Surrogate indices of insulin resistance using the Matsuda index as reference in adult men-a computational approach.

Background: Overweight and obesity, high blood pressure, hyperglycemia, hyperlipidemia, and insulin resistance (IR) are strongly associated with non-communicable diseases (NCDs), including type 2 diabetes, cardiovascular disease, stroke, and cancer. Different surrogate indices of IR are derived and validated with the euglycemic-hyperinsulinemic clamp (EHC) test. Thus, using a computational approach to predict IR with Matsuda index as reference, this study aimed to determine the optimal cutoff value and diagnosis accuracy for surrogate indices in non-diabetic young adult men. Methods: A cross-sectional descriptive study was carried out with 93 young men (ages 18-31). Serum levels of glucose and insulin were analyzed in the fasting state and during an oral glucose tolerance test (OGTT). Additionally, clinical, biochemical, hormonal, and anthropometric characteristics and body composition (DEXA) were determined. The computational approach to evaluate the IR diagnostic accuracy and cutoff value using difference parameters was examined, as well as other statistical tools to make the output robust. Results: The highest sensitivity and specificity at the optimal cutoff value, respectively, were established for the Homeostasis model assessment of insulin resistance index (HOMA-IR) (0.91; 0.98; 3.40), the Quantitative insulin sensitivity check index (QUICKI) (0.98; 0.96; 0.33), the triglyceride-glucose (TyG)-waist circumference index (TyG-WC) (1.00; 1.00; 427.77), the TyG-body mass index (TyG-BMI) (1.00; 1.00; 132.44), TyG-waist-to-height ratio (TyG-WHtR) (0.98; 1.00; 2.48), waist-to-height ratio (WHtR) (1.00; 1.00; 0.53), waist circumference (WC) (1.00; 1.00; 92.63), body mass index (BMI) (1.00; 1.00; 28.69), total body fat percentage (TFM) (%) (1.00; 1.00; 31.07), android fat (AF) (%) (1.00; 0.98; 40.33), lipid accumulation product (LAP) (0.84; 1.00; 45.49), leptin (0.91; 1.00; 16.08), leptin/adiponectin ratio (LAR) (0.84; 1.00; 1.17), and fasting insulin (0.91; 0.98; 16.01). Conclusions: The computational approach was used to determine the diagnosis accuracy and the optimal cutoff value for IR to be used in preventive healthcare.

Irisin levels during pregnancy and changes associated with the development of preeclampsia.

CONTEXT: Irisin is a recently discovered adipomyokine that regulates the differentiation and phenotype of adipose tissue. OBJECTIVE: In this study, we investigated the levels of irisin over the three trimesters of gestation in healthy and preeclamptic women and during the follicular and luteal phase of the menstrual cycle in a cohort of healthy eumenoherric women. METHODS: Serum irisin was measured by an ELISA in a longitudinal prospective cohort study in 40 healthy pregnant women, 10 mild preeclamptic women, and 20 healthy eumenoherric women during the menstrual cycle to assess irisin levels and correlations with other metabolic parameters. We identified the protein expression of fibronectin type III domain-containing protein 5, the irisin precursor, in human placenta using immunohistochemical approaches in humans. RESULTS: Serum irisin levels are higher in the luteal than in the follicular phase in eumenorrheic women. Fibronectin type III domain-containing protein 5, the irisin precursor, is expressed in human placenta, and its serum levels are higher during the entire pregnancy when compared with nonpregnant women. Serum irisin correlates positively with the homeostasis model assessment of estimated insulin resistance in the first trimester of normal pregnancy. Serum irisin levels do not change throughout gestation in preeclamptic women; however, there were lower irisin levels during the third trimester when compared with the normal pregnant group. CONCLUSION: Our results suggest that irisin may be involved in reproductive function and in the pregnancy-associated metabolic changes, and this condition may be an irisin-resistant state during gestation.

Leptin and fasting regulate rat gastric glucose-regulated protein 58.

