Ethanol Extract of Citrus grandis 'Tomentosa' Exerts Anticancer Effects by Targeting Skp2/p27 Pathway in Non-Small Cell Lung Cancer.

SCOPE: This study aims to investigate the anticancer properties of Citrus grandis 'Tomentosa' (CGT) in non-small cell lung cancer (NSCLC). METHODS AND RESULTS: The ethanol extract of CGT (CGTE) is prepared by using anhydrous ethanol and analyzed by ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS), revealing that the main chemical components in CGTE are flavonoids and coumarins, such as naringin, rhoifolin, apigenin, bergaptol, and osthole. CGTE at concentrations without inducing cell death significantly inhibits cell proliferation via inducing cell cycle G1 phase arrest by MTT, colony formation, and flow cytometry assays, implying that CGT has anticancer potential. CGTE markedly inhibits the activity of Skp2-SCF E3 ubiquitin ligase, decreases the protein level of Skp2, and promotes the accumulation of p27 by co-immunoprecipitation (co-IP) and in vivo ubiquitination assay; whereas Skp2 overexpression rescues the effects of CGTE in NSCLC cells. In subcutaneous LLC allograft and A549 xenograft mouse models, CGTE, without causing obvious side effects in mice, significantly inhibits lung tumor growth by targeting the Skp2/p27 signaling pathway. CONCLUSION: These findings demonstrate that CGTE efficiently inhibits NSCLC proliferation both in vitro and in vivo by targeting the Skp2/p27 signaling pathway, suggesting that CGTE may serve as a therapeutic candidate for NSCLC treatment.

Comparing genomes of Fructus Amomi-producing species reveals genetic basis of volatile terpenoid divergence.

Wurfbainia longiligularis and Wurfbainia villosa are both rich in volatile terpenoids and are 2 primary plant sources of Fructus Amomi used for curing gastrointestinal diseases. Metabolomic profiling has demonstrated that bornyl diphosphate (BPP)-related terpenoids are more abundant in the W. villosa seeds and have a wider tissue distribution in W. longiligularis. To explore the genetic mechanisms underlying the volatile terpenoid divergence, a high-quality chromosome-level genome of W. longiligularis (2.29 Gb, contig N50 of 80.39 Mb) was assembled. Functional characterization of 17 terpene synthases (WlTPSs) revealed that WlBPPS, along with WlTPS 24/26/28 with bornyl diphosphate synthase (BPPS) activity, contributes to the wider tissue distribution of BPP-related terpenoids in W. longiligularis compared to W. villosa. Furthermore, transgenic Nicotiana tabacum showed that the GCN4-motif element positively regulates seed expression of WvBPPS and thus promotes the enrichment of BPP-related terpenoids in W. villosa seeds. Systematic identification and analysis of candidate TPS in 29 monocot plants from 16 families indicated that substantial expansion of TPS-a and TPS-b subfamily genes in Zingiberaceae may have driven increased diversity and production of volatile terpenoids. Evolutionary analysis and functional identification of BPPS genes showed that BPP-related terpenoids may be distributed only in the Zingiberaceae of monocot plants. This research provides valuable genomic resources for breeding and improving Fructus Amomi with medicinal and edible value and sheds light on the evolution of terpenoid biosynthesis in Zingiberaceae.

Protective Effect of Ferulic Acid on Lipopolysaccharide-Induced BV2 Microglia Inflammation via AMPK/mTOR Signaling Pathway.

