Naphthylisoquinoline alkaloids, a new structural template inhibitor of Nav1.7 sodium channel.

Voltage-gated sodium channel 1.7 (Nav1.7) remains one of the most promising drug targets for pain relief. In the current study, we conducted a high-throughput screening of natural products in our in-house compound library to discover novel Nav1.7 inhibitors, then characterized their pharmacological properties. We identified 25 naphthylisoquinoline alkaloids (NIQs) from Ancistrocladus tectorius to be a novel type of Nav1.7 channel inhibitors. Their stereostructures including the linkage modes of the naphthalene group at the isoquinoline core were revealed by a comprehensive analysis of HRESIMS, 1D, and 2D NMR spectra as well as ECD spectra and single-crystal X-ray diffraction analysis with Cu Kα radiation. All the NIQs showed inhibitory activities against the Nav1.7 channel stably expressed in HEK293 cells, and the naphthalene ring in the C-7 position displayed a more important role in the inhibitory activity than that in the C-5 site. Among the NIQs tested, compound 2 was the most potent with an IC50 of 0.73 ± 0.03 µM. We demonstrated that compound 2 (3 µM) caused dramatical shift of steady-state slow inactivation toward the hyperpolarizing direction (V1/2 values were changed from -39.54 ± 2.77 mV to -65.53 ± 4.39 mV, which might contribute to the inhibition of compound 2 against the Nav1.7 channel. In acutely isolated dorsal root ganglion (DRG) neurons, compound 2 (10 µM) dramatically suppressed native sodium currents and action potential firing. In the formalin-induced mouse inflammatory pain model, local intraplantar administration of compound 2 (2, 20, 200 nmol) dose-dependently attenuated the nociceptive behaviors. In summary, NIQs represent a new type of Nav1.7 channel inhibitors and may act as structural templates for the following analgesic drug development.

MNK2-eIF4E axis promotes cardiac repair in the infarcted mouse heart by activating cyclin D1.

Adult mammals have limited potential for cardiac regeneration after injury. In contrast, neonatal mouse heart, up to 7 days post birth, can completely regenerate after injury. Therefore, identifying the key factors promoting the proliferation of endogenous cardiomyocytes (CMs) is a critical step in the development of cardiac regeneration therapies. In our previous study, we predicted that mitogen-activated protein kinase (MAPK) interacting serine/threonine-protein kinase 2 (MNK2) has the potential of promoting regeneration by using phosphoproteomics and iGPS algorithm. Here, we aimed to clarify the role of MNK2 in cardiac regeneration and explore the underlying mechanism. In vitro, MNK2 overexpression promoted, and MNK2 knockdown suppressed cardiomyocyte proliferation. In vivo, inhibition of MNK2 in CMs impaired myocardial regeneration in neonatal mice. In adult myocardial infarcted mice, MNK2 overexpression in CMs in the infarct border zone activated cardiomyocyte proliferation and improved cardiac repair. In CMs, MNK2 binded to eIF4E and regulated its phosphorylation level. Knockdown of eukaryotic translation initiation factor (eIF4E) impaired the proliferation-promoting effect of MNK2 in CMs. MNK2-eIF4E axis stimulated CMs proliferation by activating cyclin D1. Our study demonstrated that MNK2 kinase played a critical role in cardiac regeneration. Over-expression of MNK2 promoted cardiomyocyte proliferation in vitro and in vivo, at least partly, by activating the eIF4E-cyclin D1 axis. This investigation identified a novel target for heart regenerative therapy.

Secondary Metabolites and Their Cytotoxic Activity of Artemisia nitrosa Weber. and Artemisia marschalliana Spreng.

As a promising source of biologically active substances, the Artemisia species from Kazakhstan have not been investigated efficiently. Considering the rich history, medicinal values, and availability of the Artemisia plants, systematic investigations of two Artemisia species growing in the East Kazakhstan region were conducted. In this study, one new germacrane-type sesquiterpene lactone (11), together with 10 known sesquiterpenes and its dimer, were characterized from A. nitrosa Weber. Additionally, one new chromene derivative (1') with another 12 known compounds, including coumarins, sesquiterpene diketones, phenyl propanoids, polyacetylenics, dihydroxycinnamic acid derivatives, fatty acids, naphthalene derivatives, flavones, and caffeic acid derivatives were isolated from A. marschalliana Spreng. All compounds were isolated and identified for the first time from these two Artemisia species. The structures of new compounds (11, 1') were established by using UV, TOFMS, LC-MS, 1D and 2D NMR spectroscopic analyses. The cytotoxicity of all isolated compounds was evaluated. As a result, all compounds did not show significant inhibition against HL-60 and A-549 cell lines. The sesquiterpenoids isolated from A. nitrosa were tested for their inhibitory activity against the LPS-induced NO release from the RAW624.7 cells, and neither of them exhibited significant activity.

