Blood gene expression reveal pathway differences between diet-sensitive and resistant obese subjects prior to caloric restriction.

Weight loss in response to caloric restriction displays significant interindividual heterogeneity. To develop early predictors of weight-loss success, we have compared whole-blood gene expression profiles of obese, diet-sensitive vs. obese, diet-resistant subjects prior to the initiation of clinically supervised caloric restriction. Pathway enrichment analysis of gene expression profiles by multiple applications converged on the "oxidative phosphorylation" (OXPHOS) pathway, and to a lesser extent the "proteasome" pathway, as statistically significantly upregulated in obese, diet-sensitive subjects compared to the diet-resistant subjects. The finding of increased OXPHOS is consistent with earlier observations of increased proton leak, increased expression of OXPHOS genes, and increased oxidative muscle fibers in skeletal muscle of obese, diet-sensitive subjects. The current study further highlights the utility of blood as a sentinel tissue reflecting systemic states and provides a potential modality to predict future weight-loss success, relevant to the design of individualized bariatric treatment programs.

Gene expression profiling in whole blood identifies distinct biological pathways associated with obesity.

BACKGROUND: Obesity is reaching epidemic proportions and represents a significant risk factor for cardiovascular disease, diabetes, and cancer. METHODS: To explore the relationship between increased body mass and gene expression in blood, we conducted whole-genome expression profiling of whole blood from seventeen obese and seventeen well matched lean subjects. Gene expression data was analyzed at the individual gene and pathway level and a preliminary assessment of the predictive value of blood gene expression profiles in obesity was carried out. RESULTS: Principal components analysis of whole-blood gene expression data from obese and lean subjects led to efficient separation of the two cohorts. Pathway analysis by gene-set enrichment demonstrated increased transcript levels for genes belonging to the "ribosome", "apoptosis" and "oxidative phosphorylation" pathways in the obese cohort, consistent with an altered metabolic state including increased protein synthesis, enhanced cell death from proinflammatory or lipotoxic stimuli, and increased energy demands. A subset of pathway-specific genes acted as efficient predictors of obese or lean class membership when used in Naive Bayes or logistic regression based classifiers. CONCLUSION: This study provides a comprehensive characterization of the whole blood transcriptome in obesity and demonstrates that the investigation of gene expression profiles from whole blood can inform and illustrate the biological processes related to regulation of body mass. Additionally, the ability of pathway-related gene expression to predict class membership suggests the feasibility of a similar approach for identifying clinically useful blood-based predictors of weight loss success following dietary or surgical interventions.

Distinct skeletal muscle fiber characteristics and gene expression in diet-sensitive versus diet-resistant obesity.

Inter-individual variability in weight gain and loss under energy surfeit and deficit conditions, respectively, are well recognized but poorly understood phenomena. We documented weight loss variability in an intensively supervised clinical weight loss program and assessed skeletal muscle gene expression and phenotypic characteristics related to variable response to a 900 kcal regimen. Matched pairs of healthy, diet-compliant, obese diet-sensitive (ODS) and diet-resistant (ODR) subjects were defined as those in the highest and lowest quintiles for weight loss rate. Physical activity energy expenditure was minimal and comparable. Following program completion and weight stabilization, skeletal muscle biopsies were obtained. Gene expression analysis of rectus femoris and vastus lateralis indicated upregulation of genes and gene sets involved in oxidative phosphorylation and glucose and fatty acid metabolism in ODS compared with ODR. In vastus lateralis, there was a higher proportion of oxidative (type I) fibers in ODS compared with ODR women and lean controls, fiber hypertrophy in ODS compared with ODR women and lean controls, and lower succinate dehydrogenase in oxidative and oxidative-glycolytic fibers in all obese compared with lean subjects. Intramuscular lipid content was generally higher in obese versus lean, and specifically higher in ODS vs. lean women. Altogether, our findings demonstrate differences in muscle gene expression and fiber composition related to clinical weight loss success.

ERRgamma regulates cardiac, gastric, and renal potassium homeostasis.

Energy production by oxidative metabolism in kidney, stomach, and heart, is primarily expended in establishing ion gradients to drive renal electrolyte homeostasis, gastric acid secretion, and cardiac muscle contraction, respectively. In addition to orchestrating transcriptional control of oxidative metabolism, the orphan nuclear receptor, estrogen-related receptor gamma (ERRgamma), coordinates expression of genes central to ion homeostasis in oxidative tissues. Renal, gastric, and cardiac tissues subjected to genomic analysis of expression in perinatal ERRgamma null mice revealed a characteristic dysregulation of genes involved in transport processes, exemplified by the voltage-gated potassium channel, Kcne2. Consistently, ERRgamma null animals die during the first 72 h of life with elevated serum potassium, reductions in key gastric acid production markers, and cardiac arrhythmia with prolonged QT intervals. In addition, we find altered expression of several genes associated with hypertension in ERRgamma null mice. These findings suggest a potential role for genetic polymorphisms at the ERRgamma locus and ERRgamma modulators in the etiology and treatment of renal, gastric, and cardiac dysfunction.

