Astragaloside IV is the major active constituent of Astragalus membranaceus, which has been widely used for the treatment of cardiovascular diseases in China. However, the effects of astragaloside IV on myocardial ischemia and its mechanisms of action remain largely unknown. In this study, we have examined the effects of astragaloside IV on myocardial infarction and coronary flow in vivo and in vitro. The possible roles of its antioxidative and nitric oxide-inducing properties were also explored. Astragaloside IV significantly reduced infarct size in dogs subjected to coronary ligation in vivo. Astragaloside IV also improved post-ischemic heart function and ameliorated reperfusion arrhythmias in rat hearts in vitro. The cardioprotection of astragaloside IV was accompanied by a significant increase in coronary flow both in vivo and in vitro. The nitric oxide synthase inhibitor, Nomega-nitro- L-arginine methyl ester partially abrogated astragaloside IV's protective effect on heart function. Myocardial antioxidative enzyme superoxide dismutase activity increased with astragaloside IV administration. These data suggest the potential roles of antioxidative and nitric oxide-inducing properties of astragaloside IV in its protection from myocardial ischemia.

Astragalus membranaceus (AM) has been widely used for treating liver diseases in traditional Chinese medicine. Experimental evidence indicates that it has antitumor potential. In this study, the effect of AM on hepatocarcinogenesis induced by diethylnitrosamine (DEN), two-thirds partial hepatectomy, and 2-acetylaminofluorene (2-AAF) (DEN-PH-AAF) was evaluated using glutathione S-transferase placenta form (GST-P) as marker. First, rats were injected intraperitoneally (i.p.) with DEN (200 mg/kg in saline), a two-thirds partial hepatectomy was carried out 2 weeks later, and the rats were then placed on a basal diet containing 0.02% AAF from week 3 to week 8 to induce hepatocarcinogenesis. The rats were given AM (90 mg/kg or 180 mg/kg body weight) by gavage from week 3 to week 8 (treatment groups). The formation of GST-P-positive foci and the expression of GST-P protein and mRNA caused by DEN-PH-AAF were reduced in the treatment groups, which clearly suggests that AM is effective in delaying DEN-PH-AAF-induced hepatocarcinogenesis.

In the present study, we investigated the antitumor and immunomodulatory activity of Astragalus membranaceus polysaccharide (AMP) on liver cancer using murine H22 hepatocarcinoma model. The results showed that AMP (100 and 400 mg/kg) could effectively inhibit the solid tumor growth of H22 hepatocarcinoma transplanted in BALB/c mice. Besides, the body weight, spleen/thymus indexes and phagocytotic function of macrophage of H22 tumor bearing mice were also improved in two AMP treated groups. Furthermore, AMP treatment could promote the secretion of IL-2, IL-12 and TNF-α and decreased IL-10 level in serum. Taken together, these findings indicate that AMP has antitumor activity in vivo at least partly via improving immune responses of host organism, and seems to be safe and effective for the use of anti-tumor therapy.

Chinese medicine is a complex system guided by traditional Chinese medicine (TCM) theories, which has proven to be especially effective in treating chronic and complex diseases. However, the underlying modes of action (MOA) are not always systematically investigated. Herein, a systematic study was designed to elucidate the multi-compound, multi-target and multi-pathway MOA of a Chinese medicine, QiShenYiQi (QSYQ), on myocardial infarction. QSYQ is composed of Astragalus membranaceus (Huangqi), Salvia miltiorrhiza (Danshen), Panax notoginseng (Sanqi), and Dalbergia odorifera (Jiangxiang). Male Sprague Dawley rat model of myocardial infarction were administered QSYQ intragastrically for 7 days while the control group was not treated. The differentially expressed genes (DEGs) were identified from myocardial infarction rat model treated with QSYQ, followed by constructing a cardiovascular disease (CVD)-related multilevel compound-target-pathway network connecting main compounds to those DEGs supported by literature evidences and the pathways that are functionally enriched in ArrayTrack. 55 potential targets of QSYQ were identified, of which 14 were confirmed in CVD-related literatures with experimental supporting evidences. Furthermore, three sesquiterpene components of QSYQ, Trans-nerolidol, (3S,6S,7R)-3,7,11-trimethyl-3,6-epoxy-1,10-dodecadien-7-ol and (3S,6R,7R)-3,7,11-trimethyl-3,6-epoxy-1,10-dodecadien-7-ol from Dalbergia odorifera T. Chen, were validated experimentally in this study. Their anti-inflammatory effects and potential targets including extracellular signal-regulated kinase-1/2, peroxisome proliferator-activated receptor-gamma and heme oxygenase-1 were identified. Finally, through a three-level compound-target-pathway network with experimental analysis, our study depicts a complex MOA of QSYQ on myocardial infarction.