In neurodegenerative diseases, microglial activation and neuroinflammation are essential for the control and progression of neurodegenerative diseases. Mitigating microglium-induced inflammation is one strategy for hindering the progression of neurodegenerative diseases. Ferulic acid (FA) is an effective anti-inflammatory agent, but its potential role and regulation mechanism in neuroinflammatory reactions have not been fully studied. In this study, the neuroinflammation model was established by lipopolysaccharide (LPS), and the inhibitory effect of FA on neuroinflammation of BV2 microglia was studied. The results showed that FA significantly reduced the production and expression of reactive oxygen species (ROS), tumor necrosis factor-α (TNF-α), leukocyte-6 (IL-6) and interleukin-1ß (IL-1ß). We further studied the mechanism of FA's regulation of LPS-induced BV2 neuroinflammation and found that FA can significantly reduce the expression of mTOR in BV2 microglia induced by LPS, and significantly increase the expression of AMPK, indicating that FA may have an anti-inflammatory effect by activating the AMPK/mTOR signaling pathway to regulate the release of inflammatory mediators (such as NLRP3, caspase-1 p20 and IL-1ß). We further added an autophagy inhibitor (3-MA) and an AMPK inhibitor (compound C, CC) for reverse verification. The results showed that FA's inhibitory effects on TNF-α, IL-6 and IL-1ß and its regulatory effect on AMPK/mTOR were destroyed by 3-MA and CC, which further indicated that FA's inhibitory effect on neuroinflammation is related to its activation of the AMPK/mTOR autophagy signaling pathway. In a word, our experimental results show that FA can inhibit LPS-induced neuroinflammation of BV2 microglia by activating the AMPK/mTOR signaling pathway, and FA may be a potential drug for treating neuroinflammatory diseases.

The Underling Mechanisms Exploration of Rubia cordifolia L. Extract Against Rheumatoid Arthritis by Integrating Network Pharmacology and Metabolomics.

Purpose: Rubia cordifolia L. (RC) is a classic herbal medicine for the treatment of rheumatoid arthritis (RA) and has been used since ancient times. The ethanol extract of Rubia cordifolia L. (RCE) showed obvious anti-RA effects in our previous study. However, further potential mechanisms require more exploration. We aimed to investigate the mechanism of RCE for the treatment of RA by integrating metabolomics and network pharmacology in this study. Methods: An adjuvant-induced arthritis (AIA) rat model was established, and we evaluated the therapeutic effects of RCE. Metabolomics of serum and urine was used to identify the differential metabolites. Network pharmacology was applied to determine the key metabolites and potential targets. Finally, the potential targets and compounds of RCE were verified by molecular docking. Results: The results indicated that RCE suppressed foot swelling and alleviated joint damage and also had anti-inflammatory properties by inhibiting the expressions of tumor necrosis factor (TNF)-α, Interleukin (IL)-1ß, prostaglandin E2 (PGE2), and P65. Ten and seven differential metabolites were found in the serum and urine, respectively, of rats. Six key targets, ie, phospholipase A2 group IIA (PLA2G2A), phospholipase A2 group X (PLA2G10), cytidine deaminase (CDA), uridine-cytidine kinase 2 (UCK2), charcot-leyden crystal galectin (CLC), and 5',3'-nucleotidase, mitochondrial (NT5M), were discovered by network pharmacology and metabolite analysis and were found to be related to glycerophospholipid metabolism and pyrimidine metabolism. Molecular docking confirmed that the favorable compounds showed affinities with the key targets, including alizarin, 6-hydroxyrubiadin, ruberythric acid, and munjistin. Conclusion: This study revealed the underlying mechanisms of RCE and provided evidence that will allow researchers to further investigate the functions and components of RCE against RA.

The Effective Treatment of Purpurin on Inflammation and Adjuvant-Induced Arthritis.

Rubia cordifolia L. (Rubiaceae), one of the traditional anti-rheumatic herbal medicines in China, has been used to treat rheumatoid arthritis (RA) since ancient times. Purpurin, an active compound of Rubia cordifolia L., has been identified in previous studies and exerts antibacterial, antigenotoxic, anticancer, and antioxidant effects. However, the efficacy and the underlying mechanism of purpurin to alleviate RA are unclear. In this study, the effect of purpurin on inflammation was investigated using macrophage RAW264.7 inflammatory cells, induced by lipopolysaccharide (LPS), and adjuvant-induced arthritis (AIA) rat was established to explore the effect of purpurin on joint damage and immune disorders; the network pharmacology and molecular docking were integrated to dig out the prospective target. Purpurin showed significantly anti-inflammatory effect by reducing the content of IL-6, TNF-α, and IL-1ß and increasing IL-10. Besides, purpurin obviously improved joint injury and hypotoxicity in the liver and spleen and regulated the level of FOXP3 and CD4+/CD8+. Furthermore, purpurin reduced the MMP3 content of AIA rats. Network pharmacology and molecular docking also suggested that MMP3 may be the key target of purpurin against RA. The results of this study strongly indicated that purpurin has a potential effect on anti-RA.

