Type-2 diabetes (T2D)-associated SNPs are more likely to be eQTLs. The allelic expression imbalance (AEI) analysis is the measure of relative expression between two allelic transcripts and is the most sensitive measurement to detect cis-regulatory effects. We performed AEI screening to detect cis-regulators for genes expressed in transformed lymphocytes of 190 Caucasian (CA) and African American (AA) subjects to identify functional variants for T2D susceptibility in the chromosome1q21-24 region of linkage. Among transcribed SNPs studied in 115 genes, significant AEI (p<0.001) occurred in 28 and 30 genes in CA and AA subjects, respectively. Analysis of the effect of selected AEI-SNPs (≥10% mean AEI) on total gene expression further established the cis-eQTLs in THEM4 (rs13320, p=0.027), and IGSF8 (rs1131891, p=0.02). Examination of published genome-wide association data identified significant associations (p<0.01) of three AEI-SNPs with T2D in the DIAGRAM-v3 dataset. Six AEI-SNPs, including rs13320 (p=1.35E-04) in THEM4, were associated with glucose homeostasis traits in the MAGIC dataset. Evaluation of AEI-SNPs for association with glucose homeostasis traits in 611 nondiabetic subjects showed lower AIRG (p=0.005) in those with TT/TC-genotype for rs13320. THEM4 expression in adipose was higher (p=0.005) in subjects carrying the T-allele; in vitro analysis with luciferase construct confirmed the higher expression of the T-allele. Resequencing of THEM4 exons in 192 CA subjects revealed four coding non-synonymous variants, but did not explain transmission of T2D in 718 subjects from 67 Caucasian pedigrees. Our study indicates the role of a cis-regulatory SNP in THEM4 that may influence T2D predisposition by modulating glucose homeostasis.

The spontaneously hypertensive rat (SHR) has been widely used as a model for studies of hypertension and attention deficit/hyperactivity disorder (ADHD). The inbred Wistar-Kyoto (WKY) rat, derived from the same ancestral outbred Wistar rat as the SHR, are normotensive and have been used as the closest genetic control for the SHR, although the WKY has also been used as a model for depression. Notably, however, substantial behavioral and genetic differences among the WKY substrains, usually from the different vendors and breeders, have been observed. These differences have often been overlooked in prior studies, leading to inconsistent and even contradictory findings. The complicated breeding history of the SHR and WKY rats and the lack of a comprehensive understanding of the genetic background of different commercial substrains make the selection of control rats a daunting task, even for researchers who are mindful of their genetic heterogeneity. In this study, we examined the genetic relationship of 16 commonly used WKY and SHR rat substrains using genome-wide SNP genotyping data. Our results confirmed a large genetic divergence and complicated relationships among the SHR and WKY substrains. This understanding, although incomplete without the genome sequence, provides useful guidance in selecting substrains and helps to interpret previous reports when the source of the animals was known. Moreover, we found two closely related, yet distinct WKY substrains that may provide novel opportunities in modeling psychiatric disorders.

Fetuses respond to transient hypoxia (a common stressor in utero) with cellular responses that are appropriate for promoting survival of the fetus. The present experiment was performed to identify the acute genomic responses of the fetal hypothalamus to transient hypoxia. Three fetal sheep were exposed to 30 min of hypoxia and hypothalamic mRNA extracted from samples collected 30 min after return to normoxia. These samples were compared to those from 4 normoxic control fetuses using the Agilent 019921 ovine array. Differentially-regulated genes were analyzed by network analysis and by gene ontology analysis, identifying statistically significant overrepresentation of biological processes. Real-time PCR of selected genes supported the validity of the array data. Hypoxia induced increased expression of genes involved in response to oxygen stimulus, RNA splicing, anti-apoptosis, vascular smooth muscle proliferation, and positive regulation of Notch receptor target. Downregulated genes were involved in metabolism, antigen receptor-mediated immunity, macromolecular complex assembly, S-phase, translation elongation, RNA splicing, protein transport, and post-transcriptional regulation. We conclude that these results emphasize that the cellular response to hypoxia involves reduced metabolism, the involvement of the fetal immune system, and the importance of glucocorticoid signaling.

