bluuebunny/crossref_metadata_2025_split huggingface dataset:
Autoantibodies recognizing citrullinated rat filaggrin in an ELISA using citrullinated and non-citrullinated recombinant proteins as antigens are highly diagnostic for rheumatoid arthritis
Authors: VITTECOQ O , INÇAURGARAT B , JOUEN-BEADES F , LEGOEDEC J , LETOURNEUR O , ROLLAND D , GERVASI G , MÉNARD J F , GAYET A , FARDELLONE P , DARAGON A , JOLIVET M , LE LOËT X , TRON F 
Published: December 2003

SUMMARYThe objective of the study was to determine the diagnostic value for rheumatoid arthritis (RA) of anti-filaggrin autoantibodies (autoAb) recognizing citrullinated recombinant rat filaggrin (ACRF) in community cases of very early arthritis. To evaluate the diagnostic value of ACRF, were studied sera from patients with different classified rheumatic diseases and healthy subjects (group 1, n= 422) and 314 community cases of very early arthritis (group 2) that were classified as RA (n = 176), non-RA (n = 63) and undifferentiated (n = 75) arthritides after 1 years of follow-up. ACRF were measured using a new ELISA, with results expressed as the difference between the OD value obtained on citrullinated minus that on noncitrullinated rat filaggrin (differential ACRF; dACRF). For both groups, rheumatoid factors (RF), anti-keratin autoAb (AKA) and anti-perinuclear factor (APF) were tested; for group 2, anti-CCP autoAb were also tested. Different reactivity patterns against citrullinated and noncitrullinated filaggrin were observed. Almost all sera reacting with citrullinated but not noncitrullinated filaggrin were from RA patients. Among RA and non-RA sera that recognized both forms of filaggrin, a positive result was obtained only with RA sera. For groups 1 and 2, dACRF sensitivity was 58·4% and 30·7%, and specificity for RA was 99·5% and 98·4%, respectively. In group 2, dACRF specificity for RA was better than that of RF (92·1%), APF (95·2%), AKA (96·8%) and anti-CCP (95·2%). dACRF positive predictive value was high (98·2) and close to that given by the concomitant positivity of RF and anti-CCP autoAb. Despite a high positive correlation between AKA, APF, anti-CCP and dACRF test results, they were complementary since some sera were positive for only one test. Thus, in a community setting, anti-citrullinated rat filaggrin reactivity detected by a new ELISA, whose originality is based on the difference between serum's reactivities on the citrullinated and native forms of filaggrin, had a higher diagnostic value for RA than other autoAb.
Molecular mechanisms of interleukin-10-mediated inhibition of NF-κB activity: a role for p50
Authors: DRIESSLER F , VENSTROM K , SABAT R , ASADULLAH K , SCHOTTELIUS A J 
Published: December 2003

SUMMARYNuclear factor kappa B (NF-κB) is a transcription factor pivotal for the development of inflammation. A dysregulation of NF-κB has been shown to play an important role in many chronic inflammatory diseases including rheumatoid arthritis, inflammatory bowel disease and psoriasis. Although classical NF-κB, a heterodimer composed of the p50 and p65 subunits, has been well studied, little is known about gene regulation by other hetero- and homodimeric forms of NF-κB. While p65 possesses a transactivation domain, p50 does not. Indeed, p50/p50 homodimers have been shown to inhibit transcriptional activity. We have recently shown that Interleukin-10 exerts its anti-inflammatory activity in part through the inhibition of NF-κB by blocking IκB kinase activity and by inhibiting NF-κB already found in the nucleus. Since the inhibition of nuclear NF-κB could not be explained by an increase of nuclear IκB, we sought to further investigate the mechanisms involved in the inhibition of NF-κB by IL-10. We show here that IL-10 selectively induced nuclear translocation and DNA-binding of p50/p50 homodimers in human monocytic cells. TNF-α treatment led to a strong translocation of p65 and p50, whereas pretreatment with IL-10 followed by TNF-α blocked p65 translocation but did not alter the strong translocation of p50. Furthermore, macrophages of p105/p50-deficient mice exhibited a significantly decreased constitutive production of MIP-2α and IL-6 in comparison to wild type controls. Surprisingly, IL-10 inhibited high constitutive levels of these cytokines in wt macrophages but not in p105/p50 deficient cells. Our findings suggest that the selective induction of nuclear translocation and DNA-binding of the repressive p50/p50 homodimer is an important anti-inflammatory mechanism utilized by IL-10 to repress inflammatory gene transcription.
Modulation of antigen presentation by autoreactive B cell clones specific for GAD65 from a type I diabetic patient
Authors: BANGA J P , MOORE J K , DUHINDAN N , MADEC A M , VAN ENDERT P M , ORGIAZZI J , ENDL J 
Published: December 2003

