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.

Integrated phenotypic, transcriptomics and metabolomics: growth status and metabolite accumulation pattern of medicinal materials at different harvest periods of Astragalus Membranaceus Mongholicus.

BACKGROUND: Astragalus membranaceus var. mongholicus (Astragalus), acknowledged as a pivotal "One Root of Medicine and Food", boasts dual applications in both culinary and medicinal domains. The growth and metabolite accumulation of medicinal roots during the harvest period is intricately regulated by a transcriptional regulatory network. One key challenge is to accurately pinpoint the harvest date during the transition from conventional yield content of medicinal materials to high and to identify the core regulators governing such a critical transition. To solve this problem, we performed a correlation analysis of phenotypic, transcriptome, and metabolome dynamics during the harvesting of Astragalus roots. RESULTS: First, our analysis identified stage-specific expression patterns for a significant proportion of the Astragalus root genes and unraveled the chronology of events that happen at the early and later stages of root harvest. Then, the results showed that different root developmental stages can be depicted by co-expressed genes of Astragalus. Moreover, we identified the key components and transcriptional regulation processes that determine root development during harvest. Furthermore, through correlating phenotypes, transcriptomes, and metabolomes at different harvesting periods, period D (Nov.6) was identified as the critical period of yield and flavonoid content increase, which is consistent with morphological and metabolic changes. In particular, we identified a flavonoid biosynthesis metabolite, isoliquiritigenin, as a core regulator of the synthesis of associated secondary metabolites in Astragalus. Further analyses and experiments showed that HMGCR, 4CL, CHS, and SQLE, along with its associated differentially expressed genes, induced conversion of metabolism processes, including the biosynthesis of isoflavones and triterpenoid saponins substances, thus leading to the transition to higher medicinal materials yield and active ingredient content. CONCLUSIONS: The findings of this work will clarify the differences in the biosynthetic mechanism of astragaloside IV and calycosin 7-O-ß-D-glucopyranoside accumulation between the four harvesting periods, which will guide the harvesting and production of Astragalus.

Astragalus membranaceus Extract Induces Apoptosis via Generation of Reactive Oxygen Species and Inhibition of Heat Shock Protein 27 and Androgen Receptor in Prostate Cancers.

Although Astragalus membranaceus is known to have anti-inflammatory, anti-obesity, and anti-oxidant properties, the underlying apoptotic mechanism of Astragalus membranaceus extract has never been elucidated in prostate cancer. In this paper, the apoptotic mechanism of a water extract from the dried root of Astragalus membranaceus (WAM) was investigated in prostate cancer cells in association with heat shock protein 27 (HSP27)/androgen receptor (AR) signaling. WAM increased cytotoxicity and the sub-G1 population, cleaved poly (ADP-ribose) polymerase (PARP) and cysteine aspartyl-specific protease 3 (caspase 3), and attenuated the expression of B-cell lymphoma 2 (Bcl-2) in LNCaP cells after 24 h of exposure. Consistently, WAM significantly increased the number of terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL)-positive LNCaP cells. WAM decreased the phosphorylation of HSP27 on Ser82 and inhibited the expression of the AR and prostate-specific antigen (PSA), along with reducing the nuclear translocation of p-HSP27 and the AR via the disturbed binding of p-HSP27 with the AR in LNCaP cells. WAM consistently inhibited the expression of the AR and PSA in dihydrotestosterone (DHT)-treated LNCaP cells. WAM also suppressed AR stability, both in the presence and absence of cycloheximide, in LNCaP cells. Taken together, these findings provide evidence that WAM induces apoptosis via the inhibition of HSP27/AR signaling in prostate cancer cells and is a potent anticancer candidate for prostate cancer treatment.

The effect of different LED wavelengths on the components and biosynthesis of isoflavonoid in sprout Astragalus membranaceus.

