Expression and function of alpha-adrenoceptors in zebrafish: drug effects, mRNA and receptor distributions

The alpha2-adrenoceptors are G-protein-coupled receptors that mediate many of the physiological effects of norepinephrine and epinephrine. Mammals have three subtypes of alpha2-adrenoceptors, alpha2A, alpha2B and alpha2C. Zebrafish, a teleost fish used widely as a model organism, has five distinct alpha2-adrenoceptor genes. The zebrafish has emerged as a powerful tool to study development and genetics, with many mutations causing diseases reminiscent of human diseases. Three of the zebrafish adra2 genes code for orthologues of the mammalian alpha2-adrenoceptors, while two genes code for alpha2Da- and alpha2Db- adrenoceptors, representing a duplicated, fourth alpha2-adrenoceptor subtype. The three different mammalian alpha2-adrenoceptor subtypes have distinct expression patterns in different organs and tissues, and mediate different physiological functions. The zebrafish alpha2-adrenergic system, with five different alpha2-adrenoceptors, appears more complicated. In order to deduce the physiological functions of the zebrafish alpha2-adrenoceptors, we localized the expression of the five different alpha2-adrenoceptor subtypes using RT-PCR, mRNA in situ hybridization, and receptor autoradiography using the radiolabelled alpha2-adrenoceptor antagonist [ethyl-3H]RS-79948-197. Localization of the alpha2A-, alpha2B- and alpha2C-adrenoceptors in zebrafish shows marked conservation when compared with mammals. The zebrafish alpha2A, alpha2Da, and alpha2Db each partially follow the distribution pattern of the mammalian alpha2A: a possible indication of subfunction partitioning between these subtypes. The alpha2-adrenergic system is functional in zebrafish also in vivo, as demonstrated by marked locomotor inhibition, similarly to mammals, and lightening of skin colour induced by the specific alpha2-adrenoceptor agonist, dexmedetomidine. Both effects were antagonized by the specific alpha2-adrenoceptor antagonist atipamezole.     

Analysis of 8-hydroxydeoxyguanosine among workers exposed to diesel particulate exhaust: comparison with urinary metabolites and PAH air monitoring

Oxidative DNA damage and repair, as measured by 8-hydroxy-2'-deoxyguanosine (8-OHdG) in urine and DNA samples were studied in association with work-related diesel exhaust exposure among garage and waste collection workers. Seasonal variations of the urinary 8-OHdG levels in pre- and two post-workshift urine samples of 29 exposed workers and 36 control persons were evaluated. The mean±SE levels of post-workshift 8-OHdG (μmol/mol crea) were 1.52±0.44 in winter and 1.61±0.33 in summer for the exposed workers, and 1.56±0.61 in winter and 1.43±0.49 in summer for the controls, respectively. No significant difference in the urinary 8-OHdG levels between exposed workers and control subjects in winter (p=0.923) and summer (p=0.350) was observed. A linear mixed model, adjusted for years of employment, age, ex/non-smoking and BMI, indicated no significant dose exposure-relationships between the urinary 8-OHdG and 15 PAH air concentrations nor between the 8-OHdG and 7 PAH monohydroxy-metabolites analyzed in the same workers. 8-OHdG was also analyzed in the mononuclear cell DNA of 19 exposed and 18 control subjects. The mean value of 8-OHdG/non-modified 2'-deoxyguanosine (8-OHdG/105 dG±SE) were 4.89±0.17 for the exposed and 4.11±0.16 for the control persons, which showed no correlation with the urinary 8-OHdG levels (r=0.01, n=28, P=0.96). The PAH exposure at workplaces was mainly composed of volatile compounds, particularly naphthalene, suggesting low exposure through the respiratory tract and a low effect of PAH in ROS induction.     

Formation of biofilms in drinking water distribution networks,a case study in two cities in Finland and Latvia

The formation of biofilms in drinking water distribution networks is a significant technical, aesthetic and hygienic problem. In this study, the effects of assimilable organic carbon, microbially available phosphorus (MAP), residual chlorine, temperature and corrosion products on the formation of biofilms were studied in two full-scale water supply systems in Finland and Latvia. Biofilm collectors consisting of polyvinyl chloride pipes were installed in several waterworks and distribution networks, which were supplied with chemically precipitated surface waters and groundwater from different sources. During a 1-year study, the biofilm density was measured by heterotrophic plate counts on R2A-agar, acridine orange direct counting and ATP-analyses. A moderate level of residual chorine decreased biofilm density, whereas an increase of MAP in water and accumulated cast iron corrosion products significantly increased biofilm density. This work confirms, in a full-scale distribution system in Finland and Latvia, our earlier in vitro finding that biofilm formation is affected by the availability of phosphorus in drinking water.     

