Molecular diagnosis for heterogeneous genetic diseases with high-throughput DNA sequencing applied to retinitis pigmentosa

BACKGROUND:
The genetic heterogeneity of many Mendelian disorders, such as retinitis pigmentosa which results from mutations in over 40 genes, is a major obstacle to obtaining a molecular diagnosis in clinical practice. Targeted high-throughput DNA sequencing offers a potential solution and was used to develop a molecular diagnostic screen for patients with retinitis pigmentosa.
METHODS:
A custom sequence capture array was designed to target the coding regions of all known retinitis pigmentosa genes and used to enrich these sequences from DNA samples of five patients. Enriched DNA was subjected to high-throughput sequencing singly or in pools, and sequence variants were identified by alignment of up to 10 million reads per sample to the normal reference sequence. Potential pathogenicity was assessed by functional predictions and frequency in controls.
RESULTS AND CONCLUSIONS:
Known homozygous PDE6B and compound heterozygous CRB1 mutations were detected in two patients. A novel homozygous missense mutation (c.2957A?T; p.N986I) in the cyclic nucleotide gated channel ß1 (CNGB1) gene predicted to have a deleterious effect and absent in 720 control chromosomes was detected in one case in which conventional genetic screening had failed to detect mutations. The detection of known and novel retinitis pigmentosa mutations in this study establishes high-throughput DNA sequencing with DNA pooling as an effective diagnostic tool for heterogeneous genetic diseases.

RCS CONTROL USING CASCADED CIRCULARLY POLARIZED FREQUENCY SELECTIVE SURFACES AND AN AMC STRUCTURE AS A SWITCHABLE TWIST POLARIZER

This article presents a cascaded arrangement comprising a double-layer frequency selective surface circularly polarizing (CPFSS) and a second screen that can be switched between artificial magnetic conduction (AMC) or perfect electric conducting. (PEC) states. The CPFSS consists of two stacked aluminium sheets patterned with periodic split ring structures While the AMC is a PCB sheet patterned with metallic squares interconnected by links By either open or short circuiting these links it is shown that the cascade of screens can be made to twist, or not to twist, an incident 45 degrees linearly polirized signal through 90 degrees upon reflection from the assembly The system was designed and optimized using CST software and predictions were validated experimentally and measured monostatic reflection loss results (C) 2010 Wiley Periodicals, Inc Microwave Opt Technol Lett 52 577-580, 2010. Published online in Wiley InterScience (www.interscience.wiley.com) DOI 10.1002/mop.24979

Through-thickness control of polymer bioresorption via electron beam irradiation

Predicable and controlled degradation is not only central to the accurate delivery of bioactive agents and drugs, it also plays a vital role in key aspects of bone tissue engineering. The work addressed in this paper investigates the utilisation of e-beam irradiation in order to achieve a controlled (surface) degradation profile. This study focuses on the modification of commercially and clinically relevant materials, namely poly(L-lactic acid) (PLLA), poly(L-lactide-hydroxyapatite) (PLLA-HA), poly(L-lactide-glycolide) co-polymer (PLG) and poly(L-lactide-DL-lactide) co-polymer (PLDL). Samples were subjected to irradiation treatments using a 0.5 MeV electron beam with delivered surface doses of 150 and 500 kGy. In addition, an acrylic attenuation shield was used for selected samples to control the penetration of the e-beam. E-beam irradiation induced chain scission in all polymers, as characterized by reduced molecular weights and glass transition temperatures (T-g). Irradiation not only produced changes in the physical properties of the polymers but also had associated effects on surface erosion of the materials during hydrolytic degradation. Moreover, the extent to which both mechanical and hydrolytic degradation was observed is synonymous with the estimated penetration of the beam (as controlled by the employment of an attenuation shield). (C) 2010 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Somatic point mutations in mtDNA control region are influenced by genetic background and associated with healthy aging: a GEHA stud

