Regulation of the Aurora-A gene following topoisomerase I inhibition: implication of the Myc transcription factor.

During the G2 phase of the cell cycle, the Aurora-A kinase plays an important role in centrosome maturation and progression to mitosis. In this study, we show in colorectal cell lines that Aurora-A expression is downregulated in response to topoisomerase I inhibition. Using chromatin immunoprecipitation assays, we have observed that the Myc transcription factor and its Max binding partner are associated with the Aurora-A promoter during the G2 phase of the cell cycle. RNA interference experiments indicated that Myc is involved in the regulation of the Aurora-A gene. Following topoisomerase I inhibition, the expression of Myc decreased whereas Mad was upregulated, and the association of Myc and Max with the promoter of the kinase was inhibited. In parallel, an increased association of Mad and Miz-1 was detected on DNA, associated with an inhibition of the recruitment of transcriptional coactivators. Interestingly, a gain of H3K9 trimethylation and HP1gamma recruitment was observed on the Aurora-A promoter following sn38 treatment, suggesting that this promoter is located within SAHF foci following genotoxic treatment. Since Aurora-A is involved in centrosome maturation, we observed as expected that topoisomerase I inhibition prevented centrosome separation but did not affect their duplication. As a consequence, this led to G2 arrest and senescence induction.These results suggest a model by which the Aurora-A gene is inactivated by the G2 checkpoint following topoisomerase I inhibition. We therefore propose the hypothesis that the coordinated overexpression of Myc and Aurora-A, together with a downregulation of Mad and Miz-1 should be tested as a prognosis signature of poor responses to topoisomerase I inhibitors.

Leptin expression in Peripheral Blood Mononuclear Cells (PBMCs) is related with blood pressure variability.

BACKGROUND: Leptin is an adipokine initially considered as a molecule related exclusively to obesity but advances in research revealed its multiple roles in other physio-pathological mechanisms and particularly in the inflammatory ones. The aim of the present study was to demonstrate the presence of leptin in human Peripheral Blood Mononuclear Cells (PBMCs) and to quantify its mRNA in this type of tissue, closely related to inflammation. METHODS: Leptin mRNA was present in PBMCs of healthy individuals. Its expression was further studied in 83 individuals in relation to constitutional factors, anthropometric variables, blood pressure, lipid profile, glucose and markers of inflammation (C-reactive protein, lymphocyte count). RESULTS: Expression levels were significantly associated with systolic blood pressure (SBP) (p = 0.03) and diastolic blood pressure (DBP) (p = 0.003). Using a multiple regression analysis model, we showed that leptin mRNA levels explained 11% of the variation of SBP (p = 0.007) and of DBP (p = 0.003). These percentages remained at the same magnitude for SBP (9%) and for DBP (10%), after introducing BMI in the model. CONCLUSION: We report here for the first time, leptin expression in human PBMCs of healthy individuals. The associations found with blood pressure suggest a possible role of leptin in blood pressure regulation via PBMCs.

Visfatin, low-grade inflammation and body mass index (BMI).

OBJECTIVE: Visfatin is an adipokine with revealing roles in inflammatory mechanisms but its implication in inflammation related to excessive adiposity/obesity is not studied yet. Our aim was to investigate the relations of visfatin with inflammation markers and body mass index (BMI) in the peripheral blood mononuclear cells (PBMCs), a type of cells closely related to inflammatory mechanisms. DESIGN: Cross-sectional study, quantification of visfatin, TNF-alpha, IL-6 mRNA in PBMCs. PATIENTS: Eighty-three supposed healthy individuals from the STANISLAS cohort, belonging in three BMI categories: BMI < 25 kg/m(2) (lean), 25 kg/m(2) or= 30 kg/m(2) (obese). MEASUREMENTS: We measured visfatin gene expression (by real-time quantitative PCR), in relation to gene expression of the pro-inflammatory cytokines TNF-alpha, IL-6 in PBMCs and to anthropometric parameters (weight, BMI, waist : hip ratio), blood pressure, lipid profile, glucose and inflammatory markers (C-reactive protein, lymphocyte count). RESULTS: Visfatin expression in PBMCs was significantly associated with BMI in a negative way (r = -0.21, P = 0.05). Global anova analysis test for lean and over-weight/obese individuals showed a negative significant association between visfatin expression in PBMCs and BMI both for men and women (P = 0.05 and P = 0.01, respectively) and these associations remained significant after separating subjects in three groups (lean, overweight, obese) for men and women (P = 0.02 and P = 0.05, respectively). Correlation analysis between levels of expression of visfatin and TNF-alpha showed a significant positive linear association (r(2) = 0.27, P < 0.0001). CONCLUSION: These findings reveal a probable new role of visfatin in inflammation reflected in PBMCs, in the context of obesity.

