A comprehensive sensitivity analysis of microarray breast cancer classification under feature variability

Background  

Large discrepancies in signature composition and outcome concordance have been observed between different microarray breast cancer expression profiling studies. This is often ascribed to differences in array platform as well as biological variability. We conjecture that other reasons for the observed discrepancies are the measurement error associated with each feature and the choice of preprocessing method. Microarray data are known to be subject to technical variation and the confidence intervals around individual point estimates of expression levels can be wide. Furthermore, the estimated expression values also vary depending on the selected preprocessing scheme. In microarray breast cancer classification studies, however, these two forms of feature variability are almost always ignored and hence their exact role is unclear.     

Study of the proinflammatory role of human differentiated omental adipocytes

Infiltration of monocyte‐derived macrophages into adipose tissue has been associated with tissue and systemic inflammation. It has been suggested that macrophage infiltration affects fat expansion through a paracrine action on adipocyte differentiation. Our working hypothesis is that factors released by monocytes/macrophages may also affect mature adipocyte biology. Human differentiated omental adipocytes were incubated with LPS and conditioned media obtained from human macrophage‐like cell line THP‐1, previously activated or not with LPS. We show that LPS greatly increased the secretion levels of pro‐inflammatory adipokines including IL‐6, IL‐8, GRO, and MCP‐1. Macrophage‐conditioned medium also upregulated IL‐6, IL‐8, GRO, and MCP‐1 mRNA expression and protein levels and led to the novo secretion of ICAM‐1, IL‐1β, IP‐10, MIP‐1α, MIP‐1β, VEGF, and TNFα. Human differentiated adipocytes treated by macrophage‐conditioned medium displayed marked reduction of adipocyte function as assessed by decreased phosphorylation levels of ERK1, ERK2, and p38α and reduced gene expression of lipogenic markers including PPAR‐γ and fatty acid synthase. These data show that macrophage‐secreted factors not only inhibit the formation of mature adipocytes but alter their function, suggesting that human differentiated omental adipocytes might also contribute to systemic chronic low‐grade inflammation associated with human obesity. J. Cell. Biochem. 107: 1107–1117, 2009. © 2009 Wiley‐Liss, Inc.     

Structural requirements for efficient prion protein conversion: Cofactors may promote a conversion-competent structure for PrPC

To understand why cross-species infection of prion disease often results in inefficient transmission and reduced protein conversion, most research has focused on defining the effect of variations in PrP primary structures, including sequence compatibility of substrate and seed. By contrast, little research has been aimed at investigating structural differences between different variants of PrP^C and secondary structural requirements for efficient conversion. This is despite a clear role for molecular chaperones in formation of prions in non-mammalian systems, indicating the importance of secondary/tertiary structure during the conversion process. Recent data from our laboratory on the cellular location of disease-specific prion cofactors supports the critical role of specific secondary structural motifs and the stability of these motifs in determining the efficiency of disease-specific prion protein conversion. In this paper we summarize our recent results and build on the hypothesis previously suggested by Wuthrich and colleagues, that stability of certain regions of the prion protein is crucial for protein conversion to abnormal isoforms in vivo. It is suggested that one role for molecular cofactors in the conversion process is to stabilize PrP^C structure in a form that is amenable for conversion to PrP^Sc.Key words: cofactor, structure, cell-free conversion assay, fibrillization, stability, loop region     

Fat Overload Induces Changes in Circulating Lactoferrin That Are Associated With Postprandial Lipemia and Oxidative Stress in Severely Obese Subjects

