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Introduction. One of the challenges in the management of patients with cancer of the digestive tract is to prevent relapse. Epigenetic modifications (i.e. DNA methylation and histone marks) have been proven to be involved in the generation and maintenance of a subset of cells with stemness properties, that resist to conventional therapies. Our aim is to identify complexes of chromatin modifier enzymes (epienzymes) that are involved in acquired chemoresistance/aggressiveness of colon and pancreatic cancer cells. Experimental procedures/Methods. A meta-analysis performed on public transcriptomics data from colorectal cancer (CRC) patients cross-referenced with our own data including basal-like pancreatic cancer samples identified a limited number of epienzymes associated with aggressive subtypes of digestive cancers. Among them, the demethylase KDM4B, specific of the lysine 9 of Histone 3, and the transcriptional repressor PRDM1 where found correlated to the expression of several typical markers of cancer stemness. To confirm the role of these epienzymes in colon and pancreatic cancer cell chemoresistance, we have developed several in vivo and in vitro 2D and 3D models of acquired resistance to oxaliplatin-based combined chemotherapies, including FOLFIRINOX and FOLFOX. In this work, these models as well as patient samples have been used to study (i) the dynamic expression profile of these epienzymes (RT-qPCR, immunofluorescent multiplex staining, single-cell analysis and spatial transcriptomics), (ii) the gene regulons that are bound by these epienzymes (CUT&RUN-seq) and (iii) the functional role of these epienzymes in cancer stemness (organoid and sphere-forming assays). Results. We have shown that PRDM1 and KDM4B are expressed heterogeneously in cancer cell populations and overexpressed during specific time frames along the acquisition of resistance to oxaliplatin-based combined therapies both in colon and pancreatic cancer cells. Furthermore, we have obtained promising results regarding the binding of PRDM1 and KDM4B to key players of pluripotency/differentiation in chemoresistant colon cancer cells. Conclusions. Altogether, our data indicate that specific expression of chromatin modifier enzymes, most likely acting as epigenetic complexes, are key milestones of acquired resistance to conventional therapies. This work will pave the way for considering epienzyme-epienzyme interactions as potent drug targets that could circumvent cancer cell aggressiveness. Citation Format: Elsa Hadj Bachir, Charles Poiraud, Hugo Claux, Soumaya El Moghrabi, Sonia Paget, Belinda Duchêne, Nicolas Jonckheere, Bernadette Neve, Emmanuelle Leteurtre, Isabelle Van Seuningen, Audrey Vincent. Epigenetic mechanisms involved in acquired resistance to combined chemotherapies in digestive cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2965.

International audience; Twenty mucin genes have been identified and classified in two groups (encoding secreted and membrane-bound proteins). Secreted mucins participate in mucus formation by assembling a 3-dimensional network via oligomerization, whereas membrane-bound mucins are anchored to the outer membrane mediating extracellular interactions and cell signaling. Both groups have been associated with carcinogenesis progression in epithelial cancers, and are therefore considered as potential therapeutic targets. In the present review, we discuss the link between mucin expression patterns and patient survival and propose mucins as prognosis biomarkers of epithelial cancers (esophagus, gastric, pancreatic, colorectal, lung, breast or ovarian cancers). We also investigate the relationship between mucin expression and overall survival in the TCGA dataset. In particular, epigenetic mechanisms regulating mucin gene expression, such as aberrant DNA methylation and histone modification, are interesting as they are also associated with diagnosis or prognosis significance. Indeed, mucin hypomethylation has been shown to be associated with carcinogenesis progression and was linked to prognosis in colon cancer or pancreatic cancer patients. Finally we describe the relationship between mucin expression and non-coding RNAs that also may serve as biomarkers. Altogether the concomitant knowledge of specific mucin-pattern expression and epigenetic regulation could be translated as biomarkers with a better specificity/sensitivity performance in several epithelial cancers.

A major challenge in the treatment of cancer is dealing with intrinsic and/or acquired chemoresistance and with the development of metastatic lesions. The underlying mechanisms of chemoresistance are complex as a consequence of cancer heterogeneity, such as different tumour tissue origin, inter-tumour heterogeneity between patients and intra-tumour heterogeneity within cell populations of tumours. Classifying tumours according to gene expression-based molecular subtypes predicts the response to therapy and has been proposed to innovate towards personalized therapy. We here highlight the molecular subtypes observed in gastrointestinal cancers. In addition, we discuss the implication of long non-coding RNAs (lncRNAs), a class of RNAs without protein-coding sequence and over 200 nucleotides long, in chemoresistance of gastrointestinal cancers.

International audience; The recent discovery of cancer cell plasticity, i.e. their ability to reprogram into cancer stem cells (CSCs) either naturally or under chemotherapy and/or radiotherapy, has changed, once again, the way we consider cancer treatment. If cancer stemness is a reversible epigenetic state rather than a genetic identity, opportunities will arise for therapeutic strategies that remodel epigenetic landscapes of CSCs. However, the systematic use of DNA methyltransferase and histone deacetylase inhibitors, alone or in combination, in advanced solid tumors including colorectal cancers, regardless of their molecular subtypes, does not seem to be the best strategy. In this review, we first summarize the knowledge researchers have gathered on the epigenetic signatures of CSCs with the difficulty of isolating rare populations of cells. We raise questions about the relevant use of currently available epigenetic inhibitors (epidrugs) while the expression of numerous cancer stem cell markers are often repressed by epigenetic mechanisms. These markers include the three cluster of differentiation CD133, CD44 and CD166 that have been extensively used for the isolation of colon CSCs.and . Finally, we describe current treatment strategies using epidrugs, and we hypothesize that, using correlation tools comparing associations of relevant CSC markers with chromatin modifier expression, we could identify better candidates for epienzyme targeting.

