Metabolism of guarana (Paullinia cupana Kunth var. sorbilis) plants and fruit production subjected to glyphosate doses.

Guarana (Paullinia cupana Kunth var. sorbilis) is a typically Amazonian plant of high economic value due to the compounds found in its seed. For guarana to reach the maximum productive potential, management practices such as weed control are necessary. The use of herbicides is a viable alternative, however, its drift may lead to adverse effects on the primary and secondary plant metabolisms and cause losses in crop production. This study evaluated the differential drift effects of glyphosate doses on the physiology of guarana plants and the production of compounds of economic interest in their seeds. Glyphosate doses (57.6, 115.2, 230.4, 460.8 g ae ha-1) were applied to adult guarana plants after the flowering period. The photosynthetic functions and metabolism effects were evaluated. Herbicide treatments led to oxidative stress due to increased lipid peroxidation and increased carbohydrate and amino acid in their leaflets. Despite this, glyphosate showed no effect on fruit production or the content of secondary metabolites of commercial interest in seeds.

Investigation of the implementation of a Communication Enhanced Environment model on an acute/slow stream rehabilitation and a rehabilitation ward: A before-and-after pilot study.

OBJECTIVES: Develop and implement a Communication Enhanced Environment model and explore its effect on language activities for patients early after stroke. METHOD AND DESIGN: Before-and-after pilot study. SETTING: An acute/slow stream rehabilitation and a rehabilitation ward in a private hospital in Perth, Western Australia. PARTICIPANTS: Fourteen patients recruited within 21 days of stroke. Seven recruited during the before-phase (control group: patients with aphasia = 3, patients without aphasia = 4) and seven recruited in the after-phase (intervention group: patients with aphasia = 4, patients without aphasia = 3). INTERVENTION: The intervention group exposed to a Communication Enhanced Environment model had access to equipment, resources, planned social activities and trained communication partners. Both groups received usual stroke care. DATA COLLECTION: Hospital site champions monitored the availability of the intervention. Behavioural mapping completed during the first minute of each 5-minute interval over 12 hours (between 7 am and 7 pm) determined patient engagement in language activities. RESULTS: Seventy-one percent of the Communication Enhanced Environment model was available to the intervention group who engaged in higher, but not significant (95% CI), levels of language activities (600 of 816 observation time points, 73%) than the control group (551 of 835 observation time points, 66%). Unforeseen reorganisation of the acute ward occurred during the study. CONCLUSIONS: Implementation of a Communication Enhanced Environment model was feasible in this specific setting and may potentially influence patients' engagement in language activities. The unforeseen contextual challenges that occurred during the study period demonstrate the challenging nature of the hospital environment and will be useful in future research planning.

The sugarcane ShMYB78 transcription factor activates suberin biosynthesis in Nicotiana benthamiana.

KEY MESSAGE: A sugarcane MYB present in the culm induces suberin biosynthesis and is involved both with fatty acid and phenolics metabolism. Few transcription factors have been described as regulators of cell wall polymers deposition in C4 grasses. Particularly, regulation of suberin biosynthesis in this group of plants remains poorly understood. Here, we showed that the sugarcane MYB transcription factor ShMYB78 is an activator of suberin biosynthesis and deposition. ShMYB78 was identified upon screening genes whose expression was upregulated in sugarcane internodes undergoing suberization during culm development or triggered by wounding. Agrobacterium-mediated transient expression of ShMYB78 in Nicotiana benthamiana leaves induced the ectopic deposition of suberin and its aliphatic and aromatic monomers. Further, the expression of suberin-related genes was induced by ShMYB78 heterologous expression in Nicotiana benthamiana leaves. ShMYB78 was shown to be a nuclear protein based on its presence in sugarcane internode nuclear protein extracts, and protoplast transactivation assays demonstrated that ShMYB78 activates the promoters of the sugarcane suberin biosynthetic genes ß-ketoacyl-CoA synthase (ShKCS20) and caffeic acid-O-methyltransferase (ShCOMT). Our results suggest that ShMYB78 may be involved in the transcriptional regulation of suberin deposition, from fatty acid metabolism to phenylpropanoid biosynthesis, in sugarcane internodes.

Zinc toxicity in seedlings of three trees from the Fabaceae associated with arbuscular mycorrhizal fungi.