Discovery and characterization of natural products as novel indoleamine 2,3-dioxygenase 1 inhibitors through high-throughput screening.

Indoleamine 2,3-dioxygenase 1 (IDO1) is emerging as a promising therapeutic target for the treatment of malignant tumors characterized by dysregulated tryptophan metabolism. However, the antitumor efficacy of existing small-molecule IDO1 inhibitors is still unsatisfactory, and the underlying mechanism remains largely undefined. To identify novel IDO1 inhibitors, an in-house natural product library of 2000 natural products was screened for inhibitory activity against recombinant human IDO1. High-throughput fluorescence-based screening identified 79 compounds with inhibitory activity > 30% at 20 µM. Nine natural products were further confirmed to inhibit IDO1 activity by > 30% using Ehrlich's reagent reaction. Compounds 2, 7, and 8 were demonstrated to inhibit IDO1 activity in a cellular context. Compounds 2 and 7 were more potent against IDO1 than TDO2 in the enzymatic assay. The kinetic studies showed that compound 2 exhibited noncompetitive inhibition, whereas compounds 7 and 8 were graphically well matched with uncompetitive inhibition. Compounds 7 and 8 were found to bind to the ferric-IDO1 enzyme. Docking stimulations showed that the naphthalene ring of compound 8 formed "T-shaped" π-π interactions with Phe-163 and that the 6-methyl-naphthalene group formed additional hydrophobic interactions with IDO1. Compound 8 was identified as a derivative of tanshinone, and preliminary SAR analysis indicated that tanshinone derivatives may be promising hits for the development of IDO1 inhibitors. This study provides new clues for the discovery of IDO1/TDO2 inhibitors with novel scaffolds.

Metabolomic profiling of rat urine after oral administration of the prescription antipyretic Hao Jia Xu Re Qing Granules by UPLC/Q-TOF-MS.

Hao Jia Xu Re Qing Granules (HJ), is an effective clinically used antipyretic based on traditional Chinese medicine. Although its antipyretic therapeutic effectiveness is obvious, its therapeutic mechanism has not been comprehensively explored yet. In this research, we first identified potential biomarkers which may be relevant for the antipyretic effect of HJ based on urine metabolomics using ultra-performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF-MS). A rat model of fever was established using the yeast-induced febrile response. Total-ion-current metabolic profiles of different groups were acquired and the data were processed by multivariate statistical analysis-partial least-squares discriminant analysis. As envisioned, the results revealed changes of urine metabolites related to the antipyretic effect. Fourteen potential biomarkers were selected from the urine samples based on the results of Student's t-test, "shrinkage t", variable importance in projection and partial least-squares discriminant analysis. N-Acetylleucine, kynurenic acid, indole-3-ethanol, nicotinuric acid, pantothenic acid and tryptophan were the most significant biomarkers found in the urine samples, and may be crucially related to the antipyretic effect of HJ. Consequently, we propose the hypothesis that the significant antipyretic effect the HJ may be related to the inhibition of tryptophan metabolism. This research thus provides strong theoretical support and further direction to explain the antipyretic mechanism of HJ, laying the foundation for future studies.

Novel Diterpenoids from the Twigs of Podocarpus nagi.

