Positive Lusitropic Effect of Quercetin on Isolated Ventricular Myocardia from Normal and Streptozotocin-Induced Diabetic Mice.

The lusitropic effect of quercetin was examined on isolated ventricular myocardial tissue preparations from normal and streptozotocin-induced diabetic mice. The time required for 90% relaxation of the myocardium, which was prolonged in the diabetic mice, was shortened by quercetin in both normal and diabetic myocardia. This effect of quercetin was completely inhibited by cyclopiazonic acid but not by SEA0400. These results indicated that quercetin accelerates myocardial relaxation through activation of the sarco-endoplasmic reticulum Ca2+-ATPase.

Anthraquinones, Diphenyl Ethers, and Their Derivatives from the Culture of the Marine Sponge-Associated Fungus Neosartorya spinosa KUFA 1047.

Previously unreported anthraquinone, acetylpenipurdin A (4), biphenyl ether, neospinosic acid (6), dibenzodioxepinone, and spinolactone (7) were isolated, together with (R)-6-hydroxymellein (1), penipurdin A (2), acetylquestinol (3), tenellic acid C (5), and vermixocin A (8) from the culture of a marine sponge-associated fungus Neosartorya spinosa KUFA1047. The structures of the previously unreported compounds were established based on an extensive analysis of 1D and 2D NMR spectra as well as HRMS data. The absolute configurations of the stereogenic centers of 5 and 7 were established unambiguously by comparing their calculated and experimental electronic circular dichroism (ECD) spectra. Compounds 2 and 5-8 were tested for their in vitro acetylcholinesterase and tyrosinase inhibitory activities as well as their antibacterial activity against Gram-positive and Gram-negative reference, and multidrug-resistant strains isolated from the environment. The tested compounds were also evaluated for their capacity to inhibit biofilm formation in the reference strains.

Isotschimgine alleviates nonalcoholic steatohepatitis and fibrosis via FXR agonism in mice.

Farnesoid X receptor (FXR) agonist obeticholic acid (OCA) has emerged as a potential therapy for nonalcoholic fatty liver disease (NAFLD). However, the side effects of OCA may limit its application in clinics. We identified previously that isotschimgine (ITG) is a non-steroidal FXR selective agonist and has potent therapeutic effects on NAFLD in mice. Here, we aimed to evaluate the therapeutic effects of ITG on nonalcoholic steatohepatitis (NASH) and fibrosis in mice. We used methionine and choline deficient (MCD) diet-induced NASH mice, bile duct ligation (BDL), and carbon tetrachloride (CCl4 )-treated hepatic fibrosis mice to investigate the effects of ITG on NASH, fibrosis, and cholestatic liver injury. Our results showed that ITG improved steatosis and inflammation in the liver of MCD diet-fed mice, as well as alleviated fibrosis and inflammation in the liver of CCl4 -treated mice. Furthermore, ITG attenuated serum bile acid levels, and reduced vacuolization, inflammatory infiltration, hepatic parenchymal necrosis, and collagen accumulation in the liver of BDL mice. Mechanistically, ITG increased the expression of FXR target genes. These data suggest that ITG is an FXR agonist and may be developed as a novel therapy for NASH, hepatic fibrosis, or primary biliary cholangitis.

New obovatol trimeric neolignans with NO inhibitory activity from the leaves of Magnolia officinalis var. biloba.

Six new obovatol trimeric neolignans, houpulignans A-F (1-6) were isolated from the leaves of Magnolia officinalis var. biloba. Their structures were determined on the basis of the interpretation of HRESIMS, NMR data, and electronic circular dichroism (ECD) calculations. Compounds 1 and 2 are the first examples of neolignans derived from three units of obovatol bearing a rare 1,4-benzodioxepane moiety. Compound 3 possesses a benzodihydropyran ring, meanwhile three units of obovatol in 4-6 are connected by an alkyl chain. Compounds 1-3 inhibited NO production in LPS-stimulated RAW264.7 cells with IC50 values of 8.01, 20.21, and 4.05 µM, respectively.

