Tannins in Phyllanthus emblica L. improves cisplatin efficacy in lung cancer cells by boosting endoplasmic reticulum stress to trigger immunogenic cell death.

BACKGROUND: Lung cancer is one of the deadliest cancers world-wide and immunotherapy has been considered as a promising therapeutic strategy. Previously, our study found that tannins in Phyllanthus emblica L. (PTF) could inhibit the growth of tumor by activating the immune response in liver cancer, and also exhibited a cytotoxicity on human lung cancer cells A549, H460, H1703 in vitro. OBJECTIVE: To explore whether PTF inhibited the growth of lung cancer through its immune-regulating function and to clarify underlying mechanisms. METHODS: The induction of immunogenic cell death (ICD) were characterized by calreticulin exposure, extracellular ATP secretion, and High Mobility Group Box 1(HMGB1) release both in vivo using LLC-derived xenograft tumor model and in vitro using both mouse LLC and human A549 cancer cells. RESULTS: PTF inhibited lung cancer cells growth and tumorigenesis in vivo/vitro and promoted anti-tumor immune responses. We further found that PTF could induce ICD, which then activated Type I interferon responses and CXCL9/10-mediated chemotaxis. Mechanistically, PTF induced the formation of intracellular protein aggregates and following activation of PERK/ATF4/CHOP-dependent endoplasmic reticulum stress-related ICD. Moreover, PTF improved the antitumor efficacy of cisplatin by inducing ICD both in vitro and in vivo. Finally, we screened out 5 components from PTF, including gallocatechin, gallic acid, methyl gallate, ethyl gallate and ellagic acid, which could induce ICD in vitro and might be considered as the potential antitumor pharmacodynamic substances. CONCLUSION: In conclusion, PTF inhibits the growth of lung cancer by triggering ICD and remodeling the tumor microenvironment, suggesting that PTF may have promising prospects as an adjacent immunotherapy for cancers.

Genomic, transcriptomic, and epigenomic analysis of a medicinal snake, Bungarus multicinctus, to provides insights into the origin of Elapidae neurotoxins.

The many-banded krait, Bungarus multicinctus, has been recorded as the animal resource of JinQianBaiHuaShe in the Chinese Pharmacopoeia. Characterization of its venoms classified chief phyla of modern animal neurotoxins. However, the evolutionary origin and diversification of its neurotoxins as well as biosynthesis of its active compounds remain largely unknown due to the lack of its high-quality genome. Here, we present the 1.58 Gbp genome of B. multicinctus assembled into 18 chromosomes with contig/scaffold N50 of 7.53 Mbp/149.8 Mbp. Major bungarotoxin-coding genes were clustered within genome by family and found to be associated with ancient local duplications. The truncation of glycosylphosphatidylinositol anchor in the 3'-terminal of a LY6E paralog released modern three-finger toxins (3FTxs) from membrane tethering before the Colubroidea divergence. Subsequent expansion and mutations diversified and recruited these 3FTxs. After the cobra/krait divergence, the modern unit-B of ß-bungarotoxin emerged with an extra cysteine residue. A subsequent point substitution in unit-A enabled the ß-bungarotoxin covalent linkage. The B. multicinctus gene expression, chromatin topological organization, and histone modification characteristics were featured by transcriptome, proteome, chromatin conformation capture sequencing, and ChIP-seq. The results highlighted that venom production was under a sophisticated regulation. Our findings provide new insights into snake neurotoxin research, meanwhile will facilitate antivenom development, toxin-driven drug discovery and the quality control of JinQianBaiHuaShe.

Similarities and differences between Ziqin and Kuqin in anti-inflammatory, analgesic, and antioxidant activities and their core chemical composition based on the zebrafish model and spectrum-effect relationship.

