Familial adenomatous polyposis is associated with a marked decrease in alkaline sphingomyelinase activity: a key factor to the unrestrained cell proliferation?

The hydrolysis of sphingomyelin generates key molecules regulating cell growth and inducing apoptosis. Data from animal cancer models support an inhibitory role for this pathway in the malignant transformation of the colonic mucosa. In the intestinal tract, a sphingomyelinase with an optimum alkaline pH has been identified. We recently found that the activity of alkaline sphingomyelinase is significantly decreased in colorectal adenocarcinomas, indicating a potential anticarcinogenic role of this enzyme. To further examine whether the reduction of sphingomyelinase is present already in the premalignant state of neoplastic transformation, we measured sphingomyelinase activities in patients with familial adenomatous polyposis (FAP) and in sporadic colorectal tubulovillous adenomas. Tissue samples were taken from adenomas and surrounding macroscopically normal mucosa from 11 FAP patients operated with ileorectal anastomosis, from three FAP patients with intact colon, from 13 patients with sporadic colorectal adenomas and from 12 controls. Activities of acid, neutral and alkaline sphingomyelinase were measured together with alkaline phosphatase. In FAP adenoma tissue, alkaline sphingomyelinase activity was reduced by 90% compared to controls (P < 0.0001), acid sphingomyelinase by 66% (P < 0.01) and neutral sphingomyelinase by 54% (P < 0.05). Similar reductions were found in the surrounding mucosa. In sporadic adenoma tissue, only alkaline sphingomyelinase was reduced significantly, by 57% (P < 0.05). Alkaline phosphatase was not changed in FAP adenomas, but decreased in the sporadic adenomas. We conclude that the markedly reduced levels of alkaline sphingomyelinase activities in FAP adenomas and in the surrounding mucosa may be a pathogenic factor that can lead to unrestrained cell proliferation and neoplastic transformation. © 1999 Cancer Research Campaig

False negative rates in Drosophila cell-based RNAi screens: a case study

<p>Abstract</p> <p>Background</p> <p>High-throughput screening using RNAi is a powerful gene discovery method but is often complicated by false positive and false negative results. Whereas false positive results associated with RNAi reagents has been a matter of extensive study, the issue of false negatives has received less attention.</p> <p>Results</p> <p>We performed a meta-analysis of several genome-wide, cell-based <it>Drosophila </it>RNAi screens, together with a more focused RNAi screen, and conclude that the rate of false negative results is at least 8%. Further, we demonstrate how knowledge of the cell transcriptome can be used to resolve ambiguous results and how the number of false negative results can be reduced by using multiple, independently-tested RNAi reagents per gene.</p> <p>Conclusions</p> <p>RNAi reagents that target the same gene do not always yield consistent results due to false positives and weak or ineffective reagents. False positive results can be partially minimized by filtering with transcriptome data. RNAi libraries with multiple reagents per gene also reduce false positive and false negative outcomes when inconsistent results are disambiguated carefully.</p

The Aedes aegypti Toll Pathway Controls Dengue Virus Infection

Aedes aegypti, the mosquito vector of dengue viruses, utilizes its innate immune system to ward off a variety of pathogens, some of which can cause disease in humans. To date, the features of insects' innate immune defenses against viruses have mainly been studied in the fruit fly Drosophila melanogaster, which appears to utilize different immune pathways against different types of viruses, in addition to an RNA interference–based defense system. We have used the recently released whole-genome sequence of the Ae. aegypti mosquito, in combination with high-throughput gene expression and RNA interference (RNAi)-based reverse genetic analyses, to characterize its response to dengue virus infection in different body compartments. We have further addressed the impact of the mosquito's endogenous microbial flora on virus infection. Our findings indicate a significant role for the Toll pathway in regulating resistance to dengue virus, as indicated by an infection-responsive regulation and functional assessment of several Toll pathway–associated genes. We have also shown that the mosquito's natural microbiota play a role in modulating the dengue virus infection, possibly through basal-level stimulation of the Toll immune pathway

Forebrain NR2B Overexpression Facilitating the Prefrontal Cortex Long-Term Potentiation and Enhancing Working Memory Function in Mice

Prefrontal cortex plays an important role in working memory, attention regulation and behavioral inhibition. Its functions are associated with NMDA receptors. However, there is little information regarding the roles of NMDA receptor NR2B subunit in prefrontal cortical synaptic plasticity and prefrontal cortex-related working memory. Whether the up-regulation of NR2B subunit influences prefrontal cortical synaptic plasticity and working memory is not yet clear. In the present study, we measured prefrontal cortical synaptic plasticity and working memory function in NR2B overexpressing transgenic mice. In vitro electrophysiological data showed that overexpression of NR2B specifically in the forebrain region resulted in enhancement of prefrontal cortical long-term potentiation (LTP) but did not alter long-term depression (LTD). The enhanced LTP was completely abolished by a NR2B subunit selective antagonist, Ro25-6981, indicating that overexpression of NR2B subunit is responsible for enhanced LTP. In addition, NR2B transgenic mice exhibited better performance in a set of working memory paradigms including delay no-match-to-place T-maze, working memory version of water maze and odor span task. Our study provides evidence that NR2B subunit of NMDA receptor in prefrontal cortex is critical for prefrontal cortex LTP and prefrontal cortex-related working memory

