Leukotrienes cause eosinophil emigration into conjunction tissue

The ability of LTB₄, LTC₄, the 5S,6R and 5R,6S LTD₄ stereoisomers, and LTE₄ to evoke leukocyte infiltration into the conjunctiva was demonstrated in the guinea pig by histological andl ight microscopy techniques. LTD₄ and LTE₄ demonstrated a dose-dependent and predominantly eosinophilic infiltrate over the selected dose range (10ng to 1000ng), while there was only a minimal response to LTC₄. LTB₄ produced marked eosinophil inflitrates only at the highest dose; scattered neutrophil infiltrates were also noted at the high dose of LTB₄. The 5R,6S LTD₄ stereoisomer did not evoke any leukocyte infiltrantion. The SRS-A antagonist, FPL 55712, abolished pre-treatment had no inhibitory effect, indicating direct mediation of this response by LTs. Histamine caused a comparable eosinophilia over a dose range of 10μg to 1000μg. LT-induced eosinophil emigration was directed to the conjunctival epithelium; the cells appeared intact and no tissue damage was observed. These results may have relevance in the areas of allergic conjunctivitis and asthma research.     

Morphological and ultrastructural studies on protoplast production and reversion of the higher basidiomyceteAgaricus bisporus

Protoplasts have been isolated from young vegetative mycelia ofAgaricus bisporus by an enzyme mixture of novozym and chitinase. Protoplasts were released through ruptures in the wall, initially at the apices, but also later from older parts of the hyphae, indicating that they may lack the cell wall. The process of regeneration of these protoplasts has been investigated in liquid medium in which the protoplasts produced short chains of convoluted cells that finally produced a hypha. Electron microscopy has shown that at the start of regeneration two different kinds of fibrils were produced at the external surface of the protoplasts. Later, the thickness of the cell wall increased, and there was a deposit of amorphous material giving rise to a complete new wall.     

Dopamine and α-synuclein dysfunction in Smad3 null mice

Background

Recent studies suggest that the pathogenic process in neurodegenerative disorders may disrupt mature neuronal circuitries and neurogenesis in the adult brain. Abnormal activation of CDK5 is associated with neurodegenerative disorders, and recently a critical role for CDK5 in adult neurogenesis has been identified. We have developed an in vitro model of abnormal CDK5 activation during adult hippocampal neurogenesis, and here we used this model to investigate aberrantly phosphorylated downstream targets of CDK5.

Results

Abnormal CDK5 activation in an in vitro model of adult neurogenesis results in hyperphosphorylation of collapsin-response mediator protein-2 (CRMP2) and impaired neurite outgrowth. Inhibition of CDK5, or expression of a non-phosphorylatable (S522A) CRMP2 construct reduced CRMP2 hyperphosphorylation, and reversed neurite outgrowth deficits. CRMP2 plays a role in microtubule dynamics; therefore we examined the integrity of microtubules in this model using biochemical and electron microscopy techniques. We found that microtubule organization was disrupted under conditions of CDK5 activation. Finally, to study the relevance of these findings to neurogenesis in neurodegenerative conditions associated with HIV infection, we performed immunochemical analyses of the brains of patients with HIV and transgenic mice expressing HIV-gp120 protein. CDK5-mediated CRMP2 phosphorylation was significantly increased in the hippocampus of patients with HIV encephalitis and in gp120 transgenic mice, and this effect was rescued by genetic down-modulation of CDK5 in the mouse model.

Conclusions

These results reveal a functional mechanism involving microtubule destabilization through which abnormal CDK5 activation and CRMP2 hyperphosphorylation might contribute to defective neurogenesis in neurodegenerative disorders such as HIV encephalitis.     

Four distinct patterns of memory CD8 T cell responses to chronic murine cytomegalovirus infection

CMVs are beta herpesviruses that establish lifelong latent infection of their hosts. Acute infection of C57BL/6 mice with murine CMV elicits a very broad CD8 T cell response, comprising at least 24 epitopes from 18 viral proteins. In contrast, we show here that the CD8 T cell response in chronically infected mice was dominated by only five epitopes. Altogether, four distinct CD8 T cell kinetic patterns were evident. Responses to some epitopes, including M45, which dominates the acute response, contracted sharply after day 7 and developed into stable long-term memory. The response to m139 underwent rapid expansion and contraction, followed by a phase of memory inflation, whereas the response to an M38 epitope did not display any contraction phase. Finally, responses against two epitopes encoded by the immediate early gene IE3 were readily detectable in chronically infected mice but near the limit of detection during acute infection. CD8 T cells specific for the noninflationary M45 epitope displayed a classic central memory phenotype, re-expressing the lymph node homing receptor CD62L and homeostatic cytokine receptors for IL-7 and IL-15, and produced low levels of IL-2. Responses to two inflationary epitopes, m139 and IE3, retained an effector memory surface phenotype (CD62L(low), IL-7Ralpha(-), IL-15Rbeta(-)) and were unable to produce IL-2. We suggest that immunological choices are superimposed on altered viral gene expression profiles to determine immunodominance during chronic murine CMV infection.     

