Bcl-2 protects cells from cytokine-induced nitric-oxide-dependent apoptosis

 Cytokine-mediated cell death in tumor cells can be achieved through endogenous nitric oxide (NO) from within tumor cells or exogenous NO from either activated macrophages or endothelial cells. The purpose of this study was to determine the role of Bcl-2 in NO-mediated apoptosis. The incubation of murine L929 and NIH3T3 cells with interleukin-1α (IL-1α) and interferon γ (IFNγ) induced high endogenous NO production only in the L929 cells that also underwent apoptosis. NIH3T3 cells were not resistant to NO-mediated apoptosis. In fact, the incubation of L929 and NIH3T3 cells with exogenous NO derived from NO donors, sodium nitroprusside, or S-nitroso-N-acetyl-DL-penicillamine (SNAP) induced death, characterized by typical apoptotic morphology and DNA fragmentation, in both cell types, but to a higher degree in NIH3T3 cells than in the L929 cells. We then measured the effect of Bcl-2 expression on exogenous NO-induced apoptosis. At both the mRNA and protein levels, L929 fibroblasts expressed higher levels of endogenous mouse Bcl-2 than did NIH3T3 cells. At the same time, L929 cells were much more resistant to exogenous NO-induced cell death than were NIH3T3 cells. The inverse correlation between mouse Bcl-2 expression and sensitivity to exogenous NO-mediated cell death was also found in the murine K-1735 melanoma C-23 and X-21 clonal populations. Transfection of both NIH3T3 cells and L929 cells with the human bcl-2 gene led to resistance to both exogenous and endogenous NO-mediated apoptosis. These data demonstrate that NO-mediated apoptosis can be suppressed by expression of Bcl-2, suggesting that abnormal expression of Bcl-2 may influence the efficacy of tumor immunotherapy. Received: 28 June 1998 / Accepted: 23 August 1996     

Rapid warming and drought negatively impact population size and reproductive dynamics of an avian predator in the arid southwest

Avian communities of arid ecosystems may be particularly vulnerable to global climate change due to the magnitude of projected change for desert regions and the inherent challenges for species residing in resource limited ecosystems. How arid‐zone birds will be affected by rapid increases in air temperature and increased drought frequency and severity is poorly understood because avian responses to climate change have primarily been studied in the relatively mesic northern temperate regions. We studied the effects of increasing air temperature and aridity on a Burrowing Owl (Athene cunicularia) population in the southwestern United States from 1998 to 2013. Over 16 years, the breeding population declined 98.1%, from 52 pairs to 1 pair, and nest success and fledgling output also declined significantly. These trends were strongly associated with the combined effects of decreased precipitation and increased air temperature. Arrival on the breeding grounds, pair formation, nest initiation, and hatch dates all showed significant delays ranging from 9.4 to 25.1 days over 9 years, which have negative effects on reproduction. Adult and juvenile body mass decreased significantly over time, with a loss of 7.9% mass in adult males and 10.9% mass in adult females over 16 years, and a loss of 20.0% mass in nestlings over 8 years. Taken together, these population and reproductive trends have serious implications for local population persistence. The southwestern United States has been identified as a climate change hotspot, with projections of warmer temperatures, less winter precipitation, and an increase in frequency and severity of extreme events including drought and heat waves. An increasingly warm and dry climate may contribute to this species' decline and may already be a driving force of their apparent decline in the desert southwest.     

Molecular histology of arteries: mass spectrometry imaging as a novel ex vivo tool to investigate atherosclerosis

Atherosclerosis is usually the underlying cause of a fatal event such as myocardial infarction or ictus. The atherome plaque develops silently and asymptomatically within the arterial intima layer. In this context, the possibility to analyze the molecular content of arterial tissue while preserving each molecule’s specific localization is of great interest as it may reveal further insights into the physiopathological changes taking place. Mass spectrometry imaging (MSI) enables the spatially resolved molecular analysis of proteins, peptides, metabolites, lipids and drugs directly in tissue, with a resolution sufficient to reveal molecular features specific to distinct arterial structures. MSI represents a novel ex vivo imaging tool still underexplored in cardiovascular diseases. This review focuses on the MSI technique applied to cardiovascular disease and covers the main contributions to date, ongoing efforts, the main challenges and current limitations of MSI.     