Transcription factor and drug-metabolizing enzyme gene expression in lymphocytes from healthy human subjects.

We aimed to measure simultaneously the expression of drug-metabolizing enzymes (DME) and transcription factors (TF) with high importance in cardiovascular physiopathology in lymphocytes from healthy subjects. RNA was isolated from peripheral blood mononuclear cells (PBMC) of 20 subjects from the Stanislas Cohort. We used a microarray approach to measure 16 DME and 13 TF. Cytochromes P450 (P450s), including CYP2C19, CYP2C9, CYP2J2, CYP2D6, CYP1A1, CYP4F2, CYP4A11, CYP2E1, CYP11B2, CYP2C18, and CYP2A6, were expressed in all the subjects. CYP3A4 and CYP3A5 were not expressed. Glutathione S-transferases (GST) were expressed, but GSTM1 was seen only in some subjects. Pregnane X receptor (PXR), myocyte enhancer factor 2, vitamin D receptor, liver X receptor (LXR)-alpha, aryl hydrocarbon receptor (AHR), T-cell factor 7, constitutive androstane receptor, and aryl hydrocarbon receptor nuclear translocator (ARNT) were expressed in the majority of the subjects. Glucocorticoid receptor, peroxisome proliferator-activated receptor (PPAR)-gamma, and LXRbeta were expressed only in some individuals. PPARalpha mRNA was found in one subject only, and farnesoid X-activated receptor was not expressed. In addition, we found significant correlations between the expression of AHR, ARNT, and CYP1A1 and between PXR and P450 involved in leukotriene metabolism (CYP2C, CYP4F2, CYP4A11, CYP2J2, and CYP11B2). We describe here for the first time the presence of the majority of TF and DME in PBMC of healthy subjects without previous induction. The expression of these genes in lymphocytes could be a useful tool for further studying the physiological and pathological variations of DME and TF related to environment, to drug intake, and to cardiovascular metabolic cycles.

Genetic profiling in healthy subjects from the Stanislas cohort based on 24 polymorphisms: effects on biological variables.

BACKGROUND: The association of genetic profiles with biological or clinical assessments is not clearly established especially among apparently healthy subjects. METHODS: A multivariate statistical analysis was performed on 24 polymorphisms related to the main metabolic pathways involved in cardiovascular diseases (CVDs). They were collected among 1551 healthy subjects of the Stanislas cohort to obtain genetic profiles. Association with biological variables was then studied at baseline (t0) and 5 years later (t5). RESULTS: Six genetic clusters were identified with relevant profiles and five polymorphisms from the selectin, apolipoprotein C3 and lipoprotein lipase genes (SELE-98G/T, APOC3-3175C/G, APOC3-482C/T, APOC3-1100C/T, LPL-93T/G) were sufficiently characteristic to associate 99.6% of the subjects with their corresponding cluster. A 5-year follow-up showed that clinical and biological measurements in relation to CVD risk factors already differ with triglyceride (p=0.009 for t0 and p=0.005 for t5) and high-density lipoprotein cholesterol (p=0.014 for t0 and p=0.003 for t5) for these previous genetic clusters. CONCLUSIONS: This study presents the hypothesis that SELE could be protective, whereas APOC3 could be associated with risk. It remains to be seen whether these polymorphisms will be predictive of CVD events among the selected clusters of different metabolic subtypes after a 10-year follow-up.

Peripheral blood mononuclear cells (PBMCs): a possible model for studying cardiovascular biology systems.