Patterns of growth and tract formation during the early development of secondary lineages in the Drosophila larval brain

The Drosophila brain consists of a relatively small number of invariant, genetically determined lineages which provide a model to study the relationship between gene function and neuronal architecture. In following this long‐term goal, we reconstruct the morphology (projection pattern and connectivity) and gene expression patterns of brain lineages throughout development. In this article, we focus on the secondary phase of lineage morphogenesis, from the reactivation of neuroblast proliferation in the first larval instar to the time when proliferation ends and secondary axon tracts have fully extended in the late third larval instar. We have reconstructed the location and projection of secondary lineages at close (4 h) intervals and produced a detailed map in the form of confocal z‐projections and digital three‐dimensional models of all lineages at successive larval stages. Based on these reconstructions, we could compare the spatio‐temporal pattern of axon formation and morphogenetic movements of different lineages in normal brain development. In addition to wild type, we reconstructed lineage morphology in two mutant conditions. (1) Expressing the construct UAS‐p35 which rescues programmed cell death we could systematically determine which lineages normally lose hemilineages to apoptosis. (2) so‐Gal4‐driven expression of dominant‐negative EGFR ablated the optic lobe, which allowed us to conclude that the global centrifugal movement normally affecting the cell bodies of lateral lineages in the late larva is causally related to the expansion of the optic lobe, and that the central pattern of axonal projections of these lineages is independent of the presence or absence of the optic lobe. © 2015 Wiley Periodicals, Inc. Develop Neurobiol 76: 434–451, 2016     

A BchD (magnesium chelatase) mutant of rhodobacter sphaeroides synthesizes zinc bacteriochlorophyll through novel zinc-containing intermediates

Heme and bacteriochlorophyll a (BChl) biosyntheses share the same pathway to protoporphyrin IX, which then branches as follows. Fe(2+) chelation into the macrocycle by ferrochelatase results in heme formation, and Mg(2+) addition by Mg-chelatase commits the porphyrin to BChl synthesis. It was recently discovered that a bchD (Mg-chelatase) mutant of Rhodobacter sphaeroides produces an alternative BChl in which Mg(2+) is substituted by Zn(2+). Zn-BChl has been found in only one other organism before, the acidophilic Acidiphilium rubrum. Our objectives in this work on the bchD mutant were to 1) elucidate the Zn-BChl biosynthetic pathway in this organism and 2) understand causes for the low amounts of Zn-BChl produced. The bchD mutant was found to contain a Zn-protoporphyrin IX pool, analogous to the Mg-protoporphyrin IX pool found in the wild type strain. Inhibition of ferrochelatase with N-methylprotoporphyrin IX caused Zn-protoporphyrin IX and Zn-BChl levels to decline by 80-90% in the bchD mutant, whereas in the wild type strain, Mg-protoporphyrin IX and Mg-BChl levels increased by 170-240%. Two early metabolites of the Zn-BChl pathway were isolated from the bchD mutant and identified as Zn-protoporphyrin IX monomethyl ester and divinyl-Zn-protochlorophyllide. Our data support a model in which ferrochelatase synthesizes Zn-protoporphyrin IX, and this metabolite is acted on by enzymes of the BChl pathway to produce Zn-BChl. Finally, the low amounts of Zn-BChl in the bchD mutant may be due, at least in part, to a bottleneck upstream of the step where divinyl-Zn-protochlorophyllide is converted to monovinyl-Zn-protochlorophyllide.     

Arterial oxygen saturation in healthy newborns delivered at term in Cerro de Pasco (4340 m) and Lima (150 m)

Background  

High altitude is associated with both low pulse oxygen saturation at birth and more pre-term deliveries. The present study was performed to determine pulse oxygen saturation in newborns at term in Cerro de Pasco (4340 m) and Lima (150 m) to test the hypothesis that low pulse oxygen saturation at birth at high altitudes was not observed at term deliveries.     

Production of the gaseous signal molecule hydrogen sulfide in mouse tissues

The gaseous molecule hydrogen sulfide (H₂S) has been proposed as an endogenous signal molecule and neuromodulator in mammals. Using a newly developed method, we report here for the first time the ability of intact and living brain and colonic tissue in the mouse to generate and release H₂S. This production occurs through the activity of two enzymes, cystathionine-γ-lyase and cystathionine-β-synthase. The quantitative expression of messenger RNA and protein localization for both enzymes are described in the liver, brain, and colon. Expression levels of the enzymes vary between tissues and are differentially distributed. The observation that, tissues that respond to exogenously applied H₂S can endogenously generate the gas, strongly supports its role as an endogenous signal molecule.