Isolation and characterization of a spontaneously immortalized bovine retinal pigmented epithelial cell line

<p>Abstract</p> <p>Background</p> <p>The Retinal Pigmented Epithelium (RPE) is juxtaposed with the photoreceptor outer segments of the eye. The proximity of the photoreceptor cells is a prerequisite for their survival, as they depend on the RPE to remove the outer segments and are also influenced by RPE cell paracrine factors. RPE cell death can cause a progressive loss of photoreceptor function, which can diminish vision and, over time, blindness ensues. Degeneration of the retina has been shown to induce a variety of retinopathies, such as Stargardt's disease, Cone-Rod Dystrophy (CRD), Retinitis Pigmentosa (RP), Fundus Flavimaculatus (FFM), Best's disease and Age-related Macular Degeneration (AMD). We have cultured primary bovine RPE cells to gain a further understanding of the mechanisms of RPE cell death. One of the cultures, named tRPE, surpassed senescence and was further characterized to determine its viability as a model for retinal diseases.</p> <p>Results</p> <p>The tRPE cell line has been passaged up to 150 population doublings and was shown to be morphologically similar to primary cells. They have been characterized to be of RPE origin by reverse transcriptase PCR and immunocytochemistry using the RPE-specific genes <it>RPE65 </it>and <it>CRALBP </it>and RPE-specific proteins RPE65 and Bestrophin. The tRPE cells are also immunoreactive to vimentin, cytokeratin and zonula occludens-1 antibodies. Chromosome analysis indicates a normal diploid number. The tRPE cells do not grow in suspension or in soft agar. After ³H thymidine incorporation, the cells do not appear to divide appreciably after confluency.</p> <p>Conclusion</p> <p>The tRPE cells are immortal, but still exhibit contact inhibition, serum dependence, monolayer growth and secrete an extra-cellular matrix. They retain the <it>in-vivo </it>morphology, gene expression and cell polarity. Additionally, the cells endocytose exogenous melanin, A2E and purified lipofuscin granules. This cell line may be a useful <it>in-vitro </it>research model for retinal maculopathies.</p

Ordering of localized moments in Kondo lattice models

We describe the transition from a ferromagnetic phase, to a disordered para- magnetic phase, which occurs in one-dimensional Kondo lattice models with partial conduction band filling. The transition is the quantum order-disorder transition of the transverse-field Ising chain, and reflects double-exchange ordered regions of localized spins being gradually destroyed as the coupling to the conduction electrons is reduced. For incommensurate conduction band filling, the low-energy properties of the localized spins near the transition are dominated by anomalous ordered (disordered) regions of localized spins which survive into the paramagnetic (ferromagnetic) phase. Many interesting properties follow, including a diverging susceptibility for a finite range of couplings into the paramagnetic phase. Our critical line equation, together with numerically determined transition points, are used to determine the range of the double-exchange interaction. Models we consider are the spin 1/2 Kondo lattices with antiferromagnetic (Kondo) coupling, with ferromagnetic (Hund's rule) coupling, and the Kondo lattice with repulsive interactions between the conduction electrons.Comment: 18 pages, 6 embedded eps figures. To appear in Phys Rev

Manifestation of classical wave delays in a fully quantized model of the scattering of a single photon

We consider a fully quantized model of spontaneous emission, scattering, and absorption, and study propagation of a single photon from an emitting atom to a detector atom both with and without an intervening scatterer. We find an exact quantum analog to the classical complex analytic signal of an electromagnetic wave scattered by a medium of charged oscillators. This quantum signal exhibits classical phase delays. We define a time of detection which, in the appropriate limits, exactly matches the predictions of a classically defined delay for light propagating through a medium of charged oscillators. The fully quantized model provides a simple, unambiguous, and causal interpretation of delays that seemingly imply speeds greater than c in the region of anomalous dispersion.Comment: 18 pages, 4 figures, revised for clarity, typos corrrecte

Islands of linkage in an ocean of pervasive recombination reveals two-speed evolution of human cytomegalovirus genomes

