Heterogeneity in direct cytotoxic function of L3T4 T cells. TH1 clones express higher cytotoxic activity to antigen-presenting cells than TH2 clones

In the process of generating culture supernatant from T cell clones, with anti-CD3 antibodies and the B lymphoma A20 as APC, a striking difference in the stimulation of TH1 and TH2 clones was observed, i.e., TH2 clones produced higher levels of lymphokines than TH1 clones. This prompted us to test the hypothesis that differential killing of APC (thus the removal of stimuli) by T cells led to differential T cell activation. By studying a panel of five TH1 and seven TH2 clones, it was demonstrated that TH1 clones mediated significantly higher levels of cytotoxicity toward A20 cells in the presence of soluble anti-CD3 antibody (as opposed to immobilized anti-CD3). Although T cell clones could, when activated with immobilized anti-CD3, produce lymphokines cytotoxic to A20 cells, experiments in which lymphokine production was blocked indicated that T cell clones, in the presence of soluble anti-CD3, mediated killing of A20 through direct cytotoxicity. A higher level of cytotoxicity, by TH1 compared with TH2 clones, was not restricted to anti-CD3 or a particular target cell type, because it also occurred with Con A- or Ag-dependent killing (a monocyte-macrophage cell line), and LPS blasts. Furthermore, the higher cytotoxic activity of TH1 clones compared with TH2 clones was independent of the stage of T cell activation and was unlikely a result of the length of in vitro culture. High levels of killing of APC led to low levels of T cell activation, the significance of which may be as a negative feedback mechanism in the immune response. Other biologic relevancies of higher cytotoxic activity in TH1 vs TH2 cells were also discussed.     

Ionized and wobble base-pairing for bromouracil-guanine in equilibrium under physiological conditions. A nuclear magnetic resonance study on an oligonucleotide containing a bromouracil-guanine base-pair as a function of pH

A one and two-dimensional nuclear magnetic resonance study of a non-selfcomplementary oligonucleotide containing a central 5-bromouracil-guanine pair is reported. For these two bases three types of hydrogen bonding schemes could exist; wobble, rare tautomer and ionized. The two-dimensional spectra of non-exchangeable protons together with one-dimensional spectra recorded in water show that at pH 7.0 the predominant species is a right-handed B-form DNA in which the brU.G pair has wobble geometry. On raising the pH we observe a transition monitored by proton chemical shift changes for the brU.G and adjacent base-pairs. The mid-point of the transition was observed at pH 8.6. Spectra recorded at pH 9.8 show that the helix remains intact with B form conformation. It is shown that this high pH form has an ionized brU.G base-pair now in Watson-Crick geometry. Thus under physiological conditions an equilibrium exists between wobble and ionized structures.     

Physical and genetic mapping of the Rhodobacter sphaeroides 2.4.1 genome: presence of two unique circular chromosomes.

A macrorestriction map representing the complete physical map of the Rhodobacter sphaeroides 2.4.1 chromosomes has been constructed by ordering the chromosomal DNA fragments from total genomic DNA digested with the restriction endonucleases AseI, SpeI, DraI, and SnaBI. Junction fragments and multiple restriction endonuclease digestions of the chromosomal DNAs derived from wild-type and various mutant strains, in conjunction with Southern hybridization analysis, have been used to order all of the chromosomal DNA fragments. Our results indicate that R. sphaeroides 2.4.1 carries two different circular chromosomes of 3,046 +/- 95 and 914 +/- 17 kilobases (kb). Both chromosome I (3,046 kb) and chromosome II (914 kb) contain rRNA cistrons. It appears that only a single copy of the rRNA genes is contained on chromosome I (rrnA) and that two copies are present on chromosome II (rrnB, rrnC). Additionally, genes for glyceraldehyde 3-phosphate dehydrogenase (gapB) and delta-aminolevulinic acid synthase (hemT) are found on chromosome II. In each instance, there appears to be a second copy of each of these genes on chromosome I, but the extent of the DNA homology is very low. Genes giving rise to enzymes involved in CO2 fixation and linked to the gene encoding the form I enzyme (i.e., the form I region) are on chromosome I, whereas those genes representing the form II region are on chromosome II. The complete physical and partial genetic maps for each chromosome are presented.     

