Effect of Panax notoginseng saponins on the expression of beta-amyloid protein in the cortex of the parietal lobe and hippocampus, and spatial learning and memory in a mouse model of senile dementia

BACKGROUND： The pharmacological actions of Panax notoginseng saponins （PNS） lie in removing free radicals, anti-inflammation and anti-oxygenation. It can also improve memory and behavior in rat models of Alzheimer＇s disease. OBJECTIVE： Using the Morris water maze, immunohistochemistry, real-time PCR and RT-PCR, this study aimed to measure improvement in spatial learning, memory, expression of amyloid precursor protein （App） and β -amyloid （A β ）, to investigate the mechanism of action of PNS in the treatment of AD in the senescence accelerated mouse-prone 8 （SAMP8） and compare the effects with huperzine A. DESIGN, TIME AND SETTING： A completely randomized grouping design, controlled animal experiment was performed in the Center for Research ＆ Development of New Drugs, Guangxi Traditional Chinese Medical University from July 2005 to April 2007. MATERIALS： Sixty male SAMP8 mice, aged 3 months, purchased from Tianjin Chinese Traditional Medical University of China, were divided into four groups： PNS high-dosage group, PNS low-dosage group, huperzine A group and control group. PNS was provided by Weihe Pharmaceutical Co., Ltd. （batch No.： Z53021485, Yuxi, Yunan Province, China）. Huperzine A was provided by Zhenyuan Pharmaceutical Co., Ltd. （batch No.： 20040801, Zhejiang, China）. METHODS： The high-dosage group and low-dosage group were treated with 93.50 and 23.38 mg/kg PNS respectively per day and the huperzine A group was treated with 0.038 6 mg/kg huperzine A per day, all by intragastric administration, for 8 consecutive weeks. The same volume of double distilled water was given to the control group. MAIN OUTCOME MEASURES： After drug administration, learning and memory abilities were assessed by place navigation and spatial probe tests. The recording indices consisted of escape latency （time-to-platform）, and the percentage of swimming time spent in each quadrant. The number of A β 1-40, A β 1-42 and App immunopositive neurons in the brains of SAMP8 mice was analyzed by immunohistochemistry. The mRNA content ofApp, tau, acetylcholinesterase, and synaptophysin （Syp） was tested by real time PCR and RT-PCR. RESULTS： The PCR results show that PNS can downregulate the expression of the App gene and upregulate the expression of the Syp gene in the parietal cortex and hippocampus of SAMP8 mice. The therapeutic effects of the PNS high-dosage group were greater than those of the PNS low-dosage group and the huperzine A group （P 〈 0.05）. The results of the Morris water maze and immunohistochemistry indicated that PNS can improve the capacity for spatial learning and memory in SAMP8 mice, and reduce the content of A β 1-40, A β 1-42 and expression of App in the brains of SAMP8 mice. The therapeutic effects of the PNS high-dosage group were greater than that of the PNS low-dosage group and the huperzine A group （P 〈 0.05）. CONCLUSION： These results support the hypothesis that PNS plays a therapeutic and protective role on the pathological lesions and learning dysfunction of Alzheimer＇s disease. The therapeutic effects of PNS for Alzheimer＇s disease are possibly achieved through downregulating the expression of the App gene and upregulating the expression of the Syp gene. The therapeutic effects of PNS are dose-dependent and are greater than the effect of huperzine A.

Panax notoginseng saponins preconditioning protects rat liver grafts from ischemia/reper- fusion injury via an antiapoptotic pathway

BACKGROUND: Ischemia/reperfusion (I/R) injury is a major cause of primary graft dysfunction and renders an al- lograft more immunogenic in orthotopic liver transplanta- tion (OLT). Panax notoginseng saponins (PNS) has been re- ported to exert protective effects against I/R injury to vari- ous organs. The objective of this study is to investigate whether PNS preconditioning protects rat liver grafts from I/R injury via an antiapoptotic pathway. METHODS: Male Sprague-Dawley rats were used as donors and recipients of orthotopic liver transplantation ( OLT) and were divided into PNS preconditioning group (group P) and normal saline control group (group N) randomly according to whether PNS (50 mg/kg) was injected intra- venously 1 hour before liver grafts harvesting, and sham group (group S). The animals were separately killed 2, 6 and 24 hours after reperfusion. Plasma samples were collect- ed for test of alanine amino-transferase (ALT) and aspartate aminotransferase (AST). Liver tissues were collected to de- tect histological changes, apoptosis and the expression of TNF-α, Bcl-2 and Caspase-3 mRNA. RESULTS: The serum levels of ALT and AST and the apop- tosis index (AI) of liver tissue in group P were lower than in group N significantly 2, 6 and 24 hours after reperfusion. Compared with group N, the expression of TNF-a and Caspase-3 mRNA was reduced significantly in group P 2 and 6 hours after reperfusion and the expression of Bcl-2 mRNA was enhanced significantly in group P 6 and 24 hours after reperfusion. CONCLUSIONS: PNS preconditioning protects liver grafts from I/R injury effectively in rat OLT via an antiapoptotic pathway. The antiapoptotic mechanisms of PNS may in- clude inhibiting the expression of TNF-a and Caspase-3 and enhancing the expression of Bcl-2.

