Coexistence of structurally similar but electronically distinct isomers of delocalized cation radicals as a basis for the development of functional materials.

Certain low-symmetry aromatic molecules with a small HOMO/HOMO-1 energy gap might co-exist as electronic isomers in their cation radical states that differ only in bond lengths yet have distinct optical and electronic properties. These electronic isomers are sensitive to the environment and might be used for the development of novel functional materials.

The heparan sulfate proteoglycan agrin contributes to barrier properties of mouse brain endothelial cells by stabilizing adherens junctions.

Barrier characteristics of brain endothelial cells forming the blood-brain barrier (BBB) are tightly regulated by cellular and acellular components of the neurovascular unit. During embryogenesis, the accumulation of the heparan sulfate proteoglycan agrin in the basement membranes ensheathing brain vessels correlates with BBB maturation. In contrast, loss of agrin deposition in the vasculature of brain tumors is accompanied by the loss of endothelial junctional proteins. We therefore wondered whether agrin had a direct effect on the barrier characteristics of brain endothelial cells. Agrin increased junctional localization of vascular endothelial (VE)-cadherin, ß-catenin, and zonula occludens-1 (ZO-1) but not of claudin-5 and occludin in the brain endothelioma cell line bEnd5 without affecting the expression levels of these proteins. This was accompanied by an agrin-induced reduction of the paracellular permeability of bEnd5 monolayers. In vivo, the lack of agrin also led to reduced junctional localization of VE-cadherin in brain microvascular endothelial cells. Taken together, our data support the notion that agrin contributes to barrier characteristics of brain endothelium by stabilizing the adherens junction proteins VE-cadherin and ß-catenin and the junctional protein ZO-1 to brain endothelial junctions.

Loss of astrocyte polarization upon transient focal brain ischemia as a possible mechanism to counteract early edema formation.

Brain edema is the main cause of death from brain infarction. The polarized expression of the water channel protein aquaporin-4 (AQP4) on astroglial endfeet surrounding brain microvessels suggests a role in brain water balance. Loss of astrocyte foot process anchoring to the basement membrane (BM) accompanied by the loss of polarized localization of AQP4 to astrocytic endfeet has been shown to be associated with vasogenic/extracellular edema in neuroinflammation. Here, we asked if loss of astrocyte polarity is also observed in cytotoxic/intracellular edema following focal brain ischemia after transient middle cerebral artery occlusion (tMCAO). Upon mild focal brain ischemia, we observed diminished immunostaining for the BM components laminin α4, laminin α2, and the proteoglycan agrin, in the core of the lesion, but not in BMs in the surrounding penumbra. Staining for the astrocyte endfoot anchorage protein ß-dystroglycan (DG) was dramatically reduced in both the lesion core and the penumbra, and AQP4 and Kir4.1 showed a loss of polarized localization to astrocytic endfeet. Interestingly, we observed that mice deficient for agrin expression in the brain lack polarized localization of ß-DG and AQP4 at astrocytic endfeet and do not develop early cytotoxic/intracellular edema following tMCAO. Taken together, these data indicate that the binding of DG to agrin embedded in the subjacent BM promotes polarized localization of AQP4 to astrocyte endfeet. Reduced DG protein levels and redistribution of AQP4 as observed upon tMCAO might therefore counteract early edema formation and reflect a beneficial mechanism operating in the brain to minimize damage upon ischemia.

Porcine circovirus type 2 DNA influences cytoskeleton rearrangements in plasmacytoid and monocyte-derived dendritic cells.

Functional disruption of dendritic cells (DC) is an important strategy for viral pathogens to evade host defences. In this context, porcine circovirus type 2 (PCV2), a single-stranded DNA virus, impairs plasmacytoid DC (pDC) and conventional DC activation by certain viruses or Toll-like receptor (TLR) ligands. This inhibitory capacity is associated with the viral DNA, but the impairment does not affect all signalling cascades; TLR7 ligation by small chemical molecules will still induce interleukin-6 (IL-6) and tumour necrosis factor-α secretion, but not interferon-α or IL-12. In this study, the molecular mechanisms by which silencing occurs were investigated. PP2, a potent inhibitor of the Lyn and Hck kinases, produced a similar profile to the PCV2 DNA interference with cytokine secretion by pDC, efficiently inhibiting cell activation induced through TLR9, but not TLR7, ligation. Confocal microscopy and cytometry analysis strongly suggested that PCV2 DNA impairs actin polymerization and endocytosis in pDC and monocyte-derived DC, respectively. Altogether, this study delineates for the first time particular molecular mechanisms involved in PCV2 interference with DC danger recognition, which may be responsible for the virus-induced immunosuppression observed in infected pigs.

Loss of astrocyte polarity marks blood-brain barrier impairment during experimental autoimmune encephalomyelitis.

In multiple sclerosis (MS), and its animal model experimental autoimmune encephalomyelitis (EAE), dysfunction of the blood-brain barrier (BBB) leads to edema formation within the central nervous system. The molecular mechanisms of edema formation in EAE/MS are poorly understood. We hypothesized that edema formation is due to imbalanced water transport across the BBB caused by a disturbed crosstalk between BBB endothelium and astrocytes. Here, we demonstrate at the light microscopic and ultrastructural level, the loss of polarized localization of the water channel protein aquaporin-4 (AQP4) in astrocytic endfeet surrounding microvessels during EAE. AQP4 was found to be redistributed over the entire astrocytic cell surface and lost its arrangement in orthogonal arrays of intramembranous particles as seen in the freeze-fracture replica. In addition, immunostaining for the astrocytic extracellular matrix receptor beta-dystroglycan disappeared from astroglial membranes in the vicinity of inflammatory cuffs, whereas immunostaining for the dystroglycan ligands agrin and laminin in the perivascular basement membrane remained unchanged. Our data suggest that during EAE, loss of beta-dystroglycan-mediated astrocyte foot process anchoring to the basement membrane leads to loss of polarized AQP4 localization in astrocytic endfeet, and thus to edema formation in EAE.