ETHNOPHARMACOLOGICAL RELEVANCE: Scutellaria baicalensis (SB) is a traditional Chinese medicine (TCM). In the clinical application of TCM, SB has been divided into two specifications (Ziqin and Kuqin) for a long time. At present, the Chinese Pharmacopoeia Commission no longer distinguishes between the two. However, the two specifications of medicinal materials and pieces are still in circulation in the market. AIM OF THE STUDY: This work aimed at investigating the similarities and differences between Ziqin and Kuqin in anti-inflammatory, analgesic, and antioxidant activities and their material basis. It will provide a new angle for relevant regulations to formulate the specifications and standards of SB. MATERIALS AND METHODS: Here we investigated the similarities and differences between Ziqin and Kuqin in anti-inflammatory, analgesic, and antioxidant activities related to four zebrafish models and three chemical tests. The chemical fingerprints of SB (Ziqin and Kuqin) were profiled by HPLC. Meanwhile, UHPLC-Q-TOF/MS was used to identify the chemical constituents of Ziqin and Kuqin. The main effect-related compounds of SB, Ziqin, and Kuqin were screened out by spectrum-effect relationship. Finally, six monomeric compounds were validated experimentally using the zebrafish inflammation model induced by CuSO4. RESULTS: Both Ziqin and Kuqin had significant anti-inflammatory, analgesic, and antioxidant activities. Kuqin had better anti-inflammatory and analgesic activities, while Ziqin had better antioxidant activity. HPLC fingerprint and UHPLC-Q-TOF/MS evaluation showed that the chemical composition types and main components of Ziqin and Kuqin were basically the same, while the contents and proportions of chemical components in Ziqin and Kuqin were different. By spectrum-effect relationship, compounds X1, X2 (luteoloside), X3, X4 (baicalin), X6 (wogonoside), X7 (baicalein), X8 (wogonin), and X9 (oroxylin A) were the same active chemical constituents of Ziqin and Kuqin. The core components of anti-inflammatory and analgesia activities in Kuqin were compounds X1, X2, X3, X5, X6, X7, X8, and X9. The antioxidant core active components of Ziqin were compounds X2, X3, X4, X6, X7, and X9. Among them, luteoloside, baicalin, wogonoside, baicalein, wogonin, and oroxylin A were validated successfully with good anti-inflammatory effects. CONCLUSIONS: This study revealed that Ziqin and kuqin have high similarity in chemical composition, but their proportions and active core components are different. This may be one of the main reasons why they have the same activity but different activity trends. These findings will help to improve the understanding of the different clinical applications of Ziqin and Kuqin, and provide a reference for the formulation of quality standards and their further research.

A Comparative Analysis of the Chloroplast Genomes of Four Polygonum Medicinal Plants.

Polygonum is a generalized genus of the Polygonaceae family that includes various herbaceous plants. In order to provide aid in understanding the evolutionary and phylogenetic relationship in Polygonum at the chloroplast (cp) genome-scale level, we sequenced and annotated the complete chloroplast genomes of four Polygonum species using next-generation sequencing technology and CpGAVAS. Then, repeat sequences, IR contractions, and expansion and transformation sites of chloroplast genomes of four Polygonum species were studied, and a phylogenetic tree was built using the chloroplast genomes of Polygonum. The results indicated that the chloroplast genome construction of Polygonum also displayed characteristic four types of results, comparable to the published chloroplast genome of recorded angiosperms. The chloroplast genomes of the four Polygonum plants are highly consistent in genome size (159,015 bp-163,461 bp), number of genes (112 genes, including 78 protein-coding genes, 30 tRNA genes, and 4 rRNA genes), gene types, gene order, codon usage, and repeat sequence distribution, which identifies the high preservation among the Polygonum chloroplast genomes. The Polygonum phylogenetic tree was recreated by a full sequence of the chloroplast genome, which illustrates that the P. bistorta, P. orientale, and P. perfoliatum are divided into the same branch, and P. aviculare belongs to Fallopia. The precise system site of lots base parts requires further verification, but the study would provide a basis for developing the available genetic resources and evolutionary relationships of Polygonum.

Licorice Germplasm Resources Identification Using DNA Barcodes Inner-Variants.

Based on the gradual transformation from wild growth to artificial cultivation, the accurate authentication of licorice seeds contributes to the first committed step of its quality control and is pivotal to ensure the clinical efficacy of licorice. However, it is still challenging to obtain genetically stable licorice germplasm resources due to the multi-source, multi-heterozygous, polyploid, and hybrid characteristics of licorice seeds. Here, a new method for determining the heterozygosity of licorice seed mixture, based on the various sites, and finding the composition characteristics of licorice seed is preliminarily designed and proposed. Namely, high-throughput full-length multiple DNA barcodes(HFMD), based on ITS multi-copy variation exist, the full-length amplicons of ITS2, psbA-trnH and ITS are directly sequenced by rDNA through the next-generation sequence(NGS) and single-molecule real-time (SMRT) technologies. By comparing the three sequencing methods, our results proved that SMRT sequencing successfully identified the complete gradients of complex mixed samples with the best performance. Meanwhile, HFMD is a brilliant and feasible method for evaluating the heterozygosity of licorice seeds. It shows a perfect interpretation of DNA barcoding and can be applied in multi-base multi-heterozygous and polyploid species.

Diversity and composition of bacterial endophytes among plant parts of Panax notoginseng.