Identification of isotschimgine as a novel farnesoid X receptor agonist with potency for the treatment of obesity in mice.

Obesity and its associated non-alcoholic fatty liver disease (NAFLD) have become epidemic medical problems worldwide; however, the current available therapeutic options are limited. Farnesoid X receptor (FXR) has recently emerged as an attractive target for obesity treatment. Here we demonstrate that isotschimgine (ITG), a constituent in genus Ferula, as a novel FXR agonist with anti-obesity and anti-hepatic steatosis effects. The results showed that ITG activated the FXR transactivity and bound with the ligand binding dormain (LBD) of FXR with gene reporter assays and AlphaScreen assays. In high-fat diet-induced obese (DIO) mice, ITG lowered body weight and fat mass, improved insulin resistance and hepatic steatosis. Mechanistic studies showed that ITG altered the expression levels of FXR downstream genes, lipid synthesis and energy metabolism genes in the liver of mice. Our findings suggest that ITG is a novel FXR agonist and may be a potential therapeutic choice for obesity associated with NAFLD.

Point of View: Electrophysiological Endpoints Differ When Comparing the Mode of Action of Highly Successful Anti-arrhythmic Drugs in the CAVB Dog Model With TdP.

In the anaesthetized, chronic atrioventricular block (CAVB) dog, ventricular ectopic beats and Torsade de pointes arrhythmias (TdP) are believed to ensue from an abrupt prolongation of ventricular repolarization and increased temporal dispersion of repolarization, quantified as short-term variability (STV). These TdP stop spontaneously or, when supported by substantial spatial dispersion of repolarization (SDR), degenerate into ventricular fibrillation. However, most studies involving ventricular arrhythmias do not quantify SDR by means of an electrophysiological parameter. Therefore, we reviewed the effects of 4 highly effective anti-arrhythmic drugs (flunarizine, verapamil, SEA-0400, and GS-458967) on the repolarization duration and associated STV. All drugs were tested as anti-arrhythmic strategies against TdP in CAVB dogs, their high anti-arrhythmic efficacy was defined as suppressing drug-induced TdP in 100% of the experiments. This comparison demonstrates that even though the anti-arrhythmic outcome was similar for all drugs, distinct responses of repolarization duration and associated STV were observed. Moreover, the aforementioned and commonly adopted electrophysiological parameters were not always sufficient in predicting TdP susceptibility, and additional quantification of the SDR proved to be of added value in these studies. The variability in electrophysiological responses to the different anti-arrhythmic drugs and their inconsistent adequacy in reflecting TdP susceptibility, can be explained by distinct modes of interference with TdP development. As such, this overview establishes the separate involvement of temporal and spatial dispersion in ventricular arrhythmogenesis in the CAVB dog model and proposes SDR as an additional parameter to be included in future fundamental and/or pharmaceutical studies regarding TdP arrhythmogenesis.

Obovatol inhibits NLRP3, AIM2, and non-canonical inflammasome activation.

BACKGROUND: Obovatol, a biphenolic chemical originating from Magnolia obovata, has been utilized as a traditional medicine for the treatment of inflammatory diseases. Inflammasome induces maturation of inflammatory cytokines in response to intracellular danger signals, and its dysregulation induces inflammatory diseases. PURPOSE: The effect of obovatol on inflammasome activation has not been reported, although its anti-inflammatory properties have been studied. STUDY DESIGN/METHODS: Obovatol was treated to macrophages with inflammasome triggers, and secretions of interleukin (IL)-1ß, IL-18, and caspase-1 were measured as readouts of inflammasome activation. In addition, Asc pyroptosome formation, caspase-1 activity, and mitochondrial reactive oxygen species (ROS) production were analyzed in mechanical studies. Anti-inflammasome properties of obovatol were confirmed in an animal model. RESULTS: Obovatol inhibited NLRP3, AIM2, and non-canonical inflammasomes through inhibition of Asc pyroptosome formation and mitochondrial ROS generation. In addition, obovatol disrupted the priming step of inflammasome activation and inhibited transcription of inflammatory cytokines. In mice, obovatol attenuated serum IL-1ß elevation in response to monosodium urate crystals. CONCLUSION: Obovatol is suggested as an inhibitor of NLRP3, AIM2, and non-canonical inflammasomes.

