CCR5Δ32 genotype leads to a Th2 type directed immune response in ESRD patients

Background

In patients with end stage renal disease (ESRD) we observed protection from inflammation-associated mortality in CCR5Δ32 carriers, leading to CCR5 deficiency, suggesting impact of CCR5Δ32 on inflammatory processes. Animal studies have shown that CCR5 deficiency is associated with a more pronounced Th2 type immune response, suggesting that in human CCR5Δ32 carriers the immune response may be more Th2 type directed. So, in the present study we determined the Th1-Th2 type directed immune response in ESRD patients carrying and not carrying the CCR5Δ32 genetic variant after stimulation.

Methodology/Principal Findings

We tested this hypothesis by determining the levels of IFN-γ and IL-4 and the distribution of Th1, Th2 and Th17 directed circulating CD4+ and CD8+ T cells and regulatory T cells (Tregs) after stimulation in ESRD patients with (n = 10) and without (n = 9) the CCR5Δ32 genotype. The extracellular levels of IFN-γ and IL-4 did not differ between CCR5Δ32 carriers and non carriers. However, based on their intracellular cytokine profile the percentages IL-4 secreting CD4+ and CD8+ T cells carrying the CCR5Δ32 genotype were significantly increased (p = 0.02, respectively p = 0.02) compared to non carriers, indicating a more Th2 type directed response. Based on their intracellular cytokine profile the percentages IFN-γ and IL-17 secreting T cells did not differ between carriers and non-carriers nor did the percentage Tregs, indicating that the Th1, Th17 and T regulatory response was not affected by the CCR5Δ32 genotype.

Conclusions/Significance

This first, functional human study shows a more pronounced Th2 type immune response in CCR5Δ32 carriers compared to non carriers. These differences may be involved in the previously observed protection from inflammation-associated mortality in ESRD patients carrying CCR5Δ32.     

The crucial role of particle surface reactivity in respirable quartz-induced reactive oxygen/nitrogen species formation and APE/Ref-1 induction in rat lung

Persistent inflammation and associated excessive oxidative stress have been crucially implicated in quartz-induced pulmonary diseases, including fibrosis and cancer. We have investigated the significance of the particle surface reactivity of respirable quartz dust in relation to the in vivo generation of reactive oxygen and nitrogen species (ROS/RNS) and the associated induction of oxidative stress responses in the lung. Therefore, rats were intratracheally instilled with 2 mg quartz (DQ12) or quartz whose surface was modified by either polyvinylpyridine-N-oxide (PVNO) or aluminium lactate (AL). Seven days after instillation, the bronchoalveolar lavage fluid (BALF) was analysed for markers of inflammation (total/differential cell counts), levels of pulmonary oxidants (H₂O₂, nitrite), antioxidant status (trolox equivalent antioxidant capacity), as well as for markers of lung tissue damage, e.g. total protein, lactate dehydrogenase and alkaline phosphatase. Lung homogenates as well as sections were investigated regarding the induction of the oxidative DNA-lesion/oxidative stress marker 8-hydroxy-2''-deoxyguanosine (8-OHdG) using HPLC/ECD analysis and immunohistochemistry, respectively. Homogenates and sections were also investigated for the expression of the bifunctional apurinic/apyrimidinic endonuclease/redox factor-1 (APE/Ref-1) by Western blotting and immunohistochemistry. Significantly increased levels of H₂O₂ and nitrite were observed in rats treated with non-coated quartz, when compared to rats that were treated with either saline or the surface-modified quartz preparations. In the BALF, there was a strong correlation between the number of macrophages and ROS, as well as total cells and RNS. Although enhanced oxidant generation in non-coated DQ12-treated rats was paralleled with an increased total antioxidant capacity in the BALF, these animals also showed significantly enhanced lung tissue damage. Remarkably however, elevated ROS levels were not associated with an increase in 8-OHdG, whereas the lung tissue expression of APE/Ref-1 protein was clearly up-regulated. The present data provide further in vivo evidence for the crucial role of particle surface properties in quartz dust-induced ROS/RNS generation by recruited inflammatory phagocytes. Our results also demonstrate that quartz dust can fail to show steady-state enhanced oxidative DNA damage in the respiratory tract, in conditions were it elicits a marked and persistent inflammation with associated generation of ROS/RNS, and indicate that this may relate to compensatory induction of APE/Ref-1 mediated base excision repair.     