Structural and spectroscopic characterization of exogenous ligand binding to isolated factor F430 and its configurational isomers

Binding of axial ligands to the nickel(II) of isolated factor F430 from the methyl reductase enzyme of Methanobacterium thermoautotrophicum is demonstrated. Evidence of bis-ligand coordination is obtained from the x-ray absorption, optical, and resonance Raman spectral characterization of F430 and its 12,13-diepimeric isomer in the presence of a large excess of cyanide, pyridine, or 1-methylimidazole. Significant broadening and 5-10-nm red shifts of the main 430-nm optical absorption band and shifts of up to 30 cm-1 for the high-frequency Raman lines are observed upon coordination of these axial ligands. The Raman spectra of native F430 and the diepimer with a particular axial ligand are nearly identical. Nickel x-ray absorption edge spectra of the diepimer in the absence and presence of these exogenous ligands are indicative of conversion from a square-planar to a tetragonally distorted octahedral geometry. Analyses of the nickel extended x-ray absorption fine structure data for the ligated diepimer complexes yield detailed structural information for these complexes. Implications of these data with respect to the enzymatic mechanism and the structure of the enzyme-bound factor are discussed.     

The immune response and the eye. III. Anterior chamber-associated immune deviation can be adoptively transferred by serum

After the anterior chamber (AC) injection of trinitrophenol-coupled (TNP) spleen cells, it is observed that systemic delayed-type hypersensitivity responses to TNP are inhibited by Ag-specific suppressor T cells. We recently reported that suppression is initiated by viable TNP-coupled T cells within the inoculum and upon further analysis we found that these cells have the surface phenotype of CD4+ Ts inducer cells. We report here that treatment of these TNP-T cells with cycloheximide or cytochalasin-B before to AC injection abolishes suppression, whereas treatment with 2000 rad radiation does not. This indicates that protein synthesis and secretion are required to initiate suppression but proliferation is not. Further, we demonstrate the adoptive transfer of suppression by serum of AC inoculated animals. Detection of the component in serum in adoptive transfer assays, however, requires removal of the spleen before AC injection. We establish that the material in serum is a Ts cell product (T suppressor-inducer factor) based on three criteria: it is Ag specific, genetically restricted, and reactive with a mAb that specifically identifies these molecules. These results suggest that the signal leaving the eye to induce suppression of delayed-type hypersensitivity is T cell derived and that molecules mediating immune regulation for this organ are made within the eye and transported via the serum to the spleen.     

Rate of release of Ca2+ following laser photolysis of the DM-nitrophen-Ca2+ complex.

The determination of the rate of release of Ca2+ by pulsed photolysis of the photolabile chelator DM-nitrophen is important for its use in time-resolved physiological studies: the rate of substrate or effector release should be faster than the processes they initiate. Flash photolysis of DM-nitrophen using a 50-ns pulse from a frequency-doubled ruby laser (with emission at 347 nm having energy of ca. 10-20 mJ) yields short-lived photochromic or aci-nitro intermediates. At pH 6.9, double-exponential decay of a photochromic intermediate was observed for DM-nitrophen itself and its Ca2+ complex (tau 1/2 values of 24 and 570 microseconds, and 32 and 220 microseconds respectively), while only monoexponential decay of the DM-nitrophen-Mg2+ complex was detected (tau 1/2 = 31 microseconds). Only the photochemistry of DM-nitrophen-Ca2+ was found to be pH sensitive (monoexponential decay, tau 1/2 approximately 115 microseconds at pH 7.9 and 8.9). Use of the Ca(2+)-sensitive metallochromic dye antipyrylazo III in conjunction with pulsed photolysis of DM-nitrophen-Ca2+ enabled an upper limit of the half-time of release of Ca2+ to be established of ca. 180 microseconds (the rate of association of Ca2+ with the dye was probably rate determining). The rate of Ca2+ photorelease may, however, be faster than this. Thus, the DM-nitrophen-Ca2+ complex releases Ca2+ on photolysis sufficiently rapidly for the study of many Ca(2+)-dependent physiological processes with improved kinetic resolution over conventional mixing methods.     

Ecotoxicological contamination processes: Interaction with vegetation (a review)

Up-to-date crop production concentrates on the maximisation of plant matter production, both by optimum utilization of solar energy by plants and by the application of additional energy. Therefore a natural strategy is being developed aimed at combining the physiological functions of the assimilation organs with their external shaping and strengthening their variety in space and area. As demonstrated by many deposition studies, this effect is contradictory to the factors conditioning the low deposition capacity of plant covers. A high level of particle, gas and solution deposition increases the danger of food chain contamination and has a negative impact on plant production. Data concentrated on transport values of pollutants and their long-term dynamics in stands could be used for the selection of ideal crop types; therefore every effort should be directed towards selection and breeding.     

Stream vegetation in different landscape types

A methodology is proposed for assessing the ecological value of streams in the catchment of the Moravská Dyje River. It is concluded that by using a wide range of parameters that a more objective assessment is achieved than if only one were used. The landscape of the catchment studied contains excessive amounts of nutrients and, in comparison to its natural state, has become too uniform.