A pathway through interferon-gamma is the main pathway for induction of nitric oxide upon stimulation with bacterial lipopolysaccharide in mouse peritoneal cells.

Production of nitric oxide (NO) in response to bacterial lipopolysaccharide (LPS) was investigated using cultures of mouse peritoneal exudate cells (PEC) and the macrophage cell line RAW264.7. In the presence of anti-(interferon-gamma) (IFN-gamma), NO production was markedly suppressed in the PEC culture but not in the RAW264.7 culture. In the PEC culture, LPS induced both IFN-gamma production and activation of IFN response factor-1, which leads to the gene expression of inducible NO synthase, but neither was induced in the culture of RAW264.7 cells. In addition to anti-(IFN-gamma), antibodies against interleukin (IL)-12 and IL-18 showed a suppressive effect on LPS-induced NO production in the PEC culture, and these antibodies in synergy showed strong suppression. Stimulation of the PEC culture with IL-12 or IL-18 induced production of IFN-gamma and NO, and these cytokines, in combination, exhibited marked synergism. Stimulation of the culture with IFN-gamma induced production of NO, but not IL-12. The macrophage population in the PEC, prepared as adherent cells, responded well to LPS for IL-12 production, but weakly for production of IFN-gamma and NO. The macrophages also responded well to IFN-gamma for NO production. For production of IFN-gamma by stimulation with LPS or IL-12 + IL-18, nonadherent cells were required in the PEC culture. Considering these results overall, the indirect pathway, through the production of intermediates (such as IFN-gamma-inducing cytokines and IFN-gamma) by the cooperation of macrophages with nonadherent cells, was revealed to play the main role in the LPS-induced NO production pathway, as opposed to the direct pathway requiring only a macrophage population.     

Stable transformation of cultured cells of the liverwortMarchantia polymorpha by particle bombardment

Suspension-cultured cells (A-18 line) of the liverwortMarchanta polymorpha were bombarded by a pneumatic particle gun with plasmid pCH harbouring the hygromycin phosphotransferase (HPT) gene (hpt) under the control of the cauliflower mosaic virus (CaMV) 35 S promoter and the nopaline synthase polyadenylation region. Nine weeks after bombardments, 128 hygromycin-resistant calluses were obtained from an approximate total of 7×10⁶ cells. Ten cell lines chosen randomly were analysed further. Southern blot analysis showed that all of the ten lines contain thehpt gene in the genome, demonstrating that these lines are transformants. An HPT enzyme activity assay confirmed the expression of the gene in all of the transformant lines.     

FISH analysis of the telomere sequences of bulldog ants (Myrmecia: Formicidae)

Chromosomes from several species of ants from the genus Myrmecia were hybridized with deoxyoligomer probes of either (T₂AG₂)₇, the putative insect telomere repeat sequence, or (T₂AG₃)₇, the vertebrate telomere repeat sequence. While both sequence hibridized over a range of stringency conditions, (T₂AG₂)_n was clearly the predominant sequence at the termini of the Myrmecia chromosomes. No interstitial sites of either sequence were detected. The genus Myrmecia has a wide range of karyotypes, with chromosome numbers ranging from 2n=2–84. It has been hypothesized that the ancestral karyotype was 2n=4 and karyotype evolution proceeded with an increase in chromosome number. In the absence of detectable interstitial sites of telomere sequence, it is interesting to speculate on the origin of the new telomeres as the chromosome numbers increased.     

Lack of invaginations of the plasma membrane during budding and cell division of Saccharomyces cerevisiae and Schizosaccharomyces pombe

Abstract The plasma membrane of Saccharomyces cerevisiae and Schizosaccharomyces pombe in stationary phase had abundant invaginations. A round uninvaginated area emerged before budding when S. cerevisiae cells were given fresh medium. Middle-sized buds had some invaginations, whereas the neck between the bud and mother had very few. S. pombe which has neither the neck nor the predetermined position to divide had no uninvaginated ring area even in long cells during elongation in fresh medium. However, an uninvaginated ring area emerged as the earliest noticeable stage of cytokinesis. The uninvaginated state of the plasma membrane appeared to be correlated with budding and cell division.     

Cloning and nucleotide sequences of the linear DNA killer plasmids from yeast.

The linear DNA killer plasmids (pGKL1 and pGKL2) isolated from a Kluyveromyces lactis killer strain are also maintained and expressed its killer character in Saccharomyces cerevisiae. After these killer plasmid DNAs isolated from S. cerevisiae were treated with alkali, four terminal fragments from each plasmid DNAs were cloned separately. Using these and other cloned DNA fragments, the terminal nucleotide sequences of pGKL2 and the complete nucleotide sequence of pGKL1 were determined. The inverted terminal repetitions of 202 bp and 182 bp were found in pGKL1 and pGKL2, respectively. The pGKL1 sequence showed an extremely high A + T content of 73.2% and it contained five large open reading frames. The largest of these open reading frame was suggested to code for a membrane-bound precursor of glycoprotein subunit of the killer toxin.     