Two natural molecules preferentially inhibit azole-resistant Candida albicans with MDR1 hyperactivation.

Antifungal drug resistance is a significant clinical problem, and antifungal agents that can evade resistance are urgently needed. In infective niches, resistant organisms often co-existed with sensitive ones, or a subpopulation of antibiotic-susceptible organisms may evolve into resistant ones during antibiotic treatment and eventually dominate the whole population. In this study, we established a co-culture assay in which an azole-resistant Candida albicans strain was mixed with a susceptible strain labeled with green fluorescent protein to mimic in vivo conditions and screen for antifungal drugs. Fluconazole was used as a positive control to verify the validity of this co-culture assay. Five natural molecules exhibited antifungal activity against both susceptible and resistant C. albicans. Two of these compounds, retigeric acid B (RAB) and riccardin D (RD), preferentially inhibited C. albicans strains in which the efflux pump MDR1 was activated. This selectivity was attributed to greater intracellular accumulation of the drugs in the resistant strains. Changes in sterol and lipid compositions were observed in the resistant strains compared to the susceptible strain, and might increase cell permeability to RAB and RD. In addition, RAB and RD interfered with the sterol pathway, further aggregating the decrease in ergosterol in the sterol synthesis pathway in the MDR1-activated strains. Our findings here provide an alternative for combating resistant pathogenic fungi.

Repurposing of an old drug: In vitro and in vivo efficacies of buparvaquone against Echinococcus multilocularis.

The metacestode stage of the fox tapeworm Echinococcus multilocularis causes the lethal disease alveolar echinococcosis. Current chemotherapeutic treatment options are based on benzimidazoles (albendazole and mebendazole), which are insufficient and hence alternative drugs are needed. In this study, we screened the 400 compounds of the Medicines for Malaria Venture (MMV) Pathogen Box against E. multilocularis metacestodes. For the screen, we employed the phosphoglucose isomerase (PGI) assay which assesses drug-induced damage on metacestodes, and identified ten new compounds with activity against the parasite. The anti-theilerial drug MMV689480 (buparvaquone) and MMV671636 (ELQ-400) were the most promising compounds, with an IC50 of 2.87 µM and 0.02 µM respectively against in vitro cultured E. multilocularis metacestodes. Both drugs suggested a therapeutic window based on their cytotoxicity against mammalian cells. Transmission electron microscopy revealed that treatment with buparvaquone impaired parasite mitochondria early on and additional tests showed that buparvaquone had a reduced activity under anaerobic conditions. Furthermore, we established a system to assess mitochondrial respiration in isolated E. multilocularis cells in real time using the Seahorse XFp Analyzer and demonstrated inhibition of the cytochrome bc1 complex by buparvaquone. Mice with secondary alveolar echinococcosis were treated with buparvaquone (100 mg/kg per dose, three doses per week, four weeks of treatment), but the drug failed to reduce the parasite burden in vivo. Future studies will reveal whether improved formulations of buparvaquone could increase its effectivity.

[Design, synthesis and antiproliferative activity in cancer cells of novel dihydropyrazolyl and pyrazolyl artemisinin-phenyl ethers].

To explore potent anticancer agent based on artemisinin scaffold, a series of 10-O-phenyl ethers derivatives containing dihydropyrazolyl or pyrazolyl moiety have been designed and synthesized. Their structures were determined by LC-MS and ¹H-NMR date. Inhibitory effects of the target compounds in human breast cancer MCF-7, MCF/Adr, MDA-MB-231 cells and prostate cell line PC-3 were determined by MTT assay. Those derivatives displayed good antiproliferative activity against the tested cancer cells. Particularly, target compounds exhibited significant cytotoxicity against drug-resistance cells MCF/Adr, which was worthy for further investigation.

