Nitrogen-fixing bacteria promote growth and bioactive components accumulation of Astragalus mongholicus by regulating plant metabolism and rhizosphere microbiota.

BACKGROUND: The excessive application of chemical fertilizers in the cultivation of Astragalus mongholicus Bunge results in a reduction in the quality of the medicinal plant and compromises the sustainable productivity of the soil. PGPB inoculant is a hot topic in ecological agriculture research. In the cultivation of Astragalus mongholicus, the screened nitrogen-fixing bacteria can promote plant growth, however, whether it can promote the accumulation of main bioactive components remains unknown. In this study, mixed inoculants containing 5 strains of growth promoting bacteria (Rhizobium T16 , Sinorhizobium T21 , Bacillus J1 , Bacillus G4 and Arthrobacter J2) were used in the field experiment. The metabolic substances in the root tissues of Astragalus mongholicus were identified during the harvest period by non-targeted metabolomics method, and the differential metabolites between groups were identified by statistical analysis. Meanwhile, high-throughput sequencing was performed to analyze the changes of rhizosphere soil and endophytic microbial community structure after mixed microbial treatment. RESULTS: The results of non-targeted metabolism indicated a significant increase in the levels of 26 metabolites after treatment including 13 flavonoids, 3 saponins and 10 other components. The contents of three plant hormones (abscisic acid, salicylic acid and spermidine) also increased after treatment, which presumed to play an important role in regulating plant growth and metabolism. Studies on endosphere and rhizosphere bacterial communities showed that Rhzobiaceae, Micromonosporaceae, and Hypomicrobiaceae in endophytic, and Oxalobactereae in rhizosphere were significantly increased after treatment. These findings suggest their potential importance in plant growth promotion and secondary metabolism regulation. CONCLUSIONS: This finding provides a basis for developing nitrogen-fixing bacteria fertilizer and improving the ecological planting efficiency of Astragalus mongholicus.

Legume nodulation and nitrogen fixation require interaction of DnaJ-like protein and lipid transfer protein.

The lipid transport protein (LTP) product of the AsE246 gene of Chinese milk vetch (Astragalus sinicus) contributes to the transport of plant-synthesized lipids to the symbiosome membranes (SMs) that are required for nodule organogenesis in this legume. However, the mechanisms used by nodule-specific LTPs remain unknown. In this study, a functional protein in the DnaJ-like family, designated AsDJL1, was identified and shown to interact with AsE246. Immunofluorescence showed that AsDJL1 was expressed in infection threads (ITs) and in nodule cells and that it co-localized with rhizobium, and an immunoelectron microscopy assay localized the protein to SMs. Via co-transformation into Nicotiana benthamiana cells, AsDJL1 and AsE246 displayed subcellular co-localization in the cells of this heterologous host. Co-immunoprecipitation assays confirmed that AsDJL1 interacted with AsE246 in nodules. The essential interacting region of AsDJL1 was determined to be the zinc finger domain at its C-terminus. Chinese milk vetch plants transfected with AsDJL1-RNAi had significantly decreased numbers of ITs, nodule primordia and nodules as well as reduced (by 83%) nodule nitrogenase activity compared with the controls. By contrast, AsDJL1 overexpression led to increased nodule fresh weight and nitrogenase activity. RNAi-AsDJL1 also significantly affected the abundance of lipids, especially digalactosyldiacylglycerol, in early-infected roots and transgenic nodules. Taken together, the results of this study provide insights into the symbiotic functions of AsDJL1, which may participate in lipid transport to SMs and play an essential role in rhizobial infection and nodule organogenesis.

Genome-wide identification and expression pattern analysis of the GRF transcription factor family in Astragalus mongholicus.