An artificial light source is the optimal element for studying the usability of the medicinal plant Astragalus membranaceus as a sprout vegetable. Based on artificial light source conditions, formononetin (FO) level was the highest (2.6 mg/L) in A. membranaceus exposed to white light emitting diode (LED) light, and calycosin (CA) level was the highest (3.09 mg/L) in the plant exposed to red LED light. According to the publicly available transcriptome data of LED-exposed sprout A. membranaceus LED, reference genes related to the content enhancement of FO, an isoflavone compound, and those related to the content enhancement of CA were selected. The expression patterns of these genes were assayed using qPCR. Among the genes related to FO enhancement, Gene-225190T showed the highest mRNA levels in cells of LED-white light-exposed sprout A. membranaceus; among the genes related to CA enhancement, Gene_042770T showed the highest expression under red LED light. Most genes related to the overall biosynthesis regulation of flavonoids of the upper concept of isoflavone were highly expressed in response to red LED light, and the transcriptional level of 4CL in response to red LED light was the highest. Based on these results, the artificial light sources that regulated the FO and CA contents in sprouts A. membranaceus were white and red LED lights, and the selected reference genes were capable of regulating isoflavone biosynthesis.

Dark septate endophyte inoculation enhances antioxidant activity in Astragalus membranaceus var. mongholicus under heat stress.

The influence of dark septate endophytic (DSE) on the antioxidant activity of Astragalus membranaceus var. mongholicus under heat stress was investigated. A. membranaceus plants, with or without DSE inoculation, were grown at 28°C for 8 weeks in a greenhouse and subsequently subjected to heat stress conditions (42°C) in an artificial climate chamber. DSE inoculation significantly decreased the malondialdehyde (MDA) content during the initial three days of heat stress. The activities of superoxide dismutase (SOD) and peroxidase (POD) of A. membranaceus leaves were significantly enhanced by DSE inoculation under heat stress, with SOD activities being 63-81% higher than in other treatments. The glutathione (GSH) and putrescine (Put) contents accumulated significantly on the third day under heat stress with DSE inoculation. Additionally, the contents of soluble sugars and proline (Pro) exhibited significant increases on the seventh day of heat stress and were 33-55% and 81-83% higher than in other treatments, respectively. Three-way ANOVA shows that DSE inoculation under heat stress exerted a significant impact on MDA. Multivariate linear regression and structural equality modelling (SEM) further show that the interaction among these antioxidants significantly decreased MDA content and maintained the normal function of cell membranes. In conclusion, DSE inoculation enhanced the heat tolerance of A. membranaceus by boosting its antioxidant capacity and reducing MDA production. This study highlights the potential of utilizing DSE as a strategy to enhance plant heat tolerance.

Simultaneous determination and pharmacokinetics study of three triterpenoid saponins in rat plasma by ultra-high-performance liquid chromatography tandem mass-spectrometry after oral administration of Astragalus Membranaceus leaf extract.

A selective and sensitive ultra-high-performance liquid chromatography-tandem mass spectrometry method was developed and validated for the determination of three triterpenoid saponins isolated from Astragalus membranaceus leaf extract. In this article, a method for simultaneous determination of Huangqiyenin A, Huangqiyenin E, and Huangqiyenin K was established for the first time. The method was successfully applied to the pharmacokinetic study of Astragalus membranaceus leaf extract after oral administration. Liquid-liquid extraction was applied to plasma sample preparation. Multiple reaction monitoring mode with an electrospray ion source in positive electrospray ionization was chosen to quantify the analytes. Chromatographic separation was performed on a Waters HSS T3 column, using gradient elution with a mobile phase composed of acetonitrile and 5 mM ammonium acetate/water. The pharmacokinetic results showed that all three compounds had the characteristics of rapid absorption-slow metabolism trend. The time of maximum plasma concentration of Huangqiyenin A is higher than Huangqiyenin E and Huangqiyenin K. And the maximum plasma concentration of Huangqiyenin A is higher as well. The pharmacokinetic results revealed the pharmacokinetic characteristics of the three analytes in rat plasma, which could provide a helpful reference for the further study of Astragalus membranaceus leaf extract.

Characterization and protein engineering of glycosyltransferases for the biosynthesis of diverse hepatoprotective cycloartane-type saponins in Astragalus membranaceus.

