Role of fatty-acid synthesis in dendritic cell generation and function.

Dendritic cells (DC) are professional APCs that regulate innate and adaptive immunity. The role of fatty-acid synthesis in DC development and function is uncertain. We found that blockade of fatty-acid synthesis markedly decreases dendropoiesis in the liver and in primary and secondary lymphoid organs in mice. Human DC development from PBMC precursors was also diminished by blockade of fatty-acid synthesis. This was associated with higher rates of apoptosis in precursor cells and increased expression of cleaved caspase-3 and BCL-xL and downregulation of cyclin B1. Further, blockade of fatty-acid synthesis decreased DC expression of MHC class II, ICAM-1, B7-1, and B7-2 but increased their production of selected proinflammatory cytokines including IL-12 and MCP-1. Accordingly, inhibition of fatty-acid synthesis enhanced DC capacity to activate allogeneic as well as Ag-restricted CD4(+) and CD8(+) T cells and induce CTL responses. Further, blockade of fatty-acid synthesis increased DC expression of Notch ligands and enhanced their ability to activate NK cell immune phenotype and IFN-γ production. Because endoplasmic reticulum (ER) stress can augment the immunogenic function of APC, we postulated that this may account for the higher DC immunogenicity. We found that inhibition of fatty-acid synthesis resulted in elevated expression of numerous markers of ER stress in humans and mice and was associated with increased MAPK and Akt signaling. Further, lowering ER stress by 4-phenylbutyrate mitigated the enhanced immune stimulation associated with fatty-acid synthesis blockade. Our findings elucidate the role of fatty-acid synthesis in DC development and function and have implications to the design of DC vaccines for immunotherapy.

Dendritic cells limit fibroinflammatory injury in nonalcoholic steatohepatitis in mice.

UNLABELLED: Nonalcoholic steatohepatitis (NASH) is the most common etiology of chronic liver dysfunction in the United States and can progress to cirrhosis and liver failure. Inflammatory insult resulting from fatty infiltration of the liver is central to disease pathogenesis. Dendritic cells (DCs) are antigen-presenting cells with an emerging role in hepatic inflammation. We postulated that DCs are important in the progression of NASH. We found that intrahepatic DCs expand and mature in NASH liver and assume an activated immune phenotype. However, rather than mitigating the severity of NASH, DC depletion markedly exacerbated intrahepatic fibroinflammation. Our mechanistic studies support a regulatory role for DCs in NASH by limiting sterile inflammation through their role in the clearance of apoptotic cells and necrotic debris. We found that DCs limit CD8(+) T-cell expansion and restrict Toll-like receptor expression and cytokine production in innate immune effector cells in NASH, including Kupffer cells, neutrophils, and inflammatory monocytes. Consistent with their regulatory role in NASH, during the recovery phase of disease, ablation of DC populations results in delayed resolution of intrahepatic inflammation and fibroplasia. CONCLUSION: Our findings support a role for DCs in modulating NASH. Targeting DC functional properties may hold promise for therapeutic intervention in NASH.

Toll-like receptor 7 regulates pancreatic carcinogenesis in mice and humans.

Pancreatic ductal adenocarcinoma is an aggressive cancer that interacts with stromal cells to produce a highly inflammatory tumor microenvironment that promotes tumor growth and invasiveness. The precise interplay between tumor and stroma remains poorly understood. TLRs mediate interactions between environmental stimuli and innate immunity and trigger proinflammatory signaling cascades. Our finding that TLR7 expression is upregulated in both epithelial and stromal compartments in human and murine pancreatic cancer led us to postulate that carcinogenesis is dependent on TLR7 signaling. In a mouse model of pancreatic cancer, TLR7 ligation vigorously accelerated tumor progression and induced loss of expression of PTEN, p16, and cyclin D1 and upregulation of p21, p27, p53, c-Myc, SHPTP1, TGF-ß, PPARγ, and cyclin B1. Furthermore, TLR7 ligation induced STAT3 activation and interfaced with Notch as well as canonical NF-κB and MAP kinase pathways, but downregulated expression of Notch target genes. Moreover, blockade of TLR7 protected against carcinogenesis. Since pancreatic tumorigenesis requires stromal expansion, we proposed that TLR7 ligation modulates pancreatic cancer by driving stromal inflammation. Accordingly, we found that mice lacking TLR7 exclusively within their inflammatory cells were protected from neoplasia. These data suggest that targeting TLR7 holds promise for treatment of human pancreatic cancer.

Dendritic cell populations with different concentrations of lipid regulate tolerance and immunity in mouse and human liver.

BACKGROUND & AIMS: Immune cells of the liver must be able to recognize and react to pathogens yet remain tolerant to food molecules and other nonpathogens. Dendritic cells (DCs) are believed to contribute to hepatic tolerance. Lipids have been implicated in dysfunction of DCs in cancer. Therefore, we investigated whether high lipid content in liver DCs affects induction of tolerance. METHODS: Mouse and human hepatic nonparenchymal cells were isolated by mechanical and enzymatic digestion. DCs were purified by fluorescence-activated cell sorting or with immunomagnetic beads. DC lipid content was assessed by flow cytometry, immune fluorescence, and electron microscopy and by measuring intracellular component lipids. DC activation was determined from surface phenotype and cytokine profile. DC function was assessed in T-cell, natural killer (NK) cell, and NKT cell coculture assays as well as in vivo. RESULTS: We observed 2 distinct populations of hepatic DCs in mice and humans based on their lipid content and expression of markers associated with adipogenesis and lipid metabolism. This lipid-based dichotomy in DCs was unique to the liver and specific to DCs compared with other hepatic immune cells. However, rather than mediate tolerance, the liver DC population with high concentrations of lipid was immunogenic in multiple models; they activated T cells, NK cells, and NKT cells. Conversely, liver DCs with low levels of lipid induced regulatory T cells, anergy to cancer, and oral tolerance. The immunogenicity of lipid-rich liver DCs required their secretion of tumor necrosis factor α and was directly related to their high lipid content; blocking DC synthesis of fatty acids or inhibiting adipogenesis (by reducing endoplasmic reticular stress) reduced DC immunogenicity. CONCLUSIONS: Human and mouse hepatic DCs are composed of distinct populations that contain different concentrations of lipid, which regulates immunogenic versus tolerogenic responses in the liver.