Characterization of two novel highly active glycoside hydrolase family 53 endo-1,4-ß-galactanases and their synergism with other carbohydrases in plant polysaccharide decomposition.

Two novel Bacillus glycoside hydrolase family 53 (GH53) endo-1,4-ß-galactanases (Bs936 and Bs4828) were identified. The recombinant Bs936 and Bs4828 displayed maximal activities at pH 5.5/55 °C and pH 6.5/65 °C, respectively. Stability analyses revealed that the enzymes were stable between pH 4.5-10, retaining over 80% activities after 12 h incubation at 25 °C. Moreover, Bs936 maintained about 75% activity after being treated at 45 °C for 2 h, while Bs4828 kept full activity after 4 h of incubation at 50 °C. Importantly, Bs936 and Bs4828 exhibited good activity towards ß-1,4-galactan, showing specific activities of 1859.46- and 3110.79 U/mg towards potato galactan, respectively. The corresponding Km and kcat/Km were 4.93 mg/mL and 296.35 mL/mg/s for Bs936, and 6.60 mg/mL and 707.12 mL/mg/s for Bs4828. Extensive synergy assays revealed that each endo-1,4-ß-galactanase acted synergistically with the other ß-galactosidase (Bs937 or Bs4829), endo-1,5-α-L-arabinosidase (BsAbn2) and an endoxylanase/endoglucanase+cellulase mixture (BaXynA + LsCel) towards the hydrolysis of galactan, debranched arabinan/arabinan, and pretreated corn stover, respectively. The corresponding degrees of synergy were 1.28, 1.17/1.09, and 1.15 for Bs486 and 1.55, 1.04/1.08, and 1.11 for Bs4828, respectively. This study expands the repertoire of GH53 enzymes, and demonstrates the role of two highly active endo-1,4-ß-galactanases in polysaccharides hydrolysis.

Anti-Inflammatory Activation of Phellodendri Chinensis Cortex is Mediated by Berberine Erythrocytes Self-Assembly Targeted Delivery System.

Background: Berberine (BBR) is the primary active component of Phellodendri Chinensis Cortex (PCC), which has been traditionally used to treat inflammatory diseases. However, the discrepancy between its low bioavailability and significant therapeutic effect remains obscure. The purpose of this study was to explore the previously unsolved enigma of the low bioavailability of BBR and its appreciable anti-inflammatory effect to reveal the action mechanism of BBR and PCC. Methods: The quantitative analysis of BBR and its metabolite oxyberberine (OBB) in blood and tissues was performed using high-performance liquid chromatography to investigate the conversion and distribution of BBR/OBB mediated by erythrocytes. Routine blood tests and immunohistochemical staining were used to explore the potential relationship between the amounts of monocyte/macrophage and the drug concentration in erythrocytes and tissues (liver, heart, spleen, lung, kidney, intestine, muscle, brain and pancreas). To comparatively explore the anti-inflammatory effects of BBR and OBB, the acetic acid-induced vascular permeability mice model and lipopolysaccharide-induced RAW 264.7 macrophages were employed. Results: Nearly 92% of BBR existed in the erythrocytes in rats. The partition coefficient of BBR between plasma and erythrocytes (Kp/b) decreased with time. OBB was found to be the oxidative metabolite of BBR in erythrocytes. Proportion of BBR/OBB in erythrocytes changed from 9.38% to 16.30% and from 13.50% to 46.24%, respectively. There was a significant relationship between the BBR/OBB concentration in blood and monocyte depletion after a single administration of BBR. BBR/OBB was transported via erythrocytes to various tissues (liver, kidney, spleen, lung, and heart, etc), with the liver achieving the highest concentration. OBB exhibited similar anti-inflammatory effect in vitro and in vivo as BBR with much smaller dosage. Conclusion: BBR was prodominantly found in erythrocytes, which was critically participated in the biodistribution, pharmacokinetics, metabolism and target delivery of BBR and its metabolite. The anti-inflammatory activity of BBR and PCC was intimately associated with the metabolism into the active congener OBB and the targeted delivery to monocytes/macrophages mediated by the erythrocytes.