BACKGROUND: Bacterial endophytes are widespread inhabitants inside plant tissues that play crucial roles in plant growth and biotransformation. This study aimed to offer information for the exploitation of endophytes by analyzing the bacterial endophytes in different parts of Panax notoginseng. METHODS: We used high-throughput sequencing methods to analyze the diversity and composition of bacterial endophytes from different parts of P. notoginseng. RESULTS: A total of 174,761 classified sequences were obtained from the analysis of 16S ribosomal RNA in different parts of P. notoginseng. Its fibril displayed the highest diversity of bacterial endophytes. Principal coordinate analysis revealed that the compositions of the bacterial endophytes from aboveground parts (flower, leaf, and stem) differed from that of underground parts (root and fibril). The abundances of Conexibacter, Gemmatimonas, Holophaga, Luteolibacter, Methylophilus, Prosthecobacter, and Solirubrobacter were significantly higher in the aboveground parts than in the underground parts, whereas the abundances of Bradyrhizobium, Novosphingobium, Phenylobacterium, Sphingobium, and Steroidobacter were markedly lower in the aboveground parts. CONCLUSIONS: Our results elucidated the comprehensive diversity and composition profiles of bacterial endophytes in different parts of 3-year-old P. notoginseng. Our data offered pivotal information to clarify the role of endophytes in the production of P. notoginseng and its important metabolites.

Integrated metabolomic and transcriptomic analyses revealed the distribution of saponins in Panax notoginseng.

Panax notoginseng is famous for its important therapeutic effects. Saponins are bioactive compounds found in different parts and developmental stages of P. notoginseng plants. Thus, it is urgently to study saponins distribution in different parts and growth ages of P. notoginseng plants. In this study, potential biomarkers were found, and their chemical characteristic differences were revealed through metabolomic analysis. High-performance liquid chromatography data indicated the higher content of saponins (i.e., Rg1, Re, Rd, and Rb1) in the underground parts than that in the aerial parts. 20(S)-Protopanaxadiol saponins were mainly distributed in the aerial parts. Additionally, the total saponin content in the 3-year-old P. notoginseng plant (188.0 mg/g) was 1.4-fold higher than that in 2-year-old plant (130.5 mg/g). The transcriptomic analysis indicated the tissue-specific transcription expression of genes, namely, PnFPS, PnSS, PnSE1, PnSE2, and PnDS, which encoded critical synthases in saponin biosyntheses. These genes showed similar expression patterns among the parts of P. notoginseng plants. The expression levels of these genes in the flowers and leaves were 5.2fold higher than that in the roots and fibrils. These results suggested that saponins might be actively synthesized in the aerial parts and transformed to the underground parts. This study provides insights into the chemical and genetic characteristics of P. notoginseng to facilitate the synthesis of its secondary metabolites and a scientific basis for appropriate collection and rational use of this plant.

Rhizospheric microbial communities are driven by Panax ginseng at different growth stages and biocontrol bacteria alleviates replanting mortality.

The cultivation of Panax plants is hindered by replanting problems, which may be caused by plant-driven changes in the soil microbial community. Inoculation with microbial antagonists may efficiently alleviate replanting issues. Through high-throughput sequencing, this study revealed that bacterial diversity decreased, whereas fungal diversity increased, in the rhizosphere soils of adult ginseng plants at the root growth stage under different ages. Few microbial community, such as Luteolibacter, Cytophagaceae, Luteibacter, Sphingomonas, Sphingomonadaceae, and Zygomycota, were observed; the relative abundance of microorganisms, namely, Brevundimonas, Enterobacteriaceae, Pandoraea, Cantharellales, Dendryphion, Fusarium, and Chytridiomycota, increased in the soils of adult ginseng plants compared with those in the soils of 2-year-old seedlings. Bacillus subtilis 50-1, a microbial antagonist against the pathogenic Fusarium oxysporum, was isolated through a dual culture technique. These bacteria acted with a biocontrol efficacy of 67.8%. The ginseng death rate and Fusarium abundance decreased by 63.3% and 46.1%, respectively, after inoculation with B. subtilis 50-1. Data revealed that microecological degradation could result from ginseng-driven changes in rhizospheric microbial communities; these changes are associated with the different ages and developmental stages of ginseng plants. Biocontrol using microbial antagonists alleviated the replanting problem.

Panax ginseng genome examination for ginsenoside biosynthesis.

