Allele-specific disparity in breast cancer

Background

Molecular alterations critical to development of cancer include mutations, copy number alterations (amplifications and deletions) as well as genomic rearrangements resulting in gene fusions. Massively parallel next generation sequencing, which enables the discovery of such changes, uses considerable quantities of genomic DNA (> 5 ug), a serious limitation in ever smaller clinical samples. However, a commonly available microarray platforms such as array comparative genomic hybridization (array CGH) allows the characterization of gene copy number at a single gene resolution using much smaller amounts of genomic DNA. In this study we evaluate the sensitivity of ultra-dense array CGH platforms developed by Agilent, especially that of the 1 million probe array (1 M array), and their application when whole genome amplification is required because of limited sample quantities.

Methods

We performed array CGH on whole genome amplified and not amplified genomic DNA from MCF-7 breast cancer cells, using 244 K and 1 M Agilent arrays. The ADM-2 algorithm was used to identify micro-copy number alterations that measured less than 1 Mb in genomic length.

Results

DNA from MCF-7 breast cancer cells was analyzed for micro-copy number alterations, defined as measuring less than 1 Mb in genomic length. The 4-fold extra resolution of the 1 M array platform relative to the less dense 244 K array platform, led to the improved detection of copy number variations (CNVs) and micro-CNAs. The identification of intra-genic breakpoints in areas of DNA copy number gain signaled the possible presence of gene fusion events. However, the ultra-dense platforms, especially the densest 1 M array, detect artifacts inherent to whole genome amplification and should be used only with non-amplified DNA samples.

Conclusions

This is a first report using 1 M array CGH for the discovery of cancer genes and biomarkers. We show the remarkable capacity of this technology to discover CNVs, micro-copy number alterations and even gene fusions. However, these platforms require excellent genomic DNA quality and do not tolerate relatively small imperfections related to the whole genome amplification.     

Silencing of renal DNaseI in murine lupus nephritis imposes exposure of large chromatin fragments and activation of Toll like receptors and the Clec4e

Recent studies demonstrate that transformation of mild lupus nephritis into end-stage disease is imposed by silencing of renal DNaseI gene expression in (NZBxNZW)F1 mice. Down-regulation of DNaseI results in reduced chromatin fragmentation, and in deposition of extracellular chromatin-IgG complexes in glomerular basement membranes in individuals that produce IgG anti-chromatin antibodies. The main focus of the present study is to describe the biological consequences of renal DNaseI shut-down and reduced chromatin fragmentation with a particular focus on whether exposed large chromatin fragments activate Toll like receptors and the necrosis-related Clec4e receptor in murine and human lupus nephritis. Furthermore, analyses where performed to determine if matrix metalloproteases are up-regulated as a consequence of chromatin-mediated Toll like receptors/Clec4e stimulation. Mouse and human mRNA expression levels of DNaseI, Toll like receptors 7-9, Clec4e, pro-inflammatory cytokines and MMP2/MMP9 were determined and compared with in situ protein expression profiles and clinical data. We demonstrate that exposure of chromatin significantly up-regulate Toll like receptors and Clec4e in mice, and also but less pronounced in patients with lupus nephritis treated with immunosuppresants. In conclusion, silencing of renal DNaseI gene expression initiates a cascade of inflammatory signals leading to progression of both murine and human lupus nephritis. Principal component analyses biplot of data from murine and human lupus nephrits demonstrate the importance of DNaseI gene shut down for progression of the organ disease.     

Beneficial effects on arterial stiffness and pulse-wave reflection of combined enalapril and candesartan in chronic kidney disease - a randomized trial

Background

Cardiovascular disease (CVD) is highly prevalent in patients with chronic kidney disease (CKD). Inhibition of the renin-angiotensinsystem (RAS) in hypertension causes differential effects on central and brachial blood pressure (BP), which has been translated into improved outcome. The objective was to examine if a more complete inhibition of RAS by combining an angiotensin converting enzyme inhibitor (ACEI) and an angiotensin receptor antagonist (ARB) compared to monotherapy has an additive effect on central BP and pulse-wave velocity (PWV), which are known markers of CVD.