Targeted Structure-Activity Analysis of Endochin-like Quinolones Reveals Potent Qi and Qo Site Inhibitors of Toxoplasma gondii and Plasmodium falciparum Cytochrome bc1 and Identifies ELQ-400 as a Remarkably Effective Compound against Acute Experimental Toxoplasmosis.

Cytochrome bc1 inhibitors have been broadly studied as human and veterinary medicines and agricultural fungicides. For the most part, cytochrome bc1 inhibitors compete with ubiquinol at the ubiquinol oxidation (Qo) site or with ubiquinone at the quinone reduction (Qi) site. 4(1 H)-Quinolones with 3-position substituents may inhibit either site based on quinolone ring substituents. 4(1 H)-Quinolones that inhibit the Qi site are highly effective against toxoplasmosis, malaria, and babesiosis and do not inhibit human cytochrome bc1. We tested a series of 4(1 H)-Quinolones against wild-type and drug resistant strains of Toxoplasma gondii and Plasmodium falciparum. These experiments identified very potent compounds that inhibit T. gondii proliferation at picomolar concentrations. The most potent compounds target the Qo site, and for these compounds, an alkyl side chain confers potency against T. gondii greater than that of bulkier side chains. Our experiments also show that substituents on the quinolone ring influenced selectivity between T. gondii and P. falciparum and between Qo and Qi site-mediated activity. Comparison of the parasite cytochrome b sequences identified amino acids that are associated with drug resistance in P. falciparum that exist naturally in wild-type T. gondii. These underlying differences may influence drug susceptibility. Finally, a Qo site active 4(1 H)-quinolone-3-diarylether tested in a murine model of toxoplasmosis was superior to atovaquone, resulting in survival from Type I strain T. gondii infection. These experiments identify highly effective compounds for toxoplasmosis and provide valuable insight into the structure-activity relationship of cytochrome bc1 inhibitors.

Retinoid isomerase inhibitors impair but do not block mammalian cone photoreceptor function.

Visual function in vertebrates critically depends on the continuous regeneration of visual pigments in rod and cone photoreceptors. RPE65 is a well-established retinoid isomerase in the pigment epithelium that regenerates rhodopsin during the rod visual cycle; however, its contribution to the regeneration of cone pigments remains obscure. In this study, we use potent and selective RPE65 inhibitors in rod- and cone-dominant animal models to discern the role of this enzyme in cone-mediated vision. We confirm that retinylamine and emixustat-family compounds selectively inhibit RPE65 over DES1, the putative retinoid isomerase of the intraretinal visual cycle. In vivo and ex vivo electroretinography experiments in Gnat1-/- mice demonstrate that acute administration of RPE65 inhibitors after a bleach suppresses the late, slow phase of cone dark adaptation without affecting the initial rapid portion, which reflects intraretinal visual cycle function. Acute administration of these compounds does not affect the light sensitivity of cone photoreceptors in mice during extended exposure to background light, but does slow all phases of subsequent dark recovery. We also show that cone function is only partially suppressed in cone-dominant ground squirrels and wild-type mice by multiday administration of an RPE65 inhibitor despite profound blockade of RPE65 activity. Complementary experiments in these animal models using the DES1 inhibitor fenretinide show more modest effects on cone recovery. Collectively, these studies demonstrate a role for continuous RPE65 activity in mammalian cone pigment regeneration and provide further evidence for RPE65-independent regeneration mechanisms.

Suppression of the NF­κB signaling pathway in colon cancer cells by the natural compound Riccardin D from Dumortierahirsute.

