Cord blood hemopoietic progenitor cell toll like receptor expression and function: a mechanism underlying allergic inflammation in early life?

RATIONALE: Neonatal immune responses to environmental stimuli, mediated via TLR, may determine the development of atopy in childhood. Since hemopoietic mechanisms are involved in development and maintenance of allergic inflammation, we investigated alterations in progenitor expression and differentiation profiles after stimulation with TLR agonists.<p></p> METHODS: Freshly isolated, CD34-enriched human CB cells were stimulated with 10μg/mL lipopolysaccharide (LPS) or 5μM CpG ODN overnight. Flow cytometric analyses were used to evaluate surface and intracellular expression of TLR-2, TLR-4, TLR-9, as well as the hemopoietic cytokine receptors (HCR) IL-5R, IL-3R and GM-CSFR; methylcellulose cultures were performed to assess CD34+ cell differentiation capacity into Eo/B CFU.<p></p> RESULTS: After TLR agonist stimulation, CD34+ cell TLR-2, -4 and TLR-9 percentage expression increased significantly (p=0.005), whereas HCR expression decreased (p=0.01); however, mean fluorescence intensity of all receptors was found to be increased. Stimulation with a combination of TLR agonists and hemopoietic cytokines induced increased IL-5- and IL-3-responsive Eo/B CFU (p=0.02), when compared to hemopoietic cytokine stimulation alone.<p></p> CONCLUSIONS: CB CD34+ progenitor cells significantly express TLR, and TLR ligation directly affects both TLR and HCR expression. These receptor alterations allow modulation of progenitor cell differentiation capacity into eosinophils and basophils, key cells involved in allergic inflammation. These findings may highlight an alternate innate immune pathway of microbial influence on the development of allergic inflammation in early life.<p></p&gt

Regulation of Hsp70 function by a eukaryotic DnaJ homolog

We report that a purified cytoplasmic Hap70 homolog from Saccharomyces cerevisiae, Hsp70SSA1, exhibits a weak ATPase activity, which is stimulated by a purified eukaryotic dnaJp homolog (YDJ1p). Stable complex formation between Hsp70SSA1and the permanently unfolded protein carboxymethylated α-lactalbumin (CMLA) was assayed by native gel electrophoresis.We report that a purified cytoplasmic Hap70 homolog from Saccharomyces cerevisiae, Hsp70SSA1, exhibits a weak ATPase activity, which is stimulated by a purified eukaryotic dnaJp homolog (YDJ1p). Stable complex formation between Hsp70SSA1and the permanently unfolded protein carboxymethylated α-lactalbumin (CMLA) was assayed by native gel electrophoresis

In Vitro Effects of Budesonide on Eosinophil-Basophil Lineage Commitment

IL-5 is the primary cytokine that stimulates the production and survival of eosinophils and basophils from progenitor cells. The inhaled glucocorticoid, budesonide, has been shown to exert a therapeutic effect via suppression of eosinophil/basophil progenitors in vivo. Since various steroids have exhibited the ability to enhance eosinophil/basophil progenitor differentiation, we examined the effects of budesonide in vitro. Bone marrow and cord blood samples were obtained and cultured in the presence of IL-5 alone or IL-5 plus budesonide. Eosinophil/basophil colony-forming units were enumerated from cultured nonadherent mononuclear cells and from purified CD34+ cells. CD34+ cells with and without budesonide were also examined for up-regulation of ERK1/2, MAPK and GATA-1 using real time-PCR. Results: i) up-regulation of eosinophil/basophil colony-forming units is due to the direct effects of budesonide on IL-5-stimulated progenitors; ii) GATA-1 is likely involved in the early amplification of eosinophil/basophil progenitor commitment leading to increased differentiation. A potential transcriptional pathway has been identified which may mediate the effects of budesonide on eosinophil/basophil lineage commitment

Europe as a multilevel federation

Peer reviewedPublisher PD

Analyzing the carbon dioxide emissions of R134a alternatives in water-cooled centrifugal chillers using the life cycle climate performance framework

