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.

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.

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 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.

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.

Prevention of Deep Vein Thrombosis by Panax Notoginseng Saponins Combined with Low-Molecular-Weight Heparin in Surgical Patients

Objective:To evaluate the efficacy of deep vein thrombosis(DVT)prevention among realworld surgical inpatients who received panax notoginseng saponins(PNS)combined with low-molecularweight heparin(LMWH).Methods:A prospective cohort study was conducted among surgical patients between January 2016 and November 2018 in Xuanwu Hospital,Capital Medical University,Beijing,China.Participants received LMWH alone or PNS combined with LMWH for preventing DVT.The primary outcome was incidence of lower extremity DVT,which was screened once a week.Participants in the LMWH group were given LMWH(enoxaparin)via hypodermic injection,4000-8000 AxalU once daily.Participants in the exposure group received PNS(Xuesaitong oral tablets,100 mg,3 times daily)combined with LMWH given the same as LMWH group.Results:Of the 325 patients screened for the study,281 participants were included in the final analysis.The cohort was divided into PNS+LMWH group and LMWH group with 134 and 147 participants,respectively.There was a significant difference of DVT incidence between two groups(P=0.01),with 21(15.7%)incident DVT in the PNS+LMWH group,and 41(27.9%)incident DVT in the LMWH group.Compared with participants without DVT,the participants diagnosed with DVT were older and had higher D-dimer level.The multivariate logistic regression moclel showed a significant lower risk of incident DVT among participants in the PNS+LMWH group compared with the LMWH group(odds ratio 0.46,95%confidence interval,0.25-0.86).There were no significant differences in thromboelaslography values(including R,K,Angle,and MA)and differences in severe bleeding between two groups.No symptomatic pulmonary embolism occurred during the study.Conclusion:Combined application of PNS and LMWH can effectively reduce the incidence of DVT among surgical inpatients compared with LMWH monotherapy,without increased risk of bleeding.

Panax Notoginseng Saponins Increases the Blood Concentration of Nifedipine by Inhibiting CYP3A4 Enzyme through PXR-and CAR-Mediated Pathway

Objective:To explore the mechanism underlying the effect of Panax notoginseng saponins(PNS)on the pharmacokinetics of nifedipine(NF)in rats.Materials and Methods:Twenty-four rats were randomly divided into blank(BL)group,PNS group,NF group,and PNS+NF group,with six rats in each group.Noncompartmental analysis and t-test were carried out to determine the difference between the pharmacokinetic parameters of NF in different groups.CYP3 A4 enzyme activity was calculated using the probe drug method.The mRNA and protein contents of CYP3 A4,nuclear receptor CAR,and PXR in rat liver were quantitatively analyzed by qRT-PCR and Western blot.Results:After the rats were treated with the combination of PNS and NF,the plasma concentration,half-life,peak time,and area under the concentration-time curve of NF increased,whereas the clearance rate decreased.The inhibitory effect on CYP3 A4 enzyme activity was in the following order:PNS+NF group(strongest)>PNS group>NF group,and BL group(weakest).Similar changes were observed for the inhibitory effect on CYP3 A4,CAR,and PXR mRNA and protein content,and the order was as follows:PNS+NF group(weakest)<PNS group<NF group,and BL group(strongest).Conclusion:In combination with NF,PNS may inhibit the mRNA and protein expression of nuclear receptor CAR and PXR and the activity of CYP3 A4 enzyme,slowing down the pharmacokinetics of NF in rats,increasing its blood concentration,and enhancing the therapeutic effect of NF.

SIMULTANEOUS QUANTITATION OF FIVE PANAX NOTOGINSENG SAPONINS IN EIGHT CHINESE PATENT MEDICINES BY MULTI HEART-CUTTING TWODIMENSIONAL LIQUID CHROMATOGRAPHY

Current China Pharmacopoeia(ChP)standards employ diversified and case-dependent assay methods to evaluate the quality of different Chinese patent medicines(CPM)that contain Panax notoginseng as the monarch drug.This study is aimed to develop a two-dimensional liquid chromatography(2D LC)based assay approach to unify and simplify the quality assessment of Notoginseng containing CPMs.Multi heart-cutting 2D LC(MHC-2D LC)was used to separate and quantify five major

Transformation of Compound K from Saponins in Leaves of Panax notoginseng by Immobilized β-Glucanase

Objective To prepare an active anti-tumor component,compound K(C-K),from saponins in leaves of Panax notoginseng(SLPN) using immobilized β-glucanase.Methods Two entrapments,alginate gel-1(Alg 1) and alginate gel-2(Alg 2),were evaluated for their ability to immobilize β-glucanase.The amount and purity of C-K obtained from the transformation process were analyzed by HPLC,and the immobilizing parameters were optimized.Results β-Glucanase can be immobilized and reused with either of the entrapment.However,using Alg 1 resulted in higher enzyme activity than Alg 2.The optimal concentration of the immobilized enzyme was 10%;The optimal crosslinking time was 4-6 h;and the optimal concentration of the crosslinking agent was 6%- 7%.Conclusion Immobilized β-glucanase shows sustained enzyme activity,good ethanol tolerance,and was reusable for the preparation of C-K from SLPN.