BACKGROUND: Astragalus membranaceus is a plant of the Astragalus genus, which is used as a traditional Chinese herbal medicine with extremely high medicinal and edible value. Astragalus mongholicus, as one of the representative medicinal materials with the same origin of medicine and food, has a rising market demand for its raw materials, but the quality is different in different production areas. Growth-regulating factors (GRF) are transcription factors unique to plants that play important roles in plant growth and development. Up to now, there is no report about GRF in A. mongholicus. METHODS AND RESULTS: This study conducted a genome-wide analysis of the AmGRF gene family, identifying a total of nine AmGRF genes that were classified into subfamily V based on phylogenetic relationships. In the promoter region of the AmGRF gene, we successfully predicted cis-elements that respond to abiotic stress, growth, development, and hormone production in plants. Based on transcriptomic data and real-time quantitative polymerase chain reaction (qPCR) validation, the results showed that AmGRFs were expressed in the roots, stems, and leaves, with overall higher expression in leaves, higher expression of AmGRF1 and AmGRF8 in roots, and high expression levels of AmGRF1 and AmGRF9 in stems. CONCLUSIONS: The results of this study provide a theoretical basis for the further exploration of the functions of AmGRFs in plant growth and development.

Identification and Expression Analysis of Cytochrome P450 Genes Probably Involved in Triterpenoid Saponins Biosynthesis in Astragalus mongholicus.

Cytochromes P450 (P450s) are one of the largest enzymatic protein families and play critical roles in the synthesis and metabolism of plant secondary metabolites. Astragaloside IV (AS-IV) is one of the primary active components in Astragalus herbs, exhibiting diverse biological activities and pharmacological effects. However, P450s involved in the astragaloside biosynthesis have not been systematically analyzed in Astragalus mongholicus (A. mongholicus). In this study, we identified 209 P450 genes from the genome of A. mongholicus (AmP450s), which were classified into nine clans and 47 families and performed a systematic overview of their physical and chemical properties, phylogeny, gene structures and conserved motifs. Weighted gene co-expression network analysis (WGCNA) revealed that AmP450s are critical in the astragaloside biosynthesis pathway. The expression levels of these AmP450s were verified by quantitative real-time PCR (qRT-PCR) analysis in the root, stem and leaf, showing that most AmP450s are abundant in the root. Additionally, the correlation analysis between gene expressions and AS-IV content showed that twelve AmP450s, especially CYP71A28, CYP71D16 and CYP72A69, may have significant potential in the biosynthesis of astragaloside. This study systematically investigates the P450s of A. mongholicus and offers valuable insights into further exploring the functions of CYP450s in the astragaloside biosynthesis pathway.

Astragalus injection ameliorates lipopolysaccharide-induced cognitive decline via relieving acute neuroinflammation and BBB damage and upregulating the BDNF-CREB pathway in mice.

CONTEXT: Post-sepsis cognitive impairment is one of the major sequelae observed in survivors of sepsis. Astragalus injection is the normally preferred treatment in sepsis in clinical settings. OBJECTIVE: This study evaluated the benefits and related mechanism of Astragalus injection on post-sepsis cognitive impairment. MATERIALS AND METHODS: C57BL/6J mice were divided into three groups: Control, LPS (2.5 mg/kg, i.p.), and LPS + Astragalus injection (5.0 mL/kg). The surviving mice from sepsis were injected with material named Astragalus injection continuously for 13 days. Behavioural tests were first conducted to evaluate the benefits. Second, inflammatory cytokines secretion, BBB integrity, neurodegeneration, and protein expression was evaluated in vivo and in vitro. RESULTS: Compared with the LPS group, mice in Astragalus injection group exhibited shorter escape latency (34.6 s versus 24.5 s) in the Morris water maze test. Treatment with Astragalus injection could reverse LPS-induced neuroinflammation in mice and BV2 cells. Continuous Astragalus injection treatment not only prevented blood-brain barrier dysfunction, but also prevented neurodegeneration. Further molecular docking tests and western blot results reflected that the main constituents of Astragalus injection could interact with TrkB (the estimated binding energy values were -7.0 to -5.0 kcal/mol) and upregulate the protein expression of BDNF/TrkB/CREB signalling pathway during the chronic stage in mice. DISCUSSION: Astragalus injection treatment could reduce neuroinflammation, reverse BBB dysfunction, prevent neurodegeneration, and upregulate BDNF-CREB pathway during LPS-induced sepsis, ultimately preventing the development of cognitive decline. CONCLUSION: Astragalus injection could be a potential preventive and therapeutic strategy for sepsis survivors in clinical settings.

[Molecular mechanism underlying difference of astragaloside â £ content in imitating wild and cultivated Astragalus mongholicus].

