A humanized model for multiple sclerosis using HLA-DR2 and a human T-cell receptor.

Multiple sclerosis (MS) is a complex chronic neurologic disease with a suspected autoimmune pathogenesis. Although there is evidence that the development of MS is determined by both environmental influences and genes, these factors are largely undefined, except for major histocompatibility (MHC) genes. Linkage analyses and association studies have shown that susceptibility to MS is associated with genes in the human histocompatibility leukocyte antigens (HLA) class II region, but the contribution of these genes to MS disease development less compared with their contribution to disorders such as insulin-dependent diabetes mellitus. Due to the strong linkage disequilibrium in the MHC class II region, it has not been possible to determine which gene(s) is responsible for the genetic predisposition. In transgenic mice, we have expressed three human components involved in T-cell recognition of an MS-relevant autoantigen presented by the HLA-DR2 molecule: DRA*0101/DRB1*1501 (HLA-DR2), an MHC class II candidate MS susceptibility genes found in individuals of European descent; a T-cell receptor (TCR) from an MS-patient-derived T-cell clone specific for the HLA-DR2 bound immunodominant myelin basic protein (MBP) 4102 peptide; and the human CD4 coreceptor. The amino acid sequence of the MBP 84-102 peptide is the same in both human and mouse MBP. Following administration of the MBP peptide, together with adjuvant and pertussis toxin, transgenic mice developed focal CNS inflammation and demyelination that led to clinical manifestations and disease courses resembling those seen in MS. Spontaneous disease was observed in 4% of mice. When DR2 and TCR double-transgenic mice were backcrossed twice to Rag2 (for recombination-activating gene 2)-deficient mice, the incidence of spontaneous disease increased, demonstrating that T cells specific for the HLA-DR2 bound MBP peptide are sufficient and necessary for development of disease. Our study provides evidence that HLA-DR2 can mediate both induced and spontaneous disease resembling MS by presenting an MBP self-peptide to T cells.

"To have and then lose the safety net" - Female cancer survivors' experiences of the process of becoming ready to return to work.

BACKGROUND: Validation studies have not been able to confirm the stage-specific understanding as operationalised in the readiness for return to work (RRTW) questionnaire. OBJECTIVE: To explore retrospectively how working female cancer survivors experienced the process of becoming ready to RTW during and beyond participation in an occupational rehabilitation intervention and thereby expand the understanding of the RRTW construct. METHODS: A qualitative research design was employed. Thirteen female cancer survivors were included for semi-structured interviews one to two years after they had completed active treatment and returned to work. The RRTW construct guided data generation and analysis. Content analysis was performed in four analytical steps that combined a concept-driven and a data-driven analytic strategy. RESULTS: Three themes were identified; "To have and then lose the safety net", "Realise a changed life situation", "Strive to balance work and everyday life". In a time span of approximately one to two years (from receiving treatment, being enrolled in an intervention and to gradually returning to work); the identified themes were interdependent of each other as one theme gradually evolved to the next theme in the process of engaging in sustained work participation. CONCLUSIONS: The present study points towards continuous development of the RRTW construct and whether the addition of a preparedness dimension would improve validity.

High quality anti-relaxation coating material for alkali atom vapor cells.

We present an experimental investigation of alkali atom vapor cells coated with a high quality anti-relaxation coating material based on alkenes. The prepared cells with single compound alkene based coating showed the longest spin relaxation times which have been measured up to now with room temperature vapor cells. Suggestions are made that chemical binding of a cesium atom and an alkene molecule by attack to the C = C bond plays a crucial role in such improvement of anti-relaxation coating quality.

Assessing the polarization of a quantum field from stokes fluctuations.

We propose an operational degree of polarization in terms of the variance of the Stokes vector minimized over all the directions of the Poincaré sphere. We examine the properties of this second-order definition and carry out its experimental determination. Quantum states with the same standard (first-order) degree of polarization are correctly discriminated by this new measure. We argue that a comprehensive quantum characterization of polarization properties requires a whole hierarchy of higher-order degrees.

Generation of two-mode squeezed and entangled light in a single temporal and spatial mode.

We analyse a novel squeezing and entangling mechanism which is due to correlated Stokes and anti-Stokes photon forward scattering in a multi-level atom vapour. We develop a full quantum model for an alkali atomic vapour including quantized collective atomic states which predicts high degree of squeezing for attainable experimental conditions. Following the proposal we present an experimental demonstration of 3.5 dB pulsed frequency nondegenerate squeezed (quadrature entangled) state of light using room temperature caesium vapour. The source is very robust and requires only a few milliwatts of laser power. The squeezed state is generated in the same spatial mode as the local oscillator and in a single temporal mode. The two entangled modes are separated by twice the Zeeman frequency of the vapour which can be widely tuned. The narrow-band squeezed light generated near an atomic resonance can be directly used for atom-based quantum information protocols. Its single temporal mode characteristics make it a promising resource for quantum information processing.

Selective positive modulation of the SK3 and SK2 subtypes of small conductance Ca2+-activated K+ channels.