Aberrant reactions of signal transducer and transcriptional activator (STAT) are frequently detected in multiple myeloma (MM) cancers and can upregulate the expression of multiple genes related to cell proliferation, survival, metastasis, and angiogenesis. Therefore, agents capable of inhibiting STAT activation can form the basis of novel therapies for MM patients. In the present study, we investigated whether the potential anti-cancer effects of Formononetin (FT), a naturally occurring isoflavone derived from Astragalus membranaceus, Trifolium pratense, Glycyrrhiza glabra, and Pueraria lobata, against MM cell lines and human multiple myeloma xenograft tumors in athymic nu/nu mice model are mediated through the negative regulation of STAT3 and STAT5 pathways. Data from the in vitro studies indicated that FT could significantly inhibit cell viability, and induce apoptosis. Interestingly, FT also suppressed constitutive STAT3 (tyrosine residue 705 and serine residue 727) and STAT5 (tyrosine residue 694/699) activation, which correlated with the suppression of the upstream kinases (JAK1, JAK2, and c-Src) in MM cells, and this effect was found to be mediated via an increased production of reactive oxygen species (ROS) due to GSH/GSSG imbalance. Also, FT abrogated STAT3 and STAT5 DNA binding capacity and nuclear translocation. FT induced cell cycle arrest, downregulated the expression of STAT3-regulated anti-apoptotic, angiogenetic, and proliferative gene products; and this correlated with induction of caspase-3 activation and cleavage of PARP. Intraperitoneal administration of FT significantly suppressed the tumor growth in the multiple myeloma xenograft mouse model without exhibiting any significant adverse effects. Overall, our findings indicate that FT exhibits significant anti-cancer effects in MM that may be primarily mediated through the ROS-regulated inhibition of the STAT3 and STAT5 signaling cascade.

Oxidative stress plays a key role in the pathogenesis of various diseases. Antioxidants protect the cells and tissues from oxidative stress by scavenging free radicals and reactive oxygen species. These antioxidants may be endogenous or exogenous. Plants are considered as potential and powerful exogenous source of antioxidants. Astragalus species (spp.), especially Astragalus membranaceus, have a long history of medicinal use in traditional Chinese medicine. Specifically, constituents of the dried roots of Astragalus spp. (Radix Astragali) provide significant protection against heart, brain, kidney, intestine, liver and lung injury in various models of oxidative stress-related disease. Different isolated constituents of Astragalus spp., such as astragalosides, flavonoids and polysaccharides also displayed significant prevention of tissue injury via antioxidant mechanisms. In this article, the antioxidant benefits of Astragalus spp. and its isolated components in protecting tissues from injury are reviewed, along with identification of the various constituents that possess antioxidant activity.

OBJECTIVE

To investigate the protective effects of Astragalus membranaceus (Am) against hapten-induced colitis in male Sprague-Dawley rats as well as its underlying mechanism.

METHODS

Experimental colitis was induced in rats by enema administration of 2,4-dinitrobenzene sulfonic acid (DNBS). Rats were either pretreated with Am extract (2 or 4 g/kg, p.o. once daily) starting from 10 d before DNBS enema, or received Am post-treatment (2 or 4 g/kg, p.o. twice daily) on the three consecutive days following DNBS administration. Colonic lesion area and histological damage were determined, while the activities of myeloperoxidase (MPO) and xanthine oxidase, as well as reduced glutathione (GSH) content were measured in the excised colonic tissues. Besides, protein expression of inducible nitrite oxide synthase (iNOS), intercellular adhesion molecule-1 (ICAM-1) and P-selectin was also detected by Western blot analysis.

RESULTS

Our findings had shown that both macroscopic lesion area and histological colonic damage induced by DNBS were significantly reduced by both Am pre- and post-treatments. These were accompanied by attenuation of the elevated colonic MPO activity and downregulation of the iNOS, P-selectin, and ICAM-1 protein expression. Besides, deprivation of colonic GSH level under colitis condition was also preserved.