The difference of astragaloside Ⅳ content and the expression of its biosynthesis related genes in imitating wild Astragalus mongolicus(IWA) and cultivated A.mongolicus(CA) under different growth years were systematically compared and analyzed.Then the key enzyme genes affected the difference of astragaloside Ⅳ content in the above two A.mongolicus were screened.High-perfo-rmance liquid chromatography(HPLC)was used to determine the content of astragaloside Ⅳ in A.mongolicusunderthe above two diffe-rent growth patterns.Based on the Illumina HiSeq and PacBio high-throughput sequencing platforms, thesecond-and third-generation transcriptome sequencing(RNA-Seq)databaseof the two A.mongolicuswas constructed.The related enzyme genes in the biosynthetic pathway of astragaloside Ⅳ were screened and verified byquantitative reverse transcriptase polymerase chain reaction(RT-qPCR).The RNA-sequencing(RNA-Seq) and RT-qPCR data of each gene were subjected to correlation analysis and trend analysis.The results showed that the variation trend of astragaloside Ⅳ contentby HPLC wasthe same as that of genes by RNA-Seq and RT-qPCR in 1-4 year IWA and 1-2 year CA.The trend level of astragaloside Ⅳ contentwas lower in 2-year IWA than 1-year IWA.Compared with 2-year IWA, 3-year IWA had an upward trend, while 4-year IWA hada downward trend versus 3-year IWA.Additionally, 1-year CA had increased trendthan 2-year CA.However, the content of astragaloside Ⅳ in 5-year IWA was higher than that of 6-year IWA, which wasinconsistent with the findings of RNA-Seq and RT-qPCR.This study preliminarily clarifiedthat the difference of astragaloside Ⅳ contentin 1-4 year IWA and 1-2 year CA wasclosely related to the expression of the upstream and midstream genes(MVK, CMK, PMK, MVD, SS) in the biosynthetic pathway.The results facilitate the production and planting of Radix Astragali seu Hedysari.

[Mechanism of Astragali Radix-Puerariae Lobatae Radix combination in regulating type 2 diabetes mellitus through AMPK signaling pathway: based on network pharmacology and experimental verification].

This study aims to explore the mechanism of Astragali Radix-Puerariae Lobatae Radix(AP) combination in the treatment of type 2 diabetes mellitus(T2 DM) based on network pharmacology and experiment. The effective components and targets of the pair were retrieved from the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform(TCMSP) and targets of T2 DM from each disease database. On this basis, the common targets of the medicinals and the disease were screened out. The protein-protein interaction(PPI) network was established based on STRING. Then Cytoscape 3.7.1 was employed for visualization of the common targets and the network topology analysis of key targets, followed by Gene Ontology(GO) term enrichment and Kyoto Encyclopedia of Genes and Genomes(KEGG) pathway enrichment of core targets by DAVID. Thereby, the possible molecular mechanism was unveiled. High-fat diet was combined with streptozotocin(STZ, injected into tail vein) for T2 DM rat modeling. Rats were classified into the normal group, model group, positive control group(metformin hydrochloride), AP high-dose, medium-dose, and low-dose groups. After 4 weeks of intragastric administration, serum fasting blood glucose(FBG), fasting insulin(FINS), aspartate aminotransferase(AST), alanine aminotransferase(ALT), triglyceride(TG), total cholesterol(TC), low-density lipoprotein cholesterol(LDL-C), high-density lipoprotein cholesterol(HDL-C), interleukin(IL)-6, and tumor necrosis factor(TNF)-α of rats in each group were measured. The expression of insulin receptor substrate-2(IRS-2), adenosine monophosphate-activated protein kinase(AMPK), phosphorylated AMPK(p-AMPK), glucose 6 phosphatase(G6 Pase), and phosphoenolpyruvate carboxy kinase(Pepck) in rat liver was detected by Western blot. A total of 131 core targets of the combination in the treatment of T2 DM were screened out, among which protein kinase B(AKT) 1, mitogen-activated protein kinase(MAPK) 1, TNF-α, IL-6 were more critical. KEGG enrichment analysis suggested that the combination decreased blood glucose mainly through PI3 K/AKT signaling pathway, AMPK signaling pathway, TNF signaling pathway, and MAPK signaling pathway. The levels of FBG and FINS were lower and the glycogen level was higher in the AP high-dose and medium-dose groups than in the model group. The levels of AST, ALT, TG, and LDL-C in the three AP groups and the level of TC in AP high-dose and low-dose groups decreased compared with those in the model group. Levels of IL-6 and TNF-α were lower in AP high-dose and medium-dose groups than in the model group. The expression of IRS-2, AMPK, and p-AMPK was higher and that of G6 Pase and Pepck was lower in AP high-dose group than in the model group. Thus, the combination had multi-component, multi-target, and multi-pathway characteristics in the treatment of T2 DM. It may regulate AMPK signaling pathway through IL-6 and TNF-α to influence insulin resistance, glycogen synthesis, gluconeogenesis, islet ß cell transport, and inflammatory response, thereby exerting therapeutic effect on T2 DM.

