Examination of the Cytotoxic and Embryotoxic Potential and Underlying Mechanisms of Next-Generation Synthetic Trioxolane and Tetraoxane Antimalarials

Semisynthetic artemisinin-based therapies are the first-line treatment for P. falciparum malaria, but next-generation synthetic drug candidates are urgently required to improve availability and respond to the emergence of artemisinin-resistant parasites. Artemisinins are embryotoxic in animal models and induce apoptosis in sensitive mammalian cells. Understanding the cytotoxic propensities of antimalarial drug candidates is crucial to their successful development and utilization. Here, we demonstrate that, similarly to the model artemisinin artesunate (ARS), a synthetic tetraoxane drug candidate (RKA182) and a trioxolane equivalent (FBEG100) induce embryotoxicity and depletion of primitive erythroblasts in a rodent model. We also show that RKA182, FBEG100 and ARS are cytotoxic toward a panel of established and primary human cell lines, with caspase-dependent apoptosis and caspase-independent necrosis underlying the induction of cell death. Although the toxic effects of RKA182 and FBEG100 proceed more rapidly and are relatively less cell-selective than that of ARS, all three compounds are shown to be dependent upon heme, iron and oxidative stress for their ability to induce cell death. However, in contrast to previously studied artemisinins, the toxicity of RKA182 and FBEG100 is shown to be independent of general chemical decomposition. Although tetraoxanes and trioxolanes have shown promise as next-generation antimalarials, the data described here indicate that adverse effects associated with artemisinins, including embryotoxicity, cannot be ruled out with these novel compounds, and a full understanding of their toxicological actions will be central to the continuing design and development of safe and effective drug candidates which could prove important in the fight against malaria.     

Becoming Multicellular by Aggregation; The Morphogenesis of the Social Amoebae Dicyostelium discoideum

The organisation and form of most organisms is generated during theirembryonic development and involves precise spatial and temporal controlof cell division, cell death, cell differentiation and cell movement.Differential cell movement is a particularly important mechanism in thegeneration of form. Arguably the best understood mechanism of directedmovement is chemotaxis. Chemotaxis plays a major role in the starvationinduced multicellular development of the social amoebae Dictyostelium.Upon starvation up to 10⁵ individual amoebae aggregate to form afruiting body. In this paper we review the evidence that the movement ofthe cells during all stages of Dictyostelium development is controlled bypropagating waves of cAMP which control the chemotactic movement ofthe cells. We analyse the complex interactions between cell-cell signallingresulting in cAMP waves of various geometries and cell movement whichresults in a redistribution of the signalling sources and therefore changes thegeometry of the waves. We proceed to show how the morphogenesis,including aggregation stream and mound formation, slug formation andmigration, of this relatively simple organism is beginning to be understoodat the level of rules for cell behaviour, which can be tested experimentallyand theoretically by model calculations.     

Seeing through the static: the temporal dimension of plant–animal mutualistic interactions

Most studies of plant–animal mutualistic networks have come from a temporally static perspective. This approach has revealed general patterns in network structure, but limits our ability to understand the ecological and evolutionary processes that shape these networks and to predict the consequences of natural and human‐driven disturbance on species interactions. We review the growing literature on temporal dynamics of plant–animal mutualistic networks including pollination, seed dispersal and ant defence mutualisms. We then discuss potential mechanisms underlying such variation in interactions, ranging from behavioural and physiological processes at the finest temporal scales to ecological and evolutionary processes at the broadest. We find that at the finest temporal scales (days, weeks, months) mutualistic interactions are highly dynamic, with considerable variation in network structure. At intermediate scales (years, decades), networks still exhibit high levels of temporal variation, but such variation appears to influence network properties only weakly. At the broadest temporal scales (many decades, centuries and beyond), continued shifts in interactions appear to reshape network structure, leading to dramatic community changes, including loss of species and function. Our review highlights the importance of considering the temporal dimension for understanding the ecology and evolution of complex webs of mutualistic interactions.     

Aurothiomalate as an ultrastructural marker. Electron microscopy and x-ray microanalysis of various tissues after in vivo gold injections.

