Polyoxypregnane Glycosides from Root of Marsdenia tenacissima and Inhibited Nitric Oxide Levels in LPS Stimulated RAW 264.7 Cells.

Six new polyoxypregnane glycosides, marstenacisside F1−F3 (1−3), G1−G2 (4−5) and H1 (6), as well as 3-O-ß-D-glucopyranosyl-(1→4)-6-deoxy-3-O-methyl-ß-D-allopyranosyl-(1→4)-ß-D-oleandropyranosyl-11α,12ß-di-O-benzoyl-tenacigenin B (7), were isolated from the roots of Marsdenia tenacissima. Their structures were established by an extensive interpretation of their 1D and 2D NMR and HRESIMS data. Compounds 1−7 were tenacigenin B derivatives with an oligosaccharide chain at C-3. This was the first time that compound 7 had been isolated from the title plant and its 1H and 13C NMR data were reported. Compounds 4 and 5 were the first examples of C21 steroid glycoside bearing unique ß-glucopyranosyl-(1→4)-ß-glucopyranose sugar moiety. All the isolated compounds were evaluated for anti-inflammatory activity by inhibiting nitric oxide (NO) production in the lipopolysaccharide-induced RAW 264.7 cells. The results showed that marstenacisside F1 and F2 exhibited significant NO inhibitory activity with an inhibition rate of 48.19 ± 4.14% and 70.33 ± 5.39%, respectively, at 40 µM, approximately equal to the positive control (L-NMMA, 68.03 ± 0.72%).

Lagotis brachystachya maxim attenuates chronic alcoholic liver injury combined with gouty arthritis in rats via its anti-inflammatory activity.

Lagotis brachystachya Maxim, a common herb in Tibetan medicine, is mainly used to treat pneumonia, hepatitis, yellow water disease (gouty arthritis). Since long-term heavy drinking is also a risk factor for gouty arthritis, the present study aimed to evaluate the underlying protective role and mechanism of extracts of Lagotis brachystachya (ELB) in chronic alcoholic liver injury combined with gouty arthritis. The rat chronic alcoholic liver injury combined with gouty arthritis model was established by long-term alcohol consumption and monosodium urate (MSU) injection. The therapeutical action of ELB was then evaluated by biochemical measurement, histopathological examination, ankle swelling assessment, and protein detection. According to biochemical measurements and histopathological evaluation, ELB could alleviate the symptoms of alcoholic liver injury combined with gouty arthritis. In addition, chronic alcohol consumption and MSU activated inflammatory-related signaling such as TLR4/MyD88/NF-κB, NLRP3, and JAK2/STAT3 pathways in the liver and synovial tissues, while ELB significantly inhibited the activation of the inflammatory signaling pathway. In conclusion, ELB is protective in rats with chronic alcoholic liver injury and gouty arthritis, possibly mediated by the inhibition of TLR4/MyD88/NF-κB, NLRP3, and JAK2-STAT3 signaling pathways in both the hepatic and synovial tissues.

Gypenoside XVII, an Active Ingredient from Gynostemma Pentaphyllum, Inhibits C3aR-Associated Synaptic Pruning in Stressed Mice.

Gynostemma pentaphyllum is a herbal medicine widely used in Asian countries, and its saponin extracts have been shown to possess potent anti-inflammatory effects. Gypenoside XVII, an active ingredient isolated from Gynostemma pentaphyllum, has been found to alleviate the inflammation induced by LPS in the BV2 microglia, according to our preliminary study. This study aims to evaluate whether Gypenoside XVII could attenuate depression-like symptoms in vivo and tries to demonstrate the involvement of the complement regulation in its antidepressant-like effect. The results showed that Gypenoside XVII significantly attenuated depression-like behaviors in the forced swimming test, tail suspension test and sucrose preference test. It also alleviated the acute stress-induced hyperactivity of serum corticosterone levels. Additionally, Gypenoside XVII significantly inhibited the activation of microglia and the expression of C3 in mice exposed to chronic unpredictable mild stress (CUMS). Meanwhile, the activation of C3aR/STAT3 signaling and the expression of proinflammatory cytokines was reversed by Gypenoside XVII. Moreover, CUMS induced excessive synaptic pruning by activating microglia, while Gypenoside XVII restored it in the prefrontal cortex. Our data demonstrated that Gypenoside XVII, the active ingredient of Gynostemma pentaphyllum, produced the antidepressant-like effects in mice, which was mediated by the inhibition of complement C3/C3aR/STAT3/cytokine signaling in the prefrontal cortex.

