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31.
32.
Infection of CD127+ (interleukin-7 receptor+) CD4+ cells and overexpression of CTLA-4 are linked to loss of antigen-specific CD4 T cells during primary human immunodeficiency virus type 1 infection 下载免费PDF全文
Zaunders JJ Ip S Munier ML Kaufmann DE Suzuki K Brereton C Sasson SC Seddiki N Koelsch K Landay A Grey P Finlayson R Kaldor J Rosenberg ES Walker BD Fazekas de St Groth B Cooper DA Kelleher AD 《Journal of virology》2006,80(20):10162-10172
We recently found that human immunodeficiency virus (HIV)-specific CD4+ T cells express coreceptor CCR5 and activation antigen CD38 during early primary HIV-1 infection (PHI) but then rapidly disappear from the circulation. This cell loss may be due to susceptibility to infection with HIV-1 but could also be due to inappropriate apoptosis, an expansion of T regulatory cells, trafficking out of the circulation, or dysfunction. We purified CD38+++CD4+ T cells from peripheral blood mononuclear cells, measured their level of HIV-1 DNA by PCR, and found that about 10% of this population was infected. However, a small subset of HIV-specific CD4+) T cells also expressed CD127, a marker of long-term memory cells. Purified CD127+CD4+ lymphocytes contained fivefold more copies of HIV-1 DNA per cell than did CD127-negative CD4+ cells, suggesting preferential infection of long-term memory cells. We observed no apoptosis of antigen-specific CD4+ T cells in vitro and only a small increase in CD45RO+CD25+CD127dimCD4+ T regulatory cells during PHI. However, 40% of CCR5+CD38+++ CD4+ T cells expressed gut-homing integrins, suggesting trafficking through gut-associated lymphoid tissue (GALT). Furthermore, 80% of HIV-specific CD4+ T cells expressed high levels of the negative regulator CTLA-4 in response to antigen stimulation in vitro, which was probably contributing to their inability to produce interleukin-2 and proliferate. Taken together, the loss of HIV-specific CD4+ T cells is associated with a combination of an infection of CCR5+ CD127+ memory CD4+ T cells, possibly in GALT, and a high expression of the inhibitory receptor CTLA-4. 相似文献
33.
Thuvaraka Thavayogarajah Preetish Gangopadhyay Stefan Rahlfs Katja Becker Klaus Lingelbach Jude M. Przyborski Anthony A. Holder 《PloS one》2015,10(4)
Plasmodium falciparum invades human red blood cells, residing in a parasitophorous vacuole (PV), with a parasitophorous vacuole membrane (PVM) separating the PV from the host cell cytoplasm. Here we have investigated the role of N-myristoylation and two other N-terminal motifs, a cysteine potential S-palmitoylation site and a stretch of basic residues, as the driving force for protein targeting to the parasite plasma membrane (PPM) and subsequent translocation across this membrane. Plasmodium falciparum adenylate kinase 2 (Pf AK2) contains these three motifs, and was previously proposed to be targeted beyond the parasite to the PVM, despite the absence of a signal peptide for entry into the classical secretory pathway. Biochemical and microscopy analyses of PfAK2 variants tagged with green fluorescent protein (GFP) showed that these three motifs are involved in targeting the protein to the PPM and translocation across the PPM to the PV. It was shown that the N-terminal 37 amino acids of PfAK2 alone are sufficient to target and translocate GFP across the PPM. As a control we examined the N-myristoylated P. falciparum ADP-ribosylation factor 1 (PfARF1). PfARF1 was found to co-localise with a Golgi marker. To determine whether or not the putative palmitoylation and the cluster of lysine residues from the N-terminus of PfAK2 would modulate the subcellular localization of PfARF1, a chimeric fusion protein containing the N-terminus of PfARF1 and the two additional PfAK2 motifs was analysed. This chimeric protein was targeted to the PPM, but not translocated across the membrane into the PV, indicating that other features of the N-terminus of PfAK2 also play a role in the secretion process. 相似文献
34.