Introduction: To reduce greenhouse gases, the Kigali Amendment to the Montreal Protocol seeks a phasedown of hydrofluorocarbons. R134a alternatives were analyzed for use in a water-cooled chiller: R450A, R513A, R516A, R1234ze (E), R515A, and R515B.Methods: A thermodynamic model of the chiller was employed to calculate compressor power, an input to the life cycle climate performance (LCCP) framework to estimate total equivalent carbon dioxide emissions, CO2eq. Emissions were calculated for an 809 kW [230 Tons of refrigeration (RT) nameplate] water-cooled centrifugal chiller at constant cooling capacity using five power sources (i.e., coal, distillate fuel oil, natural gas, nuclear, and wind) for a median chiller lifetime of 27 years. Two chiller operating profiles were considered: one using operational data from a university campus and a second from literature based on the Atlantic Fleet operation.Results and discussion: When powered via fossil fuels, over 90% of emissions were due to the indirect emissions from energy; therefore, the global warming potential (GWP) of the refrigerant was not the primary contributor to the total CO2eq emissions. With natural gas, total LCCP emissions were reduced for R450A (7.8%), R513A (4.7%), R516A (9.4%), R1234ze (E) (10%), R515A (8.4%), and R515B (6.4%) compared to R134a for the university campus load profile. For the round-the-clock Atlantic Fleet profile, there were emission reductions for R450A (3.6%), R513A (0.25%), R516A (2.3%), R1234ze (E) (2.4%), R515A (1.5%) and R515B (2.4%) compared to R134a. When coupled with renewable energy, the indirect emissions from the chillers substantially decreased, and GWP-dependent leakage emissions accounted for up to 74% or 40% of emissions from R134a alternatives powered by wind and nuclear, respectively. For operation using the load profile from the university campus chillers, R450A had the highest coefficient of performance (COP) of 5.802, while R513A had the lowest COP (5.606). Tradeoffs between alternative refrigerants exist in terms of operation, temperature glide, size of heat exchangers, system design, flammability, cost, availability, and material compatibility. In terms of flammability, R134a, R513A, R450A, R515B and R515A are A1 (nonflammable) fluids. R450A and R516A also have temperature glides of 0.4 K and 0.056 K, respectively, which can affect condenser design. In terms of equipment modification (sizing), R513A require fewer modifications

Quality Control Autophagy Degrades Soluble ERAD-Resistant Conformers of the Misfolded Membrane Protein GnRHR

Molecular chaperones triage misfolded proteins via action as substrate selectors for quality control (QC) machines that fold or degrade clients. Herein, the endoplasmic reticulum (ER) associated Hsp40 JB12 is reported to participate in partitioning mutant conformers of GnRHR, a G-protein coupled receptor, between ER-associated degradation (ERAD) and a novel ERQC-autophagy pathway for membrane proteins. ERQC-autophagy degrades E90K-GnRHR because pools of its partially folded and detergent soluble degradation intermediates are resistant to ERAD. S168R-GnRHR is globally misfolded and disposed of via ERAD, but inhibition of p97, the protein retrotranslocation motor, shunts S168R-GnRHR from ERAD to ERQC autophagy. Partially folded and grossly misfolded forms of GnRHR associate with JB12 and Hsp70. Elevation of JB12 promotes ERAD of S168R-GnRHR, with E90K-GnRHR being resistant. E90K-GnRHR elicits association of the Vps34 autophagy initiation complex with JB12. Interaction between ERassociated Hsp40s and the Vps34 complex permits the selective degradation of ERAD-resistant membrane proteins via ERQC-autophagy

Long-term effects of chronic light pollution on seasonal functions of European blackbirds (turdus merula)

Light pollution is known to affect important biological functions of wild animals, including daily and annual cycles. However, knowledge about long-term effects of chronic exposure to artificial light at night is still very limited. Here we present data on reproductive physiology, molt and locomotor activity during two-year cycles of European blackbirds (Turdus merula) exposed to either dark nights or 0.3 lux at night. As expected, control birds kept under dark nights exhibited two regular testicular and testosterone cycles during the two-year experiment. Control urban birds developed testes faster than their control rural conspecifics. Conversely, while in the first year blackbirds exposed to light at night showed a normal but earlier gonadal cycle compared to control birds, during the second year the reproductive system did not develop at all: both testicular size and testosterone concentration were at baseline levels in all birds. In addition, molt sequence in light-treated birds was more irregular than in control birds in both years. Analysis of locomotor activity showed that birds were still synchronized to the underlying light-dark cycle. We suggest that the lack of reproductive activity and irregular molt progression were possibly the results of i) birds being stuck in a photorefractory state and/or ii) chronic stress. Our data show that chronic low intensities of light at night can dramatically affect the reproductive system. Future studies are needed in order to investigate if and how urban animals avoid such negative impact and to elucidate the physiological mechanisms behind these profound long-term effects of artificial light at night. Finally we call for collaboration between scientists and policy makers to limit the impact of light pollution on animals and ecosystems

Sequential Quality-Control Checkpoints Triage Misfolded Cystic Fibrosis Transmembrane Conductance Regulator

Cystic fibrosis arises from the misfolding and premature degradation of CFTR Delta F508, a Cl- ion channel with a single amino acid deletion. Yet, the quality-control machinery that selects CFTR Delta F508 for degradation and the mechanism for its misfolding are not well defined. We identified an ER membrane-associated ubiquitin ligase complex containing the E3 RMA1, the E2 Ubc6e, and Derlin-1 that cooperates with the cytosolic Hsc70/CHIP E3 complex to triage CFTR and CFTR Delta F508. Derlin-1 serves to retain CFTR in the ER membrane and interacts with RMA1 and Ubc6e to promote CFTR's proteasomal degradation. RMA1 is capable of recognizing folding defects in CFTR Delta F508 coincident with translation, whereas the CHIP E3 appears to act posttranslationally. A folding defect in CFTR Delta F508 detected by RMA1 involves the inability of CFTR's second membrane-spanning domain to productively interact with amino-terminal domains. Thus, the RMA1 and CHIP E3 ubiquitin ligases act sequentially in ER membrane and cytosol to monitor the folding status of CFTR and CFTR Delta F508