Dynamic content changes of cordycepin and adenosine and transcriptome in Cordyceps kyushuensis Kob at different fermentation stages.

20% (w/w) Astragali radix was added to the rice medium to cultivate C. kyushuensis Kob. The fermentation product was collected at mycelium stage, coloring stage, stromata-forming initial stage and fruiting body stage of C. kyushuensis Kob. The dynamic content changes of cordycepin and adenosine were detected at different fermentation stages. In the rice medium with Astragalus radix, both cordycepin and adenosine reached the highest content value on the 30th day of fermentation, 17.31 mg/g and 0.94 mg/g, respectively, which were 8.6 times and 2.0 times of that in rice medium at the same stage. At the same time, transcriptomics technology was used to analyze C. kyushuensis Kob during these four periods.

Astragalus root extract improved average daily gain, immunity, antioxidant status and ruminal microbiota of early weaned yak calves.

BACKGROUND: Early weaning in yak calves is being attempted to improve yak reproduction rate. However, this has to be done with caution because of the high mortality rate of calves due to the lack of nutrients and the harsh environmental conditions. Twenty-four weaned male yak calves were used in a 60 day feeding trial in which astragalus root extract (ARE) was supplemented. They were assigned randomly to one of four dietary treatments (n = six per treatment) that differed in ARE level: 0 g kg-1 (control), ARE0 ; 20 g kg-1 , ARE20 ; 50 g kg-1 , ARE50; and 80 g kg-1 dry matter intake (DMI), ARE80 . RESULTS: Final bodyweight and average daily gain (ADG) were significantly higher and the DMI/ADG ratio was significantly lower in calves with ARE supplementation than control (ARE0 ) calves. Ruminal concentrations of acetate and propionate and serum concentration of superoxide dismutase in ARE80 calves were higher than in the other groups and serum concentration of insulin was higher in ARE80 calves than in ARE20 calves. Serum immunoglobulin G (IgG) and interleukin-2 (IL-2) concentrations in ARE-fed calves were higher than in controls. Serum tumor necrosis factor (TNF-α) concentration was higher in ARE50 and ARE80 groups than ARE0 calves and serum interleukin-6 (IL-6) concentration was higher in ARE80 than in ARE0 calves. Serum immunoglobulin A (IgA), IgG and immunoglobulin M (IgM) concentrations increased with age in ARE-fed calves. ARE supplementation increased the abundance of fiber degrading bacteria. CONCLUSION: ARE at a dosage of 5% to 8% DMI can be supplemented to early weaned yak calves to improve growth performance, antioxidant capacity and immunity. © 2020 Society of Chemical Industry.

The auxin-inducible phosphate transporter AsPT5 mediates phosphate transport and is indispensable for arbuscule formation in Chinese milk vetch at moderately high phosphate supply.