CONCLUSIONS

These results demonstrate that Am possesses both preventive and therapeutic potential in experimental colitis. The anti-inflammatory actions involve anti-oxidation along with inhibition of adhesion molecule synthesis in the colonic tissues.

In recent decades, the polysaccharides from the medicinal plants have attracted a lot of attention due to their significant bioactivities, such as anti-tumor activity, antioxidant activity, anticoagulant activity, antidiabetic activity, radioprotection effect, anti-viral activity, hypolipidemic and immunomodulatory activities, which make them suitable for medicinal applications. Previous studies have also shown that medicinal plant polysaccharides are non-toxic and show no side effects. Based on these encouraging observations, most researches have been focusing on the isolation and identification of polysaccharides, as well as their bioactivities. A large number of bioactive polysaccharides with different structural features and biological effects from medicinal plants have been purified and characterized. This review provides a comprehensive summary of the most recent developments in physiochemical, structural features and biological activities of bioactive polysaccharides from a number of important medicinal plants, such as polysaccharides from Astragalus membranaceus, Dendrobium plants, Bupleurum, Cactus fruits, Acanthopanax senticosus, Angelica sinensis (Oliv.) Diels, Aloe barbadensis Miller, and Dimocarpus longan Lour. Moreover, the paper has also been focused on the applications of bioactive polysaccharides for medicinal applications. Recent studies have provided evidence that polysaccharides from medicinal plants can play a vital role in bioactivities. The contents and data will serve as a useful reference material for further investigation, production, and application of these polysaccharides in functional foods and therapeutic agents.

Astragalus membranaceus is widely used to treat stroke and chronic debilitating diseases in China, but the mechanism has not been fully demonstrated to data. In the present study, we, using astragaloside IV, a purified extract from astragalus membranaceus, to a focal cerebral ischemia/reperfusion rat model, aimed to investigate the effect of astragaloside IV on the permeability of the blood-brain barrier since disruption of blood-brain barrier induced by ischemia/reperfusion leads to serious brain injuries. We found that astragaloside IV (10, 20 mg/kg) significantly attenuated the permeability of blood-brain barrier in comparison with vehicle group after ischemia/reperfusion assessed via Evans blue leakage (P<0.05). This was further confirmed by examination of blood-brain barrier permeability under the electron microscope, using lanthanum as a tracer of blood vessel permeability. Lanthanum was usually found within the blood vessel in sham group, rather than in perivascular tissues as shown in vehicle group. In drug groups, lanthanum stain was mainly restricted within the cerebral capillary, indicating the potential protective effect of astragaloside IV on the integrity of blood-brain barrier in ischemia/reperfusion rats. Furthermore, we found that expression of occludin and zonae occludens-1 (ZO-1), the tight junction proteins, was decreased in endothelial cells in vehicle group, which, however, could be reversed by astragaloside IV administration. We propose that regulation of tight junctional proteins in the endothelial cells may be one mechanism astragaloside IV-mediated in attribution to blood-brain barrier protection in the ischemia/reperfusion rats.

Cerebral edema is a critical complication after intravascular thrombolysis post-acute stroke. However, clinical options remained limited for treating cerebral edema after cerebral ischemia/reperfusion (I/R) injury. In the present study, astragaloside IV, a purified extract from astragalus membranaceus, was used in the focal I/R rat model, aimed to investigate its effect on the cerebral edema. We found that astragaloside IV (10 and 20mg/kg) significantly attenuated the cerebral water content (P<0.05) and improved neurological outcomes (P<0.05) in comparison with vehicle group. Moreover, we investigate the effect of astragaloside IV on the (blood-brain barrier) BBB since cerebral edema was closely related to the permeability of the BBB. We found that the permeability of BBB was improved significantly in astragaloside IV groups compared with vehicle group via Evans blue leakage (P<0.05). This was further confirmed under the electron microscope, using lanthanum as a tracer of blood vessel permeability. Lanthanum was usually found within the blood vessel in sham group, rather than in perivascular tissues as shown in vehicle group. In drug groups, lanthanum stain was mainly restricted within the cerebral capillary, indicating the potential BBB-protective effect of astragaloside IV. Furthermore, we found that expressions of Matrix metalloproteinase-9 (MMP-9) and aquaporin 4 (AQP4) were increased in vehicle group, which were related to cerebral vasogenic edema or cytotoxic edema. The up-regulations of MMP-9 and AQP4 were inhibited significantly by astragaloside IV administration. We propose that the anti-edema potential of astragaloside IV was correlated with its regulation of MMP-9 and AQP4.

