Metabolomic landscape of macrophage discloses an anabolic signature of dengue virus infection and antibody-dependent enhancement of viral infection.

Dengue virus (DENV) infection causes dengue fever, the most prevalent arthropod-transmitted viral disease worldwide. Viruses are acellular parasites and obligately rely on host cell machinery for reproduction. Previous studies have indicated metabolomic changes in endothelial cell models and sera of animal models and patients with dengue fever. To probe the immunometabolic mechanism of DENV infection, here, we report the metabolomic landscape of a human macrophage cell model of DENV infection and its antibody-dependent enhancement. DENV infection of THP-1-derived macrophages caused 202 metabolic variants, of which amino acids occupied 23.7%, fatty acids 21.78%, carbohydrates 10.4%, organic acids 13.37%, and carnitines 10.4%. These metabolomic changes indicated an overall anabolic signature, which was characterized by the global exhaustion of amino acids, increases of cellular fatty acids, carbohydrates and pentoses, but decreases of acylcarnitine. Significant activation of metabolic pathways of glycolysis, pentose phosphate, amino acid metabolism, and tricarboxylic acid cycle collectively support the overall anabolism to meet metabolic demands of DENV replication and immune activation by viral infection. Totally 88 of 202 metabolic variants were significantly changed by DENV infection, 36 of which met the statistical standard (P<0.05, VIP>1.5) of differentially expressed metabolites, which were the predominantly decreased variants of acylcarnitine and the increased variants of fatty acids and carbohydrates. Remarkably, 11 differentially expressed metabolites were significantly distinct between DENV only infection and antibody-dependent enhancement of viral infection. Our data suggested that the anabolic activation by DENV infection integrates the viral replication and anti-viral immune activation.

Proteomic and metabonomic analysis uncovering Enterovirus A71 reprogramming host cell metabolic pathway.

Enterovirus A71 (EV71) infection can cause hand, foot, and mouth disease (HFMD) and severe neurological complications in children. However, the biological processes regulated by EV71 remain poorly understood. Herein, proteomics and metabonomics studies were conducted to uncover the mechanism of EV71 infection in rhabdomyosarcoma (RD) cells and identify potential drug targets. Differential expressed proteins from enriched membrane were analyzed by isobaric tags for relative and absolute quantitation (iTRAQ)-based proteomics technology. Twenty-six differential proteins with 1.5-fold (p < 0.05) change were detected, including 14 upregulated proteins and 12 downregulated proteins. The upregulated proteins are mainly involved in metabolic process, especially in the glycolysis pathway. Alpha-enolase (ENO1) protein was found to increase with temporal dependence following EV71 infection. The targeted metabolomics analysis revealed that glucose absorption and glycolysis metabolites were increased after EV71 infection. The glycolysis pathway was inhibited by knocking down ENO1 or the use of a glycolysis inhibitor (dichloroacetic acid [DCA]); and we found that EV71 infection was inhibited by depleting ENO1 or using DCA. Our study indicates that EV71 may reprogram glucose metabolism by activating glycolysis, and EV71 infection can be inhibited by interrupting the glycolysis pathway. ENO1 may be a potential target against EV71, and DCA could act as an inhibitor of EV71.

Sensitivity and specificity of magnetic resonance imaging in routine diagnosis of pulmonary lesions: a comparison with computed tomography.

