U mobilization and associated U isotope fractionation by sulfur-oxidizing bacteria.

Uranium (U) contamination of the environment causes high risk to health, demanding for effective and sustainable remediation. Bioremediation via microbial reduction of soluble U(VI) is generating high fractions (>50%) of insoluble non-crystalline U(IV) which, however, might be remobilized by sulfur-oxidizing bacteria. In this study, the efficacy of Acidithiobacillus (At.) ferrooxidans and Thiobacillus (T.) denitrificans to mobilize non-crystalline U(IV) and associated U isotope fractionation were investigated. At. ferrooxidans mobilized between 74 and 91% U after 1 week, and U mobilization was observed for both, living and inactive cells. Contrary to previous observations, no mobilization by T. denitrificans could be observed. Uranium mobilization by At. ferrooxidans did not cause U isotope fractionation suggesting that U isotope ratio determination is unsuitable as a direct proxy for bacterial U remobilization. The similar mobilization capability of active and inactive At. ferrooxidans cells suggests that the mobilization is based on the reaction with the cell biomass. This study raises doubts about the long-term sustainability of in-situ bioremediation measures at U-contaminated sites, especially with regard to non-crystalline U(IV) being the main component of U bioremediation.

Holo-APP and G-protein-mediated signaling are required for sAPPα-induced activation of the Akt survival pathway.

Accumulating evidence indicates that loss of physiologic amyloid precursor protein (APP) function leads to reduced neuronal plasticity, diminished synaptic signaling and enhanced susceptibility of neurons to cellular stress during brain aging. Here we investigated the neuroprotective function of the soluble APP ectodomain sAPPα (soluble APPα), which is generated by cleavage of APP by α-secretase along the non-amyloidogenic pathway. Recombinant sAPPα protected primary hippocampal neurons and SH-SY5Y neuroblastoma cells from cell death induced by trophic factor deprivation. We show that this protective effect is abrogated in neurons from APP-knockout animals and APP-depleted SH-SY5Y cells, but not in APP-like protein 1- and 2- (APLP1 and APLP2) depleted cells, indicating that expression of membrane-bound holo-APP is required for sAPPα-dependent neuroprotection. Trophic factor deprivation diminished the activity of the Akt survival pathway. Strikingly, both recombinant sAPPα and the APP-E1 domain were able to stimulate Akt activity in wild-type (wt) fibroblasts, SH-SY5Y cells and neurons, but failed to rescue in APP-deficient neurons or fibroblasts. The ADAM10 (a disintegrin and metalloproteinase domain-containing protein 10) inhibitor GI254023X exacerbated neuron death in organotypic (hippocampal) slice cultures of wt mice subjected to trophic factor and glucose deprivation. This cell death-enhancing effect of GI254023X could be completely rescued by applying exogenous sAPPα. Interestingly, sAPPα-dependent Akt induction was unaffected in neurons of APP-ΔCT15 mice that lack the C-terminal YENPTY motif of the APP intracellular region. In contrast, sAPPα-dependent rescue of Akt activation was completely abolished in APP mutant cells lacking the G-protein interaction motif located in the APP C-terminus and by blocking G-protein-dependent signaling with pertussis toxin. Collectively, our data provide new mechanistic insights into the physiologic role of APP in antagonizing neurotoxic stress: they suggest that cell surface APP mediates sAPPα-induced neuroprotection via G-protein-coupled activation of the Akt pathway.

A differential capacitive electrical field sensor array for contactless measurement of respiratory rate.

Although respiratory rate is an important vital sign for early detection of deterioration, on general wards it is not routinely monitored. Since patients may not tolerate cables attached to their chest, we developed an unobtrusive and contactless measurement method which can be placed under a mattress. The sensor array uses the Maxwell-Wagner relaxation effect by capacitive injection of a high-frequency voltage into the torso and subsequent measurement of respiratory-induced phase shift. Simulations of the entire measurement scenario indicate an improved signal-to-noise ratio if a differential method is applied with specific positioning of the electrodes. A prototype was designed and the simulations were confirmed with measurements on a phantom and in a human self-experiment by the authors using self-constructed hardware. Movement artifacts were detected using an artifact detection algorithm which allows reliable estimation of the respiratory rate.

238U/235U variations in meteorites: extant 247Cm and implications for Pb-Pb dating.

