Atmospheric deposition and soil water chemistry in Swedish forests since 1985 - Effects of reduced emissions of sulphur and nitrogen.

Trends for the atmospheric deposition of sulphur (S) and inorganic nitrogen (inorg-N) to forests and changes in the forest soil water chemistry in Sweden have been assessed since 1985, with special focus on the last 25 years, based on measurements within the Swedish Throughfall Monitoring Network (SWETHRO). The reductions in the deposition of S and inorg-N in the southern part of Sweden corresponded relatively well with the pollutant emission reductions for S and inorg-N from both EU27 + UK and Sweden during 1996/97-2021/22. For northern Sweden the deposition of S and inorg-N decreased to a lesser extent than both European and Swedish emissions. The bulk deposition of NO3-N has decreased more than the deposition of NH4-N over the last 25-year period, which is consistent with the much larger emission reductions for NOx compared to NH3 from EU27 + UK and Sweden. The S concentrations in the soil water, at 50 cm below soil surface, have decreased during the last 25 years, however somewhat less than the S deposition. At sites with low ANC and pH in the beginning of the period, the increase in ANC was generally greater and the increase in pH was smaller, but at sites with high pH and ANC above zero, the increase in pH was dominant, in line with the nonlinear relationship between pH and ANC in the soil water. The incidence of elevated concentrations of NO3-N in the soil water was highest in southwest Sweden, ranging between 4 and 19 % of all measuring occasions since 1985/86. The reduced deposition of N over the 35-year period was not reflected in the incidence of elevated concentrations of NO3-N in the soil water over time.

Twenty years of nitrogen deposition to Norway spruce forests in Sweden.

The yearly, total (dry+wet) deposition of inorganic nitrogen (inorg-N) to Norway spruce forests was estimated with a full spatial coverage over Sweden for a twenty-year period, 2001-2020, based on combined measurements with Teflon string samplers, throughfall deposition and bulk deposition to the open field. The results were based on a novel method to apply estimates of the dry deposition based on measurements at a limited number of sites, to a larger number of sites with only bulk deposition measurements, in turn based on the existence of a strong geographical gradient in the dry deposition of inorg-N from southwest to northeast Sweden. The method should be applicable for other geographical regions where gaseous NH3, NO2 and HNO3 are not main drivers of N dry deposition and where geographical gradients in dry deposition could be defined. It was shown that Norway spruce forests in south Sweden receive more N from deposition than has been previously estimated, based on modelling. Clear time trends were demonstrated for decreased deposition of inorg-N to Norway spruce forests in all parts of Sweden. The decreases were somewhat larger than what could be expected from the decrease in the reported emissions of inorg-N from Europe. The results emphasize that estimates of the total deposition are necessary in order to map levels and follow the development of N deposition in forests.

Optimization of droplet digital PCR assays for the type-specific detection and quantification of five HPV genotypes, including additional data on viral loads of nine different HPV genotypes in cervical carcinomas.

The droplet digital PCR (ddPCR) system enables high-sensitivity detection of nucleic acids and direct absolute quantification of the targets. The aim of this research was to evaluate this system for viral load (VL) analysis of the human papillomavirus (HPV) genotypes HPV31, 35, 39, 51 and 56 measured in number of viral particles per cell. The sample types used for the optimization of the ddPCR assay were formalin-fixed paraffin-embedded (FFPE) tissues and cervical liquid cytology samples. The presently optimized ddPCR assays, together with assays optimized previously for HPV16, 18, 33 and 45, with the same ddPCR method, were used for the VL analysis of cervical tumor samples. Results published previously on the present study cohort showed that women with a cervical tumor containing multiple high-risk HPV genotypes had a worse prognosis compared to women with single-genotype-infected tumors. The VL was therefore analyzed in this study for the same cohort, as a possible explanatory factor to the prognostic differences. The results of the optimization part of the study, with analysis of VL using ddPCR in DNA from varying sample types (FFPE and liquid cytology samples), showed that each of the five assays demonstrated good inter- and intra-assay means with a coefficient of variation (CV) under 8% and 6% respectably. The cohort results showed no difference in VL between tumors with multiple and single HPV infections, and therefore did most likely not constitute a contributing factor for prognostic differences observed previously. However, tumors from women aged 60 years or older or containing certain HPV genotypes and genotype genera were associated with a higher VL.