Dendritic cells promote pancreatic viability in mice with acute pancreatitis.

BACKGROUND & AIMS: The cellular mediators of acute pancreatitis are incompletely understood. Dendritic cells (DCs) can promote or suppress inflammation, depending on their subtype and context. We investigated the roles of DC in development of acute pancreatitis. METHODS: Acute pancreatitis was induced in CD11c.DTR mice using caerulein or L-arginine; DCs were depleted by administration of diphtheria toxin. Survival was analyzed using Kaplan-Meier method. RESULTS: Numbers of major histocompatibility complex II(+)CD11c(+) DCs increased 100-fold in pancreata of mice with acute pancreatitis to account for nearly 15% of intrapancreatic leukocytes. Intrapancreatic DCs acquired a distinct immune phenotype in mice with acute pancreatitis; they expressed higher levels of major histocompatibility complex II and CD86 and increased production of interleukin-6, membrane cofactor protein-1, and tumor necrosis factor-α. However, rather than inducing an organ-destructive inflammatory process, DCs were required for pancreatic viability; the exocrine pancreas died in mice that were depleted of DCs and challenged with caerulein or L-arginine. All mice with pancreatitis that were depleted of DCs died from acinar cell death within 4 days. Depletion of DCs from mice with pancreatitis resulted in neutrophil infiltration and increased levels of systemic markers of inflammation. However, the organ necrosis associated with depletion of DCs did not require infiltrating neutrophils, activation of nuclear factor-κB, or signaling by mitogen-activated protein kinase or tumor necrosis factor-α. CONCLUSIONS: DCs are required for pancreatic viability in mice with acute pancreatitis and might protect organs against cell stress.

Dendritic cell depletion exacerbates acetaminophen hepatotoxicity.

UNLABELLED: Acetaminophen (APAP) overdose is one of the most frequent causes of acute liver failure in the United States and is primarily mediated by toxic metabolites that accumulate in the liver upon depletion of glutathione stores. However, cells of the innate immune system, including natural killer (NK) cells, neutrophils, and Kupffer cells, have also been implicated in the centrilobular liver necrosis associated with APAP. We have recently shown that dendritic cells (DCs) regulate intrahepatic inflammation in chronic liver disease and, therefore, postulated that DC may also modulate the hepatotoxic effects of APAP. We found that DC immune-phenotype was markedly altered after APAP challenge. In particular, liver DC expressed higher MHC II, costimulatory molecules, and Toll-like receptors, and produced higher interleukin (IL)-6, macrophage chemoattractant protein-1 (MCP-1), and tumor necrosis factor alpha (TNF-α). Conversely, spleen DC were unaltered. However, APAP-induced centrilobular necrosis, and its associated mortality, was markedly exacerbated upon DC depletion. Conversely, endogenous DC expansion using FMS-like tyrosine kinase 3 ligand (Flt3L) protected mice from APAP injury. Our mechanistic studies showed that APAP liver DC had the particular capacity to prevent NK cell activation and induced neutrophil apoptosis. Nevertheless, the exacerbated hepatic injury in DC-depleted mice challenged with APAP was independent of NK cells and neutrophils or numerous immune modulatory cytokines and chemokines. CONCLUSION: Taken together, these data indicate that liver DC protect against APAP toxicity, whereas their depletion is associated with exacerbated hepatotoxicity.

In hepatic fibrosis, liver sinusoidal endothelial cells acquire enhanced immunogenicity.

The normal liver is characterized by immunologic tolerance. Primary mediators of hepatic immune tolerance are liver sinusoidal endothelial cells (LSECs). LSECs block adaptive immunogenic responses to Ag and induce the generation of T regulatory cells. Hepatic fibrosis is characterized by both intense intrahepatic inflammation and altered hepatic immunity. We postulated that, in liver fibrosis, a reversal of LSEC function from tolerogenic to proinflammatory and immunogenic may contribute to both the heightened inflammatory milieu and altered intrahepatic immunity. We found that, after fibrotic liver injury from hepatotoxins, LSECs become highly proinflammatory and secrete an array of cytokines and chemokines. In addition, LSECs gain enhanced capacity to capture Ag and induce T cell proliferation. Similarly, unlike LSECs in normal livers, in fibrosis, LSECs do not veto dendritic cell priming of T cells. Furthermore, whereas in normal livers, LSECs are active in the generation of T regulatory cells, in hepatic fibrosis LSECs induce an immunogenic T cell phenotype capable of enhancing endogenous CTLs and generating potent de novo CTL responses. Moreover, depletion of LSECs from fibrotic liver cultures mitigates the proinflammatory milieu characteristic of hepatic fibrosis. Our findings offer a critical understanding of the role of LSECs in modulating intrahepatic immunity and inflammation in fibro-inflammatory liver disease.