Human cytomegalovirus (HCMV) infects most of the population worldwide, persisting throughout the host's life in a latent state with periodic episodes of reactivation. While typically asymptomatic, HCMV can cause fatal disease among congenitally infected infants and immunocompromised patients. These clinical issues are compounded by the emergence of antiviral resistance and the absence of an effective vaccine, the development of which is likely complicated by the numerous immune evasins encoded by HCMV to counter the host's adaptive immune responses, a feature that facilitates frequent super-infections. Understanding the evolutionary dynamics of HCMV is essential for the development of effective new drugs and vaccines. By comparing viral genomes from uncultivated or low-passaged clinical samples of diverse origins, we observe evidence of frequent homologous recombination events, both recent and ancient, and no structure of HCMV genetic diversity at the whole-genome scale. Analysis of individual gene-scale loci reveals a striking dichotomy: while most of the genome is highly conserved, recombines essentially freely and has evolved under purifying selection, 21 genes display extreme diversity, structured into distinct genotypes that do not recombine with each other. Most of these hyper-variable genes encode glycoproteins involved in cell entry or escape of host immunity. Evidence that half of them have diverged through episodes of intense positive selection suggests that rapid evolution of hyper-variable loci is likely driven by interactions with host immunity. It appears that this process is enabled by recombination unlinking hyper-variable loci from strongly constrained neighboring sites. It is conceivable that viral mechanisms facilitating super-infection have evolved to promote recombination between diverged genotypes, allowing the virus to continuously diversify at key loci to escape immune detection, while maintaining a genome optimally adapted to its asymptomatic infectious lifecycle

The novel RASSF6 and RASSF10 candidate tumour suppressor genes are frequently epigenetically inactivated in childhood leukaemias.

BACKGROUND: The Ras-association family (RASSF) of tumour suppressor genes (TSGs) contains 10 members that encode proteins containing Ras-association (RA) domains. Several members of the RASSF family are frequently epigenetically inactivated in cancer, however, their role in leukaemia has remained largely uninvestigated. Also, RASSF10 is a predicted gene yet to be experimentally verified. Here we cloned, characterised and demonstrated expression of RASSF10 in normal human bone marrow. We also determined the methylation status of CpG islands associated with RASSF1-10 in a series of childhood acute lymphocytic leukaemias (ALL) and normal blood and bone marrow samples. RESULTS: COBRA and bisulphite sequencing revealed RASSF6 and RASSF10 were the only RASSF members with a high frequency of leukaemia-specific methylation. RASSF6 was methylated in 94% (48/51) B-ALL and 41% (12/29) T-ALL, whilst RASSF10 was methylated in 16% (8/51) B-ALL and 88% (23/26) T-ALL. RASSF6 and RASSF10 expression inversely correlated with methylation which was restored by treatment with 5-aza-2'deoxycytidine (5azaDC). CONCLUSION: This study shows the hypermethylation profile of RASSF genes in leukaemias is distinct from that of solid tumours and represents the first report of inactivation of RASSF6 or RASSF10 in cancer. These data show epigenetic inactivation of the candidate TSGs RASSF6 and RASSF10 is an extremely frequent event in the pathogenesis of childhood leukaemia. This study also warrants further investigation of the newly identified RASSF member RASSF10 and its potential role in leukaemia.RIGHTS : This article is licensed under the BioMed Central licence at http://www.biomedcentral.com/about/license which is similar to the 'Creative Commons Attribution Licence'. In brief you may : copy, distribute, and display the work; make derivative works; or make commercial use of the work - under the following conditions: the original author must be given credit; for any reuse or distribution, it must be made clear to others what the license terms of this work are

Ground State Magnetization of Polymerized Spin Chains

We investigate the ground state magnetization plateaus appearing in spin 1/2 polymerized Heisenberg chains under external magnetic fields. The associated fractional quantization scenario and the exponents which characterize the opening of gapful excitations are analyzed by means of abelian bosonization methods. Our conclusions are fully supported by the extrapolated results obtained from Lanczos diagonalizations of finite systems.Comment: 5 pages, 6 figures, final version to appear in Phys.Rev.

Non-standard embedding and five-branes in heterotic M-Theory

We construct vacua of M-theory on S^1/Z_2 associated with Calabi-Yau three-folds. These vacua are appropriate for compactification to N=1 supersymmetry theories in both four and five dimensions. We allow for general E_8 x E_8 gauge bundles and for the presence of five-branes. The five-branes span the four-dimensional uncompactified space and are wrapped on holomorphic curves in the Calabi-Yau space. Properties of these vacua, as well as of the resulting low-energy theories, are discussed. We find that the low-energy gauge group is enlarged by gauge fields that originate on the five-brane world-volumes. In addition, the five-branes increase the types of new E_8 x E_8 breaking patterns allowed by the non-standard embedding. Characteristic features of the low-energy theory, such as the threshold corrections to the gauge kinetic functions, are significantly modified due to the presence of the five-branes, as compared to the case of standard or non-standard embeddings without five-branes.Comment: 34 pages, Latex 2e with amsmath, typos removed, factors corrected, refs improve