Due to diverse human activities zinc (Zn) may reach phytotoxic levels in the soil. Here, we evaluated the differential sensibility of three Brazilian tree species from the Fabaceae to increasing soil Zn concentrations and its physiological response to cope with excess Zn. A greenhouse experiment was conducted with the species: Mimosa caesalpiniaefolia, Erythrina speciosa and Schizolobium parahyba, and the addition of 0, 200, 400 and 600 mg Zn kg-1 to the soil. Plants were harvested after three months of cultivation, and growth, root symbiosis, biochemical markers and elemental composition were analyzed. Soil Zn addition reduced seedling growth, irrespective of the species, with a strong reduction in M. caesalpiniaefolia. Regarding root symbiosis, in N2-fixing species, nitrogenase activity was reduced by the highest Zn concentrations. Zn addition caused plants nutritional imbalances, mainly in roots. The content of photosynthetic pigments in leaves decreased up to 40%, suggesting that high Zn contents interfered with its biosynthesis, and altered the content of foliar polyamines and free amino acids, depending on the species and the soil Zn concentration. Zn toxicity in M. caesalpiniaefolia plants was observed at available soil Zn concentrations greater than 100 mg kg-1 (DTPA-extractable), being the most sensitive species and E. speciosa was moderately sensitive. S. parahyba was a moderately tolerant species, which seems to be related to polyamines accumulation and to mycorrhizal association. This last species has the potential for revegetation of areas with moderately high soil Zn concentration and for phytostabilization purposes. Future research evaluating the tolerance to multiple metal stress under field conditions should confirm S. parayba suitability in Zn contaminated areas of tropical regions.

Encapsulation of Variabilin in Stearic Acid Solid Lipid Nanoparticles Enhances Its Anticancer Activity in Vitro.

The use of natural products as chemotherapeutic agents is well established; however, many of these are associated with undesirable side effects, including high toxicity and instability. Furthermore, the development of drug resistant cancers makes the search for new anticancer lead compounds a priority. In this study, the extraction of an Ircinia sp. sponge resulted in the isolation of an inseparable mixture of (7E,12E,20Z)-variabilin (1) and (7E,12Z,20Z)-variabilin (2) and structural assignment was established using standard 1D and 2D NMR experiments. The cytotoxic activity of the compound against three solid tumour cell lines displayed moderate anti-cancer activity through apoptosis, together with a general lack of selectivity among the cancer cell lines studied. Structural assignment and cytotoxic evaluation of variabilin was complicated and further aggravated by its inherent instability. Variabilin was therefore incorporated into solid lipid nanoparticles (SLNs) and the stability and cytotoxic activity evaluated. Encapsulation of variabilin into SLNs led to a marked improvement in stability of the natural product coupled with enhanced cytotoxic activity, particularly against the prostate (PC-3) cancer cell line, with IC50 values of 87.74 µM vs. 8.94 µM for the variabilin alone and Var-SLN, respectively. Both variabilin and Var-SLN revealed comparable activity to Ceramide against the MCF-7 breast cancer cell line, revealing IC50 values of 34.8, 38.1 and 33.6 µM for variabilin, Var-SLN and Ceramide, respectively. These samples revealed no activity (>100 µM for all) against HT-29 (colon) cell lines and MCF-12 (normal breast) cell lines. Var-SLNs induced 47, 48 and 59% of apoptosis in HT-29, MCF-7 and PC-3 cells, respectively, while variabilin alone revealed 38, 29 and 29% apoptotic cells for HT-29, MCF-7 and PC-3 cell lines, respectively. The encapsulation of natural products into SLNs may provide a promising approach to overcome some of the issues hindering the development of new anticancer drugs from natural products.

Design, synthesis and biological evaluation of indane derived GPR40 agoPAMs.

GPR40 (FFAR1 or FFA1) is a G protein-coupled receptor, primarily expressed in pancreatic islet ß-cells and intestinal enteroendocrine cells. When activated by fatty acids, GPR40 elicits increased insulin secretion from islet ß-cells only in the presence of elevated glucose levels. Towards this end, studies were undertaken towards discovering a novel GPR40 Agonist whose mode of action is via Positive Allosteric Modulation of the GPR40 receptor (AgoPAM). Efforts were made to identify a suitable GPR40 AgoPAM tool molecule to investigate mechanism of action and de-risk liver toxicity of GPR40 AgoPAMs due to reactive acyl-glucuronide (AG) metabolites.

