Rapid patterning of Dictyostelium discoideum cells under confined geometry and its relation to differentiation

The following was recently reported by Bonner et al. (1995): (1) Rapid differentiation occurred into two zones in Dictyostelium discoideum cells confined in a fine glass capillary. The cells in the anterior zone exposed to the air appear similar to prestalk cells, while the posterior zone isolated from the air mimics prespore cells. (2) The volumes of the two zones are proportional to each other for different sized cell masses, and the proportion is the same as that in normal migrating slugs. We investigated the nature of this newly discovered rapid differentiation in a slightly modified geometry. Exponentially growing cells were harvested, washed to remove external nutrients, and pelleted by centrifugation. Subsequently, a small drop of the pelleted (starved) cells was placed on a slide glass and then confined in a two-dimensional space between the slide glass and a coverslip, with help of spacers whose thickness varied from 25 to 100 μm. As a result, a dark zone, which looked optically different, emerged within several minutes in the periphery of the disc of the confined cells, corresponding to the zonation in a capillary as previously reported. When the width of the peripheral zone was measured for more than 30 samples of different diameters for each thickness of the spacers, the width was found to be always about 100 μm, irrespective of the size difference of the cell mass placed. This seems to be contradictory to the previous observation made by Bonner et al. (1995). We also examined oxygen concentration dependence on the zone width. The zone width was found to be independent of the oxygen concentration at low concentrations, but increased rapidly at high concentrations. A reaction-diffusion mechanism for formation of the zone and possible involvement of atmospheric oxygen (O₂) in the initial steps of cell differentiation and pattern formation is discussed.     

Induction of globin mRNA accumulation by hemin in cultured erythroleukemic cells

The role of heme in erythroid development is investigated in erythroleukemic (Friend) cells. Exogenous hemin induces the accumulation of globin mRNA and globin protein in T3-Cl2 erythroleukemia cells to levels comparable to those induced by polar solvents, such as dimethylsulfoxide (DMSO). The hemin concentration required for maximal induction (10^?4 M) is the same as that which stimulates globin message translation in reticulocytes or cell-free reticulocyte lysates. Hemin and DMSO together cause T3-Cl2 cells to accumulate 8–9 fold more globin mRNA than either inducer individually. The kinetics of globin mRNA induction in hemin as compared to DMSO are very different: globin message accumulation begins 4 hr after hemin addition, but not until 30–40 hr after DMSO addition. Biliverdin induces 20–40 fold less hemoglobin than hemin; delta-aminolevulinic acid and porphobilinogen do not induce.     

Response Suppression Delays the Planning of Subsequent Stimulus-Driven Saccades

The completion of an antisaccade selectively increases the reaction time (RT) of a subsequent prosaccade: a result that has been interpreted to reflect the residual inhibition of stimulus-driven saccade networks [1], [2]. In the present investigation we sought to determine whether the increase in prosaccade RT is contingent on the constituent antisaccade planning processes of response suppression and vector inversion or is limited to response suppression. To that end, in one block participants alternated between pro- and antisaccades after every second trial (task-switching block), and in another block participants completed a series of prosaccades that were randomly (and infrequently) interspersed with no-go catch-trials (go/no-go block). Notably, such a design provides a framework for disentangling whether response suppression and/or vector inversion delays the planning of subsequent prosaccades. As expected, results for the task-switching block showed that antisaccades selectively increased the RTs of subsequent prosaccades. In turn, results for the go/no-go block showed that prosaccade RTs were increased when preceded by a no-go catch-trial. Moreover, the magnitude of the RT ‘cost’ was equivalent across the task-switching and go/no-go blocks. That prosaccades preceded by an antisaccade or a no-go catch-trial produced equivalent RT costs indicates that the conjoint processes of response suppression and vector inversion do not drive the inhibition of saccade planning mechanisms. Rather, the present findings indicate that a general consequence of response suppression is a residual inhibition of stimulus-driven saccade networks.     

Surface Tensiometry of Apolipoprotein B Domains at Lipid Interfaces Suggests a New Model for the Initial Steps in Triglyceride-rich Lipoprotein Assembly

Apolipoprotein B (apoB) is the principal protein component of triacylglyceride (TAG)-rich lipoproteins, including chylomicrons and very low density lipoprotein, which is the precursor to LDL (the “bad cholesterol”). TAG-rich lipoprotein assembly is initiated by the N-terminal βα1 superdomain of apoB, which co-translationally binds and remodels the luminal leaflet of the rough endoplasmic reticulum. The βα1 superdomain contains four domains and is predicted to interact directly with lipids. Using drop tensiometry, we examined the interfacial properties of the α-helical and C-sheet domains and several subdomains to establish a detailed structure-function relationship at the lipid/water interface. The adsorption, stress response, exchangeability, and pressure (Π)-area relationship were studied at both triolein/water and triolein/1-palmitoyl, 2-oleoylphosphatidylcholine/water interfaces that mimic physiological environments. The α-helical domain spontaneously adsorbed to a triolein/water interface and formed a viscoelastic surface. It was anchored to the surface by helix 6, and the other helices were ejected and/or remodeled on the surface as a function of surface pressure. The C-sheet instead formed an elastic film on a triolein/water interface and was irreversibly anchored to the lipid surface, which is consistent with the behavior of amphipathic β-strands. When both domains were adsorbed together on the surface, the C-sheet shielded a portion of the α-helical domain from the surface, which retained its globular structure. Overall, the unique secondary and tertiary structures of the N-terminal domains of apoB support the intrinsic capability of co-translational lipid recruitment. The evidence presented here allows the construction of a detailed model of the initiation of TAG-rich lipoprotein assembly.     