Large datasets from -omics studies need to be deeply investigated. The aim of this paper is to provide a new method (LEM method) for the search of transcriptome and metabolome connections. The heuristic algorithm here described extends the classical canonical correlation analysis (CCA) to a high number of variables (without regularization) and combines well-conditioning and fast-computing in 'R'. Reduced CCA models are summarized in PageRank matrices, the product of which gives a stochastic matrix that resumes the self-avoiding walk covered by the algorithm. Then, a homogeneous Markov process applied to this stochastic matrix converges the probabilities of interconnection between genes, providing a selection of disjointed subsets of genes. This is an alternative to regularized generalized CCA for the determination of blocks within the structure matrix. Each gene subset is thus linked to the whole metabolic or clinical dataset that represents the biological phenotype of interest. Moreover, this selection process reaches the aim of biologists who often need small sets of genes for further validation or extended phenotyping. The algorithm is shown to work efficiently on three published datasets, resulting in meaningfully broadened gene networks.

Using genomic microarray analysis, our objective was to identify and annotate differences in the pre-training skeletal muscle transcriptomes among human subjects clustered as non-responders (Non), modest responders (Mod), and extreme responders (Xtr) based on differential magnitudes of myofiber hypertrophy in response to progressive resistance training (RT) (Non -16μm(2), Mod 1,111μm(2), or Xtr 2,475μm(2)). In prior work, we noted differences among clusters in the prevalence of myogenic stem cells prior to and during RT (35), and in the translational signaling responses to the first bout of resistance exercise (30). Here we identified remarkable differences in the pre-training transcript profiles among clusters (8026 genes differentially expressed between Xtr and Non; 2463 between Xtr and Mod; and 1294 between Mod and Non). Annotated functions and networks of differentially expressed genes suggest Xtr were "primed" to respond to RT through transcriptional regulation, along with a uniquely expressed network of genes involved in skeletal muscle development, while the failed response in Non may have been driven by excessive pro-inflammatory signaling. Protein follow-up analysis revealed higher basal levels of acetylated histone H3 (K36) in the two responder clusters (Mod, Xtr) compared to Non, and only the responders experienced alterations in the muscle content of select proteins (e.g., α-tubulin, p27(kip)) in response to the first resistance exercise stimulus. Overall, the widely disparate transcriptomes identified prior to RT among the three clusters support the notion that at least some of the inter-individual heterogeneity in propensity for RT-induced myofiber hypertrophy is likely pre-determined.

Temperature is one of the most prominent abiotic factors affecting ectotherms. Most fish species, as ectotherms, have extraordinary ability to deal with a wide range of temperature changes. While the molecular mechanism underlying temperature adaptation has long been of interest, it is still largely unexplored with fish. Understanding of the fundamental mechanisms conferring tolerance to temperature fluctuations is a topic of increasing interest as temperature may continue to rise as a result of global climate change. Catfish have a wide natural habitat and possess great plasticity in dealing with environmental variations in temperature. However, no studies have been conducted at the transcriptomic level to determine heat stress induced gene expression. In the present study, we conducted a RNA-Seq analysis to identify heat stress induced genes in catfish at the transcriptome level. Expression analysis identified a total of 2,260 differentially expressed genes with a cutoff of two-fold change. qRT-PCR validation suggested the high reliability of the RNA-Seq results. Gene ontology, enrichment and pathway analyses were conducted to gain insight into physiological and gene pathways. Specifically, genes involved in oxygen transport, protein folding and degradation, and metabolic process were highly induced while general protein synthesis was dramatically repressed in response to the lethal temperature stress. This is the first RNA-Seq-based expression study in catfish in response to heat stress. The candidate genes identified should be valuable for further targeted studies on heat tolerance, thereby assisting the development of heat-tolerant catfish lines for aquaculture.

