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Acute pancreatitis is an inflammatory process of the pancreatic gland that may lead to dysregulation of the trans-sulfuration pathway. The aims of this work were firstly to study the methionine cycle as well as the trans-sulfuration pathway using metabolomic and proteomic approaches identifying the causes of this dysregulation in an experimental model of acute pancreatitis; and secondly to reveal the effects of S-adenosylmethionine administration on these pathways. Acute pancreatitis was induced by cerulein in mice, and a group of animals received S-adenosylmethionine treatment. Cerulein-induced acute pancreatitis rapidly caused marked depletion of methionine, S-adenosylmethionine, 5'-methylthioadenosine, cystathionine, cysteine, and glutathione levels in pancreas, but S-adenosylhomocysteine and homocysteine remained unchanged. Protein steady-state levels of S-adenosylhomocysteine-hydrolase and cystathionine gamma-lyase diminished but methylthioadenosine phosphorylase levels increased in pancreas with acute pancreatitis. Although cystathionine beta-synthase protein levels did not change with acute pancreatitis, Nos2 mRNA and protein levels were markedly up-regulated and caused tyrosine nitration of cystathionine beta-synthase in pancreas. S-adenosylmethionine administration enhanced Nos2 mRNA expression and cystathionine beta-synthase nitration and triggered homocysteine accumulation in acute pancreatitis. Furthermore, S-adenosylmethionine administration promoted enrichment of the euchromatin marker H3K4me3 in the promoters of Tnf-alpha, Il-6, and Nos2 and enhanced the mRNA up-regulation of these genes. Accordingly, S-adenosylmethionine administration increased inflammatory infiltrate and edema in pancreas with acute pancreatitis. In conclusion, tyrosine-nitration of cystathionine beta-synthase blockades the trans-sulfuration pathway in acute pancreatitis promoting homocysteine accumulation upon S-adenosylmethionine treatment. Copyright © 2019. Published by Elsevier B.V.

Chromatin remodeling seems to regulate the patterns of proinflammatory genes. Our aim was to provide new insights into the epigenetic mechanisms that control transcriptional activation of early-and late-response genes in initiation and development of severe acute pancreatitis as a model of acute inflammation. Chromatin changes were studied by chromatin immunoprecipitation analysis, nucleosome positioning, and determination of histone modifications in promoters of proinflammatory genes in vivo in the course of taurocholate-induced necrotizing pancreatitis in rats and in vitro in rat pancreatic AR42J acinar cells stimulated with taurocholate or TNF-alpha. Here we show that the upregulation of early and late inflammatory genes rely on histone acetylation associated with recruitment of histone acetyltransferase CBP. Chromatin remodeling of early genes during the inflammatory response in vivo is characterized by a rapid and transient increase in H3K14ac, H3K27ac, and H4K5ac as well as by recruitment of chromatin-remodeling complex containing BRG-1. Chromatin remodeling in late genes is characterized by a late and marked increase in histone methylation, particularly in H3K4. JNK and p38 MAPK drive the recruitment of transcription factors and the subsequent upregulation of early and late inflammatory genes, which is associated with nuclear translocation of the early gene Egr-1. In conclusion, specific and strictly ordered epigenetic markers such as histone acetylation and methylation, as well as recruitment of BRG-1-containing remodeling complex are associated with the upregulation of both early and late proinflammatory genes in acute pancreatitis. Our findings highlight the importance of epigenetic regulatory mechanisms in the control of the inflammatory cascade.

KRAS mutational status is considered a negative predictive marker of the response to anti-EGFR therapies in colorectal cancer (CRC) patients. However, conflicting data exist regarding the variable response to EGFR-targeted therapy. The effects of oncogenic KRAS on downstream targets were studied in cell lines with different KRAS mutations. Cells harboring a single KRAS(G13D) allele showed the most tumorigenic profile, with constitutive activation of the downstream pathway, rendering them EGF-unresponsive. Conversely, KRAS(A146T) cells showed a full EGF-response in terms of signal transduction pathways, cell proliferation, migration or adhesion. Moreover, the global acetylome of CRC cells was also dependent on KRAS mutational status. Several hnRNP family members were identified within the 36 acetylated-proteins. Acetylation status is known to be involved in the modulation of EGF-response. In agreement with results presented herein, hnRNPA1 and L acetylation was induced in response to EGF in KRAS(A146T) cells, whereas acetyl-hnRNPA1 and L levels remained unchanged after growth factor treatment in KRASG13D unresponsive cells. Our results showed that hnRNPs induced-acetylation is dependent on KRAS mutational status. Nevertheless hnRNPs acetylation might also be the point where different oncogenic pathways converge.

