Hydrogen sulfide attenuates carbon tetrachloride-induced hepatotoxicity, liver cirrhosis and portal hypertension in rats

BACKGROUND : Hydrogen sulfide (H(2)S) displays vasodilative, anti-oxidative, anti-inflammatory and cytoprotective activities. Impaired production of H(2)S contributes to the increased intrahepatic resistance in cirrhotic livers. The study aimed to investigate the roles of H(2)S in carbon tetrachloride (CCl(4))-induced hepatotoxicity, cirrhosis and portal hypertension.

METHODS AND FINDINGS : Sodium hydrosulfide (NaHS), a donor of H(2)S, and DL-propargylglycine (PAG), an irreversible inhibitor of cystathionine γ-lyase (CSE), were applied to the rats to investigate the effects of H(2)S on CCl(4)-induced acute hepatotoxicity, cirrhosis and portal hypertension by measuring serum levels of H(2)S, hepatic H(2)S producing activity and CSE expression, liver function, activity of cytochrome P450 (CYP) 2E1, oxidative and inflammatory parameters, liver fibrosis and portal pressure. CCl(4) significantly reduced serum levels of H(2)S, hepatic H(2)S production and CSE expression. NaHS attenuated CCl(4)-induced acute hepatotoxicity by supplementing exogenous H(2)S, which displayed anti-oxidative activities and inhibited the CYP2E1 activity. NaHS protected liver function, attenuated liver fibrosis, inhibited inflammation, and reduced the portal pressure, evidenced by the alterations of serum alanine aminotransferase (ALT), aspartate aminotransferase (AST), hyaluronic acid (HA), albumin, tumor necrosis factor (TNF)-α, interleukin (IL)-1β, IL-6 and soluble intercellular adhesion molecule (ICAM)-1, liver histology, hepatic hydroxyproline content and α-smooth muscle actin (SMA) expression. PAG showed opposing effects to NaHS on most of the above parameters.

CONCLUSIONS :  Exogenous H2S attenuates CCl4-induced hepatotoxicity, liver cirrhosis and portal hypertension by its multiple functions including anti-oxidation, anti-inflammation, cytoprotection and anti-fibrosis, indicating that targeting H2S may present a promising approach, particularly for its prophylactic effects, against liver cirrhosis and portal hypertension.

Caveolin-1 orchestrates the balance between glucose and lipid-dependent energy metabolism : implications for liver regeneration

Caveolin-1 (CAV1) is a structural protein of caveolae involved in lipid homeostasis and endocytosis. Using newly generated pure Balb/C CAV1 null (Balb/CCAV1−/−) mice, CAV1−/− mice from Jackson Laboratories (JAXCAV1−/−), and CAV1−/− mice developed in the Kurzchalia Laboratory (KCAV1−/−), we show that under physiological conditions CAV1 expression in mouse tissues is necessary to guarantee an efficient progression of liver regeneration and mouse survival after partial hepatectomy. Absence of CAV1 in mouse tissues is compensated by the development of a carbohydrate-dependent anabolic adaptation. These results were supported by extracellular flux analysis of cellular glycolytic metabolism in CAV1-knockdown AML12 hepatocytes, suggesting cell autonomous effects of CAV1 loss in hepatic glycolysis. Unlike in KCAV1−/− livers, in JAXCAV1−/− livers CAV1 deficiency is compensated by activation of anabolic metabolism (pentose phosphate pathway and lipogenesis) allowing liver regeneration. Administration of 2-deoxy-glucose in JAXCAV1−/− mice indicated that liver regeneration in JAXCAV1−/− mice is strictly dependent on hepatic carbohydrate metabolism. Moreover, with the exception of regenerating JAXCAV1−/− livers, expression of CAV1 in mice is required for efficient hepatic lipid storage during fasting, liver regeneration, and diet-induced steatosis in the three CAV1−/− mouse strains. Furthermore, under these conditions CAV1 accumulates in the lipid droplet fraction in wildtype mouse hepatocytes. Conclusion: Our data demonstrate that lack of CAV1 alters hepatocyte energy metabolism homeostasis under physiological and pathological conditions.

Fructose containing sugars modulate mRNA of lipogenic genes ACC and FAS and protein levels of transcription factors ChREBP and SREBPIc with no effect on body weight or liver fat

The aim of this study was to determine the effects of high-glucose, high-fructose and high-sucrose diets on weight gain, liver lipid metabolism and gene expression of proteins involved with hepatic fat metabolism. Rats were fed a diet containing either 60% glucose, 60% fructose, 60% sucrose, or a standard chow for 28 days. Results indicated that high-fructose and high-sucrose diets were associated with higher mRNA levels of gene transcripts involved with fat synthesis; ACC, FAS and ChREBP, with no change in SREBP-1C mRNA. The protein level of ChREBP and SREBP1c was similar in liver homogenates from all groups, but were higher in nuclear fractions from the liver of high-fructose and high-sucrose fed rats. The mRNA level of gene transcripts involved with fat oxidation was the same in all three diets, whilst a high-fructose diet was associated with greater amount of mRNA of the fat transporter CD36. Despite the changes in mRNA of lipogenic proteins, the body weight of animals from each group was the same and the livers from rats fed high-fructose and high-sucrose diets did not contain more fat than control diet livers. In conclusion, changing the composition of the principal monosaccharide in the diet to a fructose containing sugar elicits changes in the level of hepatic mRNA of lipogenic and fat transport proteins and protein levels of their transcriptional regulators; however this is not associated with any changes in body weight or liver fat content.