Ginseng, which contains ginsenosides as bioactive compounds, has been regarded as an important traditional medicine for several millennia. However, the genetic background of ginseng remains poorly understood, partly because of the plant's large and complex genome composition. We report the entire genome sequence of Panax ginseng using next-generation sequencing. The 3.5-Gb nucleotide sequence contains more than 60% repeats and encodes 42 006 predicted genes. Twenty-two transcriptome datasets and mass spectrometry images of ginseng roots were adopted to precisely quantify the functional genes. Thirty-one genes were identified to be involved in the mevalonic acid pathway. Eight of these genes were annotated as 3-hydroxy-3-methylglutaryl-CoA reductases, which displayed diverse structures and expression characteristics. A total of 225 UDP-glycosyltransferases (UGTs) were identified, and these UGTs accounted for one of the largest gene families of ginseng. Tandem repeats contributed to the duplication and divergence of UGTs. Molecular modeling of UGTs in the 71st, 74th, and 94th families revealed a regiospecific conserved motif located at the N-terminus. Molecular docking predicted that this motif captures ginsenoside precursors. The ginseng genome represents a valuable resource for understanding and improving the breeding, cultivation, and synthesis biology of this key herb.

Comprehensive Characterization for Ginsenosides Biosynthesis in Ginseng Root by Integration Analysis of Chemical and Transcriptome.

Herbgenomics provides a global platform to explore the genetics and biology of herbs on the genome level. Panax ginseng C.A. Meyer is an important medicinal plant with numerous pharmaceutical effects. Previous reports mainly discussed the transcriptome of ginseng at the organ level. However, based on mass spectrometry imaging analyses, the ginsenosides varied among different tissues. In this work, ginseng root was separated into three tissues-periderm, cortex and stele-each for five duplicates. The chemical analysis and transcriptome analysis were conducted simultaneously. Gene-encoding enzymes involved in ginsenosides biosynthesis and modification were studied based on gene and molecule data. Eight widely-used ginsenosides were distributed unevenly in ginseng roots. A total of 182,881 unigenes were assembled with an N50 contig size of 1374 bp. About 21,000 of these unigenes were positively correlated with the content of ginsenosides. Additionally, we identified 192 transcripts encoding enzymes involved in two triterpenoid biosynthesis pathways and 290 transcripts encoding UDP-glycosyltransferases (UGTs). Of these UGTs, 195 UGTs (67.2%) were more highly expressed in the periderm, and that seven UGTs and one UGT were specifically expressed in the periderm and stele, respectively. This genetic resource will help to improve the interpretation on complex mechanisms of ginsenosides biosynthesis, accumulation, and transportation.

Survey of commercial Rhodiola products revealed species diversity and potential safety issues.

The adulteration of herbal products is a threat to consumer safety. Here we surveyed the species composition of commercial Rhodiola products using DNA barcoding as a supervisory method. A Rhodiola dietary supplement DNA barcode database was successfully constructed using 82 voucher samples from 10 Rhodiola species. Based on the DNA barcoding standard operating procedure (SOP), we used this database to identify 100 Rhodiolae Crenulatae Radix et Rhizoma decoction piece samples that were purchased from drug stores and hospitals. The results showed that only 36 decoction piece sequences (40%) were authentic R. crenulata, which is recorded in Chinese Pharmacopeia, whereas the other samples were all adulterants and may indicate a potential safety issue. Among the adulterants, 35 sequences (38.9%) were authenticated as R. serrata, nine sequences (10%) were authenticated as R. rosea, which is documented in the United States Pharmacopeia, and the remaining samples were authenticated as other three Rhodiola species. This result indicates decoction pieces that are available in the market have complex origins and DNA barcoding is a convenient tool for market supervision.

Stability and accuracy assessment of identification of traditional Chinese materia medica using DNA barcoding: a case study on Flos Lonicerae Japonicae.

DNA barcoding is a novel molecular identification method that aids in identifying traditional Chinese materia medica using traditional identification techniques. However, further study is needed to assess the stability and accuracy of DNA barcoding. Flos Lonicerae Japonicae, a typical medicinal flower, is widely used in China, Korea, and other Southeast Asian countries. However, Flos Lonicerae Japonicae and its closely related species have been misused and traded at varying for a wide range of prices. Therefore, Flos Lonicerae Japonicae must be accurately identified. In this study, the ITS2 and psbA-trnH regions were amplified by polymerase chain reaction (PCR). Sequence assembly was performed using CodonCode Aligner V 3.5.4. The intra- versus inter-specific variations were assessed using six metrics and "barcoding gaps." Species identification was conducted using BLAST1 and neighbor-joining (NJ) trees. Results reveal that ITS2 and psbA-trnH exhibited an average intraspecific divergence of 0.001 and 0, respectively, as well as an average inter-specific divergence of 0.0331 and 0.0161. The identification efficiency of ITS2 and psbA-trnH evaluated using BLAST1 was 100%. Flos Lonicerae Japonicae was formed into one clade through the NJ trees. Therefore, Flos Lonicerae Japonicae can be stably and accurately identified through the ITS2 and psbA-trnH regions, respectively.