Multiple sclerosis (MS) is a complex debilitating disease of the central nervous system (CNS) perceived to result from the autoimmune effect of T cells in damaging myelin sheath. However, the exact pathogenesis of the disease remains elusive. Initial studies describing the possibility of defective pyruvate metabolism in MS were performed in 1950s. The group observed elevated blood pyruvate level in both fasting and postprandial times in MS patients with relapse. Similarly, other investigators also reported increased fasting pyruvate level in this disease. These reports hint to a possible abnormality of pyruvate metabolism in MS patients. In addition, increase in levels of Krebs cycle acids like alpha-ketoglutarate in fasting and citrate after glucose intake in MS patients further strengthened the connection of disturbed pyruvate metabolism with MS progression. These studies led the investigators to explore the role of disturbed glucose metabolism in pathophysiological brain function. Under normal circumstances, complex molecules are metabolized into simpler molecules through their respective pathways. Differential expression of genes encoding enzymes of the glucose metabolic pathway in CNS may result in neurological deficits. In this review article, we discuss the studies related to disturbed carbohydrate metabolism in MS and other neurodegenerative diseases. These observations open new perspectives for the understanding of metabolic dynamics in MS yet many puzzling aspects and critical questions need to be addressed. Much more research is required to fully unravel the disease mechanism, and a proper understanding of the disease could eventually lead to new treatments.

The inhibitor of differentiation Id2, a protein lacking the basic DNA-binding domain, is involved in the modulation of a number of biological processes. The molecular mechanisms explaining Id2 pleiotropic functions are poorly understood. Id2 and E2F4 are known to bind simultaneously to c-myc promoter. To study whether Id2 plays a global role on transcriptional regulation, we performed in vivo genome-wide ChIP/chip experiments for Id2 and E2F4 in adult mouse liver. An Id2-containing complex was bound to a common sequence downstream from the TSS on a subset of 442 E2F4 target genes mainly related to cell development and chromatin structure. We found a positive correlation between Id2 protein levels and the expression of E2F4/Id2 targets in fetal and adult liver. Id2 protein stability increased in fetal liver by interaction with USP1 de-ubiquitinating enzyme, which was induced during development. In adult liver, USP1 and Id2 levels dramatically decreased. In differentiated liver tissue, when Id2 concentration was low, E2F4/Id2 was bound to the same region as paused Pol II and target genes remained transcriptionally inactive. Conversely, in fetal liver when Id2 levels were increased, Id2 and Pol II were released from gene promoters and target genes up-regulated. During liver regeneration after partial hepatectomy, we obtained the same results as in fetal liver. Our results suggest that Id2 might be part of a reversible development-related program involved in the paused-ON/OFF state of Pol II on selected genes that would remain responsive to specific stimuli.

The epigenetic identity of oligodendrocytes is modulated by posttranslational modifications of histones. Acetylation of histone H3 results from the balance between the activity of histone acetyltransferases (HATs) and histone deacetylases and modulates transcriptional activation. We have previously shown that, in rodents, histone deacetylation favors oligodendrocyte differentiation, whereas acetylation is associated with increased levels of transcriptional inhibitors of oligodendrocyte differentiation. Here, we report, in humans brains, a shift toward histone acetylation in the white matter of the frontal lobes of aged subjects and in patients with chronic multiple sclerosis (MS). Increased immunoreactivity for acetylated histone H3 was observed in the nuclei of NogoA+ oligodendrocytes in a subset of MS samples. These changes were associated with high levels of transcriptional inhibitors of oligodendrocyte differentiation (i.e., TCF7L2, ID2, and SOX2) and higher HAT transcript levels (i.e., CBP, P300) in female MS patients compared with non-neurological controls and correlated with disease duration. Chromatin immunoprecipitation from samples of MS patients revealed enrichment of acetyl-histone H3 at the promoter of the increased target genes (i.e., TCF7L2). The data in chronic lesions contrasted with findings in early MS lesions, where a marked oligodendroglial histone deacetylation was observed. Together, these data suggest that histone deacetylation is a process that occurs at the early stages of the disease and whose efficiency decreases with disease duration.

Modifications on specific histone residues and DNA methylation play an essential role in lineage choice and cellular reprogramming. We have previously shown that histone modifications or combinatorial codes of transcription factors (TFs) are critical for the differentiation of multipotential progenitors into myelinating oligodendrocytes. In this study we asked whether combining global manipulation of DNA methylation and histone acetylation together with the expression of oligodendrocyte-specific TFs, was sufficient to switch the identity of fibroblasts into myelin gene-expressing cells.