Two orphan seven-transmembrane segment receptors which are expressed in CD4-positive cells support simian immunodeficiency virus infection.

Clinical isolates of primate immunodeficiency viruses, including human immunodeficiency virus type 1 (HIV-1), enter target cells by sequential binding to CD4 and the chemokine receptor CCR5, a member of the seven-transmembrane receptor family. HIV-1 variants which use additional chemokine receptors are present in the central nervous system or emerge during the course of infection. Simian immunodeficiency viruses (SIV) have been shown to use CCR5 as a coreceptor, but no other receptors for these viruses have been identified. Here we show that two orphan seven-transmembrane segment receptors, gpr1 and gpr15, serve as coreceptors for SIV, and are expressed in human alveolar macrophages. The more efficient of these, gpr15, is also expressed in human CD4(+) T lymphocytes and activated rhesus macaque peripheral blood mononuclear cells. The gpr15 and gpr1 proteins lack several hallmarks of chemokine receptors, but share with CCR5 an amino-terminal motif rich in tyrosine residues. These results underscore the potential diversity of seven-transmembrane segment receptors used as entry cofactors by primate immunodeficiency viruses, and may contribute to an understanding of viral variation and pathogenesis.

The envelope glycoprotein ectodomains determine the efficiency of CD4+ T lymphocyte depletion in simian-human immunodeficiency virus-infected macaques.

CD4+ T lymphocyte depletion in human immunodeficiency virus type 1 (HIV-1)-infected humans underlies the development of acquired immune deficiency syndrome. Using a model in which rhesus macaques were infected with chimeric simian-human immunodeficiency viruses (SHIVs), we show that both the level of viremia and the structure of the HIV-1 envelope glycoprotein ectodomains individually contributed to the efficiency with which CD4(+) T lymphocytes were depleted. The envelope glycoproteins of recombinant SHIVs that efficiently caused loss of CD4(+) T lymphocytes exhibited increased chemokine receptor binding and membrane-fusing capacity compared with those of less pathogenic viruses. These studies identify the HIV-1 envelope glycoprotein ectodomains as determinants of CD4(+) T lymphocyte loss in vivo and provide a foundation for studying pathogenic mechanisms.

Phase II study of oral fingolimod (FTY720) in multiple sclerosis: 3-year results.

In a 6-month, placebo-controlled trial, oral fingolimod (FTY720) 1.25 or 5.0 mg, once daily, significantly reduced MRI inflammatory activity and annualized relapse rate compared with placebo in patients with relapsing multiple sclerosis (MS). The objectives were to monitor the 36-month, interim efficacy and safety results of the ongoing extension of this study. In the extension (months 7-36), placebo-treated patients were re-randomized to either dose of fingolimod; fingolimod-treated patients continued at the same dose. During months 15-24, all patients receiving fingolimod 5.0 mg switched to 1.25 mg. Of the 250 patients who entered the extension study, 173 (69%) continued to month 36. Most patients were free from gadolinium-enhanced lesions (88-89%) or new T2 lesions (70-78%) at month 36. Patients receiving continuous fingolimod treatment had sustained low annualized relapse rates of 0.20-0.21, and 68-73% remained relapse-free at month 36. Over 36 months, nasopharyngitis (34%), headache (30%), fatigue (19%) and influenza (18%) were the most commonly reported adverse events. Pulmonary function remained stable and blood pressure was stable after an initial increase (3-5 mmHg) during the first 6 months of fingolimod treatment; serious adverse events included infections and skin cancer. The low MRI and clinical disease activity at 6 months were maintained at 36 months with fingolimod, which was generally well tolerated by most patients. The efficacy and safety of oral fingolimod are being further evaluated in a large phase III MS study programme.

