Marie-Paule Gustin

The microRNA Signature in Response to Insulin Reveals Its Implication in the Transcriptional Action of Insulin in Human Skeletal Muscle and the Role of a Sterol Regulatory Element–Binding Protein-1c/Myocyte Enhancer Factor 2C Pathway

OBJECTIVE Factors governing microRNA expressions in response to changes of cellular environment are still largely unknown. Our aim was to determine whether insulin, the major hormone controlling whole-body energy homeostasis, is involved in the regulation of microRNA expressions in human skeletal muscle. RESEARCH DESIGN AND METHODS We carried out comparative microRNA (miRNA) expression profiles in human skeletal muscle biopsies before and after a 3-h euglycemic-hyperinsulinemic clamp, with TaqMan low-density arrays. Then, using DNA microarrays, we determined the response to insulin of the miRNA putative target genes in order to determine their role in the transcriptional action of insulin. We further characterized the mechanism of action of insulin on two representative miRNAs, miR-1 and miR-133a, in human muscle cells. RESULTS Insulin downregulated the expressions of 39 distinct miRNAs in human skeletal muscle. Their potential target mRNAs coded for proteins that were mainly involved in insulin signaling and ubiquitination-mediated proteolysis. Bioinformatic analysis suggested that combinations of different downregulated miRNAs worked in concert to regulate gene expressions in response to insulin. We further demonstrated that sterol regulatory element–binding protein (SREBP)-1c and myocyte enhancer factor 2C were involved in the effect of insulin on miR-1 and miR-133a expression. Interestingly, we found an impaired regulation of miRNAs by insulin in the skeletal muscle of type 2 diabetic patients, likely as consequences of altered SREBP-1c activation. CONCLUSIONS This work demonstrates a new role of insulin in the regulation of miRNAs in human skeletal muscle and suggests a possible implication of these new modulators in insulin resistance.

Depressive feelings in children with narcolepsy.

OBJECTIVES We aimed to evaluate depressive feelings and their correlations in children and adolescents with narcolepsy collected in national reference centers for narcolepsy. METHODS We compared clinical and sleep characteristics of patients with and without depressive symptoms evaluated on the Childrens Depression Inventory (CDI). RESULTS Our study sample included 88 children (44 boys; 44 de novo patients) with a mean age of 11.9 ± 3.1 years at diagnosis (37.5% were aged ⩽ 10 years). Obesity was found in 59% of the sample and cataplexy was present in 80.7%. The DQB1*0602 allele was positive in 93.5% of our sample. There were 25% of children who had clinically depressive feelings (CDI>16), especially girls older than the age of 10 years. Bivariate associations indicated that depressive feelings were associated with fatigue (48%), hyperactivity (31%), insomnia (16%), and excessive daytime sleepiness (EDS) (14-24%). In the multivariate model adjusted for gender and age, only fatigue explained the variability of the depression score. CONCLUSION In our large cohort, high levels of depressive symptoms essentially expressed by fatigue affected 25% of children with narcolepsy. The girls older than 10 years of age were especially vulnerable. The similar prevalence of depressive feelings in treated vs never-treated patients suggests a specific need for diagnosing and managing this symptom in young patients with narcolepsy.