Effects of Panax notoginseng saponins in a rat model of Alzheimer's disease

BACKGROUND： Modem pharmacological studies have demonstrated that Panax notoginseng saponins （PNS） can ameliorate and protect from neuropathological impairment. Whether PNS can improve the abnormality in memory and behavior of rats with Alzheimer＇s disease （AD） remains unclear. OBJECTIVE： Based on a Morris water maze test, this study aimed to measure improvements of spatial learning and memory by PNS in a rat model of AD, and to compare effects with huperzine A. DESIGN： A completely randomized grouping design, controlled animal experiment. SETTING： Center of Research ＆ Development of New Drugs, Guangxi Traditional Chinese Medical University. MATERIALS： Ninety healthy Wistar rats of both genders, 15-month-old （n =75） and 3-month-old rats as young controls （n =15）, were used for this study. The study was performed in accordance with animal ethics guidelines for the use and care of animals. PNS was provided by Weihe Pharmaceutical Co., Ltd （permission No. Z53021485, Yuxi, Yunan Province, China）. Morris water maze equipment was provided by the Institute of Physiology, Chinese Academy of Science. METHODS： This study was performed at the Center of Research ＆ Development of New Drugs, Guangxi Traditional Chinese Medical University from June 2003 to April 2005. Of the included rats, 15 healthy aged rats were randomly chosen as aged controls, and the remaining 60 aged rats were randomly divided into 4 groups with 15 rats in each： model group, PNS high- and low-dose groups, and an huperzine A group. Rats in the model group and the 3 treated groups were treated with intraperitoneal infusion of 9.6 g/L D-galactose （5 mL/kg） every day for 6 weeks successively to induce a subacute aging model. During week 7, animals received 1 μ L ibotenic acid （5 g/L） bilaterally into the nucleus basalis of Meynert to create a rat model of AD. The young and old rat controls received, in parallel, a corresponding volume of saline. Two weeks later, rats in the PNS high- and low-dose groups were gavaged with 200 and 100 mg/kg PNS suspension, respectively. Huperzine A suspension （0.3 mg/kg） was used in the huperzine A group. Rats in the other 3 groups were gavaged with a corresponding volume of normal saline. In each group, administration was carried out once per day for 4 consecutive weeks. MAIN OUTCOME MEASURES： After administration, learning and memory abilities were measured by place navigation and spatial probe tests. Recording indices consisted of escape latency （time-to-platform）, number of times to find the platform within 2 minutes, number of times the animal crosses the original platform location, and the percent of swimming time in each quadrant. RESULTS： Several rats died due to inflammatory reactions following brain lesion or intragastric administration; therefore, 61 rats were included in the final analysis. Results of spatial navigation test： Escape latency of rats in the model group was significantly prolonged, and number of times to find the platform within 2 minutes were significantly reduced compared with other groups （both P 〈 0.05）. No significant differences in these two indices were measured among the administration groups （all P 〉 0.05）. Results of spatial probe test： Times for crossing the original platform location and percent of time spent in the quadrant of original platform location were significantly less in the model group than in the other groups （P 〈 0.05）. There were no significant differences in these two indices among the administration groups （P 〉 0.05）. CONCLUSION： PNS can remarkably improve spatial learning and memory abilities of rats with AD. The therapeutic effect of PNS is not dose-dependent and is equivalent to the effect of huperzine A.

Panax notoginseng saponins influence on transplantation of neural stem cell-derived dopaminergic neurons in a rat model of Parkinson’s disease