SUMMARY
We used a GAD65-specific human B–T cell line cognate system in vitro to investigate the modulation of GAD65 presentation by autoantibody, assessed in a proliferation assay. Generally, if the T cell determinant overlaps or resides within the antibody epitope, effects of presentation are blunted while if they are distant can lead to potent presentation. For three different autoreactive B–T cell line cognate pairs, the modulation of GAD65 presentation followed the mode of overlapping or distant epitopes with resultant potent or undetectable presentation. However, other cognate pairs elicited variability in this pattern of presentation. Notably, one B cell line, DPC, whose antibody epitope did not overlap with the T cell determinants, was consistently poor in presenting GAD65. Using the fluorescent dye Alexa Fluor 647 conjugated to GAD65 to study receptor-mediated antigen endocytosis showed that all the antigen-specific B cell clones were efficient in intracellular accumulation of the antigen. Additionally, multicolour immunofluorescence microscopy showed that the internalized GAD65/surface IgG complexes were rapidly targeted to a perinuclear compartment in all GAD-specific B cell clones. This analysis also demonstrated that HLA-DM expression was reduced strongly in DPC compared to the stimulatory B cell clones. Thus the capability of antigen-specific B cells to capture and present antigen to human T cell lines is dependent on the spatial relationship of B and T cell epitopes as well other factors which contribute to the efficiency of presentation.
The cardioprotector dexrazoxane augments therapeutic efficacy of mitoxantrone in experimental autoimmune encephalomyelitis
Authors: WEILBACH F X , CHAN A , TOYKA K V , GOLD R 
Published: December 2003

SUMMARYThe present study investigates the immunological effects of a combination treatment of mitoxantrone and the cardioprotector dexrazoxane in experimental autoimmune encephalomyelitis (EAE). Mitoxantrone, an anthracycline-derived immunosuppressive drug has been approved recently for treatment of very active multiple sclerosis (MS). Its prolonged use is limited due to its cardiotoxic properties. Dexrazoxane (DZR (S)-(+)-1,2-bis (3,5.dioxopiperazinyl)propane, ICRF-187) is an iron III chelator which in animal models and in cancer patients reduces anthracycline and mitoxantrone induced cardiotoxicity when given immediately before these agents. We examined the immunological effects of dexrazoxane in combination with mitoxantrone in experimental autoimmune encephalomyelitis (EAE) in Lewis rats. EAE was induced by active immunization with myelin basic protein (MBP) or by adoptive transfer of MBP specific T cells (AT-EAE). The clinical course, spinal cord pathology, activity of metalloproteinases (MMP-2 and MMP-9) and T cell apoptosis were assessed. Monotherapy with DZR ameliorated slightly the course of actively induced EAE and AT-EAE. The combination of DZR and mitoxantrone was superior to mitoxantrone given alone. Clinical amelioration ran in parallel with the marked reduction of inflammatory infiltration which was nearly abolished by the combination treatment. DZR did not affect the activity of metalloproteinase 9 and did not increase the proportion of apoptotic lymph node cells ex vivo or T cells in situ. We conclude that in addition to its cardioprotective role, DZR augments mitoxantrone-mediated immunosuppressive effects in animal models of human central nervous system (CNS) autoimmune disease. Clinical trials in MS patients are warranted to evaluate the unexpected immunosuppressive efficacy of DZR as add-on treatment.
Anti-C1q autoantibodies in murine lupus nephritis
Authors: TROUW L A , SEELEN M A , VISSEREN R , DUIJS J M G J , BENEDIKTSSON H , DE HEER E , ROOS A , VAN KOOTEN C , DAHA M R 
Published: December 2003

