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OBJECTIVES: Nonalcoholic steatohepatitis (NASH) is the most severe form of a hepatic condition known as nonalcoholic fatty liver disease (NAFLD). NASH is histologically characterized by hepatic fat accumulation, inflammation, and ballooning, and eventually coupled with fibrosis that, in turn, may progress to end-stage liver disease even in young individuals. Hence, there is a critical need for specific noninvasive markers to predict hepatic inflammation at an early age. We investigated whether plasma levels of cathepsin D (CatD), a lysosomal protease, correlated with the severity of liver inflammation in pediatric NAFLD. METHODS: Liver biopsies from children (n=96) with NAFLD were histologically evaluated according to the criteria of Kleiner (NAFLD activity score) and the Brunt's criteria. At the time of liver biopsy, blood was taken and levels of CatD, alanine aminotransferase (ALT), and cytokeratin-18 (CK-18) were measured in plasma. RESULTS: Plasma CatD levels were significantly lower in subjects with liver inflammation compared with steatotic subjects. Furthermore, we found that CatD levels were gradually reduced and corresponded with increasing severity of liver inflammation, steatosis, hepatocellular ballooning, and NAFLD activity score. CatD levels correlated with pediatric NAFLD disease progression better than ALT and CK-18. In particular, CatD showed a high diagnostic accuracy (area under receiver operating characteristic curve (ROC-AUC): 0.94) for the differentiation between steatosis and hepatic inflammation, and reached almost the maximum accuracy (ROC-AUC: 0.998) upon the addition of CK-18. CONCLUSIONS: Plasma CatD holds a high diagnostic value to distinguish pediatric patients with hepatic inflammation from children with steatosis.

Item does not contain fulltext, Recent investigations have suggested that inflammasome activation plays an important role during atherosclerosis. Upon activation, the inflammasome induces processing and release of pro-inflammatory cytokines interleukin 1beta (IL-1beta) and interleukin 18 (IL-18) via activation of caspase-1/11. Previously, it was shown that complete caspase-1 deficiency is protective against atherosclerosis development. However, while macrophages are the main inflammatory cells involved in atherosclerosis, the exact role of macrophage-specific caspase-1/11 activation during development of cardiovascular disease has never been investigated. We hypothesized that hematopoietic caspase-1/11 deficiency leads to reduced atherosclerosis development. To investigate the specific contribution of hematopoietic caspase-1/11 activation to atherosclerosis development, Ldlr(-/-) mice received a transplant (tp) of wild-type (WT) or caspase-1/11(-/-) bone marrow, to create WT-tp mice and caspase-1/11(-/-) -tp mice, and fed a high-fat, high-cholesterol diet for 12 weeks. Our results showed an increase in anti-inflammatory blood leukocytes in caspase-1/11(-/-) -tp mice compared with WT-tp mice, as indicated by a decreased level of Ly6C(high) monocytes and an increased level of Ly6C(low) monocytes. In line with our hypothesis, hematopoietic deletion of caspase-1/11 resulted in a strong reduction in atherosclerotic plaque size. Furthermore, necrotic core content was dramatically decreased in caspase-1/11(-/-) -tp mice. Our data indicate that hematopoietic caspase-1/11 activation is involved in vascular inflammation and atherosclerosis, and plays an important role in cardiovascular disease progression.

Contains fulltext : 125609.pdf (publisher's version ) (Open Access), BACKGROUND & AIMS: While non-alcoholic steatohepatitis (NASH) is characterized by hepatic steatosis combined with inflammation, the mechanisms triggering hepatic inflammation are unknown. In Ldlr(-/-) mice, we have previously shown that lysosomal cholesterol accumulation in Kupffer cells (KCs) correlates with hepatic inflammation and cholesterol crystallization. Previously, cholesterol crystals have been shown to induce the activation of inflammasomes. Inflammasomes are protein complexes that induce the processing and release of pro-inflammatory cytokines IL-1b and IL-18 via caspase-1 activation. Whereas caspase-1 activation is independent of caspase-11 in the canonical pathway of inflammasome activation, caspase-11 was found to trigger caspase-1-dependent IL-1b and IL-18 in response to non-canonical inflammasome activators. So far, it has not been investigated whether inflammasome activation stimulates the formation of cholesterol crystals. We hypothesized that inflammasome activation in KCs stimulates cholesterol crystallization, thereby leading to hepatic inflammation. METHODS: Ldlr (-/-) mice were transplanted (tp) with wild-type (Wt) or caspase-1/11(-/-) (dKO) bone marrow and fed either regular chow or a high-fat, high-cholesterol (HFC) diet for 12 weeks. In vitro, bone marrow derived macrophages (BMDM) from wt or caspase-1/11(-/-) mice were incubated with oxLDL for 24h and autophagy was assessed. RESULTS: In line with our hypothesis, caspase-1/11(-/-)-tp mice had less severe hepatic inflammation than Wt-tp animals, as evident from liver histology and gene expression analysis in isolated KCs. Mechanistically, KCs from caspase-1/11(-/-)-tp mice showed less cholesterol crystals, enhanced cholesterol efflux and increased autophagy. In wt BMDM, oxLDL incubation led to disturbed autophagy activity whereas BMDM from caspase-1/11(-/-) mice had normal autophagy activity. CONCLUSION: Altogether, these data suggest a vicious cycle whereby disturbed autophagy and decrease