Phytochemical investigation of the twigs of Podocarpus nagi (Podocarpaceae) led to the isolation of two new abietane-type diterpenoids, named 1ß,16-dihydroxylambertic acid (1) and 3ß,16-dihydroxylambertic acid (2), along with two new ent-pimarane-type diterpenoids, named ent-2ß,15,16,18-tetrahydroxypimar-8(14)-ene (3) and ent-15-oxo-2ß,16,18-trihydroxypimar-8(14)-ene (4). Their respective structures were elucidated on the basis of spectroscopic analyses, including 1D- and 2D-NMR, IR, CD, and HR-ESI-MS. This is the first time ent-pimarane-type diterpenoids from the genus Podocarpus has been reported. All four new compounds were tested for cytotoxic activity. The MTT assay results showed that compounds 3 and 4 significantly inhibited the proliferation of human cervical cancer Hela cells, human lung cancer A549 cells, and human breast cancer MCF-7 cells at a concentration of 10 µM. Furthermore, using the lipopolysaccharide (LPS)-stimulated RAW264.7 cells, compounds 2 and 4 were found to significantly inhibit nitrogen oxide (NO) production with IC50 values of 26.5 ± 6.1 and 17.1 ± 1.5 µM, respectively.

[Research status of natural compounds combine with antifungal agents against drug-resistent Candida albicans].

To against the emergence of drug-resistent candidiasis, the studys of synergism of natural compounds combine with antifungal agents in vitro showed a continuous growth in recent years. The paper reviewed recent progresses to compare the synergetic effect by FICI method, and to conclude the synergetic mechanisms which have been confirmed as a reference for futher study.

Humulane-type sesquiterpenoids from Pilea cavaleriei subsp. crenata.

Nine new, uncommon humulane-type sesquiterpenoids (1, 2, 4, 6-11), together with two known derivatives, were isolated from extracts of the plant Pilea cavaleriei subsp. crenata. The structures of these compounds were fully elucidated by extensive analyses of spectroscopic data (MS, 1D- and 2D-NMR), use of the Mosher method, and by X-ray crystallographic analysis, in combination with chemical conversions. An ene reaction was discovered during the chemical transformations, which might provide an explanation for the wide distribution of the allylic hydroperoxide group in natural products.

Triterpenoids from the stem bark of Melia toosendan and determination of their absolute configurations at C(24).

Six new triterpenoids, meliasenins S-X (1-6, resp.), were isolated from the stem bark of Melia toosendan. Their structures were elucidated by mass spectrometry, NMR experiments, and comparison with the known compounds. Particularly, the absolute configuration at C(24) in new compounds was determined through their CD spectra of the [Pr(FOD)3 ] complex (fod=1,1,1,2,2,3,3,7,7,7-decafluoroheptane-4,6-dione) in CCl4 , as well as by using Mosher's method.

[Study on antifungal susceptibility of different extract of Dryopteris fragrans].

OBJECTIVE: To examine the antifungal effect of different extract of Dryopteris fragrans (L.) Schott. in vitro, and screen the effective fraction from those extracts. METHODS: Separated the Dryopteris fragrans extract and got four parts by refluxing extraction,and determined the contents of total phloroglucinol. Disc agar diffusion method and solid agar dilution method were used to determine inhibitory effect. Minimum inhibitory concentration (MIC) and minimal fungicidal concentration (MFC) of different parts of Dryopteris fragrans extract against four strains of common clinical dermatophytes were investigated. RESULTS: The data showed that the contents sequence of total phloroglucinol was in the following order: 95% -ethanol extract > water extract > diethyl ether extract > petroleum ether extract, and the antimicrobial activities against the four dermatophytes were as following order: 95% -ethanol extract > water extract > di-ethyl ether extract > petroleum ether extract. CONCLUSION: The contents of total phloroglucinol in 95% -ethanol extract of Dryopteris fragrans is the highest, and the antifungal activity against dermatophytes in vitro is the strongest. The effective fraction of Dryopteris fragrans is the 95%-ethanol extract.

[Effect of Huangqi gegen decoction (HGD) on TGF-beta1/Smad3 pathway in diabetic cardiomyopathy rats].

OBJECTIVE: To study the effect of HGD on diabetic cardiomyopathy and its mechanism. METHODS: The T2-DM rats model was established by combining high fat diet with STZ. The blood glucose, insulin, myocardial fibrosis and TGF-beta1/Smad3 signaling pathway were observed; TGF-beta1 and Smad3 mRNA expression were detected by RT-PCR method, protein expression detected by immunohistochemical method. RESULTS: HGD obviously reduced fasting blood glucose, insulin, improved insulin resistance, reduced myocardial hydroxyproline contents, lowered cardiac index, significantly inhibited over-expression of TGF-beta1/SMAD3 mRNA and protein in diabetic rats cardiac. CONCLUSION: HGD can obviously prevent experimental diabetic myocardial fibrosis through the regulation effect on TGFbeta1/Smad3 signaling pathway.

