Thymic CCL2 influences induction of T-cell tolerance.

Thymic epithelial cells (TEC) and dendritic cells (DC) play a role in T cell development by controlling the selection of the T cell receptor repertoire. DC have been described to take up antigens in the periphery and migrate into the thymus where they mediate tolerance via deletion of autoreactive T cells, or by induction of natural regulatory T cells. Migration of DC to thymus is driven by chemokine receptors. CCL2, a major ligand for the chemokine receptor CCR2, is an inflammation-associated chemokine that induces the recruitment of immune cells in tissues. CCL2 and CCR2 are implicated in promoting experimental autoimmune encephalomyelitis (EAE), a mouse model for multiple sclerosis. We here show that CCL2 is constitutively expressed by endothelial cells and TEC in the thymus. Transgenic mice overexpressing CCL2 in the thymus showed an increased number of thymic plasmacytoid DC and pronounced impairment of T cell development. Consequently, CCL2 transgenic mice were resistant to EAE. These findings demonstrate that expression of CCL2 in thymus regulates DC homeostasis and controls development of autoreactive T cells, thus preventing development of autoimmune diseases.

Lentiviral-mediated administration of IL-25 in the CNS induces alternative activation of microglia.

Interleukin-25 (IL-25) is the only anti-inflammatory cytokine of the IL-17 family, and it has been shown to be efficacious in inhibiting neuroinflammation. Known for its effects on cells of the adaptive immune system, it has been more recently described to be effective also on cells of the innate immune system, namely macrophages. We used a lentiviral-mediated gene therapy approach to deliver IL-25 to the central nervous system (CNS) in two mouse models of neuroinflammation, entorhinal cortex lesion and experimental autoimmune encephalomyelitis. In both, we found that IL-25 gene therapy was able to modulate CNS myeloid cells, either infiltrating macrophages or resident microglia, towards an anti-inflammatory, tissue-protective phenotype, as testified by the increase in markers such as Arginase-1 (Arg1), Mannose receptor 1 (CD206) and Chitinase 3-like 3 (Ym1). As a consequence, neuroinflammation was partly inhibited and the CNS protected from immune-mediated damage. To our knowledge, this is the first example of M2 shift (alternative activation) induced in vivo on CNS-resident myeloid cells by gene therapy, and may constitute a promising strategy to investigate the potential role of protective microglia in neurological disorders.

Transgressive physiological and transcriptomic responses to light stress in allopolyploid Glycine dolichocarpa (Leguminosae).

Allopolyploidy is often associated with increased photosynthetic capacity as well as enhanced stress tolerance. Excess light is a ubiquitous plant stress associated with photosynthetic light harvesting. We show that under chronic excess light, the capacity for non-photochemical quenching (NPQ(max)), a photoprotective mechanism, was higher in a recently formed natural allotetraploid (Glycine dolichocarpa, designated 'T2') than in its diploid progenitors (G. tomentella, 'D3'; and G. syndetika, 'D4'). This enhancement in NPQ(max) was due to an increase in energy-dependent quenching (qE) relative to D3, combined with an increase in zeaxanthin-dependent quenching (qZ) relative to D4. To explore the genetic basis for this phenotype, we profiled D3, D4 and T2 leaf transcriptomes and found that T2 overexpressed genes of the water-water cycle relative to both diploid progenitors, as well as genes involved in cyclic electron flow around photosystem I (CEF-PSI) and the xanthophyll cycle, relative to D4. Xanthophyll pigments have critical roles in NPQ, and the water-water cycle and CEF-PSI are non-photosynthetic electron transport pathways believed to facilitate NPQ formation. In the absence of CO(2), T2 also exhibited greater quantum yield of photosystem II than either diploid, indicating a greater capacity for non-photosynthetic electron transport. We postulate that, relative to its diploid progenitors, T2 is able to achieve higher NPQ(max) due to an increase in xanthophyll pigments coupled with enhanced electron flow through the water-water cycle and CEF-PSI.

Angiotensin II Type 1 receptor (AT1) signaling in astrocytes regulates synaptic degeneration-induced leukocyte entry to the central nervous system.

