Rapid immunoblot and kinase assay tests for a syndromal form of X linked mental retardation: Coffin-Lowry syndrome

Coffin-Lowry syndrome (CLS) is a syndromal form of X linked mental retardation, in which some associated facial, hand, and skeletal abnormalities are diagnostic features. Accurate diagnosis, critical for genetic counselling, is often difficult, especially in early childhood. We have recently shown that Coffin-Lowry syndrome is caused by mutations in the gene encoding RSK2, a growth factor regulated protein kinase. RSK2 mutations are very heterogeneous and most of them lead to premature termination of translation or to loss of phosphotransferase activity or both. In the present study, we have evaluated immunoblot and RSK2 kinase assays as a rapid and simple diagnostic test for CLS, using cultured lymphoblastoid or fibroblast cell lines. Western blot analysis failed to detect RSK2 in six patients, suggesting the presence of truncated proteins in these patients. This conclusion was confirmed in four patients, in whom the causative mutations, all leading to premature termination of translation, were identified. Of four patients showing a normal amount of RSK2 protein on western blot and tested for RSK2 phosphotransferase activity, one had a dramatically impaired activity. Analysis of the RSK2 cDNA sequence in this patient showed a mutation of a putative phosphorylation site that would be critical for RSK2 activity. Preliminary results show that, at least, the western blot protocol can be successfully applied to lymphocyte protein extracts prepared directly from blood samples. These assays promise to become important diagnostic tools for CLS, particularly with regard to very young patients with no family history of the condition.     

Myelinosome Organelles in the Retina of R6/1 Huntington Disease (HD) Mice: Ubiquitous Distribution and Possible Role in Disease Spreading

Visual deficit is one of the complications of Huntington disease (HD), a fatal neurological disorder caused by CAG trinucleotide expansions in the Huntingtin gene, leading to the production of mutant Huntingtin (mHTT) protein. Transgenic HD R6/1 mice expressing human HTT exon1 with 115 CAG repeats recapitulate major features of the human pathology and exhibit a degeneration of the retina. Our aim was to gain insight into the ultrastructure of the pathological HD R6/1 retina by electron microscopy (EM). We show that the HD R6/1 retina is enriched with unusual organelles myelinosomes, produced by retinal neurons and glia. Myelinosomes are present in all nuclear and plexiform layers, in the synaptic terminals of photoreceptors, in the processes of retinal neurons and glial cells, and in the subretinal space. In vitro study shows that myelinosomes secreted by human retinal glial Müller MIO-M1 cells transfected with EGFP-mHTT-exon1 carry EGFP-mHTT-exon1 protein, as revealed by immuno-EM and Western-blotting. Myelinosomes loaded with mHTT-exon1 are incorporated by naive neuronal/neuroblastoma SH-SY5Y cells. This results in the emergence of mHTT-exon1 in recipient cells. This process is blocked by membrane fusion inhibitor MDL 28170. Conclusion: Incorporation of myelinosomes carrying mHTT-exon1 in recipient cells may contribute to HD spreading in the retina. Exploring ocular fluids for myelinosome presence could bring an additional biomarker for HD diagnostics.     

New wireless connection between user and VE using speech processing

This paper presents a novel speak-to-VR virtual-reality peripheral network (VRPN) server based on speech processing. The server uses a microphone array as a speech source and streams the results of the process through a Wi-Fi network. The proposed VRPN server provides a handy, portable and wireless human machine interface that can facilitate interaction in a variety interfaces and application domains including HMD- and CAVE-based virtual reality systems, flight and driving simulators and many others. The VRPN server is based on a speech processing software development kits and VRPN library in C++. Speak-to-VR VRPN works well even in the presence of background noise or the voices of other users in the vicinity. The speech processing algorithm is not sensitive to the user’s accent because it is trained while it is operating. Speech recognition parameters are trained by hidden Markov model in real time. The advantages and disadvantages of the speak-to-VR server are studied under different configurations. Then, the efficiency and the precision of the speak-to-VR server for a real application are validated via a formal user study with ten participants. Two experimental test setups are implemented on a CAVE system by using either Kinect Xbox or array microphone as input device. Each participant is asked to navigate in a virtual environment and manipulate an object. The experimental data analysis shows promising results and motivates additional research opportunities.     