Efficacy of using cancer stem cell markers in isolating and characterizing liver cancer stem cells

Recent evidence suggests that a subset of hepatocellular carcinomas (HCCs) are derived from liver cancer stem cells (LCSCs). In order to isolate and characterize LCSCs, reliable markers that are specific to these cells are required. We evaluated the efficacy of a range of cancer stem cell (CSC) markers in isolating and characterizing LCSCs. We show that the most widely used CSC markers are not specific to LCSCs. By western analysis, protein expression of the common markers showed no significant difference between HCC tumor tissues and adjacent non-cancerous liver. Further, isolation of LCSCs from common HCC cell lines using FACScan and microbeads showed no consistent marker expression pattern. We also show that LCSCs have unique subtypes. Immunohistochemistry of HCC tissues showed that different HCCs express unique combinations of LCSC markers. Quantitative real-time polymerase chain reaction analysis showed that LCSCs isolated using different markers in the same HCC phenotype had different expression profiles. Likewise, LCSCs isolated from different HCC phenotypes with the same marker also had unique expression profiles and displayed varying resistance profiles to Sorafenib. Thus, using a range of commonly used CSC markers in HCCs and cell lines, we demonstrate that currently available markers are not specific for LCSCs. LCSCs have unique subtypes that express distinctive combinations of LCSC markers and altered drug resistance profiles, making their identification problematic.

A global approach to dissecting lipid-induced impairment of liver gluconeogenesis

This thesis identified: 1) novel aspects of cholesterol metabolism, 2) a novel signalling lipid in the phosphatidylinositol class, and 3) a gene involved in regulating lipid breakdown, as being regulated in the reversal of a fatty acid induced diabetic liver. The identification of these targets may allow for future development of targeted therapies against type 2 diabetes with a specific focus on the liver.

Overexpression of sphingosine kinase 1 in liver reduces triglyceride content in mice fed a low but not high-fat diet

 Hepatic insulin resistance is a major risk factor for the development of type 2 diabetes and is associated with the accumulation of lipids, including diacylglycerol (DAG), triacylglycerols (TAG) and ceramide. There is evidence that enzymes involved in ceramide or sphingolipid metabolism may have a role in regulating concentrations of glycerolipids such as DAG and TAG. Here we have investigated the role of sphingosine kinase (SphK) in regulating hepatic lipid levels. We show that mice on a high-fat high-sucrose diet (HFHS) displayed glucose intolerance, elevated liver TAG and DAG, and a reduction in total hepatic SphK activity. Reduced SphK activity correlated with downregulation of SphK1, but not SphK2 expression, and was not associated with altered ceramide levels. The role of SphK1 was further investigated by overexpressing this isoform in the liver of mice in vivo. On a low-fat diet (LFD) mice overexpressing liver SphK1, displayed reduced hepatic TAG synthesis and total TAG levels, but with no change to DAG or ceramide. These mice also exhibited no change in gluconeogenesis, glycogenolysis or glucose tolerance. Similarly, overexpression of SphK1 had no effect on the pattern of endogenous glucose production determined during a glucose tolerance test. Under HFHS conditions, normalization of liver SphK activity to levels observed in LFD controls did not alter hepatic TAG concentrations. Furthermore, DAG, ceramide and glucose tolerance were also unaffected. In conclusion, our data suggest that SphK1 plays an important role in regulating TAG metabolism under LFD conditions.

CD8+ T cells from a novel T cell receptor transgenic mouse induce liver-stage immunity that can be boosted by blood-stage infection in rodent malaria

To follow the fate of CD8+ T cells responsive to Plasmodium berghei ANKA (PbA) infection, we generated an MHC I-restricted TCR transgenic mouse line against this pathogen. T cells from this line, termed PbT-I T cells, were able to respond to blood-stage infection by PbA and two other rodent malaria species, P. yoelii XNL and P. chabaudi AS. These PbT-I T cells were also able to respond to sporozoites and to protect mice from liver-stage infection. Examination of the requirements for priming after intravenous administration of irradiated sporozoites, an effective vaccination approach, showed that the spleen rather than the liver was the main site of priming and that responses depended on CD8α+ dendritic cells. Importantly, sequential exposure to irradiated sporozoites followed two days later by blood-stage infection led to augmented PbT-I T cell expansion. These findings indicate that PbT-I T cells are a highly versatile tool for studying multiple stages and species of rodent malaria and suggest that cross-stage reactive CD8+ T cells may be utilized in liver-stage vaccine design to enable boosting by blood-stage infections.