Expression change in Angiopoietin-1 underlies change in relative brain size in fish

Brain size varies substantially across the animal kingdom and is often associated with cognitive ability; however, the genetic architecture underpinning natural variation in these key traits is virtually unknown. In order to identify the genetic architecture and loci underlying variation in brain size, we analysed both coding sequence and expression for all the loci expressed in the telencephalon in replicate populations of guppies (Poecilia reticulata) artificially selected for large and small relative brain size. A single gene, Angiopoietin-1 (Ang-1), a regulator of angiogenesis and suspected driver of neural development, was differentially expressed between large- and small-brain populations. Zebra fish (Danio rerio) morphants showed that mild knock down of Ang-1 produces a small-brained phenotype that could be rescued with Ang-1 mRNA. Translation inhibition of Ang-1 resulted in smaller brains in larvae and increased expression of Notch-1, which regulates differentiation of neural stem cells. In situ analysis of newborn large- and small-brained guppies revealed matching expression patterns of Ang-1 and Notch-1 to those observed in zebrafish larvae. Taken together, our results suggest that the genetic architecture affecting brain size in our population may be surprisingly simple, and Ang-1 may be a potentially important locus in the evolution of vertebrate brain size and cognitive ability.     

Assessment of occupational exposure to PAHs in an Estonian coke oven plant- correlation of total external exposure to internal dose measured as 1-hydroxypyrene concentration

The exposure of cokery workers to polynuclear aromatic hydrocarbons at an Estonian oil shale processing plant was assessed by using occupational hygiene and biomonitoring measurements which were carried out twice, in midwinter and in the autumn. To assess the external dose of polynuclear aromatic hydrocarbons, pyrene and benzo[a]pyrene concentrations were measured from the breathing zone of workers during a workshift. Skin contamination with pyrene and benzo[a]pyrene was assessed by skin wipe sampling before and after the workshift. As a biomarker of overall exposure to polynuclear aromatic hydrocarbons, and as an integral of all absorption routes of pyrene, 1-hydroxypyrene concentration was measured from post shift urine samples. Of the personal air samples, 18% exceeded the Finnish threshold limit value of benzo[a]pyrene (10 μg m^-3). Mean value (two separate measurements together) for benzo[a]pyrene was 5.7 μg m^-3 and for pyrene, 8.1 μg m^-3. Based on skin wipe sample analyses, the skin contamination was also obvious. The mean value of benzo[a]pyrene in the samples collected after the shift was 1.2 ng cm^-2. Benzo[a]pyrene was not found in control samples. The mean value of urinary 1-hydroxypyrene concentration was 6.0 μmol mol^-1 creatinine for the exposed workers and 0.5 μmol mol^-1 creatinine for the controls. This study undoubtedly shows the usefulness of 1-hydroxypyrene as an indicator of internal dose of polynuclear aromatic hydrocarbons. It can be concluded that the cokery workers at the Kohtla-Järve plant are exposed to high concentrations of polynuclear aromatic compounds, and the exposure level is considerably higher during the winter measurements.     

Pentraxin 3 (PTX3) Is Associated with Severe Sepsis and Fatal Disease in Emergency Room Patients with Suspected Infection: A Prospective Cohort Study

Background

Early diagnostic and prognostic stratification of patients with suspected infection is a difficult clinical challenge. We studied plasma pentraxin 3 (PTX3) upon admission to the emergency department in patients with suspected infection.

Methods

The study comprised 537 emergency room patients with suspected infection: 59 with no systemic inflammatory response syndrome (SIRS) and without bacterial infection (group 1), 67 with bacterial infection without SIRS (group 2), 54 with SIRS without bacterial infection (group 3), 308 with sepsis (SIRS and bacterial infection) without organ failure (group 4) and 49 with severe sepsis (group 5). Plasma PTX3 was measured on admission using a commercial solid-phase enzyme-linked immunosorbent assay (ELISA).

