Mechanism of Active Ingredients of Astragalus Membranacus in Treating Heart Failure： A Review / 中国实验方剂学杂志

Heart failure is one of the main cardiovascular system diseases at present， and it is a clinical syndrome caused by changes in cardiac structure and function， resulting in impaired ejection function or ventricular filling. Therefore， heart failure has become the most important cardiovascular disease in the 21st century. In recent years， the incidence of heart failure is increasing， and the survival rate of patients with heart failure is very low. Traditional Chinese medicine has rich experience in preventing and treating heart failure. With the modernization of traditional Chinese medicine， more and more attention has been paid to the research， development， and application of active ingredients in traditional Chinese medicine. Traditional Chinese medicine has unique advantages in improving the heart function of patients with heart failure by treating multiple targets and multiple pathways through syndrome differentiation. Astragalus membranacus， a traditional Chinese medicine， is a kind of medicine that benefits Qi and blood circulation and removes evil spirits. It has the functions of improving myocardial energy metabolism and hemodynamics， protecting myocardial muscle， and promoting angiogenesis. Astragalus membranaceus is often used to treat patients with heart failure， yielding remarkable results. In recent years， it has been found that astragaloside， Astragalus polysaccharide， quercetin， calyx isoflavones， and other main active ingredients of Astragalus membranacus can improve cardiac function and treat heart failure by inhibiting inflammatory response， myocardial apoptosis， and myocardial fibrosis. This paper reviewed the research progress of the action and mechanism of the active ingredients of Astragalus membranacus in the treatment of heart failure by studying relevant literature， with a view to providing a reference for its further research， development， and application in the prevention and treatment of heart failure.

Preparation and polarization activity research of astragalus polysaccharide-superparamagnetic iron oxide nanocomposite / 药学学报

Size and surface modification are the two key factors affecting the effect of macrophages polarization induced by superparamagnetic iron oxide nanoparticles (SPIONs). The smaller the particle size, the better the polarization effect of SPIONs. Besides, the reasonable SPIONs surface modification method can also be used to enhance the polarization effect. In this study, SPIONs was prepared by solvothermal method and optimized by Box-Benhnken center combination design and response surface method. Furthermore, astragalus polysaccharide-superparamagnetic iron oxide nanocomplex (APS-SPIONs) was successfully constructed by EDC/NHS esterification method. The structure of APS-SPIONs was confirmed by dynamic light scatter and infrared spectrometer, and the contents of iron and polysaccharide were characterized by spectrophotometry. The effect of APS-SPIONs on inducing mouse macrophages RAW264.7 polarization was investigated by flow cytometry. The RAW264.7 macrophages-HepG2 human hepatoma cancer cells Transwell co-culture system was established to investigate APS-SPIONs improve anti-tumor function of macrophages in vitro, and the proliferation activity of APS-SPIONs on RAW264.7 detected by cell counting kit-8 (CCK-8) method. The results showed that the average particle size and zeta potential of APS-SPIONs were (82.93 ± 1.47) nm and (-24.00 ± 0.47) mV. Polysaccharide and Fe content were 8.69% and 7.04%, respectively. APS-SPIONs effectively induced the polarization of RAW264.7 into M1 type in vitro, improving the anti-tumor ability of macrophages in a co-culture system, without effecting the proliferation of macrophages. Our study provides a drug development strategy and preliminary research results to educate macrophages and reshape the tumor immune microenvironment to achieve tumor-killing effects.

Modulating effects of Astragalus polysaccharide on immune disorders via gut microbiota and the TLR4/NF-κB pathway in rats with syndrome of dampness stagnancy due to spleen deficiency / 浙江大学学报（英文版）（B辑：生物医学和生物技术）

The syndrome of dampness stagnancy due to spleen deficiency (DSSD) is relatively common globally. Although the pathogenesis of DSSD remains unclear, evidence has suggested that the gut microbiota might play a significant role. Radix Astragali, used as both medicine and food, exerts the effects of tonifying spleen and qi. Astragalus polysaccharide (APS) comprises a macromolecule substance extracted from the dried root of Radix Astragali, which has many pharmacological functions. However, whether APS mitigates the immune disorders underlying the DSSD syndrome via regulating gut microbiota and the relevant mechanism remains unknown. Here, we used DSSD rats induced by high-fat and low-protein (HFLP) diet plus exhaustive swimming, and found that APS of moderate molecular weight increased the body weight gain and immune organ indexes, decreased the levels of interleukin-1β (IL-1β), IL-6, and endotoxin, and suppressed the Toll-like receptor 4/nuclear factor-‍κB (TLR4/NF-‍κB) pathway. Moreover, a total of 27 critical genera were significantly enriched according to the linear discriminant analysis effect size (LEfSe). APS increased the diversity of the gut microbiota and changed its composition, such as reducing the relative abundance of Pseudoflavonifractor and Paraprevotella, and increasing that of Parasutterella, Parabacteroides, Clostridium XIVb, Oscillibacter, Butyricicoccus, and Dorea. APS also elevated the contents of short-chain fatty acids (SCFAs). Furthermore, the correlation analysis indicated that 12 critical bacteria were related to the body weight gain and immune organ indexes. In general, our study demonstrated that APS ameliorated the immune disorders in DSSD rats via modulating their gut microbiota, especially for some bacteria involving immune and inflammatory response and SCFA production, as well as the TLR4/NF-κB pathway. This study provides an insight into the function of APS as a unique potential prebiotic through exerting systemic activities in treating DSSD.