Anosognosia versus unilateral neglect. Coexistence and their relations to age, stroke severity, lesion site and cognition.

The aim of this study was to examine the association between anosognosia and unilateral neglect (UN), with special focus on age, stroke severity, lesion location and pre-stroke dementia. The basis of this investigation was a population-based stroke incidence study. Anosognosia was assessed using a questionnaire, and UN using a three-item version of the Behaviour Inattention Test, the Baking Tray Task and a test of personal neglect. Stroke severity was assessed using the NIH stroke scale. Patients with anosognosia were older, and they more often had pre-stroke dementia than patients having UN only. No particular lesion localization was associated with anosognosia, while UN was strongly associated with previously defined lesion sites, often in the parietal lobe. There was a borderline significance regarding stroke severity in patients having anosognosia compared with those with UN only. Patients with anosognosia had higher mortality than patients without, but when controlled for age and stroke severity, this effect was not independent. While UN is closely associated with 'classical' lesion sites, anosognosia is a condition that more often occurs in a previously impaired brain. For anosognosia, lesion location appears to be less important. Anosognosia also tends to occur with larger strokes.

Complexes between cationic liposomes and DNA visualized by cryo-TEM.

The association structures formed by cationic liposomes and DNA-plasmids have been successfully employed as gene carriers in transfection assays. In the present study such complexes was studied by cryo-TEM (cryo-transmission electron microscopy). Cationic liposomes made up by DOPE (dioleoylphosphatidylethanolamine) and various amounts of three different cationic surfactants were investigated. The cryo-TEM analysis suggests that an excess of lipid in terms of charge, leads to entrapment of the DNA molecules between the lamellas in clusters of aggregated multilamellar structures. With increasing amounts of DNA free or loosely bound plasmids were found in the vicinity of the complexes. The importance of the choice of surfactant, as reported from many transfection assays, was not reflected in changes of the type of DNA-vesicle association. A tendency towards polymorphism of the lipid mixtures is reported and its possible implications are discussed.

A multi-step lipid mixing assay to model structural changes in cationic lipoplexes used for in vitro transfection.

Formation of liposome/polynucleotide complexes (lipoplexes) involves electrostatic interactions, which induce changes in liposome structure. The ability of these complexes to transfer DNA into cells is dependent on the physicochemical attributes of the complexes, therefore characterization of binding-induced changes in liposomes is critical for the development of lipid-based DNA delivery systems. To clarify the apparent lack of correlation between membrane fusion and in vitro transfection previously observed, we performed a multi-step lipid mixing assay to model the sequential steps involved in transfection. The roles of anion charge density, charge ratio and presence of salt on lipid mixing and liposome aggregation were investigated. The resonance-energy transfer method was used to monitor lipid mixing as cationic liposomes (DODAC/DOPE and DODAC/DOPC; 1:1 mole ratio) were combined with plasmid, oligonucleotides or Na(2)HPO(4). Cryo-transmission electron microscopy was performed to assess morphology. As plasmid or oligonucleotide concentration increased, lipid mixing and aggregation increased, but with Na(2)HPO(4) only aggregation occurred. NaCl (150 mM) reduced the extent of lipid mixing. Transfection studies suggest that the presence of salt during complexation had minimal effects on in vitro transfection. These data give new information about the effects of polynucleotide binding to cationic liposomes, illustrating the complicated nature of anion induced changes in liposome morphology and membrane behavior.

Characterization of the binding epitope of the monoclonal antibody DMAb-1 to ganglioside GM2.

