Polymorphonuclear leucocytes selectively produce anti-inflammatory interleukin-1 receptor antagonist and chemokines, but fail to produce pro-inflammatory mediators

The role of neutrophils in the immune response has long been regarded as mainly phagocytic, but recent publications have indicated the production of several cytokines by polymorphonuclear leucocytes (PMN). The results of the individual reports, however, vary considerably. In this study, we established a cytokine profile of pure human neutrophils and demonstrated that minor contamination of peripheral blood mononuclear cells (PBMCs) in PMN preparations can lead to false-positive results. In our hands, peripheral blood PMN fail to produce the pro-inflammatory cytokines interleukin (IL)-1beta, IL-6 and tumour necrosis factor-alpha (TNF-alpha). Instead, they secrete large amounts of the chemokine IL-8 and the anti-inflammatory IL-1 receptor antagonist (IL-1ra). Additionally, PMN preparations of a high purity show production of the chemokines macrophage inflammatory protein (MIP)-1alpha, MIP-1beta and growth-related oncogene-alpha (GRO-alpha), as well as macrophage colony-stimulating factor (M-CSF). The neutrophil therefore represents a novelty by producing the antagonist of IL-1beta (i.e. IL-1ra) in the absence of IL-1beta itself. To support our results, we differentiated stem cells from human cord blood into PMN and monocytes, respectively. These in vitro-differentiated PMN showed the same cytokine profile as peripheral blood PMN lacking IL-1beta, while differentiated monocytes produced the expected IL-1beta in addition to IL-1ra. The clear anti-inflammatory nature of their cytokine profile enables PMN to antagonize pro-inflammatory signals in experimental conditions. It is therefore possible that PMN play a key role in immune regulation by counteracting a dysregulation of the inflammatory process. Clinical studies, in which administration of recombinant G-CSF had a favourable effect on the outcome of severe infections and even sepsis without worsening inflammation, could thus be explained by our results.     

Interleukin 27 negatively regulates the development of interleukin 17-producing T helper cells during chronic inflammation of the central nervous system

Studies have focused on the events that influence the development of interleukin 17 (IL-17)-producing T helper cells (T(H)-17 cells) associated with autoimmunity, such as experimental autoimmune encephalitis, but relatively little is known about the cytokines that antagonize T(H)-17 cell effector responses. Here we show that IL-27 receptor-deficient mice chronically infected with Toxoplasma gondii developed severe neuroinflammation that was CD4+ T cell dependent and was associated with a prominent IL-17 response. In vitro, treatment of naive primary T cells with IL-27 suppressed the development T(H)-17 cells induced by IL-6 and transforming growth factor-beta, which was dependent on the intracellular signaling molecule STAT1 but was independent of inhibition of IL-6 signaling mediated by the suppressor protein SOCS3. Thus IL-27, a potent inhibitor of T(H)-17 cell development, may be a useful target for treating inflammatory diseases mediated by these cells.     

Proteomics reveals a global phenotypic shift of NK cells in HCV patients treated with direct-acting antivirals

Chronic hepatitis C virus (HCV) infections compromise natural killer (NK)-cell immunity. Direct-acting antivirals (DAA) effectively eliminate HCV, but the long-term effects on NK cells in cured patients are debated. We conducted a proteomic study on CD56⁺ NK cells of chronic HCV-infected patients before and 1 year after DAA therapy. Donor-variation was observed in NK-cell proteomes of HCV-infected patients, with 46 dysregulated proteins restored after DAA therapy. However, 30% of the CD56⁺ NK-cell proteome remained altered 1 year post-therapy, indicating a phenotypic shift with low donor-variation. NK cells from virus-negative cured patients exhibited global regulation of RNA-processing and pathways related to “stimuli response”, “chemokine signaling”, and “cytotoxicity regulation”. Proteomics identified downregulation of vesicle transport components (CD107a, COPI/II complexes) and altered receptor expression profiles, indicating an inhibited NK-cell phenotype. Yet, activated NK cells from HCV patients before and after therapy effectively upregulated IFN-γ and recruited CD107a. Conversely, reduced surface expression levels of Tim-3 and 2B4 were observed before and after therapy. In conclusion, this study reveals long-term effects on the CD56⁺ NK-cell compartment in convalescent HCV patients 1 year after therapy, with limited abundance of vesicle transport complexes and surface receptors, associated with a responsive NK-cell phenotype.     