Chaetochromones A – C,Three New Polyketides from Mangrove Plant Derived Endophytic Fungus Phomopsis sp. SCSIO 41006

Three new polyketides, chaetochromones A – C ( 1  –  3 ), together with a chromone ( 4 ), were isolated from the ethyl acetate extract of mangrove‐derived fungus Phomopsis sp. SCSIO 41006. Their structures were elucidated by means of spectroscopic techniques (UV, IR, MS, 1D‐ and 2D‐NMR). The absolute configurations of the new compounds were established by CD data.     

Normalization,Social Bonding,and Emotional Support— A Dog’s Effect within a Prison Workshop for Women

A fundamental tenet of the Danish Prison System is the principle of normalization, meaning that prisons are organized in such a way that the conditions within the walls more or less resemble the conditions outside them. When prison conditions differ as little as possible from normal daily life on the outside, it underpins rehabilitation efforts. To have contact with animals during incarceration can be seen as a part of normalization and thus contributing to rehabilitation. However, in Danish prisons, animal-based programs are not usually offered, nor are prisoners allowed to keep a pet. In an open prison, a women’s prison workshop was established in 2014. In response to prisoners’ requests for contact with animals, an employee brought her own dog during the hours of the workshop, from Monday to Friday. In Denmark and the other Scandinavian countries, not much attention has been given to the effect of the human–animal bond within prisons. To document how well it might work, qualitative methods for data collection were used, including interviews with incarcerated women (n = 12) and staff (n = 3) and participant observation (67 hours) within the women’s workshop. The dog contributed to normalize the prison setting, and participants revealed that the dog improved social relations between inmates and between staff and inmates. Finally, the dog provided comfort to the incarcerated women when they had to deal with difficult personal feelings. A recommendation for policy makers and prison officials arising from this study is that animals should be a normal part of the prison setting.     

The cellular modifier MOAG‐4/SERF drives amyloid formation through charge complementation

While aggregation‐prone proteins are known to accelerate aging and cause age‐related diseases, the cellular mechanisms that drive their cytotoxicity remain unresolved. The orthologous proteins MOAG‐4, SERF1A, and SERF2 have recently been identified as cellular modifiers of such proteotoxicity. Using a peptide array screening approach on human amyloidogenic proteins, we found that SERF2 interacted with protein segments enriched in negatively charged and hydrophobic, aromatic amino acids. The absence of such segments, or the neutralization of the positive charge in SERF2, prevented these interactions and abolished the amyloid‐promoting activity of SERF2. In protein aggregation models in the nematode worm Caenorhabditis elegans, protein aggregation and toxicity were suppressed by mutating the endogenous locus of MOAG‐4 to neutralize charge. Our data indicate that MOAG‐4 and SERF2 drive protein aggregation and toxicity by interactions with negatively charged segments in aggregation‐prone proteins. Such charge interactions might accelerate primary nucleation of amyloid by initiating structural changes and by decreasing colloidal stability. Our study points at charge interactions between cellular modifiers and amyloidogenic proteins as potential targets for interventions to reduce age‐related protein toxicity.     

