopsia hepática tiene otras limitaciones bien conocidas, ya que la técnica es un procedimiento costoso e invasivo, no bien aceptado por los pacientes y que no se puede usar libremente para el seguimiento. Además, es propenso a errores de submuestreo y sufre de variabilidad interobservador en la puntuación patológica. Más recientemente, se ha propuesto el uso del análisis de imagen digital asistido por computadora (DIA) de las secciones histológicas obtenidas en diferentes etiologías de EPC para una cuantificación más precisa de algunas características histológicas como la esteatosis y la fibrosis., [EN] Chronic liver disease (CLD) is a global problem with a growing demand for medical care. Inflammatory activity and derived fibrosis are common and relevant histopathological features in CLD. Traditionally, liver biopsy has been considered the gold standard for its diagnosis and classification of its level of severity. The established histological evaluation is based on semi-quantitative scoring systems based on well-defined but interpretive criteria related to the severity of the underlying CLD. This subjective categorical evaluation is based on architectural and cellular changes, but not on numerical continuous pathological changes. Therefore, standard biopsy reports are prone to great variability, limiting critical evaluation of histological features in the research setting and reducing reproducibility of results. On the other hand, liver biopsy has other well-known limitations, since the technique is an expensive and invasive procedure, little accepted by patients and cannot be freely used for follow-up. Furthermore, it is prone to subsampling errors and suffers from interobserver variability in the pathological score. More recently, the use of computer-aided digital image analysis (DIA) of histological sections obtained in different EPC etiologies has been proposed for a more precise quantification of some histological characteristics such as steatosis and fibrosis.

Background & aimsHow benign liver steatosis progresses to nonalcoholic steatohepatitis (NASH), fibrosis, and hepatocellular carcinoma (HCC) remains elusive. NASH progression entails diverse pathogenic mechanisms and relies on complex cross-talk between multiple tissues such as the gut, adipose tissues, liver, and the brain. Using a hyperphagic mouse fed with a Western diet (WD), we aimed to elucidate the cross-talk and kinetics of hepatic and extrahepatic alterations during NASH-HCC progression, as well as regression.MethodsHyperphagic mice lacking a functional Alms1 gene (Foz/Foz) and wild-type littermates were fed WD or standard chow for 12 weeks for NASH/fibrosis and for 24 weeks for HCC development. NASH regression was modeled by switching back to normal chow after NASH development.ResultsFoz+WD mice were steatotic within 1 to 2 weeks, developed NASH by 4 weeks, and grade 3 fibrosis by 12 weeks, accompanied by chronic kidney injury. Foz+WD mice that continued on WD progressed to cirrhosis and HCC within 24 weeks and had reduced survival as a result of cardiac dysfunction. However, NASH mice that were switched to normal chow showed NASH regression, improved survival, and did not develop HCC. Transcriptomic and histologic analyses of Foz/Foz NASH liver showed strong concordance with human NASH. NASH was preceded by an early disruption of gut barrier, microbial dysbiosis, lipopolysaccharide leakage, and intestinal inflammation. This led to acute-phase liver inflammation in Foz+WD mice, characterized by neutrophil infiltration and increased levels of several chemokines/cytokines. The liver cytokine/chemokine profile evolved as NASH progressed, with subsequent predominance by monocyte recruitment.ConclusionsThe Foz+WD model closely mimics the pathobiology and gene signature of human NASH with fibrosis and subsequent HCC. Foz+WD mice provide a robust and relevant preclinical model of NASH, NASH-associated HCC, chronic kidney injury, and heart failure.