Research progress of effective components of Astragalus on prevention and treatment of organ fibrosis / 药学学报

Astragalus, which was first documented in Shennong Bencao Jing, is the dried root of Astragalus membranaceus (Fisch.) Bge. or Astragalus membranaceus (Fisch.) Bge. var. mongholicus (Bge.) Hsiao. The active ingredients astragalus membranaceus saponins (AMS), astragalus polysaccharides (APS) and astragalus flavonoids (AFS) have pharmacological effects such as anti-tumor properties, lowering blood sugar, regulating lipid metabolism, cardiovascular protection, anti-oxidation, bone protection, anti-fibrosis, etc. Fibrosis affects almost all organs, particularly vital organs such as the lungs, liver, heart and kidneys. The primary pathological changes of fibrosis involve abnormal increase of myofibroblasts and excessive deposition of extracellular matrix (ECM) components, which lead to the formation of scar tissue, ultimately resulting in fibrosis and even functional loss or failure of organs, which seriously threatens human health and life. Recent, studies have shown that Astragalus membranaceus has a good therapetuic effect on organ fibrosis. This article reviews the current advances of Astragalus in the prevention and treatment of fibrosis of lungs, liver, heart, kidneys and other important organs.

Astragalus polysaccharides affects multidrug resistance gene 1 and P-glycoprotein 170 in adriamycin nephropathy rats via regulating microRNA-16/NF-κB axis / 中南大学学报(医学版)

OBJECTIVES@#Nephrotic syndrome is a common disease of the urinary system. The aim of this study is to explore the effect of astragalus polysaccharides (APS) on multidrug resistance gene 1 (MDR1) and P-glycoprotein 170 (P-gp170) in adriamycin nephropathy rats and the underlying mechanisms.@*METHODS@#A total of 72 male Wistar rats were divided into a control group, a model group, an APS low-dose group, an APS high-dose group, an APS+micro RNA (miR)-16 antagomir group and an APS+miR-16 antagomir control group, with 12 rats in each group. Urine protein (UP) was detected by urine analyzer, and serum cholesterol (CHOL), albumin (ALB), blood urea nitrogen (BUN), and creatinine (SCr) were detected by automatic biochemical analyzer; serum interleukin-6 (IL-6), IL-1β, tumor necrosis factor α (TNF-α) levels were detected by ELISA kit; the morphological changes of kidney tissues were observed by HE staining; the levels of miR-16 and MDR1 mRNA in kidney tissues were detected by real-time RT-PCR; the expression levels of NF-κB p65, p-NF-κB p65, and P-gp170 protein in kidney tissues were detected by Western blotting; and dual luciferase was used to verify the relationship between miR-16 and NF-κB.@*RESULTS@#The renal tissue structure of rats in the control group was normal without inflammatory cell infiltration. The renal glomeruli of rats in the model group were mildly congested, capillary stenosis or occlusion, and inflammatory cell infiltration was obvious. The rats in the low-dose and high-dose APS groups had no obvious glomerular congestion, the proliferation of mesangial cells was significantly reduced, and the inflammatory cells were reduced. Compared with the high-dose APS group and the APS+miR-16 antagomir control group, there were more severe renal tissue structure damages in the APS + miR-16 antagomir group. Compared with the control group, the levels of UP, CHOL, BUN, SCr, IL-6, IL-1β, TNF-α, and MDR1 mRNA, and the protein levels of p-NF-κB p65 and P-gp170 in the model group were significantly increased (all P<0.05); the levels of ALB and miR-16 were significantly decreased (both P<0.05). Compared with the model group, the levels of UP, CHOL, BUN, SCr, IL-6, IL-1β, TNF-α, and MDR1 mRNA, and the protein levels of pNF-κB p65 and P-gp170 in the low-dose and high-dose APS groups were significant decreased (all P<0.05); and the levels of ALB and miR-16 were significantly increased (both P<0.05). Compared with APS+miR-16 antagomir control group, the UP, CHOL, BUN, SCr, IL-6, IL-1β, and TNF-α levels, MDR1 mRNA, and the protein levels of p-NF-κB p65 and P-gp170 were significantly increased (all P<0.05). The levels of ALB and miR-16 were significantly decreased in the APS+miR-16 antagomir group compared with the APS+miR-16 antagomir control group (both P<0.05).@*CONCLUSIONS@#APS can regulate the miR-16/NF-κB signaling pathway, thereby affecting the levels of MDR1 and P-gp170, and reducing the inflammation in the kidney tissues in the adriamycin nephropathy rats.

Screening for anti-inflammatory components of Astragalus polysaccharide and metabolomics research based on molecular weight distribution / 药学学报