Background: State-of-the-art thoracic magnetic resonance imaging (MRI) plays a complementary role in the assessment of pulmonary nodules/masses which potentially indicate to cancer. We aimed to evaluate the sensitivity and specificity of MRI in diagnosis of pulmonary nodules/masses. Methods: Sixty-eight patients with computed tomography (CT)-detected pulmonary nodules/masses underwent 3T MRI (T1-VIBE, T1-starVIBE, T2-fBLADE turbo spin-echo, and T2-SPACE). The detection rate was calculated for each of the different subgroups of pulmonary nodules according to lung imaging reporting and data system (Lung-RADS). The four MRI sequences were compared in terms of detection rate and image quality-signal to noise ratio (SNR), contrast to noise ratio (CNR) and 5-point scoring scale. Agreement of lesion size measurement between CT and MRI was assessed by intraclass correlation coefficient (ICC). The picture-SNR, lesion-SNR and CNR of each sequence were analyzed by Mann-Whitney U test. Results: In total, 232 pulmonary lesions were detected by CT. The CT showed 86 solid nodules (SNs) <6 mm, 15 SNs between 6-8 mm, 35 SNs between 8-15 mm, and 52 SNs between 15-30 mm. The T1-VIBE, T1-starVIBE, T2-fBLADE TSE and T2-SPACE sequences accurately detected 141 SNs (141/188, 75%/83.3%), 150 SNs (150/188, 79.8%/100%), 166 SNs (166/188, 88.3%/66.7%) and 169 SNs (169/188, 89.9%/53.3%), respectively. Four ground glass nodules (GGNs) (4/6) were detected by T2-fBLADE TSE. Twelve part-solid nodules (PSNs) (12/22) were detected by T1-VIBE and 20 PSNs (20/22) by T2-SPACE. A total of 100 lesions (2.2±1.4 cm, 0.8-7.3 cm) were accurately detected and measured by the four MRI sequences with ICC >0.96. The picture-SNR, lesion-SNR and CNR by T1-starVIBE were higher than those by T1-VIBE (P<0.001). The lesion-SNR and CNR by T2-fBLADE TSE were higher than those by T2-SPACE (P=0.006, 0.038). 86% of images by T1-starVIBE, 92% by T2-fBLADE TSE, 90% by T2-SPACE and 93% by T1-VIBE were scored 3 or more. Conclusions: MRI achieves high sensitivity and specificity for different type of pulmonary nodules detection and is an effective alternative to CT as a diagnostic tool for pulmonary nodules.

PDIA2 Bridges Endoplasmic Reticulum Stress and Metabolic Reprogramming During Malignant Transformation of Chronic Colitis.

Background: Chronic inflammation contributes to approximately 20% of cancers; the underlying mechanisms are still elusive. Here, using an animal model of colitis to colon-cancerous transformation, we demonstrated that endoplasmic reticulum (ER) stress couples with metabolic reprogramming to promote a malignant transformation of chronic inflammation. Methods: The animal model for chronic colitis to colon-cancerous transformation was established in C57BL/6N mice by azoxymethane (AOM) and dextran sodium sulfate (DSS) treatments. The differential proteins in control and AOM/DSS-treated colon mucosa were determined using proteomic analysis; the kinetics of metabolic modifications were monitored by mitochondrial oxygen flux, extracellular acidification, and targeted metabolomics; the molecule linker between ER stress and metabolic modifications were identified by coimmunoprecipitation, KEGG pathway analysis, and the subcutaneous tumor model using gene-specific knockdown colon cancer cells. Tissue array analysis were used to evaluate the differential protein in cancer and cancer-adjacent tissues. Results: AOM/DSS treatment induced 38 tumors in 10 mice at the 14th week with the mean tumor size 9.35 ± 3.87 mm2, which was significantly decreased to 5.85 ± 0.95 mm2 by the ER stress inhibitor 4-phenylbutyric acid (4PBA). Seven differential proteins were determined from control (1,067 ± 48) and AOM/DSS-treated mucosa (1,077 ± 59); the level of ER protein PDIA2 (protein disulfide isomerase-associated 2) was increased over 7-fold in response to AOM/DSS treatment. PDIA2 interacted with 420 proteins that were involved in 8 signaling pathways, in particular with 53 proteins in metabolic pathways. PDIA2 translocated from ER to mitochondria and interacted with the components of complexes I and II to inhibit oxophosphorylation but increase glycolysis. Knockdown PDIA2 in colon cancer cells restored the metabolic imbalance and significantly repressed tumor growth in the xenograft animal model. 4PBA therapy inhibited the AOM/DSS-mediated overexpression of PDIA2 and metabolic modifications and suppressed colon cancer growth. In clinic, PDIA2 was overexpressed in colon cancer tissues rather than cancer-adjacent tissues and was related with the late stages and lymph node metastasis of colon cancer. Conclusions: Persistent ER stress reprograms the metabolism to promote the malignant transformation of chronic colitis; PDIA2 serves as a molecule linker between ER stress and metabolic reprogramming. The inhibition of ER stress restores metabolic homeostasis and attenuates the cancerous transformation of chronic inflammation.