Biodegradable core-shell electrospun nanofibers containing bevacizumab to treat age-related macular degeneration.

Age-related macular degeneration (AMD) is a degenerative ocular disease that affects the central retina. It is considered the main cause of blindness and loss of vision worldwide. Angiogenic factors are associated with AMD, which has led to the use of antiangiogenic drugs, such as bevacizumab, to treat the disease using frequent intravitreal injections. In the present study, biodegradable core shell nanofibers containing bevacizumab were prepared by the coaxial electrospinning technique. It is thought that the shell could control the release of the drug, while the core would protect and store the drug. Poly(caprolactone) (PCL) and gelatin were used to form the shell of the nanofibers, while poly(vinyl alcohol) (PVA) and bevacizumab comprised the core. The nanofibers were characterized using microscopy techniques, thermal analysis, and FTIR. The results showed that core-shell nanofibers were produced as designed. Bevacizumab activity was evaluated using a chicken embryo chorioallantoic membrane (CAM) assay. An enzyme-linked immunosorbent assay was used to quantify the amount of the drug released from the different nanofibers in vitro. The toxicity of the nanofibers was evaluated in human retinal pigment epithelial (ARPE) cells. The CAM results demonstrated that bevacizumab maintained its antiangiogenic activity when incorporated into the nanofibers. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) tests revealed that the nanofibers showed no cellular toxicity, even in the presence of bevacizumab. The core-shell structure of the nanofibers reduced the release rate of bevacizumab compared with PVA nanofibers. The bevacizumab-loaded biodegradable nanofibers presented interesting properties that would potentially constitute an alternative therapy to intravitreal injections to treat AMD.

Electrospun poly(ε-caprolactone) matrices containing silver sulfadiazine complexed with ß-cyclodextrin as a new pharmaceutical dosage form to wound healing: preliminary physicochemical and biological evaluation.

Cooperation between researchers in the areas of medical, pharmaceutical and materials science has facilitated the development of pharmaceutical dosage forms that elicit therapeutic effects and protective action with a single product. In addition to optimizing pharmacologic action, such dosage forms provide greater patient comfort and increase success and treatment compliance. In the present work, we prepared semipermeable bioactive electrospun fibers for use as wound dressings containing silver sulfadiazine complexed with ß-cyclodextrin in a poly(Ɛ-caprolactone) nanofiber matrix aiming to reduce the direct contact between silver and skin and to modulate the drug release. Wound dressings were prepared by electrospinning, and were subjected to ATR-FT-IR and TG/DTG assays to evaluate drug stability. The hydrophilicity of the fibrous nanostructure in water and PBS buffer was studied by goniometry. Electrospun fibers permeability and swelling capacity were assessed, and a dissolution test was performed. In vitro biological tests were realized to investigate the biological compatibility and antimicrobial activity. We obtained flexible matrices that were each approximately 1.0 g in weight. The electrospun fibers were shown to be semipermeable, with water vapor transmission and swelling indexes compatible with the proposed objective. The hydrophilicity was moderate. Matrices containing pure drug modulated drug release adequately during 24 h but presented a high hemolytic index. Complexation promoted a decrease in the hemolytic index and in the drug release but did not negatively impact antimicrobial activity. The drug was released predominantly by diffusion. These results indicate that electrospun PCL matrices containing ß-cyclodextrin/silver sulfadiazine inclusion complexes are a promising pharmaceutical dosage form for wound healing.

GPR40 reduces food intake and body weight through GLP-1.

