Rôle modulateur de la glutathion transférase Pi dans la prolifération et la mort des cellules normales et transformées

L'expression élevée de la GSTP1 est fréquemment observée dans les cancers et est positivement corrélée à la résistance aux chimiothérapies. Cette enzyme de détoxication de phase II peut aussi réguler l'activité de protéines comme JNK et TRAF2 et, par conséquent, peut moduler les voies de prolifération et de mort cellulaire. Ce projet a donc consisté à étudier le rôle de la GSTP1 dans la prolifération des hépatocytes normaux ou transformés. L'étude de la régénération hépatique chez des souris Gstp1/2 / a permis de démontrer le rôle des protéines GSTP1 et GSTP2 dans le contrôle de la progression des hépatocytes normaux dans le cycle cellulaire. Après hépatectomie partielle chez les souris Gstp1/2 / , une diminution importante du nombre d'hépatocytes dans les phases S, G2 et M est observée comparativement à des foies de souris contrôle. Cette réduction est associée à des retards d'expression de protéines impliquées dans l'initiation de la prolifération, le contrôle du point de restriction dépendant des mitogènes et dans la transition G1/S. Ces modifications sont associées à une réduction de l'expression de TRAF2 et de l'activation de JNK et ERK, alors que les taux de p21 et de p53 sont élevés. Parallèlement, un décalage dans l'expression d'enzymes qui régulent l'homéostasie redox et participent à l'activation des MAPK est observé. L'utilisation de cellules cancéreuses de différentes origines dont le foie, a également permis de corréler l'absence de GSTP1 à une diminution de prolifération cellulaire sans altération de la suivie cellulaire. Cependant dans ces conditions, nous observons une augmentation de l'expression de TRAF2, pJNK, pATF2, ATF3 associée à une induction de p21. Nous avons également montré que les effets de la GSTP1 sur la prolifération cellulaire sont régulés par l'activation de JNK. L'évidence du lien entre l'expression de la GSTP1 et la prolifération hépatocytaire nous a conduit à analyser l'expression d'enzymes de détoxication dans des carcinomes hépatocellulaires (CHC) et nous avons constaté une induction d'expression de GSTP1 dans le tissu péritumoral des CHC par rapport au foie normal.Increased GSTP1 expression is frequently observed in cancers and is positively correlated with chemotherapy resistance. This phase II detoxifying enzyme can also regulate JNK and TRAF2 activities and, consequently, can modulate proliferation and cell death pathways. This project aimed at studying the role of GSTP1 during proliferation in normal and transformed hepatocytes. Liver regeneration study in Gstp1/2 / mice showed the involvement of GSTP1 and GSTP2 proteins in the cell cycle progression control of normal hepatocytes. After partial hepatectomy in Gstp1/2 / mice, the number of cells in S, G2 and M phases was decreased compared to livers of wildtype mice. This reduction is associated with the delay in the expression of proteins involved in proliferation initiation, mitogen restriction point control and G1/S transition. These modifications are associated with the decrease in TRAF2 expression and the activation of JNK and ERK, whereas p21 and p53 levels are high. Furthermore, expression of enzymes involved in redox homeostasis and MAPK activation is delayed. Study of cells derived from various cancers, including HCC, highlighted a correlation between low expression of GSTP1 and decrease in cell proliferation without cell survival alteration. However in these conditions, we observed the increase in TRAF2, pJNK, pATF2 and ATF3 expression together with the induction of p21. We also showed that GSTP1 effects are regulated by JNK activation. These results showed a link between GSTP1 expression and hepatocyte proliferation and led us to investigate the GSTP1 expression in HCC. We noticed an induction of GSTP1 expression in peritumoral tissue compared to normal liver.RENNES1-Bibl. électronique (352382106) / SudocSudocFranceF

Glutathione transferase pi modulatory role in proliferation and death of normal and transformed cells