SUMMARY
Autoantibodies against C1q can be found in the circulation of patients with several autoimmune diseases including systemic lupus erythematosus (SLE). In SLE there is an association between the occurrence of these antibodies and renal involvement. How anti-C1q autoantibodies contribute to renal disease is currently unknown. Cohorts of MRL-lpr mice, which are known to develop age-dependent SLE-like disease, were used to study the relationship between levels of anti-C1q autoantibodies and renal disease. We collected serum, urine and renal tissue and analysed autoantibodies, complement levels and renal deposition as well as renal function. At 2 months of age all mice already had elevated levels of anti-C1q autoantibodies, and elution of kidneys revealed the presence of these antibodies in renal immune deposits in MRL-lpr mice and not in control MRL+/+ mice. In conclusion, anti-C1q antibodies are already present in serum and immune deposits of the kidney early in life and therefore can play a role in nephritis during experimental SLE-like disease in mice.
Multiple leucocyte activation markers to detect neonatal infection
Authors: HODGE G , HODGE S , HAN P , HASLAM R 
Published: December 2003

SUMMARY
Diagnosis of congenital or neonatal infection is often based on clinical signs. However, clinical symptoms of infections may not be specific, and for this reason early diagnosis is often determined on results of laboratory tests, which may not currently be adequate. A more reliable method of detection of infection may be the demonstration of activated lymphocytes, which can be conducted rapidly and before the isolation of the infected organism. We have shown that detection of up-regulation of CD45RO, an activated/memory isoform of CD45 present on T cells, provides a reasonably sensitive screening test for neonatal infection. We also showed that dual expression of CD45RA/CD45RO was up-regulated early during the infective process in neonates with documented infection. However, other leucocytes are also activated during the infective process. To improve the sensitivity of the neonatal infection screening test and to identify the types of leucocytes involved in the immune response to the infective organism, we studied further the up-regulation of a comprehensive range of surface activation markers on T cells, monocytes and natural killer (NK) cells from a group of 17 newborn patients with positive culture, a group of 40 possibly infected patients based on clinical signs and a control group. ‘Normal’ ranges were established for each activation marker for each leucocyte subset from 1 to 7 and 7-14-day-old newborns &lt;35 weeks’ gestation and 35-40 weeks’ gestation. There was a significant increase in the percentage of T cells expressing CD25 in the peripheral blood from infants at 2 weeks of age. Expression of HLA-DR on T cells, CD25 and CD69 on monocytes and HLA-DR on NK cells was also increased significantly in the peripheral blood from infants at 2 weeks of age and may reflect a maturation of these functional surface molecules. Up-regulation of CD69 on NK cells was the most sensitive marker for neonatal sepsis (positive in 13/16 patients). CD69 and CD25 expression was increased significantly on T cells in 11/17 and 10/17 patients, respectively. A combination of CD45RA/CD45RO and CD45RO identified 11/16 infected patients. Measurement of CD69 expression on NK cells with CD45RA, CD45RO, CD25 and CD69 expression on T cells resulted in a significant increase in at least two leucocyte activation markers from infected patients. In conclusion, this is the first report of the up-regulation of CD69 on NK cells as a sensitive marker of neonatal infection. A combination of this marker with CD45RA, CD45RO, CD25 and CD69 expression on peripheral blood derived T cells is the most sensitive and specific for neonatal infection.
Oriented immobilization of Pseudomonas putida putidaredoxin at a gold (111)–buffer interface: a real time scanning tunnelling microscopy study
Authors: Mukhopadhyay R. , Lo K. K. , Wong L. L. , Hill H. A. O. 
Published: December 2003

SummaryA scanning tunnelling microscopy study of adsorption of wild‐type Pseudomonas putida putidaredoxin at a gold (111)–buffer interface has been made in real time. Reversible adsorption has been observed reflecting weak interaction of the wild‐type protein with a gold (111) electrode. A genetically engineered mutant, C73S‐D58C, which contains a surface thiol, has been used for ‘immobilization’ and ‘orientated adsorption’ on the gold surface. The implication of such orientated immobilization in development of a bio‐electrode surface has been predicted.
φFLIM: a new method to avoid aliasing in frequency‐domain fluorescence lifetime imaging microscopy
Authors: Van Munster E. B. , Gadella T. W. J. 
Published: December 2003