Molecular mass distribution of Astragalus polysaccharides is wide. Astragalus polysaccharides prepared by conventional water extraction and alcohol precipitation are mostly mixture of macromolecules. Although studies have shown that Astragalus polysaccharides have two-sided immunomodulation, the relationship between anti-inflammatory components and molecular mass distribution of Astragalus polysaccharides is not clear. Therefore, Astragalus polysaccharides were extracted by water extraction and alcohol precipitation. The relative molecular weight of them was determined by high performance gel permeation chromatography (HPGPC). Astragalus polysaccharides with different molecular weights were separated and prepared by membrane separation. RAW 264.7 cells were induced by lipopolysaccharide (LPS) to establish an inflammatory cell model in vitro and the anti-inflammatory polysaccharide were screened. The anti-inflammatory regulation mechanism of Astragalus polysaccharides was analyzed by the LC-MS/MS metabonomics technology. The results showed that APS was composed of APS-Ⅰ ( > 2 000 kDa) and APS-Ⅱ (10 kDa). APS-Ⅰ was composed of mannose, rhamnose, galacturonic acid, glucose, galactose, arabinose and the molar ratios of these monosaccharide of APS-I were 0.54∶0.26∶12.24∶17.24∶8.46∶1. APS-II was composed of rhamnose, galacturonic acid, glucose, galactose, arabinose and the molar ratios of these monosaccharide of APS-II were 0.26∶0.14∶24.04∶0.62∶1. APS-Ⅰ and APS-Ⅱ had no cell toxicity to RAW 264.7 macrophage in the range of 0-100 μg·mL-1. Compared with the model group, APS-I at a concentration of 0-100 μg·mL-1could significantly inhibit the secretion of NO and TNF-α by RAW 264.7, and can significantly promote the secretion of IL-10. APS-I had better anti-inflammatory activity than APS-II in vitro. The metabolomics results showed that 32 different metabolites were found between the model group and blank group; APS-I group can significantly callback 18 different metabolites; mainly related to arginine biosynthesis, arginine and proline metabolism, pyrimidine metabolism, citric acid cycle (TCA cycle), cysteine and methionine acid metabolism, tryptophan metabolism. This study found that APS-I had better anti-inflammatory activity than APS-II in vitro, and its mechanism may be closely related to amino acid metabolism and energy metabolism, which indicated the direction for further clarifying the pharmacodynamic material basis of Astragalus polysaccharides.

Effect Enhancing and Toxicity-reducing Activities of Astragalus Polysaccharide Injection on U14 Cervical Cancer in Model Mice Receiving X-Ray Treatment / 中国实验方剂学杂志

ObjectiveTo study the effect-enhancing and toxicity-reducing activities of astragalus polysaccharide injection （APS） on U14 cervical cancer in model mice receiving X-ray treatment. MethodU14 mouse cervical cancer cells were cultured in vitro and injected into the right forelimb armpit of Kunming mice for constructing a subcutaneous tumor-bearing model of cervical cancer. The tumor-bearing mice were randomly divided into the model group， X-ray intervention（IR， 6 Gy） group， APS （10 mL·kg-1·d-1） group， and IR + APS group. Following the observation of the state， body mass， and food intake of mice in each group， the volume of the tumor was measured. The tumor cell cycle and apoptosis were determined by flow cytometry. The protein and mRNA expression levels of apoptosis-related proteins p53， B-cell lymphoma-2 （Bcl-2）， Bcl-2-associated X protein （Bax）， and cleaved cysteine-dependent aspartate-directed protease-3 （Caspase-3） in tumor tissues were assayed by Western blot and real-time fluorescence quantitative polymerase chain reaction （Real-time PCR）. ResultThe comparison with the model group showed that mice in the IR group had poor mental status and reduced mobility. The IR group and IR + APS group exhibited reduced food intake and body mass since the 8th d （P<0.05， P<0.01） and narrowed tumor volume since the 9th d （P<0.01）. In the IR group， the proportion of cells in the G1 phase was increased， while the proportion of those in the S phase was decreased （P<0.01）. In the IR + APS group， the proportion of cells in the G1 phase rose， whereas the proportion of those in the G2 and S phases cells declined （P<0.05， P<0.01）. The apoptotic rates in both the IR group and IR + APS group were elevated significantly （P<0.01）. Compared with the model group， the IR group and IR + APS group displayed up-regulated cleaved Caspase-3 and Bax protein and mRNA expression in tumor tissues， but down-regulated Bcl-2 and p53 protein and mRNA expression （P<0.05， P<0.01）. Compared with the IR group， the mice in the IR + APS group had better mobility and hair， normal body mass， and increased food intake （P<0.05）. The tumor volume in the IR + APS group was reduced （P<0.05）. The proportion of cells in the G2 phase was reduced， but the proportion of those in the S phase was raised （P<0.05）. The apoptosis rate was increased （P<0.05）. The apoptosis-related protein Bax protein expression in the tumor tissue was up-regulated， while the protein expression levels of Bcl-2 and p53 were down-regulated （P<0.05， P<0.01）. ConclusionAPS maintains the life state of U14 cervical cancer model mice treated with X-ray and promotes tumor cell apoptosis， thus enhancing the efficiency and reducing toxicity.

Effect and Mechanism of Lily Polysaccharide Combined with Astragalus Polysaccharide on Depressive Behavior in Chronic Stress Mice / 中国实验方剂学杂志