Quantitative Bias in Illumina TruSeq and a Novel Post Amplification Barcoding Strategy for Multiplexed DNA and Small RNA Deep Sequencing

Here we demonstrate a method for unbiased multiplexed deep sequencing of RNA and DNA libraries using a novel, efficient and adaptable barcoding strategy called Post Amplification Ligation-Mediated (PALM). PALM barcoding is performed as the very last step of library preparation, eliminating a potential barcode-induced bias and allowing the flexibility to synthesize as many barcodes as needed. We sequenced PALM barcoded micro RNA (miRNA) and DNA reference samples and evaluated the quantitative barcode-induced bias in comparison to the same reference samples prepared using the Illumina TruSeq barcoding strategy. The Illumina TruSeq small RNA strategy introduces the barcode during the PCR step using differentially barcoded primers, while the TruSeq DNA strategy introduces the barcode before the PCR step by ligation of differentially barcoded adaptors. Results show virtually no bias between the differentially barcoded miRNA and DNA samples, both for the PALM and the TruSeq sample preparation methods. We also multiplexed miRNA reference samples using a pre-PCR barcode ligation. This barcoding strategy results in significant bias

For whom does it work? Moderators of outcome on the effect of a transdiagnostic internet-based maintenance treatment after inpatient psychotherapy: Randomized controlled trial

Background: Recent studies provide evidence for the effectiveness of Internet-based maintenance treatments for mental disorders. However, it is still unclear which participants might or might not profit from this particular kind of treatment delivery. Objective: The study aimed to identify moderators of treatment outcome in a transdiagnostic Internet-based maintenance treatment (TIMT) offered to patients after inpatient psychotherapy for mental disorders in routine care. Methods: Using data from a randomized controlled trial (N=400) designed to test the effectiveness of TIMT, we performed secondary analyses to identify factors moderating the effects of TIMT (intervention) when compared with those of a treatment-as-usual control condition. TIMT involved an online self-management module, asynchronous patient-therapist communication, a peer support group, and online-based progress monitoring. Participants in the control condition had unstructured access to outpatient psychotherapy, standardized outpatient face-to-face continuation treatment, and psychotropic management. Self-reports of psychopathological symptoms and potential moderators were assessed at the start of inpatient treatment (T1), at discharge from inpatient treatment/start of TIMT (T2), and at 3-month (T3) and 12-month follow-up (T4). Results: Education level, positive outcome expectations, and diagnoses significantly moderated intervention versus control differences regarding changes in outcomes between T2 and T3. Only education level moderated change differences between T2 and T4. The effectiveness of the intervention (vs control) was more pronounced among participants with a low (vs high) education level (T2-T3: B=-0.32, SE 0.16, P=.049; T2-T4: B=-0.42, SE 0.21, P=.049), participants with high (vs low) positive outcome expectations (T2-T3: B=-0.12, SE 0.05, P=.02) and participants with anxiety disorder (vs mood disorder) (T2-T3: B=-0.43, SE 0.21, P=.04). Simple slope analyses revealed that despite some subgroups benefiting less from the intervention than others, all subgroups still benefited significantly. Conclusions: This transdiagnostic Internet-based maintenance treatment might be suitable for a wide range of participants differing in various clinical, motivational, and demographic characteristics. The treatment is especially effective for participants with low education levels. These findings may generalize to other Internet-based maintenance treatments. Trial Registration: International Standard Randomized Controlled Trial Number (ISRCTN): 28632626; http://www.controlled-trials.com/isrctn/pf/ 28632626 (Archived by WebCite at http://www.webcitation.org/6IqZjTLrx). © David Daniel Ebert, Mario Gollwitzer, Heleen Riper, Pim Cuijpers, Harald Baumeister, Matthias Berking

Measurement of the ttbar Production Cross Section in ppbar Collisions at sqrt{s}=1.96 TeV using Lepton + Jets Events with Secondary Vertex b-tagging

We present a measurement of the ttbar production cross section using events with one charged lepton and jets from ppbar collisions at a center-of-mass energy of 1.96 TeV. In these events, heavy flavor quarks from top quark decay are identified with a secondary vertex tagging algorithm. From 162 pb-1 of data collected by the Collider Detector at Fermilab, a total of 48 candidate events are selected, where 13.5 +- 1.8 events are expected from background contributions. We measure a ttbar production cross section of 5.6^{+1.2}_{-1.1} (stat.) ^{+0.9}_{0.6} (syst.) pb.Comment: 28 pages, 20 figures. Published in Physical Review