Supramolecular Encapsulation and Bioactivity Modulation of a Halonium Ion by Cucurbit[<i>n</i>]uril (<i>n</i> = 7, 8)

This is the first time that cucurbit[7]­uril and cucurbit[8]­uril have been demonstrated to serve as synthetic receptors for a halonium guest species, diphenyleneiodonium, modulating its bioactivities and alleviating its cardiotoxicity, which further expands the onium family of guest molecules for the cucurbit­[n]­uril family and provides new insights for halonium-cucurbit­[n]­uril host–guest chemistry and its potential applications in pharmaceutical chemistry

SU5416 directly inhibits nNOS enzyme activity in a concentration-dependent manner.

<p>The inhibitory effects of SU5416 on rat cerebellum nNOS were shown in the graph. The IC₅₀ value was also indicated. Each individual point was an average from three independent experiments.</p

Table_1.DOCX

<p>In response to the microenvironment, microglia may polarize into either an M1 pro-inflammatory phenotype, exacerbating neurotoxicity, or an M2 anti-inflammatory phenotype, conferring neuroprotection. Betulinic acid (BA) is a naturally pentacyclic triterpenoid with considerable anti-inflammatory properties. Here, we aim to investigate the potential effects of BA on microglial phenotype polarization and to reveal the underlying mechanisms of action. First, we confirmed that BA promoted M2 polarization and inhibited M1 polarization in lipopolysaccharide (LPS)-stimulated BV-2 microglial cells. Then, we demonstrated that the effect of BA on microglial polarization was dependent on AMP-activated protein kinase (AMPK) activation, as evidenced by the fact that both AMPK inhibitor compound C and AMPK siRNA abolished the M2 polarization promoted by BA. Moreover, we found that calmodulin-dependent protein kinase kinase β (CaMKKβ), but not liver kinase B1, was the upstream kinase required for BA-mediated AMPK activation and microglial M2 polarization, via the use of both the CaMKKβ inhibitor STO-609 and CaMKKβ siRNA. Finally, BA enhanced AMPK phosphorylation and promoted M2 microglial polarization in the cerebral cortex of LPS-injected mice brains, which was attenuated by pre-administration of the AMPK inhibitor. This study demonstrated that BA promoted M2 polarization of microglia, thus conferring anti-neuroinflammatory effects via CaMKKβ-dependent AMPK activation.</p

SU5416 attenuates the deficit of locomotion behavior on zebrafish larval induced by MPTP.

<p>One dpf zebrafish embryos were treated with 200 µM MPTP for 2 days, and then co-incubated with 10 µM MPTP and SU5416 or VRI at the indicated concentrations for 72 hours, and zebrafish larval co-treated with MPTP and 150 µM L-dopa or 20 µM L-deprenyl were used as positive controls. After treatment, zebrafish were collected to perform locomotion behavior test using Viewpoint Zebrabox system and total distances travelled in 10 min were calculated. Data, expressed as percentage of control, were the mean ± SEM of 12 fish larvae per group from 3-time independent experiments. ^##<i>p</i><0.01 <i>versus</i> control group; **<i>p</i><0.01 <i>versus</i> MPTP group (Turkey’s test).</p

SU5416 prevents MPP⁺-induced apoptosis in a concentration-dependent manner.

<p>(A) SU5416, but not VRI, prevented MPP⁺-induced cell death in a concentration-dependent manner. CGNs were treated with SU5416, VRI, EPTU, 7-nitroindazole (7-NI), 1400 W or DMSO (vehicle control) at the indicated concentrations for 2 hours and then exposed to 35 µM MPP⁺. Cell viability was measured by MTT assay at 24 hours after MPP⁺ challenge. (B) SU5416 blocked neuronal loss induced by MPP⁺. CGNs were pre-incubated with or without 20 µM SU5416 and exposed to 35 µM MPP⁺2 hours later. At 24 hour after MPP⁺ challenge, CGNs were assayed with FDA/PI double staining. (C) SU5416 reversed the morphological alteration induced by MPP⁺. CGNs were pre-incubated with or without 20 µM SU5416 and exposed to 35 µM MPP⁺2 hours later. At 24 hour after MPP⁺ challenge, CGNs were assayed with nNOS and Hoechst double staining. (D) The number of apoptotic nuclei with condensed chromatin was counted from representative Hoechst staining photomicrographs and represented as a percentage of the total number of nuclei counted. Data, expressed as percentage of control, were the mean ± SEM of three separate experiments; *<i>p</i><0.05 and **<i>p</i><0.01 <i>versus</i> MPP⁺ group in (A) or <i>versus</i> control in (D); ^##<i>p</i><0.01 <i>versus</i> MPP⁺ group in (D) (Turkey’s test).</p

Anti-angiogenic effects of SU5416 and VRI in zebrafish.