Expression of human immunodeficiency virus type 1gag gene using genetically engineered herpes simplex virus type 1 recombinants

Infectious herpes simplex virus type 1 (HSV-1) recombinants were constructed by inserting the cDNA sequence of the human immunodeficiency virus type 1 (HIV-1)gag gene (from nucleotide position 675 [SacI] to 3859 [Asp 718] of the cDNA sequences of HIV-1 strain BH-10) within the DNA sequences of theBamHI DNA fragment B of the genome of an apathogenic HSV-1 strain HFEM. This HSV-1 strain possesses a 4.1-kbp deletion within theBamHI DNA fragment B between 0.762 and 0.789 map units of the viral genome, which allows the insertion of at least 4 kbp of foreign genetic material into this particular region. The DNA sequences of the immediate early promoter (IE4) of HSV-1 that were inserted upstream from thegag gene were used as a promoter. The screening of 205 virus stocks derived from individual plaques revealed that 46 recombinant viruses harbor HIV-1gag-specific DNA sequences. However, it was found that only six of the recombinant viruses are able to express thegag gene product of HIV-1. This indicates that the ratio of the positive recombination events is about 2.9%.     

Circulating leucocyte subpopulations in sedentary subjects following graded maximal exercise with hypoxia

Summary Ten healthy sedentary subjects [age, 27.5 (SD 3.5) years; height, 180 (SD 5) cm; mass, 69.3 (SD 6.3) kg] performed two periods of maximal incremental graded cycle ergometer exercise in a supine position. Randomly ordered and using an open spirometric system, one exercise was carried out during normoxia [maximal oxygen consumption ( O_2max)=38.6 (SD 3.5) ml·min^–1·kg^–1; maximal blood lactate concentration, 9.86 (SD 1.85) mmol·l^–1; test duration, 22.6 (SD 2.7) min], the other during hypoxia [ O_2max=33.2 (SD 3.2) ml·min^–1· kg^–1; maximal blood lactate concentration, 10.38 (SD 2.02) mmol·l^–1; test duration, 19.7 (SD 2.8) min]. At rest, immediately (0 p) and 60 min (60 p) after exercise, counts of leucocyte subpopulations (flow cytometry), cortisol and catecholamine concentrations were determined. At 0 p in contrast to normoxia, during hypoxia there was no significant increase of granulocytes. There were no significant differences between normoxia and hypoxia in the increases from rest to 0 p in counts of monocytes, total lymphocytes and lymphocyte subpopulations [clusters of differentiation (CD), CD3⁺, CD4⁺CD45RO^–, CD4⁺CD45RO⁺, CD8⁺CD45RO^–, CD8⁺CD45RO⁺, CD3⁺HLA-DR⁺, CD3^–CD16/CD56⁺, CD3⁺CD16/CD56⁺, CD 19⁺] as well as adrenaline, noradrenaline and cortisol concentrations. The counts of CD3 ^–CD16/CD56⁺-and CD8 ⁺CD45RO⁺-cells increased most. At 60 p, CD3^–CD16/CD56⁺ and CD3⁺CD16/CD56⁺-cell counts were below pre-exercise levels and under hypoxia slightly but significantly lower than under normoxia. We concluded that the exercise-induced mobilization and redistribution of most leucocyte and lymphocyte subpopulations were unimpaired under acute hypoxia at sea level. Reduced increases of granulocyte counts during the study and reduced cell numbers of natural killer cells and cytotoxic, not major histocompatibility complex-restricted T-cells, only indicated marginal effects on the immune system.     