Lactoferrin is an innate immune system protein with anti‐inflammatory and antioxidant activities. We aimed to evaluate circulating lactoferrin levels in association with lipid concentrations, and parameters of oxidative stress and inflammation in subjects with morbid obesity after an acute fat intake. The effects of a 60 g fat overload on circulating lactoferrin and antioxidant activities were evaluated in 45 severely obese patients (15 men and 30 women, BMI 53.4 ± 7.2 kg/m²). The change in circulating lactoferrin after fat overload was significantly and inversely associated with the free fatty acid (FFA) change. In those subjects with the highest increase in lactoferrin (in the highest quartile), high‐density lipoprotein (HDL)‐cholesterol decreased after fat overload to a lesser extent (P = 0.03). In parallel to lipid changes, circulating lactoferrin concentrations were inversely linked to the variations in catalase (CAT) and glutathione reductase (GSH‐Rd). Baseline circulating lactoferrin concentration was also inversely associated with the absolute change in antioxidant activity after fat overload, and with the change in C‐reactive protein (CRP). Furthermore, those subjects with higher than the median value of homeostasis model assessment of insulin secretion (HOMA_IS) had significantly increased lactoferrin concentration after fat load (885 ± 262 vs. 700 ± 286 ng/ml, P = 0.03). Finally, we further explored the action of lactoferrin in vitro. Lactoferrin (10 µmol/l) led to significantly lower triglyceride (TG) concentrations and lactate dehydrogenase activity (as expression of cell viability) in the media from adipose explants obtained from severely obese subjects. In conclusion, circulating lactoferrin concentrations, both at baseline and fat‐stimulated, were inversely associated with postprandial lipemia, and parameters of oxidative stress and fat‐induced inflammation in severely obese subjects.     

The Gene Expression of the Main Lipogenic Enzymes is Downregulated in Visceral Adipose Tissue of Obese Subjects

Contradictory findings regarding the gene expression of the main lipogenic enzymes in human adipose tissue depots have been reported. In this cross‐sectional study, we aimed to evaluate the mRNA expression of fatty acid synthase (FAS) and acetyl‐CoA carboxilase (ACC) in omental and subcutaneous (SC) fat depots from subjects who varied widely in terms of body fat mass. FAS and ACC gene expression were evaluated by real time‐PCR in 188 samples of visceral adipose tissue which were obtained during elective surgical procedures in 119 women and 69 men. Decreased sex‐adjusted FAS (?59%) and ACC (?49%) mRNA were found in visceral adipose tissue from obese subjects, with and without diabetes mellitus type 2 (DM‐2), compared with lean subjects (both P < 0.0001). FAS mRNA was also decreased (?40%) in fat depots from overweight subjects (P < 0.05). Indeed, FAS mRNA was significantly and positively associated with ACC gene expression (r = 0.316, P < 0.0001) and negatively with BMI (r = ?0.274), waist circumference (r = ?0.437), systolic blood pressure (r = ?0.310), serum glucose (r = ?0.277), and fasting triglycerides (r = ?0.226), among others (all P < 0.0001). Similar associations were observed for ACC gene expression levels. In a representative subgroup of nonobese (n = 4) and obese women (n = 6), relative FAS gene expression levels significantly correlated (r = 0.657, P = 0.034; n = 10) with FAS protein values. FAS protein levels were also inversely correlated with blood glucose (r = ?0.640, P = 0.046) and fasting triglycerides (r = ?0.832, P = 0.010). In conclusion, the gene expression of the main lipogenic enzymes is downregulated in visceral adipose tissue from obese subjects.     

Environmental and Genetic Factors Influence the Relationship Between Circulating IL‐10 and Obesity Phenotypes

Interleukin‐10 (IL‐10) is a centrally operating anti‐inflammatory cytokine that plays a crucial role in the regulation of the innate immune system. It has strong inactivating properties on the inflammatory host response and has been related with viral persistence. We aimed to evaluate the association among circulating IL‐10, obesity phenotypes, IL‐10 and IL‐10R1 gene polymorphisms, and the environmental exposure to viral infection. IL‐10 ?819C/T gene promoter and IL‐10 receptor‐1 ?243A/G gene polymorphisms were studied in 760 subjects, whereas the former was also investigated in a replication study of 676 subjects. The association of circulating IL‐10 levels (enzyme‐linked immunosorbent assay) with the serum IgG against adenoviruses and enteroviruses was evaluated in a subset of 189 subjects. Circulating levels of IL‐10 were increased in obese people and were positively associated with weight, BMI, waist, waist‐to‐hip ratio, fat mass, systolic pressure, and, interestingly, the titer of adenoviruses and enteroviruses. Obese subjects with adenovirus titer over the median had the highest circulating IL‐10 concentration. Both obesity and adenovirus titer were independently associated with IL‐10 variance. Nonmorbid obese T carriers for the ?819CT IL‐10 gene polymorphism had significantly higher BMI and waist circumference, and those with normal fasting glucose had increased fasting triglycerides. G carriers for the ?536AG IL‐10R1 gene polymorphism had higher systolic and diastolic pressures, and IL‐10 levels; and obese G carriers had an increased waist‐to‐hip ratio. In summary, circulating IL‐10 levels were associated not only with obesity status but also with genetic factors and with the exposure to environmental pathogens.     