Comparative Analysis of the Metabolites and Biological Activity of Cultivated and Wild Lignosus rhinocerotis.

In this paper, Lignosus rhinocerotis (Cooke) Ryvarden (L. rhinocerotis) cultivated in rice medium (LRR) and in sawdust medium (LRS) was harvested. Then, in terms of the LRR, LRS, and wild L. rhinocerotis (LRW), the total flavonoid contents, total polyphenol contents, total polysaccharide contents, and metabolites were detected; antioxidants of their aqueous extracts and anti-inflammatory of their polysaccharides were performed. In addition, the possible mechanism of the polysaccharides of L. rhinocerotis inhibiting lung damage was elucidated. The results showed that 32 compounds were characterized in L. rhinocerotis, including flavonoids, terpenoids, lignans, and steroids and there were 20 compounds in cultivated and wild L. rhinocerotis; LRR has the highest total polyphenol and flavonoid contents, as well as ABTS and DPPH scavenging capacity. The total polysaccharide contents and the FRAP scavenging capacity of wild L. rhinocerotis were higher than those of cultivated L. rhinocerotis. The inhibition of polysaccharides of LRW (PLRW) on LPS-induced MRC-5 damage was stronger than that of the polysaccharides from cultivated L. rhinocerotis. The PLRW may alleviate lung damage by inhibiting the NLRP3 pathway and thereby suppressing the inflammatory response. In summary, both cultivated and wild L. rhinocerotis are abundant in bioactive components and have antioxidant and anti-inflammatory activities.

Functional analysis of Pogostemon cablin farnesyl pyrophosphate synthase gene and its binding transcription factor PcWRKY44 in regulating biosynthesis of patchouli alcohol.

Farnesyl pyrophosphate synthase (FPPS) plays an important role in the synthesis of plant secondary metabolites, but its function and molecular regulation mechanism remain unclear in Pogostemon cablin. In this study, the full-length cDNA of the FPP synthase gene from P. cablin (PcFPPS) was cloned and characterized. The expressions of PcFPPS are different among different tissues (highly in P. cablin flowers). Subcellular localization analysis in protoplasts indicated that PcFPPS was located in the cytoplasm. PcFPPS functionally complemented the lethal FPPS deletion mutation in yeast CC25. Transient overexpression of PcFPPS in P. cablin leaves accelerated terpene biosynthesis, with an ~47% increase in patchouli alcohol. Heterologous overexpression of PcFPPS in tobacco plants was achieved, and it was found that the FPP enzyme activity was significantly up-regulated in transgenic tobacco by ELISA analysis. In addition, more terpenoid metabolites, including stigmasterol, phytol, and neophytadiene were detected compared with control by GC-MS analysis. Furthermore, with dual-LUC assay and yeast one-hybrid screening, we found 220 bp promoter of PcFPPS can be bound by the nuclear-localized transcription factor PcWRKY44. Overexpression of PcWRKY44 in P. cablin upregulated the expression levels of PcFPPS and patchoulol synthase gene (PcPTS), and then promote the biosynthesis of patchouli alcohol. Taken together, these results strongly suggest the PcFPPS and its binding transcription factor PcWRKY44 play an essential role in regulating the biosynthesis of patchouli alcohol.

Rhamnosyltransferases involved in the biosynthesis of flavone rutinosides in Chrysanthemum species.

