Genetic, environmental and stochastic factors in monozygotic twin discordance with a focus on epigenetic differences

PMCID: PMC3566971This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited

Cryo-electron microscopy of viruses

Thin vitrified layers of unfixed, unstained and unsupported virus suspensions can be prepared for observation by cryo-electron microscopy in easily controlled conditions. The viral particles appear free from the kind of damage caused by dehydration, freezing or adsorption to a support that is encountered in preparing biological samples for conventional electron microscopy. Cryo-electron microscopy of vitrified specimens offers possibilities for high resolution observations that compare favourably with any other electron microscopical method

Transcriptome Analysis of the Octopus vulgaris Central Nervous System

Background: Cephalopoda are a class of Mollusca species found in all the world's oceans. They are an important model organism in neurobiology. Unfortunately, the lack of neuronal molecular sequences, such as ESTs, transcriptomic or genomic information, has limited the development of molecular neurobiology research in this unique model organism. Results: With high-throughput Illumina Solexa sequencing technology, we have generated 59,859 high quality sequences from 12,918,391 paired-end reads. Using BLASTx/BLASTn, 12,227 contigs have blast hits in the Swissprot, NR protein database and NT nucleotide database with E-value cutoff 1e(-5). The comparison between the Octopus vulgaris central nervous system (CNS) library and the Aplysia californica/Lymnaea stagnalis CNS ESTs library yielded 5.93%/13.45% of O. vulgaris sequences with significant matches (1e(-5)) using BLASTn/tBLASTx. Meanwhile the hit percentage of the recently published Schistocerca gregaria, Tilapia or Hirudo medicinalis CNS library to the O. vulgaris CNS library is 21.03%-46.19%. We constructed the Phylogenetic tree using two genes related to CNS function, Synaptotagmin-7 and Synaptophysin. Lastly, we demonstrated that O. vulgaris may have a vertebrate-like Blood-Brain Barrier based on bioinformatic analysis. Conclusion: This study provides a mass of molecular information that will contribute to further molecular biology research on O. vulgaris. In our presentation of the first CNS transcriptome analysis of O. vulgaris, we hope to accelerate the study of functional molecular neurobiology and comparative evolutionary biology.National fund for oceanography research in Public Interest [201005013]; National Key Technology RD Program [2011BAD13

Unconventional Low-Cost Fabrication and Patterning Techniques for Point of Care Diagnostics

The potential of rapid, quantitative, and sensitive diagnosis has led to many innovative ‘lab on chip’ technologies for point of care diagnostic applications. Because these chips must be designed within strict cost constraints to be widely deployable, recent research in this area has produced extremely novel non-conventional micro- and nano-fabrication innovations. These advances can be leveraged for other biological assays as well, including for custom assay development and academic prototyping. The technologies reviewed here leverage extremely low-cost substrates and easily adoptable ways to pattern both structural and biological materials at high resolution in unprecedented ways. These new approaches offer the promise of more rapid prototyping with less investment in capital equipment as well as greater flexibility in design. Though still in their infancy, these technologies hold potential to improve upon the resolution, sensitivity, flexibility, and cost-savings over more traditional approaches

Rest-Mediated Regulation of Extracellular Matrix Is Crucial for Neural Development

Neural development from blastocysts is strictly controlled by intricate transcriptional programmes that initiate the down-regulation of pluripotent genes, Oct4, Nanog and Rex1 in blastocysts followed by up-regulation of lineage-specific genes as neural development proceeds. Here, we demonstrate that the expression pattern of the transcription factor Rest mirrors those of pluripotent genes during neural development from embryonic stem (ES) cells and an early abrogation of Rest in ES cells using a combination of gene targeting and RNAi approaches causes defects in this process. Specifically, Rest ablation does not alter ES cell pluripotency, but impedes the production of Nestin+ neural stem cells, neural progenitor cells and neurons, and results in defective adhesion, decrease in cell proliferation, increase in cell death and neuronal phenotypic defects typified by a reduction in migration and neurite elaboration. We also show that these Rest-null phenotypes are due to the dysregulation of its direct or indirect target genes, Lama1, Lamb1, Lamc1 and Lama2 and that these aberrant phenotypes can be rescued by laminins