A novel phenolic compound from Phyllanthus emblica.

A new compound, mucic acid 3-O-gallate (1), was isolated from the fruit of Phyllanthus emblica L, together with 5 known compounds (2-6). Their structures were elucidated on the basis of spectroscopic analysis, including 1D and 2D NMR techniques, and by comparison with literature data.

Transcriptome analysis of buds and leaves using 454 pyrosequencing to discover genes associated with the biosynthesis of active ingredients in Lonicera japonica Thunb.

BACKGROUND: Lonicera japonica Thunb. is a plant used in traditional Chinese medicine known for its anti-inflammatory, anti-oxidative, anti-carcinogenic, and antiviral pharmacological properties. The major active secondary metabolites of this plant are chlorogenic acid (CGA) and luteoloside. While the biosynthetic pathways of these metabolites are relatively well known, the genetic information available for this species, especially the biosynthetic pathways of its active ingredients, is limited. METHODOLOGY/PRINCIPAL FINDINGS: We obtained one million reads (average length of 400 bp) in a whole sequence run using a Roche/454 GS FLX titanium platform. Altogether, 85.69% of the unigenes covering the entire life cycle of the plant were annotated and 325 unigenes were assigned to secondary metabolic pathways. Moreover, 2039 unigenes were predicted as transcription factors. Nearly all of the possible enzymes involved in the biosynthesis of CGA and luteoloside were discovered in L. japonica. Three hydroxycinnamoyl transferase genes, including two hydroxycinnamoyl-CoA quinate hydroxycinnamoyl transferase genes and one hydroxycinnamoyl-CoA shikimate/quinate hydroxycinnamoyl transferase (HCT) gene featuring high similarity to known genes from other species, were cloned. The HCT gene was discovered for the first time in L. japonica. In addition, 188 candidate cytochrome P450 unigenes and 245 glycosyltransferase unigenes were found in the expressed sequence tag (EST) dataset. CONCLUSION: This study provides a high quality EST database for L. japonica by 454 pyrosequencing. Based on the EST annotation, a set of putative genes involved in CGA and luteoloside biosynthetic pathways were discovered. The database serves as an important source of public information on genetic markers, gene expression, genomics, and functional genomics in L. japonica.

Elsholtzia: phytochemistry and biological activities.

Plants of the genus Elsholtzia (Lamiaceae) have a long history of medicinal use in folk. The phytochemical investigations revealed the presence of flavonoids, phenylpropanoids, terpenoids, and other compounds. Abundant volatile components are also identified. Pure compounds, volatile constituents and crude extracts from the genus exhibited a wide spectrum of in vitro and in vivo pharmacological activities. The aims of this review hopefully provide comprehensive information on the distribution, phytochemistry, volatile components, and pharmacological research of Elsholtzia for exploring the potential and advance researches.

Phytochemicals and biological studies of plants from the genus Balanophora.

This review focus on the phytochemical progress and biological studies of plants from the genus Balanophora (Balanophoraceae) over the past few decades, in which most plants growth in tropical and subtropical regions of Asia and Oceania, and nearly 20 species ranged in southwest China. These dioeciously parasitic plants are normally growing on the roots of the evergreen broadleaf trees, especially in the family of Leguminosae, Ericaceae, Urticaceae, and Fagaceae. The plants are mainly used for clearing away heat and toxic, neutralizing the effect of alcoholic drinks, and as a tonic for the treatment of hemorrhoids, stomachache and hemoptysis. And it has been used widely throughtout local area by Chinese people.Cinnamic acid derivative tannins, possessing a phenylacrylic acid derivative (e. g. caffeoyl, coumaroyl, feruloyl or cinnamoyl), which connected to the C(1) position of a glucosyl unit by O-glycosidic bond, are the characteristic components in genus Balanophora. In addition, several galloyl, caffeoyl and hexahydroxydiphenoyl esters of dihydrochalcone glucosides are found in B. tobiracola, B. harlandii, and B. papuana. Other compounds like phenylpropanoids, flavonoids, terpenoids and sterols are also existed. And their biological activities, such as radical scavenging activities, HIV inhibiting effects, and hypoglycemic effects are highlighted in the review.