[ES] Las enfermedades hepáticas crónicas (EPC) constituyen un problema mundial con una demanda creciente de atención médica. La actividad inflamatoria y la fibrosis derivada son características histopatológicas comunes y relevantes en la EPC. Tradicionalmente, la biopsia hepática se ha considerado el gold standard para su diagnóstico y la clasificación de su nivel gravedad. La evaluación histológica establecida se basa en sistemas de puntuación semicuantitativos basados ¿¿en criterios bien definidos, aunque interpretativos relacionados con la gravedad de la EPC subyacente. Esta evaluación categórica subjetiva se basa en cambios arquitectónicos y celulares, pero no en alteraciones patológicas continuas numéricas. Por lo tanto, los informes de biopsia estándar son propensos a una gran variabilidad, lo que limita una evaluación crítica de las características histológicas en el entorno de la investigación y reduce la reproducibilidad de los resultados. Por otra parte, la biopsia hepática tiene otras limitaciones bien conocidas, ya que la técnica es un procedimiento costoso e invasivo, no bien aceptado por los pacientes y que no se puede usar libremente para el seguimiento. Además, es propenso a errores de submuestreo y sufre de variabilidad interobservador en la puntuación patológica. Más recientemente, se ha propuesto el uso del análisis de imagen digital asistido por computadora (DIA) de las secciones histológicas obtenidas en diferentes etiologías de EPC para una cuantificación más precisa de algunas características histológicas como la esteatosis y la fibrosis., [EN] Chronic liver disease (CLD) is a global problem with a growing demand for medical care. Inflammatory activity and derived fibrosis are common and relevant histopathological features in CLD. Traditionally, liver biopsy has been considered the gold standard for its diagnosis and classification of its level of severity. The established histological evaluation is based on semi-quantitative scoring systems based on well-defined but interpretive criteria related to the severity of the underlying CLD. This subjective categorical evaluation is based on architectural and cellular changes, but not on numerical continuous pathological changes. Therefore, standard biopsy reports are prone to great variability, limiting critical evaluation of histological features in the research setting and reducing reproducibility of results. On the other hand, liver biopsy has other well-known limitations, since the technique is an expensive and invasive procedure, little accepted by patients and cannot be freely used for follow-up. Furthermore, it is prone to subsampling errors and suffers from interobserver variability in the pathological score. More recently, the use of computer-aided digital image analysis (DIA) of histological sections obtained in different EPC etiologies has been proposed for a more precise quantification of some histological characteristics such as steatosis and fibrosis.

[ES] Las enfermedades hepáticas crónicas (EPC) constituyen un problema mundial con una demanda creciente de atención médica. La actividad inflamatoria y la fibrosis derivada son características histopatológicas comunes y relevantes en la EPC. Tradicionalmente, la biopsia hepática se ha considerado el gold standard para su diagnóstico y la clasificación de su nivel gravedad. La evaluación histológica establecida se basa en sistemas de puntuación semicuantitativos basados ¿¿en criterios bien definidos, aunque interpretativos relacionados con la gravedad de la EPC subyacente. Esta evaluación categórica subjetiva se basa en cambios arquitectónicos y celulares, pero no en alteraciones patológicas continuas numéricas. Por lo tanto, los informes de biopsia estándar son propensos a una gran variabilidad, lo que limita una evaluación crítica de las características histológicas en el entorno de la investigación y reduce la reproducibilidad de los resultados. Por otra parte, la biopsia hepática tiene otras limitaciones bien conocidas, ya que la técnica es un procedimiento costoso e invasivo, no bien aceptado por los pacientes y que no se puede usar libremente para el seguimiento. Además, es propenso a errores de submuestreo y sufre de variabilidad interobservador en la puntuación patológica. Más recientemente, se ha propuesto el uso del análisis de imagen digital asistido por computadora (DIA) de las secciones histológicas obtenidas en diferentes etiologías de EPC para una cuantificación más precisa de algunas características histológicas como la esteatosis y la fibrosis., [EN] Chronic liver disease (CLD) is a global problem with a growing demand for medical care. Inflammatory activity and derived fibrosis are common and relevant histopathological features in CLD. Traditionally, liver biopsy has been considered the gold standard for its diagnosis and classification of its level of severity. The established histological evaluation is based on semi-quantitative scoring systems based on well-defined but interpretive criteria related to the severity of the underlying CLD. This subjective categorical evaluation is based on architectural and cellular changes, but not on numerical continuous pathological changes. Therefore, standard biopsy reports are prone to great variability, limiting critical evaluation of histological features in the research setting and reducing reproducibility of results. On the other hand, liver biopsy has other well-known limitations, since the technique is an expensive and invasive procedure, little accepted by patients and cannot be freely used for follow-up. Furthermore, it is prone to subsampling errors and suffers from interobserver variability in the pathological score. More recently, the use of computer-aided digital image analysis (DIA) of histological sections obtained in different EPC etiologies has been proposed for a more precise quantification of some histological characteristics such as steatosis and fibrosis.