IL-1R/C3aR signaling regulates synaptic pruning in the prefrontal cortex of depression.

BACKGROUND: Major depressive disorder is characterized by not only monoamine neurotransmitters deficiencies but also persistent neuroinflammation. The complement system is an attractive therapeutic target for various inflammation-related diseases due to its early activation in inflammatory processes. RESULTS: In the present study, the dynamic alteration of complement C3 and its receptor C3aR during the occurrence of depression and the mechanism of astrocyte-microglia IL-1R/C3/C3aR on synaptic pruning were investigated. The proteomic analysis firstly showed that chronic stress caused an elevation of C3. GO analysis indicated that complement system-mediated synaptic pruning signaling was involved in depression. The dynamic observation indicated that C3/C3aR was activated in the early onset and throughout the course of depression induced by lipopolysaccharide (LPS) and chronic stress. In contrast, C3aR blockade inhibited the hyperactivation of microglial APT2/DHHC7 palmitoylation cycle, which mediated the translocation of STAT3 and the expression of proinflammatory cytokines. Meanwhile, C3aR blockade also attenuated the synaptic pruning and enhanced the synaptogenesis in the prefrontal cortex of mice. Moreover, the blockade of IL-1R/NF-κB signaling pathway reduced the release of C3 from astrocyte. CONCLUSIONS: The current study demonstrates that astrocyte-microglia IL-1R/C3/C3aR activation causes the abnormal synaptic pruning in depression, and suggests that the activation of complement C3/C3aR may be particularly helpful in predicting the onset stage of depression.

Inhibition of Microglial NLRP3 with MCC950 Attenuates Microglial Morphology and NLRP3/Caspase-1/IL-1ß Signaling In Stress-induced Mice.

Major depressive disorder is characterized by the deficiencies of monoamine neurotransmitters, neurotrophic factors and persistent neuroinflammation. Microglial activation has been associated with neuroinflammation-related mental diseases, accompanied by NLR family pyrin domain containing 3 (NLRP3) inflammasome. Here, we investigated the effect of NLRP3 inhibition by its small molecular inhibitor MCC950 on inflammatory activity and depressive-like mice induced by chronic unpredictable mild stress (CUMS), followed by the behavioral tests including sucrose preference test and forced swimming test. NLRP3/caspase-1/IL-1ß signaling and microglial morphology in the prefrontal cortex were measured. The results showed that CUMS caused a decrease in sucrose preference and an increase in immobility time, which were reversed by NLRP3 inhibitor MCC950. In addition, NLRP3 inhibition decreased the number of microglia and changed the activated state of microglia to a resting state by morphology 3D reconstruction. Moreover, NLRP3 inhibition inactivated NLRP3/caspase-1/IL-1ß signaling in the prefrontal cortex. The results from immunofluorescence demonstrated that NLRP3 and IL-1ß expression was decreased in microglia in response to MCC950 treatment. Accordingly, proinflammatory cytokines were also decreased by NLRP3 inhibition. In conclusion, this study demonstrates that microglial NLRP3 inhibition prevents stress-induced neuroinflammation in the prefrontal cortex and suggests that microglial NLRP3 could be one of the potential therapeutic targets for depression treatment.

Schwann Cells Accelerate Osteogenesis via the Mif/CD74/FOXO1 Signaling Pathway In Vitro.

Schwann cells have been found to promote osteogenesis by an unclear molecular mechanism. To better understand how Schwann cells accelerate osteogenesis, RNA-Seq and LC-MS/MS were utilized to explore the transcriptomic and metabolic response of MC3T3-E1 to Schwann cells. Osteogenic differentiation was determined by ALP staining. Lentiviruses were constructed to alter the expression of Mif (macrophage migration inhibitory factor) in Schwann cells. Western blot (WB) analysis was employed to detect the protein expression. The results of this study show that Mif is essential for Schwann cells to promote osteogenesis, and its downstream CD74/FOXO1 is also involved in the promotion of Schwann cells on osteogenesis. Further, Schwann cells regulate amino acid metabolism and lipid metabolism in preosteoblasts. These findings unveil the mechanism for Schwann cells to promote osteogenesis where Mif is a key factor.

Two new monoterpenoid indole alkaloids from Mappianthus iodoides stems.

Two previously unreported monoterpenoid indole alkaloids, 19(20)E-5-carboxymethylvallesiachotamine (1) and 19(20)Z-5-carboxymethylvallesiachotamine (2), along with five known indole alkaloids (3-7) were isolated from Mappianthus iodoides stems. Their structures were characterized by extensive spectroscopic data. Among these isolates, compounds 1 and 2 were the two new Δ19(20) geometric isomers. The anti-inflammatory activities in vitro of all isolated compounds were evaluated using NO assay.[Formula: see text].