Phosphorus is a macronutrient that is essential for plant survival. Most land plants have evolved the ability to form a mutualistic symbiosis with arbuscular mycorrhizal (AM) fungi, which enhances phosphate (Pi) acquisition. Modulation of Pi transporter systems is the master strategy used by mycorrhizal plants to adapt to ambient Pi concentrations. However, the specific functions of PHOSPHATE TRANSPORTER 1 (PHT1) genes, which are Pi transporters that are responsive to high Pi availability, are largely unknown. Here, we report that AsPT5, an Astragalus sinicus (Chinese milk vetch) member of the PHT1 gene family, is conserved across dicotyledons and is constitutively expressed in a broad range of tissues independently of Pi supply, but is remarkably induced by indole-3-acetic acid (auxin) treatment under moderately high Pi conditions. Subcellular localization experiments indicated that AsPT5 localizes to the plasma membrane of plant cells. Using reverse genetics, we showed that AsPT5 not only mediates Pi transport and remodels root system architecture but is also essential for arbuscule formation in A. sinicus under moderately high Pi concentrations. Overall, our study provides insight into the function of AsPT5 in Pi transport, AM development and the cross-talk between Pi nutrition and auxin signalling in mycorrhizal plants.

Mesorhizobium huakuii HtpG Interaction with nsLTP AsE246 Is Required for Symbiotic Nitrogen Fixation.

Plant nonspecific lipid transfer proteins (nsLTPs) are involved in a number of biological processes including root nodule symbiosis. However, the role of nsLTPs in legume-rhizobium symbiosis remains poorly understood, and no rhizobia proteins that interact with nsLTPs have been reported to date. In this study, we used a bacteria two-hybrid system and identified the high temperature protein G (HtpG) from Mesorhizobium huakuii that interacts with the nsLTP AsE246. The interaction between HtpG and AsE246 was confirmed by far-Western blotting and bimolecular fluorescence complementation. Our results indicated that the heat shock protein 90 (HSP90) domain of HtpG mediates the HtpG-AsE246 interaction. Immunofluorescence assay showed that HtpG was colocalized with AsE246 in infected nodule cells and symbiosome membranes. Expression of the htpG gene was relatively higher in young nodules and was highly expressed in the infection zones. Further investigation showed that htpG expression affects lipid abundance and profiles in root nodules and plays an essential role in nodule development and nitrogen fixation. Our findings provide further insights into the functional mechanisms behind the transport of symbiosome lipids via nsLTPs in root nodules.

Reduction mechanism of Cd accumulation in rice grain by Chinese milk vetch residue: Insight into microbial community.

Chinese milk vetch is an efficient approach to reduce Cd accumulation in rice, nevertheless, its reduction mechanism is not well understood. In this study, we investigated the rice grain Cd, soil properties and microbial community in a Cd-polluted paddy field amended with milk vetch residue (MV) or without (CK) during rice growth period. We found that milk vetch residue averagely decreased the Cd content in rice grain by 45%. Decrease of Cd in rice mainly attributed to the inhibition of Cd activation by milk vetch residue at heading stage probably by the formation of HA-Cd (Humic Acid) and CdS. Increased pH and organic matter (OM) promoted the reduction of available Cd. In addition, nonmetric multidimensional scaling (NMDS) analysis revealed that microbial community structure was significantly different between MV and CK treatment (r = 0.187, p = 0.002), and the core functions of differentially abundant genera were mainly associated with N-cycling, organic matter degradation and sulfate-reducing. The application of milk vetch residue increased the abundance of sulfate-reducing bacteria (SRB) by 8-112% during the rice growth period, which may involve in promoting the transformation of Cd to a more stably residual Cd (CdS). Canonical correspondence analysis (CCA) and mantel test analysis indicated that available K (p = 0.004) and available N (p = 0.005) were the key environmental factors of shaping the SRB. Altogether, changes in soil properties affected microbial structure and functional characteristics, especially the response of SRB in MV treatment would provide valuable insights into reducing the bioavailability of Cd in soil.

Benefit of an association of an antioxidative substrate and a traditional chinese medicine on telomere elongation.