G protein-coupled receptor 40 (GPR40) partial agonists lower glucose through the potentiation of glucose-stimulated insulin secretion, which is believed to provide significant glucose lowering without the weight gain or hypoglycemic risk associated with exogenous insulin or glucose-independent insulin secretagogues. The class of small-molecule GPR40 modulators, known as AgoPAMs (agonist also capable of acting as positive allosteric modulators), differentiate from partial agonists, binding to a distinct site and functioning as full agonists to stimulate the secretion of both insulin and glucagon-like peptide-1 (GLP-1). Here we show that GPR40 AgoPAMs significantly increase active GLP-1 levels and reduce acute and chronic food intake and body weight in diet-induced obese (DIO) mice. These effects of AgoPAM treatment on food intake are novel and required both GPR40 and GLP-1 receptor signaling pathways, as demonstrated in GPR40 and GLP-1 receptor-null mice. Furthermore, weight loss associated with GPR40 AgoPAMs was accompanied by a significant reduction in gastric motility in these DIO mice. Chronic treatment with a GPR40 AgoPAM, in combination with a dipeptidyl peptidase IV inhibitor, synergistically decreased food intake and body weight in the mouse. The effect of GPR40 AgoPAMs on GLP-1 secretion was recapitulated in lean, healthy rhesus macaque demonstrating that the putative mechanism mediating weight loss translates to higher species. Together, our data indicate effects of AgoPAMs that go beyond glucose lowering previously observed with GPR40 partial agonist treatment with additional potential for weight loss.

Flooding of the root system in soybean: biochemical and molecular aspects of N metabolism in the nodule during stress and recovery.

Nitrogen fixation of the nodule of soybean is highly sensitive to oxygen deficiency such as provoked by waterlogging of the root system. This study aimed to evaluate the effects of flooding on N metabolism in nodules of soybean. Flooding resulted in a marked decrease of asparagine (the most abundant amino acid) and a concomitant accumulation of γ-aminobutyric acid (GABA). Flooding also resulted in a strong reduction of the incorporation of (15)N2 in amino acids. Nodule amino acids labelled before flooding rapidly lost (15)N during flooding, except for GABA, which initially increased and declined slowly thereafter. Both nitrogenase activity and the expression of nifH and nifD genes were strongly decreased on flooding. Expression of the asparagine synthetase genes SAS1 and SAS2 was reduced, especially the former. Expression of genes encoding the enzyme glutamic acid decarboxylase (GAD1, GAD4, GAD5) was also strongly suppressed except for GAD2 which increased. Almost all changes observed during flooding were reversible after draining. Possible changes in asparagine and GABA metabolism that may explain the marked fluctuations of these amino acids during flooding are discussed. It is suggested that the accumulation of GABA has a storage role during flooding stress.

Fluorescence Behaviour and Singlet Oxygen Production of Aluminium Phthalocyanine in the Presence of Upconversion Nanoparticles.

NaYF4:Yb/Er/Gd upconversion nanoparticles (UCNP) were synthesised and the photoemission stabilised by embedding them in electrospun fibers. The photophysical behaviour of chloro aluminium tetrasulfo phthalocyanine (ClAlTSPc) was studied in the presence of UCNPs when the two are mixed in solution. The fluorescence quantum yield value of ClAlTSPc decreased in the presence of UCNPs due to the heavy atom effect of UCNPs. This effect also resulted in increase in triplet quantum yields for ClAlTSPc in the presence of UCNPs. The fluorescence lifetimes for UCNPs were shortened at 658 nm in the presence of ClAlTSPc when the former was embedded in fiber and suspended in a dimethyl sulfoxide solution of the latter. A clear singlet oxygen generation by ClAlTSPc though Förster resonance energy transfer was demonstrated using a singlet oxygen quencher, 1,3-diphenylisobenzofuran.

[Incidence and management of chemotherapy-induced nausea and vomiting in women with breast cancer]. / Ocorrência e manejo de náusea e vômito no tratamento quimioterápico em mulheres com câncer de mama.

The objective of this study was to analyze the incidence of chemotherapy-induced nausea and vomiting in women with breast cancer and identify strategies used by them to control these signs and symptoms. Data for this cross-sectional study were collected through interviews during the last cycle of chemotherapy, between August 2011 and March 2012, in a university hospital in the State of São Paulo. The sample consisted of 22 women between the ages of 31 and 70, of whom 77.3% reported nausea and 50% vomiting during treatment. Regarding symptom management, 82% of the women reported having received some information centered on the use of prescribed medication. However, 27.3% did not know what medication they had taken. We concluded that there is a lack of systematic care and institutional protocol to guide professionals in providing standardized information to women so they can better control nausea and vomiting.

Attention Deficits in Healthcare Workers with Non-Clinical Burnout: An Exploratory Investigation.

