An electron microscopic study of neuronal degeneration and glial cell reaction in the retina of glaucomatous rats

The present investigation was focused on the ultrastructural changes in the neurons and glial cells in the retina of rats with experimentally-induced glaucoma. An experimental glaucoma model was created by limbal-derived vein cauterization. Animals were sacrificed at 1, 3 weeks and 3 months post-operation. Retinae were dissected and processed for electron microscopy. Neuronal degeneration was observed in all the different layers of the retina at both 1 and 3 weeks post-operation. Some degenerating neurons were found in the ganglion cell layer (GCL), inner nuclear layer (INL) and outer nuclear layer (ONL). And the dying neurons presented apoptotic-like more than necrotic neurons. Many degenerating axons and axon terminals were observed between neurons in the GCL, inner plexiform layer (IPL), INL, and outer plexiform layer (OPL). Activated astrocytes and microglial cells were present in close association with degenerating neurons and axons. The Müller cells in the INL also presented longer and darker processes with more microfilaments than in normal cells. Degenerating neuronal debris, degenerating axonal profiles and electron-dense bodies were often found in the cytoplasm of macrophages. The results suggest that both microglial cells and astrocytes are activated in the process of neuronal degeneration in the retina of experimentally-induced glaucomatous rats. It is hypothesized that they may play a protective role in removing degenerating neuronal elements in the retina after the onset of glaucoma.     

Bioaugmentation and coexistence of two functionally similar bacterial strains in aerobic granules

The survival of the inoculated microbial culture is critical for successful bioaugmentation but impossible to predict precisely. As an alternative strategy, bioaugmentation of a group of microorganisms may improve reliability of bioaugmentation. This study evaluated simultaneous bioaugmentation of two functionally similar bacterial strains in aerobic granules. The two strains, Pandoraea sp. PG-01 and Rhodococcus erythropolis PG-03, showed high phenol degradation and growth rates in phenol medium, but they were characterized as having a poor aggregation activity and weak bioflocculant-producing and biofilm-forming abilities. In the spatially homogeneous batch conditions, strain PG-01 with higher growth rates outcompeted strain PG-03. However, the two strains could stably coexist in the spatially heterogeneous conditions. Then the two strains were mixed and bioaugmented into activated sludge in two sequencing batch reactors, which were operated with the different settling times of 5 and 30 min, respectively. Aerobic granules were developed only in the reactor with a settling time of 5 min. Fluorescence in situ hybridization and denaturing gradient gel electrophoresis showed that the two strains could coexist in aerobic granules but not in activated sludge. These findings suggested that the compact structure of aerobic granules provided spatial isolation for coexistence of competitively superior and inferior strains with similar functions.     

Effect of human chorionic gonadotrophin on 7,12-dimethylbenz[a]anthracene-induced DNA binding and repair synthesis by rat mammary epithelial cells

The administration of the placental hormone human chorionic gonadotrophin (HCG) to 50-day-old virgin Sprague--Dawley rats has been shown to reduce the incidence of 7,12-dimethylbenz[a]anthracene (DMBA)-induced mammary cancer. We now report from studies using rat mammary epithelial cells in culture that the anti-carcinogenic effect of HCG may be related to its effect on DNA binding of DMBA and on DNA repair. The results showed that the level of excision repair in cells derived from young virgin (YV) rats grown in the presence of HCG (10 units/ml) was 2.5-4.0 times higher than the level exhibited by control YV cells and 1.5-2.5 times over that obtained for cells from old virgin and parous rats. The effect of HCG on DMBA-DNA binding was also determined in YV cells cultured in the presence of HCG (10 units/ml). Results from this study indicated that DMBA-DNA binding was inhibited by 30-40% in HCG-treated cells as compared to control cells. DNA binding of DMBA was also determined with mammary epithelial cells from YV rats which were given subcutaneous injections of HCG (5 units/rat) 5 times per week for 4 weeks. Using this in vivo-in vitro protocol, DMBA-DNA binding was 17-51% lower in cells from HCG-treated rats than in cells derived from control saline-treated rats. These results suggest that the protective effect provided by HCG against DMBA-induced mammary tumorigenesis may be attributed to its ability to inhibit binding of the carcinogen to mammary cell DNA and to its ability to increase the level of excision repair in the cells.     