Chukwunonso Onyilagha Ping Jia Nipun Jayachandran Sen Hou Ifeoma Okwor Shiby Kuriakose Aaron Marshall Jude E. Uzonna 《PLoS neglected tropical diseases》2015,9(4)
Background
Bam32, a 32 kDa adaptor molecule, plays important role in B cell receptor signalling, T cell receptor signalling and antibody affinity maturation in germinal centres. Since antibodies against trypanosome variant surface glycoproteins (VSG) are critically important for control of parasitemia, we hypothesized that Bam32 deficient (Bam32-/-) mice would be susceptible to T. congolense infection.Methodology/Principal Findings
We found that T. congolense-infected Bam32-/- mice successfully control the first wave of parasitemia but then fail to control subsequent waves and ultimately succumb to their infection unlike wild type (WT) C57BL6 mice which are relatively resistant. Although infected Bam32-/- mice had significantly higher hepatomegaly and splenomegaly, their serum AST and ALT levels were not different, suggesting that increased liver pathology may not be responsible for the increased susceptibility of Bam32-/- mice to T. congolense. Using direct ex vivo flow cytometry and ELISA, we show that CD4+ T cells from infected Bam32-/- mice produced significantly increased amounts of disease-exacerbating proinflammatory cytokines (including IFN-γ, TNF-α and IL-6). However, the percentages of regulatory T cells and IL-10-producing CD4+ cells were similar in infected WT and Bam32-/- mice. While serum levels of parasite-specific IgM antibodies were normal, the levels of parasite-specific IgG, (particularly IgG1 and IgG2a) were significantly lower in Bam32-/- mice throughout infection. This was associated with impaired germinal centre response in Bam32-/- mice despite increased numbers of T follicular helper (Tfh) cells. Adoptive transfer studies indicate that intrinsic B cell defect was responsible for the enhanced susceptibility of Bam32-/- mice to T. congolense infection.Conclusions/Significance
Collectively, our data show that Bam32 is important for optimal anti-trypanosome IgG antibody response and suppression of disease-promoting proinflammatory cytokines and its deficiency leads to inability to control T. congolense infection in mice. 相似文献35.
Emanuela Carlotti David Wrench Guglielmo Rosignoli Jacek Marzec Ajanthah Sangaralingam Lena Hazanov Miri Michaeli Simon Hallam Tracy Chaplin Sameena Iqbal Maria Calaminici Bryan Young Ramit Mehr Peter Campbell Jude Fitzgibbon John G. Gribben 《PloS one》2015,10(9)
Understanding the dynamics of evolution of Follicular Lymphoma (FL) clones during disease progression is important for monitoring and targeting this tumor effectively. Genetic profiling of serial FL biopsies and examples of FL transmission following bone marrow transplant suggest that this disease may evolve by divergent evolution from a common ancestor cell. However where this ancestor cell resides and how it evolves is still unclear. The analysis of the pattern of somatic hypermutation of the immunoglobulin gene (Ig) is traditionally used for tracking the physiological clonal evolution of B cells within the germinal center and allows to discriminate those cells that have just entered the germinal center and display features of ancestor cells from those B cells that keep re-circulating across different lymphoid organs. Here we investigated the pattern of somatic hypermutation of the heavy chain of the immunoglobulin gene (IgH-VH) in 4 flow-sorted B cells subpopulations belonging to different stages of differentiation, from sequential lymph node biopsies of cases displaying diverse patterns of evolution, using the GS-FLX Titanium sequencing platform. We observed an unexpectedly high level of clonality, with hundreds of distinct tumor subclones in the different subpopulations from the same sample, the majority detected at a frequency <10−2. By using a lineage trees analysis we observed in all our FL and t-FL cases that the oligoclonal FL population was trapped in a narrow intermediate stage of maturation that maintains the capacity to undergo SHM, but was unable to further differentiate. The presence of such a complex architecture highlights challenges currently encountered in finding a cure for this disease. 相似文献
36.
Othman I. Alomair Ian M. Brereton Maree T. Smith Graham J. Galloway Nyoman D. Kurniawan 《PloS one》2015,10(6)
Magnetic Resonance Imaging (MRI) of the rodent brain at ultra-high magnetic fields (> 9.4 Tesla) offers a higher signal-to-noise ratio that can be exploited to reduce image acquisition time or provide higher spatial resolution. However, significant challenges are presented due to a combination of longer T
1 and shorter T
2/T2* relaxation times and increased sensitivity to magnetic susceptibility resulting in severe local-field inhomogeneity artefacts from air pockets and bone/brain interfaces. The Stejskal-Tanner spin echo diffusion-weighted imaging (DWI) sequence is often used in high-field rodent brain MRI due to its immunity to these artefacts. To accurately determine diffusion-tensor or fibre-orientation distribution, high angular resolution diffusion imaging (HARDI) with strong diffusion weighting (b >3000 s/mm2) and at least 30 diffusion-encoding directions are required. However, this results in long image acquisition times unsuitable for live animal imaging. In this study, we describe the optimization of HARDI acquisition parameters at 16.4T using a Stejskal-Tanner sequence with echo-planar imaging (EPI) readout. EPI segmentation and partial Fourier encoding acceleration were applied to reduce the echo time (TE), thereby minimizing signal decay and distortion artefacts while maintaining a reasonably short acquisition time. The final HARDI acquisition protocol was achieved with the following parameters: 4 shot EPI, b = 3000 s/mm2, 64 diffusion-encoding directions, 125×150 μm2 in-plane resolution, 0.6 mm slice thickness, and 2h acquisition time. This protocol was used to image a cohort of adult C57BL/6 male mice, whereby the quality of the acquired data was assessed and diffusion tensor imaging (DTI) derived parameters were measured. High-quality images with high spatial and angular resolution, low distortion and low variability in DTI-derived parameters were obtained, indicating that EPI-DWI is feasible at 16.4T to study animal models of white matter (WM) diseases. 相似文献
37.