Gene polymorphisms of TCF7L2 are associated with increased risk of type 2 diabetes and transcription factor 7-like 2 (TCF7L2) plays a role in hepatic glucose metabolism. We therefore addressed the impact of TCF7L2 isoforms on hepatocyte nuclear factor 4α (HNF4α) and the regulation of gluconeogenesis genes.Liver TCF7L2 transcripts were analysed by quantitative PCR in 33 non-diabetic and 31 type 2 diabetic obese individuals genotyped for TCF7L2 rs7903146. To analyse transcriptional regulation by TCF7L2, small interfering RNA transfection, luciferase reporter and co-immunoprecipitation assays were performed in human hepatoma HepG2 cells.In livers of diabetic compared with normoglycaemic individuals, five C-terminal TCF7L2 transcripts showed increased expression. The type 2 diabetes risk allele of rs7903146 positively correlated with TCF7L2 expression in livers from normoglycaemic individuals only. In HepG2 cells, transcript and TCF7L2 protein levels were increased upon incubation in high glucose and insulin. Of the exon 13 transcripts, six were increased in a glucose dose-responsive manner. TCF7L2 transcriptionally regulated 29 genes related to glucose metabolism, including glucose-6-phosphatase. In cultured HepG2 cells, TCF7L2 did not regulate HNF4Α and FOXO1 transcription, but did affect HNF4α protein expression. The TCF7L2 isoforms T6 and T8 (without exon 13 and with exon 15/14, respectively) specifically interacted with HNF4α.The different levels of expression of alternative C-terminal TCF7L2 transcripts in HepG2 cells, in livers of normoglycaemic individuals carrying the rs7901346 type 2 diabetes risk allele and in livers of diabetic individuals suggest that these transcripts play a role in the pathophysiology of type 2 diabetes. We also report for the first time a protein interaction in HepG2 cells between HNF4α and the T6 and T8 isoforms of TCF7L2, which suggests a distinct role for these specific alternative transcripts.

Aims/hypothesis Transcription factor 7-like 2 (TCF7L2) is a Wnt-signalling-associated transcription factor. Genetic studies have clearly demonstrated that DNA polymorphisms within TCF7L2 confer the strongest known association with increased risk of type 2 diabetes. However, the impact of the TCF7L2 type-2-diabetes-associated rs7903146 T allele on biological function and morphology of human pancreatic islets is unknown.

Kruppel-like transcription factors (KLFs) have elicited significant attention because of their regulation of essential biochemical pathways and, more recently, because of their fundamental role in the mechanisms of human diseases. Neonatal diabetes mellitus is a monogenic disorder with primary alterations in insulin secretion. We here describe a key biochemical mechanism that underlies neonatal diabetes mellitus insulin biosynthesis impairment, namely a homozygous mutation within the insulin gene (INS) promoter, c.-331C>G, which affects a novel KLF-binding site. The combination of careful expression profiling, electromobility shift assays, reporter experiments, and chromatin immunoprecipitation demonstrates that, among 16 different KLF proteins tested, KLF11 is the most reliable activator of this site. Congruently, the c.-331C>G INS mutation fails to bind KLF11, thus inhibiting activation by this transcription factor. Klf11(-/-) mice recapitulate the disruption in insulin production and blood levels observed in patients. Thus, these data demonstrate an important role for KLF11 in the regulation of INS transcription via the novel c.-331 KLF site. Lastly, our screening data raised the possibility that other members of the KLF family may also regulate this promoter under distinct, yet unidentified, cellular contexts. Collectively, this study underscores a key role for KLF proteins in biochemical mechanisms of human diseases, in particular, early infancy onset diabetes mellitus.

Context: The transcription factor Krüppel-like zinc finger 11 (KLF11) has been suggested to contribute to genetic risk of type 2 diabetes (T2D). Our previous results showed that four KLF11 variants, in strong linkage disequilibrium (LD block including +185 A>G/Gln62Arg and −1659 G>C) were associated with T2D in a north European case-control study. Here we further analyzed these variants for T2D association in a general Danish population and assess their possible effect on gene function. Methods: We genotyped Gln62Arg variant, representative for the LD block, in 5864 subjects of the INTER99 study to assess association to T2D and glucose metabolism-related quantitative traits. We studied effects of LD-block variants on KLF11 function and in particular, the effect of −1659G>C on transcriptional regulation of KLF11 using EMSA, chromatin immunoprecipitation, gene reporter assays, and small interfering RNA transfection. Results: We could not confirm T2D association of the KLF11 LD block, however, in glucose-tolerant subjects; it was significantly associated with higher fasting serum insulin and C-peptide levels and increased homeostasis model assessment insulin resistance indexes (P = 0.00004, P = 0.006, and P = 0.00002, respectively). In addition, binding of signal transducer and activator of transcription (STAT)-3 to the wild-type (−1659G>C) allele stimulated gene transcription, whereas STAT3 did not bind onto the mutant allele. Conclusions: We showed that KLF11 may interfere with glucose homeostasis in a Danish general population and that STAT3-mediated up-regulation of KLF11 transcription was impaired by the −1659G>C variant. Overall, KLF11 variants may have a deleterious effect on insulin sensitivity, although that may not be sufficient to lead to T2D.

Individual genotyping of single nucleotide polymorphisms (SNPs) remains expensive, especially for linkage disequilibrium mapping strategies involving high-throughput SNP genotyping. On one hand, current methods may suit scientific and laboratory needs in regard to accuracy, reproducibility/robustness, and large-scale application. On the other hand, a cheaper and less time-consuming alternative to individual genotyping is the use of SNP allele frequencies determined in DNA pools. We have developed an accurate and reproducible protocol for allele frequency determination using PyrosequencingTM technology in large genomic DNA pools (374 individuals). The measured correlation (R2) in large DNA pools was 0.980. In the context of disease-associated SNPs studies, we compared the allele frequencies between the disease (e.g., type 2 diabetes and obesity) and control groups detected by either individual genotyping or Pyrosequencing of DNA pools. In large pools, the variation between the two methods was 1.5 ± 0.9%. It may be concluded that the allele frequency determination protocol could reliably detect over 4% differences between populations. The method is economical in regard to amounts of DNA, PCR, and primer extension reagents required. Furthermore, it allows the rapid determination of allele frequency differences in case/control groups for association studies and susceptibility gene discovery in complex diseases.