Beat-to-beat estimation of windkessel model parameters in conscious rats

A windkessel model was applied on a beat-to-beat basis to evaluate the arterial mechanical characteristics in seven conscious rats. Ascending aortic arterial pressure (AP) and blood flow were recorded during steady-state in basal conditions, during infusions of isoprenaline, sodium nitroprusside, and phenylephrine, and after intravenous atenolol injection. For each cardiac cycle the exponential decay time constant (τ) was estimated from the aortic AP curve, peripheral resistances ( R) were taken as the ratio of mean AP to cardiac output, and systemic arterial compliance ( C) was calculated as τ/ R. In all conditions, mean correlation coefficients of the exponential regression and ∼70% of values in each rat were >0.99, demonstrating the model validity. In all conditions τ and C exhibited a large spontaneous variability over time, and beat-to-beat correlations were high between τ and C (0.83 ± 0.03). C was increased by sodium nitroprusside, decreased by isoprenaline, but not significantly decreased by phenylephrine [5.1 ± 0.2, 3.2 ± 0.3, and 3.9 ± 0.2 μl/mmHg, respectively, vs. 4.2 ± 0.3 μl/mmHg (baseline)]. In conclusion, the windkessel model enables τ and C to be reliably estimated in conscious rats during spontaneous and drug-induced hemodynamic variations.A windkessel model was applied on a beat-to-beat basis to evaluate the arterial mechanical characteristics in seven conscious rats. Ascending aortic arterial pressure (AP) and blood flow were recorded during steady-state in basal conditions, during infusions of isoprenaline, sodium nitroprusside, and phenylephrine, and after intravenous atenolol injection. For each cardiac cycle the exponential decay time constant (tau) was estimated from the aortic AP curve, peripheral resistances (R) were taken as the ratio of mean AP to cardiac output, and systemic arterial compliance (C) was calculated as tau/R. In all conditions, mean correlation coefficients of the exponential regression and approximately 70% of values in each rat were > 0.99, demonstrating the model validity. In all conditions tau and C exhibited a large spontaneous variability over time, and beat-to-beat correlations were high between tau and C (0.83 +/- 0.03). C was increased by sodium nitroprusside, decreased by isoprenaline, but not significantly decreased by phenylephrine [5.1 +/- 0.2, 3.2 +/- 0.3, and 3.9 +/- 0.2 microliters/mmHg, respectively, vs. 4.2 +/- 0.3 microliters/mmHg (baseline)]. In conclusion, the windkessel model enables tau and C to be reliably estimated in conscious rats during spontaneous and drug-induced hemodynamic variations.

Functional meta-analysis of double connectivity in gene coexpression networks in mammals.

In functional genomics, the high-throughput methods such as microarrays 1) allow analysis of the relationships between genes considering them as elements of a network and 2) lead to biological interpretations thanks to Gene Ontology. But up to now it has not been possible to find relationships between the functions and the connectivity of the genes in coexpression networks. To achieve this aim, we have defined a double connectivity for each gene by the numbers of its significant negative and positive correlations with the other genes within a given biological condition, or group. Here, based on the analysis of 1,260 DNA microarrays, we show that this double connectivity clearly separates two types of genes, those with a predominantly strong negative connectivity, hub- genes, and those with a predominantly strong positive connectivity, hub+ genes. Interestingly, the hub+ genes concerned transcription factors more often than by chance and, similarly, for the hub- genes concerning miRNA predicted targets. Furthermore, a meta-analysis of GO annotations carried out on 67 groups in humans and rats shows that these two types of genes correspond to a functional biological duality. The hub- genes were mainly involved in basic functions common to all eukaryote cells, whereas the hub+ genes were mainly involved in specialized functions related to cell differentiation and communication. The separation and the biological role of these hub- and hub+ genes provide a powerful new tool for a better understanding of the control and regulation of the key genes involved in cellular differentiation and physiopathological conditions.

Quality of Life in Children with Narcolepsy

To evaluate the health‐related quality of life (HRQL) and its correlates in children and adolescents with narcolepsy.

Critical Study of Observation of the Sternal End of the Right 4th Rib

Abstract:  Studies of the method of estimating age at death by the 4th rib exclusively concerned the phase method without fundamentally challenging the method as such. The present study analyzed observation of the variables on which the İşcan method is based. Ten observers made two assessments of the stage of pit depth, pit shape, rim and wall configurations of 59 right 4th ribs harvested from males (mean age: 49 years; range: 47–94 years). Observation showed poor reproducibility and repeatability for all three variables (Wilcoxon test, κ‐coefficient). Analysis of problem ribs revealed difficulty in measuring and imprecision in describing pit depth and failure to take account of continual aging for the other two variables. Despite these results, İşcan’s variables provide objective information on age at death. It is recommended that the method be improved by better description of the variables and use of multivariate statistical analysis.