Astrocytes are the major cellular component of the blood-brain barrier glia limitans and act as regulators of leukocyte infiltration via chemokine expression. We have studied angiotensin-II receptor Type 1 (AT1) and related NF-κB signaling in astrocytes. Angiotensin II derives from cleavage of angiotensin I by angiotensin converting enzyme (ACE), angiotensin I deriving from angiotensinogen via cleavage by renin. Level of expression of ACE was slightly increased in transgenic mice that express dominant-negative IκBα in astrocytes (GFAP-IκBα-dn mice), whereas angiotensinogen and renin, also constitutively expressed in the CNS, were unaffected by NF-κB inhibition. Leukocytes infiltrate the hippocampus of mice after unilateral stereotactic lesion of afferent perforant path axons in the entorhinal cortex. Upregulation of the chemokine CXCL10 that normally occurs in response to synaptic degeneration in the dentate gyrus following axonal transection was totally abrogated in GFAP-IκBα-dn mice. Whereas angiotensin II was upregulated in microglia and astrocytes in the dentate gyrus post-lesion, AT1 was exclusively expressed on astrocytes. Blocking AT1 with Candesartan led to significant increase in numbers of infiltrating macrophages in the hippocampus 2days post-lesion. Lesion-induced increases in T-cell infiltration and morphologic glial response were unaffected, and the blood-brain barrier remained intact to horseradish peroxidase. These findings show that angiotensin II signaling to astrocytes via AT1 plays an important role in regulation of leukocyte infiltration to the CNS in response to a neurodegenerative stimulus, and identify potential targets for therapies directed at adaptive immune responses in the CNS.

Genetic models for CNS inflammation.

The use of transgenic technology to over-express or prevent expression of genes encoding molecules related to inflammation has allowed direct examination of their role in experimental disease. This article reviews transgenic and knockout models of CNS demyelinating disease, focusing primarily on the autoimmune disease multiple sclerosis, as well as conditions in which an inflammatory response makes a secondary contribution to tissue injury or repair, such as neurodegeneration, ischemia and trauma.

Neuromyelitis optica (NMO)--an autoimmune disease of the central nervous system (CNS).

In the past 10 years, neuromyelitis optica (NMO) has evolved from Devic's categorical clinical description into a broader disease spectrum. Serum IgG antibodies have been identified in NMO patients with the water channel aquaporin-4 (AQP4) as their main target antigen. AQP4 antibodies/NMO-IgG have been shown to be a highly specific and moderately sensitive serum biomarker for NMO. The immunopathology of NMO lesions supports that anti-AQP4 antibodies/NMO-IgG are involved in the pathogenesis of NMO. In vitro studies have demonstrated that human NMO-IgG induce necrosis and impair glutamate transport in astrocytes. Certain ethnic groups, notably of Asian and African origin, seem to be more susceptible to NMO than others. The genetic background for these putative differences is not known, a weak human leucocyte antigen association has been identified. AQP4 gene variants could represent a genetic susceptibility factor for different clinical phenotypes within the NMO spectrum. Experimental models have been described including a double-transgenic myelin-specific B- and T-cell mouse. NMO-like disease has been induced with passive transfer of human anti-AQP4 antibodies to the plasma of mice with pre-established experimental autoimmune encephalomyelitis or by intrathecal administration to naive mice. NMO may be characterized as a channelopathy of the central nervous system with autoimmune characteristics.

CD54/intercellular adhesion molecule 1 and major histocompatibility complex II signaling induces B cells to express interleukin 2 receptors and complements help provided through CD40 ligation.

We have examined signaling roles for CD54 intercellular adhesion molecule 1 and major histocompatibility complex (MHC) II as contact ligands during T help for B cell activation. We used a T helper 1 (Th1)-dependent helper system that was previously shown to be contact as well as interleukin 2 (IL-2) dependent to demonstrate the relative roles of CD54, MHC II, and CD40 signaling in the events leading to the induction of B cell proliferation and responsiveness to IL-2. Paraformaldehyde-fixed activated Th1-induced expression of IL-2R alpha, IL-2R beta, and B7, and upregulated MHC II and CD54 on B cells. Anti-CD54 and MHC II mAbs as well as a CD8 alpha-CD40 ligand (L) soluble construct inhibited both the T-dependent induction of Ig secretion, and B cell phenotypic changes. We then compared the effects of activated Th1 cells with that of cross-linking these molecules. Cross-linking of CD54 and MHC II resulted in the upregulated expression of MHC II and of CD54 and B7, respectively, analogous to the effect of fixed activated Th1 cells. B7 expression was further enhanced by co-cross-linking CD54 and MHC II. Cross-linking of CD40 achieved comparable effects. Strikingly, cross-linking ligation of CD54 and MHC II in the presence of IL-5 induced expression of a functional IL-2R on small resting B cells. By contrast CD40 ligation, which induced B cell proliferation, did not induce IL-2 responsiveness. These data show that CD40 ligation is necessary but may not be sufficient for B cell differentiation and identify CD54 and MHC II as contact ligands that can complement CD40 signaling in the generation of T-dependent B cell responses to IL-2.