Background & aimsHow benign liver steatosis progresses to nonalcoholic steatohepatitis (NASH), fibrosis, and hepatocellular carcinoma (HCC) remains elusive. NASH progression entails diverse pathogenic mechanisms and relies on complex cross-talk between multiple tissues such as the gut, adipose tissues, liver, and the brain. Using a hyperphagic mouse fed with a Western diet (WD), we aimed to elucidate the cross-talk and kinetics of hepatic and extrahepatic alterations during NASH-HCC progression, as well as regression.MethodsHyperphagic mice lacking a functional Alms1 gene (Foz/Foz) and wild-type littermates were fed WD or standard chow for 12 weeks for NASH/fibrosis and for 24 weeks for HCC development. NASH regression was modeled by switching back to normal chow after NASH development.ResultsFoz+WD mice were steatotic within 1 to 2 weeks, developed NASH by 4 weeks, and grade 3 fibrosis by 12 weeks, accompanied by chronic kidney injury. Foz+WD mice that continued on WD progressed to cirrhosis and HCC within 24 weeks and had reduced survival as a result of cardiac dysfunction. However, NASH mice that were switched to normal chow showed NASH regression, improved survival, and did not develop HCC. Transcriptomic and histologic analyses of Foz/Foz NASH liver showed strong concordance with human NASH. NASH was preceded by an early disruption of gut barrier, microbial dysbiosis, lipopolysaccharide leakage, and intestinal inflammation. This led to acute-phase liver inflammation in Foz+WD mice, characterized by neutrophil infiltration and increased levels of several chemokines/cytokines. The liver cytokine/chemokine profile evolved as NASH progressed, with subsequent predominance by monocyte recruitment.ConclusionsThe Foz+WD model closely mimics the pathobiology and gene signature of human NASH with fibrosis and subsequent HCC. Foz+WD mice provide a robust and relevant preclinical model of NASH, NASH-associated HCC, chronic kidney injury, and heart failure.

Tissue resident memory CD8 T (TRM) cells provide effective tissue surveillance and can respond rapidly to infection due to their strategic location. Within the liver, TRM cells can induce effective protection against liver-stage Plasmodium infection. Recently, members from our group identified a highly immunogenic peptide (named Pb 1) within the putative 60S ribosomal protein L6 of P. berghei ANKA. Experiments conducted and presented in this thesis aimed to assess the suitability of Pb 1 for the induction of endogenous liver TRM cells that confer sterilizing protection in B6 mice. To this end, a series of different immunisation strategies targeting the Pb 1 epitope were implemented and specific CD8 T cell responses were assessed. Results revealed that the number of naive specific CD8 T cell precursors for the Pb 1 epitope was very large. Substantial expansion and formation of specific liver TRM cells was achieved by two different immunisation strategies: i) Single injection with Clec9A mAb plus adjuvant and ii) Prime and trap, both targeting the Pb 1 epitope. While mice vaccinated with Clec9A mAb developed partial protection, almost all mice vaccinated with prime-and-trap targeting Pb 1 were sterilely protected against liver stage challenge. Inflammation favours the formation TRM cells and adjuvants can affect their numbers. Accordingly, a second focus of this thesis sought to investigate how to enhance liver TRM cell formation by using TLR and RIG I like receptors agonists as adjuvants. For this, eight different agonists were assessed for the generation of liver TRM cells induced by Clec9A targeted immunisation with the Pb 1 epitope. Data from this screen showed that CpG based adjuvants were most effective at inducing the formation of TRM cells in the livers of vaccinated mice and that the transfection reagent DOTAP enhanced this effect. Based on this understanding, we then investigated the potential of CpG and its encapsulation in DOTAP to improve TRM cell generat

