[Show abstract][Hide abstract] ABSTRACT: Liver monocytes play a major role in the development of NASH (non-alcoholic steatohepatitis). In inflamed tissues, monocytes can differentiate in both macrophages and dendritic cells. In the present study, we investigated the role of moDCs (monocyte-derived inflammatory dendritic cells) in experimental steatohepatitis induced in C57BL/6 mice by feeding on a MCD (methionine/choline-deficient) diet. The evolution of steatohepatitis was characterized by an increase in hepatic CD45(+)/CD11b(+) myeloid cells displaying the monocyte/macrophage marker F4-80(+). In the early phases (4 weeks of treatment), Ly6C(high)/CD11b(+)/F4-80(+) inflammatory macrophages predominated. However, their frequency did not grow further with the disease progression (8 weeks of treatment), when a 4-fold expansion of CD11b(+)/F4-80(+) cells featuring the fractalkine receptor (CX3CR1) was evident. These CX3CR1(+) cells were also characterized by the combined expression of inflammatory monocyte (Ly6C, CD11b) and dendritic cell (CD11c, MHCII) markers as well as by a sustained TNFα (tumour necrosis factor α) production, suggesting monocyte differentiation into inflammatory moDCs. The expansion of TNFα-producing CX3CR1(+) moDCs was associated with an elevation in hepatic and circulating TNFα level and with the worsening of parenchymal injury. Hydrogen sulfide (H2S) has been shown to interfere with CX3CR1 up-regulation in monocyte-derived cells exposed to pro-inflammatory stimuli. Treating 4-week-MCD-fed mice with the H2S donor NaHS while continuing on the same diet prevented the accumulation of TNFα-producing CX3CR1(+) moDCs without interfering with hepatic macrophage functions. Furthermore, NaHS reduced hepatic and circulating TNFα levels and ameliorated transaminase release and parenchymal injury. Altogether, these results show that inflammatory CX3CR1(+) moDCs contributed in sustaining inflammation and liver injury during steatohepatitis progression.
[Show abstract][Hide abstract] ABSTRACT: The growing diffusion of nonalcoholic fatty liver disease (NAFLD) is a consequence of the worldwide increase in the prevalence of obesity. Oxidative stress is widely recognized to play a pivotal role in NAFLD evolution to nonalcoholic steatohepatitis (NASH). Here we review recent evidence suggesting that oxidative stress-derived antigens originating within fatty livers stimulate both humoral and cellular adaptive immune responses and the possible mechanisms involved in sustaining hepatic inflammation in NASH.
World Journal of Hepatology 07/2015; 7(13):1725-9. DOI:10.4254/wjh.v7.i13.1725
[Show abstract][Hide abstract] ABSTRACT: Unlabelled:
Annexin A1 (AnxA1) is an effector of the resolution of inflammation and is highly effective in terminating acute inflammatory responses. However, its role in chronic settings is less investigated. Because changes in AnxA1 expression within adipose tissue characterize obesity in mice and humans, we queried a possible role for AnxA1 in the pathogenesis of nonalcoholic steatohepatitis (NASH), a disease commonly associated with obesity. NASH was induced in wild-type (WT) and AnxA1 knockout (AnxA1 KO) C57BL/6 mice by feeding a methionine-choline deficient (MCD) diet up to 8 weeks. In MCD-fed WT mice, hepatic AnxA1 increased in parallel with progression of liver injury. This mediator was also detected in liver biopsies from patients with NASH and its degree of expression inversely correlated with the extent of fibrosis. In both humans and rodents, AnxA1 production was selectively localized in liver macrophages. NASH in AnxA1 KO mice was characterized by enhanced lobular inflammation resulting from increased macrophage recruitment and exacerbation of the M1 phenotype. Consistently, in vitro addition of recombinant AnxA1 to macrophages isolated from NASH livers down-modulated M1 polarization through stimulation of interleukin-10 production. Furthermore, the degree of hepatic fibrosis was enhanced in MCD-fed AnxA1 KO mice, an effect associated with augmented liver production of the profibrotic lectin, galectin-3. Accordingly, AnxA1 addition to isolated hepatic macrophages reduced galectin-3 expression.
Macrophage-derived AnxA1 plays a functional role in modulating hepatic inflammation and fibrogenesis during NASH progression, suggesting the possible use of AnxA1 analogs for therapeutic control of this disease.