L'expression élevée de la GSTP1 est fréquemment observée dans les cancers et est positivement corrélée à la résistance aux chimiothérapies. Cette enzyme de détoxication de phase II peut aussi réguler l'activité de protéines comme JNK et TRAF2 et, par conséquent, peut moduler les voies de prolifération et de mort cellulaire. Ce projet a donc consisté à étudier le rôle de la GSTP1 dans la prolifération des hépatocytes normaux ou transformés. L'étude de la régénération hépatique chez des souris Gstp1/2‐/‐ a permis de démontrer le rôle des protéines GSTP1 et GSTP2 dans le contrôle de la progression des hépatocytes normaux dans le cycle cellulaire. Après hépatectomie partielle chez les souris Gstp1/2‐/‐, une diminution importante du nombre d'hépatocytes dans les phases S, G2 et M est observée comparativement à des foies de souris contrôle. Cette réduction est associée à des retards d'expression de protéines impliquées dans l'initiation de la prolifération, le contrôle du point de restriction dépendant des mitogènes et dans la transition G1/S. Ces modifications sont associées à une réduction de l'expression de TRAF2 et de l'activation de JNK et ERK, alors que les taux de p21 et de p53 sont élevés. Parallèlement, un décalage dans l'expression d'enzymes qui régulent l'homéostasie redox et participent à l'activation des MAPK est observé. L'utilisation de cellules cancéreuses de différentes origines dont le foie, a également permis de corréler l'absence de GSTP1 à une diminution de prolifération cellulaire sans altération de la suivie cellulaire. Cependant dans ces conditions, nous observons une augmentation de l'expression de TRAF2, pJNK, pATF2, ATF3 associée à une induction de p21. Nous avons également montré que les effets de la GSTP1 sur la prolifération cellulaire sont régulés par l'activation de JNK. L'évidence du lien entre l'expression de la GSTP1 et la prolifération hépatocytaire nous a conduit à analyser l'expression d'enzymes de détoxication dans des carcinomes hépatocellulaires (CHC) et nous avons constaté une induction d'expression de GSTP1 dans le tissu péritumoral des CHC par rapport au foie normal.Increased GSTP1 expression is frequently observed in cancers and is positively correlated with chemotherapy resistance. This phase II detoxifying enzyme can also regulate JNK and TRAF2 activities and, consequently, can modulate proliferation and cell death pathways. This project aimed at studying the role of GSTP1 during proliferation in normal and transformed hepatocytes. Liver regeneration study in Gstp1/2‐/‐ mice showed the involvement of GSTP1 and GSTP2 proteins in the cell cycle progression control of normal hepatocytes. After partial hepatectomy in Gstp1/2‐/‐ mice, the number of cells in S, G2 and M phases was decreased compared to livers of wildtype mice. This reduction is associated with the delay in the expression of proteins involved in proliferation initiation, mitogen restriction point control and G1/S transition. These modifications are associated with the decrease in TRAF2 expression and the activation of JNK and ERK, whereas p21 and p53 levels are high. Furthermore, expression of enzymes involved in redox homeostasis and MAPK activation is delayed. Study of cells derived from various cancers, including HCC, highlighted a correlation between low expression of GSTP1 and decrease in cell proliferation without cell survival alteration. However in these conditions, we observed the increase in TRAF2, pJNK, pATF2 and ATF3 expression together with the induction of p21. We also showed that GSTP1 effects are regulated by JNK activation. These results showed a link between GSTP1 expression and hepatocyte proliferation and led us to investigate the GSTP1 expression in HCC. We noticed an induction of GSTP1 expression in peritumoral tissue compared to normal liver

Regulation of signal transduction by glutathione transferases.

International audienceGlutathione transferases (GST) are essentially known as enzymes that catalyse the conjugation of glutathione to various electrophilic compounds such as chemical carcinogens, environmental pollutants, and antitumor agents. However, this protein family is also involved in the metabolism of endogenous compounds which play critical roles in the regulation of signaling pathways. For example, the lipid peroxidation product 4-hydroxynonenal (4-HNE) and the prostaglandin 15-deoxy-Δ12,14-prostaglandin J(2) (15d-PGJ(2)) are metabolized by GSTs and these compounds are known to influence the activity of transcription factors and protein kinases involved in stress response, proliferation, differentiation, or apoptosis. Furthermore, several studies have demonstrated that GSTs are able to interact with different protein partners such as mitogen activated protein kinases (i.e., c-jun N-terminal kinase (JNK) and apoptosis signal-regulating kinase 1 (ASK1)) which are also involved in cell signaling. New functions of GSTs, including S-glutathionylation of proteins by GSTs and ability to be a nitric oxide (NO) carrier have also been described. Taken together, these observations strongly suggest that GST might play a crucial role during normal or cancer cells proliferation or apoptosis

Borrelia miyamotoi: 43 Cases Diagnosed in France by Real-Time PCR in Patients With Persistent Polymorphic Signs and Symptoms