SummaryIn conventional wide‐field frequency‐domain fluorescence lifetime imaging microscopy (FLIM), excitation light is intensity‐modulated at megahertz frequencies. Emitted fluorescence is recorded by a CCD camera through an image intensifier, which is modulated at the same frequency. From images recorded at various phase differences between excitation and intensifier gain modulation, the phase and modulation depth of the emitted light is obtained. The fluorescence lifetime is determined from the delay and the decrease in modulation depth of the emission relative to the excitation. A minimum of three images is required, but in this case measurements become susceptible to aliasing caused by the presence of higher harmonics. Taking more images to avoid this is not always possible owing to phototoxicity or movement. A method is introduced, φFLIM, requiring only three recordings that is not susceptible to aliasing. The phase difference between the excitation and the intensifier is scanned over the entire 360° range following a predefined phase profile, during which the image produced by the intensifier is integrated onto the CCD camera, yielding a single image. Three different images are produced following this procedure, each with a different phase profile. Measurements were performed with a conventional wide‐field frequency‐domain FLIM system based on an acousto‐optic modulator for modulation of the excitation and a microchannel‐plate image intensifier coupled to a CCD camera for the detection. By analysis of the harmonic content of measured signals it was found that the third harmonic was effectively the highest present. Using the conventional method with three recordings, phase errors due to aliasing of up to ± 29° and modulation depth errors of up to 30% were found. Errors in lifetimes of YFP‐transfected HeLa cells were as high as 100%. With φFLIM, using the same specimen and settings, systematic errors due to aliasing did not occur.
Analysis of boron‐10 in soft tissue by dynamic secondary ion mass spectrometry
Authors: Oyedepo A. C. , Brooke S. L. , Heard P. J. , Day J. C. C. , Allen G. C. , Patel H. 
Published: December 2003

SummaryWe report here a preliminary study in which dynamic secondary ion mass spectrometry (SIMS) has provided images of boron‐10 (10B) in biological tissue as used in research into boron neutron capture therapy. Cultured tumour cells incubated in media containing known concentrations of a 10B‐containing compound, p‐boronophenylalanine (BPA), and intracranial tumour tissue from animals previously injected with BPA were analysed by an in‐house constructed SIMS. Investigations were conducted in positive secondary ion detection mode using a 25‐keV, 5‐nA gallium primary ion source. For calibration purposes, tissue standards were also analysed and their boron‐to‐carbon signal ratios correlated to bulk boron concentrations measured by inductively coupled plasma atomic emission spectroscopy (ICP‐AES). Ion maps of 10B, 12C, 23Na and 39K showing gross tissue and cell features were acquired. SIMS and ICP‐AES standard measurements were in good agreement. Tissue regions with high or low 10B concentrations were identified along with 10B hotspots in normal brain areas. Cultured cells revealed the intracellular localization of 10B. SIMS is capable of producing images showing the distribution of 10B at p.p.m. levels in cells and in normal and tumour‐bearing brain tissue.
Optimizing imaging parameters for the separation of multiple labels in a fluorescence image
Authors: Neher R. , Neher E. 
Published: December 2003

SummaryA theoretical analysis is presented on how to separate the contributions from individual, simultaneously present fluorophores in a spectrally resolved image. Equations are derived that allow the calculation of the signal‐to‐noise ratio of the estimates for such contributions, given the spectral information on the individual fluorophores, the excitation wavelengths and intensities, and the number and widths of the spectral detection channels. We then ask how such imaging parameters have to be chosen for optimal fluorophore separation. We optimize the signal‐to‐noise ratio or optimize a newly defined ‘figure of merit’, which is a measure of efficiency in the use of emitted photons. The influence of photobleaching on the resolution and on the choice of imaging parameters is discussed, as well as the additional resolution gained by including fluorescence lifetime information. A surprisingly small number of spectral channels are required for an almost optimal resolution, if the borders of these channels are optimally selected. The detailed consideration of photobleaching is found to be essential, whenever there is significant bleaching. Consideration of fluorescence lifetime information (in addition to spectral information) improves results, particularly when lifetimes differ by more than a factor of two.
Freeze‐substitution and low‐temperature embedding of dairy products for transmission electron microscopy
Authors: Smith A. K. , Goff H. D. , Sun B. D. 
Published: December 2003