Overexpression of lunatic fringe does not affect epithelial cell differentiation in the developing mouse lung

The Notch/Notch-ligand pathway regulates cell fate decisions and patterning in various tissues. Several of its components are expressed in the developing lung, suggesting that this pathway is important for airway cellular patterning. Fringe proteins, which modulate Notch signaling, are crucial for defining morphogenic borders in several organs. Their role in controlling cellular differentiation along anterior-posterior axis of the airways is unknown. Herein, we report the temporal-spatial expression patterns of Lunatic fringe (Lfng) and Notch-regulated basic helix-loop-helix factors, Hes1 and Mash-1, during murine lung development. Lfng was only expressed during early development in epithelial cells lining the larger airways. Those epithelial cells also expressed Hes1, but at later gestation Hes1 expression was confined to epithelium lining the terminal bronchioles. Mash-1 displayed a very characteristic expression pattern. It followed neural crest migration in the early lung, whereas at later stages Mash-1 was expressed in lung neuroendocrine cells. To clarify whether Lfng influences airway cell differentiation, Lfng was overexpressed in distal epithelial cells of the developing mouse lung. Overexpression of Lfng did not affect spatial or temporal expression of Hes1 and Mash-1. Neuroendocrine CGRP and protein gene product 9.5 expression was not altered by Lfng overexpression. Expression of proximal ciliated (beta-tubulin IV), nonciliated (CCSP), and distal epithelial cell (SP-C, T1alpha) markers also was not influenced by Lfng excess. Overexpression of Lfng had no effect on mesenchymal cell marker (alpha-sma, vWF, PECAM-1) expression. Collectively, the data suggest that Lunatic fringe does not play a significant role in determining cell fate in fetal airway epithelium.     

The activity of the TRP-like channel depends on its expression system

The Drosophila light activated TRP and TRPL channels have been a model for TRPC channel gating. Several gating mechanisms have been proposed following experiments conducted on photoreceptor and tissue cultured cells. However, conclusive evidence for any mechanism is still lacking. Here, we show that the Drosophila TRPL channel expressed in tissue cultured cells is constitutively active in S2 cells but is silent in HEK cells. Modulations of TRPL channel activity in different expression system by pharmacology or specific enzymes, which change the lipid content of the plasma membrane, resulted in conflicting effects. These findings demonstrate the difficulty in elucidating TRPC gating, as channel behavior is expression system dependent. However, clues on the gating mechanism may arise from understanding how different expression systems affect TRPC channel activation.     

Fluorescent cell barcoding as a tool to assess the age-related development of intracellular cytokine production in small amounts of blood from infants

Fluorescent Cell Barcoding (FCB) is a flow cytometric technique which has been used for assessing signaling proteins. This FCB technique has the potential to be applied in other multiparameter analyses. Since data on antigen (Ag)-specific T-cell immune responses, like intracellular cytokine production, are still lacking in infants because limited blood volumes can be obtained for analysis, the FCB technique could be very useful for this purpose. The objectives of this study were to modify the FCB method to be able to measure multiple Ag-specific cytokine reponses in T-cells upon simultaneous stimulation by various antigens and mitogens in small amounts of blood and to investigate the cytokine pattern of T-cell subsets in healthy infants aged six and twelve months. Blood samples, collected from 20 healthy infants aged six and twelve months, were stimulated in vitro with the antigens: phorbol-myristate-acetate (PMA), purified-protein-derivative (PPD), Tetanus-toxoid (TT), Staphylococcal-enterotoxin-B (SEB), and phytohemagglutinin (PHA). Each stimulus was barcoded by labelling with different intensities of fluorescent cell barcoding (FCB) markers. Intracellular production of interleukin-2, interferon-gamma, and tumor necrosis factor-alpha was measured simultaneously in just one blood sample of 600 μl whole blood. Significant age-related differences in cytokine production were shown for PMA, PHA, and TT in CD4(+) T-cells, and for PMA, PHA, SEB, and TT in CD8(+) T-cells. The intracellular cytokine production by CD4(+) and CD8(+) T-cells was higher at twelve months compared to six months of age for all antigens, except for PMA, which was lower at the age of twelve months. Based on the consistency in both T-cell subsets, we conclude that the new FCB method is a promising tool to investigate the age-related development of intracellular cytokine production in infants.