[Show abstract][Hide abstract] ABSTRACT: In recent years, an increasing number of reports have shown the involvement of osteopontin (OPN), a pleiotropic cytokine and an important component of the extracellular matrix (ECM), in the pathogenesis of liver injury and the development of fibrosis.1 ,2 OPN is also frequently overexpressed in hepatocellular carcinoma (HCC), where it modulates HCC growth, invasion and metastasis,2 and in cholangiocarcinoma, where its expression bears prognostic significance. Previous studies3 have shown that in injured livers OPN is expressed by hepatic stellate cells (HSC) and upregulates collagen I production. Interestingly, in HSC, OPN expression appears to be downstream of SOX9, a Hedgehog- and Notch-controlled transcription factor that is expressed also in biliary cells and hepatic progenitor cells (HPC)/hepatocytes committed to the biliary fate.4-6 In this issue of Gut, two papers investigate the role of OPN in HPC-driven ductular reaction (DR) in relation to the progression of liver fibrosis.7 ,8 DR, a histological lesion present in most chronic liver diseases, is a dynamic, multicellular complex characterised by the presence of ‘reactive’ ductular epithelial cells, arranged in irregular strings along the margins of the portal tract in close contact with mesenchymal, inflammatory and endothelial cells. Reactive ductular cells acquire novel functions including the secretion of cytokines, chemokines, growth factors and angiogenic factors, enabling them to establish intense paracrine communications with … [Full text of this article]
Gut 07/2014; 63(11). DOI:10.1136/gutjnl-2014-307712 · 14.66 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Unlabelled:
Previous studies have shown that human nonalcoholic steatohepatitis (NASH) is often associated with the presence of circulating antibodies against protein adducted by lipid peroxidation products. Here we used the methionine-choline deficient (MCD) model of NASH to characterize the possible involvement of adaptive immunity in NASH. In mice fed up to 8 weeks with the MCD diet the extension of liver injury and lobular inflammation paralleled the development of immunoglobulin G (IgG) against malonyldialdehyde (MDA) and 4-hydroxynonenal (4-HNE)-derived antigens as well as with the hepatic recruitment of CD4(+) and CD8(+) T-lymphocytes responsive to the same antigens. Moreover, in these animals the individual IgG reactivity against MDA-adducts positively correlated with transaminase release and hepatic tumor necrosis factor alpha (TNF-α) expression. To substantiate the role of immune responses triggered by oxidative stress in the progression of NASH, mice were immunized with MDA-adducted bovine serum albumin (MDA-BSA) before feeding the MCD diet. MDA-BSA immunization did not affect control mice livers, but further stimulated transaminase release, lobular inflammation, and the hepatic expression of proinflammatory cytokine in MCD-fed mice. The increased severity of NASH in immunized MCD-fed mice involved liver recruitment and the T helper (Th)-1 activation of CD4(+) T cells that, in turn, further stimulated macrophage M1 responses. Moreover, hepatic fibrosis was also evident in these animals in relation with an IL-15-mediated increase of natural killer T-cells (NKT) and the up-regulation in liver production of osteopontin by NKT cells and hepatic macrophages.
These results indicate that oxidative stress can contribute to the progression of NASH by stimulating both humoral and cellular immune responses, pointing to the possible role of adaptive immunity in the pathogenesis of the disease.
[Show abstract][Hide abstract] ABSTRACT: Growing evidence indicates that NF-kB activation contributes to the pathogenesis of non-alcoholic steatohepatisis (NASH). Among the NF-kB sub-units, p50/NFkB1 has regulatory activities down-modulating NF-kB-mediated responses. In this study we investigated the effects of NFKB1 deficiency on the progression of NASH induced by feeding mice with a methionine-choline deficient (MCD) diet. Following 4 weeks on the MCD diet, steatosis, ALT release, hepatocyte apoptosis, lobular inflammation and TNF-α production were higher in NFKB1-ko than in wild-type (WT) mice. NFkB1-/- mice also showed appreciable centrilobular collagen deposition, increased number of activated hepatic stellate cells and higher type-I procollagen-α and TIMP-1 mRNA expression. Despite NF-kBp50 homodimers regulate macrophage activation, the number of hepatic macrophages and liver mRNAs for inducible NO synthase, IL-12p40, CCL2, CXCL10 were comparable in the two strains. NASH was associated with an increase in liver infiltrating T-lymphocytes that was more evident in MCD-fed NFKB1-/- than in similarly treated WT mice. Flow-cytorimetry showed that T cell recruitment involved effector CD8+-T cells without changes in the helper CD4+ T-cell fraction. Furthermore, while NASH lowered hepatic natural killer T cells (NKT) in WT mice, the NKT pool was selectively increased in the livers of MCD-fed NFkB1-/- mice. Such a NKT recruitment was associated with an early over-expression of IL-15, a cytokine controlling NKT cell survival and maturation. In the livers of MCD-fed NFkB1-/- mice, but not in those of WT littermates, we also observed an up-regulation in the production of NKT-related cytokines IFN-γ and osteopontin. Altogether these results indicated that NF-KB1 down-modulation enhanced NASH progression to fibrosis by favoring NKT cell recruitment, stressing the contribution of NKT cells in the pathogenesis of NASH.