Although plant secondary metabolites are important source of new drugs, obtaining these compounds is challenging due to their high structural diversity and low abundance. The roots of Astragalus membranaceus are a popular herbal medicine worldwide. It contains a series of cycloartane-type saponins (astragalosides) as hepatoprotective and antivirus components. However, astragalosides exhibit complex sugar substitution patterns which hindered their purification and bioactivity investigation. In this work, glycosyltransferases (GT) from A. membranaceus were studied to synthesize structurally diverse astragalosides. Three new GTs, AmGT1/5 and AmGT9, were characterized as 3-O-glycosyltransferase and 25-O-glycosyltransferase of cycloastragenol respectively. AmGT1G146V/I variants were obtained as specific 3-O-xylosyltransferases by sequence alignment, molecular modelling and site-directed mutagenesis. A combinatorial synthesis system was established using AmGT1/5/9, AmGT1G146V/S and the reported AmGT8 and AmGT8A394F . The system allowed the synthesis of 13 astragalosides in Astragalus root with conversion rates from 22.6% to 98.7%, covering most of the sugar-substitution patterns for astragalosides. In addition, AmGT1 exhibited remarkable sugar donor promiscuity to use 10 different donors, and was used to synthesize three novel astragalosides and ginsenosides. Glycosylation remarkably improved the hepatoprotective and SARS-CoV-2 inhibition activities for triterpenoids. This is one of the first attempts to produce a series of herbal constituents via combinatorial synthesis. The results provided new biocatalytic tools for saponin biosynthesis.

Identification of oxidosqualene cyclases associated with saponin biosynthesis from Astragalus membranaceus reveals a conserved motif important for catalytic function.

INTRODUCTION: Triterpenoids and saponins have a broad range of pharmacological activities. Unlike most legumes which contain mainly oleanane-type scaffold, Astragalus membranaceus contains not only oleanane-type but also cycloartane-type saponins, for which the biosynthetic pathways are unknown. OBJECTIVES: This work aims to study the function and catalytic mechanism of oxidosqualene cyclases (OSCs), one of the most important enzymes in triterpenoid biosynthesis, in A. membranaceus. METHODS: Two OSC genes, AmOSC2 and AmOSC3, were cloned from A. membranaceus. Their functions were studied by heterologous expression in tobacco and yeast, together with in vivo transient expression and virus-induced gene silencing. Site-directed mutagenesis and molecular docking were used to explain the catalytic mechanism for the conserved motif. RESULTS: AmOSC2 is a ß-amyrin synthase which showed higher expression levels in underground parts. It is associated with the production of ß-amyrin and soyasaponins (oleanane-type) in vivo. AmOSC3 is a cycloartenol synthase expressed in both aerial and underground parts. It is related to the synthesis of astragalosides (cycloartane-type) in the roots, and to the synthesis of cycloartenol as a plant sterol precursor. From AmOSC2/3, conserved triad motifs VFM/VFN were discovered for ß-amyrin/cycloartenol synthases, respectively. The motif is a critical determinant of yield as proved by 10 variants from different OSCs, where the variant containing the conserved motif increased the yield by up to 12.8-fold. Molecular docking and mutagenesis revealed that Val, Phe and Met residues acted together to stabilize the substrate, and the cation-π interactions from Phe played the major role. CONCLUSION: The study provides insights into the biogenic origin of oleanane-type and cycloartane-type triterpenoids in Astragalus membranaceus. The conserved motif offers new opportunities for OSC engineering.

The combination of Astragalus membranaceus and ligustrazine mitigates cerebral ischemia-reperfusion injury via regulating NR2B-ERK/CREB signaling.