Telomere shortening is involved in age-related disorders, such as cancer and cardiovascular diseases. Recently, telomerase re-activation strategies have been proposed to counteract telomere shortening and its consequences. Here, we investigated the benefit of dietary supplementation with a mix of S-adenosyl-methionine (SAMe) and a polysaccharide extract of Astragalus (APS) on telomere length of circulating lymphocytes of healthy volunteers. Blood lymphocytes of a cohort of 26 healthy volunteers who were administrated the mix of SAMe and APS in a food supplement for one year were collected. In vitro treatment of blood lymphocytes of healthy volunteers with the mix was also performed. A cohort of 150 healthy volunteers was used as a control. Telomere length was measured by Q-FISH. The micronucleus assay was performed to detect genotoxicity of the mix. The telomeres of circulating lymphocytes of the cohort of 26 donors supplemented with the mix were significantly longer than those of matched controls (p < 10-4). This elongation was essentially observed in the lymphocytes of older donors. Similarly, in vitro treatment of circulating lymphocytes with the mix significantly increased telomere length and decrease the proportion of cells with short telomeres. Here, we observed an increase in telomere length after in vivo and in vitro administration of a mix with SAMe and APS.  The benefit of dietary supplementation with this mix opens a new horizon for the battle against aging and could be used in the treatment of chronic age-related disorders.

Neuroprotective Effect of Astragalus Polysacharin on Streptozotocin (STZ)-Induced Diabetic Rats.

BACKGROUND In the recent years, there has been increasing interest in traditional Chinese medicine as a neuroprotective nutrient in the management of chronic neurodegenerative disease, such as diabetic cognitive decline. Astragalus polysacharin (APS), a Chinese herb extract, is a biologically active treatment for neurodegenerative diseases. Therefore, in the present study, we investigated the neuroprotective effects of APS (20 mg/kg) on diabetes-induced memory impairments in Sprague-Dawley (SD) rats and explored its underlying mechanisms of action. MATERIAL AND METHODS Thirty SD rats were randomly divided into a control group (CON group, n=10), a diabetic model (DM) group (n=10), and an APS group (n=10). We administered 55 mg/kg streptozotocin (STZ, Sigma) by intraperitoneal injection to induce a diabetic model. Food and water intake, body weight, and blood fasting plasma glucose (FPG) were measured. The Morris water maze test (MWM) was used to assess learning and memory ability, and we measured levels of N-methyl-D-aspartate receptor (NMDA), calcium/calmodulin-dependent protein kinase II (CaMKII), and cAMP response element-binding protein (CREB) in the hippocampus. RESULTS APS (20 mg/kg) administration decreased the rats' fasting plasma glucose (FPG) levels and body weight. APS (20 mg/kg) administration improved the cognitive performance of diabetes-induced rats in the Morris water maze test. APS (20 mg/kg) administration reduced the number of dead cells in the CA1 region of the hippocampus. Furthermore, APS (20 mg/kg) administration obviously upregulated the phosphorylation levels CREB, NMDA, and CaMK II. CONCLUSIONS These results suggest that APS has the neuroprotective effects, and it may be a candidate for treatment of neurodegenerative diseases such as diabetic cognitive impairment.

The Effects of Astragalus Polysaccharide on Bone Marrow-Derived Mesenchymal Stem Cell Proliferation and Morphology Induced by A549 Lung Cancer Cells.

BACKGROUND The tumor microenvironment in lung cancer plays an important role in tumor progression and metastasis. Bone marrow-derived mesenchymal stem cells (MSCs) co-cultured with A549 lung cancer cells show changes in morphology, increase cell proliferation, and cell migration. This study aimed to investigate the effects of Astragalus polysaccharide (APS), a traditional Chinese herbal medicine, on the changes induced in bone marrow-derived MSCs by A549 lung cancer cells in vitro. MATERIAL AND METHODS Bone marrow-derived MSCs were co-cultured with A549 cells (Co-BMSCs). Co-cultured bone marrow-derived MSCs and A549 cells treated with 50 µg/ml of APS (Co-BMSCs + APS) were compared with untreated Co-BMSCs. Cell proliferation was measured using the cell counting kit-8 (CCK-8) assay. Flow cytometry evaluated the cell cycle. Microarray assays for mRNA expression and Western blot for protein expression were used. RESULTS Compared with untreated Co-BMSCs, APS treatment of Co-BMSCs improved cell morphology, reduced cell proliferation, and inhibited cell cycle arrest. The mitogen-activated protein kinase (MAPK)/nuclear factor-kappa B (NF-kappaB) pathway, TP53, caspase-3, acetylated H4K5, acetylated H4K8, and acetylated H3K9 were involved in the regulatory process. CONCLUSIONS APS treatment reduced cell proliferation and morphological changes in bone marrow-derived MSCs that were co-cultured with A549 lung cancer cells in vitro.