Astragalus membranaceus is an important medicinal plant in Asia. Several of its varieties have been used interchangeably as raw materials for commercial production. High resolution genetic markers are in urgent need to distinguish these varieties. Here, we sequenced and analyzed the chloroplast genome of A. membranaceus (Fisch.) Bunge var. mongholicus (Bunge) P.K. Hsiao using the next generation DNA sequencing technology. The genome was assembled using Abyss and then subjected to gene prediction using CPGAVAS and repeat analysis using MISA, Tandem Repeats Finder, and REPuter. Finally, the genome was subjected phylogenetic and comparative genomic analyses. The complete genome is 123,582 bp long, containing only one copy of the inverted repeat. Gene prediction revealed 110 genes encoding 76 proteins, 30 tRNAs, and four rRNAs. Five intra-specific hypermutation loci were identified, three of which are heteroplasmic. Furthermore, three gene losses and two large inversions were identified. Comparative genomic analyses demonstrated the dynamic nature of the Papilionoideae chloroplast genomes, which showed occurrence of numerous hypermutation loci, frequent gene losses, and fragment inversions. Results obtained herein elucidate the complex evolutionary history of chloroplast genomes and have laid the foundation for the identification of genetic markers to distinguish A. membranaceus varieties.

Natural products are rich sources of gene modulators that may be useful in prevention and treatment of cancer. Recently, nonsteroidal anti-inflammatory drug (NSAID) activated gene-1 (NAG-1) has been focused as a target of action against diverse cancers like colorectal, pancreatic, prostate, and breast. A variety of natural agents have been reported to play a pivotal role in regulation of NAG-1 through multiple transcriptional mechanisms. The aim of this paper is to review the NAG-1 modulators derived from natural products including plants, marine organisms, and microorganisms. Plant extracts belonging to the families of Fabaceae (Astragalus membranaceus), Ranunculaceae (Coptis chinensis), Menispermaceae (Coscinium fenestratum), Umbelliferae (Pleurospermum kamtschaticum), Lamiaceae (Marubium vulgare), and Rosaceae (Prunus serotina) increased the protein expression of NAG-1 in human colon cancer or hepatocarcinoma cells. Phytochemicals in the class of flavonoids (apigenin, quercetin, isoliquiritigenin, and 2'-hydroxyflavanone), isoflavonoids (formononetin and genistein), catechins (epigallocatechin gallate and epicatechin gallate), stilbenoids (resveratrol and pinosylvin), phenolics (6-gingerol), phloroglucinols (rottlerin and aspidin PB), terpenoids (18 α-glycyrrhetinic acid, platycodin D, pseudolaric acid B, and xanthorrhizol), alkaloids (berberine, capsaicin, and indole-3-carbinol), lignans (isochaihulactone), anthraquinones (damnacanthal), and allyl sulfides (diallyl disulfide) elicited NAG-1 overexpression in various cancer cells. Pectenotoxin-2 from marine organisms and prodigiosin and anisomycin from microorganisms were also reported as NAG-1 modulators. Several transcription factors including EGR-1, p53, ATF-3, Sp1 and PPARγ were involved in natural products-induced NAG-1 transcriptional signaling pathway.

OBJECTIVE

Diabetic nephropathy (DN) is the leading cause of the end-stage failure of kidney, but the efficacy of currently available strategies for the prevention of DN remains poor. An activation of the transcription factor, nuclear factor-kappaB (NF-kappaB), has been suggested to be a key step in the pathogenesis of DN. In the present study, we investigated the effect of Astragalus polysaccharide (APS), an aqueous extract from the Astragalus membranaceus roots, on gene expressions of NF-kappaB and an inhibitory protein of nuclear factor-kappaB (IkappaB) in experimental DN induced by streptozotocin in male Sprague-Dawley rats.

METHODS

Rats with DN were treated with APS (1g/kg p.o.) or benazepril (1.5mg/kg p.o.), an angiotensin-converting enzyme inhibitor, using as positive control. The biochemical parameters such as blood glucose, plasma lipid and microalbuminuria were measured. Also, the mRNA level of NF-kappaB or IkappaB in renal cortex was determined using reverse transcription-polymerase chain reaction.