The "mechanistic target of rapamycin" (mTOR) is a central controller of growth, proliferation and/or motility of various cell-types ranging from adipocytes to immune cells, thereby linking metabolism and immunity. mTOR signaling is overactivated in obesity, promoting inflammation and insulin resistance. Therefore, great interest exists in the development of mTOR inhibitors as therapeutic drugs for obesity or diabetes. However, despite a plethora of studies characterizing the metabolic consequences of mTOR inhibition in rodent models, its impact on immune changes associated with the obese condition has never been questioned so far. To address this, we used a mouse model of high-fat diet (HFD)-fed mice with and without pharmacologic mTOR inhibition by rapamycin. Rapamycin was weekly administrated to HFD-fed C57BL/6 mice for 22 weeks. Metabolic effects were determined by glucose and insulin tolerance tests and by indirect calorimetry measures of energy expenditure. Inflammatory response and immune cell populations were characterized in blood, adipose tissue and liver. In parallel, the activities of both mTOR complexes (e. g. mTORC1 and mTORC2) were determined in adipose tissue, muscle and liver. We show that rapamycin-treated mice are leaner, have enhanced energy expenditure and are protected against insulin resistance. These beneficial metabolic effects of rapamycin were associated to significant changes of the inflammatory profiles of both adipose tissue and liver. Importantly, immune cells with regulatory functions such as regulatory T-cells (Tregs) and myeloid-derived suppressor cells (MDSCs) were increased in adipose tissue. These rapamycin-triggered metabolic and immune effects resulted from mTORC1 inhibition whilst mTORC2 activity was intact. Taken together, our results reinforce the notion that controlling immune regulatory cells in metabolic tissues is crucial to maintain a proper metabolic status and, more generally, comfort the need to search for novel pharmacological inhibitors of the mTOR signaling pathway to prevent and/or treat metabolic diseases.

Peroxisome proliferator-activated receptors (PPARs) are nuclear receptors that regulate lipid and glucose metabolism and cellular differentiation. PPAR-alpha and PPAR-gamma are both expressed in human macrophages where they exert anti-inflammatory effects. The activation of PPAR-alpha may promote foam-cell formation by inducing expression of the macrophage scavenger receptor CD36. This prompted us to investigate the influence of different PPAR-activators on cholesterol metabolism and foam-cell formation of human primary and THP-1 macrophages. Here we show that PPAR-alpha and PPAR-gamma activators do not influence acetylated low density lipoprotein-induced foam-cell formation of human macrophages. In contrast, PPAR-alpha and PPAR-gamma activators induce the expression of the gene encoding ABCA1, a transporter that controls apoAI-mediated cholesterol efflux from macrophages. These effects are likely due to enhanced expression of liver-x-receptor alpha, an oxysterol-activated nuclear receptor which induces ABCA1-promoter transcription. Moreover, PPAR-alpha and PPAR-gamma activators increase apoAI-induced cholesterol efflux from normal macrophages. In contrast, PPAR-alpha or PPAR-gamma activation does not influence cholesterol efflux from macrophages isolated from patients with Tangier disease, which is due to a genetic defect in ABCA1. Here we identify a regulatory role for PPAR-alpha and PPAR-gamma in the first steps of the reverse-cholesterol-transport pathway through the activation of ABCA1-mediated cholesterol efflux in human macrophages.

Hepatic lipase (HL) is anN-glycoprotein that acquires triglyceridase activity somewhere during maturation and secretion. To determine where and how HL becomes activated, the effect of drugs that interfere with maturation and intracellular transport of HL protein was studied using freshly isolated rat hepatocytes. Carbonyl cyanidem-chlorophenyl hydrazone (CCCP), castanospermine, monensin, and colchicin all inhibited secretion of HL without affecting its specific enzyme activity. The specific enzyme activity of intracellular HL was decreased by 25–50% upon incubation with CCCP or castanospermine, and increased 2-fold with monensin and colchicin. Glucose trimming of HL protein was not affected by CCCP, as indicated by digestion of immunoprecipitates with jack bean α-mannosidase. Pulse labeling experiments with [35S]methionine indicated that conversion of the 53-kDa precursor to the 58-kDa form, nor the development of endoglycosidase H-resistance, were essential for acquisition of enzyme activity. In sucrose gradients, HL protein from secretion media sedimented as a homogeneous band of about 5.8 S, whereas HL protein from the cell lysates migrated as a broad band extending from 5.8 S to more than 8 S. With both sources, HL activity was exclusively associated with the 5.8 S HL protein form. We conclude that glucose trimming of HL protein in the endoplasmic reticulum is not sufficient for activation; full activation occurs during or after transport from the endoplasmic reticulum to the Golgi and is associated with a decrease in sedimentation velocity.

Human hepatic lipase (HL) is a glycoprotein with four N-linked oligosaccharide side chains. The importance of glycosylation for the secretion of catalytically active HL was studied in HepG2 cells by using inhibitors of intracellular trafficking, N-glycosylation and oligosaccharide processing. Secretion of HL was inhibited by carbonyl cyanide m-chlorophenylhydrazone (CCCP), monensin, brefeldin A (BFA), tunicamycin, castanospermine and N-methyldeoxynojirimycin, but not by 1-deoxymannojirimycin. Secretion of α1-antitrypsin, an unrelated N-glycoprotein, was also inhibited by monensin, BFA and tunicamycin, but not by CCCP, castanospermine or N-methyldeoxynojirimycin. Intracellular HL activity decreased with CCCP, tunicamycin, castanospermine and N-methyldeoxynojirimycin, but increased with monensin and BFA. In the absence of protein synthesis de novo, HL activity secreted into the medium was 7.8±2.1-fold higher (mean±S.D., n = 7) than the simultaneous fall in intracellular HL activity. In cells pretreated with monensin or BFA, this factor decreased to 1.3±0.5, indicating that the apparent increase in HL activity had already occurred within these cells. After chromatography on Sepharose–heparin, the specific triacylglycerol hydrolase activity of secreted HL was only 1.7±0.3-fold higher than that of intracellular HL, indicating that the secretion-coupled increase in HL activity is only partly explained by true activation. We conclude that oligosaccharide processing by glucosidases in the endoplasmic reticulum is necessary for the transport of newly synthesized human HL, but not α1-antitrypsin, to the Golgi, where the catalytic activity of HL is unmasked.

