Allotopic Expression of ATP6 in Mouse as a Transgenic Model of Mitochondrial Disease.

Progress in animal modeling of polymorphisms and mutations in mitochondrial DNA (mtDNA) is not as developed as nuclear transgenesis due to a host of cellular and physiological distinctions. mtDNA mutation modeling is of critical importance as mutations in the mitochondrial genome give rise to a variety of pathological conditions and play a contributing role in many others. Nuclear localization and transcription of mtDNA genes followed by cytoplasmic translation and transport into mitochondria (allotopic expression, AE) provide an opportunity to create in vivo modeling of a targeted mutation in mitochondrial genes.

Testicular germ line cell identification, isolation, and transplantation in two North American catfish species.

Our aim was to transplant blue catfish germ line stem cells into blastulae of triploid channel catfish embryos to produce interspecific xenogenic catfish. The morphological structure of the gonads of blue catfish (Ictalurus furcatus) in ~ 90- to 100-day-old juveniles, two-year-old juveniles, and mature adults was studied histologically. Both oogonia (12-15 μm, diameter with distinct nucleus 7-8 μm diameter) and spermatogonia (12-15 μm, with distinct nucleus 6-7.

Spermatogonial stem cells specific marker identification in channel catfish, Ictalurus punctatus and blue catfish, I. furcatus.

Testicular germ cells of channel catfish, Ictalurus punctatus, and blue catfish, I. furcatus were separated into four layers with Percoll density gradient centrifugation, containing different cell types (40% in the first layer were spermatogonial stem cells, SSCs). Expression of seventeen genes was analyzed for cells from different layers by real-time quantitative PCR.

Allotopic expression of ATP6 in the mouse as a transgenic model of mitochondrial disease.

Progress in animal modeling of polymorphisms and mutations in mitochondrial DNA (mtDNA) is not as developed as nuclear transgenesis due to a host of cellular and physiological distinctions. mtDNA mutation modeling is of critical importance as mutations in the mitochondrial genome give rise to a variety of pathological conditions and play a contributing role in many others. Nuclear localization and transcription of mtDNA genes followed by cytoplasmic translation and transport into mitochondria (allotopic expression, AE) provide an opportunity to create in vivo modeling of a targeted mutation in mitochondrial genes and has been suggested as a strategy for gene replacement therapy in patients harboring mitochondrial DNA mutations.

Characterization of mitochondrial populations during stem cell differentiation.

Mitochondrial dynamics play an important role in numerous physiological and pathophysiological phenomena in the developing and adult human heart. Alterations in structural aspects of cellular mitochondrial composition as a function of changes in physiology can easily be visualized using fluorescence microscopy. Commonly, mitochondrial location, number, and morphology are reported qualitatively due to the lack of automated and user-friendly computer-based analysis tools.

Biomimetic materials design for cardiac tissue regeneration.

Cardiovascular disease is the leading cause of death worldwide. In the absence of sufficient numbers of organs for heart transplant, alternate approaches for healing or replacing diseased heart tissue are under investigation. Designing biomimetic materials to support these approaches will be essential to their overall success.

Nuclear expression of a mitochondrial DNA gene: mitochondrial targeting of allotopically expressed mutant ATP6 in transgenic mice.

Nuclear encoding of mitochondrial DNA transgenes followed by mitochondrial targeting of the expressed proteins (allotopic expression; AE) represents a potentially powerful strategy for creating animal models of mtDNA disease. Mice were created that allotopically express either a mutant (A6M) or wildtype (A6W) mt-Atp6 transgene. Compared to non-transgenic controls, A6M mice displayed neuromuscular and motor deficiencies (wire hang, pole, and balance beam analyses; P < 0.

Animal models of human mitochondrial DNA mutations.

Background: Mutations in mitochondrial DNA (mtDNA) cause a variety of pathologic states in human patients. Development of animal models harboring mtDNA mutations is crucial to elucidating pathways of disease and as models for preclinical assessment of therapeutic interventions.

Scope Of Review: This review covers the knowledge gained through animal models of mtDNA mutations and the strategies used to produce them.

Taking a systems approach to the identification of novel therapeutic targets and biomarkers.

Systems biology focuses on the roles of cellular pathways and networks rather than single biomolecules to describe biological function. A systems view of biology requires technology that can generate and quantitatively analyze, large multi-dimensional data sets from many different sources. New technology has made this approach to drug discovery increasingly feasible.

The mitochondrial genome sequence of Mus terricolor: comparison with Mus musculus domesticus and implications for xenomitochondrial mouse modeling.

Knowledge of the mitochondrial DNA (mtDNA) sequence of divergent murine species is critical from both a phylogenetic perspective and in understanding nuclear-mitochondrial interactions, particularly as the latter influences our xenocybrid models of mitochondrial disease. To this end, the sequence of the mitochondrial genome of the murine species Mus terricolor (formerly Mus dunni) is reported and compared with the published sequence for the common laboratory mouse Mus musculus domesticus strain C57BL/6J. These species are of interest because xenomitochondrial cybrid mice were created that harbor M.

Foundation Review: Transgenic animals and their impact on the drug discovery industry.

The ability to direct genetic changes at the molecular level has resulted in a revolution in biology. Nowhere has this been more apparent than in the production of transgenic animals. Transgenic technology lies at the junction of several enabling techniques in such diverse fields as embryology, cell biology and molecular genetics.

Miniaturized, ultra-high throughput screening of tyrosine kinases using homogeneous, competitive fluorescence immunoassays.

Assay miniaturization and the implementation of high-density 1,536-microwell screening increase the speed and efficiency of screening and lead discovery. To serve this need, a platform of miniaturizable assay technologies has been assembled for specific biological targets. This platform will enable initiation and completion of uHTS screens in a straightforward and expeditious manner.

Evaluation of compound interference in immobilized metal ion affinity-based fluorescence polarization detection with a four million member compound collection.

IMAP is a non-separation-based, antibody-independent, FP assay that can be applied to many types of protein kinases and phosphatases. This technology is currently being used in many high-throughput screening campaigns throughout the industry. In this technology, a fluorescently labeled peptide substrate is phosphorylated and then captured on immobilized metal (M(III)) nanoparticles, an interaction that is enhanced at low pH (pH 5.

1,536-well assay development and screening using whole cell displacement binding and laser scanning imaging.

A screen of a GPCR against Pharmacopeia's combinatorial libraries was performed using 1,536-well plates in a 1.5-microl assay volume with an LSI that was specially modified to enable detection at these volumes. The screen encompassed approximately 4 x 10(6) compounds.