A novel pentacyclic triterpene from the canes of Uncaria sessilifructus (Rubiaceae).

One novel pentacyclic triterpene, 24-dimethoxymethyl-3ß,6ß,19α- trihydroxy -12-en-28-oic acid (1), along with six known compounds 2-7, were isolated from the canes of Uncaria sessilifructus Roxb. Their structures were determined according to spectroscopic and spectrometric analysis. The anti-inflammatory activities of the isolated compounds (1-7) were scanned against NO production in LPS-activated RAW 264.7 macrophages by MTS assay, however no activities were observed.

Active Flavonoids From Lagotis brachystachya Attenuate Monosodium Urate-Induced Gouty Arthritis via Inhibiting TLR4/MyD88/NF-κB Pathway and NLRP3 Expression.

Lagotis brachystachya Maxim is a characteristic herb commonly used in Tibetan medicine. Tibetan medicine records it as an important medicine for the clinical treatment of "Yellow Water Disease," the symptoms of which are similar to that of arthritis. Our previous study showed that the flavonoid fraction extracted from L. brachystachya could attenuate hyperuricemia. However, the effects of the active flavonoids on gouty arthritis remain elusive, and the underlying mechanism is not understood. In the present study, the effects of the active flavonoids were evaluated in rats or Raw264.7 cells with gouty arthritis induced by monosodium urate (MSU) crystal, followed by the detection of TLR4, MyD88, pNF-κB, and NLR family pyrin domain-containing 3 (NLRP3) expression. The swelling of the ankle joint induced by MSU crystal began to be relieved 6 h post the administration with the active flavonoids. In addition, the active flavonoids not only alleviated MSU crystal-induced inflammation in synovial tissues by histopathological examination but also reduced tumor necrosis factor alpha (TNF-α) and interleukin-1 beta (IL-1ß) levels in the joint tissue fluid of MSU crystal-induced rats. Furthermore, Western blot analysis indicated that the active flavonoids reduced the production of these cytokines by inhibiting the TLR4/MyD88/NF-κB pathway and decreasing NLRP3 expression in synovial tissues of rats. More importantly, the inhibition of TLR4/MyD88/NF-κB pathway and NLRP3 expression was also confirmed in MSU-induced Raw264.7 cells. In conclusion, these results indicated that the active flavonoids from L. brachystachya could effectively attenuate gouty arthritis induced by MSU crystal through the TLR4/MyD88/NF-κB pathway and NLRP3 expression in vivo and in vitro, suggesting several potential candidates for the treatment of gouty arthritis.

Development of a strategy for a quick process for separation of volatile compounds in counter-current chromatography: purification of cinnamaldehyde from Cinnamomum cassia by high performance counter-current chromatography.

It is a challenge for many researchers to separate volatile compounds. In this study, we introduce a rapid and efficient method of separating target compound from the twigs of Cinnamomum cassia by high performance counter-current chromatography. Under the bioassay guidance, the total extract exhibited a potential activity against NO production in RAW 264.7 macrophages and the total extract was further separated by high performance counter-current chromatography. Cinnamaldehyde (1) was enriched by counter-current chromatography (CCC) with reversed-phase mode using n-hexane-ethyl acetate-methanol-water (1:1:1:1,v/v/v/v) as the solvent system. Further identification was achieved by high performance liquid chromatography (HPLC).

[Preparation of brazilein from Caesalpinia sappan by high performance countercurrent chromatography].