International audienceBackground: Borrelia species are divided into three groups depending on the induced disease and the tick vector. Borrelia miyamotoi is a relapsing fever Borrelia but can induce symptoms related to Lyme disease. Discovered in 1995, it is found in ticks around the world. In France, this species of Borrelia has been isolated in ticks and rodents, but was not yet observed in humans. Objective: The aim of the study was to look for B. miyamotoi in symptomatic patients. Methods: Real-time PCR was performed on 824 blood samples from patients presenting symptoms of persistent polymorphic syndrome possibly due to tick bite, a syndrome recognized by the French Authority for Health, which is close to the post-treatment Lyme disease syndrome. PCR was also performed on 24 healthy control persons. The primers were specifically designed for this particular species of Borrelia. The sequence of interest of 94 bp is located on the glpQ gene. Sequencing of amplification products, randomly chosen, confirmed the amplification specificity. To better investigate cases, a clinical questionnaire was sent to the patients PCR-positive for B. miyamotoi and to their physician. Results: This search revealed a positive PCR for B. miyamotoi in the blood from 43 patients out of 824 (5.22%). PCR was negative in all control persons. A clinical chart was obtained from 31 of the 43 patients. A history of erythema migrans was reported in five of these 31 patients (16%). All patients complained about fatigue, joint pain and neuro-cognitive disorders. Some patients complained about respiratory problems (chest tightness and/or lack of air in 41.9%). Episodes of relapsing fever were reported by 11 of the 31 patients (35.5%). Chilliness, hot flushes and/or sweats were reported by around half of the patients. B. miyamotoi may not cross-react with B. burgdorferi serology. Conclusion: This study is the first to detect B. miyamotoi in human blood in France. This series of human B. miyamotoi infection is the largest in patients with long term persistent syndrome. Our data suggest that this infection may be persistent, even on the long term

Pretreatment with Mangafodipir Improves Liver Graft Tolerance to Ischemia/Reperfusion Injury in Rat

Ischemia/reperfusion injury occurring during liver transplantation is mainly due to the generation of reactive oxygen species (ROS) upon revascularization. Thus, delivery of antioxidant enzymes might reduce the deleterious effects of ROS and improve liver graft initial function. Mangafodipir trisodium (MnDPDP), a contrast agent currently used in magnetic resonance imaging of the liver, has been shown to be endowed with powerful antioxidant properties. We hypothesized that MnDPDP could have a protective effect against liver ischemia reperfusion injury when administrated to the donor prior to harvesting. Livers from Sprague Dawley rats pretreated or not with MnDPDP were harvested and subsequently preserved for 24 h in CelsiorH solution at 4uC. Organs were then perfused ex vivo for 120 min at 37uC with Krebs Henseleit solution. In MnDPDP (5 mmol/kg) group, we observed that ATP content was significantly higher at the end of the cold preservation period relative to untreated group. After reperfusion, livers from MnDPDP-treated rats showed better tissue integrity, less hepatocellular and endothelial cell injury. This was accompanied by larger amounts of bile production and higher ATP recovery as compared to untreated livers. The protective effect of MnDPDP was associated with a significant decrease of lipid peroxidation, mitochondrial damage, and apoptosis. Interestingly, MnDPDP-pretreated livers exhibited activation of Nfr2 and HIF-1a pathways resulting in a higher catalase and HO-1 activities. MnDPDP also increased total nitric oxide (NO) production which derived from higher expression of constitutive NO synthase and lower expression of inducible N

Carbon tetrachloride-mediated lipid peroxidation induces early mitochondrial alterations in mouse liver.

International audienceAlthough carbon tetrachloride (CCl(4))-induced acute and chronic hepatotoxicity have been extensively studied, little is known about the very early in vivo effects of this organic solvent on oxidative stress and mitochondrial function. In this study, mice were treated with CCl(4) (1.5 ml/kg ie 2.38 g/kg) and parameters related to liver damage, lipid peroxidation, stress/defense and mitochondria were studied 3 h later. Some CCl(4)-intoxicated mice were also pretreated with the cytochrome P450 2E1 inhibitor diethyldithiocarbamate or the antioxidants Trolox C and dehydroepiandrosterone. CCl(4) induced a moderate elevation of aminotransferases, swelling of centrilobular hepatocytes, lipid peroxidation, reduction of cytochrome P4502E1 mRNA levels and a massive increase in mRNA expression of heme oxygenase-1 and heat shock protein 70. Moreover, CCl(4) intoxication induced a severe decrease of mitochondrial respiratory chain complex IV activity, mitochondrial DNA depletion and damage as well as ultrastructural alterations. Whereas DDTC totally or partially prevented all these hepatic toxic events, both antioxidants protected only against liver lipid peroxidation and mitochondrial damage. Taken together, our results suggest that lipid peroxidation is primarily implicated in CCl(4)-induced early mitochondrial injury. However, lipid peroxidation-independent mechanisms seem to be involved in CCl(4)-induced early hepatocyte swelling and changes in expression of stress/defense-related genes. Antioxidant therapy may not be an efficient strategy to block early liver damage after CCl(4) intoxication

Differences in early acetaminophen hepatotoxicity between obese ob/ob and db/db mice.