BACKGROUND AND PURPOSE: Cerebral ischemia-reperfusion (I/R) injury is a major factor underlying the high mortality and morbidity rates in stroke patients. Our previous study found that the combination of Astragalus membranaceus extract and ligustrazine (Ast+Lig) treatment could protect brain tissues against inflammation in rats with thrombolytic cerebral ischemia. Activation of N-methyl-D-aspartate receptors (NMDAR) is implicated in brain damage induced by cerebral I/R injury. METHODS: We used in vivo and in vitro models of cerebral I/R injury for middle cerebral artery occlusion/reperfusion in mice and oxygen-glucose deprivation/reoxygenation in primary rat cerebral cortical neurons to evaluate the protective effects of Ast+Lig on cerebral I/R injury, and whether the protective mechanism was related to the regulation of NMDAR-ERK/CREB signaling. RESULTS: Treatment with Ast+Lig, or MK-801 (an inhibitor of NMDAR) significantly ameliorated neurological deficits, decreased infarct volumes, suppressed neuronal damage and Ca2+ influx, and maintained the mitochondrial membrane potential in vivo and in vitro following cerebral I/R injury based on 2,3,5-triphenyl tetrazolium chloride staining, immunohistochemistry, and immunofluorescent staining. Furthermore, treatment with Ast+Lig evidently prevented the upregulation of NR2B, but not NR2A, in vivo and in vitro following cerebral I/R injury based on western blotting and reverse transcription-quantitative PCR analyses. Moreover, treatment with Ast+Lig significantly increased the phosphorylation of ERK and CREB, as well as increasing their mRNA expression levels in vivo and in vitro following cerebral I/R injury. CONCLUSIONS: The overall results thus suggest that the Ast+Lig combination conferred neuroprotective properties against cerebral I/R injury via regulation of the NR2B-ERK/CREB signaling pathway.

Multi-omics analysis reveals therapeutic effects of Bacillus subtilis-fermented Astragalus membranaceus in hyperuricemia via modulation of gut microbiota.

At present, uncovering how to preventandcontrol hyperuricemia has become an important public health issue. Fermented traditionalChinesemedicine has exhibited promising applications in the clinical management of hyperuricemia. In this study, we generated a hyperuricemic mouse model to explore the potent therapeutic ability of Bacillus subtilis-fermented Astragalus membranaceus (BFA) on this condition by multi-omics analysis. We found that the serum uric acid level was decreased in hyperuricemic mice after BFA treatment. BFA effectively attenuated renal inflammation and regulated the expression of urate transporters. Additionally, we found that BFA could increase the abundances of butyrate-producing bacteria, including Butyricimonas synergistica, Odoribacter splanchnicus, and Collinsella tanakaei, and probiotics, including Lactobacillus intestinalis and Bacillus mycoides, in hyperuricemic mice. Therefore, we believe that BFA has the potential to become a novel safe and valid functional food for addressing hyperuricemia.

A Protective Role of Phenylalanine Ammonia-Lyase from Astragalus membranaceus against Saline-Alkali Stress.

Phenylalanine ammonia-lyase (PAL, E.C.4.3.1.5) catalyzes the benzene propane metabolism and is the most extensively studied enzyme of the phenylpropanoid pathway. However, the role of PAL genes in Astragalus membranaceus, a non-model plant showing high capability toward abiotic stress, is less studied. Here, we cloned AmPAL and found that it encodes a protein that resides in the cytoplasmic membrane. The mRNA of AmPAL was strongly induced by NaCl or NaHCO3 treatment, especially in the root. Overexpressing AmPAL in Nicotiana tabacum resulted in higher PAL enzyme activities, lower levels of malondialdehyde (MDA), and better root elongation in the seedlings under stress treatment compared to the control plants. The protective role of AmPAL under saline-alkali stress was also observed in 30-day soil-grown plants, which showed higher levels of superoxide dismutase (SOD), proline, and chlorophyll compared to wild-type N. Tabacum. Collectively, we provide evidence that AmPAL is responsive to multiple abiotic stresses and that manipulating the expression of AmPAL can be used to increase the tolerance to adverse environmental factors in plants.

Biosynthetic mechanisms of isoflavone accumulation affected by different growth patterns in Astragalus mongholicus products.