Hepatomas are exquisitely sensitive to pharmacologic ascorbate (P-AscH-).

Rationale: Ascorbate is an essential micronutrient known for redox functions at normal physiologic concentrations. In recent decades, pharmacological ascorbate has been found to selectively kill tumour cells. However, the dosing frequency of pharmacologic ascorbate in humans has not yet been defined. Methods: We determined that among five hepatic cell lines, Huh-7 cells were the most sensitive to ascorbate. The effects of high-dose ascorbate on hepatoma were therefore assessed using Huh-7 cells and xenograft tumour mouse model. Results: In Huh-7 cells, ascorbate induced a significant increase in the percentage of cells in the G0/G1 phase, apoptosis and intracellular levels of ROS. High doses of ascorbate (4.0 pmol cell-1), but not low doses of ascorbate (1.0 pmol cell-1), also served as a pro-drug that killed hepatoma cells by altering mitochondrial respiration. Furthermore, in a Huh-7 cell xenograft tumour mouse model, intraperitoneal injection of ascorbate (4.0 g/kg/3 days) but not a lower dose of ascorbate (2.0 g/kg/3 days) significantly inhibited tumour growth. Gene array analysis of HCC tumour tissue from xenograft mice given IP ascorbate (4.0 g/kg/3 days) identified changes in the transcript levels of 192 genes/ncRNAs involved in insulin receptor signalling, metabolism and mitochondrial respiration. Consistent with the array data, gene expression levels of AGER, DGKK, ASB2, TCP10L2, Lnc-ALCAM-3, and Lnc-TGFBR2-1 were increased 2.05-11.35 fold in HCC tumour tissue samples from mice treated with high-dose ascorbate, and IHC staining analysis also verified that AGER/RAGE and DGKK proteins were up-regulated, which implied that AGER/RAGE and DGKK activation might be related to oxidative stress, leading to hepatoma cell death. Conclusions: Our studies identified multiple mechanisms are responsible for the anti-tumour activity of ascorbate and suggest high doses of ascorbate with less frequency will act as a novel therapeutic agent for liver cancer in vivo.

Frontline Science: Monocytes sequentially rewire metabolism and bioenergetics during an acute inflammatory response.

Metabolism directs the severe acute inflammatory reaction of monocytes to guard homeostasis. This occurs by sequentially activating anabolic immune effector mechanisms, switching to immune deactivation mechanisms and then restoring immunometabolic homeostasis. Nuclear sirtuin 1 and mitochondrial pyruvate dehydrogenase kinase metabolically drive this dynamic and are druggable targets that promote immunometabolic resolution in septic mice and increase survival. We used unbiased metabolomics and a validated monocyte culture model of activation, deactivation, and partial resolution of acute inflammation to sequentially track metabolic rewiring. Increases in glycogenolysis, hexosamine, glycolysis, and pentose phosphate pathways were aligned with anabolic activation. Activation transitioned to combined lipid, protein, amino acid, and nucleotide catabolism during deactivation, and partially subsided during early resolution. Lipid metabolic rewiring signatures aligned with deactivation included elevated n-3 and n-6 polyunsaturated fatty acids and increased levels of fatty acid acylcarnitines. Increased methionine to homocysteine cycling increased levels of s-adenosylmethionine rate-limiting transmethylation mediator, and homocysteine and cysteine transsulfuration preceded increases in glutathione. Increased tryptophan catabolism led to elevated kynurenine and de novo biosynthesis of nicotinamide adenine dinucleotide from quinolinic acid. Increased branched-chain amino acid catabolism paralleled increases in succinyl-CoA. A rise in the Krebs cycle cis-aconitate-derived itaconate and succinate with decreased fumarate and acetyl-CoA levels occurred concomitant with deactivation and subsided during early resolution. The data suggest that rewiring of metabolic and mitochondrial bioenergetics by monocytes sequentially activates, deactivates, and resolves acute inflammation.