BACKGROUND： Dopaminergic neurons differentiated from neural stem cells have been successfully used in the treatment of rat models of Parkinson＇s disease; however, the survival rate of transplanted cells has been low. Most cells die by apoptosis as a result of overloaded intracellular calcium and the formation of oxygen free radicals. OBJECTIVE： To observe whether survival of transplanted cells, transplantation efficacy, and dopaminergic differentiation from neural stem cells is altered by Panax notoginseng saponins （PNS） in a rat model of Parkinson＇s disease. DESIGN, TIME AND SETTING： Cellular and molecular biology experiments with randomized group design. The experiment was performed at the Animal Experimental Center, First Hospital of Sun Yat-sen University from April to October 2007. MATERIALS： Thirty-two adult, healthy, male Sprague Dawley rats, and four healthy Sprague Dawley rat embryos at gestational days 14-15 were selected. The right ventral mesencephalon was injected with 6-hydroxydopamine to establish a model of Parkinson＇s disease. 6-hydroxydopamine and apomorphine were purchased from Sigma, USA. METHODS： Neural stem cells derived from the mesencephalon of embryonic rats were cultivated and passaged in serum-free culture medium. Lesioned animals were randomly divided into four groups （n = 8）： dopaminergic neuron, dopaminergic neuron ＋ PNS, PNS, and control. The dopaminergic neuron group was injected with 3 μL cell suspension containing dopaminergic neurons differentiated from neural stem cells. The dopaminergic neurons ＋ PNS group received 3 μ L dopaminergic cell suspension combined with PNS （250 mg/L）. The PNS group received 3 μL PNS （250 mg/L）, and the control group received 3 μL DMEM/F12 culture medium. MAIN OUTCOME MEASURES： The rats were transcardially perfused with 4% paraformaldehyde at 60 days post-grafting for immunohistochemistry. The rats were intraperitoneally injected with apomorphine （0.5 mg/kg） to induce rotational behavior. RESULTS： Cell counts of tyrosine hydroxylase-positive neurons in the dopaminergic neuron ＋ PNS group were （732±82.6） cells/400-fold field. This was significantly greater than the dopaminergic neuron group [（326 ± 34.8） cells/400-fold field, P 〈 0.01]. Compared to the control group, the rotational asymmetry of rats that received dopaminergic neuron transplants was significantly decreased, beginning at 20 days after operation （P 〈 0.01）. Rotational asymmetry was further reduced between 10-60 days post-surgery in the dopaminergic neuron ＋ PNS group, compared to the dopaminergic neuron group （P 〈 0.01）. CONCLUSION： Panax notoginseng saponins can increase survival and effectiveness of dopaminergic neurons differentiated from neural stem cells for transplantation in a rat model of Parkinson＇s disease.

Yizhijiannao Granule and a combination of its effective monomers,icariin and Panax notoginseng saponins,inhibit early PC12 cell apoptosis induced by beta-amyloid(25-35)

One of our previous studies showed that Yizhijiannao Granule,a compound Chinese medicine, effectively improved the clinical symptoms of Alzheimer’s disease.In the present study,we established a model of Alzheimer’s disease using beta-amyloid（25-35）in PC12 cells,and treated the cells with Yizhijiannao Granule and its four monomers,i.e.,icariin,catechin,Panax notoginseng saponins,and eleutheroside E.Flow cytometry showed that Yizhijiannao Granule-containing serum, icariin,Panax notoginseng saponins,and icariin＋Panax notoginseng saponins were protective against beta-amyloid（25-35）-induced injury in PC12 cells.Icariin in combination with Panax notoginseng saponins significantly inhibited early apoptosis of PC12 cells with beta-amyloid （25-35）-induced injury compared to icariin or Panax notoginseng saponins alone.The effects of icariin＋Panax notoginseng saponins were similar to the effects of Yizhijiannao Granule.The findings indicate that two of the effective monomers of Yizhijiannao Granule,icariin and Panax notoginseng saponins,can synergistically inhibit early apoptosis of PC12 cells induced by beta-amyloid（25-35）.

Effects of Panax notoginseng saponins on hydrogen peroxide-induced apoptosis in cultured rabbit bone marrow stromal cells

Objective To investigate the effects of Panax notoginseng saponins(PNS)on hydrogen peroxide(H2O2)-induced apoptosis in cultured rabbit bone marrow stromal cells(BMSCs).Methods BMSCs from 3-month-old New Zealand rabbits were isolated and cultured by the density gradient centrifugation combined with adherent method.The cultured BMSCs were divided into three groups:normal control,H2O2 treatment(100μmol/L),and PNS pretreatment(0.1g/L).Intracellular reactive oxygen species(ROS)levels as the index of oxidative stress were measured by using 2’7’-dichlorodihydrofluorescein diacetate.Flow cytometry was used to observe the apoptosis of BMSCs by staining with annexinV-FITC/PI.The protein expression of Bax in BMSCs was analyzed by Western blotting.Activity of caspase-3 enzyme was measured by spectrofluorometry.Results Pretreatment with PNS significantly decreased intracellular ROS level induced by H2O2(P<0.01).PNS markedly attenuated H2O2-induced apoptosis rate from 38.68% to 19.24%(P<0.01).PNS reversed H2O2-induced augmentation of Bax expression.Furthermore,PNS markedly reduced the altered in activity of caspase-3 enzyme induced by H2O2(P<0.01).Conclusion PNS has a protective effect on hydrogen peroxide-induced apoptosis in cultured rabbit BMSCs by scavenging ROS and decreasing Bax expression and caspase-3 activity.

