30 - Identifying the presence of ​Scn4a, Scn5a, ​ and​ Scn9a ​ in the Mammalian Clitoris

Between 40 and 50% of women report experiences of sexual dysfunction (SD). This includes an inability to achieve sexual arousal, experience sexual pleasure and/or achieve an orgasm. Though there is no evidence of SD interfering with the ability to conceive, the conditions impacts affected individuals quality of life. Of particular interest are the mechanisms involved in the process of female sexual arousal. SD is often viewed as a psychological condition and limited information is known about the biological underpinnings of this condition. The goal of our laboratory is to identify the role of voltage gated ion channels in the mammalian clitoris. These candidate genes are of interest based on studies examining patients with epilepsy reporting decreased ability to achieve sexual arousal after taking common antiepileptic drugs carbamazepine or phenytoin. These drugs function by blocking sodium current from voltage gated ion channels in excitable cells. This suggests that voltage gated sodium channels may be present in the clitoris, and thusly play a role in sexual function. In this project, we sought to identify if voltage gated ion channels commonly found in the peripheral nervous system where expressed in the mammalian clitoris. This includes Scn4a, Scn5a, and Scn9a. Utilizing a RT-PCR profiler assay for neuronal ion channels (Qiagen, inc), we examined the expression pattern of these transcripts in the C57B/6J mouse clitoris tissue. Expression patterns were then confirmed with single RT-PCR. Transcripts with a ≥ 2 fold difference were considered significant. We found that Scn9a was expressed in this excitable tissue at significant levels, suggesting its role in arousal and the potential for therapeutic intervention

Successful Amelioration of Mitochondrial Optic Neuropathy Using the Yeast NDI1 Gene in a Rat Animal Model

Background: Leber’s hereditary optic neuropathy (LHON) is a maternally inherited disorder with point mutations in mitochondrial DNA which result in loss of vision in young adults. The majority of mutations reported to date are within the genes encoding the subunits of the mitochondrial NADH-quinone oxidoreductase, complex I. Establishment of animal models of LHON should help elucidate mechanism of the disease and could be utilized for possible development of therapeutic strategies. Methodology/Principal Findings: We established a rat model which involves injection of rotenone-loaded microspheres into the optic layer of the rat superior colliculus. The animals exhibited the most common features of LHON. Visual loss was observed within 2 weeks of rotenone administration with no apparent effect on retinal ganglion cells. Death of retinal ganglion cells occurred at a later stage. Using our rat model, we investigated the effect of the yeast alternative NADH dehydrogenase, Ndi1. We were able to achieve efficient expression of the Ndi1 protein in the mitochondria of all regions of retinal ganglion cells and axons by delivering the NDI1 gene into the optical layer of the superior colliculus. Remarkably, even after the vision of the rats was severely impaired, treatment of the animals with the NDI1 gene led to a complete restoration of the vision to the normal level. Control groups that received either empty vector or the GFP gene had no effects

Patient-Specific Prosthetic Fingers by Remote Collaboration - A Case Study

The concealment of amputation through prosthesis usage can shield an amputee from social stigma and help improve the emotional healing process especially at the early stages of hand or finger loss. However, the traditional techniques in prosthesis fabrication defy this as the patients need numerous visits to the clinics for measurements, fitting and follow-ups. This paper presents a method for constructing a prosthetic finger through online collaboration with the designer. The main input from the amputee comes from the Computer Tomography (CT) data in the region of the affected and the non-affected fingers. These data are sent over the internet and the prosthesis is constructed using visualization, computer-aided design and manufacturing tools. The finished product is then shipped to the patient. A case study with a single patient having an amputated ring finger at the proximal interphalangeal joint shows that the proposed method has a potential to address the patient's psychosocial concerns and minimize the exposure of the finger loss to the public.Comment: Open Access articl

I-TRAP: A method to identify transcriptional regulator activated promoters

BACKGROUND: The differential expression of virulence genes is often used by microbial pathogens in adapting to the environment of their host. The differential expression of such sets of genes can be regulated by RNA polymerase sigma factors. Some sigma factors are differentially expressed, which can provide a means to identifying other differentially expressed genes such as those whose expression are controlled by the sigma factor. METHODS: To identify sigma factor-regulated genes, we developed a method, termed I-TRAP, for the identification of transcriptional regulator activated promoters. The I-TRAP method is based on the fact that some genes will be differentially expressed in the presence and absence of a transcriptional regulator. I-TRAP uses a DNA library in a promoter-trap vector that contains two reporter genes, one to allow the selection of active promoters in the presence of the transcriptional regulator and a second to allow screening for promoter activity in the absence of the transcriptional regulator. RESULTS: To illustrate the development and use of the I-TRAP approach, the construction of the vectors, host strains, and library necessary to identify SigmaE-regulated genes of Mycobacterium tuberculosis is described. CONCLUSION: The I-TRAP method should be a versatile and useful method for identifying and characterizing promoter activity under a variety of conditions and in response to various regulatory proteins. In our study, we isolated 360 clones that may contain plasmids carrying SigmaE-regulated promoters genes of M. tuberculosis