Greater loss of axons in primary progressive multiple sclerosis plaques compared to secondary progressive disease.

The pathological substrate of progressive disability in multiple sclerosis is hypothesized to be axonal loss. Differences in the demographic, pathological and radiological features of patients with primary progressive compared with secondary progressive multiple sclerosis raise the question as to whether they actually represent separate clinical entities. So far, large pathological studies comparing axonal damage between primary progressive and secondary progressive multiple sclerosis have not been reported. In this clinico-pathological study we examined the cervical spinal cord in patients with primary and secondary progressive multiple sclerosis. Human cervical spinal cord was derived at autopsy from 54 patients (17 primary progressive, 30 secondary progressive and 7 controls). Tissue was stained immunohistochemically and examined to determine: (i) the number of surviving corticospinal tract axons; (ii) the extent of grey and white matter demyelination; (iii) the degree of inflammation inside and outside of lesions; and (iv) the relationship between demyelination and axonal loss. Associated clinical data was used to calculate expanded disability status scale for each patient preceding death. Motor disability in the primary progressive and secondary progressive groups was similar preceding death. Secondary progressive multiple sclerosis patients showed considerably more extensive demyelination of both the white and grey matter of the cervical spinal cord. The total number of corticospinal axons was equally low in primary progressive and secondary progressive multiple sclerosis groups versus controls. The reduction of axonal density in demyelinated regions compared to normal appearing white matter was significantly more extensive in primary progressive versus secondary progressive patients (33% reduction versus 16% reduction, P < 0.001). These findings suggest axonal loss is the pathological substrate of progressive disability in both primary progressive and secondary progressive multiple sclerosis with a common plaque-centred mechanism. More extensive axonal loss within areas of demyelination in primary progressive multiple sclerosis could explain high levels of axonal loss observed in these patients despite low levels of demyelination.

Regional variations in the extent and pattern of grey matter demyelination in multiple sclerosis: a comparison between the cerebral cortex, cerebellar cortex, deep grey matter nuclei and the spinal cord.

BACKGROUND: Substantial grey matter (GM) demyelination occurs in both the cerebral cortex and spinal cord in multiple sclerosis (MS). GM demyelination also occurs in the cerebellar cortex and the deep GM nuclei of the brain. However, no study has made a direct "within subject" comparison of the extent of GM pathology between these regions. AIM: To examine the extent and pattern of GM demyelination in the motor cortex, cingulate gyrus, cerebellum, thalamus and spinal cord in MS. METHODS: Postmortem study using material from 14 MS cases and three controls. Sections were taken from the five predetermined areas and stained for proteolipid protein. The extent of GM and white matter (WM) demyelination was assessed in each region. RESULTS AND CONCLUSION: Overall, 28.8% of the GM was demyelinated compared with 15.6% of the WM (p<0.001), with demyelination being greater in the GM than in the WM at each of the anatomical sites. There was substantial variation in the extent of demyelination between the different CNS regions. GM demyelination was most extensive in the spinal cord and cerebellum while WM demyelination was most prominent in the spinal cord.

T-cell costimulation: T cells themselves call the shots.

The CD40L molecule expressed by CD4+ regulatory T lymphocytes is known to deliver signals that activate B cells and macrophages. It now appears that CD40L regulates T cells themselves, during both their development and their participation in adaptive immune responses.

Microglia and macrophages express tumor necrosis factor receptor p75 following middle cerebral artery occlusion in mice.

The proinflammatory and potential neurotoxic cytokine tumor necrosis factor (TNF) is produced by activated CNS resident microglia and infiltrating blood-borne macrophages in infarct and peri-infarct areas following induction of focal cerebral ischemia. Here, we investigated the expression of the TNF receptors, TNF-p55R and TNF-p75R, from 1 to 10 days following permanent occlusion of the middle cerebral artery in mice. Using quantitative polymerase chain reaction (PCR), we observed that the relative level of TNF-p55R mRNA was significantly increased at 1-2 days and TNF-p75R mRNA was significantly increased at 1-10 days following arterial occlusion, reaching peak values at 5 days, when microglial-macrophage CD11b mRNA expression was also increased. In comparison, the relative level of TNF mRNA was significantly increased from 1 to 5 days, with peak levels 1 day after arterial occlusion. In situ hybridization revealed mRNA expression of both receptors in predominantly microglial- and macrophage-like cells in the peri-infarct and subsequently in the infarct, and being most marked from 1 to 5 days. Using green fluorescent protein-bone marrow chimeric mice, we confirmed that TNF-p75R was expressed in resident microglia and blood-borne macrophages located in the peri-infarct and infarct 1 and 5 days after arterial occlusion, which was supported by Western blotting. The data show that increased expression of the TNF-p75 receptor following induction of focal cerebral ischemia in mice can be attributed to expression in activated microglial cells and blood-borne macrophages.