Antitussive indole alkaloids from Kopsia hainanensis.

Three new indole alkaloids, named kopsihainins A-C (1-3), and two known compounds, kopsinine (4) and methyl demethoxycarbonylchanofruticosinate (5), were isolated from the stems of Kopsia hainanensis. Their structures were determined using extensive spectroscopic methods. The two main constituents 4 and 5 exhibited significant antitussive activity in a citric acid induced guinea pig cough model. The antitussive effect of 4 was demonstrated to interact with the δ-opioid receptor. This is the first report of antitussive effects of aspidofractinine type and chanofruticosinate type alkaloids.

Phenol esters and other constituents from the stem barks of Stereospermum acuminatissimum.

A new ester, 2-(4'-hydroxyphenyl)ethyl dotriacontanoate (1), and a new inseparable mixture of octacosan-1,28-dioldiferulate and triacontan-1,30-dioldiferulate (2) were isolated from the stem barks of Stereospermum acuminatissimum, along with 24 known compounds including 4 triterpenoids, 11 anthraquinones, 2 lignans, 3 phenylpropanoids, 2 4-hydroxyphenethyl esters, 1 methoxyphenol, and 1 iridoid. The structures of the new metabolites were determined with the help of spectroscopic data including extensive 2D NMR spectroscopy. The known compounds were identified by comparison of their physical and spectroscopic data with those reported in the literature. The compounds were tested against Candida albicans ATCC 24433, C. albicans ATCC 90028, Candida glabrata ATCC 90030, Candida krusei ATCC 6258, and Candida parapsilosis ATCC 22019. Some of them were moderately active.

[Effect of modified zhenzhu tiaozhi capsule on lipid peroxidation and endothelial functions in atherosclerotic rabbits].

OBJECTIVE: To observe the anti-atherosclerotic effect and its possible mechanisms of Modified Zhenzhu Tiaozhi Capsule (MZTC) in rabbits. METHODS: 50 rabbits were divided into five groups, i.e., the normal group, the model group, the simvastatin group (3 mg/kg), the high dose MZTC group (1.6 g crude drug/kg), and the low dose MZTC group (1.6 g crude drug/kg), respectively, ten in each group. The atherosclerosis (AS) model was established by feeding rabbits with high fat diet. Corresponding medicines were administrated on the basis of high fat diet for twelve successive weeks. Levels of blood lipids, serum superoxide dismutase (SOD), malondialdehyde (MDA), and nitric oxide (NO) were detected. The area of the aortic tunica intima plaque was determined. The pathological changes of the aorta were observed. RESULTS: Compared with the model group, serum levels of total cholesterol (TC), triglyceride (TG), low-density lipoprotein cholesterol (LDL-C), MDA, and atherosclerotic index (AI) were significantly lowered in the MZTC groups (P<0.05, P<0.01), while serum levels of high-density lipoprotein cholesterol (HDL-C), SOD, and NO obviously increased (P<0.05, P<0.01). Serum levels of TC, TG, LDL-C, and in the MZTC groups were obviously higher than those of the simvastatin group (P<0.01), and there was insignificant difference in other indices. The ratio of the aortic tunica intima plaque percentage to the total tunica intima area was also markedly lower than that of the model group (P<0.05, P<0.01). Results under light microscope indicated the pathological changes of the aorta was obviously attenuated. CONCLUSIONS: MZTC could inhibit the formation and development of AS plaque. Its mechanism might be associated with regulating lipids metabolism, antioxidation, and improving endothelial functions.

Three new carabrane sesquiterpenoid derivatives from the whole plant of Carpesium abrotanoides L.

Dicarabrols B and C (1 and 2), two new carabrane sesquiterpenoid dimers, along with one new carabrane sesquiterpenoid (3) were isolated from the whole plant of Carpesium abrotanoides L. Their full structures were established by extensive analysis of HR-ESI-MS and NMR spectroscopic data, and time-dependent density functional theory (TDDFT) electronic circular dichroism (ECD) calculations. Dicarabrol B possesses a novel C30 skeleton featuring a methylene-tethered bridge between two sesquiterpene moieties, while dicarabrol C presents the unique linkage of a cyclopentane ring in the molecule. Dicarabrol C exhibited potent inhibitory effects on HL-60 cells with an IC50 value of 3.7 µmol·L-1.