RESULTS

Eight weeks after the treatment, symptoms including shineless, bristly hair, polyuria, polydipsia, lethargy, physical inactivity, loss of body weight, kyphosis and decubitus position were ameliorated by APS. The levels of blood glucose, plasma lipid and microalbuminuria were lowered in APS-treated rats compared with control rats. The ratio of kidney weight over body weight was reduced and the renal function was improved after APS treatment. The mRNA level of NF-kappaB in renal cortex was decreased and IkappaB mRNA expression was raised by APS. These results suggest that APS has prophylactic and therapeutic effects on the progress of DN;

CONCLUSIONS

therefore, APS is helpful for the prevention and/or treatment of DN at early stage.

Astragalus polysaccharides (APS), one of main bioactive components in Astragalus membranaceus Bunge, has been reported to possess anti-inflammatory activities, but the molecular mechanisms behind this activity are largely unknown. This study aimed to investigate expression of inflammatory cytokines and the MAPK/NF-κB pathway in human THP-1 macrophages induced by lipopolysaccharide (LPS). The results showed that the concentrations of TNF-a and IL-1β released from LPS stimulated THP-1 cells increased significantly compared to control (p < 0.01). After treatment with APS, the TNF-a and IL-1β levels were significantly lower than those in the LPS group (p < 0.05). The mRNA expression of TNF-a and IL-1β were also inhibited. Mechanistic studies indicated that APS strongly suppressed NF-κB activation and down-regulated the phosphorylation of ERK and JNK, which are important signaling pathways involved in the production of TNF-a and IL-1β, demonstrating that APS could suppress the production of TNF-a and IL-1β by LPS stimulated macrophages by inhibiting NF-κB activation and ERK and JNK phosphorylation.

BACKGROUND

The persistent renal hemodynamic maladjustment caused by imbalances between vasoactivators predisposes the kidney to tubulointerstitial injury and ultimate interstitial fibrosis. The decoction (A&A) of a combination of roots of two Chinese herbs, Astragalus membranaceus var. mongholicus and Angelica sinensis, has shown antifibrotic effects in rats with chronic kidney diseases and improvement of renal blood flow in rats with acute ischemic renal injury. In the present study, we investigated the effects and possible mechanisms of A&A on vasoactivators in the process of renal interstitial fibrosis.

METHODS

Male Wistar rats were randomly divided into sham, unilateral ureteral obstruction (UUO) and UAA (UUO plus A&A administration) groups. After oral administration of A&A (14 g/kg/d) for 3, 7 and 10 days, morphological changes were evaluated by HE, Masson and Sirius red staining technique. The levels of Ang-II, ET-1, and the activities of different nitric oxide synthases (NOSs) in renal homogenate were measured by radioimmunoassay. The nitrite concentration as nitric oxide (NO) production was measured using the Griess reagent. Western blot analysis and immunohistochemical staining were performed to determine the expressions of eNOS, nNOS, and iNOS in the kidney. The ability of scavenging reactive oxygen species (ROS) was evaluated by spectrophotometry.

RESULTS

Morphological analysis showed severe interstitial mononuclear cells infiltration, tubular atrophy, renal fibrosis and collagen expression in kidneys of UUO group, which reduced by A&A administration (p<0.05, UAA vs. UUO group). The levels of Ang-II and ET-I were increased in obstructed kidneys, but not significantly changed after A&A administration. NO production did not change in obstructed kidney at day 3 but increased in day 7 and day 10. Administering A&A progressively increased NO production by 2.2, 1.2, and 1.2 fold at days 3, 7 and 10, respectively. The activities of constitutive NOS and iNOS were comparable between UUO group and sham group. In contrast, the activity of constitutive NOS was much higher in UAA than that of UUO rats, which increased 78%, 68% and 78% at days 3, 7 and 10 respectively, although the protein expression of eNOS, nNOS and iNOS in renal tissue had no change in UAA rats. The activities of scavenging ROS in UUO group were not significantly different from the sham group at days 3 and 7, but increased at day 10 (24.1+/-15.0 vs. 10.1+/-0.8 U/min/mg protein, p<0.05). After A&A administration, the activities of scavenging ROS were significantly increased at days 3 and 7 (51.5+/-17.9 vs. 11.7+/-7.4 U/min/mg protein, p<0.05; and 16.1+/-5.6 vs. 7.7+/-1.4 U/min/mg protein, p<0.05) respectively, comparing with the UUO group.