The 238U/235U isotope ratio has long been considered invariant in meteoritic materials (equal to 137.88). This assumption is a cornerstone of the high-precision lead-lead dates that define the absolute age of the solar system. Calcium-aluminum-rich inclusions (CAIs) of the Allende meteorite display variable 238U/235U ratios, ranging between 137.409 +/- 0.039 and 137.885 +/- 0.009. This range implies substantial uncertainties in the ages that were previously determined by lead-lead dating of CAIs, which may be overestimated by several million years. The correlation of uranium isotope ratios with proxies for curium/uranium (that is, thorium/uranium and neodymium/uranium) provides strong evidence that the observed variations of 238U/235U in CAIs were produced by the decay of extant curium-247 to uranium-235 in the early solar system, with an initial 247Cm/235U ratio of approximately 1.1 x 10(-4) to 2.4 x 10(-4).

Fe isotope variations in natural materials measured using high mass resolution multiple collector ICPMS.

We present the first measurements of Fe isotope variations in chemically purified natural samples using high mass resolution multiple-collector inductively coupled plasma source mass spectrometry (MC-ICPMS). High mass resolution allows polyatomic interferences at Fe masses to be resolved (especially, (40)Ar(14)N(+), (40)Ar(16)O(+), and (40)Ar(16)OH(+)). Simultaneous detection of Fe isotope ion beams using multiple Faraday collectors facilitates high-precision isotope ratio measurements. Fe in basalt and paleosol samples was extracted and purified using a simple, single-stage anion chemistry procedure. A Cu "element spike" was used as an internal standard to correct for variations in mass bias. Using this procedure, we obtained data with an external precision of 0.03-0.11 per thousand and 0.04-0.15 per thousand for delta(56/54)Fe and delta(57/54)Fe, respectively (2sigma). Use of Cu was necessary for such reproducibility, presumably because of subtle effects of residual sample matrix on mass bias. These findings demonstrate the utility of high-resolution MC-ICPMS for high-precision Fe isotope analysis in geologic and other natural materials. They also highlight the importance of internal monitoring of mass bias, particularly when using routine methods for Fe extraction and purification.

Individual and common antigen-recognition sites of liver-derived T cells in patients with autoimmune hepatitis.

Autoimmune hepatitis (AIH) is characterized by dense T-cell infiltrations in the liver tissue, but little is known how T cells influence the pathogenesis. To address this question, the distribution of T-cell receptor variable beta-chain (TCR Vbeta) transcripts of peripheral blood and liver-infiltrating T cells from previously untreated patients with newly diagnosed acute exacerbated AIH was investigated. Furthermore, the lengths and sequences of complementary-determining region 3 (CDR3) were studied. Reverse transcriptase-polymerase chain reaction (RT-PCR) analysis and CDR3 spectratyping revealed multiple clonal expansions of liver-infiltrating T cells but not peripheral T cells within various TCR Vbeta families. Further analysis of overexpressed TCR Vbeta transcripts using TCR beta-chain-joining element (TCR Jbeta)-specific primers in a nested PCR showed characteristic Vbeta/Jbeta combinations. Subsequent sequencing of CDR3 regions from PCR products confirmed the clonality of T-cell expansions and the usage of common and individual CDR3 motifs. In conclusion, the clonality of expanded T cells within the liver tissue during early clinical manifestation of untreated AIH indicated that autoantigen-specific T cells accumulate at the inflammation site. Individual and common CDR3 motifs argued for predominant epitopes that were recognized by liver-infiltrating T cells in AIH patients.

A clinical trial of tailored office systems for preventive service delivery. The Study to Enhance Prevention by Understanding Practice (STEP-UP).

BACKGROUND: The potential of primary care practice settings to prevent disease and morbidity through health habit counseling, screening for asymptomatic disease, and immunizations has been incompletely met. This study was designed to test a practice-tailored approach to increasing preventive service delivery with particular emphasis on health habit counseling. DESIGN: Group randomized clinical trial and multimethod process assessment. SETTING/PARTICIPANTS: Seventy-seven community family practices in northeast Ohio. INTERVENTION: After a 1-day practice assessment, a nurse facilitator met with practice clinicians and staff and assisted them with choosing and implementing individualized tools and approaches aimed at increasing preventive service delivery. MAIN OUTCOME MEASURE: Summary scores of the health habit counseling, screening and immunization services recommended by the U.S. Preventive Services Task Force up to date for consecutive patients during randomly selected chart review days. RESULTS: A significant increase (p=0.015) in global preventive service delivery rates at the 1-year follow-up was found in the intervention group (31% to 42%) compared to the control group (35% to 37%). Rates specifically for health habit counseling (p=0.007) and screening services (p=0.048) were increased, but not for immunizations. CONCLUSIONS: An approach to increasing preventive service delivery that is individualized to meet particular practice needs can increase global preventive service delivery rates.