CD95-CD95L: can the brain learn from the immune system?

Members of the tumor necrosis factor/nerve growth factor receptor superfamily of cell-surface molecules can play the dual role of mediating either cytotoxicity or cell survival, both in the immune system and in the nervous system. A member of this superfamily, CD95 (also known as ApoI or Fas), was initially identified in the immune system and has been shown to mediate receptor-dependent programmed cell death and to be expressed in the nervous system. In neurodegenerative disorders, CD95-CD95 ligand expression on glial cells might precede receptor-mediated apoptosis by cells of the CNS. It is now being recognized that CD95 signaling by immune cells mediates effects other than apoptosis, such as cell survival and under inflammatory conditions expression of this protein promotes neural-immune interactions. Both neuroscientists and immunologists can contribute to defining the mechanisms underlying these divergent effects and utilize such knowledge to aid understanding of cell death and survival.

The effects of gold and Co-adsorbed carbon on the adsorption and thermal decomposition of acetic acid on Pd{111}.

The growth and annealing behavior of ultrathin Au films on Pd{111} were monitored with scanning tunneling microscopy (STM) and medium energy ion scattering (MEIS). The adsorption of acetic acid on both clean and deliberately carbon-contaminated bimetallic surfaces was investigated with reflection absorption infrared spectroscopy (RAIRS) and temperature-programmed desorption (TPD). We report that the surface chemistry of acetic acid is strongly modified by the presence of Au in the bimetallic surface which acts both to stabilize adsorbed acetate and to decrease the tendency of acetic acid to decompose on adsorption to produce adsorbed carbon. The adsorption of acetic acid at 300 K is found to cause measurable segregation of Pd to the surface for all surface compositions tested.

Pathological study of spinal cord atrophy in multiple sclerosis suggests limited role of local lesions.

Imaging studies in multiple sclerosis have shown that spinal cord atrophy correlates with clinical disability. The pathological substrate of atrophy has not as yet been investigated adequately. In order to determine the cause of spinal cord atrophy in multiple sclerosis, five different sections of the spinal cord were examined histopathologically in 33 controls and 55 multiple sclerosis cases. In the multiple sclerosis cases in each section the total lesion load and the cross-sectional area of the cord were measured. Multiple regression models were estimated, controlling for sex, age, duration of the disease and location of the cord sections. The multiple sclerosis cords were found to be significantly smaller than the controls. The duration of the disease played the most important role in determining cord atrophy. The degree of atrophy varied in different parts of the cord. Individual lesions played a minor role in local atrophy. Our findings suggest that axonal degeneration, possibly caused by the cumulative number of lesions in the brain and cord, or an alternative atrophic process, is responsible for spinal cord atrophy in multiple sclerosis, rather than tissue loss within individual lesions.

IFNgamma enhances microglial reactions to hippocampal axonal degeneration.

Glial reactivity is implicated in CNS repair and regenerative responses. Microglia, the cells responding earliest to axonal injury, produce tumor necrosis factor-alpha (TNFalpha), a cytokine with both cytopathic and neuroprotective effects. We have studied activation of hippocampal microglia to produce TNFalpha in response to transection of perforant path axons in SJL/J mice. TNFalpha mRNA was produced in a transient manner, peaking at 2 d and falling again by 5 d after lesioning. This was unlike other markers of glial reactivity, such as Mac-1 upregulation, which were sustained over longer time periods. Message for the immune cytokine interferon-gamma (IFNgamma) was undetectable, and glial reactivity to axonal lesions occurred as normal in IFNgamma-deficient mice. Microglial responses to lesion-induced neuronal injury were markedly enhanced in myelin basic protein promoter-driven transgenic mice, in which IFNgamma was endogenously produced in hippocampus. The kinetics of TNFalpha downregulation 5 d after lesion was not affected by transgenic IFNgamma, indicating that IFNgamma acts as an amplifier and not an inducer of response. These results are discussed in the context of a regenerative role for TNFalpha in the CNS, which is innately regulated and potentiated by IFNgamma.

Femtosecond energy-transfer processes in the B800-850 light-harvesting complex of Rhodobacter sphaeroides 2.4.1.