Study on Hypoglycemic Effect of the Drug Pair of Astragalus Radix and Dioscoreae Rhizoma in T2DM Rats by Network Pharmacology and Metabonomics.

Type 2 diabetes mellitus (T2DM) is a metabolic disease accompanied by a series of diseases such as diabetic nephropathy. The drug pair (HS) of Astragalus Radix (HQ) and Dioscoreae Rhizoma (SY) was designed by Dr. Shi Jinmo to improve the treatment of T2DM. However, the exact mechanism involved requires further clarification. In this work, 1H-NMR-based metabonomics and network pharmacology were adopted. Metabolic profiling indicated that the metabolic perturbation was reduced after HS treatment. The results found 21 biomarkers. According to the network pharmacology, we found that the regulation of T2DM was primarily associated with 18 active compounds in HS. These active compounds mainly had an effect on 135 targets. Subsequently, combining network pharmacology and metabonomics, we found four target proteins, which indicated that HS has potential hypoglycemic effects through regulating monoamine oxidases B (MAOB), acetyl-CoA carboxylase 1 (ACACA), carbonic anhydrase 2 (CA2), and catalase (CAT). In conclusion, the result showed that these four targets might be the most relevant targets for the treatment of T2DM with HS. This study clarified the mechanism of HS in the treatment of T2DM and also confirmed the feasibility of combining metabonomics and network pharmacology to study the mechanisms of traditional Chinese medicine (TCM). In the future, this approach may be a potentially powerful tool to discovery active components of traditional Chinese medicines and elucidate their mechanisms.

[Short-term water changes response of saponin biosynthesis process in Astragalus membranceus].

The study is aimed to explore the effect of different water on the content of total saponins,astragaloside Ⅳ and gene expression in the growth of Astragalus membranceus. In this study, one-year-old A. membranaceus was used as the experimental material, by pot culture different water treatments were simulated at herbal garden in Jilin Agricultural University. The content of astragaloside Ⅳ was determined by HPLC and the total saponins by UV spectrophotometry. With 18 S RNA as a reference gene, fluorescent quantitative PCR was applied to analyze the eight key enzymes in astragalus saponin synthesis pathway AACT,HMGS,HMGR,IDI,FPS,SS,SE,CAS expression. With the decrease of soil water, the content of astragaloside Ⅳ in the root tissue of A. membranaceus showed an increasing trend, up to 1.46 mg·g~(-1). The total saponin content tended to increase, up to 6.80 mg·g~(-1). The results of relative expression of genes showed that the eight genes showed different effects at different water. With the change of soil water content, the amount of(AACT,IDI,SS) relative expression in drought stress group firstly increased and then decreased, then increased, and then decreased. The amount of(HMGS,HMGR,FPS) relative expression in drought stress group increased firstly and then decreased. The amount of(SE,CAS) relative expression in drought stress group increased firstly and then decreased, and continued to decrease after rehydration. The expression of key enzyme genes involved in the synthesis of astragaloside was influenced by each other, and the expression of key enzyme in roots showed a correlation with the content of astragaloside. Correlation analysis showed that there was a very significant positive correlation between HMGR gene and total saponins content in drought stress group and a significant negative correlation between content of CAS and total saponins. The contents of FPS,SE,CAS and astragaloside Ⅳ were very significantly and negative correlated. The relationship between other genes and quality was positive. Therefore, HMGR, FPS, SE and CAS genes have significant effects on the regulation of saponin content under water control. On the 15 th day after water regulation, the total amount of astragaloside and total saponins reached the highest value and could be harvested.

Astragalus Polysaccharide Protects Neurons and Stabilizes Mitochondrial in a Mouse Model of Parkinson Disease.