In rats, the daily changes in hepatic lipase (HL) activity in the liver follow the diurnal rhythm of the catecholamines. To study the underlying mechanism, the effect of adrenaline on maturation and secretion of HL was determined in freshly isolated rat hepatocytes. Adrenaline (10 microM) acutely inhibited the secretion of HL. This effect was abolished by 0.1 microM prazosin, but not by 1 microM propranolol, indicating the involvement of the alpha1-adrenergic pathway. Prazosin was at least 1000-fold more potent than WB4101, a selective alpha1A-antagonist. Adrenaline had no effect on HL secretion in hepatocytes pretreated with chloroethylclonidine, an irreversible alpha1B-selective antagonist. Inhibition of HL was not induced by 10 microM methoxamine, a alpha1A-selective agonist. Thus, adrenaline inhibited HL secretion through activation of the alpha1-adrenoceptors subtype B, which have been shown to signal through Ca2+ as well as cAMP. A similar reduction in HL secretion was induced by the Ca2+-mobilizing hormones angiotensin II (100 nM) and vasopressin (12 nM), the Ca2+ ionophore A23187 (2 microM), and by thapsigargin (1 microM), which inhibits the ER Ca2+-ATPase pump. HL secretion was unaffected by elevating cAMP with 10 microM forskolin or 1 microM 8-Br-cAMP. These results suggest that the alpha1B-adrenergic effects on HL expression are mainly mediated through elevation of intracellular Ca2+. Chelation of extracellular Ca2+ and subsequent lowering of intracellular Ca2+ with EGTA also inhibited HL secretion. In pulse-chase experiments, adrenaline was shown to inhibit the maturation of HL from the 53 kDa, Endo H-sensitive precursor to the Endo H-resistant, catalytically active protein of 58 kDa. In addition, adrenaline induced intracellular degradation of newly synthesized HL. Similar post-translational effects, both qualitatively and quantitatively, were observed with A23187, thapsigargin and EGTA. We conclude that the inhibition of HL maturation and increase in intracellular degradation induced by catecholamines, A23187, thapsigargin and EGTA is evoked by changes in Ca2+ homoeostasis, possibly through lowering ER Ca2+.

Objective-Dyslipidemia contributes to endothelial dysfunction in type 2 diabetes mellitus. Fenofibrate (FF), a ligand of the peroxisome proliferator-activated receptor-α (PPARα), has beneficial effects on microvascular complications. FF may act on the endothelium by regulating vasoactive factors, including endothelin-1 (ET-1). In vitro, FF decreases ET-1 expression in human microvascular endothelial cells. We investigated the molecular mechanisms involved in the effect of FF treatment on plasma levels of ET-1 in type 2 diabetes mellitus patients. Methods and Results-FF impaired the capacity of transforming growth factor-β to induce ET-1 gene expression. PPARα activation by FF increased expression of the transcriptional repressor Krüppel-like factor 11 and its binding to the ET-1 gene promoter. Knockdown of Krüppel-like factor 11 expression potentiated basal and transforming growth factor-β-stimulated ET-1 expression, suggesting that Krüppel-like factor 11 downregulates ET-1 expression. FF, in a PPARα-independent manner, and insulin enhanced glycogen synthase kinase-3β phosphorylation thus reducing glycogen synthase kinase-3 activity that contributes to the FF-mediated reduction of ET-1 gene expression. In type 2 diabetes mellitus, improvement of flow-mediated dilatation of the brachial artery by FF was associated with a decrease in plasma ET-1. Conclusion-FF decreases ET-1 expression by a PPARα-dependent mechanism, via transcriptional induction of the Krüppel-like factor 11 repressor and by PPARα-independent actions via inhibition of glycogen synthase kinase-3 activity. © 2013 American Heart Association, Inc., Région Nord-Pas de Calais/FEDER; Fondation Coeur et Artères; National Health and Medical Research Council (NHMRC) of Australia; NHMRC medical scholarship; French National Research Agency (ANR) Institut National de la Santé et de la Recherche Médicale; Centre National de la Recherche Scientifique; Institut Universitaire de France

Type 2 diabetes manifests when the β-cell fails to secrete sufficient amounts of insulin to maintain normoglycemia and undergoes apoptosis. The disease progression results from an interplay of environmental factors and genetic predisposition. Polymorphisms in T-cell factor 7-like 2 (TCF7L2) strongly correlate with type 2 diabetes mellitus (T2DM). While TCF7L2 mRNA is upregulated in islets in diabetes, protein levels are downregulated. The loss of TCF7L2 induces impaired function and apoptosis. By analyzing human isolated islets, we provide three explanations for this opposite regulation and the mechanisms of TCF7L2 on β-cell function and survival. (i) We found TCF7L2 transcripts in the human β-cell, which had opposite effects on β-cell survival, function and Wnt signaling activation. While TCF7L2 clone B1, which lacks exons 13, 14, 15 and 16 induced β-cell apoptosis, impaired function and inhibited glucagon-like peptide 1 response and downstream targets of Wnt signaling, clones B3 and B7 which both contain exon 13, improved β-cell survival and function and activated Wnt signaling. (ii) TCF7L2 mRNA is extremely unstable and is rapidly degraded under pro-diabetic conditions and (iii) TCF7L2 depletion in islets induced activation of glycogen synthase kinase 3-β, but this was independent of endoplasmic reticulum stress. We demonstrated function-specific transcripts of TCF7L2, which possessed distinct physiological and pathophysiological effects on the β-cell. The presence of deleterious TCF7L2 splice variants may be a mechanism of β-cell failure in T2DM.

Pdx-1 (pancreatic-duodenal homeobox-1), a MODY4 homeodomain transcription factor, serves as a master regulator in the pancreas because of its importance during organogenesis and in adult islet insulin-producing beta cell activity. Here, we show that KLF11, an SP/Kruppel-like (SP/KLF) transcription factor, mutated in French maturity onset diabetes of the young patients (MODY7), regulates Pdx-1 transcription in beta cells through two evolutionarily conserved GC-rich motifs in conserved Area II, a control region essential to islet beta cell-enriched expression. These regulatory elements, termed GC1 (human base pair -2061/-2055) and GC2 (-2036/-2027), are also nearly identical to the consensus KLF11 binding sequence defined here by random oligonucleotide binding analysis. KLF11 specifically associates with Area II in chromatin immunoprecipitation assays, while preventing binding to GC1- and/or GC2-compromised Pdx1-driven reporter activity in beta cell lines. Mechanistically, we find that KLF11 interacts with the coactivator p300 via its zinc finger domain in vivo to mediate Pdx-1 activation. Together, our data identified a hierarchical regulatory cascade for these two MODY genes, suggesting that gene regulation in MODY is more complex than anticipated previously. Furthermore, because KLF11 like most MODY-associated transcription factors uses p300, these data further support a role for this coactivator as a critical chromatin link in forms of type 2 diabetes.

