Metabolomics is a newly developed discipline following genomics,transcriptomics,and proteomics.It is based on the theory of systems biology,supported by modern spectroscopy analysis technology and is analyzed by the e...Metabolomics is a newly developed discipline following genomics,transcriptomics,and proteomics.It is based on the theory of systems biology,supported by modern spectroscopy analysis technology and is analyzed by the external stimulation of organisms.By analyzing the overall changes of metabolites with low molecular weight in organism after being stimulated by the outside world,we can understand the physiological and pathological state of organism,and finally clarify the essence of biological changes of organism.In recent years,metabolomics has developed rapidly.It has been widely used in early toxicity screening of lead compounds,preclinical toxicity of drugs and toxicity evaluation of clinical drugs.The in-depth interpretation of metabolomics results is also developing.This article reviewed the progress of metabolomics technology and its application in the study of toxicity evaluation of traditional Chinese medicine.展开更多
Traditional Chinese medicine (TCM) is a medical system that has collected and summarized abundant clinical experience over its long history of more than 2000 years. However, the frequent occurrence of TCM-induced adve...Traditional Chinese medicine (TCM) is a medical system that has collected and summarized abundant clinical experience over its long history of more than 2000 years. However, the frequent occurrence of TCM-induced adverse reactions has hindered the modernization and internationalization of TCM, while attracting increasing attention from around the world. Unlike chemical drugs and biological agents, the difficulties involved in research on the toxicity and safety of TCM mainly include the complexity of its components and the unpredictability of drug–body interactions. Much of TCM, which has overall therapeutic effects, has the typical mechanisms of multiple components, multiple pathways, and multiple targets. While considering the gradualness and unpredictability of TCM toxicity, the ambiguity of toxicants and safe dosage, and individual differences during long-term TCM administration, we have systematically established key techniques for the toxicity assessment of TCM. These techniques mainly include TCM toxicity discovery in an early phase, based on a combination of drug toxicology genomics and metabolomics;methods to identify dose–toxicity relationships in TCM;and integrated techniques for the exploration of TCM interactions, such as fast-screening tests based on drug-metabolizing enzymes and receptor pathways. In particular, we have developed a new technical system for TCM safety evaluation using molecular toxicology, which has been validated well in research on TCM compatibility contraindication, quality control, and allergen discovery. The application of this key technical platform is introduced here in detail. This application includes model organisms, toxicant biomarkers, a magnetic suspension technique, and the application of network toxicology and computational toxicology in research on the toxicity of Fructus toosendan, Semen cassiae, Polygonum multiflorum, and Fructus psoraleae.展开更多
AIM:To investigate the hepatoprotective effects and mechanisms of an extract of Salvia miltiorrhiza and Carthamus tinctorius in vivo.METHODS:C57BL/6J mice were randomly assigned to five groups and intraperitoneally ad...AIM:To investigate the hepatoprotective effects and mechanisms of an extract of Salvia miltiorrhiza and Carthamus tinctorius in vivo.METHODS:C57BL/6J mice were randomly assigned to five groups and intraperitoneally administered 0.9% saline,Salvia miltiorrhiza and Carthamus tinctorius extract [Danhong injection(DHI),0.75 and 3 g/kg mixed extract] or reduced glutathione for injection(RGI,300 mg/kg) for 30 min before exposure to lipopolysaccharide(LPS,16 mg/kg). After intraperitoneal LPS stimulation for 90 min or 6 h,the mice were sacrificed by ether anaesthesia,and serum and liver samples were collected. Histological analysis(H&E) and terminal