BACKGROUND Astragalus polysaccharides (APS) have a very good therapeutic effect in the treatment of neurodegenerative diseases and nerve injury disease. However, research on Parkinson disease (PD) treatment with APS is lacking. MATERIAL AND METHODS The present study was designed to explore the effects of APS on the protection of neurons and mitochondrial in a mouse model of PD using behavioral experiments, and observations of mitochondrial structure and transmembrane potential. RESULTS It was shown that APS could attenuate 1-methyl-4-pheyl-1,2,3,6-tetrahydropyridine (MPTP)-induced motor dysfunction (P<0.01), increase the proportion of TH-positive cells (P<0.01), reverse MPTP-induced mitochondrial structural damage, and reduce MPTP-induced high levels of reactive oxygen species (ROS) and increase MPTP-induced decrease in mitochondrial membrane potential. In addition, APS also decreased the bax/bcl2 ratio, and cytochrome-c and caspase-3 protein content (P<0.01) in substantia nigra in our mouse PD model. CONCLUSIONS APS provided a protective effect on neurons and mitochondrial in a mouse PD model.

Astragalus Polysaccharide Inhibits Ionizing Radiation-Induced Bystander Effects by Regulating MAPK/NF-kB Signaling Pathway in Bone Mesenchymal Stem Cells (BMSCs).

BACKGROUND This study investigated the effect of Astragalus polysaccharides (APS) on radiation-induced bystander effects (RIBE) in human bone mesenchymal stem cells (BMSCs) induced by irradiated A549 cells. MATERIAL AND METHODS A549 cells were irradiated with 2 Gy X-rays to obtain conditioned medium. BMSCs were incubated with the conditioned medium or APS. The levels of reactive oxygen species (ROS) and TGF-ß were detected by ELISA. Cell survival, genomic instability, and DNA damages were detected by CCK-8 assay, colony formation assay, the micronucleus test and immunofluorescence assay, respectively. The protein and phosphorylation protein expression of p38, c-Jun N-terminal kinase (JNK), extracellular regulated protein kinase (ERK1/2), P65, and cyclooxygenase-2 (COX-2) in bystander effect cells were detected by Western blot. RESULTS The expression of COX-2 and ROS increased following stimulation with conditioned medium; this effect was inhibited by pre-exposing the cells to APS. BMSCs growth and colony formation rate decreased following stimulation with conditioned medium; this effect was suppressed by pre-exposing the cells to APS. In addition, the micronucleus rate and 53BP1 foci number increased after treatment with conditioned medium; this increase in BMSCs was inhibited by APS. The levels of phosphorylated p38, JNK, ERK1/2, NF-κB P65, and COX-2 proteins were increased by conditioned medium but were decreased by pre-treatment with APS. CONCLUSIONS RIBE in BMSCs induced by the irradiated A549 was mediated by the ROS in the conditioned medium and might be related to MAPK/NF-κB signal pathways in BMSCs. APS may block RIBE through regulating the MAPK/NF-κB pathway.

Effects of Elevated CO2 on the Swainsonine Chemotypes of Astragalus lentiginosus and Astragalus mollissimus.

Rapid changes in the Earth's atmosphere and climate associated with human activity can have significant impacts on agriculture including livestock production. CO2 concentration has risen from the industrial revolution to the current time, and is expected to continue to rise. Climatic changes alter physiological processes, growth, and development in numerous plant species, potentially changing concentrations of plant secondary compounds. These physiological changes may influence plant population density, growth, fitness, and toxin concentrations and thus influence the risk of toxic plants to grazing livestock. Locoweeds, swainsonine-containing Astragalus species, are one group of plants that may be influenced by climate change. We evaluated how two different swainsonine-containing Astragalus species responded to elevated CO2 concentrations. Measurements of biomass, crude protein, water soluble carbohydrates and swainsonine concentrations were measured in two chemotypes (positive and negative for swainsonine) of each species after growth at CO2 levels near present day and at projected future concentrations. Biomass and water soluble carbohydrate concentrations responded positively while crude protein concentrations responded negatively to elevated CO2 in the two species. Swainsonine concentrations were not strongly affected by elevated CO2 in the two species. In the different chemotypes, biomass responded negatively and crude protein concentrations responded positively in the swainsonine-positive plants compared to the swainsonine-negative plants. Ultimately, changes in CO2 and endophyte status will likely alter multiple physiological responses in toxic plants such as locoweed, but it is difficult to predict how these changes will impact plant herbivore interactions.