Colorectal cancer (CRC) is a major cause of mortality and morbidity. Chronic inflammation is closely associated with the development, progression and prognosis of the majority of intestinal malignancies. In recent years, targeting the nuclear factor (NF)­κB signaling pathway for CRC therapy has become an attractive strategy. Riccardin D, a novel macrocyclicbis (bibenzyl) compound, was isolated from the Chinese liverwort plant. Previous studies have suggested that Riccardin D exerted chemo­preventative effects against the intestinal malignancy formation. In the present study, cell counting kit­8, Hochest 33258 staining, mitochondria membrane permeability assay, western blotting analysis, reverse transcription­polymerase chain reaction, luciferase reporter gene assay and molecular modeling analysis were performed to detect the effect and mechanisms of Riccardin D on human colon cancer cells. The results demonstrated that Riccardin D significantly inhibited the growth of HT­29 cells. In addition, the cDNA expression of cyclooxygenase­2, and the protein expression and activity of NF­κB and tumor necrosis factor­α were downregulated; however, the protein expression of cleaved caspase­3 and ­9, and cleaved poly (adenosine diphosphate­ribose) polymerase, and the B­cell lymphoma (Bcl)­2: Bcl­2­associated X protein ratio were upregulated. Furthermore, Auto Dock analysis identified binding sites between Riccardin D and NF­κB. These results indicated that Riccardin D may inhibit cell proliferation and induce apoptosis in HT­29 cells, which may be associated with the blocking of the NF­κB signaling pathway. Thus, Riccardin D should be investigated as an NF­κB inhibitor in cancer therapy.

Sinopestalotiollides A-D, cytotoxic diphenyl ether derivatives from plant endophytic fungus Pestalotiopsis palmarum.

Four new diphenyl ether derivatives, sinopestalotiollides A-D (1-4), one new natural α-pyrone product (11), as well as twelve known compounds (5-1 7), were obtained from the ethyl acetate extract of the endophytic fungus Pestalotiopsis palmarum isolated from the leaves of medicinal plant Sinomenium acutum (Thunb.) Rehd et Wils. The structures were elucidated by HR-ESI-MS and NMR spectrometry data. Bioassay experiments revealed that compounds 1-4 and 11 exhibited strong to weak cytotoxicities against three human tumor cell lines Hela, HCT116 and A549.

Isolation and structure elucidation of a new compound from the fungus Aspergillus flavipes PJ03-11.

A new diphenyl ether 3-methylpentyl-2, 4-dichloroasterrate (2), along with a known diphenyl ether butyl 2, 4-dichloroasterrate (1) were isolated from the metabolites of a wetland fungus Aspergillus flavipes. PJ03-11. The structures of 1 and 2 were determined by extensive NMR and HR-ESI-MS experiments. Compounds 1 and 2 showed weak cytotoxic activity, but both of them showed no antimicrobial activity.

Diphenyl ethers from a marine-derived isolate of Aspergillus sp. CUGB-F046.

One new diphenyl ether, diorcinol K (1), along with three known compounds, diorcinols D (2), F (3) and I (4) were isolated from the fermentation media of a marine-derived fungus Aspergillus sp. CUGB-F046 which was isolated from a sediment sample collected from the Bohai Sea, China. Their structures were elucidated by detailed spectroscopic methods. Compounds 1, 2 and 4 displayed significant antibacterial activities against Staphylococcus aureus and methicillin-resistant S. aureus with MIC values of 3.125, 6.25 and 6.25 µg/mL, respectively.

Selenium-containing analogues of WC-9 are extremely potent inhibitors of Trypanosoma cruzi proliferation.

The obligate intracellular parasite, Trypanosoma cruzi is the etiologic agent of Chagas disease or American trypanosomiasis, which is the most prevalent parasitic disease in the Americas. The present chemotherapy to control this illness is still deficient particularly in the chronic stage of the disease. The ergosterol biosynthesis pathway has received much attention as a molecular target for the development of new drugs for Chagas disease. Especially, inhibitors of the enzymatic activity of squalene synthase were shown to be effective compounds on T. cruzi proliferation in in vitro assays. In the present study we designed, synthesized and evaluated the effect of a number of isosteric analogues of WC-9 (4-phenoxyphenoxyethyl thiocyanate), a known squalene synthase inhibitor, on T. cruzi growth in tissue culture cells. The selenium-containing derivatives turned out to be extremely potent inhibitors of T. cruzi growth. Certainly, 3-phenoxyphenoxyethyl, 4-phenoxyphenoxyethyl, 4-(3-fluorophenoxy)phenoxyethyl, 3-(3-fluorophenoxy)phenoxyethyl selenocyanates and (±)-5-phenoxy-2-(selenocyanatomethyl)-2,3-dihydrobenzofuran arose as relevant members of this family of compounds, which exhibited effective ED50 values of 0.084 µM, 0.11 µM, 0.083, µM, 0.085, and 0.075 µM, respectively. The results indicate that compounds bearing the selenocyanate moiety are at least two orders of magnitude more potent than the corresponding skeleton counterpart bearing the thiocyanate group. Surprisingly, these compounds exhibited excellent selectively index values ranging from 900 to 1800 making these molecules promising candidates as antiparasitic agents.