3, 4-seco-Isopimarane and 3, 4-seco-pimarane diterpenoids from Callicarpa nudiflora.

A phytochemical investigation was carried out on the extract of a medicinal plant Callicarpa nudiflora, resulting in the characterization of five new 3, 4-seco-isopimarane (1-5) and one new 3, 4-seco-pimarane diterpenoid (6), together with four known compounds. The structures of the new compounds were fully elucidated by extensive analysis of MS, 1D and 2D NMR spectroscopic data, and time-dependent density functional theory (TDDFT) calculation of electronic circular dichroism (ECD) spectra, and DFT calculations for NMR chemical shifts and optical rotations.

Limonoids and triterpenoids from the stem bark of Melia toosendan.

Four new limonoids (1, 3, 4, 6), named meliatoosenins A-D, eight new euphane- and tirucallane-type triterpenoids (8-15), named meliasenins A-H, and 13 known compounds have been isolated from the stem bark of Melia toosendan. The structures of new compounds were established on the basis of 1D and 2D NMR experiments ((1)H-(1)H COSY, HSQC, HMBC, and ROESY).

Constituents of Trigonostemon chinensis.

Phytochemical investigation on the stem bark and wood of Trigonostemon chinensis led to the isolation of four new dinorditerpenoids, trigonostemons A-D (1, 3, 5, 6), a new phenanthrenone, trigonostemon E (7), and a new bisindole alkaloid, trigonostemon F (8). The structures were established by extensive spectroscopic methods. The absolute configurations of 1-6 were determined by X-ray crystallography, circular dichroism, quantum chemical TDDFT calculations, and chemical transformations. The relative configuration of 8 was confirmed by X-ray diffraction analysis.

Sesquiterpenoids and diterpenoids from Chloranthus anhuiensis.

A phytochemical investigation of the roots of Chloranthus anhuiensis afforded three new sesquiterpene lactones, chloraniolide A (1), (3R)-3-hydroxyatractylenolide III (2), and 8beta-hydroxy-1-oxoeudesma-3,7(11)-dien-12,8alpha-olide (3), and two new diterpenoids, (12R,13E)-15-(acetoxy)-12-hydroxylabda-8(20),13-dien-19-oic acid (4) and (12S,13E)-15-(acetoxy)-12-dihydroxylabda-8(20),13-dien-19-oic acid (5), as well as 17 known sesquiterpenoid and diterpenoid compounds. Their structures were established on the basis of 1D- and 2D-NMR, and other spectroscopic analyses.

[The effect of fufang zhenzhu tiaozhi capsule on lipid metabolism in atherosclerotic rabbits].

OBJECTIVE: To observe the effect of fufang zhenzhu tiaozhi capsule (FTZ) on lipid metabolism in atherosclerotic rabbits. METHODS: 50 rabbits were divided into five groups: normal control group, model group, simvastatin group, the high-dose group and low-dose group of FTZ. The atherosclerotic model was established by feeding the rabbits with high fat diet. The medicine interfered groups were administrated with homologous medicine on the base of high fat diet. All the rabbits were fed for 12 weeks. The blood lipids levels were determined just before the experiment and at the forth, the eighth and the twelfth weekend of the experiment. At the end of the experiment, serum apolipoprotein levels, liver lipid contents and the area of atherosclerotic plaque were determined respectively. RESULTS: Compared with model group, TC, TG, LDL-C and ApoB levels were significantly decreased in FTZ groups (P < 0.05 or P < 0.01). The level of HDL-C and the ratio of ApoA I /ApoB were raised significantly (P < 0.05). The contents of TC,TG in liver tissue and the area percentage of atherosclerotic plaque were also markedly reduced (P < 0.05 or P < 0.01) in the FTZ groups. CONCLUSION: FTZ could efficiently regulate the abnormality of lipid metabolism in experimental atherosclerotic rabbits, and this maybe one mamechanisms of its antiatherogenic effect.