Right ventricular failure (RVF) is the most frequent cause of death in patients with pulmonary arterial hypertension (PAH); however, therapies targeted to treat RVF are practically non-existent. Chronic treatment with carvedilol has been shown to reduce established maladaptive right ventricle (RV) hypertrophy and to improve RV function in experimental PAH. However, the mechanisms by which carvedilol improves RVF are still unknown. By microarray studies we have previously demonstrated that RVF is characterized by a distinct gene expression profile when compared with functional, compensatory hypertrophy. We next sought to identify the effects of carvedilol on gene expression in a genome-wide basis. PAH and RVF were induced in male Sprague-Dawley rats by the combination of VEGF-receptor blockade and chronic hypoxia. After RVF was established, rats were treated with carvedilol or vehicle for four weeks. RNA was isolated from RV tissue and hybridized for microarray analysis. An initial prediction analysis of carvedilol-treated RVs showed that the gene expression profile mostly resembled the RVF prediction set. However, a more extensive analysis revealed a small group of genes differentially expressed after carvedilol treatment. Further analysis categorized these genes in pathways involving cardiac hypertrophy, mitochondrial dysfunction, protein ubiquitination, and sphingolipid metabolism. Genes encoding proteins in the cardiac hypertrophy and protein ubiquitination pathways were downregulated in the RV by carvedilol, while genes encoding proteins in the mitochondrial dysfunction and sphingolipid metabolism pathways were upregulated by carvedilol. These gene expression changes may explain some of the mechanisms that underlie the functional improvement of the RV after carvedilol treatment.

It is well recognized that there is sex-dimorphic expression of mRNA and protein in the heart; however the underlying mechanism is poorly understood. Endothelial nitric oxide synthase (eNOS) is an important regulator of cardiac function, and the expression levels of eNOS differ between male and female hearts. The aim of this study was to examine whether expression of specific microRNA (miRNA) in males and females contributes to changes in the expression of eNOS. miRNA was extracted from the myocardium of male and female C57BL/6 mice and subjected to an Affymetrix miRNA array. Decreased expression of miR-222 was discovered in females and confirmed by qRT-PCR from whole heart or isolated cardiomyocytes. The transcription factor, V-ets erythroblastosis virus E26 oncogene homolog-1 (ets-1), was identified as a potential target of miR-222 using TargetScan, and five-fold increased ets-1 protein expression in females was confirmed by western blot. Targeting of ets-1 by miR-222 was determined in HEK293 cells overexpressing luciferase under regulation of either the ets-1 3'UTR, a null 3'UTR control or a scrambled ets-1 3'UTR and treated with a small molecule miR-222 mimic or inhibitor. Additionally qRT-PCR confirmed that mRNA levels of the ets-1 transcriptional target, endothelial nitric oxide synthase (eNOS), were 25% higher in females. Compared to untreated myocyte controls, 50% inhibition of eNOS expression was achieved by treatment with a miR-222 mimic, compared to a 25% increase due to miR-222 inhibitor. Our findings indicate that sex-dependent miR-222 regulation alters the expression of the cardiac regulatory protein eNOS.

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The bradykinin receptor B2 (BDKRB2) gene +9/-9 polymorphism has been associated with higher gene transcriptional activity, and characteristics of cardiovascular phenotypes and physical performance. We hypothesized that vasodilation and ACE activity response to exercise training is modulated by BDKRB2 gene. Seventy one healthy volunteers were genotyped for the BDKRB2 gene polymorphism. Heart rate (HR), mean blood pressure (MBP), and forearm blood flow (FBF) were evaluated. Angiotensin-I converting enzyme (ACE) activity was measured by fluorescence. Aerobic training was performed for 16 weeks. All variables were re-assessed following completion of the training period. In pre-training period, HR, MBP, FBF and forearm vascular conductance (FVC) were similar among all genotypes. After physical training, the FBF and the FVC response during handgrip exercise as area under the curve were higher in -9/-9 carriers, than the other two groups. However, there were no changes on HR and MBP for all three groups. In addition, in post-training period the decrease in ACE activity was higher in the -9/-9 group, than the other two groups. These results suggest that reflex muscle vasodilation and ACE activity in response to exercise training are modulated by BDKRB2 gene +9/-9 polymorphism in healthy individuals.