Burnout syndrome is characterized by exhaustion, cynicism, and reduced effectiveness. Workers with high burnout scores who continue their professional activities are identified as experiencing non-clinical burnout (NCB), which includes early stages where burnout symptoms (BNS) are present but not yet severe enough to necessitate work leave. This study aimed to investigate the impact of BNS on attention performance among healthcare workers (HCWs) at a COVID-19 reference hospital during the pandemic. The Maslach Burnout Inventory (MBI) was applied to assess the three burnout dimensions. The Continuous Visual Attention Test (CVAT) evaluated four different attention subdomains. Participants were divided into two groups based on their scores on the MBI: controls and NCB. Thirteen controls were matched with 13 NCB subjects based on age, sex, and HCW category. This sample (n = 26, 65% male) consisted of 11 physicians and 15 nursing professionals with a mean age of 35.3 years (standard deviation = 5.47). NCB subjects had higher impulsivity than controls. There were not any significant group differences in the other attention subdomains. We found significant correlations between impulsivity and all burnout dimensions: higher absolute scores in BNS are associated with higher impulsivity. We concluded that NCB leads to executive attention deficits.

In silico approaches for predicting natural compounds with therapeutic potential and vaccine candidates against Streptococcus equi.

Equine strangles is a prevalent disease that affects the upper respiratory in horses and is caused by the Gram-positive bacterium Streptococcus equi. In addition to strangles, other clinical conditions are caused by the two S. equi subspecies, equi and zooepidemicus, which present relevant zoonotic potential. Treatment of infections caused by S. equi has become challenging due to the worldwide spreading of infected horses and the unavailability of effective therapeutics and vaccines. Penicillin treatment is often recommended, but multidrug resistance issues arised. We explored the whole genome sequence of 18 S. equi isolates to identify candidate proteins to be targeted by natural drug-like compounds or explored as immunogens. We considered only proteins shared among the sequenced strains of subspecies equi and zooepidemicus, absent in the equine host and predicted to be essential and involved in virulence. Of these, 4 proteins with cytoplasmic subcellular location were selected for molecular docking with a library of 5008 compounds, while 6 proteins were proposed as prominent immunogens against S. equi due to their probabilities of behaving as adhesins. The molecular docking analyses revealed the best ten ligands for each of the 4 drug target candidates, and they were ranked according to their binding affinities and the number of hydrogen bonds for complex stability. Finally, the natural 5-ring compound C25H20F3N5O3 excelled in molecular dynamics simulations for the increased stability in the interaction with UDP-N-acetylenolpyruvoylglucosamine reductase (MurB). This research paves the way to developing new therapeutics to minimize the impacts caused by S. equi infections.Communicated by Ramaswamy H. Sarma.

Nanophytomedicines as Therapeutic Agents for Parkinson's Disease.

Phytochemicals are promising therapeutics for various neurodegenerative disorders, including Parkinson's disease (PD). However, their efficacy, pharmacokinetic properties, and penetration across the blood-brain barrier can be improved using delivery systems such as nanoparticles. We reviewed recently published work in which nanoparticles were used to deliver phytochemicals toward PD treatment. The studies show that nanoparticles not only improve the pharmacological effect of the phytochemicals but also enable targeting to the brain and crossing of the blood-brain barrier. Various ligands were added to the nanoparticles to improve blood-brain barrier transportation. The promising findings from the published studies reveal that more research into nanophytomedicine approaches as therapeutic targets for PD is warranted, especially since they have the potential to protect against key features of PD, including α-synuclein aggregation, mitochondrial dysfunction, and dopaminergic neuronal death. Furthermore, future directions should involve smart designs to tailor nanoparticles for improved therapeutic delivery by modifying their features, such as architecture, surface and material properties, targeting ligands, and responsiveness.

Semi-field evaluation of the space spray efficacy of Fludora Co-Max EW against wild insecticide-resistant Aedes aegypti and Culex quinquefasciatus mosquito populations from Abidjan, Côte d'Ivoire.