Total Saponins of Panax notoginseng Activate Akt/mTOR Pathway and Exhibit Neuroprotection in vitro and in vivo against Ischemic Damage

Objective:To reveal the neuroprotective effect and the underlying mechanisms of a mixture of the main components of Panax notoginseng saponins(TSPN)on cerebral ischemia-reperfusion injury and oxygenglucose deprivation/reoxygenation(OGD/R)of cultured cortical neurons.Methods:The neuroprotective effect of TSPN was evaluated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide(MTT)assay,flow cytometry and live/dead cell assays.The morphology of dendrites was detected by immunofluorescence.Middle cerebral artery occlusion(MCAO)was developed in rats as a model of cerebral ischemia-reperfusion.The neuroprotective effect of TSPN was evaluated by neurological scoring,tail suspension test,2,3,5-triphenyltetrazolium chloride(TTC)and Nissl stainings.Western blot analysis,immunohistochemistry and immunofluorescence were used to measure the changes in the Akt/mammalian target of rapamycin(mTOR)signaling pathway.Results:MTT showed that TSPN(50,25 and 12.5μg/m L)protected cortical neurons after OGD/R treatment(P<0.01 or P<0.05).Flow cytometry and live/dead cell assays indicated that 25μg/m L TSPN decreased neuronal apoptosis(P<0.05),and immunofluorescence showed that 25μg/m L TSPN restored the dendritic morphology of damaged neurons(P<0.05).Moreover,12.5μg/m L TSPN downregulated the expression of Beclin-1,Cleaved-caspase 3 and LC3 B-Ⅱ/LC3 B-Ⅰ,and upregulated the levels of phosphorylated(p)-Akt and p-mTOR(P<0.01 or P<0.05).In the MCAO model,50μg/m L TSPN improved defective neurological behavior and reduced infarct volume(P<0.05).Moreover,the expression of Beclin-1 and LC3 B in cerebral ischemic penumbra was downregulated after 50μg/m L TSPN treatment,whereas the p-mTOR level was upregulated(P<0.05 or P<0.01).Conclusions:TSPN promoted neuronal survival and protected dendrite integrity after OGD/R and had a potential therapeutic effect by alleviating neurological deficits and reversing neuronal loss.TSPN promoted p-mTOR and inhibited Beclin-1 to alleviate ischemic damage,which may be the mechanism that underlies the neuroprotective activity of TSPN.

Design of Traditional Chinese Medicine Extraction Workshop Process and Automation System

Objective: This paper takes the example of a Panax notoginseng extraction workshop and designs an automated production workshop with advanced domestic capabilities. Methods: 1) Based on the small-scale Panax notoginseng extraction process, the feasibility of the workshop production process is demonstrated. 2) The workshop process design for Panax Notoginseng saponin extraction is completed, including production organization plans and the selection of key equipment. 3) For the Panax notoginseng extraction workshop process, an automated production control system is designed. Conclusion: Through optimized design of the production process and automation system, continuous and automated production of traditional Chinese medicine extraction is achieved, leading to improvements in drug quality and production efficiency.

三七总皂苷药理作用及临床应用研究进展

三七总皂苷(Panax notoginseng saponins,PNS)是三七(Panax notoginseng)的主要活性成分,具有抗炎、降血脂、抗氧化和抗肿瘤等药理作用。在查阅近年来国内外文献的基础上,对三七总皂苷的药理作用和临床应用进行了综述,为其进一步研究和应用提供参考。

Panax Notoginseng Saponin Attenuates Gastric Mucosal Epithelial Cell Injury Induced by Dual Antiplatelet Drugs through COX and PI3K/Akt/ VEGF-GSK-3β-RhoA Network Pathway

Objective To elucidate the underlying mechanism of Panax notoginseng saponin(PNS)on gastric epithelial cell injury and barrier dysfunction induced by dual antiplatelet(DA).Methods Human gastric mucosal epithelial cell(GES-1)was cultured and divided into 4 groups:a control,a DA,a PNS+DA and a LY294002+PNS+DA group.GES-1 apoptosis was detected by flow cytometry,cell permeability were detected using Transwell,level of prostaglandins E2(PGE2),6-keto-prostaglandin F1α(6-keto-PGF1α)and vascular endothelial growth factor(VEGF)in supernatant were measured by enzyme linked immunosorbent assay(ELISA),expression of phosphatidylinositide 3-kinase(PI3K),phosphorylated-PI3K(p-PI3K),Akt,phosphorylated-Akt(p-Akt),cyclooxygenase-1(COX-1),cyclooxygenase-2(COX-2),glycogen synthase kinase-3β(GSK-3β)and Ras homolog gene family member A(RhoA)were measured by Western-blot.Results DA induced apoptosis and hyper-permeability in GES-1,reduced supernatant level of PGE2,6-keto-PGF1αand VEGF(P<0.05).Addition of PNS reduced the apoptosis of GES-1 caused by DA,restored the concentration of PGE2,6-keto-PGF1αand VEGF(P<0.05).In addition,PNS attenuated the alteration of COX-1 and COX-2 expression induced by DA,up-regulated p-PI3K/p-Akt,down-regulated RhoA and GSK-3β.LY294002 mitigated the effects of PNS on cell apoptosis,cell permeability,VEGF concentration,and expression of RhoA and GSK-3βsignificantly.Conclusions PNS attenuates the suppression on COX/PG pathway from DA,alleviates DA-induced GES-1 apoptosis and barrier dysfunction through PI3K/Akt/VEGF-GSK-3β-RhoA network pathway.