Bufalin reverses intrinsic and acquired drug resistance to cisplatin through the AKT signaling pathway in gastric cancer cells.

Cisplatin is the most common chemotherapeutic agent for gastric cancer (GC), however it activates AKT, which contributes to intrinsic and acquired resistance. Bufalin, a traditional Chinese medicine, shows significant anticancer activity by inhibiting the AKT pathway. It was therefore hypothesized that bufalin could counteract cisplatin resistance in GC cells. SGC7901, MKN­45 and BGC823 human GC cells were cultured under normoxic and hypoxic conditions. Effects of cisplatin and bufalin on GC cells were measured by a cell counting kit, apoptosis was analyzed by flow cytometry, and immunoblotting was used to detect proteins associated with the AKT signaling pathway. It was demonstrated that bufalin synergized with cisplatin to inhibit proliferation and promote apoptosis of GC cells by diminishing the activation of cisplatin-induced AKT under normoxic and hypoxic conditions. Bufalin also inhibits cisplatin-activated molecules downstream of AKT that affect proliferation and apoptosis, including glycogen synthase kinase, mammalian target of rapamycin, ribosomal protein S6 Kinase and eukaryotic translation initiation factor-4E-binding protein-1. To investigate acquired cisplatin resistance, a cisplatin­resistant cell line SGC7901­CR was used. It was demonstrated that bufalin reversed acquired cisplatin resistance and significantly induced apoptosis through the AKT pathway. These results imply that bufalin could extend the therapeutic effect of cisplatin on GC cells when administered in combination.

Sirtuins Link Inflammation and Metabolism.

Sirtuins (SIRT), first discovered in yeast as NAD+ dependent epigenetic and metabolic regulators, have comparable activities in human physiology and disease. Mounting evidence supports that the seven-member mammalian sirtuin family (SIRT1-7) guard homeostasis by sensing bioenergy needs and responding by making alterations in the cell nutrients. Sirtuins play a critical role in restoring homeostasis during stress responses. Inflammation is designed to "defend and mend" against the invading organisms. Emerging evidence supports that metabolism and bioenergy reprogramming direct the sequential course of inflammation; failure of homeostasis retrieval results in many chronic and acute inflammatory diseases. Anabolic glycolysis quickly induced (compared to oxidative phosphorylation) for ROS and ATP generation is needed for immune activation to "defend" against invading microorganisms. Lipolysis/fatty acid oxidation, essential for cellular protection/hibernation and cell survival in order to "mend," leads to immune repression. Acute/chronic inflammations are linked to altered glycolysis and fatty acid oxidation, at least in part, by NAD+ dependent function of sirtuins. Therapeutically targeting sirtuins may provide a new class of inflammation and immune regulators. This review discusses how sirtuins integrate metabolism, bioenergetics, and immunity during inflammation and how sirtuin-directed treatment improves outcome in chronic inflammatory diseases and in the extreme stress response of sepsis.

Bufalin promotes apoptosis of gastric cancer by down-regulation of miR-298 targeting bax.