Background Transgenic mice over-expressing SMAD7 in pancreatic beta-cells develop type 2 diabetes (T2D). The expression of SMAD7 is affected by KLF11, which contains gene variants that have previously been shown to be involved in genetic susceptibility to T2D, and by the highly homologous KLF10. This study aims to assess the genetic contribution of SMAD7 and KLF10 gene variants to T2D susceptibility in the French population. Methods We screened both genes to identify rare and frequent variants by direct sequencing and then genotyped these variants. Six frequent variants of SMAD7 and six of KLF10 were analyzed in 349 T2D patients and 349 normoglycaemic adult subjects. Variants with statistically significant differences in allele and/or genotype distribution were further analyzed in a population sample of 1.712 T2D patients and 1.072 normoglycaemic subjects. Results Two variants showed a significant association under a recessive model: The intronic SMAD7 IVS2 –21 had an odds ratio of 0.62 ( P = 0.007, 95% CI = 0.44–0.88; P = 0.034 when adjusting for age, sex and BMI by logistic regression), and the KLF10 3′UTR +1002 variant had an Odds Ratio of 0.81 ( P = 0.009, 95% CI = 0.69–0.95; P = 0.042 when adjusting for age, sex and BMI). Conclusion Although the observed association of SMAD7 and KLF10 gene variants with T2D is modest, they may weakly contribute to a particular genetic background that increases the susceptibility to development of T2D.

Introduction Des mutations du gene KCNJ11 sont responsables de differentes formes de diabete precoce. La mutation KCNJ11-E227K est associee a la fois au MODY et au diabete neonatal dans differentes familles, suggerant que le fonds genetique familial contribuerait a moduler le phenotype. Afin de mieux caracteriser les mecanismes induits par cette mutation, nous utilisons les nouvelles techniques de modification ciblee du genome. Materiels et methodes Nous avons derive les cellules-souches pluripotentes induites (iPSC) a partir des cellules mononucleees du sang d’individus porteurs des mutations des genes impliques dans le diabete monogenique. Grâce a la technique des TALE Nucleases et/ou Crispr/Cas9, nous introduisons des cassures double-brin dans l’ADN et la reparation faite par recombinaison homologue afin de realiser la modification nucleotidique specifique de la mutation KCNJ11-E227K. Nous etudierons les iPSC en presence ou non de la mutation E227K pendant leur differenciation en cellules beta-pancreatiques, et l’impact fonctionnel sur leur capacite a secreter l’insuline. Resultats Nous presentons notre strategie de modification ciblee dans une lignee cellulaire qui exprime le gene KCNJ11. Nous avons analyse trois ARNguide de Cas9, et grâce au clonage TOPO-TA, nous avons mis en evidence plusieurs evenements de la reparation de l’ADN par « jonction d’extre-mites non homologues » accompagne de la perte, du gain ou de l’inversion de nucleotides. Ce type de modification n’a pas ete observe aux loci non-specifiques. Pour stimuler la « recombinaison homologue », nous realisons des clonages de donneurs d’ADN avec des fragments d’ADN synthetise. Cette strategie pourra etre facilement adaptee a d’autres regions de genes cibles. Nous utilisons deux etapes de la recombinaison homologue consecutives pour realiser une correction nucleotidique specifique sans alterer l’integrite de l’ADN genomique environnant. Conclusion Les techniques de modification ciblee du genome sont performantes et permettent l’analyse d’une mutation specifique (KCNJ11-E227K) sur un fonds genetique familial lie a la severite du phenotype. Declaration d’interet Les auteurs declarent ne pas avoir d’interet direct ou indirect (financier ou en nature) avec un organisme prive, industriel ou commercial en relation avec le sujet presente.

KLF11 (TIEG2) is a pancreas-enriched transcription factor that has elicited significant attention because of its role as negative regulator of exocrine cell growth in vitro and in vivo . However, its functional role in the endocrine pancreas remains to be established. Here, we report, for the first time, to our knowledge, the characterization of KLF11 as a glucose-inducible regulator of the insulin gene. A combination of random oligonucleotide binding, EMSA, luciferase reporter, and chromatin immunoprecipitation assays shows that KLF11 binds to the insulin promoter and regulates its activity in beta cells. Genetic analysis of the KLF11 gene revealed two rare variants (Ala347Ser and Thr220Met) that segregate with diabetes in families with early-onset type 2 diabetes, and significantly impair its transcriptional activity. In addition, analysis of 1,696 type 2 diabetes mellitus and 1,776 normoglycemic subjects show a frequent polymorphic Gln62Arg variant that significantly associates with type 2 diabetes mellitus in North European populations (OR = 1.29, P = 0.00033). Moreover, this variant alters the corepressor mSin3A-binding activity of KLF11, impairs the activation of the insulin promoter and shows lower levels of insulin expression in pancreatic beta cells. In addition, subjects carrying the Gln62Arg allele show decreased plasma insulin after an oral glucose challenge. Interestingly, all three nonsynonymous KLF11 variants show increased repression of the catalase 1 promoter, suggesting a role in free radical clearance that may render beta cells more sensitive to oxidative stress. Thus, both functional and genetic analyses reveal that KLF11 plays a role in the regulation of pancreatic beta cell physiology, and its variants may contribute to the development of diabetes.

The gene GAD2 encoding the glutamic acid decarboxylase enzyme (GAD65) is a positional candidate gene for obesity on Chromosome 10p11-12, a susceptibility locus for morbid obesity in four independent ethnic populations. GAD65 catalyzes the formation of gamma-aminobutyric acid (GABA), which interacts with neuropeptide Y in the paraventricular nucleus to contribute to stimulate food intake. A case-control study (575 morbidly obese and 646 control subjects) analyzing GAD2 variants identified both a protective haplotype, including the most frequent alleles of single nucleotide polymorphisms (SNPs) +61450 C>A and +83897 T>A (OR = 0.81, 95% CI [0.681-0.972], p = 0.0049) and an at-risk SNP (-243 A>G) for morbid obesity (OR = 1.3, 95% CI [1.053-1.585], p = 0.014). Furthermore, familial-based analyses confirmed the association with the obesity of SNP +61450 C>A and +83897 T>A haplotype (chi(2) = 7.637, p = 0.02). In the murine insulinoma cell line betaTC3, the G at-risk allele of SNP -243 A>G increased six times GAD2 promoter activity (p < 0.0001) and induced a 6-fold higher affinity for nuclear extracts. The -243 A>G SNP was associated with higher hunger scores (p = 0.007) and disinhibition scores (p = 0.028), as assessed by the Stunkard Three-Factor Eating Questionnaire. As GAD2 is highly expressed in pancreatic beta cells, we analyzed GAD65 antibody level as a marker of beta-cell activity and of insulin secretion. In the control group, -243 A>G, +61450 C>A, and +83897 T>A SNPs were associated with lower GAD65 autoantibody levels (p values of 0.003, 0.047, and 0.006, respectively). SNP +83897 T>A was associated with lower fasting insulin and insulin secretion, as assessed by the HOMA-B% homeostasis model of beta-cell function (p = 0.009 and 0.01, respectively). These data support the hypothesis of the orexigenic effect of GABA in humans and of a contribution of genes involved in GABA metabolism in the modulation of food intake and in the development of morbid obesity.

