Synthesis and evaluation of potent and selective MGL inhibitors as a glaucoma treatment

Monoacylglycerol lipase (MGL) inhibition provides a potential treatment approach to glaucoma through the regulation of ocular 2-arachidonoylglycerol (2-AG) levels and the activation of CB1 receptors. Herein, we report the discovery of new series of carbamates as highly potent and selective MGL inhibitors. The new inhibitors showed potent nanomolar inhibitory activity against recombinant human and purified rat MGL, were selective (>1000-fold) against serine hydrolases FAAH and ABHD6 and lacked any affinity for the cannabinoid receptors CB1 and CB2. Protein-based ¹H NMR experiments indicated that inhibitor 2 rapidly formed a covalent adduct with MGL with a residence time of about 6?h. This interconversion process “intrinsic reversibility” was exploited by modifications of the ligand’s size (length and bulkiness) to generate analogs with “tunable’ adduct residence time (τ). Inhibitor 2 was evaluated in a normotensive murine model for assessing intraocular pressure (IOP), which could lead to glaucoma, a major cause of blindness. Inhibitor 2 was found to decrease ocular pressure by ～4.5?mmHg in a sustained manner for at least 12?h after a single ocular application, underscoring the potential for topically-administered MGL inhibitors as a novel therapeutic target for the treatment of glaucoma.     

Macrophage-derived Wnt opposes Notch signaling to specify hepatic progenitor cell fate in chronic liver disease

During chronic injury a population of bipotent hepatic progenitor cells (HPCs) become activated to regenerate both cholangiocytes and hepatocytes. Here we show in human diseased liver and mouse models of the ductular reaction that Notch and Wnt signaling direct specification of HPCs via their interactions with activated myofibroblasts or macrophages. In particular, we found that during biliary regeneration, expression of Jagged 1 (a Notch ligand) by myofibroblasts promoted Notch signaling in HPCs and thus their biliary specification to cholangiocytes. Alternatively, during hepatocyte regeneration, macrophage engulfment of hepatocyte debris induced Wnt3a expression. This resulted in canonical Wnt signaling in nearby HPCs, thus maintaining expression of Numb (a cell fate determinant) within these cells and the promotion of their specification to hepatocytes. By these two pathways adult parenchymal regeneration during chronic liver injury is promoted.     

Control of the exotic bulb, Yellow Soldier (Lachenalia reflexa) invading a Banksia woodland, Perth, Western Australia

Summary In recent times managers have become increasingly aware of the South African bulbous species Yellow Soldier ( Lachenalia reflexa ) becoming a serious weed of bushland on the Swan Coastal Plain. In 1998, trials were implemented to investigate control options for Yellow Soldier invading the understorey of a Banksia ( Banksia attenuata ) Woodland west of Perth. Our trials showed that hand removal over two seasons left all natives intact but was very labour intensive, only reducing cover of Yellow Soldier by 44%. It also triggered germination by ephemeral weeds. Wiping the leaves of individual plants with a 10% glyphosate solution was not effective and was also highly labour intensive. Spot spraying with metsulfuron methyl at 0.2 g/15 L (5 g/ha) reduced the cover of Yellow Soldier by 65%, was easier to implement and appeared to have had insignificant effects on natives. We hope that this trial will encourage other workers in the field to undertake controlled trials to refine treatments at restoration sites.     

Inhibition of neuronal nitric oxide synthase activity promotes migration of human‐induced pluripotent stem cell‐derived neural stem cells toward cancer cells

The breakthrough in derivation of human‐induced pluripotent stem cells (hiPSCs) provides an approach that may help overcome ethical and allergenic challenges posed in numerous medical applications involving human cells, including neural stem/progenitor cells (NSCs). Considering the great potential of NSCs in targeted cancer gene therapy, we investigated in this study the tumor tropism of hiPSC‐derived NSCs and attempted to enhance the tropism by manipulation of biological activities of proteins that are involved in regulating the migration of NSCs toward cancer cells. We first demonstrated that hiPSC‐NSCs displayed tropism for both glioblastoma cells and breast cancer cells in vitro and in vivo. We then compared gene expression profiles between migratory and non‐migratory hiPSC‐NSCs toward these cancer cells and observed that the gene encoding neuronal nitric oxide synthase (nNOS) was down‐regulated in migratory hiPSC‐NSCs. Using nNOS inhibitors and nNOS siRNAs, we demonstrated that this protein is a relevant regulator in controlling migration of hiPSC‐NSCs toward cancer cells, and that inhibition of its activity or down‐regulation of its expression can sensitize poorly migratory NSCs and be used to improve their tumor tropism. These findings suggest a novel application of nNOS inhibitors in neural stem cell‐mediated cancer therapy.     

Functional consequences of climate change-induced plant species loss in a tallgrass prairie

Future climate change is likely to reduce the floristic diversity of grasslands. Yet the potential consequences of climate-induced plant species losses for the functioning of these ecosystems are poorly understood. We investigated how climate change might alter the functional composition of grasslands for Konza Prairie, a diverse tallgrass prairie in central North America. With species-specific climate envelopes, we show that a reduction in mean annual precipitation would preferentially remove species that are more abundant in the more productive lowland positions at Konza. As such, decreases in precipitation could reduce productivity not only by reducing water availability but by also removing species that inhabit the most productive areas and respond the most to climate variability. In support of this prediction, data on species abundance at Konza over 16 years show that species that are more abundant in lowlands than uplands are preferentially reduced in years with low precipitation. Climate change is likely to also preferentially remove species from particular functional groups and clades. For example, warming is forecast to preferentially remove perennials over annuals as well as Cyperaceae species. Despite these predictions, climate change is unlikely to unilaterally alter the functional composition of the tallgrass prairie flora, as many functional traits such as physiological drought tolerance and maximum photosynthetic rates showed little relationship with climate envelope parameters. In all, although climatic drying would indirectly alter grassland productivity through species loss patterns, the insurance afforded by biodiversity to ecosystem function is likely to be sustained in the face of climate change.     