In vivo effects of monoclonal antibody to ICAM-1 (CD54) in nonhuman primates with renal allografts

These studies test whether allograft rejection can be blocked by interference with leukocyte adhesion, using a murine IgG2a mAb (R6.5) reactive with monkey ICAM-1 (CD54). In 16 Cynomolgus renal allograft recipients, R6.5 was administered prophylactically as the sole immunosuppressive agent for 12 days (0.01 to 2 mg/kg/day). Survival in 14 recipients with technically successful grafts was significantly prolonged (24.2 +/- 2.4 vs 9.2 +/- 0.6 days for controls; p less than 0.001). Intercellular adhesion molecule-1 (CD54) (ICAM-1) was expressed on vascular endothelium in the kidney and other organs in the monkey in a pattern similar to that in humans. During cellular rejection in controls, ICAM-1 expression increased on endothelial cells, infiltrating mononuclear leukocytes and tubular cells. Biopsies during R6.5 administration showed decreased T cell infiltration (CD2, CD8, CD4) compared with controls and decreased arterial endothelial inflammation. No changes occurred in circulating T cells, aside from variable coating with mIgG. In six of eight other recipients R6.5 administration (0.5 to 2 mg/kg/day for 10 days) reversed preexisting rejection that resulted from taper of Cyclosporine to subtherapeutic levels. Responding grafts showed decreased edema and hemorrhage but no consistent change in the infiltrate. At 1 h after the first dose, mouse IgG deposited primarily on the graft vascular endothelium without any change in the inflammatory infiltrate. Mouse IgG also deposited on the endothelium of normal organs without eliciting an inflammatory response and was cleared from the endothelium within 4 days. Inasmuch as the principal site of binding was the vascular endothelium, we hypothesize that the antibody blocks adhesion to graft ICAM-1 molecules on the vessels. Anti-ICAM-1 also binds to recipient cells and may interfere with Ag presentation and/or T cell interactions. Whatever the mechanism(s), these studies indicate that an anti-ICAM-1 antibody inhibits T cell mediated injury in vivo, and that ICAM-1 is a critical molecule in the pathogenesis of allograft rejection.     

A novel property of spider silk: chemical defence against ants

Spider webs are made of silk, the properties of which ensure remarkable efficiency at capturing prey. However, remaining on, or near, the web exposes the resident spiders to many potential predators, such as ants. Surprisingly, ants are rarely reported foraging on the webs of orb-weaving spiders, despite the formidable capacity of ants to subdue prey and repel enemies, the diversity and abundance of orb-web spiders, and the nutritional value of the web and resident spider. We explain this paradox by reporting a novel property of the silk produced by the orb-web spider Nephila antipodiana (Walckenaer). These spiders deposit on the silk a pyrrolidine alkaloid (2-pyrrolidinone) that provides protection from ant invasion. Furthermore, the ontogenetic change in the production of 2-pyrrolidinone suggests that this compound represents an adaptive response to the threat of natural enemies, rather than a simple by-product of silk synthesis: while 2-pyrrolidinone occurs on the silk threads produced by adult and large juvenile spiders, it is absent on threads produced by small juvenile spiders, whose threads are sufficiently thin to be inaccessible to ants.     

Break-induced loss of heterozygosity in fission yeast: dual roles for homologous recombination in promoting translocations and preventing de novo telomere addition

Loss of heterozygosity (LOH), a causal event in tumorigenesis, frequently encompasses multiple genetic loci and whole chromosome arms. However, the mechanisms leading to such extensive LOH are poorly understood. We investigated the mechanisms of DNA double-strand break (DSB)-induced extensive LOH by screening for auxotrophic marker loss approximately 25 kb distal to an HO endonuclease break site within a nonessential minichromosome in Schizosaccharomyces pombe. Extensive break-induced LOH was infrequent, resulting from large translocations through both allelic crossovers and break-induced replication. These events required the homologous recombination (HR) genes rad32(+), rad50(+), nbs1(+), rhp51(+), rad22(+), rhp55(+), rhp54(+), and mus81(+). Surprisingly, LOH was still observed in HR mutants, which resulted predominantly from de novo telomere addition at the break site. De novo telomere addition was most frequently observed in rad22Delta and rhp55Delta backgrounds, which disrupt HR following end resection. Further, levels of de novo telomere addition, while increased in ku70Delta rhp55Delta strains, were reduced in exo1Delta rhp55Delta and an rhp55Delta strain overexpressing rhp51. These findings support a model in which HR prevents de novo telomere addition at DSBs by competing for resected ends. Together, these results suggest that the mechanisms of break-induced LOH may be predicted from the functional status of the HR machinery.     

Iron-dependent production of a heat-modifiable, 23,000-Mr outer membrane protein in Paracoccus denitrificans.