BACKGROUND: At present, Astragalus mongholicus products on the market represent two growth patterns: imitative wild A. mongholicus (WAM) and cultivated A. mongholicus (CAM). The 6-year-old WAM (A6) and 2-year-old CAM (B2) products are often sold as commodities. This study aimed to explore the effects of the abovementioned growth patterns on the biosynthetic mechanisms of isoflavone accumulation in A. mongholicus products. RESULTS: In this paper, the content of calycosin-7-O-ß-D-glucoside in 6-year-old WAM (A6) was significantly higher than that in 2-year-old CAM (B2) based on high-performance liquid chromatography. Tissue anatomy indicated that A6 has developed phloem fibers, thickened secondary walls, and a more well-developed vascular system than B2. Thirteen differentially accumulated metabolites were found in A6 and B2 by UHPLC-ESI-Q-TOF-MS/MS, of which isoflavones were highly and significantly enriched in A6. By combining transcriptomics and metabolomics analysis, we found that the metabolomics profile was the same as the transcriptomics profile in both A6 and B2. In total, 11 novel isoflavone-related genes were isolated using BLAST and functional annotation through RNA-Seq and Iso-Seq. The results of integrated analysis, Short Time-series Expression Miner analysis, and Pearson correlation analysis showed that the regulation of four key enzymes, cinnamate 4-hydroxylase, 6-deoxychalcone synthase, chalcone reductase, and chalcone isomerase, led to the high accumulation of isoflavones in A6. In addition, AmUFGT (c778119) and AmUCGT (c303354) were predicted to be 7-O-glycosyltransferases by phylogenetic analysis; these genes catalyze formononetin and calycosin, respectively. CONCLUSIONS: The findings of this work will clarify the differences in the biosynthetic mechanism of isoflavone accumulation between A6 and B2, which will guide the cultivation of A. mongholicus.

Astragalus polysaccharide: a review of its immunomodulatory effect.

The Astragalus polysaccharide is an important bioactive component derived from the dry root of Astragalus membranaceus. This review aims to provide a comprehensive overview of the research progress on the immunomodulatory effect of Astragalus polysaccharide and provide valuable reference information. We review the immunomodulatory effect of Astragalus polysaccharide on central and peripheral immune organs, including bone marrow, thymus, lymph nodes, spleen, and mucosal tissues. Furthermore, the immunomodulatory effect of Astragalus polysaccharide on a variety of immune cells is summarized. Studies have shown that Astragalus polysaccharide can promote the activities of macrophages, natural killer cells, dendritic cells, T lymphocytes, B lymphocytes and microglia and induce the expression of a variety of cytokines and chemokines. The immunomodulatory effect of Astragalus polysaccharide makes it promising for the treatment of many diseases, including cancer, infection, type 1 diabetes, asthma, and autoimmune disease. Among them, the anticancer effect is the most prominent. In short, Astragalus polysaccharide is a valuable immunomodulatory medicine, but further high-quality studies are warranted to corroborate its clinical efficacy.

Effects of Astragalus membranaceus on systemic lupus erythematosus in a mouse model of pregnancy.

BACKGROUND: This study used astragalus membranaceus (AM) to treat systemic lupus erythematosus (SLE) model mice during pregnancy, aiming to explore the role of AM in Helper T cell 17 (Th17) differentiation and SLE during pregnancy. METHODS: We used lipopolysaccharide to constructed the SLE mouse model. AM decoction given by intragastric administration lasted from the eighth week of the mouse age until the mouse was killed. We estimated the messenger RNA  levels of IL-17a and Rorc, counted the Th17 cell number and examined the levels of cytokines including interleukin (IL)-12, tumor necrosis factor α, interferon gamma, IL-17A in mouse serum. Periodic acid-Schiff staining and renal glomerular/tubulointerstitial (TI) score were used to evaluate the progression of lupus nephritis (LN). RESULTS: AM treatment improved the conception rate and increased the number and average weight of fetuses in SLE mice. It significantly decreased the urinary albumin/creatinine ratios and reduced the glomerular scores and TI scores in the pregnant SLE mice. AM gavage significantly decreased the weight of spleen, mesenteric lymph node, total splenocytes and T cells, and the expression of proinflammatory factors. Furthermore, AM treatment reduced the ratio of Th17 cells and the expression levels of RORγt and IL-17A. CONCLUSION: AM significantly improved pregnancy outcomes and inhibited lupus nephritis during pregnancy in SLE mice.

Astragalus membranaceus Enhances Myotube Hypertrophy through PI3K-Mediated Akt/mTOR Signaling Phosphorylation.