deoxynucleotidyl transferase-mediated d UTP nick end-labelling(TUNEL) staining were performed. Alanine transferase(ALT),aspartate transaminase(AST),total bilirubin(TBil),glutathione-S-transferase(GST),malondialdehyde(MDA),tumour necrosis factor(TNF)-α,interleukin(IL)-6,and caspase-3 levels were measured. Bax,Bcl-2,P-IκBα,IκBα,P-NF-κB p65,and NF-κB p65 protein levels were determined by Western blot. TNF-α,IL-6,caspase-3,Bax and Bcl-2 m RNA expression was measured by real-time reverse transcription-polymerase chain reaction(RT-PCR).RESULTS:Hematoxylin-eosin staining and TUNEL results suggested that DHI(3 g/kg) treatment alleviated inflammatory and apoptotic(P < 0.01) injury in the liver of mice. DHI treatment dose-dependently blunted the abnormal changes in biochemical parameters such as ALT(72.53 ± 2.83 for 3 g/kg,P < 0.01),AST(76.97 ± 5.00 for 3 g/kg,P < 0.01),TBil(1.17 ± 0.10 for 3 g/kg,P < 0.01),MDA(0.81 ± 0.36 for 3 g/kg,P < 0.01),and GST(358.86 ± 12.09 for 3 g/kg,P < 0.01). Moreover,DHI(3 g/kg) remarkably decreased LPS-induced protein expression of TNF-α(340.55 ± 10.18 for 3 g/kg,P < 0.01),IL-6(261.34 ± 10.18 for 3 g/kg,P < 0.01),and enzyme activity of caspase-3(0.93 ± 0.029 for 3 g/kg,P < 0.01). The LPS-induced m RNA expression of TNF-α,IL-6 and caspase-3 was also decreased by DHI. Western blot analysis revealed that DHI antagonised LPS-stimulated decrease of Bcl-2 and increase of Bax protein expression. Furthermore,DHI inhibited LPS-induced IκBα and NF-κB p65 phosphorylation.CONCLUSION:DHI may be a multi-function protectant against acute hepatic injury in mice through its antiinflammatory,anti-oxidative and anti-apoptotic activities.展开更多
Inherent complexity of plant metabolites necessitates the use of multi-dimensional information to accomplish comprehensive profiling and confirmative identification.A dimension-enhanced strategy,by offline two-dimensi...Inherent complexity of plant metabolites necessitates the use of multi-dimensional information to accomplish comprehensive profiling and confirmative identification.A dimension-enhanced strategy,by offline two-dimensional liquid chromatography/ion mobility-quadrupole time-of-flight mass spectrometry(2 D-LC/IM-QTOF-MS)enabling four-dimensional separations(2 D-LC,IM,and MS),is proposed.In combination with in-house database-driven automated peak annotation,this strategy was utilized to characterize ginsenosides simultaneously from white ginseng(WG)and red ginseng(RG).An offline 2 DLC system configuring an Xbridge Amide column and an HSS T3 column showed orthogonality 0.76 in the resolution of ginsenosides.Ginsenoside analysis was performed by data-independent high-definition MSE(HDMSE)in the negative ESI mode on a Vion?IMS-QTOF hybrid high-resolution mass spectrometer,which could better resolve ginsenosides than MSEand directly give the CCS information.An in-house ginsenoside database recording 504 known ginsenosides and 58 reference compounds,was established to assist the identification of ginsenosides.Streamlined workflows,by applying UNIFI?to automatedly annotate the HDMSEdata,were proposed.We could separate and characterize 323 ginsenosides(including 286 from WG and 306 from RG),and 125 thereof may have not been isolated from the Panax genus.The established 2 D-LC/IM-QTOF-HDMSEapproach could also act as a magnifier to probe differentiated components between WG and RG.Compared with conventional approaches,this dimensionenhanced strategy could better resolve coeluting herbal components and more efficiently,more reliably identify the multicomponents,which,we believe,offers more possibilities for the systematic exposure and confirmative identification of plant metabolites.展开更多