Synthesis and characterization of poly[2,3,9,10,16,17,23,24-octakis(dodecyloxycarbonyl)phthalocyaninatogermoxane]

In this article we describe the preparation of poly[2,3,9,10,16,17,23,24-octakis(dodecyloxycarbonyl)phthalocyaninatogermoxane] by thermal polycondensation of 2,3,9,10,16,17,23,24-octakis(dodecyloxycarbonyl)phthalocyaninatogermanium dihydroxide, and its characterization. The average molar mass of the resulting polymer was determined. High molar masses are assessible, e.g., M?_w = 360000 g/mol, M?_n = 140000 g/mol, at moderate temperature for the polycondensation (200°C). The polymer shows a liquid-crystalline phase in an extremely large temperature range. Furthermore, Langmuir-Blodgett films (LB films) of the polymer and of the monomer were prepared and characterized. In LB films the phthalocyanine rings are preferentially oriented normal to the dipping direction.     

Chemical shift sodium imaging in a mouse model of thromboembolic stroke at 9.4 T

Purpose:

To estimate changes in the ²³Na density and in the ²³Na relaxation time T₂* in the anatomically small murine brain after stroke.

Materials and Methods:

Three‐dimensional acquisition weighted chemical shift imaging at a resolution of 0.6 × 0.6 × 1.2 mm³ was used for sodium imaging and relaxation parameter mapping. In vivo measurements of the mouse brain (n = 4) were performed 24 hours after stroke, induced by microinjection of purified murine thrombin into the right middle cerebral artery. The measurement time was 14 minutes in one mouse and 65 minutes in the other three. An exponential fit estimation of the free induction decay was calculated for each voxel enabling the reconstruction of locally resolved relaxation parameter maps.

Results:

The infarcted areas showed an increase in sodium density between 160% and 250%, while the T₂* relaxation time increased by 5%–72% compared to unaffected contralateral brain tissue.

Conclusion:

²³Na chemical shift imaging at a resolution of 0.6 × 0.6 × 1.2 mm³ enabled sodium imaging of the anatomical small mouse brain and the acquired data allowed calculating relaxation parameter maps and hence a more exact evaluation of sodium signal changes after stroke. J. Magn. Reson. Imaging 2011;. © 2011 Wiley‐Liss, Inc.     

Stereotactic radiotherapy for choroidal melanomas by means of HybridArc™

Purpose

Stereotactic radiotherapy (SRT) is suitable to treat ocular tumours. The optimal beam geometry for SRT, however, has not been defined. Here we evaluate a combination technique with dynamic conformal arcs (DCAs) and intensity-modulated static fields (IMRT), known as HybridArc? (HA).

Methods

For the first consecutive 25 cases with choroidal melanomas with volumes of 0.02 to 1.18?cm³ treated with 50?Gy in five fractions, the results with respect to dose conformity, homogeneity, and dose distributions were summarised. To describe the dose distribution at the planning target volume (PTV) boundary, we defined a spatially averaged dose gradient (SADG) and compared it with Paddick’s gradient index (GI). We made dosimetric comparisons between HA and other irradiation techniques.

Results

The PTVs ranged from 0.42 to 3.37?cm³. The conformity index (CI) was 1.25?±?0.15, and the homogeneity index (HI) 0.08?±?0.02. The SADG was (?3.5?±?0.5) Gy/mm or (?7.0?±?1.0) %/mm between the isodose levels 95 and 20%; local minima reached ?11.5?Gy/mm or ?22.9%/mm. The coefficient of determination for a nonlinear regression of GI on SADG was 0.072. After a median follow-up time of 19.6 months, local tumour control was 100% without any case of post-therapeutic enucleation. Two patients (8%) developed liver metastases.

Conclusion

SRT of ocular tumours by HA is highly appropriate, and HA is superior to intensity-modulated arc therapy (IMAT) concerning dose reduction in organs at risk (OARs). The novel gradient measure SADG is more informative than Paddick’s GI.