Peroxisome proliferator-activated receptor gamma 2 and acyl-CoA synthetase 5 polymorphisms influence diet response.

Peroxisome proliferator-activated receptor gamma (PPARgamma) and its response gene, Acyl CoA synthetase 5 (ACSL5), which has an important role in fatty acid metabolism, may affect weight loss in response to caloric restriction. Therefore, we aimed to determine whether these genes were involved in the interindividual response to dietary treatment. Genotypic/phenotypic comparisons were made between selected obese women from the quintiles losing the most (diet responsive, n = 74) and the quintiles losing the least (diet-resistant, n = 67) weight in the first 6 weeks of a 900-kcal formula diet. Two common PPARgamma single nucleotide polymorphisms, Pro(12)Ala and C1431T, and eight polymorphisms across the ACSL5 gene were selected for single locus and haplotypic association analyses. The PPARgamma Pro(12)Ala single nucleotide polymorphism was associated with diet resistance (odds ratio = 3.48, 95% confidence interval = 1.41 to 8.56, p = 0.03), and the rs2419621, located in the 5'untranslated region of the ACSL5 gene, displayed the strongest association with diet response (odds ratio = 3.45, 95% confidence interval = 1.61 to 7.69, p = 0.001). Skeletal muscle ACSL5 mRNA expression was significantly lower in carriers of the wildtype compared with the variant rs2419621 allele (p = 0.03). Our results suggest a link between PPARgamma2 and ACSL5 genotype and diet responsiveness.

Independently-addressable micron-sized biosensor elements.

With the continuing development of micro-total analysis systems and sensitive biosensing technologies, it is often desirable to immobilize biomolecules onto a surface in a small well-defined area. A novel method was developed to electrochemically attach DNA probes to micron-sized regions of a gold surface using biotin-LC-hydrazide (BH). Previously, we have found that the radical produced during the oxidation of BH will attach to a wide variety of electroactive surfaces. An array of micron-sized gold band electrodes (75 microm wide) was fabricated onto glass microscope slides and BH was deposited onto each electrode through the application of an oxidizing potential. Subsequent attachment of avidin to the biotinylated surface created the 'molecular sandwich' architecture necessary for further immobilization of biotinylated biomolecules to the surface. In this work, we utilized biotinylated DNA probes of varying sequence to illustrate the specificity of the attachment scheme. The immobilization of avidin, DNA probe, and hybridization of DNA target is visualized with fluorescence tags and the spatially selective attachment and hybridization of unique DNA sequences is demonstrated.

Barriers and influences in seeking health care among lower income minority women.

The purpose of this study was to identify and describe perceived barriers to seeking health care, determine perceptions of confidence in health care practitioners, and explore strategies to enhance, promote, and improve early health care intervention among low income minority women. Focus group methodology was used to collect data and content analysis was used to analyze the data. Results revealed four broad categories for discussion: (a) confidence in the physician, (b) frequency of engaging in screening procedures, (c) barriers and influences in seeking health care, and (d) a wish list for covered health care services. The study underscored the importance of both spirituality and family in the lives of aging minority women. The paper includes implications for public policy and suggests an agenda for public policy advocates in the new millennium.

Tissue expression profiling using real-time PCR.

The recent development of real-time PCR allows for the rapid and accurate quantitation of gene expression in cells and tissues. Real-time PCR instrumentation is designed for the simultaneous quantitation of gene expression from a few samples up to 384 samples. The normal tissue expression profile of a gene can provide useful insights into its potential role in normal physiological processes. When combined with the tissue expression profile of the gene in diseased tissues, information concerning the potential role in pathological processes can be determined. This unit describes a protocol to determine the relative abundance of mRNA in a panel of human tissues using real-time PCR.

Effects of Discrete Protein-Surface Interactions in Scanning Force Microscopy Adhesion Force Measurements.

The potential for measuring specific molecular recognition forces between probe-bound ligands and surface-bound proteins using a scanning force microscope (SFM) has recently gained much attention. Generally, observed discontinuities in the SFM force-displacement curves are attributed to the breaking of discrete, specific affinity bonds. The present study on the molecular recognition system composed of surface-immobilized antifluorescyl IgG molecules and SFM probe-bound fluorescein ligands has demonstrated that similar intermittent discontinuities in the SFM force-displacement curves may in fact be largely due to nonspecific discrete interactions between the protein and the SFM probe. The mechanical behavior of the cantilever-spherical bead system used in this study is discussed, as it appears to cause a false indication of the separation distance between the surface and probe. The strong lateral interactions which result in "stick and slip"-like discontinuities seen in the adhesion curves are likely the result of localized adhesion due to the heterogeneous nature of proteins and the lack of molecular mobility allowed in the experimental system. The effect is magnified with increasing contact time between the protein and probe. Factors which may cause such anomalous behavior in a specific ligand-protein system are discussed in order to avoid misinterpretation of SFM adhesion measurements.