Results

The median PTX3 levels in groups 1–5 were 2.6 ng/ml, 4.4 ng/ml, 5.0 ng/ml, 6.1 ng/ml and 16.7 ng/ml, respectively (p<0.001). The median PTX3 concentration was higher in severe sepsis patients compared to others (16.7 vs. 4.9 ng/ml, p<0.001) and in non-survivors (day 28 case fatality) compared to survivors (14.1 vs. 5.1 ng/ml, p<0.001). A high PTX3 level predicted the need for ICU stay (p<0.001) and hypotension (p<0.001). AUC^ROC in the prediction of severe sepsis was 0.73 (95% CI 0.66–0.81, p<0.001) and 0.69 in case fatality (95% CI 0.58–0.79, p<0.001). PTX3 at a cut-off level for 14.1 ng/ml (optimal cut-off value for severe sepsis) showed 63% sensitivity and 80% specificity. At a cut-off level 7.7 ng/ml (optimal cut-off value for case fatality) showed 70% sensitivity and 63% specificity in predicting case fatality on day 28.In multivariate models, high PTX3 remained an independent predictor of severe sepsis and case fatality after adjusting for potential confounders.

Conclusions

A high PTX3 level on hospital admission predicts severe sepsis and case fatality in patients with suspected infection.     

Life-cycle climate impacts of peat fuel: calculation methods and methodological challenges

Purpose

There has been lively debate, especially in Finland and Sweden, on the climate impacts of peat fuel. Previous studies of peat fuel's life-cycle climate impacts were controversial in their interpretation. The aim of this paper is conclusive examination of the issues of LCA methodology, derived from critical review of previous studies and recalculation based on the latest knowledge of greenhouse gas balances related to peat fuel’s utilisation and the radiative forcing impacts of greenhouse gases.

Methods

The most recent findings on emissions and the gas fluxes between soil, vegetation and atmosphere were used in calculation of the life-cycle climate impacts of the various peat fuel utilisation chains by means of LCA methodology. In the main, the calculation methods and rules were the same as in the previous studies, with the aim being to distinguish the impact of peat fuel’s utilisation from that of the natural or semi-natural situation. A dynamic method was employed for assessing changes in radiative forcing. The results of alternative peat fuel utilisation chains were compared to the corresponding result for coal.

Results

There are many steps in peat fuel LCA, where different assumptions lead to different outcomes. Determining the functional unit, reference situations and system boundaries, as well as the emission calculation methods, is important from this point of view. Determination of the initial reference situation emerged as one of the critical points in the calculations. Time scale can strongly affect the final outcomes in a study where effects of long-term land-use change are considered.

Conclusions

Each peatland area is unique. The higher the greenhouse gas emissions in the initial reference situation, the greater is the climate impact of the area and the more suitable the area is for peat extraction. The study showed that more greenhouse gas flux measurements are needed, for better assessment of the climate impacts of different potential peat extraction sites. Climate change mitigation requires quick actions, and uncertainties related to emissions are higher for longer time spans. Therefore, it can be concluded that a perspective spanning more than 100 years is inappropriate in peat fuel's life-cycle climate impact assessments.     

Load application induces changes in the expression levels of Sox-9, FGFR-3 and VEGF in condylar chondrocytes

Experimental and clinical observations have proven the modulatory effects of mechanical loading on the development and maintenance of cartilage architecture. Here we examined the involvement of Sox-9, FGFR-3 and VEGF (pivotal factors controlling cartilage development and growth) in the mechano-transduction pathway of mandibular condylar cartilage by changing the dynamics of the transmitted load via changes in food hardness. To this end, condyle cartilage tissue of rats fed with hard or soft food was analyzed immunohistochemically at various time points. Our findings demonstrate that different mechanical loading conditions in condylar chondrocytes trigger differentiation-/maturation-related processes by affecting the expression levels of these factors, ultimately influencing condylar cartilage growth.     

Abnormal behavior,striatal dopamine turnover and opioid peptide gene expression in histamine‐deficient mice

Hypothalamic histaminergic neurons regulate a variety of homeostatic, metabolic and cognitive functions. Recent data have suggested a modulatory role of histamine and histamine receptors in shaping striatal activity and connected the histaminergic system to neuropsychiatric disorders. We characterized exploratory behavior and striatal neurotransmission in mice lacking the histamine producing enzyme histidine decarboxylase (Hdc). The mutant mice showed a distinct behavioral pattern during exploration of novel environment, specifically, increased frequency of rearing seated against the wall, jumping and head/body shakes. This behavioral phenotype was associated with decreased levels of striatal dopamine and serotonin and increased level of dopamine metabolite DOPAC. Gene expression levels of dynorphin and enkephalin, opioids released by medium spiny neurons of striatal direct and indirect pathways respectively, were lower in Hdc mutant mice than in control animals. A low dose of amphetamine led to similar behavioral and biochemical outcomes in both genotypes. Increased striatal dopamine turnover was observed in Hdc KO mice after treatment with dopamine precursor l ‐Dopa. Overall, our study suggests a role for striatal dopamine and opioid peptides in formation of distinct behavioral phenotype of Hdc KO mice.