Bufalin is used to treat many patients with solid malignant tumors clinically. Bufalin could induce gastric cancer cell apoptosis via BAX. microRNA (miRNA) plays important roles in gene regulation. However, miRNA involving in bufalin inducing apoptosis of gastric cancer cells remains to futher research. To study the regulatory role of miRNA in bufalin induced cancer cell apoptosis. Firstly, we verifed that bufalin could induce gastric cancer cell apoptosis by inducing BAX expression. miR-298 was predicted as a regulator of BAX and further study verified Bax was a target gene of miR-298 by luciferase reporter assay. miR-298 could down-regulate BAX on mRNA and protein level in gastric cancer cells. miR-298 promoted cell proliferation and inhibited apoptosis of gastric cancer cells. It was also found that bufalin inhibited cell proliferation and promoted cell apoptosis by down-regualtion of miR-298. In summary, bufalin-associated miR-298 may indirectly be involved in cell proliferation and apoptosis by targeting BAX, pointing to use as a potential molecular target in gastric cancer therapy.

The identification of specific methylation patterns across different cancers.

Abnormal DNA methylation is known as playing an important role in the tumorgenesis. It is helpful for distinguishing the specificity of diagnosis and therapeutic targets for cancers based on characteristics of DNA methylation patterns across cancers. High throughput DNA methylation analysis provides the possibility to comprehensively filter the epigenetics diversity across various cancers. We integrated whole-genome methylation data detected in 798 samples from seven cancers. The hierarchical clustering revealed the existence of cancer-specific methylation pattern. Then we identified 331 differentially methylated genes across these cancers, most of which (266) were specifically differential methylation in unique cancer. A DNA methylation correlation network (DMCN) was built based on the methylation correlation between these genes. It was shown the hubs in the DMCN were inclined to cancer-specific genes in seven cancers. Further survival analysis using the part of genes in the DMCN revealed high-risk group and low-risk group were distinguished by seven biomarkers (PCDHB15, WBSCR17, IGF1, GYPC, CYGB, ACTG2, and PRRT1) in breast cancer and eight biomarkers (ZBTB32, OR51B4, CCL8, TMEFF2, SALL3, GPSM1, MAGEA8, and SALL1) in colon cancer, respectively. At last, a protein-protein interaction network was introduced to verify the biological function of differentially methylated genes. It was shown that MAP3K14, PTN, ACVR1 and HCK sharing different DNA methylation and gene expression across cancers were relatively high degree distribution in PPI network. The study suggested that not only the identified cancer-specific genes provided reference for individual treatment but also the relationship across cancers could be explained by differential DNA methylation.

SIRT1 inhibition during the hypoinflammatory phenotype of sepsis enhances immunity and improves outcome.

Mechanism-based sepsis treatments are unavailable, and their incidence is rising worldwide. Deaths occur during the early acute phase of hyperinflammation or subsequent postacute hypoinflammatory phase with sustained organ failure. The acute sepsis phase shifts rapidly, and multiple attempts to treat early excessive inflammation have uniformly failed. We reported in a sepsis cell model and human sepsis blood leukocytes that nuclear NAD+ sensor SIRT1 deacetylase remodels chromatin at specific gene sets to switch the acute-phase proinflammatory response to hypoinflammatory. Importantly, SIRT1 chromatin reprogramming is reversible, suggesting that inhibition of SIRT1 might reverse postacute-phase hypoinflammation. We tested this concept in septic mice, using the highly specific SIRT1 inhibitor EX-527, a small molecule that closes the NAD+ binding site of SIRT1. Strikingly, when administered 24 h after sepsis, all treated animals survived, whereas only 40% of untreated mice survived. EX-527 treatment reversed the inability of leukocytes to adhere at the small intestine MVI, reversed in vivo endotoxin tolerance, increased leukocyte accumulation in peritoneum, and improved peritoneal bacterial clearance. Mechanistically, the SIRT1 inhibitor restored repressed endothelial E-selectin and ICAM-1 expression and PSGL-1 expression on the neutrophils. Systemic benefits of EX-527 treatment included stabilized blood pressure, improved microvascular blood flow, and a shift toward proimmune macrophages in spleen and bone marrow. Our findings reveal that modifying the SIRT1 NAD+ axis may provide a novel way to treat sepsis in its hypoinflammatory phase.

