The fermentation stoichiometry of Thermotoga neapolitana and influence of temperature,oxygen, and pH on hydrogen production

The hyperthermophilic bacterium, Thermotoga neapolitana, has potential for use in biological hydrogen (H₂) production. The objectives of this study were to (1) determine the fermentation stoichiometry of Thermotoga neapolitana and examine H₂ production at various growth temperatures, (2) investigate the effect of oxygen (O₂) on H₂ production, and (3) determine the cause of glucose consumption inhibition. Batch fermentation experiments were conducted at temperatures of 60, 65, 70, 77, and 85°C to determine product yield coefficients and volumetric productivity rates. Yield coefficients did not show significant changes with respect to growth temperature and the rate of H₂ production reached maximum levels in both the 77°C and 85°C experiments. The fermentation stoichiometry for T. neapolitana at 85°C was 3.8 mol H₂, 2 mol CO₂, 1.8 mol acetate, and 0.1 mol lactate produced per mol of glucose consumed. Under microaerobic conditions H₂ production did not increase when compared to anaerobic conditions, which supports other evidence in the literature that T. neapolitana does not produce H₂ through microaerobic metabolism. Glucose consumption was inhibited by a decrease in pH. When pH was adjusted with buffer addition cultures completely consumed available glucose. © 2009 American Institute of Chemical Engineers Biotechnol. Prog., 2009     

Full development of Th2 immunity requires both innate and adaptive sources of CD154

The CD40-CD154 interaction is critical for Th2 response generation during helminth infection and following immunization with helminth-conditioned dendritic cells, yet the key cellular sources of these molecules have still to be defined in vivo. In this study, we demonstrate that the requirement for CD40 expression during murine Th2 response induction is restricted exclusively to the Ag-bearing dendritic cells. In contrast, development of full Th2 immunity required CD154 expression on multiple populations. In this respect, optimal production of IL-5, IL-10, and IL-13 was dependent upon CD154 expression by both CD4(+) T cells and non-lymphoid cells. IL-4 production had less stringent costimulatory requirements, with expression of CD154 on either non-lymphoid cells or T cells alone being sufficient to enable production of this archetypal Th2 cytokine. Disparities in CD154 requirements for T cell and B cell responses were revealed during experimental schistosomiasis where, even in the face of robust Th2 generation, B cell class-switching was entirely dependent upon expression of CD154 by the lymphoid compartment. These data help define the costimulatory interactions that occur during the generation of Th2 immunity, and challenge the widely held view that CD154 expressing T cells are the sole contributors in this process.     

Fine mapping of Ath6, a quantitative trait locus for atherosclerosis in mice

Ath6 is a novel quantitative trait locus associated with differences in susceptibility to atherosclerosis between C57BL/6J (B6) and C57BLKS/J (BKS) inbred mouse strains. Combining data from an intercross and a backcross (1593 meioses) between mice from B6 and BKS strains and from The Jackson Laboratory interspecific backcross panels, (C57BL/6J ×Mus spretus) F₁× C57BL/6J and (C57BL/6J × SPRET/Ei) F₁× SPRET/Ei, we constructed a consensus genetic map and narrowed Ath6 to a 1.07 ± 0.26 cM interval between the anonymous DNA marker D12Pgn4 and the gene Nmyc1. This region is near the proximal end of murine Chromosome (Chr) 12, which is homologous to the human chromosomal region 2p24-p25. Marker order in the Ath6 region was concordant among the two crosses and The Jackson Laboratory interspecific backcross panels. This high resolution map rules out candidate genes encoding apolipoprotein B, syndecan 1, and Adam17. The two Ath6 crosses have a combined potential resolution of 0.06 cM. Received: 12 September 2000 / Accepted: 22 February 2001     

Study on the dehydrating effect of the red cell Na+/K+-pump in nystatin-treated cells with varying Na+ and water contents

Using the antibiotic Nystatin, we have developed a systematic method for the preparation of red blood cells with independently selected levels of intracellular Na⁺ concentrations and water content. Such cells provided an experimental model to study the effect of Na⁺/K⁺ pump stimulation on red cell water content. Even in initially dehydrated cells, stimulation of the Na⁺/K⁺ pump by elevated intracellular Na⁺ caused subsequent further loss of cell water. Cell water loss was reflected in decreased monovalent cation content per unit mass of hemoglobin and by a shift in the density distribution of the cell populations to higher densities on discontinuous Stractan gradients. We conclude that the 3 Na_out⁺ : 2 K_in⁺ stoichiometry of the Na⁺/K⁺ pump results in a net desalting effect with increased pump activity. Under the conditions of these experiments, the cell appears to have no effective mechanism to compensate for a net loss of ions and water.     

Extracellular ATP induces the accumulation of superoxide via NADPH oxidases in Arabidopsis