Panax notoginseng saponins improve recovery after spinal cord transection by upregulating neurotrophic factors

Saponins extracted from Panax notoginseng are neuroprotective, but the mechanisms underlying this effect remain unclear. In the present study, we established a rat model of thoracic（T10） spinal cord transection, and injected Panax notoginseng saponins（100 mg/kg） or saline 30 minutes after injury. Locomotor functions were assessed using the Basso, Beattie, and Bresnahan（BBB） scale from 1 to 30 days after injury, and immunohistochemistry was carried out in the ventral horn of the spinal cord at 1 and 7 days to determine expression of nerve growth factor（NGF） and brain-derived neurotrophic factor（BDNF）. Our results show that at 7–30 days post injury, the BBB score was higher in rats treated with Panax notoginseng saponins than in those that received saline. Furthermore, at 7 days, more NGF- and BDNF-immunoreactive neurons were observed in the ventral horn of the spinal cord of rats that had received Panax notoginseng saponins than in those that received saline. These results indicate that Panax notoginseng saponins caused an upregulation of NGF and BDNF in rats with spinal cord transection, and improved hindlimb motor function.

Effects of Panax notoginseng saponins on apoptosis induced by hydrogen peroxide in cultured rabbit bone marrow stromal cells via altering the oxidative stress level and down-regulating caspase-3

Objective： To investigate the effects of Panax notoginseng saponins （PNS） on hydrogen peroxide （H2O2）-induced apoptosis in cultured rabbit bone marrow stromal cells （BMSCs）. Methods： The effects of different concentrations of PNS on proliferation and early osteoblast differentiation of BMSCs were determined by the MTT assay and an alkaline phosphatase （ALP） assay. An optimal effective concentration of PNS was determined and used in subsequent experiments. The cultured BMSCs were divided into three groups： untreated control, H2O2 treated, and PNS pretreatment of H2O2 treated. The oxidative stress level was assessed by superoxide dismutase （SOD） and malondialdehyde （MDA） assays. Flow cytometry was used to determine BMSC apoptosis by staining with annexinV-FITC/propidium iodide （PI）. The activity of caspase-3 enzyme was measured by spectrofluorometry. Results： PNS （0.1g/L） significantly increased both BMSC proliferation rate and ALP activity, while it decreased the indicators of oxidative stress, caspase-3 activity, and the apoptosis rate of BMSCs induced by H2O2.. Conclusion： PNS, acting as a biological antioxidant, had a protective effect on H2O2-induced apoptosis in cultured rabbit BMSCs by decreasing oxidative stress and down-regulating caspase-3.

The neuroprotective role of Panax notoginseng saponins in APP/PS1 transgenic mice through the modulation of cerebrovascular

Background:Panax notoginseng saponins(PNS)is extracted from Sanqi(Panax notoginseng),which is a valuable herb and has been widely used in traditional Chinese medicine for the treatment of cerebrovascular diseases and pain.PNS has been proved to promote blood circulation and angiogenesis by inhibiting platelet aggregation.In our previous study,PNS accompanied with geniposide can prevent Alzheimer’s disease(AD).However,the efficacy of PNS and its potential mechanism in AD remain unclear.Methods:Amyloid precursor protein/presenilin-1(APP/PS1)transgenic(Tg)mice were used as AD-like animal models.Wild-type mice and APP/PS1 transgenic were administrated with saline solution while mice in PNS treatment group were administrated with PNS at a dosage of 17 mg/kg/day for three months.Morris water maze(MWM)was applied to evaluate the spatial learning and memory and step-down test was used to evaluate the cognitive function.1%Thioflavin-S staining was used to calculate the average number amyloid plaques in cortex and hippocampus.CD31 staining was detected to observe the density of cerebrovascular in hippocampus areas and CD105 staining was further detected to evaluate angiogenesis.Laser Doppler PeriFlux 5000 was further measured the change of cerebrovascular blood flow.ChemDraw was used to draw the molecular structures of five main ingredients of PNS.AlzPlatform were used to estimate the potential targets of PNS.Results:By a bench of behavioral tests,PNS showed a better tendency in proving cognitive functions.In addition,the amyloid plaques in both cortex and hippocampus were significantly reduced after PNS intervention(P<0.05 and P<0.001 respectively).Furthermore,the density of cerebrovascular in the hippocampus areas was increased under PNS administration(P<0.001),which accompanied with angiogenesis in dentate gyrus areas and cerebrovascular blood flow promotion(P<0.05).By AlzPlatform docking serve,we screened five major ingredients of PNS-R1,Rd,Rb1,Re and Rg1.These screening data suggested that vascular related proteins could be the one of potential targets of PNS,such as platelet activating factor receptor and vasopressin V1a receptor.Conclusion:By modulating cerebrovascular function,PNS can reduce the deposition of amyloid plaques and exhibit the role of neuroprotection in a preventive strategy.