CONCLUSIONS

The anti-fibrosis effects of A&A might be associated with enhancing NO production via eNOS activation and scavenging ROS, and in turn might improve ischemic microvasculature and attenuate interstitial fibrosis.

Shi-Quan-Da-Bu-Tang (Ten Significant Tonic Decoction), or SQT (Juzentaihoto, TJ-48) was formulated by Taiping Hui-Min Ju (Public Welfare Pharmacy Bureau) in Chinese Song Dynasty in AD 1200. It is prepared by extracting a mixture of ten medical herbs (Rehmannia glutinosa, Paeonia lactiflora, Liqusticum wallichii, Angelica sinesis, Glycyrrhiza uralensis, Poria cocos, Atractylodes macrocephala, Panax ginseng. Astragalus membranaceus and Cinnamomum cassia) that tone the blood and vital energy, and strengthen health and immunity. This potent and popular prescription has traditionally been used against anemia, anorexia, extreme exhaustion, fatigue, kidney and spleen insufficiency and general weakness, particularly after illness. In order to restore immunity in cancer patients, potentiate the therapeutic effect and ameliorate adverse toxicity of anticancer agents, 116 Chinese herbal formularies (Kampo) have been screened and evaluated. Fifteen compounds were found to have such actions. Among these, SQT was selected as the most effective as a potent biological response modifier. During the last eight years, animal models and clinical studies have revealed that SQT demonstrates extremely low toxicity (LD50>> 15 g/kg op murine), self-regulatory and synergistic actions of its components in immunomodulatory and immunopotentiating effects (by stimulating hemopoietic factors and interleukins production in association with NK cells, etc.), potentiates therapeutic activity in chemotherapy (mitomycin, cisplatin, cyclophosphamide and fluorouracil) and radiotherapy, inhibits the recurrence of malignancies, prolongs survival, as well as ameliorate and/or prevents adverse toxicities (GI disturbances such as anorexia, nausea, vomiting, hematotoxicity, immunosuppression, leukopenia, thrombocytopenia, anemia and nephropathy, etc.) of many anticancer drugs. The application and mechanistic studies of SQT in future development have potential importance in basic and clinical research of the traditional Chinese therapeutic approach of "toning the blood and strengthening Qi (vital energy)" in cancer immunotherapy.

Several botanicals, including Crataegus oxycantha, Terminalia arjuna, Inula racemosa, and Astragalus membranaceus, have been found to have therapeutic benefit for the treatment of cardiovascular disease. Crataegus oxycantha has been used traditionally as a cardiac tonic and current uses include treatment for angina, hypertension, arrhythmias, and congestive heart failure. Animal studies have also indicated that Crataegus extracts may also have potential use as anti-ischemic and lipid-lowering agents. The bark of the Terminalia arjuna tree has a long history of use as a cardiac tonic as well, and has been indicated in the treatment of coronary artery disease, heart failure, hypercholesterolemia and for relief of anginal pain. Additionally, it has been found to have antibacterial and antimutagenic properties. Inula racemosa, also known as Pushkarmoola, is another traditional Ayurvedic botanical that has potential cardioprotective benefit. In human trials, a combination of Inula racemosa and Commiphora mukul was shown to be superior to nitroglycerin in reducing the chest pain and dyspnea associated with angina. Astragalus membranaceus, a Chinese herb, is often used as a "Qi tonifier" and has been studied for its therapeutic benefit in treatment of ischemic heart disease, myocardial infarction, heart failure, and relief of anginal pain. Clinical studies have indicated that its in vitro antioxidant activity is the mechanism by which it affords its cardioprotective benefit.