BACKGROUND: The inflammation system, alone or in relation to or interaction with other cardiovascular pathways, is suggested to be the central pathway in the development and progression of cardiovascular diseases. The aim of the present investigation was to propose a specific and informative model for exploring this hypothesis. METHODS: In a biological system approach, we studied the expression of 182 candidate cardiovascular genes in peripheral blood mononuclear cells (PBMCs), cells that provide specific information on the inflammatory pathway. We explored their expression in 20 individuals with or without risk factors (obesity, hypertension) for cardiovascular disease. RESULTS: We found that: 1) 166 among the 182 selected genes were expressed in at least one individual's PBMCs, some of them being detected for the first time in this tissue; 2) all pathways were represented by the majority of their genes selected; 3) genes were expressed at a level sufficient for further study of the inter-individual variations in their mRNA to determine their biological variation; and 4) 15 genes discriminated hypertensive from obese or controls. CONCLUSIONS: The results of the present investigation support our proposal of a promising novel strategy based on PBMC transcriptomic studies to elucidate the complexity of the cardiovascular system in relation to inflammation. Preliminary data support the usefulness of the PBMC model in hypertension/inflammation research.

Analysis of the effect of multiple genetic variants of cardiovascular disease risk on insulin concentration variability in healthy adults of the STANISLAS cohort. The role of FGB-455 G/A polymorphism.

INTRODUCTION: Given the hypothesis of a common soil for atherosclerosis, type 2 diabetes and metabolic syndrome, we tested the contribution of gene polymorphisms involved in cardiovascular diseases on fasting insulin concentration (FIC). METHODS: The polymorphisms were investigated by a multiplex assay in 308 apparently healthy French middle-aged men and women, taken from the STANISLAS cohort. FIC was measured by a microparticular enzymatic immunoassay. RESULTS: After a series of regression analyses involving 34 polymorphisms, FGB -455G/A was the only polymorphism that remained significantly associated with FIC when adjusting the analyses for multiple testing. Stepwise models showed that FGB polymorphism accounted for 4.39% of FIC variability in men. Additionally, interactions between FGB and with environmental factors (alcohol and smoking in men, and BMI in women) were found. DISCUSSION: To our knowledge, this is the first study reporting an influence of FGB polymorphism on FIC in a healthy population. Our results concord with the already shown link between fibrinogen concentration and FIC, and support the hypothesis of a relationship between fibrinogen and endothelium in FIC homeostasis whose alteration may induce several metabolic disorders. The contribution of this gene, although modest, is consistent with the polygenic nature of insulin levels.

Upregulation of the tumor suppressor gene menin in hepatocellular carcinomas and its significance in fibrogenesis.

The molecular mechanisms underlying the progression of cirrhosis toward hepatocellular carcinoma were investigated by a combination of DNA microarray analysis and literature data mining. By using a microarray screening of suppression subtractive hybridization cDNA libraries, we first analyzed genes differentially expressed in tumor and nontumor livers with cirrhosis from 15 patients with hepatocellular carcinomas. Seventy-four genes were similarly recovered in tumor (57.8% of differentially expressed genes) and adjacent nontumor tissues (64% of differentially expressed genes) compared with histologically normal livers. Gene ontology analyses revealed that downregulated genes (n = 35) were mostly associated with hepatic functions. Upregulated genes (n = 39) included both known genes associated with extracellular matrix remodeling, cell communication, metabolism, and post-transcriptional regulation gene (e.g., ZFP36L1), as well as the tumor suppressor gene menin (multiple endocrine neoplasia type 1; MEN1). MEN1 was further identified as an important node of a regulatory network graph that integrated array data with array-independent literature mining. Upregulation of MEN1 in tumor was confirmed in an independent set of samples and associated with tumor size (P = .016). In the underlying liver with cirrhosis, increased steady-state MEN1 mRNA levels were correlated with those of collagen alpha2(I) mRNA (P < .01). In addition, MEN1 expression was associated with hepatic stellate cell activation during fibrogenesis and involved in transforming growth factor beta (TGF-beta)-dependent collagen alpha2(I) regulation. In conclusion, menin is a key regulator of gene networks that are activated in fibrogenesis associated with hepatocellular carcinoma through the modulation of TGF-beta response.

Interaction between CYP1A1 T3801C and AHR G1661A polymorphisms according to smoking status on blood pressure in the Stanislas cohort.