Linarin (acacetin-7-O-rutinoside), isorhoifolin (apigenin-7-O-rutinoside), and diosmin (diosmetin-7-O-rutinoside) are chemically and structurally similar flavone rutinoside (FR) compounds found in Chrysanthemum L. (Anthemideae, Asteraceae) plants. However, their biosynthetic pathways remain largely unknown. In this study, we cloned and compared FRs and genes encoding rhamnosyltransferases (RhaTs) among eight accessions of Chrysanthemum polyploids. We also biochemically characterized RhaTs of Chrysanthemum plants and Citrus (Citrus sinensis and Citrus maxima). RhaTs from these two genera are substrate-promiscuous enzymes catalyzing the rhamnosylation of flavones, flavanones, and flavonols. Substrate specificity analysis revealed that Chrysanthemum 1,6RhaTs preferred flavone glucosides (e.g. acacetin-7-O-glucoside), whereas Cs1,6RhaT preferred flavanone glucosides. The nonsynonymous substitutions of RhaTs found in some cytotypes of diploids resulted in the loss of catalytic function. Phylogenetic analysis and specialized pathways responsible for the biosynthesis of major flavonoids in Chrysanthemum and Citrus revealed that rhamnosylation activity might share a common evolutionary origin. Overexpression of RhaT in hairy roots resulted in 13-, 2-, and 5-fold increases in linarin, isorhoifolin, and diosmin contents, respectively, indicating that RhaT is mainly involved in the biosynthesis of linarin. Our findings not only suggest that the substrate promiscuity of RhaTs contributes to the diversity of FRs in Chrysanthemum species but also shed light on the evolution of flavone and flavanone rutinosides in distant taxa.

The Critical Role of 12-Methyl Group of Anthracycline Dutomycin to Its Antiproliferative Activity.

Anthracycline dutomycin is a tetracyclic quinone glycoside produced by Streptomyces minoensis NRRL B-5482. SW91 is a C-12 demethylated dutomycin derivative, which was identified in our previous research. In vitro cytotoxicity and apoptosis assays of these two compounds were conducted to demonstrate their antiproliferation activities. The results showed that both dutomycin and SW91 block cells at the S phase, whereas dutomycin shows more significant inhibition of cell growth. Their interactions with calf thymus DNA (CT-DNA) were investigated, with dutomycin exhibiting higher binding affinity. The molecular docking demonstrated that the 12-methyl group makes dutomycin attach to the groove of DNA. These findings suggest that dutomycin has binding higher affinity to DNA and impairs DNA replication resulting in more significant antitumor activity.

Brusatol has therapeutic efficacy in non-small cell lung cancer by targeting Skp1 to inhibit cancer growth and metastasis.

Skp1-Cul1-F-box protein (SCF) ubiquitin E3 ligases play important roles in cancer development and serve as a promising therapeutic target in cancer therapy. Brusatol (Bru), a known Nrf2 inhibitor, holds promise for treating a wide range of tumors; however, the direct targets of Bru and its anticancer mode of action remain unclear. In our study, 793 Bru-binding candidate proteins were identified by using a biotin-brusatol conjugate (Bio-Bru) followed by streptavidin-affinity pull down-based mass spectrometry. We found that Bru can directly bind to Skp1 and disrupt the interactions of Skp1 with the F-box protein Skp2, leading to the inhibition of the Skp2-SCF E3 ligase. Bru inhibited both proliferation and migration via promoting the accumulation of the substrates p27 and E-cadherin; Skp1 overexpression attenuated while Skp1 knockdown enhanced these effects of Bru in non-small cell lung cancer (NSCLC) cells. Moreover, Bru binding to Skp1 also inhibited the ß-TRCP-SCF E3 ligase. In both subcutaneous and orthotopic NSCLC xenografts, Bru significantly inhibited the growth and metastasis of NSCLC through targeting SCF complex and upregulating p27 and E-cadherin protein levels. These data demonstrate that Bru is a Skp1-targeting agent that may have therapeutic potentials in lung cancer.

Gentiopicroside Produces Endothelium-Independent Vasodilation by Deactivating the PI3K/Akt/Rho-Kinase Pathway in Isolated Rat Thoracic Aorta.