Cellular adaptive immune response against porcine circovirus type 2 in subclinically infected pigs.

BACKGROUND: Porcine circovirus type 2 (PCV2) is a dominant causative agent of postweaning multisystemic wasting syndrome (PMWS), a multifactorial disease complex with putative immunosuppressive characteristics. Little is known about adaptive PCV2-specific immune responses in infected pigs. Therefore, the T and B cell responses following PCV2 infection in 3-week old SPF piglets infected with PCV2 or PCV2 plus porcine parvovirus (PPV) were studied. RESULTS: All animals were asymptomatically infected. At 7 days post infection (d p.i.), B lymphocyte and T lymphocyte numbers decreased in the dual infected, but not in the single infected piglets. At this time point a transient PCV2 viraemia was noted in the PCV2 infected groups. Antibodies against the infecting virus were detectable at day 24-28 p.i. for anti-PCV2 antibodies and at day 10 p.i. for anti-PPV antibodies, with no apparent influence of PCV2 on the early PPV antibody development. In the animals infected with PPV alone, IFN-gamma secreting cells (SC) that were not specific for PCV2 were detected by ELISPOT assay at day 7 p.i. Interestingly, this response was absent in the PCV2/PPV dual infected animals. PCV2-specific IFN-gamma SC were observed in the PCV2/PPV infected group at 7 d p.i. and in the PCV2 single infected group at 21 d p.i. A reduction in the numbers of IFN-gamma SC was observed following anti-CD4 and anti-CD8 antibody treatment, suggesting roles for both CD4+ and CD8+ T cells in the response against PCV2 infection. This was supported by an observed increase in the percentage of IFN-gamma positive CD8hi cytotoxic T cells as well as IFN-gamma positive CD8-/low helper T cells after PCV2 in vitro re-stimulation. CONCLUSIONS: Infection of weaned SPF piglets with PCV2 alone or combined with PPV does not induce disease and in both cases a relatively slow anti-PCV2 antibody response and weak T lymphocyte responses were found. Knowledge on such immunological characteristics is important for both PCV2 pathogenesis and vaccination.

Responsiveness of fibrocytes to toll-like receptor danger signals.

Circulating myeloid cells such as plasmacytoid dendritic cells (pDC), blood DC and monocytes act as blood sentinels detecting invading pathogens through a large repertoire of expression of toll-like receptors (TLRs). Activation of these receptors is crucial to detect invading pathogens by the innate immune system. In the present work, we analysed the TLR responsiveness of fibrocytes, a blood-derived cell type of myeloid origin. Fibrocytes efficiently responded to TLR2, TLR4, and TLR7 ligands as well as to poly (I:C) or viral stimulation by producing high amount of interleukin-6. Upon virus infection of fibrocytes, IFN type I was also induced. When compared to pDC or Flt3 ligand-derived DC, fibrocytes produced 5 times and 60 times more IL-6, respectively. This response was associated with a rapid and efficient translocation of the NF-kappaB transcription factor. Analysis of the expression and functionality of TLR7 in peripheral blood leukocyte subpopulations suggested that this receptor is expressed and functional in a CD163(+) monocytic cell subpopulation containing the fibrocyte precursors. Considering the rapid entry of fibrocytes into wounds, this efficient responsiveness to TLR danger signals, reflects a potentially important role of these cells in the first line of defence against pathogen invasion following traumata.

Mouse Model of Reversible Intestinal Inflammation.

Current therapies to treat inflammatory bowel disease by dampening excessive inflammatory immune responses have had limited success ( Reinisch et al., 2011 ; Rutgeerts et al., 2005 ; Sandborn et al., 2012 ). To develop new therapeutic interventions, there is a need for better understanding of the mechanisms that are operative during mucosal healing (Pineton de Chambrun et al., 2010 ). To this end, a reversible model of colitis was developed in which colitis induced by adoptive transfer of naïve CD4+ CD45RBhi T cells in lymphopenic mice can be reversed through depletion of colitogenic CD4+ T cells ( Brasseit et al., 2016 ).

Porcine circovirus type 2 displays pluripotency in cell targeting.

Porcine circovirus type 2 (PCV2) is the causative agent of a multifactorial disease associated with immunocompromisation and co-infections. In vivo, viral DNA and antigens are found in monocytic, epithelial and endothelial cells. Of these, PCV2 replication has only been studied in monocytic cells, in which little or no replication was identified. Accordingly, PCV2 infection was studied in the endothelial cell line PEDSV.15, aortic endothelial cells, gut epithelial cells, fibrocytes and dendritic cells (DC). In all cells except DC PCV2 replication was detectable, with an increase in the levels of capsid and replicase protein. Variations in endocytic activity, virus binding and uptake did not relate to the replication efficiency in a particular cell. Furthermore, replication did not correlate to cell proliferation, although a close association of viral proteins with chromatin in dividing cells was observed. No alteration in the division rate of PCV2-infected cultures was measurable, relating to replicase expression in only a small minority of the cells. In conclusion, the broad cell targeting of PCV2 offers an explanation for its widespread tissue distribution.