Several derivatives of ganglioside GM2 were synthesized for mapping of the binding epitope of a monoclonal antibody raised against this ganglioside. The GM2 ganglioside was modified in both the hydrophobic and the hydrophobilic part of the molecule. The synthesized derivatives were characterized with fast atom bombardment mass spectrometry (FAB-MS). Affinity of the monoclonal antibody for the GM2 derivatives was determined by enzyme-linked immunosorbent assay (ELISA) on microtitre plates or by TLC immunostaining. Modifying the GM2 sialic acid by deacetylation or blocking of the carboxyl moiety abolished the binding to the monoclonal antibody while the cleaving of the glycol group on the sialic acid tail led to a 70% reduced binding affinity. Removal of the fatty acid (lyso-GM2) eliminated the binding to the antibody. GM2 derivatives with fatty acid moieties of 8 carbon atoms or less showed almost no reactivity. GM2 with saturated fatty acids 16:0, 18:0 and 20:0 had binding affinity similar to natural GM2, while the 24:0 fatty acid had only half the binding affinity. The results demonstrate the importance of ganglioside fatty acid composition with regard to ligand binding between the monoclonal antibody and its specific ganglioside antigen. Thus, caution must be shown in the application of immunaffinity methods with monoclonal antibodies for the quantitative determination of glycosphingolipids from different tissues.

Effects of human plasma proteins on maturation of monocyte-derived dendritic cells.

Dendritic cells (DC) are a promising tool for vaccine therapy due to their unique properties as antigen presenting cells and their ability to prime naïve T cells. Increasing evidence suggests that maturation stage of DC critically influences the fate of the immune response. Generation of monocyte-derived DC for clinically applicable immunotherapy requires the use of well-defined components and stringent culture conditions. An alternative strategy is to use human autologous serum. However, its constituents are not stable and reflect the inflammatory condition of the donor. In order to investigate whether DC properties are influenced by proteins present in the plasma, we matured human monocyte-derived DC with four main plasma components: fibrinogen, fibronectin, plasminogen or C-reactive protein. These purified proteins were added at various concentrations on day 6 after the initial differentiation induced by IL-4 and GM-CSF. The maturation was assessed by phenotyping of maturation-associated marker (CD83) and co-stimulatory molecule CD86 as well as IL-12 production. Functional properties of DC were assessed by endocytic activity and mixed leukocyte culture. Our results indicate that fibrinogen had DC-maturation effect comparable to poly-I:C, TNF-alpha and PGE(2) as a positive control, but it failed to induce IL-12 production. The other plasma proteins had no effect on DC maturation. CRP at high concentration had rather inhibitory effect on DC induced lymphocyte function. We conclude that none of the tested plasma components and acute phase proteins sufficiently induce fully competent mature DC. This finding is important for the preparation of human DC-based vaccines supplemented by autologous sera.

Heavy metal and nitrogen concentrations in mosses are declining across Europe whilst some "hotspots" remain in 2010.

In recent decades, naturally growing mosses have been used successfully as biomonitors of atmospheric deposition of heavy metals and nitrogen. Since 1990, the European moss survey has been repeated at five-yearly intervals. In 2010, the lowest concentrations of metals and nitrogen in mosses were generally found in northern Europe, whereas the highest concentrations were observed in (south-)eastern Europe for metals and the central belt for nitrogen. Averaged across Europe, since 1990, the median concentration in mosses has declined the most for lead (77%), followed by vanadium (55%), cadmium (51%), chromium (43%), zinc (34%), nickel (33%), iron (27%), arsenic (21%, since 1995), mercury (14%, since 1995) and copper (11%). Between 2005 and 2010, the decline ranged from 6% for copper to 36% for lead; for nitrogen the decline was 5%. Despite the Europe-wide decline, no changes or increases have been observed between 2005 and 2010 in some (regions of) countries.

Outcome measurement in economic evaluations of growth factors.

There are several ways of measuring outcome in an economic evaluation of filgrastim therapy. These measures imply different types of economic evaluation. In all cases, the filgrastim therapy is compared with no filgrastim therapy. Since different therapeutic strategies are connected with uncertain consequences, decision trees are helpful analytical tools. The potential implications of different types of economic evaluations are illustrated with a simple decision tree.