Construction of pancreatic cancer double-factor regulatory network based on chip data on the transcriptional level.

Transcription factor (TF) and microRNA (miRNA) have been discovered playing crucial roles in cancer development. However, the effect of TFs and miRNAs in pancreatic cancer pathogenesis remains vague. We attempted to reveal the possible mechanism of pancreatic cancer based on transcription level. Using GSE16515 datasets downloaded from gene expression omnibus database, we first identified the differentially expressed genes (DEGs) in pancreatic cancer by the limma package in R. Then the DEGs were mapped into DAVID to conduct the kyoto encyclopedia of genes and genomes (KEGG) pathway enrichment analysis. TFs and miRNAs that DEGs significantly enriched were identified by Fisher's test, and then the pancreatic cancer double-factor regulatory network was constructed. In our study, total 1117 DEGs were identified and they significantly enriched in 4 KEGG pathways. A double-factor regulatory network was established, including 29 DEGs, 24 TFs, 25 miRNAs. In the network, LAMC2, BRIP1 and miR155 were identified which may be involved in pancreatic cancer development. In conclusion, the double-factor regulatory network was found to play an important role in pancreatic cancer progression and our results shed new light on the molecular mechanism of pancreatic cancer.

Blocking autophagy enhances the apoptosis effect of bufalin on human hepatocellular carcinoma cells through endoplasmic reticulum stress and JNK activation.

Bufalin extracts are a part of traditional Chinese medicine, Chansu. In the current study, we investigated the effect of bufalin on the proliferation of the human hepatocellular carcinoma (HCC) cell lines, Huh-7 and HepG-2, and explored the therapeutic potential of the drug. Our results demonstrated that bufalin markedly inhibited cell proliferation and promoted apoptosis in the Huh-7 and HepG-2 cells in vitro. The underlying mechanism of the bufalin-induced apoptosis was the induction of endoplasmic reticulum (ER) stress via the IRE1-JNK pathway. In addition, during the ER stress response, the autophagy pathway, characterized by the conversion of LC3-I to LC3-II, was activated, resulting in increased Beclin-1 protein levels, decreased p62 expression and stimulation of autophagic flux. Our data supported the pro-survival role of bufalin-induced autophagy when the autophagy pathway was blocked with specific chemical inhibitors; the involvement of the IRE1 pathway in the ER stress-induced autophagy was also demonstrated when the expression of IRE1 and CHOP was silenced using siRNA. These data indicate that combining bufalin with a specific autophagy inhibitor could be a promising therapeutic approach for the treatment of HCC.

Antitumor and immunomodulating activity of a polysaccharide from Sophora flavescens Ait.

The immunostimulatory activity of Sophora flavescens polysaccharide (SFPW1) was evaluated by using in vitro cell models and in vivo animal models. The results demonstrated that SFPW1 could effectively inhibit the tumor growth in H22 tumor-bearing mice and promote the splenocyte proliferation, thus resulting in a prolonged life survival. For assay in vitro, SFPW1 significantly strengthened peritoneal macrophages to devour H22 tumor cells and stimulated macrophages to produce nitric oxide (NO) via up-regulation of inducible NO synthase (iNOS) activity. However, no direct cytotoxicity against H22 tumor cells was observed in vitro. These results suggest that SFPW1 might be a strong natural immunomodulator and the antitumor effect of this polysaccharide is associated with its potent immunostimulating effect.

MicroRNA-146a regulates both transcription silencing and translation disruption of TNF-α during TLR4-induced gene reprogramming.