BACKGROUND: Space spraying of insecticides is still an important means of controlling Aedes and Culex mosquitoes and arboviral diseases. This study evaluated the space spray efficacy of Fludora Co-Max EW, (water-based insecticide space spray combining flupyradifurone and transfluthrin with film forming aqueous spray technology (FFAST)), against wild insecticide-resistant Aedes aegypti and Culex quinquefasciatus mosquitoes from Abidjan, Côte d'Ivoire, compared with K-Othrine EC (deltamethrin-only product), in small-scale field trials. METHODS: Wild Ae. aegypti and Cx. quinquefasciatus mosquito larvae were collected in Abidjan, Côte d'Ivoire from August to December 2020. Mosquito larvae were reared in the laboratory until the adult stage. Fludora Co-Max EW and K-Othrine EC were tested against emerged adult females (F0 generation) using ultra-low volume cold fogging (ULV) and thermal fogging (TF) delivery technology, both outdoors and indoors in Agboville, Côte d'Ivoire. Specifically, cages containing 20 mosquitoes each were placed at distances of 10, 25, 50, 75 and 100 m from the spraying line for outdoor spraying, and at ceiling, mid-height and floor levels for indoor house spraying. Knockdown and mortality were recorded at each checkpoint and compared by treatments. RESULTS: Overall, Fludora Co-Max EW induced significantly higher knockdown and mortality effects in the wild insecticide-resistant Ae. aegypti and Cx. quinquefasciatus compared with K-Othrine EC. In both species, mortality rates with Fludora Co-Max EW were > 80% (up to 100%) with the ULV spray outdoors at each distance checkpoint (i.e. 10-100 m), and 100% with the ULV and TF sprays indoors at all checkpoints (i.e. ceiling, mid-height and floor). K-Othrine EC induced high mortality indoors (97.9-100%), whereas mortality outdoors rapidly declined in Ae. aegypti from 96.7% (10 m) to 36.7% (100 m) with the ULV spray, and from 85.0% (10 m) to 38.3% (100 m) with the TF spray. Fludora Co-Max EW spray applied as ULV spray outdoors had higher knockdown and higher killing effects on Ae. aegypti and Cx. quinquefasciatus than when applied as TF spray. Fludora Co-Max EW performed better against Cx. quinquefasciatus than against Ae. aegypti. CONCLUSIONS: Fludora Co-Max EW induced high mortality and knockdown effects against wild insecticide-resistant Ae. aegypti and Cx. quinquefasciatus Abidjan strains and performed better than K-Othrine EC. The presence of flupyradifurone and transfluthrin (with new and independent modes of action) and FFAST technology in the current Fludora Co-Max EW formulation appears to have broadened its killing capacity. Fludora Co-Max EW is thus an effective adulticide and may be a useful tool for Aedes and Culex mosquito and arbovirus control in endemic areas.

A non-canonical mechanism of GPCR activation.

The goal of designing safer, more effective drugs has led to tremendous interest in molecular mechanisms through which ligands can precisely manipulate signaling of G-protein-coupled receptors (GPCRs), the largest class of drug targets. Decades of research have led to the widely accepted view that all agonists-ligands that trigger GPCR activation-function by causing rearrangement of the GPCR's transmembrane helices, opening an intracellular pocket for binding of transducer proteins. Here we demonstrate that certain agonists instead trigger activation of free fatty acid receptor 1 by directly rearranging an intracellular loop that interacts with transducers. We validate the predictions of our atomic-level simulations by targeted mutagenesis; specific mutations which disrupt interactions with the intracellular loop convert these agonists into inverse agonists. Further analysis suggests that allosteric ligands could regulate signaling of many other GPCRs via a similar mechanism, offering rich possibilities for precise control of pharmaceutically important targets.

SHIP1 therapeutic target enablement: Identification and evaluation of inhibitors for the treatment of late-onset Alzheimer's disease.