Mitogen-activated protein kinases (MAPKs) are considered major signal transducers early during the development of acute pancreatitis. Pentoxifylline is a phosphodiesterase inhibitor with marked anti-inflammatory properties through blockade of extracellular signal regulated kinase (ERK) phosphorylation and tumor necrosis factor alpha production. Our aim was to elucidate the mechanism of action of pentoxifylline as an anti-inflammatory agent in acute pancreatitis. Necrotizing pancreatitis induced by taurocholate in rats and taurocholate-treated AR42J acinar cells were studied. Phosphorylation of ERK and ERK kinase (MEK1/2), as well as PP2A, PP2B, and PP2C serine/threonine phosphatase activities, up-regulation of proinflammatory genes (by reverse transcription-polymerase chain reaction and chromatin immunoprecipitation), and recruitment of transcription factors and histone acetyltransferases/deacetylases to promoters of proinflammatory genes (egr-1, atf-3, inos, icam, il-6, and tnf-alpha) were determined in the pancreas during pancreatitis. Pentoxifylline did not reduce MEK1/2 phosphorylation but prevented the marked loss of serine/threonine phosphatase PP2A activity induced by taurocholate in vivo without affecting PP2B and PP2C activities. The rapid loss in PP2A activity induced by taurocholate in acinar cells was due to a decrease in cAMP levels that was prevented by pentoxifylline. Pentoxifylline also reduced the induction of early (egr-1, atf-3) responsive genes and abrogated the up-regulation of late (inos, icam, il-6, tnf-alpha) responsive genes and recruitment of transcription factors (nuclear factor kappaB and C/EBPbeta) and histone acetyltransferases to their gene promoters during pancreatitis. In conclusion, the beneficial effects of pentoxifylline--and presumably of other phosphodiesterase inhibitors--in this disease seem to be mediated by abrogating the loss of cAMP levels and PP2A activity as well as chromatin-modifying complexes very early during acute pancreatitis.

Acute pancreatitis is an acute inflammatory process localized in the pancreatic gland that frequently involves peripancreatic tissues. It is still under investigation why an episode of acute pancreatitis remains mild affecting only the pancreas or progresses to a severe form leading to multiple organ failure and death. Proinflammatory cytokines and oxidative stress play a pivotal role in the early pathophysiological events of the disease. Cytokines such as interleukin 1beta and tumor necrosis factor alpha initiate and propagate almost all consequences of the systemic inflammatory response syndrome. On the other hand, depletion of pancreatic glutathione is an early hallmark of acute pancreatitis and reactive oxygen species are also associated with the inflammatory process. Changes in thiol homestasis and redox signaling decisively contribute to amplification of the inflammatory cascade through mitogen activated protein kinase (MAP kinase) pathways. This review focuses on the relationship between oxidative stress, pro-inflammatory cytokines and MAP kinase/protein phosphatase pathways as major modulators of the inflammatory response in acute pancreatitis. Redox sensitive signal transduction mediated by inactivation of protein phosphatases, particularly protein tyrosin phosphatases, is highlighted.

Methionine adenosyltransferase (MAT) is an essential enzyme because it catalyzes the formation of S-adenosylmethionine, the main methyl donor. Two MAT-encoding genes (MAT1A, MAT2A) are found in mammals. The latter is expressed in proliferating liver, dedifferentiation and cancer, whereas MAT1A is expressed in adult quiescent hepatocytes. Here, we report studies on the molecular mechanisms controlling the induction of MAT2A in regenerating rat liver and in proliferating hepatocytes. The MAT2A is up-regulated at two discrete moments during liver regeneration, as confirmed by RNApol-ChIP analysis. The first one coincides with hepatocyte priming (i.e. G0-G1 transition), while the second one takes place at the G1-S interface. Electrophoretic mobility shift assays showed that a putative E2F sequence present in MAT2A promoter binds this factor and ChIP assays confirmed that E2F1, E2F3 and E2F4, as well as the pocket protein p130, are bound to the promoter in quiescent liver. MAT2A activation is accompanied by changes in the binding of histone-modifying enzymes to the promoter. Interestingly, p130 is not displaced from MAT2A promoter during hepatocyte priming, but it is in the late expression of the gene at the G1-S transition. Finally, the transcription factor Sp1 seems to play a decisive role in MAT2A induction, as it binds the promoter when the gene is being actively transcribed. In summary, the present work shows that the molecular mechanism of MAT2A expression is different during G0-G1 or G1-S transition and this may be related to the distinct requirements of S-adenosylmethionine during liver regeneration.