Extracellular ATP can serve as a signaling agent in animal cells, and, as suggested by recent reports, may also do so in plant cells. In animal cells it induces the production of reactive oxygen species through the mediation of NADPH oxidase. Similarly, here we report that in leaves of Arabidopsis (Arabidopsis thaliana), applied ATP, but not AMP or phosphate, induces the accumulation of superoxide (O2-) in a biphasic, dose-dependent manner, with a threshold at 500 nm ATP. This effect did not require ATP hydrolysis for it was mimicked by ATPgammaS. ATP also induced increased levels of Arabidopsis respiratory burst oxidase homolog D (AtrbohD) mRNA, but ATP-treated plants that had disrupted AtrbohD and AtrbohF genes did not accumulate O2-, indicating that NADPH oxidases are responsible for the induced O2- accumulation. Inhibitors of mammalian P2-type ATP receptors abolished ATP-induced O2- production, suggesting that the ATP effects may be mediated through P2-like receptors in plants. Cytosolic Ca2+ and calmodulin are likely to help transduce the ATP responses, as they do in animal cells, because a Ca2+ channel blocker, a Ca2+ chelator, and calmodulin antagonist all reduced ATP-induced O2- accumulation. Furthermore, ATP treatment enhanced the expression of genes that are induced by wounds and other stresses. The ATP measured at wound sites averaged 40 microm, well above the level needed to induce O2- accumulation and gene expression changes. Transgenic plants overexpressing an apyrase gene had reduced O2- production in response to applied ATP and wounding. Together, these data suggest a possible role for extracellular ATP as a signal potentially in wound and stress responses.     

Bile acid-induced secretion in polarized monolayers of T84 colonic epithelial cells: Structure-activity relationships

Bile acid epimers and side-chain homologues are present in the human colon. To test whether such bile acids possess secretory activity, cultured T84 colonic epithelial cells were used to quantify the secretory properties of synthetic epimers and homologues of deoxycholic acid (DCA) and chenodeoxycholic acid (CDCA). In our study, chloride secretion was measured as changes in short-circuit current (DeltaI(sc), in microA/cm2) with the use of voltage-clamped monolayers of T84 cells mounted in Ussing chambers. Bile acids were added at 0.5 mM, a concentration that did not alter transepithelial resistance. Data were expressed as peak DeltaI(sc) (means +/- SD). When added bilaterally, DCA stimulated a DeltaI(sc) response of 15.7 +/- 12.5 microA/cm2. The 12beta-OH epimer of DCA was less potent (DeltaI(sc) = 8.0 +/- 1.7 microA/cm2), whereas its 3beta-OH epimer had no effect. CDCA stimulated secretion (DeltaI(sc) = 8.2 +/- 5.5 microA/cm2), whereas both its 7beta-OH and 3beta-OH epimers were inactive, as was lithocholic acid. HomoDCA (1 additional side-chain carbon) was active (DeltaI(sc) = 7.8 +/- 4.8 microA/cm2), whereas norDCA (1 fewer carbon) and dinorDCA (2 fewer carbons) were not. Taurine conjugates of DCA and CDCA stimulated secretion (DeltaI(sc) = 12.3 +/- 7.5 and 8.8 +/- 4.8 microA/cm2, respectively) from the basolateral side but not the apical side. Uptake of taurine conjugates from the basolateral but not the apical side was shown by mass spectrometry. These studies indicate marked structural specificity for bile acid-induced chloride secretion and show that modification of bile acid structure by colonic bacteria modulates the secretory properties of these endogenous secretagogues.     

Capacity for protein synthesis following heat stimulus of Drosophila associates with heat tolerance but does not underlie the latitudinal tolerance cline

Across populations of Drosophila melanogaster along the Australian eastern coastline latitudinal clines occur in both heat-knockdown tolerance and hardened heat-knockdown tolerance – low latitude tropical populations being more tolerant. A latitudinal cline also occurs for rates of total protein synthesis following a mild heat stress, with tropical populations having higher rates. Since the control of protein synthesis following heat stress is an important component of the cellular heat-shock response, we hypothesised that the higher rates of synthesis that follow a heat stimulus lead to higher knockdown tolerance and underpins the cline. However, levels of heat-stimulated total protein synthesis have been negatively related to heat-hardening capacity, a somewhat conflicting result. Here we examine the relationship between these physiological and adaptive traits in a set of 40 family lines derived from a hybrid laboratory population established by crossing populations from either end of the latitudinal transect. Among these lines high levels of heat-stimulated total protein synthesis were associated with both low basal and low heat-hardened adult knockdown time, confirming the importance of a negative relationship between protein synthesis and thermal tolerance. This result, when considered along with the directions of the latitudinal clines in protein synthesis and tolerance, suggests that variation in rates of heat-stimulated total protein synthesis is not a factor contributing to the latitudinal cline in heat tolerance. Given the robustness of this negative relationship we discuss possible explanations and future experiments to elucidate how the cellular heat stress response might facilitate increased knockdown tolerance.     

Mitochondrial DNA of ancient Cumanians: culturally Asian steppe nomadic immigrants with substantially more western Eurasian mitochondrial DNA lineages

The Cumanians were originally Asian pastoral nomads who in the 13th century migrated to Hungary. We have examined mitochondrial DNA from members of the earliest Cumanian population in Hungary from two archeologically well-documented excavations and from 74 modern Hungarians from different rural locations in Hungary. Haplogroups were defined based on HVS I sequences and examinations of haplogroup-associated polymorphic sites of the protein coding region and of HVS II. To exclude contamination, some ancient DNA samples were cloned. A database was created from previously published mtDNA HVS I sequences (representing 2,615 individuals from different Asian and European populations) and 74 modem Hungarian sequences from the present study. This database was used to determine the relationships between the ancient Cumanians, modern Hungarians, and Eurasian populations and to estimate the genetic distances between these populations. We attempted to deduce the genetic trace of the migration of Cumanians. This study is the first ancient DNA characterization of an eastern pastoral nomad population that migrated into Europe. The results indicate that, while still possessing a Central Asian steppe culture, the Cumanians received a large admixture of maternal genes from more westerly populations before arriving in Hungary. A similar dilution of genetic, but not cultural, factors may have accompanied the settlement of other Asian nomads in Europe.