The B800-to-B850 energy transfer time in the purified B800-850 light-harvesting complex of Rhodobacter sphaeroides 2.4.1 is determined to be 0.7 ps at room temperature. The electronic state dynamics of the principal carotenoid of this species, spheroidene, are examined, both in vivo and in vitro, by direct femtosecond time-resolved experiments and by fluorescence emission yield studies. Evidence is presented which suggests that carotenoid-to-bacteriochlorophyll energy transfer may occur directly from the initially excited carotenoid S2 state, as well as from the carotenoid S1 state. Further support for this conjecture is obtained from calculations of energy transfer rates from the carotenoid S2 state. Previous measurements of in vivo carotenoid and B800 dynamics are discussed in light of the new results, and currently unresolved issues are described.

The changing face of cytokines in the brain: perspectives from EAE.

There has been a significant increase in the number of cytokines known to exist, over the past few years. This has led to a re-examination of the established roles of cytokines, as the functions of newly identified members are characterized. In this review, we describe how the recent discovery and characterization of interleukin (IL) -23 has led to a re-evaluation of the role of interferon (IFN) gamma and IFNgamma-inducing factors in experimental autoimmune encephalomyelitis (EAE). Recent studies suggest that IFNgamma-secreting T cells, considered the hallmark of EAE, may not be the major detrimental effector cell, and may even have a regulatory function. The impact of this on current understanding of cytokine networks underlying CNS inflammation in EAE is discussed.

Photochemical and Nonphotochemical Fluorescence Quenching Processes in the Diatom Phaeodactylum tricornutum.

Nonphotochemical fluorescence quenching was found to exist in the dark-adapted state in the diatom Phaeodactylum tricornutum. Pretreatment of cells with the uncoupler carbonylcyanide m-chlorophenylhydrazone (CCCP) or with nigericin resulted in increases in dark-adapted minimum and maximum fluorescence yields. This suggests that a pH gradient exists across the thylakoid membrane in the dark, which serves to quench fluorescence levels nonphotochemically. The physiological processes involved in establishing this proton gradient were sensitive to anaerobiosis and antimycin A. Based on these results, it is likely that this energization of the thylakoid membrane is due in part to chlororespiration, which involves oxygen-dependent electron flow through the plastoquinone pool. Chlororespiration has been shown previously to occur in diatoms. In addition, we observed that cells treated with 3-(3,4-dichlorophenyl)-1,1-dimethylurea exhibited very strong nonphotochemical quenching when illuminated with actinic light. The rate and extent of this quenching were light-intensity dependent. This quenching was reversed upon addition of CCCP or nigericin and was thus due primarily to the establishment of a pH gradient across the thylakoid membrane. Preincubation of cells with CCCP or nigericin or antimycin A completely abolished this quenching. Cyclic electron transport processes around photosystem I may be involved in establishing this proton gradient across the thylakoid membrane under conditions where linear electron transport is inhibited. At steady state under normal physiological conditions, the qualitative changes in photochemical and nonphotochemical fluorescence quenching at increasing photon flux densities were similar to those in higher plants. However, important quantitative differences existed at limiting and saturating intensities. Dissimilarities in the factors that regulate fluorescence quenching mechanisms in these organisms may account for these differences.

The Effects of Excess Irradiance on Photosynthesis in the Marine Diatom Phaeodactylum tricornutum.

The response of Phaeodactylum tricornutum to excess light was remarkably similar to that observed in higher plants and green algae and was characterized by complex changes in minimal fluorescence yields of fully dark-adapted samples and declines in maximum variable fluorescence levels and oxygen evolution rates. In our study the parallel decreases in the effective rate constant for photosystem II (PSII) photochemistry, the variable fluorescence yield of a dark-adapted sample, and light-limited O2 evolution rates after short (0-10 min) exposures to photoinhibitory conditions could not be attributed to damage or down-regulation of PSII reaction centers. Instead, these changes were consistent with the presence of nonphotochemical quenching of PSII excitation energy in the antennae. This quenching was analogous to that component of nonphotochemical quenching studied in higher plants that is associated with photoinhibition of photosynthesis and/or processes protecting against photoinhibition in that it did not relax readily in the dark and persisted in the absence of a bulk transthylakoid proton gradient. The quenching was most likely associated with photoprotective processes in the PSII antenna that reduced the extent of photoinhibitory damage, particularly after longer exposures. Our results suggest that a large population of damaged, slowly recovering PSII centers did not form in Phaeodactylum even after 60 min of exposure to excess actinic light.