A Short Receptor Downregulates JAK/STAT Signalling to Control the Drosophila Cellular Immune Response

Regulation of JAK/STAT signalling by a short, nonsignalling receptor in Drosophila modulates response to specific immune challenges such as parasitoid infestations

The Role of Glypicans in Wnt Inhibitory Factor-1 Activity and the Structural Basis of Wif1's Effects on Wnt and Hedgehog Signaling

Proper assignment of cellular fates relies on correct interpretation of Wnt and Hedgehog (Hh) signals. Members of the Wnt Inhibitory Factor-1 (WIF1) family are secreted modulators of these extracellular signaling pathways. Vertebrate WIF1 binds Wnts and inhibits their signaling, but its Drosophila melanogaster ortholog Shifted (Shf) binds Hh and extends the range of Hh activity in the developing D. melanogaster wing. Shf activity is thought to depend on reinforcing interactions between Hh and glypican HSPGs. Using zebrafish embryos and the heterologous system provided by D. melanogaster wing, we report on the contribution of glypican HSPGs to the Wnt-inhibiting activity of zebrafish Wif1 and on the protein domains responsible for the differences in Wif1 and Shf specificity. We show that Wif1 strengthens interactions between Wnt and glypicans, modulating the biphasic action of glypicans towards Wnt inhibition; conversely, glypicans and the glypican-binding “EGF-like” domains of Wif1 are required for Wif1's full Wnt-inhibiting activity. Chimeric constructs between Wif1 and Shf were used to investigate their specificities for Wnt and Hh signaling. Full Wnt inhibition required the “WIF” domain of Wif1, and the HSPG-binding EGF-like domains of either Wif1 or Shf. Full promotion of Hh signaling requires both the EGF-like domains of Shf and the WIF domains of either Wif1 or Shf. That the Wif1 WIF domain can increase the Hh promoting activity of Shf's EGF domains suggests it is capable of interacting with Hh. In fact, full-length Wif1 affected distribution and signaling of Hh in D. melanogaster, albeit weakly, suggesting a possible role for Wif1 as a modulator of vertebrate Hh signaling

Resolving the neural circuits of anxiety

Although anxiety disorders represent a major societal problem demanding new therapeutic targets, these efforts have languished in the absence of a mechanistic understanding of this subjective emotional state. While it is impossible to know with certainty the subjective experience of a rodent, rodent models hold promise in dissecting well-conserved limbic circuits. The application of modern approaches in neuroscience has already begun to unmask the neural circuit intricacies underlying anxiety by allowing direct examination of hypotheses drawn from existing psychological concepts. This information points toward an updated conceptual model for what neural circuit perturbations could give rise to pathological anxiety and thereby provides a roadmap for future therapeutic development.National Institute of Diabetes and Digestive and Kidney Diseases (U.S.) (NIH Director’s New Innovator Award DP2-DK-102256-01)National Institute of Mental Health (U.S.) (NIH) R01-MH102441-01)JPB Foundatio

History-Dependent Excitability as a Single-Cell Substrate of Transient Memory for Information Discrimination

Neurons react differently to incoming stimuli depending upon their previous history of stimulation. This property can be considered as a single-cell substrate for transient memory, or context-dependent information processing: depending upon the current context that the neuron “sees” through the subset of the network impinging on it in the immediate past, the same synaptic event can evoke a postsynaptic spike or just a subthreshold depolarization. We propose a formal definition of History-Dependent Excitability (HDE) as a measure of the propensity to firing in any moment in time, linking the subthreshold history-dependent dynamics with spike generation. This definition allows the quantitative assessment of the intrinsic memory for different single-neuron dynamics and input statistics. We illustrate the concept of HDE by considering two general dynamical mechanisms: the passive behavior of an Integrate and Fire (IF) neuron, and the inductive behavior of a Generalized Integrate and Fire (GIF) neuron with subthreshold damped oscillations. This framework allows us to characterize the sensitivity of different model neurons to the detailed temporal structure of incoming stimuli. While a neuron with intrinsic oscillations discriminates equally well between input trains with the same or different frequency, a passive neuron discriminates better between inputs with different frequencies. This suggests that passive neurons are better suited to rate-based computation, while neurons with subthreshold oscillations are advantageous in a temporal coding scheme. We also address the influence of intrinsic properties in single-cell processing as a function of input statistics, and show that intrinsic oscillations enhance discrimination sensitivity at high input rates. Finally, we discuss how the recognition of these cell-specific discrimination properties might further our understanding of neuronal network computations and their relationships to the distribution and functional connectivity of different neuronal types