Gentiopicroside (GPS), a main active secoiridoid glucoside derived from the roots of perennial herbs in the Gentianaceae family, has antispasmodic and relaxant effects. However, the vasorelaxant effects of GPS on aortic rings and the molecular mechanisms involved in these effects are not yet clear. Therefore, we investigated whether GPS inhibits phenylephrine- (PE-) or KCl-induced contractions in isolated rat thoracic aortic rings. The present study found that GPS produced a dose-dependent relaxation in aortic rings precontracted with PE or KCl and significantly reduced CaCl2-, narciclasine- (Rho-kinase activator-), and phorbol-12,13-diacetate- (PKC activator-) induced vasocontractions. Pretreatment with NG-Nitroarginine methyl ester hydrochloride (L-NAME, NOS inhibitor), methylene blue (sGC inhibitor), indomethacin (COX inhibitor), 4-aminopyridine (KV channel inhibitor), and glibenclamide (KATP channel inhibitor) had no influence on the vasorelaxant effect of GPS, while BaCl2 (Kir channel inhibitor), tetraethylammonium chloride (KCa channel inhibitor), ruthenium red (RYR inhibitor), and heparin (IP3R inhibitor) significantly reduced GPS-induced vasorelaxation. Moreover, GPS pretreatment remarkably inhibited the influx of Ca2+ in vascular smooth muscle cells stimulated using KCl or PE-containing CaCl2 solution. Western blot analysis confirmed that GPS treatment inhibited PE-induced increases in the protein levels of p-Akt, p-myosin light chain (MLC), and p-myosin-binding subunit of myosin phosphatase 1 (MYPT1) in the aortic rings. Additionally, the vasorelaxation activity of GPS was attenuated upon pretreatment with LY294002 (PI3K/Akt inhibitor), Y27632 (Rho-kinase inhibitor), and verapamil (L-type Ca2+ channel inhibitor). These findings demonstrate that GPS exhibits endothelium-independent vasorelaxant effects through inhibition of voltage-dependent, receptor-operated, and inositol triphosphate receptor (IP3R)/ryanodine receptor- (RYR-) mediated Ca2+ channels as well as the PI3K/Akt/Rho-kinase signaling pathway.

Synthesis, Biological Evaluation and Low-Toxic Formulation Development of Glycosylated Paclitaxel Prodrugs.

Paclitaxel (PTX) is a famous anti-cancer drug with poor aqueous solubility. In clinical practices, Cremophor EL (polyethoxylated castor oil), a toxic surfactant, is used for dissolution of PTX, which accounts for serious side effects. In the present study, a single glucose-conjugated PTX prodrug (SG-PTX) and a double glucose-conjugated PTX prodrug (DG-PTX) were synthesized with a glycosylated strategy via succinate linkers. Both of the two prodrugs presented significant solubility improvement and drug-like lipophilicities. Compared to DG-PTX, SG-PTX manifested more promising release of the parent drug in serum. A high percentage of PTX released from SG-PTX could be detected after enzymatic hydrolysis of ß-glucuronidase. Besides, both of the two prodrugs exhibited effective cytotoxicity against breast cancer cells and ovarian cancer cells, but presented reduced cytotoxicity against normal breast cells. Moreover, SG-PTX manifested impressive solubility in a low toxic formulation (without ethanol) with a different percentage of Cremophor EL. These results indicated that glycosylation is a promising strategy for PTX modification and SG-PTX may be a feasible and potential type of PTX prodrug. In addition, ethanol-free formulation with a low percentage of Cremophor EL might have the potential to develop a safer formulation for further studies of glycosylated PTX prodrugs.

Qualitative and Quantitative Analysis of Volatile Components of Zhengtian Pills Using Gas Chromatography Mass Spectrometry and Ultra-High Performance Liquid Chromatography.

Zhengtian pills (ZTPs) are traditional Chinese medicine (TCM) which have been commonly used to treat headaches. Volatile components of ZTPs extracted by ethyl acetate with an ultrasonic method were analyzed by gas chromatography mass spectrometry (GC-MS). Twenty-two components were identified, accounting for 78.884% of the total components of volatile oil. The three main volatile components including protocatechuic acid, ferulic acid, and ligustilide were simultaneously determined using ultra-high performance liquid chromatography coupled with diode array detection (UHPLC-DAD). Baseline separation was achieved on an XB-C18 column with linear gradient elution of methanol-0.2% acetic acid aqueous solution. The UHPLC-DAD method provided good linearity (R (2) ≥ 0.9992), precision (RSD < 3%), accuracy (100.68-102.69%), and robustness. The UHPLC-DAD/GC-MS method was successfully utilized to analyze volatile components, protocatechuic acid, ferulic acid, and ligustilide, in 13 batches of ZTPs, which is suitable for discrimination and quality assessment of ZTPs.