The stomach secretes a wide range of peptides with essential metabolic functions, and thereby plays an important role in the regulation of energy homeostasis. Disulfide isomerase glucose-regulated protein 58 (GRp58) is a molecular chaperone member of the endoplasmic reticulum (ER) stress signaling pathway, which is a marker for human gastric cancer. Since GRp58 seems to be regulated by a phosphorylation/dephosphorylation pattern shift, we used the 2DE gel methodology and peptide mass fingerprinting-protein identification by means of MALDI-TOF mass spectrometry. We show that gastric mucosa GRp58 is dephosphorylated by fasting, and this effect is blunted when fasted rats are treated with leptin. Furthermore, we assessed the gene expression of GRp58 under different physiological settings known to be associated with energy homeostasis (fasting, leptin treatment and leptin deficiency). We found that intraperitoneal administration of leptin increases whereas leptin deficiency decreases GRp58 mRNA levels. However, GRp58 expression remains unchanged after fasting, indicating that leptin actions on GRp58 are no direct sensitivity to fasting. Dissection of the molecular pathways mediating the interactions between ER stress-related factors and nutrient availability, as well as their target genes, may open a new avenue for the study of obesity and other metabolic disorders.

Medicina personalizada: farmacogenómica y farmacoepigenética: [editoriales] / Tailored medicine: pharmogenomics and pharmacogenetics: [editorial]

El proyecto genoma humano, los avances y los desarrollos de las diferentes “omicas”, apoyados en la medicina traslacional, han permitido el conocimiento y avance en muchos de los mecanismos moleculares de la fisiología y la patología en el humano. Basado enestos conocimientos se tienen muchas evidencias de que la mayor parte de la terapia con medicamentos puede ser individualizada sobre los estudios de variación del genoma humano, ya que solo entre el 30 % y 60 % de los pacientes tienen una respuesta común a dichas terapias...

Análisis proteóiico de cultivos primarios de tiroides / Proteomic nalysiis of primary cultures from thyroid / Análiseproteómicco de culturas preliminares do thyroid

En el presente estudio se identificaron proteínas de expresión constitutiva, como vimentina, actina, tubulina, proteína de choque térmico de 60 kDa, peroxirredoxina y la ATP sintasa mitocondrial, en cultivos primarios de tiroides normales y de carcinoma papilar de tiroides. Se establecieron las condiciones de extracción, solubilización, análisis cuantitativo y cualitativo de dichas proteínas, y se buscaron las mejores condiciones de isoelectroenfoque (IEF) en la electroforesis en dos dimensiones (2D). En la extracción y solubilización de las proteínas se evaluó la presencia o ausencia de anfolitos y sales, se obtuvo un mejor resultado empleando en el amortiguador de extracción sales como Tris-HCl y acetato de magnesio que incrementan la solubilidad de las proteínas. Para la cuantificación se recomienda el uso conjunto de técnicas colorimétricas con la electroforesis SDS-PAGE tiñendo con azul de Coomassie y corroborando los resultados mediante western blot, lo cual permite, además, verificar la integridad de las proteínas. Respecto a la electroforesis en dos dimensiones, se obtuvieron geles con un mayor número de manchas (spots),resueltos, enfocados y reproducibles empleando en el IEF gradientes inmovilizados de pH de 4-7 y voltaje final de 8.000 V. Las proteínas se identificaron mediante el análisis bioinformático de los geles 2D con el programa PDQuest (PDQuest 7.2, Bio-Rad®) y MALDI-TOF.

In this paper, proteins of constitutive expression such as vimentin, actin, tubulin, heat shock protein of 60 kDa, peroxiredoxin and the mitochondrial ATP sintase were identified in primary cultures of normal thyroid and papillary carcinoma. The extraction conditions, solubilization, quantitative and qualitative analysis of such proteins were established. In addition, the best conditions for isoelectrofocusing (IEF) in the two-dimensional electrophoresis (2D) were founded. In the extraction and solubilization of proteins, the presence or absence ofampholytes or salts was evaluated. The best result in the extraction was obtained using buffer salts such as Tris-HCl and magnesium acetate, which increase the solubility of these proteins. For quantification, the recommendation is to combine colorimetric techniques with SDS-PAGE electrophoresis stained with Coomasie blue and Western blot to confirm the results, which allows verifying the integrity of these proteins. In the two-dimensional electrophoresis, resolved, focused and reproducible gels with greater number of spots were obtained using in the IEF immobilized pH gradients of 4-7 and final voltage of 8000 V. The proteins were identified by the bioinformatic analysis of the 2DE gels with the PDQuest program (PDQuest 7.2 (BioRad®) and MALDI-TOF.