ObjectiveTo explore the mechanism of antidepressant effect of lily polysaccharide （LLP）and astragalus polysaccharide（APS）. MethodSixty KM mice were randomly divided into blank group， model group， fluoxetine hydrochloride （8 mg·kg-1）group， LLP （0.2 g·kg-1）group， APS （0.2 g·kg-1）group and polysaccharide combination （LLP+APS，0.1 g·kg-1+0.1 g·kg-1）group， with 10 mice in each group. Except the blank group， the other groups were given chronic unpredictable mild stress （CUMS） induced mouse depression model. On the 29th day of modeling，fluoxetine hydrochloride group was given corresponding dose of fluoxetine hydrochloride， and polysaccharide groups were given corresponding drug. The depressive behavior of mice was evaluated by behavioral indexes such as body mass change， open field test. The morphological changes of hippocampal CA1 neurons were observed by Nissl staining. The contents of 5-hydroxytryptamine （5-HT）， adrenocorticotropic hormone （ACTH）， and corticosterone （CORT）， in brain tissue and plasma were measured by enzyme-linked immunosorbent assay （ELISA）. Western blot was used to detect the expression levels of related proteins in adenylate cyclase/cyclic adenylate phosphate/protein kinase A （AC/cAMP/PKA） signal pathway. ResultCompared with the blank group， mice in the model group gained weight slowly， the total distance， central distance and sugar water preference rate decreased significantly （P<0.01）， the depressive behavior was significant， the hippocampal neurons were seriously damaged， the content of 5-HT decreased （P<0.01）， the contents of ACTH and CORT increased significantly （P<0.01）， adenylate cyclase 6（ADCY6）， PKA and cAMP response element binding protein-1 （CREB-1） and brain-derived neurotrophic factor （BDNF） protein expression decreased significantly （P<0.01）. Compared with the model group， depressive behavior of mice in LLP group， APS group and LLP+APS group was significantly improved （P<0.01）. The antidepressant effect of LLP+APS was better than that of LLP and APS. Each administration group could alleviate the damage of hippocampal neurons in varying degrees， significantly increase the content of 5-HT in brain tissue （P<0.01）， and reduce the levels of ACTH and CORT in plasma （P<0.05）. The protein levels of ADCY6， PKA， CREB-1 and BDNF were significantly increased （P<0.05）. ConclusionThe antidepressant effect of LLP+APS is significantly enhanced and has a synergistic effect. The mechanism may be closely related to affecting the content of neurotransmitters， inhibiting HPA axis activity and activating AC/cAMP/PKA signal transduction pathway.

Astragalus Polysaccharides Improve Cisplatin Resistance by Inhibiting EMT of Lung Adenocarcinoma A549/DDP Cells Transplanted into Nude Mice / 中国实验方剂学杂志

ObjectiveTo investigate the inhibitory effect of Astragalus polysaccharide （APS） on epithelial-mesenchymal transition （EMT） induced by transforming growth factor-β1 （TGF-β1） in cisplatin （DDP）-resistant lung adenocarcinoma cell line A549/DDP cells transplanted into nude mice and the molecular mechanism in improving DDP resistance. MethodBALB/c nude mice were randomly divided into a blank group， a model group， a DDP group， and a combination group （APS combined with DDP）. A549/DDP cells were infected with TGF-β1-overexpressed lentiviral vector and the negative control. The infected cells were inoculated subcutaneously in nude mice. The A549/DDP cells with TGF-β1 gene overexpression were inoculated into all groups except the control group with negative TGF-β1 gene overexpression. The drug intervention was performed eight days after cell inoculation. The mice in the combination group received intragastric administration of APS （0.3 g·kg-1·d-1） and intraperitoneal injection of cisplatin （0.003 5 g·kg-1）， and those in the cisplatin group received intraperitoneal injection of cisplatin （0.003 5 g·kg-1）. After 32 days of cell inoculation， the nude mice were killed and the tumor tissues and lungs were collected. The tumor weight was recorded and the inhibition rate was calculated. The number of metastatic nodules of the lung tumor on the whole slide was counted under the microscope. Immunohistochemistry， Western blot， and real-time fluorescence-based quantitative polymerase chain reaction （Real-time PCR） were used to detect the protein and gene expression of EMT molecular markers α-catenin and N-cadherin， and tumor drug resistance markers human lung resistance protein （LRP）， multidrug resistance-associated protein （MRP）， and P-glycoprotein （P-gp） in the transplanted tumor. ResultCompared with the blank group， the model group showed increased tumor weight and metastatic nodules of the lung tumor （P<0.05）， decreased protein and mRNA expression of α-catenin （P<0.05）， and elevated protein and mRNA expression of N-cadherin， LRP， MRP， and P-gp （P<0.05）. Compared with the model group and the cisplatin group， the combination group showed reduced tumor weight and metastatic nodules of the lung tumor （P<0.05）， increased protein and mRNA expression of α-catenin （P<0.05）， and decreased protein and mRNA expression of N-cadherin， LRP， MRP， and P-gp （P<0.05）. ConclusionAPS can inhibit the growth and metastasis of the transplanted tumor of lung adenocarcinoma and improve cisplatin resistance， which may be related to the inhibition of EMT of tumor cells.

Effect of Astragalus Polysaccharide on EMT of A549/DDP Lung Adenocarcinoma Xenograft： An Exploration Based on PI3K/Akt Signaling Pathway / 中国实验方剂学杂志