Xueshuantong for Injection(Lyophilized,注射用血栓通)Alleviates Streptozotocin-Induced Diabetic Retinopathy in Rats

Objective To investigate the ameliorate effect and underlying mechanism of Xueshuantong for Injection(Lyophilized,注射用血栓通,XST)in streptozocin(STZ)-induced diabetic retinopathy(DR)rats.Methods Diabetes mellitus(DM)model was induced by intraperitoneal(i.p.)injection of STZ(60 mg/kg)in Sprague-Dawley rats.Diabetic rats were randomized into 3 groups(n=10)according to a random number table,including DM,XST50 and XST100 groups.XST treatment groups were daily i.p.injected with 50 or 100 mg/kg XST for 60 days,respectively.The control and DM groups were given i.p.injection with saline.Blood glucose level and body weight were recorded every week.Histological changes in the retina tissues were observed with hematoxylin-eosin staining.Apoptosis and inflammation related factors,including cleaved caspase-3,glial fifibrillary acidic protein(GFAP),tumor necrosis factor-α(TNF-α)and intercellular cell adhesion molecule-1(ICAM-1)were detected by Western blot or real-time polymerase chain reaction.Then,the levels of advanced glycation end product(AGE)and its receptor(RAGE)were investigated.Tight junctions proteins(Zonula occludens-1(ZO-1),Occludin and Claudin-5)of blood-retinal barrier were detected by Western blot.The levels of retinal fifibrosis,transforming growth factor-β1(TGF-β1)-Smad2/3 signaling pathway were evaluated at last.Results There was no signifificant difference in the body weight and blood glucose level between XST and DM groups(P>0.05).Compared with the DM group,XST treatment signifificantly increased the retinal thickness of rats(P<0.05 or P<0.01),and suppressed cleaved caspase-3 expression(P<0.01).XST increased the protein expressions of ZO-1,Occludin and Claudin-5 and decreased the mRNA expressions of matrix metalloproteinase 2(MMP-2)and MMP-9(P<0.05 or P<0.01).Moreover,XST signifificantly reduced the productions of AGE and RAGE proteins in the retina of rats(P<0.05 or P<0.01),suppressed the over-expression of TNF-α,and decreased the elevated level of ICAM-1 in retina of rats(P<0.05 or P<0.01).XST signifificantly reduced the levels ofα-smooth muscle actin(α-SMA),connective tissue growth factor(CTGF),TGF-β1 and phosphorylation of Smad2/3 protein in rats(P<0.05 or P<0.01).Conclusions XST had protective effects on DR with possible mechanisms of inhibiting the inflammation and apoptosis,up-regulating the expression of tight junction proteins,suppressing the productions of AGE and RAGE proteins,and blocking the TGF-β/Smad2/3 signaling pathway.XST treatment might play a role for the future therapeutic strategy against DR.

Attenuation of Brain Inflammatory Response after Focal Cerebral Ischemia/Reperfusion with Xuesaitong Injection (血塞通注射液) in Rats

Objective: To investigate the neuro-protective effect of Xuesaitong Injection (血塞通注射液, XST) on brain inflammatory response after transient focal cerebral ischemia/reperfusion in rats. Methods: Focal cerebral ischemia/reperfusion models of male rats were induced by transient occlusion for 2 h of middle cerebral artery (MCA) which was followed by 24 h reperfusion. XST was administered through intraperitoneal injection of 25 mg/kg or 50 mg/kg at 4 h after the onset of ischemia. After reperfusion for 24 h, the neurological function score was evaluated, the brain edema was detected with dry-wet weight method, the myeloperoxidase (MPO) activity and the expression of intercellular adhesion molecule-1(ICAM-1) of ischemic cerebral cortex and caudate putamen was determined by spectrophotometry and immunohistochemistry respectively. Results: XST not only lowered neurological function score at the dose of 50 mg/kg, but reduced brain edema and inhibited MPO activity and ICAM-1 expression as compared with the ischemia/reperfusion model group (P<0.01).Conclusion: XST has a definite effect on inhibiting the expression of ICAM-1 and neutrophil infiltration in rats with cerebral ischemia/reperfusion when treatment started at 4 h after ischemia onset, and also attenuates inflammation in the infarcted cerebral area.