Recent Progress in Structural Modification and Activities on Panax notoginseng Saponins

This paper reviews recent progress in the structural modification and activities on Panax notoginseng saponins(PNS).PNS can not only improve the function of cardio-cerebral system,central nervous system and immune system,but also reveal anticancer,anti-aging and anti-oxidation activities.In order to solve the problem of low bioavailability and poor absorbability of PNS in vivo,usually,the researchers modified the structure of PNS with three methods:glycoside cleavage(including acid hydrolysis,sulfation,etc.),biotransformation method(including enzyme hydrolysis,microbial transformation)and combinatorial chemical method.It was found that the structural modification sites of PNS were single,mainly aimed at C-3,C-6 and C-20,which provided a new perspective for the structural modification of PNS.Therefore,structural modification on PNS with high yield and ready availability are significant in the discovery of new active ingredients and industrialization.Derivatives of PNS are applied to research of structure-activity relationship,which is beneficial to the development of new medicines.

Comparison of pharmacokinetics of different oral Panax notoginseng saponins using ultra-high performance liquid mass spectrometry

Objective:To discuss and compare the plasma pharmacokinetics after three oral Panax notoginseng saponin(PNS)administrations in beagle dogs.PNS is the main active ingredient of the traditional Chinese medicine(TCM)Panax notoginseng.Although its outstanding therapeutic efficacy has been demonstrated by various researchers,its broader application is restricted by the low bioavailability of PNS.Methods:An ultra-high performance liquid chromatographyetandem mass spectrometry(UPLC-MS/MS)method for the simultaneous quantification of notoginsenoside R1,ginsenoside Rg1,ginsenoside Rb1,ginsenoside Rd,and ginsenoside Re in beagle dog plasma was developed and validated.The plasma samples were subjected to liquideliquid extraction with acetone and methanol,and separated on an ACQUITY C18 column(100×2.1 mm ID,1.7 mm)using acetonitrile and water as the mobile phase with a run time of 4.5 min.Results:The analytes were detected without interference in Selected Reaction Monitoring mode with positive electrospray ionization.The validated method was successfully used in comparative pharmacokinetic studies of the five saponins in beagle dogs after oral administration of three PNS preparations.Blood samples were collected up to 192 h after administration and pharmacokinetic parameters were calculated using DAS 3.20 and SPSS 17.0.The AUC_(0-t)values of Re and R1 were significantly higher in soft capsules than in hard capsules.However,the AUC_(0-t)values between hard and soft capsules were not significantly different for the other three componentsdRb1,Rd and Rg1.Conclusion:Our intuitive analysis suggests that the bioavailability of PNS in soft capsules is greater than in hard capsules.

Targeted trace ingredients coupled with chemometric analysis for consistency evaluation of Panax notoginseng saponins injectable formulations

Evaluating the consistency of herb injectable formulations could improve their product quality and clinical safety,particularly concerning the composition and content levels of trace ingredients.Panax notoginseng Saponins Injection(PNSI),widely used in China for treating acute cardiovascular diseases,contains low-abundance(10%-25%)and trace saponins in addition to its five main constituents(notoginsenoside R1,ginsenoside Rg1,ginsenoside Re,ginsenoside Rb1,and ginsenoside Rd).This study aimed to establish a robust analytical method and assess the variability in trace saponin levels within PNSI from different vendors and formulation types.To achieve this,a liquid chromatography-triple quadrupole mass spectrometry(LC-MS/MS)method employing multiple ions monitoring(MIM)was developed.A“post-column valve switching”strategy was implemented to eliminate highly abundant peaks(NR_(1),Rg_(1),and Re)at 26 min.A total of 51 saponins in PNSI were quantified or relatively quantified using 18 saponin standards,with digoxin as the internal standard.This study evaluated 119 batches of PNSI from seven vendors,revealing significant variability in trace saponin levels among different vendors and formulation types.These findings highlight the importance of consistent content in low-abundance and trace saponins to ensure product control and clinical safety.Standardization of these ingredients is crucial for maintaining the quality and effectiveness of PNSI in treating acute cardiovascular diseases.