Objective:To investigate the effect of Astragalus polysaccharide （APS） on transforming growth factor-<italic>β</italic>₁ （TGF-<italic>β</italic>₁）-induced epithelial mesenchymal transition （EMT） of A549/DDP lung adenocarcinoma xenograft and its potential molecular mechanism. Method:BALB/c nude mice were randomly divided into the non-loading group （A549/DDP cells not loaded with TGF-<italic>β</italic>₁）， model group， cisplatin group， and combined group （A549/DDP cells overexpressing TGF-<italic>β</italic>₁）. Mice in the combined group were treated with intragastric administration of APS （0.3 g·kg^-1·d^-1） and intraperitoneal injection of cisplatin （0.003 5 g·kg^-1）， while those in the cisplatin group only received intraperitoneal injection of cisplatin （0.003 5 g·kg^-1）. After drug intervention， the nude mice were sacrificed and the xenograft and lung were harvested， followed by the weighing of tumor and the calculation of the inhibition rate. The number of tumors metastasizing to the lung was counted under the microscope. The pathological features of tumors and their metastasis to the lung tumor were observed by hematoxylin-eosin （HE） staining. The protein and mRNA expression levels of EMT molecular markers E-cadherin， Vimentin， <italic>α</italic>-smooth muscle actin （<italic>α</italic>-SMA）， and phosphatidylinositol 3-kinase/protein kinase B （PI3K/Akt） in the xenograft were detected by immunohistochemistry， Western blot， and Real-time polymerase chain reaction （Real-time PCR）. Result:Compared with the non-loading group， the model group exhibited increased tumor weight and pulmonary metastatic nodules （<italic>P</italic><0.05）， sparse tumor cell junctions， long spindle cells， massive metastatic nodules in the lung， down-regulated E-cadherin protein and mRNA expression， and up-regulated Vimentin and <italic>α</italic>-SMA protein and mRNA expression and p-PI3K and p-Akt protein expression （<italic>P</italic><0.05）. Compared with the model group and cisplatin group， the combined group displayed decreased tumor weight and pulmonary metastatic nodules （<italic>P<</italic>0.05）， tight tumor cell junctions， round or oval cells， no obvious lung metastasis， up-regulated E-cadherin protein and mRNA expression （<italic>P</italic><0.05）， and down-regulated Vimentin and <italic>α</italic>-SMA protein and mRNA expression （<italic>P</italic><0.05） and p-PI3K and p-Akt protein expression （<italic>P</italic><0.05）. There was no significant difference in PI3K or Akt protein expression among groups. Conclusion:APS has a certain inhibitory effect against EMT in lung adenocarcinoma A549/DDP cells， which may be related to the inhibition of activated PI3K/Akt protein expression.

Establishment of enzymatic hydrolysis method of active Astragalus polysaccharides APS-Ⅱ and study on the immune activity of oligosaccharides after its degradation / 药学学报

italic>Astragalus polysaccharides are the main immunomodulatory substances in Astragali Radix. The structure of polysaccharides is difficult to accurately determine, which limits the in-depth study of the molecular mechanism of Astragalus polysaccharides in vivo. "Polysaccharide receptor theory" believes that there are one or more oligosaccharide fragment "active centers" in immunologically active polysaccharide molecules. Therefore, the degradation of Astragalus polysaccharides into oligosaccharides and the study of the active centers of polysaccharides at the oligosaccharide level provide new ideas in the study of the structure and mechanism of Astragalus polysaccharides. This article adopts endo-α-1,4-glucanase enzymatic hydrolysis, and determines the best degradation conditions through single factor test and orthogonal test to degrade the immunologically active polysaccharide APS-Ⅱ (10 kDa component) into oligomers with different degrees of polymerization. Then through the preparation of polyacrylamide gel chromatography and specific immune and non-specific immune cell tests, the immune activity screening of different oligosaccharide components is carried out. The animal welfare and the experimental process in this study follow the requirements of the Animal Ethics Committee of Shanxi University. The results showed that compared with the immunologically active polysaccharide APS-Ⅱ, different oligosaccharide components have obvious differences in different immunological activities. This paper studies the different immunological activities of Astragalus polysaccharides at the level of oligosaccharides, laying a foundation for further elucidating the structure and function of Astragalus polysaccharides, enriching the theory of polysaccharide receptors, and providing new ideas for the development of Astragalus polysaccharides.

Comparative Study on the Protective Effects of Different Effective Components of Astragali Radix against Ionizing Radiation-induced BMSCs DNA Damage / 中国药房

OBJECTIVE：To compare the protective effects of different effective components of Astragali radix against DNA damage of human bone marrow mesenchymal stem cells （BMSCs）induced by ionizing radiation. METHODS ：2 Gy X-rays were used to directly irradiate BMSCs to establish a radiation model. CCK- 8 method was used to detect the effects of different mass concentrations（25，50，75，100 μg/mL）of astragalus polysaccharide ，astragalus saponin and astragalus flavonoids for 1 day before radiation + 1 to 5 days after radiation on the proliferation of BMSCs. The dose concentration and the duration of intervention after radiation were selected. The irradiated BMSCs were divided into radiation group ，astragalus polysaccharide group ，astragalus saponin group and astragalus flavonoids group. The last three groups were treated with appropriate dosage of corresponding drugs before and 2 days after radiation ，and a blank groupwas set for comparison. Cytoplasmic division arrest qq.com micronucleus method was used to detect micronucleus cell rate and cell micronucleus rate after appropriate time of was used to detect th e number of 53BP1 foci in cells after appropriare time of intervention following radiation ；the number of 53BP1 foci were compared among different time points （0.5，2，12，24 h）. RESULTS ：Compared with blank group ，OD values of BMSCs were decreased significantly in radiation group （P＜0.05 or P＜0.01）. Compared with radiation group ，the OD values of BMSCs were significantly increased when 50 μ g/mL astragalus polysaccharide，astragalus saponin and astragalus flavonoids continuously intervened radiation for 2-3 days，there was significant difference in other groups at some time point （P＜0.05 or P＜ 0.01）. After consideration ，drug concentration was determined to be 50 μg/mL，and the continuous intervention time was 2 days after radiation. Compared with blank group ，the micronucleus cell rate and cell micronucleus rate of radiation group ，astragalus polysaccharide group ，astragalus saponin group and astragalus flavonoids group increased significantly ，and the number of 53BP1 focus cluster in radiation group and astragalus polysaccharide group increased significantly （P＜0.01）. Compared with radiation group and astragalus flavonoids group ，the micronucleus cell rate ，cell micronucleus rate and the number of 53BP1 focus cluster （continued intervention for 0.5，2，12 h）in the astragalus polysaccharide group and astragalus saponin group were significantly reduced，and the micronucleus cell rate and cell micronucleus rate in the astragalus polysaccharide group were significantly lower than astragalus saponin group （P＜0.05）. 53BP1 focus cluster could not be detected 24 h later （P＜0.05）. CONCLUSIONS ： Astragalus polysaccharide and astragalus saponin both have protective effects on BMSCs DNA damage induced by radiation ，and the protective effect of astragalus polysaccharide is better than that of astragalus saponin ；astragalus flavonoids has no protective effect on radiation-induced DNA damage.