Background and Aims: GAS6 signaling, through the TAM receptor tyrosine kinases AXL and MERTK, participates in chronic liver pathologies. Here, we addressed GAS6/TAM involvement in Non-Alcoholic SteatoHepatitis (NASH) development. Methods: GAS6/TAM signaling was analyzed in cultured primary hepatocytes, hepatic stellate cells (HSC) and Kupffer cells (KCs). Axl, Mertk and wild-type C57BL/6 mice were fed with Chow, High Fat Choline-Deficient Methionine-Restricted (HFD) or methionine-choline-deficient (MCD) diet. HSC activation, liver inflammation and cytokine/chemokine production were measured by qPCR, mRNA Array analysis, western blotting and ELISA. GAS6, soluble AXL (sAXL) and MERTK (sMERTK) levels were analyzed in control individuals, steatotic and NASH patients. Results: In primary mouse cultures, GAS6 or MERTK activation protected primary hepatocytes against lipid toxicity via AKT/STAT-3 signaling, while bemcentinib (small molecule AXL inhibitor BGB324) blocked AXL-induced fibrogenesis in primary HSCs and cytokine production in LPS-treated KCs. Accordingly; bemcentinib diminished liver inflammation and fibrosis in MCD- and HFD-fed mice. Upregulation of AXL and ADAM10/ADAM17 metalloproteinases increased sAXL in HFD-fed mice. Transcriptome profiling revealed major reduction in fibrotic- and inflammatory-related genes in HFD-fed mice after bemcentinib administration. HFD-fed Mertk mice exhibited enhanced NASH, while Axl mice were partially protected. In human serum, sAXL levels augmented even at initial stages, whereas GAS6 and sMERTK increased only in cirrhotic NASH patients. In agreement, sAXL increased in HFD-fed mice before fibrosis establishment, while bemcentinib prevented liver fibrosis/inflammation in early NASH. Conclusion: AXL signaling, increased in NASH patients, promotes fibrosis in HSCs and inflammation in KCs, while GAS6 protects cultured hepatocytes against lipotoxicity via MERTK. Bemcentinib, by blocking AXL signaling and increasing GAS6 levels, redu

Dynamic 18F-FDG PET with tracer kinetic modeling has the potential to noninvasively evaluate human liver inflammation using the FDG blood-to-tissue transport rate K 1. Accurate kinetic modeling of dynamic liver PET data and K 1 quantification requires the knowledge of dual-blood input function from the hepatic artery and portal vein. While the arterial input function can be derived from the aortic region on dynamic PET images, it is difficult to extract the portal vein input function accurately from PET images. The optimization-derived dual-input kinetic modeling approach has been proposed to overcome this problem by jointly estimating the portal vein input function and FDG tracer kinetics from time activity curve fitting. In this paper, we further characterize the model properties by analyzing the structural identifiability of the model parameters using the Laplace transform and practical identifiability using computer simulation based on fourteen patient datasets. The theoretical analysis has indicated that all the kinetic parameters of the dual-input kinetic model are structurally identifiable, though subject to local solutions. The computer simulation results have shown that FDG K 1 can be estimated reliably in the whole-liver region of interest with reasonable bias, standard deviation, and high correlation between estimated and original values, indicating of practical identifiability of K 1. The result has also demonstrated the correlation between K 1 and histological liver inflammation scores is reliable. FDG K 1 quantification by the optimization-derived dual-input kinetic model is promising for assessing liver inflammation.

Despite the increasing number of patients suffering from liver inflammation, an inflammatory response commonly seen in obesity and obesity-related metabolic syndrome, the mechanisms underlying this type of inflammation are largely unknown. Currently, there is no effective treatment available. The diagnosis is made by liver biopsy, a procedure associated with serious complications. This dissertation provides new insights into the mechanisms underlying liver inflammation associated with metabolic syndrome. It is shown that the hepatic inflammatory response is caused by oxidised lipid accumulation in macrophage lysosomes, which are cellular compartments situated in immune cells. This dissertation describes a new blood test which allows for easier and quicker diagnosis of patients. It is also shown that several new treatment methods are effective. Finally, it is proven that intestinal flora plays a pivotal role in metabolic syndrome.