Astragalus membranaceus (AM) is classified as a high-class traditional herbal medicine, which has strengthened vitality and multifunctional pharmacological activities, but limited empirical evidence is available to support its effects in muscular hypertrophy. It evokes skeletal muscle hypertrophy by increasing anabolic pathway, which is essential to prevent sarcopenia in elderly population. In this study, we examined the effects of AM on skeletal muscle hypertrophy by focusing on the molecular mechanism. We employed an in vitro model to investigate whether AM-treated skeletal muscle, as represented by myotube C2C12 cells, was hypertrophic, and to further investigate the efficacy of AM-activated phosphorylation of PI3K/Akt/mTOR signaling that must occur prior to myotube hypertrophy. The results showed that the myotubes formed larger multinucleated myotubes with increased diameter and thickness (1.16-fold relative to control group, p < 0.05). Administration of PI3K and mTOR inhibitors abolished AM-induced muscular hypertrophy. Moreover, AM-induced PI3K-mediated myotube hypertrophy was accompanied by the activation of Akt and mTOR signaling. We concluded that the AM is a nutritional activator to enhance muscular hypertrophy by increasing PI3K/Akt/mTOR signaling phosphorylation. As the AM is effective in myotube hypertrophy, AM and its derivatives may be promising candidates for ergogenic aid to prevent sarcopenia.

Differential Expression of Calycosin-7-O-ß-D-glucoside Biosynthesis Genes and Accumulation of Related Metabolites in Different Organs of Astragalus membranaceus Bge. var. mongholicus (Bge.) Hsiao Under Drought Stress.

Calycosin-7-O-ß-D-glycoside (CG), as a flavonoid, plays an important role in the abiotic stress response of Astragalus membranaceus Bge. var. mongholicus (Bge.) Hsiao (A. mongholicus). CG is also an active ingredient in A. mongholicus with high medicinal value. However, the response mechanism of the CG biosynthetic pathway of drought stress is not clear. In this research, drought stress was inflicted upon A. mongholicus, and the variations in flavonoid metabolites and the correlating gene expression in CG biosynthesis were studied in roots, stems, and leaves of A. mongholicus by UHPLC-MRM-MS/MS and qRT-PCR. Drought stress reduced the dry weight and increased the content of malondialdehyde (MDA) and proline. Drought was beneficial to the accumulation of L-phenylalanine and 4-coumaric acid in leaves and promoted the accumulation of all target compounds in the roots, except calycosin. Overexpression of AmIOMT was observed in the leaves, but the content of formononetin which is the product of isoflavone O-methyltransferase (IOMT) catalysis was higher in stems than in leaves. This research aims to further understand the acclimation of abiotic stress and the regulation mechanism of flavonoid accumulation in A. mongholicus.

Effect of aqueous extract of Astragalus membranaceus on behavioral cognition of rats living at high altitude.

OBJECTIVE: To investigate the effect of aqueous extract of Astragalus membranaceus on cognitive ability of rats living at high altitude. METHODS: Rats were exposed to a simulated highaltitude hypobaric hypoxia chamber. The behavior of rats was tested by eight-arm maze. The contents of malondialdehyde (MDA), glutathione (GSH), reactive oxygen species (ROS) and activity of total superoxide dismutase (T-SOD) in hippocampus were measured. The expressions of mammalian target of rapamycin (mTOR) and cleaved capase-3 in hippocampus were determined by reverse transcription-polymerase chain reaction and Western blot. RESULTS: The behavioral cognitive ability of the hypoxic control group was significantly lower than that of the normoxic control group. Under hypoxic environment, after the administration of aqueous extract of Astragalus membranaceus, the behavioral cognitive ability of rats was significantly improved. In hippocampal tissue, the content of MDA and ROS were significantly decreased, while the content of GSH and activity of T-SOD in hippocampus were significantly increased. The mRNA expression of mTOR and P70S6K and the protein expression of p-mTOR were significantly increased; the mRNA expression of 4E-binding protein 1 (4E-BP1) and the protein expression of phosphorylated-4E-BP1 (p-4EBP1) and cleaved capase-3 were significantly decreased. CONCLUSION: When the rats are exposed to high altitude hypoxia, the behavioral cognitive ability could be significantly reduced. Aqueous extract of Astragalus membranaceus can significantly improve cognitive function in rats under hypoxia. The potential mechanism is related to improving oxidative stress, reducing the accumulation of free radicals and metabolites, and activating mTOR signaling pathway.

Calycosin, a Common Dietary Isoflavonoid, Suppresses Melanogenesis through the Downregulation of PKA/CREB and p38 MAPK Signaling Pathways.