SummaryDairy products are comprised largely of fat, air and water, which makes it difficult to preserve their ultrastructure for electron microscopy. Keeping the samples frozen throughout fixation and embedding protects the structure and distribution of the components of emulsions and foams. Therefore, dairy products were freeze‐substituted and embedded at low temperature (−20 °C) to prepare them for transmission electron microscopy. Whipped cream, ice cream mix and dairy/non‐dairy mixed systems were frozen by plunging in propane, at its boiling point (−187 °C). Ice cream, because it is already frozen, was fractured into 1‐mm3 pieces in liquid nitrogen and then added to frozen fixative (−196 °C). Fixative solution consisted of glutaraldehyde, osmium tetroxide and uranyl acetate dissolved in either methanol or acetone. When material was to be stained after sectioning the fixative was limited to glutaraldehyde in methanol. The temperature was increased step‐wise from −80 to −20 °C. Solvent was replaced with resin; the polar resin Lowicryl HM4, the non‐polar resin Lowicryl HM20, LR White and LR Gold were tested. Samples were embedded and polymerized at −20 °C using ultraviolet light to cross‐link the resin. Methanol proved to be the most effective solvent for substituting the ice; the hydrophobic resin Lowicryl HM20 was the most effective resin for retaining fat structure following osmium fixation.
Graphical representation and multicomponent analysis of single‐frequency fluorescence lifetime imaging microscopy data
Authors: Clayton A. H. A. , Hanley Q. S. , Verveer P. J. 
Published: December 2003

SummaryGraphical representation of fluorescence lifetime imaging microscopy data demonstrates that a mixture of two components with single exponential decays can be resolved by single frequency measurements. We derive a method based on linear fitting that allows the calculation of the fluorescence lifetimes of the two components. We show that introduction of proper error‐weighting results in a non‐linear method that is mathematically identical to a global analysis algorithm that was recently derived. The graphical approach was applied to cellular data obtained from a lifetime‐based phosphorylation assay for the epidermal growth factor receptor and yielded results similar to those obtained by a global analysis algorithm.
Measurement of specimen‐induced aberrations of biological samples using phase stepping interferometry
Authors: Schwertner M. , Booth M. J. , Neil M. A. A. , Wilson T. 
Published: December 2003

SummaryConfocal or multiphoton microscopes, which deliver optical sections and three‐dimensional (3D) images of thick specimens, are widely used in biology. These techniques, however, are sensitive to aberrations that may originate from the refractive index structure of the specimen itself. The aberrations cause reduced signal intensity and the 3D resolution of the instrument is compromised. It has been suggested to correct for aberrations in confocal microscopes using adaptive optics. In order to define the design specifications for such adaptive optics systems, one has to know the amount of aberrations present for typical applications such as with biological samples. We have built a phase stepping interferometer microscope that directly measures the aberration of the wavefront. The modal content of the wavefront is extracted by employing Zernike mode decomposition. Results for typical biological specimens are presented. It was found for all samples investigated that higher order Zernike modes give only a small contribution to the overall aberration. Therefore, these higher order modes can be neglected in future adaptive optics sensing and correction schemes implemented into confocal or multiphoton microscopes, leading to more efficient designs.
An easy and versatile embedding method for transverse sections
Authors: De Smet I. , Chaerle P. , Vanneste S. , De Rycke R. , Inzé D. , Beeckman T. 
Published: December 2003

SummaryIn several research areas, transverse sections are indispensable for studying structural aspects of specimens. However, the oriented embedding of small cylindrical samples can become problematic, especially when transverse sections at right angles to the main axis of the object are desired. Here, we describe an easy and low‐cost technique for oriented embedding of small (∅ < 500 µm) as well as of larger specimens (∅ > 500 µm). The usefulness of the technique is demonstrated for roots and stamens of Arabidopsis thaliana and for adventitious roots of Asplenium demerkense, as examples of small and larger cylindrical samples, respectively. Furthermore, several types of resin (glycol methacrylate, epoxy and acrylic resins) were successfully tested, showing the applicability of the technique for light and electron microscopy and for immunolocalizations. In conclusion, the principle of the technique can be extended to several resins and a wide variety of specimen types, such as stems, leaves and textile fibres. The originality of the technique lies in its simplicity combined with its high efficiency to produce well‐oriented transverse sections.