Methods

Sixty-seven CKD patients (mean GFR 30, range 13–59 ml/min/1.73 m²) participated in an open randomized study of 16 weeks of monotherapy with either enalapril or candesartan followed by 8 weeks of dual blockade aiming at a total dose of 16 mg candesartan and 20 mg enalapril o.d. Pulse-wave measurements were performed at week 0, 8, 16 and 24 by the SphygmoCor device.

Results

Significant additive BP independent reductions were found after dual blockade in aortic PWV (−0.3 m/s, P<0.05) and in augmentation index (−2%, P<0.01) compared to monotherapy. Furthermore pulse pressure amplification was improved (P<0.05) and central systolic BP reduced (−6 mmHg, P<0.01).

Conclusions

Dual blockade of the RAS resulted in an additive BP independent reduction in pulse-wave reflection and arterial stiffness compared to monotherapy in CKD patients.

Trial Registration

Clinical trial.gov {"type":"clinical-trial","attrs":{"text":"NCT00235287","term_id":"NCT00235287"}}NCT00235287 http://www.clinicaltrials.gov/ct2/show/NCT00235287?term=ras+block&rank=1     

Ablation of steroid receptor coactivator-3 resembles the human CACT metabolic myopathy

Oxidation of lipid substrates is essential for survival in fasting and other catabolic conditions, sparing glucose for the brain and other glucose-dependent tissues. Here we show Steroid Receptor Coactivator-3 (SRC-3) plays a central role in long chain fatty acid metabolism by directly regulating carnitine/acyl-carnitine translocase (CACT) gene expression. Genetic deficiency of CACT in humans is accompanied by a constellation of metabolic and toxicity phenotypes including hypoketonemia, hypoglycemia, hyperammonemia, and impaired neurologic, cardiac and skeletal muscle performance, each of which is apparent in mice lacking SRC-3 expression. Consistent with human cases of CACT deficiency, dietary rescue with short chain fatty acids drastically attenuates the clinical hallmarks of the disease in mice devoid of SRC-3. Collectively, our results position SRC-3 as a key regulator of β-oxidation. Moreover, these findings allow us to consider platform coactivators such as the SRCs as potential contributors to syndromes such as CACT deficiency, previously considered as monogenic.     

The Brain’s Dorsal Route for Speech Represents Word Meaning: Evidence from Gesture

The dual-route model of speech processing includes a dorsal stream that maps auditory to motor features at the sublexical level rather than at the lexico-semantic level. However, the literature on gesture is an invitation to revise this model because it suggests that the premotor cortex of the dorsal route is a major site of lexico-semantic interaction. Here we investigated lexico-semantic mapping using word-gesture pairs that were either congruent or incongruent. Using fMRI-adaptation in 28 subjects, we found that temporo-parietal and premotor activity during auditory processing of single action words was modulated by the prior audiovisual context in which the words had been repeated. The BOLD signal was suppressed following repetition of the auditory word alone, and further suppressed following repetition of the word accompanied by a congruent gesture (e.g. [“grasp” + grasping gesture]). Conversely, repetition suppression was not observed when the same action word was accompanied by an incongruent gesture (e.g. [“grasp” + sprinkle]). We propose a simple model to explain these results: auditory and visual information converge onto premotor cortex where it is represented in a comparable format to determine (in)congruence between speech and gesture. This ability of the dorsal route to detect audiovisual semantic (in)congruence suggests that its function is not restricted to the sublexical level.     

Aquatic Plants Stimulate the Growth of and Biofilm Formation by Mycobacterium ulcerans in Axenic Culture and Harbor These Bacteria in the Environment

Mycobacterium ulcerans is the causative agent of Buruli ulcer, one of the most common mycobacterial diseases of humans. Recent studies have implicated aquatic insects in the transmission of this pathogen, but the contributions of other elements of the environment remain largely unknown. We report here that crude extracts from two green algae added to the BACTEC 7H12B culture medium halved the doubling time of M. ulcerans and promoted biofilm formation. Using the 7H12B medium, modified by the addition of the algal extract, and immunomagnetic separation, we also demonstrate that M. ulcerans is associated with aquatic plants in an area of the Ivory Coast where Buruli ulcer is endemic. Genotype analysis showed that plant-associated M. ulcerans had the same profile as isolates recovered in the same region from both aquatic insects and clinical specimens. These observations implicate aquatic plants as a reservoir of M. ulcerans and add a new potential link in the chain of transmission of M. ulcerans to humans.