On the Prospect of Identifying Adaptive Loci in Recently Bottlenecked Populations

Identifying adaptively important loci in recently bottlenecked populations – be it natural selection acting on a population following the colonization of novel habitats in the wild, or artificial selection during the domestication of a breed – remains a major challenge. Here we report the results of a simulation study examining the performance of available population-genetic tools for identifying genomic regions under selection. To illustrate our findings, we examined the interplay between selection and demography in two species of Peromyscus mice, for which we have independent evidence of selection acting on phenotype as well as functional evidence identifying the underlying genotype. With this unusual information, we tested whether population-genetic-based approaches could have been utilized to identify the adaptive locus. Contrary to published claims, we conclude that the use of the background site frequency spectrum as a null model is largely ineffective in bottlenecked populations. Results are quantified both for site frequency spectrum and linkage disequilibrium-based predictions, and are found to hold true across a large parameter space that encompasses many species and populations currently under study. These results suggest that the genomic footprint left by selection on both new and standing variation in strongly bottlenecked populations will be difficult, if not impossible, to find using current approaches.     

CD8 T cell-initiated blood-brain barrier disruption is independent of neutrophil support

Blood-brain barrier (BBB) disruption is a common feature of numerous neurologic disorders. A fundamental question in these diseases is the extent inflammatory immune cells contribute to CNS vascular permeability. We have previously shown that CD8 T cells play a critical role in initiating BBB disruption in the peptide-induced fatal syndrome model developed by our laboratory. However, myelomonocytic cells such as neutrophils have also been implicated in promoting CNS vascular permeability and functional deficit in murine models of neuroinflammatory disease. For this reason, we evaluated neutrophil depletion in a murine model of CD8 T cell-initiated BBB disruption by employing traditionally used anti-granulocyte receptor-1 mAb RB6-8C5 and Ly-6G-specific mAb 1A8. We report that CNS-infiltrating antiviral CD8 T cells express high levels of granulocyte receptor-1 protein and are depleted by treatment with RB6-8C5. Mice treated with RB6-8C5, but not 1A8, display: 1) intact BBB tight junction proteins; 2) reduced CNS vascular permeability visible by gadolinium-enhanced T1-weighted magnetic resonance imaging; and 3) preservation of motor function. These studies demonstrate that traditional methods of neutrophil depletion with RB6-8C5 are broadly immune ablating. Our data also provide evidence that CD8 T cells initiate disruption of BBB tight junction proteins and CNS vascular permeability in the absence of neutrophil support.     

A susceptibility gene for psoriatic arthritis maps to chromosome 16q: evidence for imprinting

Several genetic loci have been reported for psoriasis, but none has been specifically linked to psoriatic arthritis (PsA), a condition that affects >10% of patients with psoriasis. A genetic component for PsA is suggested by segregation within families and high concordance among identical twins. We performed a linkage scan to map genes contributing to PsA. We identified 178 patients with PsA out of 906 patients who were included in our genetic study of psoriasis. Using a comprehensive genealogy database, we were able to connect 100 of these into 39 families. We genotyped the patients using a framework marker set of 1,000 microsatellite markers, with an average density of 3 cM, and performed multipoint, affected-only, allele-sharing linkage analysis using the Allegro program. On the basis of the initial results, we genotyped more markers for the most prominent loci. A linkage with a LOD score of 2.17 was observed on chromosome 16q. The linkage analysis, conditioned on paternal transmission to affected individuals, gave a LOD score of 4.19, whereas a LOD score of only 1.03 was observed when conditioned for maternal transmission. A suggestive locus on chromosome 16q has previously been implicated in psoriasis. Our data indicate that a gene at this locus may be involved in paternal transmission of PsA.     

Pericardial fat and myocardial perfusion in asymptomatic adults from the Multi-Ethnic Study of Atherosclerosis

Background

Pericardial fat has adverse effects on the surrounding vasculature. Previous studies suggest that pericardial fat may contribute to myocardial ischemia in symptomatic individuals. However, it is unknown if pericardial fat has similar effects in asymptomatic individuals.

Methods

We determined the association between pericardial fat and myocardial blood flow (MBF) in 214 adults with no prior history of cardiovascular disease from the Minnesota field center of the Multi-Ethnic Study of Atherosclerosis (43% female, 56% Caucasian, 44% Hispanic). Pericardial fat volume was measured by computed tomography. MBF was measured by MRI at rest and during adenosine-induced hyperemia. Myocardial perfusion reserve (PR) was calculated as the ratio of hyperemic to resting MBF.

Results

Gender-stratified analyses revealed significant differences between men and women including less pericardial fat (71.9±31.3 vs. 105.2±57.5 cm³, p<0.0001) and higher resting MBF (1.12±0.23 vs. 0.93±0.19 ml/min/g, p<0.0001), hyperemic MBF (3.49±0.76 vs. 2.65±0.72 ml/min/g, p<0.0001), and PR (3.19±0.78 vs. 2.93±0.89, p = 0.03) in women. Correlations between pericardial fat and clinical and hemodynamic variables were stronger in women. In women only (p = 0.01 for gender interaction) higher pericardial fat was associated with higher resting MBF (p = 0.008). However, this association was attenuated after accounting for body mass index or rate-pressure product. There were no significant associations between pericardial fat and hyperemic MBF or PR after multivariate adjustment in either gender. In logistic regression analyses there was also no association between impaired coronary vasoreactivity, defined as having a PR <2.5, and pericardial fat in men (OR, 1.18; 95% CI, 0.82–1.70) or women (OR, 1.11; 95% CI, 0.68–1.82).

Conclusions

Our data fail to support an independent association between pericardial fat and myocardial perfusion in adults without symptomatic cardiovascular disease. Nevertheless, these findings highlight potentially important differences between asymptomatic and symptomatic individuals with respect to the underlying subclinical disease burden.