Combination of Total Astragalus Extract and Total Panax Notoginseng Saponins Strengthened the Protective Effects on Brain Damage through Improving Energy Metabolism and Inhibiting Apoptosis after Cerebral Ischemia-Reperfusion in Mice

Objective： To explore the effects and molecular mechanisms of the combination between total Astragalus extract （TAE） and total Panax notoginseng saponins （TPNS） against cerebral ischemia- reperfusion injury. Methods： C57BL/6 mice were randomly divided into sham-operated group, model group, TAE （110 mg/kg） group, TPNS （115 mg/kg） group, TAE-TPNS combination group and Edaravone （4 mg/kg） group, treated for 4 days, then, cerebral ischemia-repeffusion injury was established by bilateral common carotid artery （CCA） ligation for 20 min followed by reperfusion for 1 and 24 h. Results： TPNS could increase adenosine triphosphate （ATP） level, TAE and TAE-TPNS combination increased ATP, adenosine diphosphate （ADP） contents and Na＋-K＋-ATPase activity, and the effects of TAE-TPNS combination were stronger than those of TAE or TPNS alone after reperfusion for 1 h. After reperfusion for 24 h, TAE, TPNS and TAE-TPNS combination significantly increased neurocyte survival rate and decreased the apoptosis rate as well as down-regulated the expression of phosphorylated c-June N-terminal kinasel/2 （p-JNK1/2）, cytochrome C （Cyt C）, cysteine aspartic acid-specific protease （Caspase）-9 and Caspase-3. Furthermore, the effects in TAE-TPNS combination were better than those in TAE or TPNS alone. Conclusion： The combination of TAE 110 mg/kg and TPNS 115 mg/kg could strengthen protective effects on cerebral ischemia injury, the mechanism underlying might be related to improving jointly the early energy metabolism, and relieving the delayed apoptosis via inhibiting the mitochondrial apoptosis pathway of JNK signal transduction.

Panax notoginseng saponins prevent colitis-associated colorectal cancer development: the role of gut microbiota

Gut microbiota dysbiosis is a risk factor for colorectal cancer(CRC) in inflammatory bowel disease(IBD).In this study, the effects of Panax notoginseng saponins(PNS) on colitis-associated CRC progression were evaluated on an azoxymethane(AOM)/dextran sulfate sodium(DSS) mouse model.In vivo, PNS significantly relieved AOM/DSS-induced colon tumorigenesis and development by reducing the disease activity index(DAI) scores and colon tumor load.The 16S rRNA data of fecal samples showed that the microbiome community was obviously destructed, while PNS could recover the richness and diversity of gut microbiota.Especially, PNS could increase the abundance of Akkermansia spp.which was significantly decreased in model group and negatively correlated with the progression of CRC.Moreover, ginsenoside compound K(GC-K) was evaluated on the effects of human CRC cells,which was the main bio-transformed metabolite of PNS by gut microbiota.Our data showed that PNS played important role in the prevention of the progression of CRC, due to their regulation on the microbiome balance and microbial bio-converted product with antiCRC activity.

Panax Notoginseng Saponins Inhibits Atherosclerotic Plaque Angiogenesis by Down-Regulating Vascular Endothelial Growth Factor and Nicotinamide Adenine Dinucleotide Phosphate Oxidase Subunit 4 Expression

Objective： To investigate the mechanism of Panax notoginseng saponins （PNS）, an effective component extracted from Panax notoginseng, on atherosclerotic plaque angiogenesis in atherosclerosis-prone apolipoprotein E-knockout （ApoE-KO） mice fed with high-fat, high-cholesterol diet. Methods： Twenty ApoE-KO mice were divided into two groups, the model group and the PNS group. Ten normal C57BL/6J mice were used as a control group. PNS （60 mg/kg） was orally administered daily for 12 weeks in the PNS group, The ratio of plaque area to vessel area was examined by histological staining. The tissue sample of aortic root was used to detect the CD34 and vascular endothelial growth factor （VEGF） expression areas by immunohistochemistry. The expression of VEGF and nicotinamide adenine dinucleotide phosphate oxidase subunit 4 （NOX4） were measured by reverse transcription polymerase chain reaction and Westem blotting respectively. Results： After treatment with PNS, the plaque areas were decreased （P〈0.05）. CD34 expressing areas and VEGF expression areas in plaques were significantly decreased （P〈0.05）. Meanwhile, VEGF and NOX4 mRNA expression were decreased after treatment with PNS, VEGF and NOX4 protein expression were also decreased by about 72% and 63%, respectively （P〈0.01）. Conclusion： PNS, which decreases VEGF and NOX4 expression, could alleviate plaque angiogenesis and attenuate atherosclerosis.