Computational identification of potential transcriptional regulators of TGF-ß1 in human atherosclerotic arteries

TGF-ß is protective in atherosclerosis but deleterious in metastatic cancers. Our aim was to determine whether TGF-ß transcriptional regulation is tissue-specific in early atherosclerosis. The computational methods included 5 steps: (i) from microarray data of human atherosclerotic carotid tissue, to identify the 10 best co-expressed genes with TGFB1 (TGFB1 gene cluster), (ii) to choose the 11 proximal promoters, (iii) to predict the TFBS shared by the promoters, (iv) to identify the common TFs co-expressed with the TGFB1 gene cluster, and (v) to compare the common TFs in the early lesions to those identified in advanced atherosclerotic lesions and in various cancers. Our results show that EGR1, SP1 and KLF6 could be responsible for TGFB1 basal expression, KLF6 appearing specific to atherosclerotic lesions. Among the TFs co-expressed with the gene cluster, transcriptional activators (SLC2A4RG, MAZ) and repressors (ZBTB7A, PATZ1, ZNF263) could be involved in the fine-tuning of TGFB1 expression in atherosclerosis.

Effect modification of aortic atheroma on the prognostic value of heart rate in hypertension.

Background: Although some epidemiological studies have advocated a prognostic value of heart rate (HR) in hypertensive patients, the influence of vascular damages on this prognostic value has not been tested yet. Methods: HRs were collected by pulse palpation in 1204 primary hypertensive patients in sinus rhythm without cardiac-slowing drugs. Aortic damages were assessed by aortography, whereas cardiac disease was assessed by medical history, symptoms and electrocardiogram. Results: In a multivariable Cox model adjusted for major confounders, HR was of prognostic significance for all-cause [hazard ratio 1.12 (1.06–1.19) for 10 bpm increment and 1.39 (1.18–1.64) for HR ≥82 vs. <82 bpm] and cardiovascular death [hazard ratio 1.10 (1.02–1.20) for 10 bpm increment and 1.37 (1.09–1.72) for HR ≥82 vs. <82 bpm] after 35 years of follow-up. This association was particularly manifested at 15 years of follow-up. At that time, with the same multivariable survival model, the association between HR and cardiovascular death was stronger in patients with aortic atheroma [2.76 (1.47–5.18) for an HR ≥82 vs. <82 bpm] than in patients without [hazard ratio 1.36 (0.76–2.43) for an HR ≥82 vs. <82 bpm, P for interaction = 0.054]. Similarly, the association between HR and cardiovascular death was stronger in patients with an overt cardiac disease than those without (P for interaction = 0.044). Conclusion: In hypertensive patients, the prognostic significance of HR for cardiovascular outcome is modulated by the presence of aortic atherosclerosis or cardiac disease. This should prompt us to a thorough examination of cardiovascular damages in hypertensive patients when HR is elevated.

Analytical investigation of salivary calculi, by mid-infrared spectroscopy

Sialolithiasis is common in salivary glands, especially in the submandibular and parotid ducts. X-Ray diffractometry was the principal technique used for their analysis, sometimes associated with scanning electron microscopy. Hydroxyapatite was the most frequently described constituent, in association with whitlockite and other calcium phosphates as brushite or octocalcium phosphate. Proteic matter was detected, as mucoproteins, albumin, nucleoproteins or as degenerative bacterial matter. This study presents the identification of constituents by mid-infrared spectrometry of 74 sialoliths. Their successive layers are analyzed from their crust to the nucleus, using absorbance measurements. Spectra are compared with reference mixtures of two or more constituents. Approximately 99% of sialoliths are constituted of calcium phosphates, under carbonated forms. More than three-quarters contain proteins, in which mucins represent the majority and albumin is found in 10% of all the specimens. Only 7% calculi are an association of two constituents, 66% are made of three and 27% have four or more components. For the 74 studied sialoliths, no specimen contains hydroxyapatite; but they are composed of carbonate apatites with irregular microcrystallized forms, even if proteins are present. Some of them have a pure protein nucleus, surrounded by carbonate apatite layers; the other stones are made of internal layers of apatites and covered with a dense and varnished crust of proteins.

Interpretation of drug concentrations in an alternative matrix: the case of meprobamate in bile

Investigating toxicological causes of death may require alternative matrices when the usual ones are lacking. Whereas forensic toxicology uses bile almost only for xenobiotic screening, a diagnostic test interpreting postmortem bile concentrations of meprobamate is reported. Based on 128 sets of autopsy data, its intrinsic qualities were good, with 0.95 sensitivity and 0.93 specificity. In a French forensic population, the positive and negative predictive factors were 0.90 and 0.97, respectively. It is a useful means of revealing overdoses where blood samples are not available or of confirming blood tests when postmortem redistribution is suspected.