Resistance Training Improves Cardiovascular Risk Factors in Obese Women Despite a Significative Decrease in Serum Adiponectin Levels

Increased circulating adiponectin and insulin sensitivity are usually observed after body fat loss induced by a weight‐loss diet. Progressive resistance training (PRT) without a concomitant weight‐loss diet significantly decreases visceral fat, thus improving insulin sensitivity. Therefore, the purpose of this study was to ascertain the effects of combined 16‐week PRT and weight‐loss diet on circulating adiponectin and insulin sensitivity index. Thirty‐four obese (BMI: 30–40 kg/m²) women, aged 40–60 year, were randomized to three groups: a control group (C; n = 9); a diet group (WL; n = 12) with a caloric restriction of 500 kcal/d; and a diet plus resistance training group (WL+RT; n = 13) with the same caloric restriction as group WL and a 16‐week supervised whole body PRT of two sessions/week. Both WL and WL+RT groups showed similar decreases in body mass (?6.3% and ?7.7%) and visceral fat (?19.9% and ?20.5%). WL resulted in an expected increase in circulating levels of adiponectin (P = 0.07) and insulin sensitivity. However, circulating total adiponectin decreased (P < 0.05) in WL+RT group, whereas an improvement in different cardiovascular risk factors (insulin sensitivity, low‐density lipoprotein cholesterol (LDL‐C), etc.) was observed. In conclusion, in obese women a 16‐week combined PRT and weight‐loss diet is accompanied by significant improvements in different cardiovascular risk factors in spite of a significant decrease of circulating adiponectin.     

Detection of a permeability factor produced byHaemophilus pleuropneumoniae

Toxigenic profiles ofHaemophilus pleuropneumoniae were determined for isolates rom serotypes 1 and 5. None of the 18 strains investigated produced hemolytic or proteolytic activity. Intradermal injection of rabbits with culture supernatants induced an edematous zone (permeability factor, PF) for strains of either serotypes. The presence of this PF seemed to depend on the component freshness of the culture media, especially for isolates of serotype 5. PF activity appeared at two different time intervals: at 2–3 h of culture and at 12 h of culture. Isolates ofH. pleuropneumoniae from serotypes 1 and 5 can produce a PF that is not related to hemolytic no proteolytic activity.     

Macroenvironmental factors as ultimate determinants of distribution of common toad and natterjack toad in the south of Spain

We have analysed the relations between the distribution areas of Bufo bufo and Bufo calamita in the south of the Iberian Peninsula. In order to characterise the localities where the species were found, we used 24 environmental variables, and we tested their influence on the distribution of both species by nonparametric methods. By means of logistic regression we calculated the odds of finding one or other species in a locality. The environmental parameters that increase the probability of presence of each species and of both species together are related with the climatic stability and with the climatic subregions. Bufo bufo is more likely to be found in areas where the climate is more predictable, probably because in these areas it may exert its competitive superiority over B. calamita . In zones with very low climatic stability B. calamita is more likely to be found than B. bufo , probably because B. bufo lacks the ability to adapt to unpredictable conditions. In areas with intermediate climatic predictability B. bufo is present, but it would be prevented from removing B. calamita , and so the odds of finding each species are equilibrated. In these areas there would be a balance between the superior competitiveness of B. bufo and the higher adaptability of B. calamita . With the same logistic equation we characterise the typical habitats of each species, and also the environments shared by both species.