Rabbits and rats were given single injections of aurothiomalate by different routes. The animals were killed at progressive intervals, and sections from various organs were examined by electron microscopic x-ray microanalysis. Ultrastructurally, characteristic material was regularly found in vacuoles, dense and heterogeneous bodies of macrophages, hepatocytes and renal epithelial cells. Occasionally, other mesenchymal cells also contained gold. Histochemical and analytical tests showed that the gold-containing organelles were devoid of acid phosphatase activity. The generalized rapid spread, retention and selectivity of localization after a single small dose make aurothiomalate a useful marker substance for ultrastructural studies.     

Prediction uncertainty of environmental change effects on temperate European biodiversity

Observed patterns of species richness at landscape scale (gamma diversity) cannot always be attributed to a specific set of explanatory variables, but rather different alternative explanatory statistical models of similar quality may exist. Therefore predictions of the effects of environmental change (such as in climate or land cover) on biodiversity may differ considerably, depending on the chosen set of explanatory variables. Here we use multimodel prediction to evaluate effects of climate, land-use intensity and landscape structure on species richness in each of seven groups of organisms (plants, birds, spiders, wild bees, ground beetles, true bugs and hoverflies) in temperate Europe. We contrast this approach with traditional best-model predictions, which we show, using cross-validation, to have inferior prediction accuracy. Multimodel inference changed the importance of some environmental variables in comparison with the best model, and accordingly gave deviating predictions for environmental change effects. Overall, prediction uncertainty for the multimodel approach was only slightly higher than that of the best model, and absolute changes in predicted species richness were also comparable. Richness predictions varied generally more for the impact of climate change than for land-use change at the coarse scale of our study. Overall, our study indicates that the uncertainty introduced to environmental change predictions through uncertainty in model selection both qualitatively and quantitatively affects species richness projections.     

The control of chemotactic cell movement during Dictyostelium morphogenesis

Differential cell movement is an important mechanism in the development and morphogenesis of many organisms. In many cases there are indications that chemotaxis is a key mechanism controlling differential cell movement. This can be particularly well studied in the starvation-induced multicellular development of the social amoeba Dictyostelium discoideum. Upon starvation, up to 10(5) individual amoebae aggregate to form a fruiting body The cells aggregate by chemotaxis in response to propagating waves of cAMP, initiated by an aggregation centre. During their chemotactic aggregation the cells start to differentiate into prestalk and prespore cells, precursors to the stalk and spores that form the fruiting body. These cells enter the aggregate in a random order but then sort out to form a simple axial pattern in the slug. Our experiments strongly suggest that the multicellular aggregates (mounds) and slugs are also organized by propagating cAMP waves and, furthermore, that cell-type-specific differences in signalling and chemotaxis result in cell sorting, slug formation and movement.     

Requirements for stress granule recruitment of fused in sarcoma (FUS) and TAR DNA-binding protein of 43 kDa (TDP-43)

Cytoplasmic inclusions containing TAR DNA-binding protein of 43 kDa (TDP-43) or Fused in sarcoma (FUS) are a hallmark of amyotrophic lateral sclerosis (ALS) and several subtypes of frontotemporal lobar degeneration (FTLD). FUS-positive inclusions in FTLD and ALS patients are consistently co-labeled with stress granule (SG) marker proteins. Whether TDP-43 inclusions contain SG markers is currently still debated. We determined the requirements for SG recruitment of FUS and TDP-43 and found that cytoplasmic mislocalization is a common prerequisite for SG recruitment of FUS and TDP-43. For FUS, the arginine-glycine-glycine zinc finger domain, which is the protein's main RNA binding domain, is most important for SG recruitment, whereas the glycine-rich domain and RNA recognition motif (RRM) domain have a minor contribution and the glutamine-rich domain is dispensable. For TDP-43, both the RRM1 and the C-terminal glycine-rich domain are required for SG localization. ALS-associated point mutations located in the glycine-rich domain of TDP-43 do not affect SG recruitment. Interestingly, a 25-kDa C-terminal fragment of TDP-43, which is enriched in FTLD/ALS cortical inclusions but not spinal cord inclusions, fails to be recruited into SG. Consistently, inclusions in the cortex of FTLD patients, which are enriched for C-terminal fragments, are not co-labeled with the SG marker poly(A)-binding protein 1 (PABP-1), whereas inclusions in spinal cord, which contain full-length TDP-43, are frequently positive for this marker protein.