Tissue specific somatic mutations occurring in the mtDNA control region have been proposed to provide a survival advantage. Data on twins and on relatives of long-lived subjects suggested that the occurrence/accumulation of these mutations may be genetically influenced. To further investigate control region somatic heteroplasmy in the elderly, we analyzed the segment surrounding the nt 150 position (previously reported as specific of Leukocytes) in various types of leukocytes obtained from 195 ultra-nonagenarians sib-pairs of Italian or Finnish origin collected in the frame of the GEHA Project. We found a significant correlation of the mtDNA control region heteroplasmy between sibs, confirming a genetic influence on this phenomenon. Furthermore, many subjects showed heteroplasmy due to mutations different from the C150T transition. In these cases heteroplasmy was correlated within sibpairs in Finnish and northern Italian samples, but not in southern Italians. This suggested that the genetic contribution to control region mutations may be population specific. Finally, we observed a possible correlation between heteroplasmy and Hand Grip strength, one of the best markers of physical performance and of mortality risk in the elderly. Our study provides new evidence on the relevance of mtDNA somatic mutations in aging and longevity and confirms that the occurrence of specific point mutations in the mtDNA control region may represent a strategy for the age-related remodelling of organismal functions.

Association Study of SNAP25 and Schizophrenia in Irish Family and Case-Control Samples

SNAP25 occurs on chromosome 20p12.2, which has been linked to schizophrenia in some samples, and recently linked to latent classes of psychotic illness in our sample. SNAP25 is crucial to synaptic functioning, may be involved in axonal growth and dendritic sprouting, and its expression may be decreased in schizophrenia. We genotyped 18 haplotype-tagging SNPs in SNAP25 in a sample of 270 Irish high-density families. Single marker and haplotype analyses were performed in FBAT and PDT. We adjusted for multiple testing by computing q values. Association was followed up in an independent sample of 657 cases and 411 controls. We tested for allelic effects on the clinical phenotype by using the method of sequential addition and 5 factor-derived scores of the OPCRIT. Nine of 18 SNPs had Pvalues

No Association of Dysbindin With Symptom Factors of Schizophrenia in an Irish Case-Control Sample

Robust associations between the dysbindin gene (DTNBP1) and schizophrenia have been demonstrated in many but not all samples, and evidence that this gene particularly predisposes to negative symptoms in this illness has been presented. The current study sought to replicate the previously reported negative symptom associations in an Irish case-control sample. Association between dysbindin and schizophrenia has been established in this cohort, and a factor analysis of the assessed symptoms yielded three factors, Positive, Negative, and Schneiderian. The sequential addition method was applied using UNPHASED to assess the relationship between these symptom factors and the high-risk haplotype. No associations were detected for any of the symptom factors indicating that the dysbindin risk haplotype does not predispose to a particular group of symptoms in this sample. Several possibilities, such as differing risk haplotypes, may explain this finding. (C) 2009 Wiley-Liss, Inc.

Association Analysis of the PIP4K2A Gene on Chromosome 10p12 and Schizophrenia in the Irish Study of High Density Schizophrenia Families (ISHDSF) and the Irish Case-Control Study of Schizophrenia (ICCSS)

Molecular studies support pharmacological evidence that phosphoinositide signaling is perturbed in schizophrenia and bipolar disorder. The phosphatidylinositol-4-phosphate-5-kinase type-II alpha (PIP4K2A) gene is located on chromosome 10p12. This region has been implicated in both diseases by linkage, and PIP4K2A directly by association. Given linkage evidence in the Irish Study of High Density Schizophrenia Families (ISHDSF) to a region including 10p12, we performed an association study between genetic variants at PIP4K2A and disease. No association was detected through single-marker or haplotype analysis of the whole sample. However, stratification into families positive and negative for the ISHDSF schizophrenia high-risk haplotype (HRH) in the DTNBP1 gene and re-analysis for linkage showed reduced amplitude of the 10p12 linkage peak in the DTNBP1 HRH positive families. Association analysis of the stratified sample showed a trend toward association of PIP4K2A SNPs rs1417374 and rs1409395 with schizophrenia in the DTNBP1 HRH positive families. Despite this apparent paradox, our data may therefore suggest involvement of PIP4K2A in schizophrenia in those families for whom genetic variation in DTNBP1 appears also to be a risk factor. This trend appears to arise from under-transmission of common alleles to female cases. Follow-up association analysis in a large Irish schizophrenia case-control control sample (ICCSS) showed significant association with disease of a haplotype comprising these same SNPs rs1417374-rs1409395, again more so in affected females, and in cases with negative family history of the disease. This study supports a minor role for PIP4K2A in schizophrenia etiology in the Irish population. (C) 2009 Wiley-Liss, Inc.