<p>One dpf Tg<i>(fli-1:EGFP)</i> transgenic zebrafish embryos were treated with SU5416, VRI or DMSO (vehicle control) at the indicated concentrations for 2 days. After treatment, intersegmental-vessel formations were observed under fluorescence microscopy. Deficit of blood vessels was indicated by yellow asterisks.</p

Transcriptome Analysis in Venom Gland of the Predatory Giant Ant <i>Dinoponera quadriceps</i>: Insights into the Polypeptide Toxin Arsenal of Hymenopterans

<div><p>Background</p><p><i>Dinoponera quadriceps</i> is a predatory giant ant that inhabits the Neotropical region and subdues its prey (insects) with stings that deliver a toxic cocktail of molecules. Human accidents occasionally occur and cause local pain and systemic symptoms. A comprehensive study of the <i>D. quadriceps</i> venom gland transcriptome is required to advance our knowledge about the toxin repertoire of the giant ant venom and to understand the physiopathological basis of Hymenoptera envenomation.</p><p>Results</p><p>We conducted a transcriptome analysis of a cDNA library from the <i>D. quadriceps</i> venom gland with Sanger sequencing in combination with whole-transcriptome shotgun deep sequencing. From the cDNA library, a total of 420 independent clones were analyzed. Although the proportion of dinoponeratoxin isoform precursors was high, the first giant ant venom inhibitor cysteine-knot (ICK) toxin was found. The deep next generation sequencing yielded a total of 2,514,767 raw reads that were assembled into 18,546 contigs. A BLAST search of the assembled contigs against non-redundant and Swiss-Prot databases showed that 6,463 contigs corresponded to BLASTx hits and indicated an interesting diversity of transcripts related to venom gene expression. The majority of these venom-related sequences code for a major polypeptide core, which comprises venom allergens, lethal-like proteins and esterases, and a minor peptide framework composed of inter-specific structurally conserved cysteine-rich toxins. Both the cDNA library and deep sequencing yielded large proportions of contigs that showed no similarities with known sequences.</p><p>Conclusions</p><p>To our knowledge, this is the first report of the venom gland transcriptome of the New World giant ant <i>D. quadriceps</i>. The glandular venom system was dissected, and the toxin arsenal was revealed; this process brought to light novel sequences that included an ICK-folded toxins, allergen proteins, esterases (phospholipases and carboxylesterases), and lethal-like toxins. These findings contribute to the understanding of the ecology, behavior and venomics of hymenopterans.</p></div

SU5416 reduces the expression of nNOS protein elevated by MPP+ in CGNs.

<p>(A) CGNs were pre-treated with 20 µM SU5416 or DMSO (vehicle control) for 2 hours, and then treated with 35 µM MPP⁺ for various durations as indicated. The total proteins were extracted for Western blot analysis with specific iNOS, nNOS and β-actin antibodies. (B) Statistical analysis of nNOS expression in each treatment group. Data are expressed as the ratio to OD values of the corresponding controls. Data, expressed as percentage of control, were the mean ± SEM of five separate experiments; *<i>p</i><0.05 <i>versus</i> MPP⁺ group at the same time (Turkey’s test).</p

SU5416 increases the number of dopaminergic neurons in MPTP-treated zebrafish larval.

<p>One dpf zebrafish embryos were co-incubated with 200 µM MPTP and 1 µM SU5416 or 0.3% DMSO (vehicle control) for 2 days. After treatment, zebrafish were collected to perform paraffin-embedding, sectioning and immunostaining. (A) Representative picture of immunostaining of zebrafish section. (B) Statistical analysis of the number of TH-positive neurons in each treatment group (n = 12 fish/group). *<i>p</i><0.05 <i>versus</i> MPTP group (Turkey’s test).</p

SU5416 reverses the elevated intracellular NO induced by MPP+ in CGNs.

<p>CGNs were pre-incubated with EPTU, 7-NI or SU5416 at the indicated concentrations for 2 hours, and exposed to 35 µM MPP⁺. Intracellular NO level was measured using DAF-FM diacetate as a probe at 8 hour after MPP⁺ challenge. Data, expressed as percentage of control, were the mean ± SEM of three separate experiments; **<i>p</i><0.01 <i>versus</i> MPP⁺ group (ANOVA and Dunnett’s test).</p