AtNAP1 represents an atypical SufB protein in Arabidopsis plastids

The assembly of iron-sulfur (Fe-S) clusters involves several pathways and in prokaryotes the mobilization of the sulfur (SUF) system is paramount for Fe-S biogenesis and repair during oxidative stress. The prokaryotic SUF system consists of six proteins: SufC is an ABC/ATPase that forms a complex with SufB and SufD, SufA acts as a scaffold protein, and SufE and SufS are involved in sulfur mobilization from cysteine. Despite the importance of Fe-S proteins in higher plant plastids, little is known regarding plastidic Fe-S cluster assembly. We have recently shown that Arabidopsis harbors an evolutionary conserved plastidic SufC protein (AtNAP7) capable of hydrolyzing ATP and interacting with the SufD homolog AtNAP6. Based on this and the prokaryotic SUF system we speculated that a SufB-like protein may exist in plastids. Here we demonstrate that the Arabidopsis plastid-localized SufB homolog AtNAP1 can complement SufB deficiency in Escherichia coli during oxidative stress. Furthermore, we demonstrate that AtNAP1 can interact with AtNAP7 inside living chloroplasts suggesting the presence of a plastidic AtNAP1.AtNAP6.AtNAP7 complex and remarkable evolutionary conservation of the SUF system. However, in contrast to prokaryotic SufB proteins with no associated ATPase activity we show that AtNAP1 is an iron-stimulated ATPase and that AtNAP1 is capable of forming homodimers. Our results suggest that AtNAP1 represents an atypical plastidic SufB-like protein important for Fe-S cluster assembly and for regulating iron homeostasis in Arabidopsis.     

The assembly of individual chaplin peptides from Streptomyces coelicolor into functional amyloid fibrils

The self-association of proteins into amyloid fibrils offers an alternative to the natively folded state of many polypeptides. Although commonly associated with disease, amyloid fibrils represent the natural functional state of some proteins, such as the chaplins from the soil-dwelling bacterium Streptomyces coelicolor, which coat the aerial mycelium and spores rendering them hydrophobic. We have undertaken a biophysical characterisation of the five short chaplin peptides ChpD-H to probe the mechanism by which these peptides self-assemble in solution to form fibrils. Each of the five chaplin peptides produced synthetically or isolated from the cell wall is individually surface-active and capable of forming fibrils under a range of solution conditions in vitro. These fibrils contain a highly similar cross-β core structure and a secondary structure that resembles fibrils formed in vivo on the spore and mycelium surface. They can also restore the growth of aerial hyphae to a chaplin mutant strain. We show that cysteine residues are not required for fibril formation in vitro and propose a role for the cysteine residues conserved in four of the five short chaplin peptides.     

Collagen and mature elastic fibre organisation as a function of depth in the human cornea and limbus

A network of circumferentially oriented collagen fibrils exists in the periphery of the human cornea, and is thought to be pivotal in maintaining corneal biomechanical stability and curvature. However, it is unknown whether or not this key structural arrangement predominates throughout the entire corneal thickness or exists as a discrete feature at a particular tissue depth; or if it incorporates any elastic fibres and how, with respect to tissue depth, the circumcorneal annulus integrates with the orthogonally arranged collagen of the central cornea. To address these issues we performed a three-dimensional investigation of fibrous collagen and elastin architecture in the peripheral and central human cornea using synchrotron X-ray scattering and non-linear microscopy. This showed that the network of collagen fibrils circumscribing the human cornea is located in the posterior one-third of the tissue and is interlaced with significant numbers of mature elastic fibres which mirror the alignment of the collagen. The orthogonal arrangement of collagen in the central cornea is also mainly restricted to the posterior stromal layers. This information will aid the development of corneal biomechanical models aimed at explaining how normal corneal curvature is sustained and further predicting the outcome of surgical procedures.     

Loss of a large grazer impacts savanna grassland plant communities similarly in North America and South Africa

Large herbivore grazing is a widespread disturbance in mesic savanna grasslands which increases herbaceous plant community richness and diversity. However, humans are modifying the impacts of grazing on these ecosystems by removing grazers. A more general understanding of how grazer loss will impact these ecosystems is hampered by differences in the diversity of large herbivore assemblages among savanna grasslands, which can affect the way that grazing influences plant communities. To avoid this we used two unique enclosures each containing a single, functionally similar large herbivore species. Specifically, we studied a bison (Bos bison) enclosure at Konza Prairie Biological Station, USA and an African buffalo (Syncerus caffer) enclosure in Kruger National Park, South Africa. Within these enclosures we erected exclosures in annually burned and unburned sites to determine how grazer loss would impact herbaceous plant communities, while controlling for potential fire-grazing interactions. At both sites, removal of the only grazer decreased grass and forb richness, evenness and diversity, over time. However, in Kruger these changes only occurred with burning. At both sites, changes in plant communities were driven by increased dominance with herbivore exclusion. At Konza, this was caused by increased abundance of one grass species, Andropogon gerardii, while at Kruger, three grasses, Themeda triandra, Panicum coloratum, and Digitaria eriantha increased in abundance.