Following the TLR-dependent initiation phase of acute systemic proinflammatory responses such as sepsis, an adaptive phase represses or activates a specific pattern of gene expression until the inflammation resolves. Here, we used the THP-1 sepsis cell model of bacterial LPS/endotoxin tolerance to show that TLR4-induced miR-146a supports the feed-forward adaptive processes that silence transcription and disrupt translation of acute proinflammatory genes. First, we found that miR-146a regulates a pathway that promotes the binding of transcription repressor RelB to the TNF-α promoter, a step known to precede histone and DNA modifications, which generate facultative heterochromatin to silence acute proinflammatory genes. However, once RelB binding occurred, miR-146a inhibition could not reverse compacted chromatin, and endotoxin tolerance persisted. Second, we observed that miR-146a regulates a pathway that supports assembly of the translation repressor complex of TNF-α by preventing the interaction of the RNA-binding protein effector Ago2 and RBM4. We also determined that once endotoxin tolerance is established, and specific genes have been reprogrammed, transcription and translation disruption can be reversed only by simultaneously depleting RelB and inhibiting miR-146a. Thus, miR-146a induction supports the TLR4-dependent shift from initiation to gene-specific repression at two levels. Our results also imply that therapies designed to reverse endotoxin tolerance as potential therapies for sepsis should be directed at the transcription and translation pathways of reprogramming.

Dynamic and selective nucleosome repositioning during endotoxin tolerance.

Sepsis is encoded by a sequel of transcription activation and repression events that initiate, sustain, and resolve severe systemic inflammation. The repression/silencing phase occurs in blood leukocytes of animals and humans following the initiation of systemic inflammation due to developing endotoxin tolerance. We previously reported that NF-kappaB transcription factor RelB and histone H3 lysine methyltransferase G9a directly interact to induce facultative heterochromatin assembly and regulate epigenetic silencing during endotoxin tolerance, which is a major feature of sepsis. The general objective of this study was to assess whether dynamic temporal, structural, and positional changes of nucleosomes influence the sepsis phenotype. We used the THP-1 sepsis cell model to isolate mononucleosomes by rapid cell permeabilization and digestion of chromatin with micrococcal nuclease and then compared tumor necrosis factor alpha (TNFalpha) proximal promoter nucleosome alignment in endotoxin-responsive and -tolerant phenotypes. We found differential and dynamic repositioning of nucleosomes from permissive to repressive locations during the activation and silencing phases of transcription reprogramming and identified the following mechanisms that may participate in the process. 1) Two proximal nucleosomes repositioned to expose the primary NF-kappaB DNA binding site in endotoxin-responsive cells, and this "promoter opening" required the ATP-independent chaperone NAP1 to replace the core histone H2A with the H2A.Z variant. 2) During RelB-dependent endotoxin tolerance, the two nucleosomes repositioned and masked the primary NF-kappaB DNA binding site. 3) Small interfering RNA-mediated inhibition of RelB expression prevented repressive nucleosome repositioning and tolerance induction, but the "open" promoter required endotoxin-induced NF-kappaB p65 promoter binding to initiate transcription, supporting the known requirement of p65 posttranslational modifications for transactivation. 4) Sustaining the permissive promoter state after RelB knockdown required ATP-dependent nucleosome remodeler BAF complex. Moreover, we found that forced expression of RelB in responsive cells induced repressive nucleosome positioning and silenced TNFalpha transcription, demonstrating the plasticity of nucleosome remodeling and its dependence on RelB. Our data suggest that nucleosome repositioning controls both the induction and epigenetic silencing phases of TNFalpha transcription associated with sepsis.

Arsenic trioxide inhibits metastatic potential of mouse hepatoma H22 cells in vitro and in vivo.