Replication of association between schizophrenia and ZNF804A in the Irish Case-Control Study of Schizophrenia sample

A recent genome-wide association study reported association between schizophrenia and the ZNF804A gene on chromosome 2q32.1. We attempted to replicate these findings in our Irish Case-Control Study of Schizophrenia (ICCSS) sample (N=1021 cases, 626 controls). Following consultation with the original investigators, we genotyped three of the most promising single-nucleotide polymorphisms (SNPs) from the Cardiff study. We replicate association with rs1344706 (trend test one-tailed P=0.0113 with the previously associated A allele) in ZNF804A. We detect no evidence of association with rs6490121 in NOS1 (one-tailed P=0.21), and only a trend with rs9922369 in RGRIP1L (one-tailed P=0.0515). On the basis of these results, we completed genotyping of 11 additional linkage disequilibrium-tagging SNPs in ZNF804A. Of 12 SNPs genotyped, 11 pass quality control criteria and 4 are nominally associated, with our most significant evidence of association at rs7597593 (P=0.0013) followed by rs1344706. We observe no evidence of differential association in ZNF804A on the basis of family history or sex of case. The associated SNP rs1344706 lies in approximately 30 bp of conserved mammalian sequence, and the associated A allele is predicted to maintain binding sites for the brain-expressed transcription factors MYT1l and POU3F1/OCT-6. In controls, expression is significantly increased from the A allele of rs1344706 compared with the C allele. Expression is increased in schizophrenic cases compared with controls, but this difference does not achieve statistical significance. This study replicates the original reported association of ZNF804A with schizophrenia and suggests that there is a consistent link between the A allele of rs1344706, increased expression of ZNF804A and risk for schizophrenia.

FBXL21 Association With Schizophrenia in Irish Family and Case-Control Samples

FBXL21 gene encodes an F-box containing protein functioning in the SCIP ubiquitin ligase complex. The role of the F-box protein is to recruit proteins designated for degradation to the ligase complex so they would be ubiquitinated. Using both family and case-control samples, we found consistent associations in and around FBXL21 gene. In the family sample (Irish study of high density schizophrenia families, ISHDSF, 1,350 subjects from 273 families), a minimal PDT P-value of 0.0011 was observed at rs31555. In the case-control sample (Irish case-control study of schizophrenia, ICCSS, 814 cases and 625 controls), significant associations were observed at two markers (rs1859427 P=0.0197, and rs6861170 P=0.0197). In haplotype analyses, haplotype 1-1 (C-T) of rs1859427-rs6861170 was overtransmitted in the ISHDSF (P=0.0437) and was over-represented in the ICCSS (P=0.0177). For both samples, the associated alleles and haplotypes were identical. These data suggested that FBXL21 maybe associated with schizophrenia in the Irish samples. (C) 2008 Wiley-Liss, Inc.

Autoimmune conditions and hairy cell leukemia: an exploratory case-control study

Background: Case reports suggest that hairy cell leukemia (HCL) may be associated with autoimmune conditions, however no systematic investigations in this area have been undertaken.

Apc(MIN) modulation of vitamin D secosteroid growth control

A central paradox of vitamin D biology is that 1alpha,25-(OH)(2) D(3) exposure inversely relates to colorectal cancer (CRC) risk despite a capacity for activation of both pro- and anti-oncogenic mediators including osteopontin (OPN)/CD44 and E-cadherin, respectively. Most sporadic CRCs arise from adenomatous polyposis coli (APC) gene mutation but understanding of its effects on vitamin D growth control is limited. Here we investigate effects of the Apc(Min/+) genotype on 1alpha,25-(OH)(2) D(3) regulation of OPN/CD44/E-cadherin signalling and intestinal tumourigenesis, in vivo. In untreated Apc(Min/+) versus Apc(+/+) intestines, expression levels of OPN and its CD44 receptor were increased, whereas E-cadherin tumour suppressor signalling was attenuated. Treatment by 1alpha,25-(OH)(2) D(3) or rationally designed analogues (QW or BTW) enhanced OPN but inhibited expression of CD44, the OPN receptor implicated in cell growth. These treatments also enhanced E-cadherin tumour suppressor activity, characterized by inhibition of beta-catenin nuclear localization, T-cell factor 1 and c-myelocytomatosis protein expression in Apc(Min/+) intestine. All secosteroids suppressed Apc(Min/+)-driven tumourigenesis although QW and BTW had lower calcium-related toxicity. Taken together, these data indicate that the Apc(Min/+) genotype modulates vitamin D secosteroid actions to promote functional predominance of E-cadherin tumour suppressor activity within antagonistic molecular networks. APC heterozygosity may promote favourable tissue- or tumour-specific conditions for growth control by vitamin D secosteroid treatment.

