Identification and characterization of potential antioxidant components in Isodon amethystoides (Benth.) Hara tea leaves by UPLC-LTQ-Orbitrap-MS.

Isodon amethystoides (Benth.) Hara (IA) tea is a commonly used dietetic Chinese herb and employed for the treatments of tumor and lung abscess. To assess chemical composition and antioxidant capacity of IA leaves extract, a UPLC-LTQ-Orbitrap-MS method and antioxidant tests were used, respectively. 17 compounds were identified including Vinyl caffeate (1), 3,4-dimethoxyphenyl-ß-D-glucopyranoside (2), Rutin (3), Quercetin (4), Loliolide (5), Caffeic acid (6), Rubesanolide D (7), Isorhamnetin (8), Lambertic acid (9), 6, 7-Dehydroroyleanone (10), Dihydrorabdokunmin C (11), Nervosin (12), Quercitrin (13), Vitexin (14), ß-sitosterol (15), Wangzaozin A (16), Amethystonoic acid (17). Among these, 1-14 compounds were novel and have not been reported ever before in IA while component 10 was a novel finding within this genus. Flavonoid components showed better free radical scavenging ability and profound correlation was observed between diterpenoid compounds content and flavonoids activity. Our results provide experimental basis for extraction and separation of chemical constituents of IA which are antioxidant in nature.

Quality evaluation of bee pollens by chromatographic fingerprint and simultaneous determination of its major bioactive components.

Bee pollens constitute a large number of flavonoids and thus possess great medicinal value. However different varieties of bee pollen flavonoids vary with different species and their content also differ greatly in different region. Herein, the aim of present research is to establish a method based on high performance liquid chromatography (HPLC) for quantitative analysis of flavonoids compounds and chemical fingerprint analysis of bee pollen. Five batches of rape bee pollen collected from different region of China and particularly six bee pollen species obtained in Anhui were used to establish the fingerprint. The feasibility and advantages of the used HPLC fingerprint were verified for its similarity evaluation by systematically comparing chromatograms with professional analytical software. The similarities of liquid chromatography fingerprints for five batches of rape bee pollen were more than 0.994 while six batches of different species of bee pollen were lower than 0.810. In quantitative analysis, the six compounds showed good regression (R ≥ 0.9964) within the test ranges, and all the values for the RSD were lower than 2%. The developed HPLC fingerprint method was found simple, reliable, and it was validated for the quality control and identification of bee pollen. Additionally, simultaneous quantification of six flavonoids ingredients in the bee pollen samples was conducted to reveal the variation in their content. The results indicated that the HPLC fingerprint, as a characteristic distinguishing method combining similarity evaluation and quantification analysis, can be successfully used to assess the quality and also to identify the authenticity of bee pollen.

Revealing the Synergistic Mechanism of Multiple Components in Compound Fengshiding Capsule for Rheumatoid Arthritis Therapeutics by Network Pharmacology.

BACKGROUND: Compound Fengshiding capsule (CFC), is a Chinese formulation from herbal origin including Alangium platanifolium, Angelicae dahurica, Cynanchum paniculatum and Glycyrrhiza uralensis. CFC is widely used as clinical therapy against rheumatoid arthritis. However, its exact mechanism of action has not been explored yet. METHODS: In order to explore the synergistic mechanism of CFC, we designed a study adopting network pharmacology scheme to screen the action targets in relation to the CFC components. The study analyses target facts of salicin, paeonol, liquiritin and imperatorin from PubMed database, and explores the potential pharmacological targets of rheumatoid arthritis, cervical neuralgia and sciatica related diseases for their interaction. RESULTS: The results of boosted metabolic pathway showed that the chemical components of CFC interrupted many immune-related pathways, thus participating in immunity regulation of the body and playing a role in the treatment of rheumatism. Collectively, CFC has apoptotic, oxidative stress modulatory and anti-inflammatory effects that accumulatively serve for its clinical application against rheumatoid arthritis. CONCLUSION: Conclusively, our findings from present study reconnoiters and compacts systematic theoretical approach by utilizing the network pharmacology mechanism of four effective components for the treatment of rheumatism indicating sufficient potential drug targets associated with CFC against rheumatism. These interesting findings entreaties for further in vitro and in vivo studies on the mechanism of compound active ingredient against rheumatism.

Liquiritin from Glycyrrhiza uralensis Attenuating Rheumatoid Arthritis via Reducing Inflammation, Suppressing Angiogenesis, and Inhibiting MAPK Signaling Pathway.

Among the various treatments, induction of synoviocyte apoptosis by natural products during a rheumatoid arthritis (RA) pathological condition can be considered to have vast potential. However, it is unclear that liquiritin, a kind of natural flavonoid extracted from the roots of Glycyrrhiza uralensis, induced the apoptosis of the synovial membrane and its molecular mechanism. In this study, interleukin-1ß (IL-1ß)-RA-FLS cells were incubated with different concentrations of liquiritin. An MTT assay, Hoechst 33342 staining, JC-1 staining, and Western blot were used to check the viability, cell apoptosis, mitochondrial membrane potential changes, and the expression of related proteins, respectively. In vivo, a TUNEL assay and HE staining of tissue were used for histopathological evaluation. Our results showed that liquiritin significantly inhibited the proliferation of IL-1ß-induced-RA-FLS, promoted nuclear DNA fragmentation, and changed the mitochondrial membrane potential to accelerate cell apoptosis. Liquiritin downregulated the ratio of Bcl-2/Bax and inhibited the VEGF expression and phosphorylation of JNK and P38. Moreover, liquiritin improved the clinical score of rheumatism, inflammatory infiltration, and angiogenesis and induced apoptosis of the synovial tissue in vivo. Hence, liquiritin ameliorates RA by reducing inflammation, blocking MAPK signaling, and restraining angiogenesis.