[Comparative Study on Effects of Anti-contusion Injury, Analgesia and Anti-inflammation of Root and Stem of Zanthoxylum nitidum].

OBJECTIVE: To provide the scientific evidence for expansion of medicinal parts of Zanthoxylum nitidum by comparing the effects of anti-contusion injury, analgesia and anti-inflammation of its root and stem. METHODS: The pharmacological effects between root and stem of Zanthoxylum nitidum were compared by observing the anti-injury effect in rats with injury struck by hammer. The analgesic effect in mice was evaluated by writhing test and hot plate test, and the anti-inflammatory effect on paw edema induced by carrageenan and granuloma induced by cotton pellet were investigated in rats. RESULTS: Both root and stem of Zanthoxylum nitidum relieved the exterior and histological symptoms of rats' injury legs struck by hammer, decreased the numbers of mice's writhing, enhanced pain threshold of mice on heat plate, inhibited the edema of rats induced by carrageenan, and suppressed the granuloma of rats induced by cotton pellet. CONCLUSION: Stem of Zanthoxylum nitidum has similar effects of anti-contusion injury, analgesia and anti-inflammation with root of Zanthoxylum nitidum.

Angoline: a selective IL-6/STAT3 signaling pathway inhibitor isolated from Zanthoxylum nitidum.

STAT3 signaling pathway is an important target for human cancer therapy. Thus, the identification of small-molecules that target STAT3 signaling will be of great interests in the development of anticancer agents. The aim of this study was to identify novel inhibitors of STAT3 pathway from the roots of Zanthoxylum nitidum (Roxb.) DC. The bioassay-guided fractionation of MeOH extract of Z. nitidum using a STAT3-responsive gene reporter assay led to the isolation of angoline (1) as a potent and selective inhibitor of the STAT3 signaling pathway (IC50=11.56 µM). Angoline inhibited STAT3 phosphorylation and its target gene expression and consequently induced growth inhibition of human cancer cells with constitutively activated STAT3 (IC50=3.14-4.72 µM). This work provided a novel lead for the development of anti-cancer agents targeting the STAT3 signaling pathway.

Characterization and functional analysis of the genes encoding 1-deoxy-D-xylulose-5-phosphate reductoisomerase and 1-deoxy-D-xylulose-5-phosphate synthase, the two enzymes in the MEP pathway, from Amomum villosum Lour.

A DXR gene, AvDXR (GenBank accession no. FJ459894), and a DXS gene, AvDXS (GenBank accession no. FJ455512), were isolated from the leaves of Amomum villosum, one of the most well-known and authentic herbs in South China. The 1,749-bp full-length cDNA of AvDXR encoded a peptide of 472 amino acids, and the 2,347-bp full-length cDNA of AvDXS encoded a peptide of 715 amino acids. The deduced amino acid sequences of the AvDXR and AvDXS proteins share high homology with DXRs and DXSs from other plant species, and AvDXS belongs to class 1 plant DXS. The characterization based on bioinformatic analysis indicated that the AvDXR and AvDXS encoded functional proteins as DXR and DXS, respectively. The functional color assay in Escherichia coli with pAC-BETA implied that AvDXR and AvDXS encoded functional proteins that manipulated the biosynthesis of isoprenoid precursors. Both AvDXR and AvDXS were expressed extensively in the leaves, stems, roots, pericarps and seeds of A. villosum. AvDXS expression was similar in all tissues investigated, whereas higher levels of AvDXR were observed in the fruits, the main part for the accumulation of volatile oil in this plant. AvDXR was transformed into tobacco to confirm its function further. Overexpression of AvDXR in transgenic T1 generation tobacco increased DXR activity, photosynthetic pigment content and volatile isoprenoid components, and the increase of photosynthetic pigment content was consistent with the AvDXR transcription level. This study demonstrated that AvDXR plays important role in isoprenoid biosynthesis and it is useful for metabolic engineering.