International audienceClinical investigations suggest that hepatotoxicity after acetaminophen (APAP) overdose could be more severe in the context of obesity and nonalcoholic fatty liver disease. The pre-existence of fat accumulation and CYP2E1 induction could be major mechanisms accounting for such hepatic susceptibility. To explore this issue, experiments were performed in obese diabetic ob/ob and db/db mice. Preliminary investigations performed in male and female wild-type, ob/ob, and db/db mice showed a selective increase in hepatic CYP2E1 activity in female db/db mice. However, liver triglycerides in these animals were significantly lower compared with ob/ob mice. Next, APAP (500 mg/kg) was administered in female wild-type, ob/ob, and db/db mice, and investigations were carried out 0.5, 2, 4, and 8 h after APAP intoxication. Liver injury 8 h after APAP intoxication was higher in db/db mice, as assessed by plasma transaminases, liver histology, and terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling assay. In db/db mice, however, the extent of hepatic glutathione depletion, levels of APAP-protein adducts, c-Jun N-terminal kinase activation, changes in gene expression, and mitochondrial DNA levels were not greater compared with the other genotypes. Furthermore, in the db/db genotype plasma lactate and β-hydroxybutyrate were not specifically altered, whereas the plasma levels of APAP-glucuronide were intermediary between wild-type and ob/ob mice. Thus, early APAP-induced hepatotoxicity was greater in db/db than ob/ob mice, despite less severe fatty liver and similar basal levels of transaminases. Hepatic CYP2E1 induction could have an important pathogenic role when APAP-induced liver injury occurs in the context of obesity and related metabolic disorders

Mangafodipir limits endothelial cells injury and favors nitric oxide production.

<p>Perfusate Purine Nucleotide Phosphorylase (PNP) activity (A) and tissue accumulation of Nitrate/nitrite levels (B) were measured in livers harvested from untreated rats and immediately submitted to 120 min of normothermic reperfusion (Control group), in livers procured from vehicle (NaCl) pretreated rats, cold stored for 24 h at 4°C and submitted to 120 min of normothermic reperfusion (I/R group) and in livers procured from 5 µmol/kg MnDPDP-treated rats, cold stored for 24 h at 4°C and submitted to 120 min of normothermic reperfusion (MnDPDP+I/R group). Western blot and densitometry analysis of constitutive nitric oxide synthase (eNOS) (C) and inducible nitric oxide synthase (iNOS) (D) in livers from control, I/R and MnDPDP+I/R groups. Western blot data represent one of three independent experiments with similar results. Western blot densitometry data are normalized for the loading control HSC70. * <i>p</i><0.05: either I/R or MnDPDP+I/R versus Control; ⁺<i>p</i><0.05: MnDPDP+I/R versus I/R.</p

Mangafodipir prevents ATP depletion.

<p>(A) Hepatic ATP content were measured in normal livers, in livers procured from sodium chloride solution pretreated rats and cold stored for 24 h at 4°C (Ischemia group) and in livers procured from 5 µmol/kg MnDPDP-treated rats and cold stored for 24 h at 4°C (MnDPDP+Ischemia group). Results were expressed as µmol/mg liver. * <i>p</i><0.05, either Ischemia or MnDPDP+Ischemia versus Control; ⁺<i>p</i><0.05, MnDPDP+Ischemia versus Ischemia. (B) Hepatic ATP content were measured in livers harvested from untreated rats and immediately submitted to 120 min of normothermic reperfusion (Control group), in livers procured from vehicle (NaCl) pretreated rats, cold stored for 24 h at 4°C and submitted to 120 min of normothermic reperfusion (I/R group) and in livers procured from 5 µmol/kg MnDPDP-treated rats, cold stored for 24 h at 4°C and submitted to 120 min of normothermic reperfusion (MnDPDP+I/R group). * <i>p</i><0.05: either I/R or MnDPDP+I/R versus Control; ⁺<i>p</i><0.05: MnDPDP+I/R versus I/R.</p