Panax Notoginseng Saponins Induced Up-Regulation,Phosphorylation and Binding Activity of MEK,ERK,AKT,PI-3K Protein Kinases and GATA Transcription Factors in Hematopoietic Cells

Objective： To investigate the effects of panax notoginseng saponins （PNS） on expression, regulation and phosphorylation of multiple protein kinases in mitogen activated protein kinase （MAPK） intracellular signal pathway and GATA transcription factors in hematopoietic cells, so as to explore its mechanism of proliferation and differentiation activity on hematopoiesis. Methods： The human granulocytic HL-60, erythrocytic K562, megakaryocytic CHRF-288 and Meg-01 cell lines were treated by PNS, the positive control of K562, CHRF-288 ceils treated by recombination human erythropoietin （Epo） and thrombopoietin （Tpo） respectively. The total cell lysate and nuclei protein were extracted after being treated by PNS, subsequently, analyzed by both Western blot and immune-precipitation. Meanwhile, the nuclei extract was performed for electrophoretic mobility shift assay （EMSA） by using 32p radio labeled double-stranded GATA consensus oligonucleotide. Results： The expression levels of kinase MEK-1, MEK-2, ERK-1, ERK-2, AKT-1, AKT-2 and PI- 3K were increased by PNS treatment to different extent in four cell lines, depending on cellular heterogeneity and sensitivity to PNS, also phosphorylation of MEK-1, ERK-1 was differentially promoted by PNS respectively （P〈0.05, 0.01, 0.001）. The expression levels of transcription factors GATA-1 and GATA-2 were increased, moreover, their DNA binding activities were raised dramatically in PNS treated K562, CHRF-288 and Meg-01 cells compared with the controls respectively （P〈0.05, 0.01, 0.001）. The positive control of K562, CHRF-288 cells treated by Epo or Tpo respectively also displayed up-regulation of protein kinases and GATA transcription factors respectively （P〈0.05, 0.01, 0.001）. Conclusion： The results indicated that intracellular signal pathway initiated by PNS was involved in MAPK pathway and transcription factors of GATA family in hematopoietic cells. PNS displayed the role to promote proliferation and differentiation, by means of increasing expression level and phosphorylation status of multiple protein kinases, also inducing synthesis of GATA transcription factors and up- regulation its DNA binding activity.

Panax Notoginseng Saponins Inhibits Ventricular Remodeling after Myocardial Infarction in Rats Through Regulating ATF3/MAP2K3/p38 MAPK and NFκB Pathway

Objective:To explore whether Panax notoginseng saponins(PNS)exhibits heart protective effect in myocardial infarction(MI)rats and to identify the potential signaling pathways involved.Methods:MI rats induced by ligating the left anterior descending(LAD)coronary artery were assigned to sham coronary artery ligation or coronary artery ligation.Totally 36 Sprague-Dawley rats were randomly divided into sham group(distilled water,n=9),MI group(distilled water,n=9),PNS group(PNS,40 mg/kg daily,n=9)and fosinopril group(FIP,1.2 mg/kg daily,n=9)according to a random number table.The left ventricular morphology and function were conducted by echocardiography.Histological alterations were evaluated by the stainings of HE and Masson.The serum levels of C reactive protein(CRP),tumor necrosis factorα(TNF-α),growth differentiation factor-15(GDF-15)and the ratio of metalloproteinase-9(MMP-9)and tissue inhibitor of MMP-9(TIMP-1)were determined by ELISA.The levels of activating transcription factor 3(ATF3),mitogen-activated protein kinase kinase 3(MAP2 K3),p38 mitogen-activated protein kinase(p38 MAPK),phosphorylation of p38 MAPK(p-p38 MAPK),transforming growth factor-β(TGF-β1),collagenⅠ,nuclear factor kappa B p65(NFκB p65),phosphorylation of NFκB p65(p-NFκB p65),and phosphorylation of inhibitory kappa Bα(p-IκBα)in hearts were measured by Western blot and immunohistochemical staining,respectively.Results:PNS improved cardiac function and fibrosis in MI rats(P<0.05).The serum levels of CRP,TNF-α,GDF-15 and the ratio of MMP9/TIMP1 were reversed by PNS in MI rats.The expressions of TGF-β1,collagenⅠ,MAP2 K3,p38 MAPK,p-p38 MAPK,NFκB p65,p-NFκB p65,and p-IκBαwere down-regulated,while ATF3 increased with the treatment of PNS(P<0.05).Conclusions:PNS may improve cardiac function and fibrosis in MI rats via regulating ATF3/MAP2 K3/p38 MAPK and NFκB signaling pathways.These results suggest the potential of PNS in preventing the development of ventricular remodeling in MI rats.