Mutation analysis of the RSK2 gene in Coffin-Lowry patients: extensive allelic heterogeneity and a high rate of de novo mutations

Coffin-Lowry syndrome (CLS) is an X-linked disorder characterized by severe psychomotor retardation, facial and digital dysmorphisms, and progressive skeletal deformations. By using a positional cloning approach, we have recently shown that mutations in the gene coding for the RSK2 serine-threonine protein kinase are responsible for this syndrome. To facilitate mutational analysis, we have now determined the genomic structure of the human RSK2 gene. The open reading frame of the RSK2 coding region is split into 22 exons. Primers were designed for PCR amplification of single exons from genomic DNA and subsequent single-strand conformation polymorphism analysis. We screened 37 patients with clinical features suggestive of CLS. Twenty-five nucleotide changes predicted to be disease-causing mutations were identified, including eight splice-site alterations, seven nonsense mutations, five frameshift mutations, and five missense mutations. Twenty-three of them were novel mutations. Coupled with previously reported mutations, these findings bring the total of different RSK2 mutations to 34. These are distributed throughout the RSK2 gene, with no clustering, and all but two, which have been found in two independent patients, are unique. A very high (68%) rate of de novo mutations was observed. It is noteworthy also that three mutations were found in female probands, with no affected male relatives, ascertained through learning disability and mild but suggestive facial and digital dysmorphisms. No obvious correlation was observed between the position or type of the RSK2 mutations and the severity or particular clinical features of CLS.     

Characterization of fibre/resin bonding in composites using a pull-out test

From the early use of glass fibres in an organic matrix to that of high strength and high modulus fibres such as carbon and Kevlar, the subject of fibre/matrix bonding has received considerable attention. This paper gives details of a pull-out test has been used to measure the adhesion between fibres and matrix. A summary of results obtained with the test technique, over a period of some ten years, is also given. Despite operational difficulties the method gave interesting results regarding the mechanical aspects of the interaction between fibres and matrix.     

Failure in radiosurgery treatment of cerebral arteriovenous malformations

OBJECTIVE: The aim of this study was to retrospectively analyze the reasons for the failure of radiosurgical treatment of cerebral arteriovenous malformations (AVMs). METHODS: Seventeen cases of noncured AVMs were reviewed 3 years after radiosurgical treatment. Follow-up ranged from 33 to 54 months (mean, 44.3 mo). Lesion dimensions varied from 9 to 55 mm (mean, 29.2 mm). The lesions were located in critical or near-critical brain regions. Angiography was performed under Talairach's stereotactic conditions. Two large AVMs bled 36 and 39 months after receiving irradiation, respectively. These two AVMs had been incompletely irradiated. RESULTS: Retrospectively, in four cases (23.5%) we observed errors in determining AVM target shape and size because of inaccurate definition of the nidus and/or because of stereoangiographic incompleteness (absence of external carotid artery injections). In five large and/or irregularly shaped AVMs (29.4%), a strategy of partial volume irradiation had been used. In one patient (5.8%), we observed the recanalization of previously embolized AVMs. In another case (5.8%), the target had been partially missed. The AVMs in one case (5.8%) had been treated with an ineffective peripheral dose. In one (5.8%), the failure occurred because of the lesion angio-architecture. In four cases (23.5%), no evident reasons for failure were determined. CONCLUSION: The results of this study suggest the necessity of complete irradiation of the nidus. The strategy of partial volume irradiation might be avoided, even if it necessitates lowering the doses to treat large AVMs. Accuracy in the target determination is required, and complete stereoangiography is necessary.     

CAVE and HMD: distance perception comparative study

Virtual Reality - This paper proposes to analyse user experience using two different immersive device categories: a cave automatic virtual environment (CAVE) and a head-mounted display (HMD). While...