BACKGROUND: It has been pointed out that only low-dose arsenic trioxide (ATO) presents therapeutic benefits outweighing the toxic side effects. Low-dose ATO can effectively alleviate acute promyelocytic leukemia (APL). However, it is quite challenging in treating solid tumors. The purpose of this study was to investigate the effect of ATO at low concentrations on the metastatic potential of mouse hepatoma H(22) cells and the anti-metastatic mechanism of ATO. METHODS: The metastatic potential of H(22) cells was evaluated by adhesion, migration and invasion assays after exposure to a low dose of ATO in vitro. The mouse lung metastatic model induced by injection of H(22) cells via the tail vein was adopted for the evaluation of metastatic potential. Different proteins in the lysate of H(22) cells exposed to ATO at different concentrations were investigated by surface-enhanced laser desorption and ionization time-of-flight mass spectrometry (SELDI-TOF-MS). Finally, Western blotting analyses were made to detect the expression pattern of MMP-2 and nm23-M1 proteins. RESULTS: Significant cell death started at ATO concentrations above 2 micromol/L. The growth and adhesion potential of H(22) cells was inhibited in a time- and dose-dependent manner, and the migration and invasion potential of H(22) cells was inhibited in a dose-dependent manner while ATO concentration was below 2 micromol/L. Mice injected with ATO at a dose of 0.5 mg/kg had fewer lung metastases. However, mice injected with ATO at a dose of 2 mg/kg or 4 mg/kg had a high mortality rate and more liver injuries. A total of 15 different protein peaks were identified between the lysate of H(22) cells treated with ATO and controls. Two proteins that peaked at m/z 5302 and 17207 coincided with MMP-2 (fragment) and nm23-M1, respectively. Western blotting analyses demonstrated that MMP-2 and MMP-2 fragments were down-regulated and nm23-M1 was up-regulated in H(22) cells treated with 2 micromol/L ATO for 48 hours. CONCLUSIONS: ATO at a low dose inhibits the metastatic potential of mouse hepatoma H(22) cells in vitro and in vivo, and involves down-regulation of MMP-2 and up-regulation of nm23-M1.

Different cell death modes of pancreatic acinar cells on macrophage activation in rats.

BACKGROUND: The pathogenesis of acute pancreatitis is complex and largely unclear. The aim of this study was to explore the relationship between modes of cell death in pancreatic acinar cells, the release of cell contents and the inflammatory response of macrophages. METHODS: Our experiment included four groups: group A (the control group), group B (AR42J cells overstimulated by caerulein), group C (AR42J cells treated with lipopolysaccharide and caerulein), and group D (AR42J cells treated with octreotide and caerulein). Apoptosis and oncosis, and the release of amylase and lactate dehydrogenase (LDH) from AR42J cells were detected. Rat macrophages were stimulated by 1 ml supernatant of culture medium of AR42J cells. Finally, NF-kappaB activation and TNF-alpha and IL-1beta secretion by macrophages were detected. RESULTS: Oncotic cells in group C increased while apoptotic cells decreased (P < 0.05); cells in group D had the inverse reaction. The release of amylase and LDH changed directly with the occurrence of oncosis. The transcription factor NF-kappaB was activated and secretion of TNF-alpha and IL-1beta were significantly higher in group C than in group B (P < 0.05); in group D, these actions were significantly lower than in group B (P < 0.05). This trend was in line with changes in amylase and LDH production. CONCLUSION: There is a close relationship between modes of pancreatic acinar cell death, the release of cell contents and the inflammatory reaction of macrophages.

A retrospective cohort study among iron-steel workers in Anshan, China: exposure assessment.

Although adequate assessment of exposure is needed in epidemiological studies among foundry workers, previous studies are often lacking in this aspect. We conducted a retrospective cohort study of a Chinese iron and steel company with a 14-yr follow up during 1980-1993. Exposure assessment was performed for a single job, i.e., the current job for the active worker and the longest job for the retired or deceased worker as of the end of the follow-up, which was allocated as the surrogate of lifetime job and was applied to a job-exposure matrix. Of the 147,062 cohort members, 52,394 males (43%) and 5,291 females (21%) were exposed to any of 15 hazardous factors such as dust, silica, PAHs (polycyclic aromatic hydrocarbons), CO (carbon monoxide) and heat. In 2,104 randomly selected samples, the exposure assessment of exposed workers based on a single job was found to be 12-14% lower than the real situation. This study suggests that the exposure assessment is valuable in evaluating the health effects among the foundry workers, despite some limitations such as underestimation of exposure assessment and the lack of data regarding smoking and drinking habits.