Calycosin, a bioactive isoflavonoid isolated from root extracts of Astragalus membranaceus, has been reported to inhibit melanogenesis, the mechanism of which remains undefined. In this study, we interrogated the mechanistic basis by which calycosin inhibits melanin production in two model systems, i.e., B16F10 melanoma cells and zebrafish embryos. Calycosin was effective in protecting B16F10 cells from α-melanocyte-stimulating hormone (α-MSH)-induced melanogenesis and tyrosinase activity. This anti-melanogenic effect was accompanied by decreased expression levels of microphthalmia-associated transcription factor (MITF), a key protein controlling melanin synthesis, and its target genes tyrosinase and tyrosinase-related protein-2 (TRP-2) in calycosin-treated cells. Mechanistically, we obtained the first evidence that calycosin-mediated MITF downregulation was attributable to its ability to block signaling pathways mediated by cAMP response element-binding protein (CREB) and p38 MAP kinase. The protein kinase A (PKA) inhibitor H-89 and p38 inhibitor SB203580 validated the premise that calycosin inhibits melanin synthesis and tyrosinase activity by regulating the PKA/CREB and p38 MAPK signaling pathways. Moreover, the in vivo anti-melanogenic efficacy of calycosin was manifested by its ability to suppress body pigmentation and tyrosinase activity in zebrafish embryos. Together, these data suggested the translational potential of calycosin to be developed as skin-lightening cosmeceuticals.

Early nutrition programming with Astragalus membranaceus polysaccharide: its effect on growth, carcasses, immunity, antioxidants, lipid profile and liver and kidney functions in broiler chickens.

This study aimed to determine the influence of in ovo injection of Astragalus membranaceus polysaccharide on growth, feed consumption, feed conversion ratio, carcasses, hematology, and blood metabolites in Cobb 500 chicks. At the 7th day of incubation, a total of 250 eggs were randomly divided into five groups with 5 replications of 10 eggs of each: negative control (normal eggs), positive control (0.5 mL saline), 1.5 mg Astragalus membranaceus, 3.0 mg Astragalus membranaceus, and 4.5 mg in ovo Astragalus membranaceus injection. Live body weight and gain were not significantly (linear and quadratic) affected by in ovo injection of Astragalus membranaceus. Kidney and liver functions were influenced by in ovo injection of Astragalus membranaceus polysaccharides in broiler chickens. Antioxidant enzymes were quadratically increased with Astragalus membranaceus polysaccharides, and the highest values achieved with 4.5 mg. The MDA concentration was linearly and quadratically decreased with in ovo injection of Astragalus membranaceus polysaccharides when compared to negative control. The highest values of IgG and IgM were achieved with 1.5 mg Astragalus membranaceus polysaccharides when compared to all other groups. In conclusion, our results indicate that in ovo injection of Astragalus membranaceus polysaccharides 1.5-4.5 mg in broiler eggs significantly improved serum ALT, AST, AP, creatinine enzymes, antioxidant activity, and immune function.

Major Chemical Compounds and Mineral Elements of Astragalus membranaceus Cultivated on the Qinghai-Tibet Plateau with Different Planting Densities.

Astragalus membranaceus is a well-known herb that is widely used in the food and pharmaceutical industries. However, its commercial development has been limited due to wild resource shortages. This study was conducted in 2018 and 2019 to assess the effect of planting density on the major chemical compounds and mineral elements and biomass yield of A. membranaceus. The biomass yield (7,700.956 kg) reached the maximum at M2 planting density in 2018. In 2019, astragaloside IV reached its maximum concentration (0.117 %) at M2 group, which was significantly different from the concentrations obtained at the other groups. Calycosin-7-O-ß-D-glucoside (0.062 %) reach its maximum concentration in 2019 at M5, but not significant with M2. The concentration of major chemical compounds among the five groups in 2018 and 2019 all conformed to the Chinese Pharmacopoeia standards. In 2018, the mineral elements (Al, Ba, Fe, Li and Mn) content was higher at M2 than other groups. However, a general decrease in the mineral elements content was observed at M2 group in 2019. Enrichment analysis demonstrated that the enrichment capacity was highest for phosphorus. In conclusion, according to the TOPSIS results, M2 planting density was recommended as the optimal application. For optimal economic benefits, A. membranaceus should be harvested when it is 2 years old.