Pharmacokinetics and correlation between in vitro release and in vivo absorption of bio-adhesive pellets of panax notoginseng saponins

The present study was designed to prepare and compare bio-adhesive pellets of panax notoginseng saponins(PNS) with hydroxy propyl methyl cellulose(HPMC), chitosan, and chitosan : carbomer, explore the influence of different bio-adhesive materials on pharmacokinetics behaviors of PNSbio-adhesive pellets, and evaluate the correlation between in vivo absorption and in vitro release(IVIVC). In order to predict the in vivo concentration-time profile by the in vitro release data of bio-adhesive pellets, the release experiment was performed using the rotating basket method in p H 6.8 phosphate buffer. The PNS concentrations in rat plasma were analyzed by HPLC-MS-MS method and the relative bioavailability and other pharmacokinetic parameters were estimated using Kinetica4.4 pharmacokinetic software. Numerical deconvolution method was used to evaluate IVIVC. Our results indicated that, compared with ordinary pellets, PNS bio-adhesive pellets showed increased oral bioavailability by 1.45 to 3.20 times, increased Cmax, and extended MRT. What's more, the release behavior of drug in HPMC pellets was shown to follow a Fickian diffusion mechanism, a synergetic function of diffusion and skeleton corrosion. The in vitro release and the in vivo biological activity had a good correlation, demonstrating that the PNS bio-adhesive pellets had a better sustained release. Numerical deconvolution technique showed the advantage in evaluation of IVIVC for self-designed bio-adhesive pellets with HPMC. In conclusion, the in vitro release data of bio-adhesive pellets with HPMC can predict its concentration-time profile in vivo.

Effects of Panax Notoginseng Saponins on Proliferation and Differentiation in NIH3T3 Cells

Objective： To investigate the effects of Panax notoginseng saponins （PNS） on the proliferation and differentiation in NIH3T3 cells. Methods： NIH3T3 cells were treated by various concentrations of PNS 0, 0.05, 0.10, 0.20, and 0.40 g/L. The vitality and proliferation potential of cells were detected by 3-（4,5-dimethylthiazol- 2-yl）-2, 5-diphenyltetrazolium bromide （MTT） assay, the alkaline phosphatase （ALP） activity was measured by p-nitrophenyl phosphate （pNPP） assay, and the mineralization formation ability was tested for the cellular differentiation toward osteoblast, as well as the expression level of phosphorylated extrecellular signal-regulated kinasel/2（P-ERK1/2）, extracellular signal-regulated kinasel/2 （ERK1/2） protein kinase was analyzed by Western blot with total cell lysate of NIH3T3 cells treated by PNS. Results： Both MTT and pNPP assay showed that optical density （OD） values were increased in response to PNS treatment at a dose-dependent pattern. The mineralization formation ability was enhanced in PNS-treated NIH3T3 cells compared with untreated cells. Meanwhile, the expression level of P-ERK1/2 protein kinase was up-regulated in PNS-treated NIH3T3 cells, while, the expression level of ERK1/2 protein kinase revealed no obvious difference with or without PNS treated cells. Conclusion： PNS could pay a role to promote the proliferation and differentiation in NIH3T3 cells by means of up-regulation of P-ERK1/2 protein kinase.

Panax notoginseng saponins prevent colitis-associated colorectal cancer via inhibition IDO1 mediated immune regulation

Colorectal cancer(CRC)is the third most lethal cancer and leading cause of cancer mortality worldwide.A key driver of CRC development is colon inflammatory responses especially in patients with inflammatory bowl disease(IBD).It has been proved that Panax notoginseng saponins(PNS)have anti-inflammatory,anti-oxidant and anti-tumor effects.The chemopreventive and immunomodulatory functions of PNS on colitis-associated colorectal cancer(CAC)have not been evaluated.This present study was designed to study the potential protective effects of PNS on AOM/DSS-induced CAC mice to explore the possible mechanism of PNS against CAC.Our study showed that PNS significantly alleviated colitis severity and prevented the occurrence of CAC.Functional assays revealed that PNS relieved immunosuppression of Treg cells in the CAC microenvironment by inhibiting the expression of IDO 1 mediated directly by signal transducer and activator of transcription 1(STAT1)rather than phosphorylated STAT1.Ultimately,Rhl,one of the PNS metabolites,exhibited the best inhibitory effect on IDO1 enzyme activity.Our study showed that PNS exerted significant chemopreventive function and immunomodulatory properties on CAC.It could reduce macrophages accumulation and Treg cells differentiation to reshape the immune microenvironment of CAC.These findings provided a promising approach for CAC intervention.