Neuroprotection in a Novel Mouse Model of Multiple Sclerosis

The authors acknowledge the support of the Barts and the London Charity, the Multiple Sclerosis Society of Great Britain and Northern Ireland, the National Multiple Sclerosis Society, USA, notably the National Centre for the Replacement, Refinement & Reduction of Animals in Research, and the Wellcome Trust (grant no. 092539 to ZA). The siRNA was provided by Quark Pharmaceuticals. The funders and Quark Pharmaceuticals had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript

Detection of subtelomere imbalance using MLPA: validation, development of an analysis protocol, and application in a diagnostic centre

BACKGROUND: Commercial MLPA kits (MRC-Holland) are available for detecting imbalance at the subtelomere regions of chromosomes; each kit consists of one probe for each subtelomere. METHODS: For validation of the kits, 208 patients were tested, of which 128 were known to be abnormal, corresponding to 8528 genomic regions overall. Validation samples included those with trisomy 13, 18 and 21, microscopically visible terminal deletions and duplications, sex chromosome abnormalities and submicroscopic abnormalities identified by multiprobe FISH. A robust and sensitive analysis system was developed to allow accurate interpretation of single probe results, which is essential as breakpoints may occur between MLPA probes. RESULTS: The validation results showed that MLPA is a highly efficient technique for medium-throughput screening for subtelomere imbalance, with 95% confidence intervals for positive and negative predictive accuracies of 0.951-0.996 and 0.9996-1 respectively. A diagnostic testing strategy was established for subtelomere MLPA and any subsequent follow-up tests that may be required. The efficacy of this approach was demonstrated during 15 months of diagnostic testing when 455 patients were tested and 27 (5.9%) abnormal cases were detected. CONCLUSION: The development of a robust, medium-throughput analysis system for the interpretation of results from subtelomere assays will be of benefit to other Centres wishing to implement such an MLPA-based service

Mapping gene associations in human mitochondria using clinical disease phenotypes

Nuclear genes encode most mitochondrial proteins, and their mutations cause diverse and debilitating clinical disorders. To date, 1,200 of these mitochondrial genes have been recorded, while no standardized catalog exists of the associated clinical phenotypes. Such a catalog would be useful to develop methods to analyze human phenotypic data, to determine genotype-phenotype relations among many genes and diseases, and to support the clinical diagnosis of mitochondrial disorders. Here we establish a clinical phenotype catalog of 174 mitochondrial disease genes and study associations of diseases and genes. Phenotypic features such as clinical signs and symptoms were manually annotated from full-text medical articles and classified based on the hierarchical MeSH ontology. This classification of phenotypic features of each gene allowed for the comparison of diseases between different genes. In turn, we were then able to measure the phenotypic associations of disease genes for which we calculated a quantitative value that is based on their shared phenotypic features. The results showed that genes sharing more similar phenotypes have a stronger tendency for functional interactions, proving the usefulness of phenotype similarity values in disease gene network analysis. We then constructed a functional network of mitochondrial genes and discovered a higher connectivity for non-disease than for disease genes, and a tendency of disease genes to interact with each other. Utilizing these differences, we propose 168 candidate genes that resemble the characteristic interaction patterns of mitochondrial disease genes. Through their network associations, the candidates are further prioritized for the study of specific disorders such as optic neuropathies and Parkinson disease. Most mitochondrial disease phenotypes involve several clinical categories including neurologic, metabolic, and gastrointestinal disorders, which might indicate the effects of gene defects within the mitochondrial system. The accompanying knowledgebase (http://www.mitophenome.org/) supports the study of clinical diseases and associated genes

Adipocyte-specific protein tyrosine phosphatase 1B deletion increases lipogenesis, adipocyte cell size and is a minor regulator of glucose homeostasis

Peer reviewedPublisher PD

An Updated Algorithm for the Generation of Neutral Landscapes by Spectral Synthesis

Background: Patterns that arise from an ecological process can be driven as much from the landscape over which the process is run as it is by some intrinsic properties of the process itself. The disentanglement of these effects is aided if it possible to run models of the process over artificial landscapes with controllable spatial properties. A number of different methods for the generation of so-called ‘neutral landscapes’ have been developed to provide just such a tool. Of these methods, a particular class that simulate fractional Brownian motion have shown particular promise. The existing methods of simulating fractional Brownian motion suffer from a number of problems however: they are often not easily generalisable to an arbitrary number of dimensions and produce outputs that can exhibit some undesirable artefacts. Methodology: We describe here an updated algorithm for the generation of neutral landscapes by fractional Brownian motion that do not display such undesirable properties. Using Monte Carlo simulation we assess the anisotropic properties of landscapes generated using the new algorithm described in this paper and compare it against a popular benchmark algorithm. Conclusion/Significance: The results show that the existing algorithm creates landscapes with values strongly correlated in the diagonal direction and that the new algorithm presented here corrects this artefact. A number of extensions of the algorithm described here are also highlighted: we describe how the algorithm can be employed to generate landscapes that display different properties in different dimensions and how they can be combined with an environmental gradient to produce landscapes that combine environmental variation at the local and macro scales