INTRODUCTION: The risk of developing Alzheimer's disease is associated with genes involved in microglial function. Inositol polyphosphate-5-phosphatase (INPP5D), which encodes Src homology 2 (SH2) domain-containing inositol polyphosphate 5-phosphatase 1 (SHIP1), is a risk gene expressed in microglia. Because SHIP1 binds receptor immunoreceptor tyrosine-based inhibitory motifs (ITIMs), competes with kinases, and converts PI(3,4,5)P3 to PI(3,4)P2, it is a negative regulator of microglia function. Validated inhibitors are needed to evaluate SHIP1 as a potential therapeutic target. METHODS: We identified inhibitors and screened the enzymatic domain of SHIP1. A protein construct containing two domains was used to evaluate enzyme inhibitor potency and selectivity versus SHIP2. Inhibitors were tested against a construct containing all ordered domains of the human and mouse proteins. A cellular thermal shift assay (CETSA) provided evidence of target engagement in cells. Phospho-AKT levels provided further evidence of on-target pharmacology. A high-content imaging assay was used to study the pharmacology of SHIP1 inhibition while monitoring cell health. Physicochemical and absorption, distribution, metabolism, and excretion (ADME) properties were evaluated to select a compound suitable for in vivo studies. RESULTS: SHIP1 inhibitors displayed a remarkable array of activities and cellular pharmacology. Inhibitory potency was dependent on the protein construct used to assess enzymatic activity. Some inhibitors failed to engage the target in cells. Inhibitors that were active in the CETSA consistently destabilized the protein and reduced pAKT levels. Many SHIP1 inhibitors were cytotoxic either at high concentration due to cell stress or they potently induced cell death depending on the compound and cell type. One compound activated microglia, inducing phagocytosis at concentrations that did not result in significant cell death. A pharmacokinetic study demonstrated brain exposures in mice upon oral administration. DISCUSSION: 3-((2,4-Dichlorobenzyl)oxy)-5-(1-(piperidin-4-yl)-1H-pyrazol-4-yl) pyridine activated primary mouse microglia and demonstrated exposures in mouse brain upon oral dosing. Although this compound is our recommended chemical probe for investigating the pharmacology of SHIP1 inhibition at this time, further optimization is required for clinical studies. Highlights: Cellular thermal shift assay (CETSA) and signaling (pAKT) assays were developed to provide evidence of src homology 2 (SH2) domain-contaning inositol phosphatase 1 (SHIP1) target engagement and on-target activity in cellular assays.A phenotypic high-content imaging assay with simultaneous measures of phagocytosis, cell number, and nuclear intensity was developed to explore cellular pharmacology and monitor cell health.SHIP1 inhibitors demonstrate a wide range of activity and cellular pharmacology, and many reported inhibitors are cytotoxic.The chemical probe 3-((2,4-dichlorobenzyl)oxy)-5-(1-(piperidin-4-yl)-1H-pyrazol-4-yl) pyridine is recommended to explore SHIP1 pharmacology.

Lead tolerance and phytoremediation potential of Brazilian leguminous tree species at the seedling stage.

A greenhouse pot experiment was conducted to evaluate the potential of three Brazilian leguminous woody species, Mimosa caesalpiniaefolia, Erythrina speciosa and Schizolobium parahyba, for the revegetation of lead- (Pb-) contaminated areas. The response of seedlings to increasing Pb concentrations (0, 250, 500 and 1000 mg kg(-1)) in the soil was studied. In addition to Pb accumulation and translocation, the following parameters were assessed: chlorophyll, nitrate, ammonia, lipid peroxidation (MDA) and free amino acid content; seedling growth; and nitrogenase activity. No differences were observed in the germination of woody species seeds sown in soils with or without Pb addition. M. caesalpiniaefolia did not show visual symptoms of Pb toxicity, while the other two species demonstrated stress symptoms, including reduced shoot biomass yield, leaf area and height. Biochemical analyses of plant tissues revealed markedly different responses to increasing Pb concentrations, such as changes in foliar soluble amino acid composition in S. parahyba; changes in ammonia and nitrate content in E. speciosa, M. caesalpiniaefolia and S. parahyba; and changes in MDA content in S. parahyba. The levels of chlorophyll a and b and carotenoid were affected in the species studied. For the Nitrogen-fixing (N(2)-fixing) species E. speciosa, an increase of Pb in the soil affected nodule formation and growth, which led to reduced nitrogenase activity in seedlings. The concentration of Pb in shoots and roots increased with the Pb concentration in soil. However, most of the Pb absorbed accumulated in the roots, and only a small fraction was translocated to aboveground parts. These findings were confirmed by the low bioconcentration factor (BCF) and translocation factor (TF) values for the three species. The tolerance index (TI) values suggested that M. caesalpiniaefolia, a N(2)-fixing tree, was the species that was most tolerant to high Pb concentrations in soil, while E. speciosa and S. parahyba showed moderate tolerance. Of the three Brazilian native woody species studied, M. caesalpiniaefolia was found to have the highest Pb tolerance and phytostabilisation potential in Pb-contaminated soils.