Study on active constituents of Astragalus polysaccharides for injection to regulate metabolism of macrophages based on LC-MS metabolomics / 中草药

Objective: To study the phagocytosis of mouse monocyte-macrophage Raw 264.7 by the active constituents of Astragalus polysaccharides for injection based on LC-MS metabolomics. Methods: Neutral red method was used to detect the phagocytosis of Raw 264.7 by different molecular weight fractions of Astragalus polysaccharides for injection. The active components were selected, and the cell culture supernatant and cell lysate were analyzed by LC-MS. Combined with multivariate statistical analysis and metabolism pathway analysis, the mechanism of action was studied. Results: The small molecular weight fraction of Astragalus polysaccharides for injection significantly promoted the phagocytosis of Raw 264.7 at a concentration of 30 μg/mL. Compared with the control group, after the active part of the injection of Astragalus polysaccharides was applied to Raw 264.7, 41 differential metabolites were found in and out of the cells, mainly related to amino acid metabolism, energy metabolism and antioxidant effects. Conclusion: The small molecular weight fraction of Astragalus polysaccharides for injection can increase the phagocytosis of Raw 264.7, and its mechanism may be closely related to amino acid metabolism, energy metabolism and antioxidant effects.

Comparative study on different areas of Astragali Radix based on oligosaccharides characteristic map and immunological activity evaluation of partial acid hydrolyzed / 中草药

Objective: The characteristic maps and immunological activities of some acid hydrolysates of polysaccharides from Astragali Radix (APS) from different areas were compared. The quality evaluation method of Astragali Radix with oligosaccharide mixture as quality control index was established. Methods: In this study, the polysaccharides from the traditional Chinese medicine Astragali Radix were used as the research object. Firstly, the optimal partial acid hydrolysis conditions were selected by orthogonal test. The polysaccharide was hydrolyzed into oligosaccharides for analysis. The characteristic map of Astragalus oligosaccharides based on partial acid hydrolysis-hydrophilic interaction chromatography was established. Multivariate statistical analysis was performed on the data by SIMCA software to distinguish Mongolian Astragalus from different areas. The partial acid hydrolysis products of Astragalus polysaccharides were characterized by hydrophilic interaction chromatography and mass spectrometry, and the activity was evaluated by mouse peritoneal macrophage phagocytosis neutral red experiment. Results: The optimal hydrolysis conditions obtained by orthogonal experiment were temperature 90 ℃, trifluoroacetic acid concentration 1 mol/L, hydrolysis time 1 h. Under this condition, Astragalus polysaccharide was hydrolyzed into characteristic oligosaccharide fragments, and the method is reproducible. The characteristic map of Astragalus oligosaccharides based on partial acid hydrolysis-hydrophilic interaction chromatography showed that the maps of the same Astragalus oligosaccharides had good consistency, and the maps of different Astragalus oligosaccharides were quite different. PCA showed that three different kinds of Mongolian Astragalus can be distinguished. It was found by mass spectrometry that the extracted Astragalus polysaccharides were mainly 1→4 linear glucan, and gluco-oligosac-charides with the degrees of polymerization 3—8 were obtained after partial acid hydrolysis. The partial acid hydrolysate of the wild Astragalus polysaccharides from Shanxi Hunyuan had higher ability to enhance the phagocytic activity of macrophages than the transplanted Astragalus and higher than the unhydrolyzed total astragalus polysaccharide. Conclusion: This study showed that the characteristics of Astragalus polysaccharides based on partial acid hydrolysis-hydrophilic interaction chromatography and the effects on cellular immune function can be used to evaluate the quality of Astragali Radix in different habitats and different planting methods, and it is also an important supplement to the quality evaluation method of Astragali Radix. At the same time, it has a certain exemplary role in the characterization of other Chinese materia medica polysaccharides.

Protective Effect of Astragulus Polysaccharide on Chromosome Damage in Human BMSCs Exposed to Formaldehyde / 中国实验方剂学杂志

Objective::To study the protective effect of astragalus polysaccharide (APS) on micronucleus and sister chromatid exchange (SCE) in human bone marrow mesenchyml stem cell (BMSCs) exposed to formaldehyde, in order to initially explore the potential mechanism. Method::BMSCs were cultured in vitro, cells were randomly divided into five groups: control group, formaldehyde group, and APS 40, 100, 400 mg·L-1 groups. BMSCs were infected with 120 μmol·L-1 formaldehyde, meanwhile, APS 40, 100, 400 mg·L-1 groups were co-cultured with 40, 100, 400 mg·L-1 APS. Cell morphology was observed by inverted phase contrast microscope, micronucleus were detected by micronucleus test, SCE was detected by SCE test, and mRNA and protein expressions of proliferating cell nuclear antigen (PCNA), xeroderma pigmentosum B, D, F, G (XPB, XPD, XPF, XPG) were detected by quantitative real-time fluorescence polymerase chain reaction(Real-time PCR)and Western blot. Result::Compared with control group, cell counts decreased, and cell morphology of BMSCs in formaldehyde group significantly changed, they were all recovered gradually in 40, 100, 400 mg·L-1 APS groups. Compared with control group, the micronucleus and SCE increased significantly (P<0.01), PCNA mRNA and protein expressions down-regulated significantly (P<0.05), while XPB, XPD, XPF, XPG mRNA and protein expressions up-regulated significantly (P<0.05, P<0.01). Compared with formaldehyde group, BMSCs were treated with APS at 40, 100, 400 mg·L-1, micronucleus and SCE decreased significantly (P<0.01), and mRNA and protein expressions of PCNA, XPB, XPD, XPF and XPG up-regulated significantly (P<0.05, P<0.01). Among them, the 100 mg·L-1 APS group had the most obvious effect. Conclusion::APS can protect formaldehyde-induced BMSCs micronucleus and SCE, especially 100 mg·L-1 APS has the most obvious effect. The mechanism may be associated with the up-regulation of expressions of PCNA, XPB, XPD, XPF and XPG in the nucleotide exicision repair pathway (NER), which promoted the damage repair.