Microbial Maintenance: A Critical Review on Its Quantification

Microbial maintenance is an important concept in microbiology. Its quantification, however, is a subject of continuous debate, which seems to be caused by (1) its definition, which includes nongrowth components other than maintenance; (2) the existence of partly overlapping concepts; (3) the evolution of variables as constants; and (4) the neglect of cell death in microbial dynamics. The two historically most important parameters describing maintenance, the specific maintenance rate and the maintenance coefficient, are based on partly different nongrowth components. There is thus no constant relation between these parameters and previous equations on this subject are wrong. In addition, the partial overlap between these parameters does not allow the use of a simple combination of these parameters. This also applies for combinations of a threshold concentration with one of the other estimates of maintenance. Maintenance estimates should ideally explicitly describe each nongrowth component. A conceptual model is introduced that describes their relative importance and reconciles the various concepts and definitions. The sensitivity of maintenance on underlying components was analyzed and indicated that overall maintenance depends nonlinearly on relative death rates, relative growth rates, growth yield, and endogenous metabolism. This quantitative sensitivity analysis explains the felt need to develop growth-dependent adaptations of existing maintenance parameters, and indicates the importance of distinguishing the various nongrowth components. Future experiments should verify the sensitivity of maintenance components under cellular and environmental conditions

Endogenous VEGF Is Required for Visual Function: Evidence for a Survival Role on Müller Cells and Photoreceptors

Vascular endothelial growth factor (VEGF) is well known for its role in normal and pathologic neovascularization. However, a growing body of evidence indicates that VEGF also acts on non-vascular cells, both developmentally as well as in the adult. In light of the widespread use of systemic and intraocular anti-VEGF therapies for the treatment of angiogenesis associated with tumor growth and wet macular degeneration, systematic investigation of the role of VEGF in the adult retina is critical.Using immunohistochemistry and Lac-Z reporter mouse lines, we report that VEGF is produced by various cells in the adult mouse retina and that VEGFR2, the primary signaling receptor, is also widely expressed, with strong expression by Müller cells and photoreceptors. Systemic neutralization of VEGF was accomplished in mice by adenoviral expression of sFlt1. After 14 days of VEGF neutralization, there was no effect on the inner and outer retina vasculature, but a significant increase in apoptosis of cells in the inner and outer nuclear layers. By four weeks, the increase in neural cell death was associated with reduced thickness of the inner and outer nuclear layers and a decline in retinal function as measured by electroretinograms. siRNA-based suppression of VEGF expression in a Müller cell line in vitro supports the existence of an autocrine role for VEGF in Müller cell survival. Similarly, the addition of exogenous VEGF to freshly isolated photoreceptor cells and outer-nuclear-layer explants demonstrated VEGF to be highly neuroprotective.These results indicate an important role for endogenous VEGF in the maintenance and function of adult retina neuronal cells and indicate that anti-VEGF therapies should be administered with caution

Development of an In Vivo RNAi Protocol to Investigate Gene Function in the Filarial Nematode, Brugia malayi

Our ability to control diseases caused by parasitic nematodes is constrained by a limited portfolio of effective drugs and a paucity of robust tools to investigate parasitic nematode biology. RNA interference (RNAi) is a reverse-genetics tool with great potential to identify novel drug targets and interrogate parasite gene function, but present RNAi protocols for parasitic nematodes, which remove the parasite from the host and execute RNAi in vitro, are unreliable and inconsistent. We have established an alternative in vivo RNAi protocol targeting the filarial nematode Brugia malayi as it develops in an intermediate host, the mosquito Aedes aegypti. Injection of worm-derived short interfering RNA (siRNA) and double stranded RNA (dsRNA) into parasitized mosquitoes elicits suppression of B. malayi target gene transcript abundance in a concentration-dependent fashion. The suppression of this gene, a cathepsin L-like cysteine protease (Bm-cpl-1) is specific and profound, both injection of siRNA and dsRNA reduce transcript abundance by 83%. In vivo Bm-cpl-1 suppression results in multiple aberrant phenotypes; worm motility is inhibited by up to 69% and parasites exhibit slow-moving, kinked and partial-paralysis postures. Bm-cpl-1 suppression also retards worm growth by 48%. Bm-cpl-1 suppression ultimately prevents parasite development within the mosquito and effectively abolishes transmission potential because parasites do not migrate to the head and proboscis. Finally, Bm-cpl-1 suppression decreases parasite burden and increases mosquito survival. This is the first demonstration of in vivo RNAi in animal parasitic nematodes and results indicate this protocol is more effective than existing in vitro RNAi methods. The potential of this new protocol to investigate parasitic nematode biology and to identify and validate novel anthelmintic drug targets is discussed