Acute pancreatitis is an inflammation initially localized in the pancreatic gland which may lead to local and systemic complications. The development of severe acute pancreatitis is mediated by pathophysiological mechanisms involved in the systemic inflammatory response, cytokines and oxidative stress being their components of major importance. Nevertheless, it is still unknown why an episode of acute pancreatitis remains mild or progresses to a severe form. Activated leukocytes are the main source of cytokines. Interleukin 1beta and tumor necrosis factor alpha (TNF-alpha) initiate and propagate almost all the consequences of the systemic inflammatory response syndrome, leading to amplification of the inflammatory response. It is noteworthy that the systemic inflammatory response is restrained and the rate of mortality decreased in acute pancreatitis when TNF-alpha is blocked with specific antibodies or in knock-out mice deficient in its receptors. A synergy between pro-inflammatory cytokines and oxidative stress occurs in the development of the inflammatory response in acute pancreatitis. Pro-inflammatory cytokines and oxidative stress trigger common signal transduction pathways that lead to amplification of the inflammatory cascade, mainly through activation of mitogen-activated protein kinases (MAPK) and nuclear factor kappaB (NF-kappaB). Furthermore, pro-inflammatory cytokines, particularly TNF-alpha, and oxidative stress promote each other generating a vicious circle in acute pancreatitis. This cross-talk that arises between pro-inflammatory cytokines and oxidative stress greatly contributes to amplification of the uncontrolled inflammatory cascade through MAPK and NF-kappaB.

The Id (inhibitor of DNA binding or inhibitor of differentiation) helix-loop-helix proteins are involved in the regulation of cell growth, differentiation and cancer. The fact that the molecular mechanisms of liver regeneration are not completely understood prompted us to study the fate of Id2 in proliferating liver. Id2 increases in liver regeneration after partial hepatectomy, following the early induction of its gene. Co-immunoprecipitation shows that Id2 forms a complex with E2F4, p130 and mSin3A in quiescent liver and all these components are present at the c-myc promoter as shown using ChIP (chromatin immunoprecipitation). Activation of c-myc during hepatocyte priming (G0-G1 transition) correlates with the dissociation of Id2 and HDAC (histone deacetylase), albeit p130 remains bound at least until 6 h. Moreover, as the G0-G1 transition progresses, Id2 and HDAC again bind the c-myc promoter concomitantly with the repression of this gene. The time course of c-myc binding to the Id2 promoter, as determined by ChIP assays is compatible with a role of the oncoprotein as a transcriptional inducer of Id2 in liver regeneration. Immunohistochemical analysis shows that Id2 also increases in proliferating hepatocytes after bile duct ligation. In this case, the pattern of Id2 presence in the c-myc promoter parallels that found in regenerating liver. Our results may suggest a control role for Id2 in hepatocyte priming, through a p130 dissociation-independent regulation of c-myc.

The dynamic state of histone acetylation is maintained by histone acetyltransferases (HATs) and deacetylases. Cellular fractionation of plasmodia of Physarum polycephalum and partial purification of subcellular fractions by chromatography revealed the existence of a cytoplasmic B-type and four nuclear A-type HATs. The cytoplasmic B-enzyme was highly specific for histone H4, causing di-acetylation of H4 in vitro. The nuclear enzymes (HAT-A1 to HAT-A4) accepted all core histones as substrates, but differed by the preference for certain histone species. Enzymes were analyzed during the naturally synchronous cell cycle of macroplasmodia. Each of the enzymes had its individual cell cycle activity pattern, indicating diverse functions in nuclear metabolism. When growing plasmodia were induced to undergo differentiation into dormant sclerotia, an additional enzyme (HAT-AS) appeared at a late stage of sclerotization which correlated with differentiation-specific histone synthesis and acetylation in the absence of DNA replication. When dormant sclerotia were induced to reenter the cell cycle, a further enzyme form (HAT-AG) appeared during a short time period prior to the first post-germination mitosis. This enzyme had a strong preference for H2B, correlating with the overproportional in vivo acetate incorporation in H2B. Both differentiation-associated HATs were undetectable in growing plasmodia. The results demonstrate that different functions of core histone acetylation are based on multiple enzyme forms that are independently regulated during the cell cycle. Transitions from one developmental stage into another are accompanied by specific enzyme forms. With respect to recent data in the literature it may be assumed that these HAT-forms are subunits of a HAT-complex whose composition changes during the cell cycle and differentiation.