Neste estudo, se identificaram proteínas da expressão constitutiva, como vimentina, actina, tubulina, proteína de choque do calor do kDa 60, peroxiredoxinaeasintase mitocondrial do ATP, em cultivos primários de tireóide normal e de carcinoma papilar de tireóide. Se estabeleceram as condições de extração e de solubilization, assim como a análise quantitativaeaqualitativa de tais proteínas, e procuraram-se asmelhores condições para isoeletroenfoque (IEF) na eletroforese bidimensional (2D). Na extração e no solubilization das proteínas, se avaliou a presença ou a ausência de anfolitos ou de sais. O melhor resultado foi obtido usando sais como Tris-HCl e acetato do magnésio no amortecedor da extração, as quais aumentam a solubilida-de das proteínas. Na quantificação, se recomenda o uso de técnicas colorimétricas com eletroforese SDS-PAGE, tingindo com azul de Coomasie. Os resultados se confirmam com Western blot, o qual permite também verificar a integridade das proteínas. Quanto à eletroforese bidimensional, se obtiveram géis com um maior número de pontos (spots) determinados, enfocados e reprodutíveis no IEF usando gradientes imobilizados de pH 4-7 e tensão final de 8000 V. As proteínas foram identificadas por análise bioinformática de géis 2D com o programa PDQuest (PDQuest 7.2 (Bio-Rad®) e MALDITOF.

Vaspin and amylin are expressed in human and rat placenta and regulated by nutritional status.

Amylin (islet amyloid polypeptide) and vaspin (visceral adipose tissue specific serpin) are gut and adipocyte hormones involved in the regulation of body weight homeostasis. The aim of this study was to examine whether amylin and vaspin are expressed in human and rat placenta, as well as their regulation by nutritional status. Our results demonstrate that amylin and vaspin are localized in both human and rat placenta. In the rat term placenta vaspin was demonstrated in the trophoblast of the fetal villi, the labyrinth. Vaspin immunostaining in human placenta was localized in cytotrophoblast and syncytiotrophoblast in the first trimester placentas while in the third trimester vaspin was localized in the syncytiotrophoblast. Regarding amylin, rat placenta of 16 days of gestational age showed an intense immunostaining, mainly localized in the labyrinth. On the other hand, in the human third trimester placenta amylin immunoreactivity was intense in the syncytiotrophoblast of the chorionic villi and in decidual cells. Furthermore, placental amylin and vaspin showed an opposite pattern of expression during pregnancy, with vaspin showing the highest expression level at the end and amylin at the beginning of pregnancy. Finally, food restriction also has contrary effects on their expression, increasing vaspin but decreasing amylin placental mRNA and protein levels. Taken together, our results demonstrate that vaspin and amylin are modulated by energy status in the placenta, which suggests that these proteins may be involved in the regulation of placental metabolic functions.

Enfermedad asociada a Clostridium difficile: nuevas amenazas de un viejo enemigo / Clostridium difficile associated disease: New threats of and old foe

La infección asociada a Clostridium difficile (EACD) es actualmente un importante problema de salud en el mundo con aumentada morbilidad y mortalidad en los adultos mayores. La EACD cada vez se está informando con mayor frecuencia en muchas partes del mundo. Desde 2002, se han informado brotes con una cepa de C. difficile resistente a fluoroquinolonas (BI/NAP1/027). Los cambios en la epidemiología incluyen la emergencia de una nueva población en riesgo, así como mayor incidencia de la enfermedad. La enfermedad se ha asociado con el uso de antibióticos, especialmente los de amplio espectro, por lo cual, éstos deben ser evitados. El tratamiento médico es con metronidazol o vancomicina y los casos severos pueden necesitar resección intestinal. Los casos recurrentes necesitan tratamiento prolongado con antibióticos, inmunoglobulinas y ocasionalmente enemas con materia fecal. Esta revisión es una actualización sobre la epidemiología, patogénesis y tratamiento de la infección por C. difficile.