The hallmark of nonalcoholic steatohepatitis is hepatocellular inflammation and injury in the setting of hepatic steatosis. Recent work has indicated that dynamic 18F-FDG PET with kinetic modeling has the potential to assess hepatic inflammation noninvasively, while static FDG-PET is less promising. Because the liver has dual blood supplies, kinetic modeling of dynamic liver PET data is challenging in human studies. This paper aims to identify the optimal dual-input kinetic modeling approach for dynamic FDG-PET of human liver inflammation. Fourteen patients with nonalcoholic fatty liver disease were included. Each patient underwent 1 h dynamic FDG-PET/CT scan and had liver biopsy within six weeks. Three models were tested for kinetic analysis: the traditional two-tissue compartmental model with an image-derived single-blood input function (SBIF), a model with population-based dual-blood input function (DBIF), and a new model with optimization-derived DBIF through a joint estimation framework. The three models were compared using Akaike information criterion (AIC), F test and histopathologic inflammation score. Results showed that the optimization-derived DBIF model improved liver time activity curve fitting and achieved lower AIC values and higher F values than the SBIF and population-based DBIF models in all patients. The optimization-derived model significantly increased FDG K1 estimates by 101% and 27% as compared with traditional SBIF and population-based DBIF. K1 by the optimization-derived model was significantly associated with histopathologic grades of liver inflammation while the other two models did not provide a statistical significance. In conclusion, modeling of DBIF is critical for dynamic liver FDG-PET kinetic analysis in human studies. The optimization-derived DBIF model is more appropriate than SBIF and population-based DBIF for dynamic FDG-PET of liver inflammation.

Despite the increasing number of patients suffering from liver inflammation, an inflammatory response commonly seen in obesity and obesity-related metabolic syndrome, the mechanisms underlying this type of inflammation are largely unknown. Currently, there is no effective treatment available. The diagnosis is made by liver biopsy, a procedure associated with serious complications. This dissertation provides new insights into the mechanisms underlying liver inflammation associated with metabolic syndrome. It is shown that the hepatic inflammatory response is caused by oxidised lipid accumulation in macrophage lysosomes, which are cellular compartments situated in immune cells. This dissertation describes a new blood test which allows for easier and quicker diagnosis of patients. It is also shown that several new treatment methods are effective. Finally, it is proven that intestinal flora plays a pivotal role in metabolic syndrome.

BACKGROUND & AIMS: Ischemia-reperfusion injury (IRI) can cause hepatic failure after liver surgery or transplantation. IRI causes oxidative stress, which injures sinusoidal endothelial cells (SECs), leading to recruitment and activation of Kupffer cells and platelets (by unknown processes) and microcirculatory impairment. We investigated whether injured SECs release microparticles (small fragments shed from outer leaf of the plasma membrane) with inflammatory effects that contribute to IRI pathogenesis. METHODS: C57BL6 mice underwent 60min of partial hepatic ischemia followed by 15min-24hrs of reperfusion. We collected blood and liver samples, isolated and measured circulating microparticles, and determined protein and lipid content. In mouse primary hepatocytes, we investigated features of microparticles and mechanisms of production. We analyzed the effects of an annexin V-homodimer (diannexin or ASP8597) on post-ischemia microparticle production and function. RESULTS: Circulating microparticles released 15-30min after ischemic reperfusion contained markers of SECs, platelets, natural killer T cells, and CD8+ cells; 4 hrs later, they contained markers of macrophages. Microparticles also contained F2- isoprostanes, markers of oxidative damage to membrane lipids. Injection of mice with TNF-alfa increased formation of microparticles, whereas injection with diannexin substantially reduced release of microparticles and prevented IRI. Hypoxia followed by re-oxygenation of primary hepatocytes generated microparticles via processes that involved oxidative stress and Ca2+. Circulating microparticles were engulfed by cultured hepatocytes, injuring them via the mitochondrial membrane permeability transition. Microparticles also activated platelets and induced migration of inflammatory cells in vitro. The inflammatory properties of microparticles involved activation of the transcription factor NF-kB, JNK and increased levels of E-selectin, P-selectin, Icam1, and Vcam1; these p