An important traditional Chinese medicine herb, Astragalus membranaceus var. Mongholicus, whose dried root is known as Radix astragali ("Huangqi" in Chinese), has high flavonoid content as an essential active constituent. Phenylalanine ammonia-lyase (PAL, EC 4.3.1.5) catalyzes the first and also a rate-limiting step in phenylpropanoid pathway, which supplies precursors for a variety of secondary metabolites including flavonoids. A PAL gene, designated AmPAL1 (GenBank accession no. AY986506), was isolated from A. membranaceus var. Mongholicus with a full-length cDNA of 2562 nucleotides and an open reading frame of 2154 bp. Northern blot analysis revealed that AmPAL1 expressed universally in different organs, and its expression was markedly induced by UV irradiation, mechanical wounding, and white light irradiation on etiolated seedlings, with some distinctive responsive properties. Content of a typical flavonoid, quercetin, in A. membranaceus var. Mongholicus of different ages correlated with PAL enzymatic activity. Transgenic tobacco plants harboring AmPAL1 under the control of the CaMV35S promoter showed significantly increased PAL activity and correlatively increased quercetin content than those in non-transformed plants. These results indicate that PAL is maybe a key point for flux into flavonoid biosynthesis in the genetic control of secondary metabolism in A. membranaceus var. Mongholicus.

The disease burden of diabetes mellitus is increasing throughout the world. The need for more potent drugs to complement the present anti-diabetic drugs has become an imperative. Astragalus membranaceus, a key component of most Chinese herbal anti-diabetic formulas, has been an important prospect for lead anti-diabetic compounds. It has been progressively studied for its anti-diabetic properties. Ethnopharmacological studies have established its potential to alleviate diabetes mellitus. Recent studies have sought to relate its chemical constituents to types 1 and 2 diabetes mellitus. Its total polysaccharides, saponins, and flavonoids fractions and several isolated compounds have been the most studied. The total polysaccharides fraction demonstrated activity to both types 1 and 2 diabetes mellitus. This paper discusses the anti-diabetic effects and pharmacological action of the chemical constituents in relation to types 1 and 2 diabetes mellitus.

BACKGROUND

Diabetic nephropathy (DN) is a major cause of Chronic Kidney Disease and End-Stage Renal Disease throughout the world; however, the reversibility of diabetic nephropathy remains controversial. Endoplasmic reticulum (ER) stress plays an important role in the pathogenesis of DN. Astragaloside IV (AS-IV) is derived from Astragalus membranaceus (Fisch) Bge, a widely used traditional herbal medicine in China, and has diverse pharmacological activities including the attenuation of podocyte injury and amelioration of proteinuria in idiopathic nephrotic syndrome. The present study aimed to investigate the effect and mechanism of AS-IV on proteinuria in the rat streptozotocin (STZ)-induced model of diabetes.

METHODS

Male Sprague-Dawley (SD) rats were randomly divided into four groups: normal control (Normal group), diabetic nephropathy (Model group), diabetic nephropathy plus AS-IV treatment (AS-IV group) and diabetic nephropathy plus 4-phenyl butyric acid treatment (PBA group). ER stress was induced in cultured human podocytes, pretreated with or without AS-IV, with tunicamycin (TM). At the end of 8 weeks, serum creatinine (Scr), blood urea nitrogen (BUN) and 24-hour urinary protein excretion rate (UAER) were determined. Renal morphology was examined after periodic acid-Schiff staining of kidney sections. Apoptosis of podocytes was measured by flow cytometry. The total expression and phosphorylation of eIF2α, PERK and JNK, and the expression of CHOP and cleaved caspase-3 were determined by western blotting. The expression of glucose-regulated protein 78 (GRP78) and 150 kDa oxygen-regulated protein (ORP150) mRNA and protein was determined by real-time PCR and western blotting respectively.

RESULTS

AS-IV treatment significantly reduced urinary albumin excretion, plasma creatinine and blood urea nitrogen levels, and prevented the mesangial matrix expansion and increase in mean mesangial induced by STZ. AS-IV also prevented the phosphorylation of eIF2α, PERK and JNK, and inhibited the expression of GRP78 and ORP150 markedly, both in vivo and in vitro. AS-IV inhibited the TM-induced apoptosis of podocytes, concomitant with decreased CHOP expression and cleaved caspase-3.

CONCLUSIONS

This study supports the hypothesis that AS-IV reduces proteinuria and attenuates diabetes, which is associated with decreased ER stress. This might be an important mechanism in the renoprotective function of AS-IV in the pathogenesis of DN.