Brazilein is among the main chemical constituents of Caesalpinia sappan. It has diverse pharmacological activities. Modern pharmacological studies have shown that the compound has antitumor, anti-inflammatory, antibacterial, antioxidant, immunomodulatory, and other pharmacological activities. Brazilein is often used as a stain in various industries. The separation of brazilein by traditional column chromatography will not only result in contamination of the chromatographic column materials, but also lead to loss of the active ingredient. Countercurrent chromatography is an advanced liquid-liquid chromatographic separation technique. It has been widely used for natural product separation and isolation as it offers several advantages, such as low solvent consumption, a highly selective solvent system, and high recoveries. Typical countercurrent chromatography techniques include centrifugal partition chromatography (CPC), high-speed countercurrent chromatography (HSCCC), and high performance countercurrent chromatography (HPCCC). It is well known that choosing a suitable solvent system is vital in countercurrent separation. Therefore, two methods were introduced for choosing a suitable solvent system. One is the generally useful estimation of solvent systems (GUESS) method, which employs thin-layer chromatography (TLC) to identify a suitable solvent system with minimal labor for the rapid purification of target compounds, and another is the Shake-Flash method. The solvent system could be determined by observing the distribution of the sample in the upper and lower phases. Two kinds of solvent systems were screened using the TLC-GUESS and Shake-Flash methods, and tested through the analysis mode of the HPCCC instrument. The results showed that chloroform-methanol-water (4:3:2, v/v/v) was the optimal solvent system for HPCCC separation. A total of 15.2 mg of brazilein and 5.7 mg of caesappanin C were obtained from an ethyl acetate extract with high purities (95.6% and 89.0%, analyzed by HPLC) in one step using the preparation mode of HPCCC, the reversed-phase liquid chromatography mode with the apparatus rotated at 1600 r/min, a flow rate of 10 mL/min, separation temperature of 25℃, and detection wavelength of 285 nm. Their structures were determined by spectroscopic and spectrometric analyses. Brazilein stained the solid packing material in the column and was difficult to elute. The results showed that the use of HPCCC for the separation of brazilein can not only prevent the loss of target active ingredients in Caesalpinia sappan, but also shorten the separation and purification times and improve the operating efficiency. Therefore, HPCCC can be used for the separation and preparation of other pigment compounds in Caesalpinia sappan and other dye plants.

One new flavonoid from Oroxylum indicum.

One new flavonoid, 5,6,7-trimethoxyflavone-8-O-ß-D-glucopyranoside (1), along with six known compounds 2-7, was isolated from Oroxylum indicum. Their structures were determined on the basis of spectral data. The antibacterial activities of compounds 1-4 were studied. Compounds 1 and 3 showed medium antibacterial activity against Staphylococcus aureus with MIC/MBC at 32-128 µg/ml.

Limonoids from the fruits of Khaya ivorensis.

Two new limonoids, namely 14,15-didehydroruageanin A (1) and 3-O-methyl- butyrylseneganolide A (2), were isolated from the fruits of Khaya ivorensis along with six known limonoids: seneganolide A (3), 1,3-dideacetylkhivorin (4), 7-deacetylkhivorin (5), 3-deacetylkhivorin (6), 1-deacetylkhivorin (7), and 3-deacetyl-7-oxokhivorin (8). All the compounds were evaluated for their cytotoxicity against five tumor cell lines.

Flavonoids from twigs of Millettia pubinervis.

A new flavone, 3-methoxy-5-hydroxy-[2",3":7,8] furanoflavone, pubinerone (1), was isolated from the twigs of Millettia pubinervis Kurz, together with ten known flavonoids, karanjin (2), kanjone (3), 3,6-dimethoxy-[2",3":7,8] furanoflavone (4), pongaglabrone (5), pongapin (6), pongaflavone (7), 3,6-dimethoxy- 6",6"-dimethylchromene-[2",3":7,8] flavone (8), pongachromene (9), 3,6-dimethoxy-3',4'-methylenedioxy-6",6"-dimethylchromene-[2",3":7,8] flavone (10) and demethoxykanugin (11). This is the first phytochemical investigation of this plant. The structure of compound 1 was elucidated on the basis of spectroscopic data interpretation, including 1D and 2D NMR and HREIMS analysis. The cytotoxicity of 1 against five human cancer cell lines; HL-60, SMMC-7721, A-549, MCF-7 and SW480, was evaluated, but it was inactive (IC50 > 40 µM).

Two new triterpenoids from Gelsemium elegans and Aglaia odorata.

Eleganoside A (1) and odoratanone A (15), a triterpenoid trisaccharide glycoside and a nortriterpenoid, together with twelve known compounds (2-13) and a mixture of cerebrosides (14) were isolated from Gelsemium elegans and Aglaia odorata. Their structures were elucidated by extensive spectroscopic and spectrometric analysis. Eleganoside A (1) features a 3-O-alpha-L-rhamnopyranosyl (1-->4)-beta-D-glucopyranosyl (1-->4)-beta-D-glucopyranoside of a peculiar 3,16-dihydroxyl-lanosta-8,24-dien-26-oic acid triterpenoid skeleton, and odoratanone A (15) is a 29-norcycloartane-type triterpenoid bearing an unusual five-membered methyl acetal ring. Anti-acetylcholinesterase/butyrylcholinesterase (AChE/BChE) assay indicated that at 50 microM, ethyl caffeate (5) was promising as a dual inhibitor of AChE and BChE, and paeonol (3) and 24-hydroperoxy-24-vinylcholesterol (9) exhibited BChE-selective inhibition.