BACKGROUND AND PURPOSE: Positive modulators of small conductance Ca(2+)-activated K(+) channels (SK1, SK2, and SK3) exert hyperpolarizing effects that influence the activity of excitable and non-excitable cells. The prototype compound 1-EBIO or the more potent compound NS309, do not distinguish between the SK subtypes and they also activate the related intermediate conductance Ca(2+)-activated K(+) channel (IK). This paper demonstrates, for the first time, subtype-selective positive modulation of SK channels. EXPERIMENTAL APPROACH: Using patch clamp and fluorescence techniques we studied the effect of the compound cyclohexyl-[2-(3,5-dimethyl-pyrazol-1-yl)-6-methyl-pyrimidin-4-yl]-amine (CyPPA) on recombinant hSK1-3 and hIK channels expressed in HEK293 cells. CyPPA was also tested on SK3 and IK channels endogenously expressed in TE671 and HeLa cells. KEY RESULTS: CyPPA was found to be a positive modulator of hSK3 (EC(50) = 5.6 +/- 1.6 microM, efficacy 90 +/- 1.8 %) and hSK2 (EC(50) = 14 +/- 4 microM, efficacy 71 +/- 1.8 %) when measured in inside-out patch clamp experiments. CyPPA was inactive on both hSK1 and hIK channels. At hSK3 channels, CyPPA induced a concentration-dependent increase in the apparent Ca(2+)-sensitivity of channel activation, changing the EC(50)(Ca(2+)) from 429 nM to 59 nM. CONCLUSIONS AND IMPLICATIONS: As a pharmacological tool, CyPPA may be used in parallel with the IK/SK openers 1-EBIO and NS309 to distinguish SK3/SK2- from SK1/IK-mediated pharmacological responses. This is important for the SK2 and SK1 subtypes, since they have overlapping expression patterns in the neocortical and hippocampal regions, and for SK3 and IK channels, since they co-express in certain peripheral tissues.

The Ca2+-activated K+ channel of intermediate conductance:a possible target for immune suppression.

The intermediate conductance Ca2+-activated K+ (IK) channel is distinguished from the functionally related Ca2+-activated K+ channels of smaller and larger unitary conductance by its molecular structure, pharmacology, tissue distribution and physiology. Like many K+ channels, IK is an assembly of four identical subunits each spanning the membrane six times and each contributing equally to the K+ selectivity pore positioned centrally in the complex. The IK channel gains its high sensitivity to intracellular Ca2+ from tightly bound calmodulin, and its activity is independent of the membrane potential. Several toxins including charybdotoxin and the more selective mutant, Glu32-charybdotoxin, maurotoxin and stichodactyla toxin potently block IK channels. Among blockers of the IK channel are also several small organic molecules including the antimycotic clotrimazole and the close analogues TRAM-34 and ICA-17043, as well as the antihypertensive, nitrendipine. The IK channel is distributed in peripheral tissues, including secretory epithelia and blood cells, but it appears absent from neuronal and muscle tissue. An important physiological role of the IK channel is to help maintain large electrical gradients for the sustained transport of ions such as Ca2+ influx that controls T lymphocyte (T cell) proliferation. In this review, special attention is given to an analysis of the use of IK blockers as potential immunosuppressants for the treatment of autoimmune disorders such as rheumatoid arthritis, inflammatory bowel disease and multiple sclerosis.

Functional characterization of HLA-DRA1*0101/DRB1*0401 molecules expressed in Drosophila melanogaster cells.

We have expressed the human MHC class II molecule, HLA-DRA1*0101/-DRB1*0401 (DRB1*0401), in Drosophila melanogaster Schneider 2 cells under control of the Drosophila metallothionein promoter. Upon induction with CuSO4, flow cytometry revealed expression of DRB1*0401 on the surface of the Drosophila cells at high levels. The membrane-bound class II molecules could present peptides specifically to DRB1*0401-restricted T cells. Drosophila-expressed DRB1*0401 molecules revealed a decreased N-linked glycosylation as compared to DRB1*0401 molecules purified from a human B-cell line. The purified DRB1*0401 molecules from Drosophila cells were dissociated into subunits in SDS-PAGE but could be stabilized with a peptide known to bind DRB1*0401 with a high affinity, indicating that the recombinant class II molecules from Drosophila cells are either empty or occupied by low affinity endogenous peptides. This assumption was further substantiated by the observation that the class II molecules from Drosophila cells had a much higher peptide-binding capacity than DRB1*0401 molecules derived from the human B-cell line. Otherwise, the two species of DRB1*0401 had similar peptide-binding specificities and affinities. The purified recombinant DRB1*0401 molecules also showed biological activity because immobilized complexes of DRB1*0401 and synthetic peptides specifically stimulated DRB1*0401-restricted T cells.

Definition of MHC and T cell receptor contacts in the HLA-DR4restricted immunodominant epitope in type II collagen and characterization of collagen-induced arthritis in HLA-DR4 and human CD4 transgenic mice.

Rheumatoid arthritis (RA) is an autoimmune disease associated with the HLA-DR4 and DR1 alleles. The target autoantigen(s) in RA is unknown, but type II collagen (CII) is a candidate, and the DR4- and DR1-restricted immunodominant T cell epitope in this protein corresponds to amino acids 261-273 (CII 261-273). We have defined MHC and T cell receptor contacts in CII 261-273 and provide strong evidence that this peptide corresponds to the peptide binding specificity previously found for RA-associated DR molecules. Moreover, we demonstrate that HLA-DR4 and human CD4 transgenic mice homozygous for the I-Abbeta0 mutation are highly susceptible to collagen-induced arthritis and describe the clinical course and histopathological changes in the affected joints.