The Effect of an Acute Bout of Moderate-Intensity Aerobic Exercise on Motor Learning of a Continuous Tracking Task

Introduction

There is evidence for beneficial effects of acute and long-term exercise interventions on several forms of memory, including procedural motor learning. In the present study we examined how performing a single bout of continuous moderate intensity aerobic exercise would impact motor skill acquisition and retention in young healthy adults, compared to a period of rest. We hypothesized that exercise would improve motor skill acquisition and retention, compared to motor practice alone.

Materials and Methods

Sixteen healthy adults completed sessions of aerobic exercise or seated rest that were immediately followed by practice of a novel motor task (practice). Exercise consisted of 30 minutes of continuous cycling at 60% peak O₂ uptake. Twenty-four hours after practice, we assessed motor learning with a no-exercise retention test (retention). We also quantified changes in offline motor memory consolidation, which occurred between practice and retention (offline). Tracking error was separated into indices of temporal precision and spatial accuracy.

Results

There were no differences between conditions in the timing of movements during practice (p = 0.066), at retention (p = 0.761), or offline (p = 0.966). However, the exercise condition enabled participants to maintain spatial accuracy during practice (p = 0.477); whereas, following rest performance diminished (p = 0.050). There were no significant differences between conditions at retention (p = 0.532) or offline (p = 0.246).

Discussion

An acute bout of moderate-intensity aerobic exercise facilitated the maintenance of motor performance during skill acquisition, but did not influence motor learning. Given past work showing that pairing high intensity exercise with skilled motor practice benefits learning, it seems plausible that intensity is a key modulator of the effects of acute aerobic exercise on changes in complex motor behavior. Further work is necessary to establish a dose-response relationship between aerobic exercise and motor learning.     

Parallel reverse genetic screening in mutant human cells using transcriptomics

Reverse genetic screens have driven gene annotation and target discovery in model organisms. However, many disease‐relevant genotypes and phenotypes cannot be studied in lower organisms. It is therefore essential to overcome technical hurdles associated with large‐scale reverse genetics in human cells. Here, we establish a reverse genetic approach based on highly robust and sensitive multiplexed RNA sequencing of mutant human cells. We conduct 10 parallel screens using a collection of engineered haploid isogenic cell lines with knockouts covering tyrosine kinases and identify known and unexpected effects on signaling pathways. Our study provides proof of concept for a scalable approach to link genotype to phenotype in human cells, which has broad applications. In particular, it clears the way for systematic phenotyping of still poorly characterized human genes and for systematic study of uncharacterized genomic features associated with human disease.     

Hypoxia-Induced Changes in DNA Methylation Alter RASAL1 and TGFβ1 Expression in Human Trabecular Meshwork Cells

PurposeFibrosis and a hypoxic environment are associated with the trabecular meshwork (TM) region in the blinding disease glaucoma. Hypoxia has been shown to alter DNA methylation, an epigenetic mechanism involved in regulating gene expression such as the pro-fibrotic transforming growth factor (TGF) β1 and the anti-fibrotic Ras protein activator like 1 (RASAL1). The purpose of this study was to compare DNA methylation levels, and the expression of TGFβ1 and RASAL1 in primary human normal (NTM) with glaucomatous (GTM) cells and in NTM cells under hypoxic conditions.MethodsGlobal DNA methylation was assessed by ELISA in cultured age-matched NTM and GTM cells. qPCR was conducted for TGFβ1, collagen 1α1 (COL1A1), and RASAL1 expression. Western immunoblotting was used to determine protein expression. For hypoxia experiments, NTM cells were cultured in a 1%O₂, 5%CO₂ and 37°C environment. NTM and GTM cells were treated with TGFβ1 (10ng/ml) and the methylation inhibitor 5-azacytidine (5-aza) (0.5μM) respectively to determine their effects on DNA Methyltransferase 1 (DNMT1) and RASAL1 expression.ResultsWe found increased DNA methylation, increased TGFβ1 expression and decreased RASAL1 expression in GTM cells compared to NTM cells. Similar results were obtained in NTM cells under hypoxic conditions. TGFβ1 treatment increased DNMT1 and COL1A1, and decreased RASAL1 expression in NTM cells. 5-aza treatment decreased DNMT1, TGFβ1 and COL1A1 expression, and increased RASAL1 expression in GTM cells.ConclusionsTGFβ1 and RASAL1 expression, global DNA methylation, and expression of associated methylation enzymes were altered between NTM and GTM cells. We found that hypoxia in NTM cells induced similar results to the GTM cells. Furthermore, DNA methylation, TGFβ1 and RASAL1 appear to have an interacting relationship that may play a role in driving pro-fibrotic disease progression in the glaucomatous TM.