Cell density-induced changes in lipid composition and intracellular trafficking

Cell density is one of the extrinsic factors to which cells adapt their physiology when grown in culture. However, little is known about the molecular changes which occur during cell growth and how cellular responses are then modulated. In many cases, inhibitors, drugs or growth factors used for in vitro studies change the rate of cell proliferation, resulting in different cell densities in control and treated samples. Therefore, for a comprehensive data analysis, it is essential to understand the implications of cell density on the molecular level. In this study, we have investigated how lipid composition changes during cell growth, and the consequences it has for transport of Shiga toxin. By quantifying 308 individual lipid species from 17 different lipid classes, we have found that the levels and species distribution of several lipids change during cell growth, with the major changes observed for diacylglycerols, phosphatidic acids, cholesterol esters, and lysophosphatidylethanolamines. In addition, there is a reduced binding and retrograde transport of Shiga toxin in high density cells which lead to reduced intoxication by the toxin. In conclusion, our data provide novel information on how lipid composition changes during cell growth in culture, and how these changes can modulate intracellular trafficking.     

Whole-genome analysis of diverse Chlamydia trachomatis strains identifies phylogenetic relationships masked by current clinical typing

Chlamydia trachomatis is responsible for both trachoma and sexually transmitted infections, causing substantial morbidity and economic cost globally. Despite this, our knowledge of its population and evolutionary genetics is limited. Here we present a detailed phylogeny based on whole-genome sequencing of representative strains of C. trachomatis from both trachoma and lymphogranuloma venereum (LGV) biovars from temporally and geographically diverse sources. Our analysis shows that predicting phylogenetic structure using ompA, which is traditionally used to classify Chlamydia, is misleading because extensive recombination in this region masks any true relationships present. We show that in many instances, ompA is a chimera that can be exchanged in part or as a whole both within and between biovars. We also provide evidence for exchange of, and recombination within, the cryptic plasmid, which is another key diagnostic target. We used our phylogenetic framework to show how genetic exchange has manifested itself in ocular, urogenital and LGV C. trachomatis strains, including the epidemic LGV serotype L2b.     

Today’s and tomorrow’s imaging and circulating biomarkers for pulmonary arterial hypertension

The pathobiology of pulmonary arterial hypertension (PAH) involves a remodeling process in distal pulmonary arteries, as well as vasoconstriction and in situ thrombosis, leading to an increase in pulmonary vascular resistance, right heart failure and death. Its etiology may be idiopathic, but PAH is also frequently associated with underlying conditions such as connective tissue diseases. During the past decade, more than welcome novel therapies have been developed and are in development, including those increasingly targeting the remodeling process. These therapeutic options modestly increase the patients' long-term survival, now approaching 60% at 5 years. However, non-invasive tools for confirming PAH diagnosis, and assessing disease severity and response to therapy, are tragically lacking and would help to select the best treatment. After exclusion of other causes of pulmonary hypertension, a final diagnosis still relies on right heart catheterization, an invasive technique which cannot be repeated as often as an optimal follow-up might require. Similarly, other techniques and biomarkers used for assessing disease severity and response to treatment generally lack specificity and have significant limitations. In this review, imaging as well as current and future circulating biomarkers for diagnosis, prognosis, and follow-up are discussed.     

Wnt signalling regulates adult hippocampal neurogenesis

The generation of new neurons from neural stem cells is restricted to two regions of the adult mammalian central nervous system: the subventricular zone of the lateral ventricle, and the subgranular zone of the hippocampal dentate gyrus. In both regions, signals provided by the microenvironment regulate the maintenance, proliferation and neuronal fate commitment of the local stem cell population. The identity of these signals is largely unknown. Here we show that adult hippocampal stem/progenitor cells (AHPs) express receptors and signalling components for Wnt proteins, which are key regulators of neural stem cell behaviour in embryonic development. We also show that the Wnt/beta-catenin pathway is active and that Wnt3 is expressed in the hippocampal neurogenic niche. Overexpression of Wnt3 is sufficient to increase neurogenesis from AHPs in vitro and in vivo. By contrast, blockade of Wnt signalling reduces neurogenesis from AHPs in vitro and abolishes neurogenesis almost completely in vivo. Our data show that Wnt signalling is a principal regulator of adult hippocampal neurogenesis and provide evidence that Wnt proteins have a role in adult hippocampal function.     

Using short-term surveys and mark–recapture to estimate diversity and population size of orchid bees in forest formations of the Brazilian savanna

Studies on the diversity and population dynamics of euglossine bees in forest formations of the Brazilian savanna can be notably informative considering that most species are forest-dependent. The current study aimed to estimate the diversity and population size of euglossine bees using short-term surveys and the mark–recapture method. We also compared short-term surveys with monthly surveys over a 1-year period (long-term surveys) to assess the species diversity. The study occurred in seven forest formations of the Brazilian savanna. We collected 14 species and marked 375 males but recaptured only 23 (6.1%). The population size of Euglossa bees ranged from 111.1 ± 34.4 to 1384.5 ± 384.6 males. The diversity indices achieved through the short-term surveys were similar to or higher than those obtained from long-term studies. The low recapture rate of euglossine males seems to be the result of their long-distance dispersal capacity. We also proposed that short-term surveys be viewed as a favourable alternative to long-term studies on diversity estimation and that euglossine bees can occur in small populations in some forest formations studied.     

Effect of light at night on the pineal rhythm in N-acetyltransferase activity in the Syrian hamsterMesocricetus auratus

Summary Pineal N-acetyltransferase activity in the male Syrian hamster exhibited a daily rhythm; the maximal night-time value was 3.5-fold higher than the day-time value. When hamsters were exposed to light at night N-acetyltransferase declined within 30 min to 1/5 of its former activity. These results indicate that in the Syrian hamster the pineal melatonin rhythm may be regulated at least partly via changes in N-acetyltransferase activity.The authors are grateful to Mrs Marie Svobodová for her skillful technical assistance.     

Blocking VEGFR-3 suppresses angiogenic sprouting and vascular network formation

Angiogenesis, the growth of new blood vessels from pre-existing vasculature, is a key process in several pathological conditions, including tumour growth and age-related macular degeneration. Vascular endothelial growth factors (VEGFs) stimulate angiogenesis and lymphangiogenesis by activating VEGF receptor (VEGFR) tyrosine kinases in endothelial cells. VEGFR-3 (also known as FLT-4) is present in all endothelia during development, and in the adult it becomes restricted to the lymphatic endothelium. However, VEGFR-3 is upregulated in the microvasculature of tumours and wounds. Here we demonstrate that VEGFR-3 is highly expressed in angiogenic sprouts, and genetic targeting of VEGFR-3 or blocking of VEGFR-3 signalling with monoclonal antibodies results in decreased sprouting, vascular density, vessel branching and endothelial cell proliferation in mouse angiogenesis models. Stimulation of VEGFR-3 augmented VEGF-induced angiogenesis and sustained angiogenesis even in the presence of VEGFR-2 (also known as KDR or FLK-1) inhibitors, whereas antibodies against VEGFR-3 and VEGFR-2 in combination resulted in additive inhibition of angiogenesis and tumour growth. Furthermore, genetic or pharmacological disruption of the Notch signalling pathway led to widespread endothelial VEGFR-3 expression and excessive sprouting, which was inhibited by blocking VEGFR-3 signals. Our results implicate VEGFR-3 as a regulator of vascular network formation. Targeting VEGFR-3 may provide additional efficacy for anti-angiogenic therapies, especially towards vessels that are resistant to VEGF or VEGFR-2 inhibitors.