Production of a 23,000-Mr major outer membrane protein of Paracoccus denitrificans ATCC 13543 was dependent upon the addition of iron to a succinate-salts medium. The 23,000-Mr protein was not produced in an iron-deficient medium, but production of five outer membrane proteins in the 85,000- to 72,000-Mr range and of catechol were induced. The 23,000-Mr protein was not produced in a complex medium even when ferric citrate was added to the medium. Production of the protein was influenced by the carbon source and was decreased by peptone.     

Identification of an integral plasma membrane pool of protein kinase C in mammalian tissues and cells

Protein kinase C (PKC) is a family of serine/threonine protein kinases that are pivotal in cellular regulation. Since its discovery in 1977, PKCs have been known as cytosolic and peripheral membrane proteins. However, there are reports that PKC can insert into phospholipids vesicles in vitro. Given the intimate relationship between the plasma membrane and the activation of PKC, it is important to determine whether such "membrane-inserted" form of PKC exists in mammalian cells or tissues. Here, we report the identification of an integral plasma membrane pool for all the 10 PKC isozymes in vivo by their ability to partition into the detergent-rich phase in Triton X-114 phase partitioning, and by their resistance to extractions with 0.2 M sodium carbonate (pH 11.5), 2 M urea and 2 M sodium chloride. The endogenous integral membrane pool of PKC in mouse fibroblasts is found to be acutely regulated by phorbol ester or diacylglycerol, suggesting that this pool of PKC may participate in cellular processes known to be regulated by PKC. At least for PKC(alpha), the C2-V3 region at the regulatory domain of the kinase is responsible for membrane integration. Further exploration of the function of this novel integral plasma membrane pool of PKC will not only shed new light on molecular mechanisms underlying its cellular functions but also provide new strategies for pharmaceutical modulation of this important group of kinases.     

Antimicrobial activity of extracts of chemical races of the lichen Pseudevernia furfuracea and their physodic acid, chloroatranorin, atranorin, and olivetoric acid constituents

The antimicrobial activity and the MIC values of the ethanol, chloroform, diethyl ether, and acetone extracts of the chemical races of Pseudevernia furfuracea (var. furfuracea and var. ceratea) and their physodic acid, chloroatranorin, atranorin, and olivetoric acid constituents have been investigated against some microorganisms. Nearly all extracts of both chemical races showed antimicrobial activity against Aeromonas hydrophila, Bacillus cereus, Bacillus subtilis, Listeria monocytogenes, Proteus vulgaris, Staphylococcus aureus, Streptococcus faecalis, Yersinia enterocolitica, Candida albicans, Candida glabrata, Alternaria alternata, Ascochyta rabiei, Aspergillus niger, Fusarium culmorum, Fusarium moniliforme, Fusarium oxysporum, Fusarium solani, and Penicillium notatum. There was no antimicrobial activity of the extracts against Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, Pseudomonas syringae, Salmonella typhimurium, Alternaria citri, Alternaria tenuissima, and Gaeumannomyces graminis. Chloroatranorin and olivetoric acid were active against the same microorganisms with few exceptions. Physodic acid was active against about the same bacteria and yeasts and inactive against all of the filamentous fungi tested. Also no activity of atranorin against the filamentous fungi was observed.     

Modulatory Effects of the Piccolo Genotype on Emotional Memory in Health and Depression

Major depressive disorder (MDD) has been associated with biased memory formation for mood-congruent information, which may be related to altered monoamine levels. The piccolo (PCLO) gene, involved in monoaminergic neurotransmission, has previously been linked to depression in a genome-wide association study. Here, we investigated the role of the PCLO risk allele on functional magnetic resonance imaging (MRI) correlates of emotional memory in a sample of 89 MDD patients (64 PCLO risk allele carriers) and 29 healthy controls (18 PCLO risk allele carriers). During negative word encoding, risk allele carriers showed significant lower activity relative to non-risk allele carriers in the insula, and trend-wise in the anterior cingulate cortex and inferior frontal gyrus. Moreover, depressed risk allele carriers showed significant lower activity relative to non-risk allele carriers in the striatum, an effect which was absent in healthy controls. Finally, amygdalar response during processing new positive words vs. known words was blunted in healthy PCLO+ carriers and in MDD patients irrespective of genotype, which may indicate that signalling of salient novel information does not occur to the same extent in PCLO+ carriers and MDD patients. The PCLO risk allele may increase vulnerability for MDD by modulating local brain function with regard to responsiveness to salient stimuli (i.e. insula) and processing novel negative information. Also, depression-specific effects of PCLO on dorsal striatal activation during negative word encoding and the absence of amygdalar salience signalling for novel positive information further suggest a role of PCLO in symptom maintenance in MDD.