Molecular mechanism of astragalus polysaccharide in alleviating insulin resistance in HepG2 cells / 预防医学

Objective@#To clarify the effect of astragalus polysaccharide （AP） on insulin resistance model of HepG2 cells induced by hyperinsulinemia and its underlying molecular mechanism in lipid metabolism and oxidative stress.@*Methods@#HepG2 cells were divided into three groups： the control group was treated without any intervention; the model group was treated with 200 μL cell culture medium containing 10-6 mol/L insulin for 48 hours to build an insulin resistance model; the AP group was treated with optimal concentration of AP based on an insulin resistance model. After 24 hours, the concentration of H2O2 and the expression of PPARγ in each group were detected. @*Results@#AP could improve the survival rate of insulin-resistant HepG2 cells in a dose-dependent manner. The highest survival rate of the cells was （118.26±1.17）% with 10 μM AP. The concentration of H2O2 in the AP group was （0.82±0.09） μM, which was lower than （1.30±0.16） μM in the model group （P<0.05）, but was close to （0.78±0.09） μM in the control group （P>0.05）. The relative mRNA expression of PPARγ in the AP group was 0.96±0.04, which was higher than 0.51±0.05 in the model group （P<0.05）, but was close to 1.00±0.11 in the control group （P>0.05）.@*Conclusions@#In the insulin resistance model in vitro, AP can significantly increase the cell survival rate, reduce intracellular H2O2 concentration, and promote the expression of PPARγ. The mechanism may be related to lipid metabolism.

Effects of astragalus polysaccharide on radiation-induced adipogenic differentiation of bonemarrow mesenchymal stem cells / 西安交通大学学报(医学版)

Objective: To study the effects of astragalus polysaccharide (APS) on adipogenic differentiation of human bone marrow mesenchymal stem cells (BMSCs) irradiated by X-ray. Methods: Human bone marrow mesenchymal stem cells irradiated by 2 Gy X-ray were intervened with 50 μg/mL of APS. The number and size of lipid droplets of stem cells were observed by oil red O staining and the expressions of CEBPα and PPAR-γ were detected by Western blot after induction with special medium. Results: The number of adipocytes differentiated from bone marrow mesenchymal stem cells decreased, and the area of lipid droplets in adipocytes decreased significantly after irradiation (P<0.05). The area of lipid droplets in the bone marrow mesenchymal stem cells treated with APS in advance increased compared with that in radiation alone (P<0.05). Similarly, the expressions of PPAR-γ and recombinant human CCAAT enhancer binding protein (CEBPα ) decreased after X-ray irradiation, while the expressions of PPAR-γ and CEBPα increased after the intervention of APS (P<0.05). Conclusion: X-ray can damage the directional adipogenic differentiation of human bone marrow mesenchymal stem cells, and APS has a protective effect on the adipogenic differentiation of human bone marrow mesenchymal stem cells after X-ray irradiation.

Effects of astragalus polysaccharide on paraquat-induced pulmonary fibrosis in rats by regulating TGF-β1/Smads signaling pathway / 西安交通大学学报(医学版)