An in vivo experiment was conducted to study the potential prebiotic effects of mushroom and herb polysaccharide extracts, Lentinus edodes extract (LenE), Tremella fuciformis extract, and Astragalus membranaceus Radix extract, on chicken growth and the cecal microbial ecosystem, as compared with the antibiotic Apramycin (APR). This investigation was carried out in terms of a dose-response study. The chickens were naturally infected with avian Mycoplasma gallisepticum prior to the experiment. The BW gain, cecal pH, viscosity, and predominant microbial populations were measured 1 wk after the extract and APR treatments. The extracts and APR significantly stimulated growth of the chickens infected with avian Mycoplasma gallisepticum. The average BW gain of the groups fed with the extracts was significantly lower than that of the antibiotic group. The extracts had no significant effect on cecal pH. However, cecal viscosity and microbial populations were significantly affected by feeding extracts and antibiotic. In contrast to APR, the extracts stimulated the number of the potentially beneficial bacteria (bifidobacteria and lactobacilli), while reducing the number of the potentially harmful bacteria (Bacteroides spp. and Escherichia coli). Of the 3 extracts, LenE was associated with the most cecal bifidobacteria and lactobacilli. With each increase in the LenE dose, birds tended to have higher BW gain and total aerobe and anaerobe counts. Numbers of predominant cecal bacteria, in particular, E. coli, bifidobacteria, and lactobacilli, were significantly increased with increases in the LenE dose. It would seem that these specific mushroom and herb polysaccharide extracts hold some promise as potential modifiers of intestinal microbiota in diseased chickens.

We show that APS, a polysaccharide isolated from the roots of Astragalus membranaceus, significantly induces nitric oxide (NO) production and inducible NO synthase (iNOS) transcription through the activation of nuclear factor-kappaB/Rel (NF-kappaB/Rel). In vivo administration of APS induced NO production by peritoneal macrophages of B6C3F1 mice. APS also dose-dependently induced the production of NO in isolated mouse peritoneal macrophages and RAW 264.7, a mouse macrophage-like cell line. Moreover, iNOS protein and mRNA transcription were strongly induced by APS in RAW 264.7 cells. To further investigate the mechanism responsible for the induction of iNOS gene expression, we investigated the effect of APS on the activation of transcription factors including NF-kappaB/Rel and Oct, whose binding sites were located in the promoter of iNOS gene. Treatment of RAW 264.7 cells with APS produced strong induction of NF-kappaB/Rel-dependent reporter gene expression, whereas Oct-dependent gene expression was not affected by APS. Nuclear translocation and DNA binding activity of NF-kappaB/Rel was significantly induced by APS. The treatment with NF-kappaB SN50, an inhibitor of NF-kappaB/Rel nuclear translocation, effectively inhibited the activation of NF-kappaB/Rel binding complexes and NO production. In conclusion, we demonstrate that APS stimulates macrophages to express iNOS gene through the activation of NF-kappaB/Rel.

Endothelial cells of cerebral microvessels are one of the components of blood-brain-barrier (BBB), which are connected by tight junctions (TJs). BBB disruption in cerebral diseases such as ischemic stroke, Alzhemer's disease, multiple sclerosis and traumatic brain injury is implicated to exacerbate the disease progression. Astragaloside IV (ASIV) isolated from Astragalus membranaceus prevents BBB breakdown in rodents induced with cerebral edema and experimental autoimmune encephalomyelitis. However, its underlying molecular mechanism has not been elucidated yet. In present study, ASIV was found to prevent the leakage of BBB in LPS-induced mice, which was accompanied with increased zo-1 and occludin but reduced VCAM-1 in brain microvessels. Similarly, in brain endothelial cell line bEnd.3 cells, ASIV mitigated the increased permeability induced by LPS, as evidenced by increased TEER and reduced sodium fluorescein extravasation. ASIV also enhanced the expression of TJ proteins such as zo-1, occludin and claudin-5 in LPS stimulated bEnd.3 cells. Meanwhile, it inhibited the inflammatory responses and prevented the monocyte adhesion onto bEnd.3 cells upon LPS stimulation. Further study disclosed that ASIV could alleviate ROS level and activate Nrf2 antioxidant pathway in bEnd.3 cells. When Nrf2 was silenced, the protective effect of ASIV was abolished. In brain microvessels of LPS-induced mice, ASIV also enhanced the expression of Nrf2 antioxidant pathway related proteins. Collectively, our results demonstrated that ASIV protected the integrity of BBB in LPS-induced mice, the mechanism of which might be mediated via activating Nrf2 signaling pathway. The findings suggested that ASIV might be a potential neuroprotective drug acting on BBB.