Effects of taurine and γ-aminobutyric acid on akinesia and analgesia induced by D-Ala2-Met-enkephalinamide in rats

Effects of taurine or γ-aminobutyric acid (GABA) on akinesia and analgesia induced by D-Ala²-Met-enkephalinamide were investigated in rats. Administration of taurine (dose range: 2.375×10^?2 M–9.5×10^?2 M/10 μl) into the left lateral ventricle 10 min prior to the injection of D-Ala²-Met-enkephalinamide (50 μg/10 μl) produced a dose-dependent reduction in the duration of akinesia and to some extent of analgesia, as estimated at 30 min and 60 min following the enkephalinamide injection; at the first estimation-time (10 min), taurine did not alter the duration of akinesia or that of analgesia. The median effective dose (ED₅₀) for akinesia determined at 60 min after D-Ala²-Met-enkephalinamide was 5 times greater and that for analgesia assessed at the same time was 1.7 times greater in taurine-treated rats than the respective doses in control animals. Administration of GABA under similar experimental conditions produced a dose-dependent reduction in the duration of analgesia from the initial estimation time (10 min) following the injection of D-Ala²-Met-enkephalinamide. The ED₅₀ for analgesia determined at 30 min after D-Ala²-Met-enkephalinamide was 3 times greater in GABA-treated rats than in control animals. Unlike the effects of taurine, GABA did not alter the duration of akinesia. Neither the duration of akinesia nor that of analgesia was modified by taurine or GABA alone in rats tested 9 min after the injection of each amino acid. These findings suggest that taurine may promote a recovery from both akinesia and analgesia, while GABA decreases only the analgesia induced by D-Ala²-Met-enkephalinamide.     

Persistence of genomes of both herpesvirus of turkeys and Marek's disease virus in a chicken T-lymphoblastoid cell line

A cell line tentatively designated as MDCC-BO1(T), was established from spleen cells of an apparently healthy chicken inoculated with herpesvirus of turkey (HVT). BO1(T) cells were T lymphoblastoid cells and the more than 95% of them had Marek's disease (MD) tumor-associated surface antigen (MATSA). However, no viral internal antigens or membrane antigens could be demonstrated in them by immunofluorescence tests using chicken anti-HVT and -MD virus (MDV) sera. The virus could be rescued from BO1(T) cells by co-cultivation with chick embryo fibroblasts (CEF). The DNA of the rescued virus was characterized as HVT DNA by its sedimentation profile in a neutral glycerol gradient and its endonuclease Hind III cleavage-pattern. Ultrastructural studies on CEF infected with the rescued virus revealed the presence of HVT-like virions. However, DNA-DNA reassociation kinetics showed that the BO1(T) cells contained a few copies of NVT and also MDV genomes.     

Further morphological characterization and structural proteins of infectious bursal disease virus.

An outer layer surrounding the capsid of infectious bursal disease virus was evident from electron micrographs of intact virus particles having diameters of 62 to 63 nm. The capsid was found to be composed of large morphological units or capsomeres, measuring about 12 nm in diameter. The architecture of the capsid appears to be that of T = 3 symmetry, with a probable 32 morphological units by rotational enhancement of image detail. Structural proteins of infectious bursal disease virus consist of seven species, two major and five minor polypeptides. These are P1 to P7, with molecular weights of 133 x 10(3), 124 x 10(3), 98 x 10(3), 51 x 10(3), 33 x 10(3), 26 x 10(3), and 23 x 10(3), respectively.     

Dose-response relations for dicentric yields in G0 lymphocytes on man and crab-eating monkey following acute and chronic γ-irradiations

A comparison has been made of dicentric yields in G₀ lymphocytes between man and crab-eating monkey, Macaca fascicularis, after acute and chronic γ-irradiations. With acute irradiation (49.6 rad/min) there was no significant difference between them, but for the chronic irradiation (17.1 rad/h) a significant difference was observed between the species. When the dose-response relations were fitted to the linear-quadratic model (Y = αD + βD²), the species-difference observed for chronic irradiation was almost entirely due to change in the value of β. In addition, after chronic irradiation the β-value for monkey was almost negligible, but that for man was significant. Post-irradiation incubation experiment showed that cells with dicentrics were partly eliminated during the course of chronic irradiation, because there were appreciable reductions of dicentric yields (ca. 25% for both man and monkey at 400 rad) together with mitotic indices (ca. 30% and 60% for man and monkey, respectively, at 400 rad). Accordingly, it would be reasonable to postulate that G₀ repair for dicentrics other than selection mechanism must play a major role in the effects of low dose rate. It can be further suggested that G₀-repair capacity for chromosal damages leading to dicentrics may be different among different primate species.