A brush method to harvest cells from the nasal mucosa for microscopic and biochemical analysis.

A method is described for the sampling of epithelial cells and other effector cells from the human airway mucosa for structural and biochemical analysis. The cell samples are obtained from the nasal mucosa using a small nylon brush which is rotated over the epithelium and soaked and shaken in a small volume of a balanced salt solution. Morphological evaluation using light microscopy and transmission electron microscopy revealed excellently preserved cytological detail. In asymptomatic individuals the cells harvested were as follows: 45 +/- 5.9% (mean +/- SEM) epithelial cells, 38 +/- 7.1% granulocytes, 16 +/- 2.3% large mononuclear cells (monocytes), and 1.3 +/- 2.3% eosinophils. Repeated measurements in the same individual revealed a coefficient of variation of the order of 40% for the proportions of cells harvested. In comparison with nasal airway lavage, a higher proportion of epithelial cells and monocytes were obtained with the brush method. The cells harvested could also be used for biochemical analysis. The histamine content of the cell pellets was found to be strongly correlated with the mast cell count (r = 0.93) and was estimated to about 10 pg/cell, which is higher than previously reported for mast cells obtained from human lung tissue dispersed by an enzymatic method. The present method appears to be appropriate for the study of cellular events in the nasal mucosal epithelium.

GABAB receptors in various in vitro and in vivo models of epilepsy: a study with the GABAB receptor blocker CGP 35348.

The effect of the GABAB receptor blocker CGP 35348 on epileptic processes in vitro and in vivo was studied. In hippocampal slices of the rat maintained in vitro, CGP 35348 (100 microM) induced a moderate increase in the frequency of extracellularly recorded spontaneous epileptiform burst discharges induced in CA3 by penicillin (1.2 mM), bicuculline (5 microM) and low Mg(2+) (0.1 mM). This effect was observed in 50-75% of the slices. A similar but less consistent increase was also observed in CA1 in bicuculline and low Mg2+. Data obtained by intracellular recordings from CA1 pyramidal cells in the presence of bicuculline (10 microM) demonstrated that CGP 35348 (100 microM) increased the duration of the paroxysmal depolarization underlying an evoked epileptiform burst and reduced the early component of the after hyperpolarization which followed the burst. In mice pretreated with isoniazid, CGP 35348 (300 mg/kg, i.p.) significantly increased the number of convulsing mice. However, convulsions induced by submaximal doses of pentylenetetrazol, picrotoxin or strychnine were not facilitated by CGP 35348. We conclude that GABAB receptors appear to exert a suppressant effect on various kinds of epileptiform discharges of hippocampal neurons in vitro. In vivo, however, the role of GABAB receptors in regulating convulsions is less prominent since only isoniazid-induced convulsions were facilitated by GABAB receptor blockade.

Clover as an indicator plant for phytotoxic ozone concentrations: visible injury in relation to species, leaf age and exposure dynamics.

Visible ozone injury on leaves of three clover species was investigated in relation to species, leaf age and exposure dynamics. It was shown that ozone episodes in south-west Sweden cause visible injury to subterranean clover (Trifolium subterraneum L.), white clover (Trifolium repens L.) and red clover (Trifolium pratense L.). Trifolium subterranean was most sensitive to ozone, whilst T. pratense was least sensitive. Application of the anti-ozonant, ethylenediurea (EDU), reduced the extent of visible ozone injury, but did not give complete protection with the concentrations used. Similarly, filtration of the air reduced the extent of visible injury in T. subterraneum enclosed in open-top chambers. EDU-treated plants of T. subterranean accumulated more biomass than the non-EDU treated plants after a period with rather high ozone concentrations, while the opposite occurred for T. pratense. Experiments with T. subterraneum in open-top chambers also showed that older leaves were more sensitive to ozone than younger leaves and that a shorter period with higher ozone concentrations produced more ozone injury than a longer period with lower ozone concentrations, although the two periods had the same number of ppb-hours.