Objective To explore the effects of astragalus polysaccharide on paraquat-induced pulmonary fibrosis in rats by regulating TGF-β1/Smads signaling pathway. Methods Rats were divided into control group, APS, PQ and PQ + APS groups. Paraquat was used to establish the rat model of pulmonary fibrosis. Lung wet/dry weight ratio and hydroxyproline content were measured; the pathological changes were observed by HE staining. The fibrosis in lung tissues was observed by Masson staining, and the concentrations of tumor necrosis factor α(TNF-α), interleukin 1β(IL-1β), and IL-6 were measured by ELISA kits. RT-PCR was used to detect the gene expressions of transforming growth factor-β1 (TGF-β1), collagen Ⅰ and collagen Ⅲ. Western blotting was used to detect the protein expressions of E-cadherin, α-smooth muscle actin (α-SMA), Vimentin, TGF-β1, Smad3, p-Smad3, and Smad7. Results Compared with the control group, in the PQ group the lung wet weight/dry weight ratio increased (t=12.922, P<0.001) and the hydroxyproline content increased (t=20.920, P<0.001). There were pathological changes and collagen deposition in lung tissues. TNF-α (t=23.932, P<0.001), IL-1β (t=34.826, P<0.001), and IL-6 (t=17.985, P<0.001) in alveolar lavage fluid all increased in concentration. TGF-β1 (t=20.934, P<0.001), collagen Ⅰ (t=26.853, P<0.001), and collagen Ⅲ (t=18.493, P<0.001) gene expressions increased. E-cadherin (t=25.456, P<0.001) protein expression decreased; α-SMA (t=26.980, P<0.001), Vimentin (t=23.862, P<0.001), and TGF-β1 (t=39.836, P<0.001) protein expressions increased. p-Smad3/Smad3 ratio (t=19.606, P<0.001) increased, Smad7 (t=30.904, P<0.001) protein expression decreased. Compared with PQ group, in PQ+APS group lung wet weight/dry weight ratio decreased (t=9.174, P<0.001), hydroxyl amino acid content (t=10.999, P<0.001) decreased, and lung tissue pathology and collagen deposition reduced. TNF-α (t=8.654, P<0.001), IL-1β (t=18.164, P<0.001), and IL-6 (t=7.573, P<0.001) concentrations in alveolar lavage fluid decreased. TGF-β1 (t=8.879, P<0.001), collagen Ⅰ (t=12.687, P<0.001) and collagen Ⅲ (t=11.333, P<0.001) gene expressions reduced. E-cadherin (t=14.255, P<0.001) protein expression increased; α-SMA (t=16.866, P<0.001), Vimentin (t=18.439, P<0.001), and TGF-β1 (t=14.688, P<0.001) protein expressions as well as p-Smad3/Smad3 ratio (t=11.384, P<0.001) were down-regulated; Smad7 (t=13.131, P<0.001) protein expression increased. Conclusion Astragalus polysaccharide can alleviate paraquat-induced pulmonary fibrosis in rats. Its mechanism may be related to inhibiting the TGF-β1/Smads signal pathway.

Astragalus polysaccharide improves anti-tumor immunity mediated by macrophages and natural killer cells in mice / 中华微生物学和免疫学杂志

Objective To evaluate the regulatory effects of Astragalus polysaccharide (APS) on macrophage polarization and NK cell-mediated anti-tumor responses in mice. Methods C57BL/ 6 mice were injected intraperitoneally with APS once a day for seven consecutive days. Activation of immune cells was then induced by intraperitoneal injection of polyinosinic-polycytidylic acid (Poly I : C) 24 h after the APS intervention. Peritoneal macrophages were collected 24 h after induction to analyze the status of polari-zation and the production of nitric oxide (NO). Cytotoxicity and exocytosis of activated NK cells were meas-ured to assess the effector functions of these cells. NK cell activities induced by NKG2D were studied in the absence of the whole JNK or JNK2 signaling pathway. Results Intraperitoneal injection of APS promoted the polarization of macrophages induced by tumor cells in mice, and enhanced the cytotoxicity of NK cells to tumor cells. However, APS was in need of the involvement of appropriate stimulatory factors to have regula-tory effects. Complete inhibition of JNK signaling pathway dramatically reduced the effector functions of NK cells, which could not be recovered by APS administration. Conclusions APS was involved in the regula-tion of anti-tumor innate immunity through enhancing the M1-polarization of macrophages and improving the effector functions of NK cells. This study might to some extent elucidate the mechanism of APS in immune regulation and anti-tumor immunity.

Effect of Astragalus Polysaccharide on Growth and Tumor-related Apoptosis Protein of Human Breast Cancer MDA-MB-231 Transplanted Tumor in Nude Mice / 中国实验方剂学杂志

Objective: To observe the inhibitory effect of astragalus polysaccharides (APS) on growth of human breast cancer MDA-MB-231 xenograft tumor in nude mice and its effect on the apoptosis of tumor cells, in order to study the effect of APS on growth and induction of apoptosis of triple negative breast cancer MDA-MB-231 and its possible molecular mechanism. Method: Human breast cancer cell MDA-MB-231 was inoculated into the right axillary subcutaneous of BALB/c-nu female nude mice to establish the transplanted tumor model of breast cancer. Eighteen nude mice were randomly divided into 3 groups:model group (saline per day), low-dose APS group (200 mg·kg-1 APS per day), and high-dose APS group (400 mg·kg-1 APS per day), with 6 rats in each group. The drug was administered by gavage (200 μL) daily for 21 days. In the experiment, the length and diameter of breast cancer transplanted tumor were measured every two days, and the tumor volume was recorded and calculated. At the end of the experiment, the changes of tumor mass and tumor volume of the low and high-dose APS groups and the model group were observed and compared, and the tumor inhibition rate was calculated. The cell morphology in tumor tissue was observed by hematoxylin-eosin (HE) staining, and Terminal-deoxynucleoitidyl transferase mediated nick end labeling (TUNEL) was used to verify the apoptosis of breast cancer tissues. The expressions of apoptosis-related proteins, such as B-cell lymphoma/leukemia-2 protein (Bcl-2), Bcl-2 associated X protein (Bax), Caspase in tumor tissues was detected by Western blot. Result: The tumor volume of breast cancer decreased in the low and high-dose APS groups, and the tumor inhibition rates were 37.9%and 57.57%, respectively, with statistically significant differences from the model group (PP0.01). HE of tumor tissue cells showed that APS led to obvious morphological changes, with apoptosis in the tissue cells. TUNEL staining showed that the apoptosis rate of tumor cells in APS intervention groups was higher than that in control group. Western blot showed that expression of Bcl-2 protein decreased(PPPPConclusion: APS can effectively inhibit the growth of MDA-MB-231 breast cancer xenografts in nude mice and induce apoptosis in human breast cancer MDA-MB-231 cells. The mechanism may be related to the effect of APS on expressions of apoptosis-related proteins Bcl-2, Bax, Caspase-9 and Caspase-7 in breast cancer cells.