Introduction L’obesite est associee a un etat inflammatoire chronique de « bas grade » dans lequel le tissu adipeux blanc joue un role cle. En effet, les cellules immunes recrutees dans le tissu - principalement les macrophages - entretiennent le foyer inflammatoire, augmentant le risque de pathologies liees au surpoids, telle que l’insulinoresistance. La voie du mammalian target of rapamycin (mTOR) est sur-activee lors de l’obesite et est impliquee dans le developpement de la resistance a l’insuline et du diabete de type 2. La rapamycine, drogue immunomodulatrice inhibant selectivement la voie mTOR, represente donc une nouvelle piste therapeutique dans le traitement des maladies metaboliques. Notre objectif est de definir si les effets metaboliques de la rapamycine sont associes a des alterations immunes et/ou inflammatoires. Materiels et methodes Des souris mâles C57BL6/J sous regime standard ou hyperlipidique ont recus des injections hebdomadaires de rapamycine (2 mg/kg, i.p.) ou d’excipient. L’evolution du poids corporel, de la prise alimentaire ainsi que la tolerance au glucose in vivo ont ete determinees. Les hormones et cytokines seriques ont ete dosees par ELISA et Multiplex Arrays. La taille des adipocytes, le recrutement cellulaire et l’etat inflammatoire du tissu adipeux ont ete analyses par histologie, puces a ADN et PCR quantitative. Les cellules immunes du sang et de la rate ont ete caracterisees par cytometrie en flux. Resultats La rapamycine limite le gain de poids corporel tout en, paradoxalement, aggravant l’intolerance au glucose. Le recrutement cellulaire massif dans le tissu adipeux blanc (principalement les macrophages), ainsi que la production accrue de cytokines inflammatoires (notamment IL-6 et MCP-1) chez les souris traitees par la rapamycine pourraient expliquer l’effet de la drogue sur l’homeostasie glucidique. Conclusion Cette etude montre que la rapamycine regule le metabolisme energetique par un relai immun, impliquant notamment les macrophages du tissu adipeux blanc

Introduction La progression du diabete de type 2 resulte des interactions entre facteurs environnementaux et predisposition genetique. Les etudes genetiques ont demontre la tres forte association des polymorphismes de TCF7L2 avec le diabete de type 2. Alors que l’expression de TCF7L2 est augmentee au niveau de l’ARNm dans des modeles de diabete ou chez les porteurs de l’allele a risque, son expression proteique est diminuee. Enfin, la diminution de l’expression de TCF7L2 par siRNA reduit la secretion d’insuline et accroit l’apoptose d’ilots isoles. Cette apparente contradiction pourrait s’expliquer par un desequilibre de l’expression de certains transcrits regulant des fonctions cellulaires distinctes. L’objectif de cette etude etait donc d’identifier les isoformes de TCF7L2 exprimees dans la cellule-beta et d’analyser leur fonction biologique. Materiels et methodes Les differents transcrits de TCF7L2 ont ete amplifies par PCR a partir d’ARN de cellules-beta humaines triees par FACS, sequences puis clones. Les differents clones ont ensuite ete surexprimes dans des ilots pancreatiques humains isoles et leurs roles sur la secretion d’insuline induite par le glucose et la survie cellulaire analyses. Resultats 9 transcrits alternatifs de TCF7L2 exprimes dans la cellule-beta ont ete identifies. Certains de ces transcrits ont des effets contraires sur la survie cellulaire, la secretion d’insuline et l’expression de genes cibles : alors que le clone #1 induit l’apoptose des cellules-beta et diminue la secretion d’insuline induite par le GLP-1, les clones #3 et #7 stimulent la secretion d’insuline et la proliferation cellulaire. Conclusion Nous avons identifie plusieurs transcrits de TCF7L2 exprimes dans la cellule-beta humaine, capables de reguler des fonctions cellulaires distinctes. La perte proteique de TCF7L2 observee dans les modeles de diabete de type 2 pourrait representer un desequilibre dans l’expression de transcrits specifiques regulant des fonctions cellulaires distinctes.

Introduction Des variants genetiques du facteur de transcription TCF7L2 sont fortement associes a la susceptibilite au diabete de type 2 (DT2) et suscitent beaucoup d’interet. Recemment, TCF7L2 a ete implique dans le metabolisme hepatique du glucose. Notre objectif a ete de determiner si les transcrits alternatifs de TCF7L2 inhibent differemment la transcription de l’enzyme glucose-6- phosphatase (G6PC). Materiels et methodes Nous avons evalue l’expression hepatique de TCF7L2 par PCR-quantitative exon-specifique dans des echantillons humains (64 patients obeses normoglycemiques ou DT2 de la cohorte ABOS) et dans les cellules HepG2. L’impact de l’inhibition de TCF7L2 par ARNi sur l’expression de G6PC a ete evalue. Par la technique du gene rapporteur luciferase, nous avons analyse les interactions possibles entre la regulation transcriptionnelle de G6PC par TCF7L2, HNF4alpha et FOXO1. En realisant des co-immunoprecipitations, nous avons analyse les interactions proteiniques de TCF7L2 avec HNF4alpha dans les cellules HepG2. Resultats Dans les cellules HepG2, tous les transcrits de TCF7L2 contenant l’exon 13 sont induits par le glucose de maniere concentration dependante. L’expression de ces transcrits est aussi augmentee dans le foie d’individus diabetiques, sauf ceux ayant un C-terminal-court (T3 et T5). La transfection d’ARNi dans les cellules HepG2, a montre que l’expression de G6PC est reprimee par TCF7L2 independamment de la presence d’insuline et de l’activite de FOXO1. Nous avons pu montrer que l’inhibition de l’expression de HNF4alpha par TCF7L2 n’est pas indispensable pour la repression transcriptionelle de G6PC. Par contre, les transcrits T6 et T8, avec un C-terminal long mais sans exon 13, interagissent avec la proteine HNF4alpha dans les cellules HepG2. Conclusion Les transcrits T3 et T5 de TCF7L2 pourraient jouer un role different dans les anomalies du metabolisme hepatique lie au DT2 en comparaison aux transcrits T6 et T8 qui interagissent specifiquement avec la proteine HNF4alpha dans les cellules HepG2.