Salicin from Alangium chinense Ameliorates Rheumatoid Arthritis by Modulating the Nrf2-HO-1-ROS Pathways.

Rheumatoid arthritis (RA) is a chronic inflammatory disorder linked to oxidative stress of rheumatoid arthritis fibroblast-like synoviocytes (RA-FLSs). The effects and potential mechanism of salicin on inflammation and oxidative stress of RA-FLSs were examined by MTT, ELISA, and Western blot methods. Salicin significantly reduced cell viability (82.03 ± 7.06, P < 0.01), cytokines (47.70 ± 1.48 ng/L for TNF-α, 30.03 ± 3.49 ng/L for IL-6) ( P < 0.01), and matrix metalloproteinases-1/-3 expression ( P < 0.01) in IL-1ß-induced RA-FLSs and inhibited ROS generation and p65 phosphorylation ( P < 0.01) as compared with IL-1ß-induced treatment. Moreover, salicin promoted Nrf2 nuclear translocation (2.15 ± 0.21) and HO-1 expression (1.12 ± 0.05) and reduced ROS production in IL-1ß-induced RA-FLSs ( P < 0.01). Salicin not only reduced the collagen-induced arthritis by reducing the clinical score ( P < 0.01), inflammatory infiltration, and synovial hyperplasia in vivo but also suppressed the oxidative damage indexes (SOD 155.40 ± 6.53 U/mg tissue, MDA 152.80 ± 5.89 nmol/g tissue, GSH 50.98 ± 3.45 nmol/g tissue, and CAT 0.92 ± 0.10 U/g protein) ( P < 0.01) of ankle joint cells. Conclusively, our findings indicate that salicin ameliorates rheumatoid arthritis, which may be associated with oxidative stress and Nrf2-HO-1-ROS pathways in RA-FLSs.

Protective effect of Rabdosia amethystoides (Benth) Hara extract on acute liver injury induced by Concanavalin A in mice through inhibition of TLR4-NF-κB signaling pathway.

Extract of Rabdosia amethystoides (Benth) Hara (ERA), a traditional Chinese medicine has antibacterial, antiviral, anti-tumor, anti-hepatitis and anti-inflammatory properties. However, the hepatoprotective effects and molecular mechanisms of ERA on acute liver injury have not been fully elucidated. This study aims to investigate the anti-inflammatory effect and liver protection of ERA against the acute liver injury induced by Concanavalin A (Con A) and its underlying molecular mechanisms in mice. Mice received ERA (50, 100, 150 mg/kg body weight) by gavage before Con A intravenous administration. We found that ERA pretreatment was able to significantly reduce the elevated serum alanine and aspartate aminotransferase levels and liver necrosis in Con A-induced hepatitis. In addition, ERA treatment significantly decreased the myeloperoxidase, malondialdehyde levels and augmented superoxide dismutase level in the liver tissue, and also suppressed the secretion of proinflammatory cytokines in the serum, compared with Con A group by enzyme linked immunosorbent assay. Furthermore, we observed that ERA pretreatment can significantly decrease the expression level of Toll-like receptor (TLR) 4 mRNA or protein in liver tissues. Further results showed that ERA pretreatment was capable of attenuating the activation of the NF-κB pathway by inhibiting IκBα kinase and p65 phosphorylation in Con A-induced liver injury. Our results demonstrate that ERA pretreatment has hepatoprotective property against Con A-induced liver injury through inhibition of inflammatory mediators in mice. The beneficial effect of ERA may be mediated by the downregulation of TLR4 expression and the inhibition of NF-κB activation.

Conserved regulatory motifs at phenylethanolamine N-methyltransferase (PNMT) are disrupted by common functional genetic variation: an integrated computational/experimental approach.

The adrenomedullary hormone epinephrine transduces environmental stressors into cardiovascular events (tachycardia and hypertension). Although the epinephrine biosynthetic enzyme PNMT genetic locus displays both linkage and association to such traits, genetic variation underlying these quantitative phenotypes is not established. Using an integrated suite of computational and experimental approaches, we elucidate a functional mechanism for common (minor allele frequencies > 30%) genetic variants at PNMT. Transcription factor binding motif prediction on mammalian PNMT promoter alignments identified two variant regulatory motifs, SP1 and EGR1, disrupted by G-367A (rs3764351), and SOX17 motif created by G-161A (rs876493). Electrophoretic mobility shifts of approximately 30-bp oligonucleotides containing ancestral versus variant alleles validated the computational hypothesis. Queried against chromaffin cell nuclear protein extracts, only the G-367 and -161A alleles shifted. Specific antibodies applied in electrophoretic gel shift experiments confirmed binding of SP1 and EGR1 to G-367 and SOX17 to -161A. The in vitro allele-specific binding was verified in cella through promoter reporter assays: lower activity for -367A haplotypes cotransfected by SP1 (p = 0.002) and EGR1 (p = 0.034); and enhanced inhibition of -161A haplotypes (p = 0.0003) cotransfected with SP1 + SOX17. Finally, we probed cis/trans regulation with endogenous factors by chromatin immunoprecipitation using SP1/EGR1/SOX17 antibodies. We describe the systematic application of complementary computational and experimental techniques to detect and document functional genetic variation in a trait-associated regulatory region. The results provide insight into cis and trans transcriptional mechanisms whereby common variation at PNMT can give rise to quantitative changes in human physiological and disease traits. Thus, PNMT variants in cis may interact with nuclear factors in trans to govern adrenergic activity.