BACKGROUND & AIMS:: Non-alcoholic steatohepatitis (NASH) is characterized by hepatic steatosis with inflammation. Although steatosis is benign and reversible, inflammation can increase liver damage. Hepatic inflammation has been associated with accumulation of cholesterol in lysosomes of Kupffer cells. 27-hydroxycholesterol (27HC), a derivative of cholesterol formed by CYP27A1, can mobilize cholesterol from the lysosomes to the cytoplasm. We investigated whether 27HC can change the intracellular distribution cholesterol and reduce hepatic inflammation in mice. METHODS:: We transplanted bone marrow from irradiated wild-type or Cyp27a1-/-mice to mice that do not express the low density lipoprotein receptor ( Ldlr-/-), which are hyperlipidemic; 9 weeks later, mice were fed either regular chow or a high-fat, high-cholesterol (HFC) diet for 3 months. In a separate experiment, Ldlr-/- mice were given subcutaneous injections of 27HC and placed on regular chow or HFC diets for 3 weeks. Blood and liver tissues samples were collected and analyzed for intracellular cholesterol distribution and inflammation. RESULTS:: In Ldlr-/-mice that received bone marrow transplants from Cyp27a1-/-mice, lysosomes of Kupfer cells had a greater accumulation of cholesterol than those of mice that received bone marrow from wild-type mice, after the HFC diet. Liver histology and gene expression analyses showed increased inflammation and liver damage in mice given bone marrow transplants from Cyp27a1-/-mice and placed on the HFC diet. Administration of 27HC to Ldlr-/-mice, following the HFC diet, reduced the accumulation of lysosomal cholesterol and hepatic inflammation, compared with mice that were not given 27HC. CONCLUSIONS:: Accumulation of cholesterol in lysosomes of Kupfer cells promotes hepatic inflammation in mice. The cholesterol derivative 27HC reduces accumulation of cholesterol in lysosomes and might be used to treat NASH.

BACKGROUND & AIMS: Non-alcoholic steatohepatitis (NASH) is characterized steatosis and inflammation. The transition from steatosis towards NASH a key step in pathogenesis, as it will set the stage for further severe damage. Under normal conditions, lipoproteins that are endocytosed by cells (KCs) are easily transferred from the lysosomes into the Oxidized LDL (oxLDL) that is taken up by the macrophages in vitro is within the lysosomes, while acetylated LDL (acLDL) is leading to normal hydrolysis, resulting in cytoplasmic storage. We have recently hepatic inflammation is correlated with lysosomal trapping of lipids. So link between lysosomal trapping of oxLDL and inflammation was not hypothesized that lysosomal trapping of oxLDL in KCs will lead to inflammation. METHODS: Ldlr-/- mice were injected with LDL, acLDL and sacrificed after 2, 6 and 24 h. RESULTS: Electron microscopy of KCs that after oxLDL injection, small lipid inclusions were present inside lysosomes after all time points and were mostly pronounced after 6 and contrast, no lipid inclusions were present inside KCs after LDL or acLDL injection. Hepatic expression of several inflammatory genes and receptors was higher after oxLDL injections compared with LDL or acLDL. CONCLUSIONS: These data suggest that trapping of oxLDL inside lysosomes vivo is causally linked to increased hepatic inflammatory gene novel observations provide new bases for prevention and treatment of