Background —Monocytic tissue factor (TF) expression may contribute to thrombogenicity associated with plaque rupture and may propagate thrombus formation at the site of vascular lesions. Induction of monocytic TF expression by endotoxin is mediated by the activation of transcription factors such as AP-1 and NF-κB. Both these signaling pathways are modulated by peroxisome proliferator–activated receptor-α (PPARα). Therefore, we have studied the effects of fibrates and other PPARα agonists on the expression of TF. Methods and Results —We show that PPARα protein, like primary human monocytes, is also expressed in the human monocytic THP-1 cell line. Fenofibric acid, WY14643, and GW2331 inhibited TF mRNA upregulation after stimulation of THP-1 cells with lipopolysaccharide or interleukin-1β. In primary human monocytes and macrophages, the lipopolysaccharide- or interleukin-1β–mediated induction of TF activity was also inhibited by fenofibric acid, WY14643, or GW2331. Conclusions —These data indicate that activation of PPARα results in the downregulation of the TF gene. Our results suggest a novel role for PPARα in the control of atherosclerotic plaque thrombogenicity through its effects on TF expression in monocytes and macrophages.

Peroxisome proliferator-activated receptors (PPAR) are ligand-activated transcription factors which form a subfamily of the nuclear receptor gene family. PPAR activators have effects on both metabolic risk factors and on vascular inflammation related to atherosclerosis. PPAR have profound effects on the metabolism of lipoproteins and fatty acids. PPAR alpha binds hypolipidemic fibrates, whereas PPAR gamma has a high affinity for antidiabetic glitazones. Both PPAR are activated by fatty acids and their derivatives. Activation of PPAR alpha increases the catabolism of fatty acids at several levels. In the liver, it increases uptake of fatty acids and activates their beta-oxidation. The effects that PPAR alpha exerts on triglyceride-rich lipoproteins is due to their stimulation of lipoprotein lipase and repression of apolipoprotein CIII expression, while the effects on high-density lipoproteins depend upon the regulation of apolipoproteins AI and AII. PPAR gamma has profound effects on the differentiation and function of adipose tissue, where it is highly expressed. PPAR are also expressed in atherosclerotic lesions. PPAR are present in vascular endothelial cells, smooth muscle cells, monocytes, and monocyte-derived macrophages. Via negative regulation of nuclear factor-kappa B and activator protein-1 signalling pathways, PPAR alpha inhibits expression of inflammatory genes, such as interleukin-6, cyclooxygenase-2, and endothelin-1. Furthermore, PPAR alpha inhibits expression of monocyte-recruiting proteins such as vascular cell adhesion molecule (VCAM)-1 and induces apoptosis in monocyte-derived macrophages. PPAR gamma activation in macrophages and foam cells inhibits the expression of activated genes such as inducible nitric oxide synthase, matrix metalloproteinase-9 and scavenger receptor A. PPAR gamma may also affect the recruitment of monocytes in atherosclerotic lesions as it is involved in the expression of VCAM-1 and intracellular adhesion molecule-1 in vascular endothelial cells. The involvement of PPAR in atherosclerosis, a disease with a chronic inflammatory character, suggests that they may play a role in other inflammatory-related diseases as well.

During hypothyroidism, hepatic lipase (HL) activity is decreased. The low HL may be due to thyroid hormone insufficiency or to the concomitant fall in growth hormone (GH) activity. We studied HL expression in hepatocytes freshly isolated from hypothyroid rats with and without additional GH-substitution. In all animals HL mRNA was detected by RT-PCR in the hepatocytes, but not in the non-parenchymal cells. In hypothyroid cells HL mRNA levels were reduced by 40%, and the in vitro secretion of HL-activity and HL-protein was decreased by about 50%. In cells from GH-substituted hypothyroid rats, HL mRNA level was normalised, but the secretion of HL remained low. The specific enzyme activity of secreted HL was similar under all conditions. The discrepancy between HL mRNA and HL secretion in GH-supplemented rats may be due to (post)translational effects. Therefore we studied the HL synthesis and maturation in hepatocytes from hypothyroid and GH-substituted rats. Pulse-labelling experiments with [(35)S]methionine showed that the incorporation of [(35)S]methionine into HL protein was lower both in hypothyroid cells and in GH-supplemented cells than in control cells. During the subsequent chase, the intracellular processing and transport of newly synthesized HL protein in the hepatocytes from hypothyroid rats, whether or not supplemented with GH, was similar to control cells. We conclude that in livers of hypothyroid, GH-substituted rats translation of HL mRNA is inhibited despite restoration of HL mRNA levels.

Catecholamines are responsible for the daily changes in hepatic lipase (HL) expression associated with feeding and fasting. We have studied the mechanism by which adrenaline decreases HL secretion in suspensions of freshly isolated rat hepatocytes. Adrenaline acutely inhibited HL activity through activation of the α 1 -adrenergic pathway. The cells had significantly less HL activity in the presence of adrenaline versus cycloheximide, where protein de novo synthesis is completely blocked. The specific enzyme activity of secreted HL was not affected. Intracellular HL activity was decreased by adrenaline treatment. Pulse-labeling with [ 35 S]methionine showed that de novo synthesis of the 53-kd endo-β-N-acetylglucosaminidase H (Endo H)—sensitive HL protein was unaffected by adrenaline. During subsequent chase of the control cells, the 53-kd form was converted to a 58-kd Endo H—resistant HL protein, which was rapidly secreted into the medium. In the presence of adrenaline, formation of the 58-kd protein was markedly reduced, whereas the 53-kd protein disappeared at a rate similar to the rate in controls. This suggests that part of the HL protein was degraded. In contrast to adrenaline, inhibition of HL secretion by colchicine was accompanied by an intracellular accumulation of HL activity and of the 58-kd protein. We conclude that adrenaline inhibits HL secretion posttranslationally by retarding the maturation of the 53-kd HL precursor to an active 58-kd protein, possibly by stimulating degradation of newly synthesized HL protein.