Electron-beam treatment of poly(lactic acid) to control degradation profiles

Bioresorbable polymers such as polylactide (PIA) and polylactide-co-glycolide (PLGA) have been used successfully as biomaterials in a wide range of medical applications. However, their slow degradation rates and propensity to lose strength before mass have caused problems. A central challenge for the development of these materials is the assurance of consistent and predictable in vivo degradation. Previous work has illustrated the potential to influence polymer degradation using electron beam (e-beam) radiation. The work addressed in this paper investigates further the utilisation of e-beam radiation in order to achieve a more surface specific effect. Variation of e-beam energy was studied as a means to control the effective penetrative depth in poly-L-lactide (PLEA). PLEA samples were exposed to e-beam radiation at individual energies of 0.5 MeV, 0.75 MeV and 1.5 MeV. The near-surface region of the PLEA samples was shown to be affected by e-beam irradiation with induced changes in molecular weight, morphology, flexural strength and degradation profile. Moreover, the depth to which the physical properties of the polymer were affected is dependent on the beam energy used. Computer modelling of the transmission of each e-beam energy level used corresponded well with these findings. (C) 2010 Elsevier Ltd. All rights reserved.

Redistribution of vibrational population in a molecular ion with nonresonant strong-field laser pulses

We present an experimental demonstration of nonresonant manipulation of vibrational states in a molecule by an intense ultrashort laser pulse. A vibrational wave packet is generated in D-2(+) through tunnel ionization of D-2 by a few-cycle pump pulse. A similar control pulse is applied as the wave packet begins to dephase so that the dynamic Stark effect distorts the electronic environment of the nuclei, transferring vibrational population. The time evolution of the modified wave packet is probed via the D-2(+) photodissociation yield that results from the application of an intense probe pulse. Comparing the measured yield with a quasiclassical trajectory model allows us to determine the redistribution of vibrational population caused by the control pulse. ©

Optimal control of atom transport for quantum gates in optical lattices

By means of optimal control techniques we model and optimize the manipulation of the external quantum state (center-of-mass motion) of atoms trapped in adjustable optical potentials. We consider in detail the cases of both noninteracting and interacting atoms moving between neighboring sites in a lattice of a double-well optical potentials. Such a lattice can perform interaction-mediated entanglement of atom pairs and can realize two-qubit quantum gates. The optimized control sequences for the optical potential allow transport faster and with significantly larger fidelity than is possible with processes based on adiabatic transport.

Cloning transformations in spin networks without external control

In this paper we present an approach to quantum cloning with unmodulated spin networks. The cloner is realized by a proper design of the network and a choice of the coupling between the qubits. We show that in the case of phase covariant cloner the XY coupling gives the best results. In the 1 -> 2 cloning we find that the value for the fidelity of the optimal cloner is achieved, and values comparable to the optimal ones in the general N -> M case can be attained. If a suitable set of network symmetries are satisfied, the output fidelity of the clones does not depend on the specific choice of the graph. We show that spin network cloning is robust against the presence of static imperfections. Moreover, in the presence of noise, it outperforms the conventional approach. In this case the fidelity exceeds the corresponding value obtained by quantum gates even for a very small amount of noise. Furthermore, we show how to use this method to clone qutrits and qudits. By means of the Heisenberg coupling it is also possible to implement the universal cloner although in this case the fidelity is 10% off that of the optimal cloner.