Divergent effects of tumor necrosis factor alpha on apoptosis of human neutrophils.

Apoptosis of neutrophils is a key mechanism to control the intensity of the acute inflammatory response. Previously, the cytokine tumor necrosis factor alpha (TNF-alpha) was reported by some to have pro-apoptotic and by others to have antiapoptotic effects on neutrophils. The aim of this study was to explain these contradictory results. We found that TNF-alpha at low concentrations strongly decreased apoptosis of neutrophils. However, at higher concentrations, TNF-alpha lost its protective effects, and also reversed the protective effects of interferon-gamma (IFN-gamma) and granulocyte-macrophage colony-stimulating factor (GM-CSF). This pro-apoptotic effect of TNF-alpha was blocked by anti-CD11b and was absent in neutrophils from patients with chronic granulomatous disease, which cannot produce toxic oxygen metabolites. Under these circumstances, we found that TNF-alpha retained its anti-apoptotic effects even at high concentrations. In conclusion, the protective effects against apoptosis of IFN-gamma, GM-CSF, and TNF-alpha itself are overruled when the concentration of TNF-alpha is high enough to produce a respiratory burst. These dual, concentration-dependent effects of TNF-alpha provide an explanation for previous controversial reports and support a dominant role for TNF-alpha in neutrophil apoptosis.

92Nb-(92)Zr and the Early Differentiation History of Planetary Bodies.

The niobium-92-zirconium-92 ((92)Nb-(92)Zr) extinct radioactive decay system (half-life of about 36 million years) can place new time constraints on early differentiation processes in the silicate portion of planets and meteorites. Zirconium isotope data show that Earth and the oldest lunar crust have the same relative abundances of (92)Zr as chondrites. (92)Zr deficits in calcium-aluminum-rich inclusions from the Allende meteorite constrain the minimum value for the initial (92)Nb/(93)Nb ratio of the solar system to 0.001. The absence of (92)Zr anomalies in terrestrial and lunar samples indicates that large silicate reservoirs on Earth and the moon (such as a magma ocean residue, a depleted mantle, or a crust) formed more than 50 million years after the oldest meteorites formed.

The viral clearance in interferon-treated chronic hepatitis C is associated with increased cytotoxic T cell frequencies.

BACKGROUND/AIMS: Cytotoxic T lymphocytes have been demonstrated in peripheral blood and liver tissue of patients with chronic hepatitis C virus infection, but their significance for viral clearance is unknown. Therefore, we analyzed hepatitis C virus-specific cytotoxic T lymphocyte precursor frequencies in chronic hepatitis C virus carriers during interferon-alpha treatment. METHODS: Blood mononuclear cells or CD8+ T cells from HLA-A2 positive and negative patients and controls were analyzed in chromium-release assays using a panel of 18 synthetic peptides from the HCV core, E1 and NS4 antigens bearing HLA-A2 binding motifs. Specific cytotoxic T lymphocyte precursor frequencies were studied within CD8+ T cells derived from interferon-alpha-treated patients using a TNF-alpha-based ELISPOT assay and compared to viremia levels. RESULTS: T cells from 16 of 24 HLA-A2+ but none of the six HLA-A2- patients with chronic hepatitis C and six HLA-A2+ healthy controls lysed targets pulsed with peptide cocktails. Fine specificity revealed four very immunogenic epitopes in the core (C36-44, C132-140) and the envelope regions (E332-340, E363-372). Cytotoxic T lymphocyte precursor frequencies were prospectively analyzed in 11 interferon-alpha-treated HLA-A2+ hepatitis C virus patients. Four sustained and two transient therapy responders showed lower pretreatment viremia levels and significantly higher specific cytotoxic T lymphocyte precursor frequencies during viral clearance compared to five therapy non-responders and untreated controls. CONCLUSIONS: The quantitative induction of HLA-class I restricted responses by interferon-alpha could contribute to a beneficial outcome of hepatitis C virus infections. Furthermore, it appears that the balance between viral load and specific cellular immune responses is critical for successful viral clearance.

Quantitative and functional analysis of core-specific T-helper cell and CTL activities in acute and chronic hepatitis B.