[Chemical constituents from stems of Dysoxylum laxiracemosum].

Twelve compounds were separated from stems of Dysoxylum laxiracemosum and their structures were identified by spectrum analysis as shoreic acid (1), cabraleahydroxylactone (2), cabralealactone (3), cinchonain (5), catechin (6), scopoletin (7), vanillic acid (8), p-hydroxybenzoic acid (9), docosanol (10), beta-sitosterol (11), daucosterol (12). Of them, compounds 1-6,8-12 were separated from this plant for the first time, and compounds 4-6 were reported from this plant genus for the first time.

Gelsemium alkaloids, immunosuppressive agents from Gelsemium elegans.

Bioassay-guided isolation of the stems of Gelsemium elegans has led to the isolation of two new Gelsemium alkaloids, 21-(2-oxopropyl)-koumine (1) and 11-methoxygelselegine (2), and two known alkaloids, koumine (3) and gelselegine (4). The structures of 1-2 were determined by spectroscopic (for both) and single-crystal X-ray diffraction (for 1) analysis. All compounds isolated were evaluated for their potential as immunosuppressive agents and the data suggested that Gelsemium alkaloids of different structural types possibly have potential as immunosuppressive agents.

A new prenylated xanthone from Garcinia xipshuanbannaensis Y.H. Li.

A new prenylated xanthone, named bannaxanthone I, has been isolated from the leaves of Garcinia xipshuanbannaensis, along with five other known compounds, bannaxanthone E, mangostinone, tovophyllin A, garcinone E, and γ-mangostin. The structure of the new compound was elucidated on the basis of high-resolution fast atom bombardment mass spectra (HRFABMS), 1D-and 2D-NMR experiments, including heteronuclear multiple bond connectivity (HMBC), heteronuclear single quantum coherence (HSQC) and ¹H-¹H-COSY (correlated spectroscopy, COSY).

[Chemical constituents from twigs of Garcinia xipshuanbannaensis].

OBJECTIVE: To study the chemical constituents of the twigs of Garcinia xipshuanbannaensis. METHOD: The compounds were isolated by column chromatography with silica gel, RP-18 and Sephadex LH-20, and their structures were elucidated by spectroscopic analysis. RESULT: Fifteen compounds were obtained and identified, which were bannaxanthone E (1), xanthochymol (2), isoxanthochymol (3), cycloxanthochymol (4), osajaxanthone (5), gentisein (6), mangostinone (7), kaempferol (8), quercetin (9), vitexin (10), 2"-O-acetylvitexin (11), 3-acetoxyoleanolic acid (12), (-)-epicatechin (13), beta-sitosterol (14) and daucosterol (15), respectively. CONCLUSION: Compounds 4-9 and 11-13 were isolated from the plant and compounds 11-13 were obtained from the genus Garcinia for the first time.

[The Y-STR polymorphisms and phylogenetic relationships of two minority populations in Liaoning province].

OBJECTIVE: To investigate the genetic polymorphisms of 11 Y-chromosomal short tandem repeats (Y-STR) loci in 484 male individuals from two minority populations, the Hui and Xibe, of Liaoning province, and to evaluate their forensic application values and genetic relationships with other 15 populations of China. METHODS: Eleven Y-STR loci in all samples were amplified with PowerPlex Y System, and the PCR products were analyzed by 310 Genetic Analyzer. Cluster analysis and neighbor-joining tree were applied to show the genetic distance among the populations. RESULTS: In Hui people, 187 haplotypes were identified, and the overall haplotype diversity value was 0.9990. The gene diversity values (GD) for each locus ranged from 0.4783(DYS437) to 0.9679(DYS385a/b); In Xibe people, 237 haplotypes were identified, and the overall haplotype diversity value was 0.9984. The GD value for each locus ranged from 0.3618(DYS391) to 0.9686(DYS385a/b). Comparing with 15 reference populations, the genetic distance between the Hui and Xibe was the nearest (0.0257), and that between the Hui and Yi was the farthest (0.1046), while the genetic distance between Xibe and Korean was also the farthest (0.0978). The NJ tree was similar to the results of clustering analysis and all the 17 populations were clustered into 3 groups. CONCLUSION: The genetic distribution of the 11 Y-STR loci in Liaoning Hui and Xibe ethnic groups showed favorable polymorphisms, therefore are suitable for forensic identification and paternity testing in the local area. The study of haplotype diversity among different populations is useful in understanding their origins, migrations and their relationships.