Objectif L’obesite est caracterisee par l’expansion excessive du tissu adipeux blanc associee a une inflammation chronique a « bas bruit » initiee par le recrutement de cellules immunes dans le tissu. L’adipogenese et la fonction des cellules immunes sont regules par la voie du « mechanistic Target Of Rapamycin » (mTOR). La voie mTOR est sur-activee lors de l’obesite, participant au developpement de la resistance a l’insuline et du diabete de type 2. Ainsi la rapamycine, inhibiteur selectif de la voie mTOR, est proposee comme une nouvelle piste therapeutique dans le traitement des maladies metaboliques. Notre objectif est de preciser les effets metaboliques de la rapamycine et de definir s’ils sont associes a des alterations immunes et/ou inflammatoires. Materiels et methodes Des souris mâles C57BL/6J mises sous regime hyperlipidique recoivent des injections hebdomadaires de rapamycine ou d’excipient. Les parametres metaboliques et immuns sont suivis par l’utilisation de techniques de biologie moleculaire (Q-PCR, transcriptomique), biochimie (ELISAs, western-blot), cytometrie en flux et histologie. Resultats Bien que limitant la prise de poids corporel, la rapamycine aggrave l’intolerance au glucose. Le recrutement massif de macrophages dans le tissu adipeux ainsi que la production accrue de cytokines inflammatoires (IL-6, MCP-1) chez les souris traitees pourraient expliquer l’effet deletere de la drogue sur l’homeostasie glucidique. Par ailleurs, l’augmentation du nombre des cellules immuno-regulatrices Gr1+CD11b+ (Myeloid-derived suppressor cells ou MDSC) dans le tissu adipeux, pourrait expliquer le maintien de la sensibilite a l’insuline des souris traitees ; les MDSCs ayant recemment ete decrites comme moderant l’inflammation et, consequemment, ameliorant la sensibilite a l’insuline de souris obeses. Conclusion Notre etude suggere que l’effet paradoxal de la rapamycine sur le metabolisme energetique pourrait resulter d’un controle exerce par la voie mTOR sur les cellules myeloides regulant l’inflammation du tissu adipeux blanc (macrophages et MDSCs).

Introduction Des variants genetiques de TCF7L2 ont ete fortement associes a la susceptibilite au diabete de type 2 (DT2). Nous avons montre que plusieurs transcrits alternatifs de TCF7L2 avaient des effets distincts sur la proliferation et la secretion d’insuline des cellules beta pancreatiques d’ilots humains isoles. Recemment, TCF7L2 a ete implique dans le metabolisme du glucose des cellules hepatiques et nous avons egalement observe une expression differentielle des transcrits TCF7L2 dans le foie humain diabetique. Etant donne que l’expression des enzymes cles de la neoglucogenese est fortement regulee par HNF4alpha et FOXO1, notre objectif a ete d’analyser l’interaction des transcrits de TCF7L2 avec ces facteurs de transcription dans les cellules hepatiques HepG2. Materiels et methodes Nous avons evalue l’expression de TCF7L2 par PCR quantitative specifique-au-exon dans les cellules HepG2 cultivees en presence de differentes concentrations de glucose et d’insuline. L’impact de l’inhibition de l’expression de TCF7L2 total ou de ses transcrits par ARNi sur l’expression des genes de la neoglucogenese a ete evalue. Par la technique du gene rapporteur luciferase, nous avons analyse le role de TCF7L2 dans la regulation de la transcription de la glucose-6-phosphatase (G6PC) et les interactions possibles entre TCF7L2, HNF4alpha et FOXO1. Resultats Notre analyse montre que l’expression des transcrits de TCF7L2, dont ceux qui sont induite dans le foie d’individus diabetiques, sont aussi induits dans les cellules HepG2 par glucose, de maniere dependante de la concentration. Grâce a la transfection d’ARNi dans les cellules HepG2, nous avons observe que la transcription de 29 genes de la neoglucogenese, dont G6PC, est reprimee par TCF7L2. La repression de la transcription de G6PC par TCF7L2 est independante de la presence de FOXO1 et de la regulation d’expression de HNF4alpha par TCF7L2. Par contre, certaines TCF7L2 transcrits interagir avec HNF4alpha dans les cellules HepG2. Conclusion Nos resultats suggerent que les transcrits alternatifs de TCF7L2 pourraient jouer un role important dans les anomalies du metabolisme hepatique du glucose dans le DT2.

Introduction Les etudes de GWAS sur l’heritabilite du DT2 ont montre le role important du gene TCF7L2. Nous avons montre que plusieurs transcrits alternatifs de TCF7L2 avaient des effets distincts sur la proliferation et la secretion d’insuline des cellules-beta pancreatiques. TCF7L2 est aussi implique dans le metabolisme du glucose des cellules hepatiques (Norton 2011, Savic 2011), d’ou l’interet d’analyser l’expression de TCF7L2 transcrits dans le foie. Patients et methodes Nous avons mesure l’ARNm de TCF7L2 par PCR-quantitative exon-specifique (d’apres Prokunina-Olsson 2009) dans des echantillons de tissu hepatique de patients obeses de la cohorte ABOS (33 normoglycemiques et 31 DT2). Dans les cellules HepG2, nous avons etudie l’expression des genes de la gluconeogenese apres diminution d’expression de TCF7L2 par siRNA. Nous avons egalement analyse les transcrits de TCF7L2 dans ces cellules cultivees avec differentes concentrations de glucose et d’insuline. Resultats Notre analyse montre que les transcrits de TCF7L2 contenant le premier site d’initiation de la transcription (TSS1) sont plus exprimes dans le foie des patients diabetiques que chez les sujets normoglycemiques (augmentation 1,3-fois du rapport TSS1/exon 8, P = 0,002). Grâce a la transfection d’ARNi dans les cellules HepG2, nous avons confirme que la transcription de plusieurs genes de la gluconeogenese, dont G6Pase, est reprimee par TCF7L2. Dans les cellules HepG2 en presence de 1 mM glucose, l’expression de toutes les transcrits de TCF7L2 contenant l’exon 8, dont les TSS1-transcrits, est augmentee d’environ 2 fois avec l’insuline (P = 0,03). D’autre part, le rapport TSS1/exon 8 est toujours augmente par l’insuline dans les cellules HepG2 cultivees avec 1 mM, 5 mM ou 11 mM glucose. Conclusion Nous avons demontre que differents transcrits de TCF7L2 sont induits par l’insuline et par un environnement diabetogene dans les hepatocytes. Ceci suggere que ces transcrits alternatifs pourraient jouer un role important dans les anomalies du metabolisme hepatique du glucose dans le DT2.