AIMS/BACKGROUND: CD4+ T-helper cell (Th) responses to hepatitis B virus (HBV) core antigen (HBc) are increased during exacerbations in acute and chronic hepatitis B (AHB, CHB) and might influence the induction of CD8+ cytotoxic T lymphocytes (CTL) that are important for viral clearance. METHODS: HBc-specific proliferative responses and cytokine release of blood mononuclear cells (PBMC) were studied in patients with AHB or CHB, as well as responders and non-responders to interferon-alpha treatment (IFN-R, IFN-NR), by [3H]-thymidine-uptake, enzyme-linked immunosorbent assay (ELISA) and Elispot assay and were compared to peptide HBc18 27-specific CTL precursor frequencies among CD8+ T cells derived from HLA-A2+ patients. RESULTS: HBc-specific proliferative PBMC responses and Th frequencies were significantly increased in AHB patients compared with untreated CHB patients. PBMC derived from IFN-R showed stronger cellular responses than IFN-NR. Stimulated PBMC from all patient groups secreted significantly more IFN-gamma than IL-4 indicating Th1/Th0 cell responses. Furthermore, in AHB and IFN-R patients, high peptide HBc18-27-specific CTL precursor frequencies closely correlated with strong HBc-specific Th responses, whereas in untreated CHB and IFN-NR patients lower CTL frequencies were observed without correlation to Th activities. CONCLUSIONS: HBV core-specific Th-cell responses appeared to support efficient CTL induction in patients with viral clearance, whereas in chronic HBV carriers quantitatively insufficient Th and CTL responses were observed. This observation could be important for future therapeutic strategies.

The cellular immune responses induced in the follow-up of interferon-alpha treated patients with chronic hepatitis C may determine the therapy outcome.

BACKGROUND/AIMS: To study whether the host's immune response determines viral clearance in chronic hepatitis C, virological markers and antigen-specific T cell reactions were analysed in 30 chronic HCV carriers followed up during interferon-alpha therapy, 11 untreated anti-HCV positive individuals and 10 healthy controls. METHODS: Proliferative T helper cell responses to recombinant HCV core and non-structural antigens were monitored by 3H-thymidine uptake assay and compared to quantitative viraemia levels and HCV genotypes. RESULTS: Of the 30 treated patients, six had sustained complete responses (20%), another six were transient therapy responders (20%) and 18 were non-responders (60%). Viral clearance was associated with the HCV genotype 3 and low pretreatment viral load. In a substantial proportion of complete and transient therapy responders, increased NS3-, helicase- and NS4-antigen-specific T cell responses were observed during interferon-alpha therapy. In non-responders and in the later clinical courses of transient therapy responders, core and NS5-specific T cell responses dominated. In addition, 11 untreated anti-HCV antibody positive individuals were studied. Two HCV-RNA negative patients who might have recovered from HCV infection showed strong persistent lymphoproliferative responses to NS3, helicase and NS4 antigens, whereas seven of the nine viraemic patients reacted with HCV core or NS5 antigens. CONCLUSIONS: Interferon-alpha treatment enhances NS3-, helicase- and NS4-antigen-specific T helper cell responses in patients with viral clearance, whereas viral persistence was associated with increased T cell reactivities against core and NS5 antigens. Immunogenetical, immunological and virological factors that may influence differential T cell induction in chronic hepatitis C are discussed.

Insertional inactivation of the gene for the meningococcal lactoferrin binding protein.

The lactoferrin binding protein (LBP) of Neisseria meningitidis (the putative meningococcal receptor for human lactoferrin, LF), has been previously characterized as an outer-membrane protein of approximately 105 kDa. Using N-terminal amino acid sequence to generate an oligonucleotide probe, a clone from a lambda gt11 phage library was isolated. This clone was subjected to shuttle mutagenesis, in which an erythromycin mini-transposon was used to interrupt the LBP coding sequence. This insertion mutation was introduced into the meningococcus. A N. meningitidis strain that carried this transposon insertion no longer produced the 105 kDa protein. The absence of this protein was correlated with the inability to bind LF or to use LF as an iron source. The LBP mutant was able to grow with other Fe sources and demonstrated no other visible membrane protein alterations. These data confirm the suggestion that LBP is the meningococcal receptor.

Genetically controlled enzymatic variation in a southern, biracial, semi-rural community: the Bogalusa heart study.

Caucasian and Afro-American schoolchildren participating in a community-based cardiovascular screening program in Bogalusa, Louisiana are characterized with respect to phenotypic and allelic frequencies at 10 enzymatic and 3 nonenzymatic polymorphic loci. Biracial gene frequencies in Bogalusa are compared with those reported for other populations of similar ethnic composition. Intra- and interracial genetic distances within and between the racial subpopulations of Bogalusa and Seattle, Washington are compared. While the comparisons do not support the concept that northern, urban blacks have more Caucasian admixture than blacks residing in the rural South, they do suggest microdifferentiation of the two white populations.