Acerogenin C from Acer nikoense exhibits a neuroprotective effect in mouse hippocampal HT22 cell lines through the upregulation of Nrf-2/HO-1 signaling pathways.

Oxidative stress contributes to neuronal death in the brain, and neuronal death can cause aging or neurodegenerative disease. Heme oxygenase 1 (HO-1) serves a vital role in the regulation of biological reactions, including oxidative stress associated with reactive oxygen species. In the present study, acerogenin C isolated from the Aceraceae plant Acer nikoense, which is used as a Japanese folk medicine for hepatic disorders and eye diseases. However, there have been no studies on the mechanisms underlying the antineurodegenerative biological activities of acerogenin C. In the present study, acerogenin C demonstrated neuroprotective action against glutamate­induced cell death in hippocampal HT22 cells through the upregulation of HO­1 expression. These effects were also associated with nuclear translocation of nuclear factor erythroid 2-related factor 2 (Nrf2) and the activation of phosphoinositide 3­kinase/protein kinase B. Taken together of the efficacy researches, this study determines that the Nrf2/HO­1 pathways denotes a biological mark and that acerogenin C might contribute to prevention of neurodegenerative disorders.

Src-homology protein tyrosine phosphatase-1 agonist, SC-43, reduces liver fibrosis.

This study aimed to investigate the role of src-homology protein tyrosine phosphatase-1 (SHP-1)-signal transducer and activator of transcription 3 (STAT3) pathway in liver fibrogenesis and the anti-fibrotic effect of SHP-1 agonist. The antifibrotic activity of SC-43, a sorafenib derivative with an enhanced SHP-1 activity, was evaluated in two fibrosis mouse models by carbon tetrachloride induction and bile duct ligation. Rat, human, and primary mouse hepatic stellate cells (HSCs) were used for mechanistic investigations. The results showed that SHP-1 protein primarily localized in fibrotic areas of human and mouse livers. SC-43 treatment reduced the activated HSCs and thus effectively prevented and regressed liver fibrosis in both fibrosis mouse models and improved mouse survival. In vitro studies revealed that SC-43 promoted HSC apoptosis, increased the SHP-1 activity and inhibited phospho-STAT3. The enhanced SHP-1 activity in HSCs significantly inhibited HSC proliferation, whereas SHP-1 inhibition rescued SC-43-induced HSC apoptosis. Furthermore, SC-43 interacted with the N-SH2 domain of SHP-1 to enhance the activity of SHP-1 as its antifibrotic mechanism. In conclusion, the SHP-1-STAT3 pathway is crucial in fibrogenesis. SC-43 significantly ameliorates liver fibrosis through SHP-1 upregulation. A SHP-1-targeted antifibrotic therapy may represent a druggable strategy for antifibrotic drug discovery.

Bioactive phenyl ether derivatives from the marine-derived fungus Aspergillus carneus.

A new phenyl ether derivative, 3-hydroxy-5-(3-hydroxy-5-methylphenoxy)-4-methoxybenzoic acid (1), along with two known analogues, 3,4-dihydroxy-5-(3-hydroxy-5-methylphenoxy)benzoic acid (2) and 3-hydroxy-5-(3-hydroxy-5-methylphenoxy)benzoic acid (3), were isolated from the fungus Aspergillus carneus collected from South China Sea. The structure elucidation of 1 was determined based on extensive NMR and MS spectroscopic analyses. Compound 2 showed a strong antioxidant activity with an IC50 value of 19.3 µM which was close to the positive control ascorbic acid (IC50 = 15.3 µM).