[Show abstract][Hide abstract] ABSTRACT: Expression of CCL2 (CC chemokine ligand 2) (or monocyte chemoattractant protein-1) regulates inflammatory cell infiltration in the liver and adipose tissue, favouring steatosis. However, its role in the pathogenesis of steatohepatitis is still uncertain. In the present study, we investigated the development of non-alcoholic steatohepatitis induced by an MCD diet (methionine/choline-deficient diet) in mice lacking the CCL2 gene on two different genetic backgrounds, namely Balb/C and C57/Bl6J. WT (wild-type) and CCL2-KO (knockout) mice were fed on a lipid-enriched MCD diet or a control diet for 8 weeks. In Balb/C mice fed on the MCD diet, a lack of CCL2 was associated with lower ALT (alanine transaminase) levels and reduced infiltration of inflammatory cells, together with a lower generation of oxidative-stress-related products. Sirius Red staining demonstrated pericellular fibrosis in zone 3, and image analysis showed a significantly lower matrix accumulation in CCL2-KO mice. This was associated with reduced hepatic expression of TGF-β (transforming growth factor-β), type I procollagen, TIMP-1 (tissue inhibitor of metalloproteinases-1) and α-smooth muscle actin. In contrast, in mice on a C57Bl/6 background, neither ALT levels nor inflammation or fibrosis were significantly different comparing WT and CCL2-KO animals fed on an MCD diet. In agreement, genes related to fibrogenesis were expressed to comparable levels in the two groups of animals. Comparison of the expression of several genes involved in inflammation and repair demonstrated that IL (interleukin)-4 and the M2 marker MGL-1 (macrophage galactose-type C-type lectin 1) were differentially expressed in Balb/C and C57Bl/6 mice. No significant differences in the degree of steatosis were observed in all groups of mice fed on the MCD diet. We conclude that, in experimental murine steatohepatitis, the effects of CCL2 deficiency are markedly dependent on the genetic background.
[Show abstract][Hide abstract] ABSTRACT: NEFA (non-esterified 'free' fatty acid)-mediated lipotoxicity plays a critical role in the pathogenesis of NASH (non-alcoholic steatohepatitis). In the light of the growing need for new therapeutic options for NASH, we investigated the action of A2aR (adenosine A(2a) receptor) stimulation against lipotoxicity. The effects of the A(2a)R agonist CGS21680 [2-p-(2-carboxyethyl)phenethylamino-5'-N-ethylcarboxyamidoadenosine] were evaluated 'in vitro' in liver cells exposed to SA (stearic acid) and 'in vivo' in rats with NASH induced by 8 weeks of feeding with an MCD diet (methionine/choline-deficient diet). In cultured hepatocytes, SA promoted apoptosis by inducing MKK4 (mitogen-activated protein kinase kinase 4)/SEK1 (stress-activated protein kinase/extracellular-signal-regulated kinase kinase-1) and JNK-1/2 (c-Jun N-terminal kinase-1/2) activation. CGS21680 addition prevented JNK-1/2 activation and reduced apoptosis without interfering with lipid accumulation. CGS21680 action required PI3K (phosphoinositide 3-kinase)/Akt-mediated block of MKK4/SEK1. Consistently, PI3K inhibition with wortmannin abolished the cytoprotective action of CGS21680 and reverted MKK4 inhibition. SA lipotoxicity was also prevented by transfecting HTC cells with a specific MKK4/SEK1 siRNA (small interfering RNA). In rats receiving the MCD diet, the development of NASH was associated with MKK4/SEK1 and JNK-1/2 activation. CGS21680 (0.5 mg/kg of body weight, intraperitoneal) administration to MCD-fed rats prevented JNK-1/2 activation by acting on MKK4/SEK1. CGS21680 also effectively reduced NASH-associated ALT (alanine aminotransferase) release, hepatocyte apoptosis, liver inflammation and fibrosis without affecting hepatic steatosis. Taken together, these results demonstrate that, by inhibiting JNK-1/2, A(2a)R stimulation reduces lipotoxicity and ameliorates NASH, giving a rationale to investigate A(2a)R agonists as possible new therapeutic agents in preventing fatty liver progression to NASH.