BACKGROUND: CYP1A1, one of the key enzymes in detoxifying toxic components produced during cigarette smoking, is regulated by aromatic hydrocarbon receptor (AHR). A CYP1A1 T3801C polymorphism, associated with a higher CYP1A1 inducibility and enhanced catalytic activity, has been linked to stroke, triple vessel disease and may, therefore, be associated with blood pressure (BP). The relation of the widely studied G1661A polymorphism of the human AHR gene with BP is unknown. OBJECTIVES: To investigate the genetic influence of CYP1A1 T3801C and AHR G1661A polymorphisms on BP in relation to tobacco consumption. DESIGN AND PARTICIPANTS: Study participants were selected from a French longitudinal cohort of volunteers for a free health check-up. These individuals (302 men and 311 women) were not taking medication that can affect blood pressure. Information about active smoking status was obtained by a self-administered questionnaire. RESULTS: After multiple regression analysis, systolic blood pressure (SBP) and diastolic blood pressure (DBP) did not differ significantly according to their tobacco status excepted for DBP in men. In addition, neither CYP1A1 T3801C nor AHR G1661A polymorphism was linked to blood pressure. However, systolic and diastolic blood pressures differed significantly according to CYP1A1 T3801C genotype between ex-smokers and smokers. Finally, the interaction between CYP1A1 T3801C and AHR G1661A polymorphisms explained a significant difference of SBP and DBP between carriers of both CYP1A1-C3801 and AHR-A1661 alleles. CONCLUSION: This study is the first to show an interaction between the CYP1A1 T3801C and AHR G1661A polymorphisms. This interaction could explain the difference in blood pressure level between smokers and non-smokers/ex-smokers but needs to be confirmed in a large sample.

A prospective study on the prevalence of metabolic syndrome among healthy french families: two cardiovascular risk factors (HDL cholesterol and tumor necrosis factor-alpha) are revealed in the offspring of parents with metabolic syndrome.

OBJECTIVE: The purpose of this study was to estimate the longitudinal variation of prevalence of metabolic syndrome within French families and to observe biological parameters involved in cardiovascular disease among their offspring. RESEARCH DESIGN AND METHODS: Three hundred seventy-one apparently healthy families (1,366 individuals) taken from the STANISLAS cohort were studied. The subjects were examined at two time points with a 5-year interval (t(0) and t(+5)). The crude prevalence of metabolic syndrome was assessed among parents according to the National Cholesterol Education Program Adult Treatment Panel III (NCEP ATP) definition. RESULTS: The prevalence of metabolic syndrome was 5.9% in men and 2.1% in women at t(0), rising to 7.2 and 5.4% in men and women, respectively, at t(+5). Children of parents having metabolic syndrome showed higher levels of tumor necrosis factor-alpha (TNF-alpha), whereas their HDL cholesterol and apolipoprotein (apo) E concentrations were lower compared with those of age- and sex-matched control subjects (P

Biological variations, genetic polymorphisms and familial resemblance of TNF-alpha and IL-6 concentrations: STANISLAS cohort.

Cytokines are involved in the development of several inflammatory diseases and atherosclerosis. Their variations in healthy individuals are not well defined. The aims of this study were: firstly, to identify factors affecting biological variation of interleukin-6 (IL-6) and tumour necrosis factor-alpha (TNF-alpha); secondly, to study their family resemblance; and thirdly, to evaluate the effect of two TNF-alpha (-308G/A and -238G/A) and two IL-6 polymorphisms (174G/C and -572G/C) on their corresponding circulating levels. A total of 171 healthy families selected from the STANISLAS cohort were studied. Age was negatively related to TNF-alpha concentrations in offspring only (both sons and daughters). Additionally, IL-6 and TNF-alpha levels were differently influenced by gender, white blood cells, tobacco consumption, and HDL-cholesterol level. A weak significant familial resemblance for TNF-alpha concentration was observed in siblings only. There was no significant familial resemblance for IL-6 levels. The TNF-alpha -308A allele was associated with decreased TNF-alpha concentrations in both offspring aged less than 18 and males without overweight (BMI<25 kg/m(2)). Fathers carrying the IL-6 -174CC genotype had higher IL-6 levels than those with the IL-6 -174G allele. Parents with the IL-6 -572GG genotype had higher IL-6 concentrations than the C allele carriers. In this sample of healthy families, plasma levels of IL-6 and TNF-alpha were differently affected by biological parameters including age, gender and smoking, and the impact of their respective polymorphisms was influenced by gender, age and BMI.