High-peak power nanosecond optical parametric amplifier with periodically poled KTP.

A two-stage optical parametric amplifier generating 5 ns 208 kW peak power pulses in the spectral region at 1.535 microm in a diffraction-limited beam was realized in a single periodically poled KTP crystal. The maximum small-signal gain for the two stages reached 75dB and the total conversion efficiency was 30%. An analysis of the small-signal gain dependence on the M2 of the pump beam is presented for the collinear and noncollinear OPA. Efficient spectral broadening of the signal was demonstrated in short pieces of single-mode telecommunication fiber.

Metachromatic, IgE-bearing and tryptase-containing cells on the nasal mucosal surface in provoked allergic rhinitis.

We studied the relationship between mast cells or basophils and symptoms in provoked allergic rhinitis. Nasal brush and lavage samples were obtained before nasal allergen challenge and every 2 h for 12 h after the challenge in 10 allergics and 3 controls. The cells were identified by their metachromatic staining properties (brush and lavage samples) or with immunohistochemical methods using specific antibodies to IgE and tryptase, a selective mast-cell marker (brush samples). Histamine was determined in the brush samples using liquid chromatography. After an initial decrease, the numbers of metachromatic, IgE-bearing and tryptase-containing cells, as well as the histamine content of the cells in the brush samples, increased during the subsequent observation hours. The prechallenge cell and histamine values correlated with the symptom score 15 min after the challenge. The prechallenge lavage samples lacked metachromatic cells, but these cells were found in increasing numbers after the provocation. Three of the patients differed from the remaining seven in that their prechallenge brush samples contained many positively stained cells. All patients showed a positive cellular response to the allergen challenge, but these three individuals showed the most vivid response. The morphology of the metachromatic cells in the prechallenge brush samples agreed with that of mast cells, but the morphology of metachromatic cells which outnumbered tryptase-containing cells in samples at 8 to 12 h rather agreed with their being basophils.

Cytological and bacteriological aspects of secretory otitis media.

Bacteriological and cytological examinations were performed on 105 middle ear secretions from 66 children with middle ear effusion (MEE) of more than 3 months' duration. The secretions were searched for granulocytes and the activity of these cells was judged by their capacity for random locomotion and their ability to reduce nitroblue tetrazolium. The functional characteristics of the granulocytes were compared with the bacteriological findings on cultures from MEE. Bacteria commonly regarded as pathogens in middle ear infections (Hemophilus influenzae, Branhamella catarrhalis or Streptococcus pneumoniae) were found in 25% of the secretions. Granulocytes with activity or lacking activity, virtually dead, were found in all secretions where these bacteria were isolated. In secretions where bacteria commonly regarded as commensals, mainly staphylococci, were isolated, about two thirds of the secretions showed phagocytes with or without activity. No relation between bacterial growth and the functional state of the granulocytes was observed. In contrast, no phagocytes were found in over 60% of MEE lacking bacterial growth. These findings suggest a role for bacteria in the development and maintenance of secretory otitis media.

A new view on chemistry of solids in solution--cryo energy-filtered transmission electron microscopy (cryo-EFTEM) imaging of aggregating palladium colloids in vitreous ice

It is shown that by using cryo-transmission electron microscopy (cryo-TEM) it is possible to image the aggregation behaviour of nanoparticles while they are still in solution. This technique has allowed the study of the arrangement of colloidal palladium particles in solution by preparing the specimen by the plunge-freezing technique. This method of rapidly cooling the specimen avoids rearrangement of the particles during specimen preparation. The palladium particles were identified by energy-filtered cryo-TEM. The aggregation of particles in solution was studied as a function of pH and ionic strength. The results can be used as recommendations for colloidal solutions intended for deposition of single particles.