38.
Single chain antibody fragments for ocular use produced at high levels in a commercial wheat variety
Brereton HM Chamberlain D Yang R Tea M McNeil S Coster DJ Williams KA 《Journal of biotechnology》2007,129(3):539-546
We are investigating the use of single chain antibody fragments (scFv) in eye drops for diagnosis and treatment of eye diseases. For ocular use, recombinant proteins must be free of bacterial endotoxin that causes inflammation in the eye. We required a means of generating high yields of scFvs with little endotoxin contamination. Using microprojectile bombardment we produced transgenic lines of the commercial wheat variety, Westonia, that express two scFvs that bind to CD4 or CD28 on the surface of rat thymocytes. A high level of expression of active scFv in the range 50-180 microg/g was measured by quantitative flow cytometry in crude extracts made from mature seeds. The levels of expression were stable over four generations of transgenic plants and mature seeds were stored for one year with little loss of scFv activity. Substantial purification of scFv was achieved by immobilised metal affinity chromatography. Compared to bacterial extracts, crude transgenic seed extracts contained only a small amount of endotoxin (150 EU/ml) that will be easily removed by purification. The transgenic wheat lines express functional scFv at levels comparable to production in bacteria and promise to be superior to bacteria for production of scFv pharmaceuticals for ocular use. 相似文献
39.
The first agmatine/cadaverine aminopropyl transferase: biochemical and structural characterization of an enzyme involved in polyamine biosynthesis in the hyperthermophilic archaeon Pyrococcus furiosus 下载免费PDF全文
Cacciapuoti G Porcelli M Moretti MA Sorrentino F Concilio L Zappia V Liu ZJ Tempel W Schubot F Rose JP Wang BC Brereton PS Jenney FE Adams MW 《Journal of bacteriology》2007,189(16):6057-6067
We report here the characterization of the first agmatine/cadaverine aminopropyl transferase (ACAPT), the enzyme responsible for polyamine biosynthesis from an archaeon. The gene PF0127 encoding ACAPT in the hyperthermophile Pyrococcus furiosus was cloned and expressed in Escherichia coli, and the recombinant protein was purified to homogeneity. P. furiosus ACAPT is a homodimer of 65 kDa. The broad substrate specificity of the enzyme toward the amine acceptors is unique, as agmatine, 1,3-diaminopropane, putrescine, cadaverine, and sym-nor-spermidine all serve as substrates. While maximal catalytic activity was observed with cadaverine, agmatine was the preferred substrate on the basis of the k(cat)/K(m) value. P. furiosus ACAPT is thermoactive and thermostable with an apparent melting temperature of 108 degrees C that increases to 112 degrees C in the presence of cadaverine. Limited proteolysis indicated that the only proteolytic cleavage site is localized in the C-terminal region and that the C-terminal peptide is not necessary for the integrity of the active site. The crystal structure of the enzyme determined to 1.8-A resolution confirmed its dimeric nature and provided insight into the proteolytic analyses as well as into mechanisms of thermal stability. Analysis of the polyamine content of P. furiosus showed that spermidine, cadaverine, and sym-nor-spermidine are the major components, with small amounts of sym-nor-spermine and N-(3-aminopropyl)cadaverine (APC). This is the first report in Archaea of an unusual polyamine APC that is proposed to play a role in stress adaptation. 相似文献
40.
Harriet Davies Tom M. Brereton David B. Roy Richard Fox 《Biodiversity and Conservation》2007,16(13):3719-3736
The UK Government has set targets for biodiversity conservation in England based on several indicators, including the status
of protected areas [e.g. Sites of Special Scientific Interest (SSSIs)]. Specifically, the Government aims to achieve favourable
condition [defined by Common Standards Monitoring (CSM)] on 95% by area of SSSIs by 2010. SSSIs are important for threatened
butterflies and management to achieve favourable condition will play a key role in determining future population levels of
these high-profile insects. Because only notified features of SSSIs are considered within CSM, we investigated the level of
notification for three threatened butterflies. We found that these species were notified at only 15–33% of SSSIs where they
occurred; though most site managers did manage for them under broader site conservation objectives. We investigated the relationship
between SSSI condition status and population trend for eight butterfly species of conservation concern to assess the benefit
to butterflies of sites attaining favourable condition status. The majority (80%) of population trends on SSSIs in favourable
condition were positive, suggesting an overall beneficial impact of SSSI management. However, four of the eight species maintained
lower populations at favourable condition SSSIs than at sites in one of the unfavourable condition categories. We suggest
that current condition assessment based chiefly on notified vegetation communities lacks the sensitivity to identify the complex
habitat conditions for these (mosaic) species. As butterflies are good indictors for a wide range of invertebrates, many species
requiring fine-scale habitat heterogeneity may be at risk from the Government’s target. 相似文献