Molecular Diagnostics of Porcine Stress Syndrome Susceptibility Associated with the Arg615Cys Mutation Using Real-Time PCR with Fluorescent Hybridization Probes / Molecular Diagnostics of Porcine Stress Syndrome Susceptibility Associated with the Arg615Cys Mutation Using Real-Time PCR with Fluorescent Hybridization Probes

Objective : The purpose of the presera study was to devélop a molecular genotyping method test by using a real time PCR hybridization probé and applying it to the analysis of C1843T mutations of the Sus scrofa RYR1 gene. Animáis population Three PSS-susceptible and PSS non-susceptible crossbred swine races were used for the experiments: Pietrain X Landrace Belga, Pietrain X Large White and Pietrain X Duroc. Methods: We have devéloped a genotyping method by using a hybridization probé and applied it to the analysis of C1843T mutations of the RYR1 gene, associated with PSS susceptibility. Genotyping results obtained by hybridization probé strategies were confirmed by restriction analysis and sequencing. In addi-tion, phenotype/genotype correlation analyses were devéloped by using the in vitro contracture test and confirmed the in vivo hálothane-succinylcholine challenge. Results: The real-time PCR with fluorescent hybridization probé methodology was designed to identify ho-mozygous PSS-resistant, PSS-susceptible animáis as well as heterozygous carriers. All cases genotyped by fluorescent hybridization probes were in agreement with PCR restriction enzyme digestión and sequencing and showed a 100% concordance between the in vivo and in vitro porcine stress syndrome (PSS) susceptibility results. Conclusions and clinical relevance: The real-time PCR with fluorescent hybridization probé method described here provides a rapid, easily interpretable and réliáble tool for genotyping the C1843T (Arg615-Cys) polymorphism of the RYR1 gene. This new methodology may be useful in the wide-scale genotyping of PSS-susceptibility and genetic selection.

Novel expression of resistin in rat testis: functional role and regulation by nutritional status and hormonal factors.

Resistin, a recently cloned adipose-secreted factor, is primarily involved in the modulation of insulin sensitivity and adipocyte differentiation. However, additional metabolic or endocrine functions of this molecule remain largely unexplored. In this study, a series of experiments were undertaken to explore the potential expression, regulation and functional role of this novel adipocytokine in rat testis. Resistin gene expression was demonstrated in rat testis throughout postnatal development, with maximum mRNA levels in adult specimens. At this age, resistin peptide was immunodetected in interstitial Leydig cells and Sertoli cells within seminiferous tubules. Testicular expression of resistin was under hormonal regulation of pituitary gonadotropins and showed stage-specificity, with peak expression values at stages II-VI of the seminiferous epithelial cycle. In addition, testicular resistin mRNA was down-regulated by the selective agonist of PPARgamma, rosiglitazone, in vivo and in vitro. Similarly, fasting and central administration of the adipocyte-derived factor, leptin, evoked a significant reduction in testicular resistin mRNA levels, whereas they remained unaltered in a model of diet-induced obesity. From a functional standpoint, resistin, in a dose-dependent manner, significantly increased both basal and choriogonadotropin-stimulated testosterone secretion in vitro. Overall, our present results provide the first evidence for the expression, regulation and functional role of resistin in rat testis. These data underscore a reproductive facet of this recently cloned molecule, which may operate as a novel endocrine integrator linking energy homeostasis and reproduction.

Effect of food restriction on ghrelin in normal-cycling female rats and in pregnancy.

OBJECTIVE: Ghrelin is a 28-amino-acid acylated peptide that was recently identified as the endogenous ligand for the growth hormone secretagogue receptor. Previous studies have shown that ghrelin potently increases growth hormone release and food intake. The aim of this study was to clarify the physiological implications of ghrelin in the regulation of energy balance, by assessing the effect of undernutrition throughout 21 days in normal-cycling and pregnant rats on ghrelin. RESEARCH METHODS AND PROCEDURES: We have determined ghrelin levels by radioimmunoassay and gastric ghrelin mRNA expression by Northern blot analysis during 21 days of chronic food restriction (30% of ad libitum available diet) in normal-cycling female rats and in pregnancy. RESULTS: Our results show that chronic food restriction led to an increase in plasmatic ghrelin levels in normal-cycling female rats. In pregnancy, ghrelin plasmatic levels were enhanced particularly during the latter part of gestation (19 and 21 days) compared with pregnant rats with free access to food. Gastric ghrelin mRNA expression showed a similar expression pattern, being higher in the food-restricted group than in the group fed ad libitum, in normal-cycling as well as in pregnant rats. DISCUSSION: These observations indicate that ghrelin plasmatic levels and ghrelin gastric mRNA are up-modulated during undernutrition in normal-cycling rats and in pregnancy. These findings suggest that increased ghrelin levels may have a role in mediating the physiological responses to undernutrition and could represent an adaptative response to prevent long-lasting alterations in energy balance and body weight homeostasis.