Baicalein(BE) is one of the main active flavonoids representing the variety of pharmacological effects including anticancer, anti-inflammatory and cardiovascular protective activities, but it's very low solubility...Baicalein(BE) is one of the main active flavonoids representing the variety of pharmacological effects including anticancer, anti-inflammatory and cardiovascular protective activities, but it's very low solubility, dissolution rate and poor oral absorption limit the therapeutic applications. In this work, a nano-cocrystal strategy was successfully applied to improve the dissolution rate and bioavailability of BE. Baicalein-nicotinamide(BE-NCT) nanococrystals were prepared by high pressure homogenization and evaluated both in vitro and in vivo. Physical characterization results including scanning electron microscopy, dynamic light scattering, powder X-ray diffraction and differential scanning calorimetry demonstrated that BE-NCT nano-cocrystals were changed into amorphous state with mean particle size of 251.53 nm. In the dissolution test, the BE-NCT nano-cocrystals performed 2.17-fold and 2.54-fold enhancement than BE coarse powder in FaSSIF-V2 and FaSSGF. Upon oral administration, the integrated AUC0-t of BE-NCT nano-cocrystals(6.02-fold) was significantly higher than BE coarse powder(1-fold), BE-NCT cocrystals(2.87-fold) and BE nanocrystals(3.32-fold). Compared with BE coarse powder, BE-NCT cocrystals and BE nanocrystals, BENCT nano-cocrystals possessed excellent performance both in vitro and in vivo evaluations.Thus, it can be seen that nano-cocrystal is an appropriate novel strategy for improving dissolution rate and bioavailability of poor soluble natural products such as BE.展开更多
OBJECTIVE To highlight the pharmacological effects of rosemary and its active compounds and eluci⁃date its related mechanisms in nonalcoholic fatty liver disease(NAFLD)management both in vitro and in vivo.METHODS In o...OBJECTIVE To highlight the pharmacological effects of rosemary and its active compounds and eluci⁃date its related mechanisms in nonalcoholic fatty liver disease(NAFLD)management both in vitro and in vivo.METHODS In orotic acid induced NAFLD rats model,rats were administrated with 100,200 and 400 mg·kg^-1 rosemary ethanol extract(RO),10,25 and 50 mg·kg^-1 rosemary acid(RA),and 5,10 and 25 mg·kg^-1 carnosic acid(CA)for three weeks respec⁃tively.Sodium oleate induced HepG2 cell model was used to study the regulation effect of rosemary ethanol extract and its main metabolites on fat accumulation.lipid metabolism related gene expression was analyzed by Western blotting and real-time PCR to clarify the specific molecular mechanism of RO,RA and CA in lipid accumulation.RESULTS RO,RA and CA significantly reduced the contents of liver triglyceride(TG),total cholesterol(TC),free fatty acids(FFA)and improved cell hypertrophy,vacuolation,and cell necrosis in liver of orotic acid induced NAFLD model rats.The mecha⁃nism and related pathways of RO and its main metabolites against lipid disorder was related to the up-regulation of the phosphorylation of adenosine 5′-monophosphate(AMP)-activated protein kinase(AMPK)and inhibition of the sterol regu⁃latory element binding protein-1c(SREBP-1c)cracking into the nuclear,following down-regulation of fatty acid synthesis.CONCLUSION The rosemary has effectively function to regulate lipid metabolism through AMPK/SREBP1c signaling pathway.展开更多
Blindness and vision impairment are the most devastating global health problems resulting in a substantial economic and social burden.Delivery of drug to particular parts of the anterior or posterior segment has been ...Blindness and vision impairment are the most devastating global health problems resulting in a substantial economic and social burden.Delivery of drug to particular parts of the anterior or posterior segment has been a major challenge due to various protective barriers and elimination mechanisms associated with the unique anatomical and physiological nature of the ocular system.Drug administration to the eye by conventional delivery systems results in poor ocular bioavailability(<5%).The designing of a novel approach for a safe,simple,and effective ocular drug delivery is a major concern and requires innovative strategies to combat the problem.Over the past decades,several novel approaches involving different strategies have been developed to improve the ocular delivery system.Among these,the ophthalmic in-situ gel has attained a great attention over the past few years.This review discussed and summarized the recent and the promising research progress of in-situ gelling in ocular drug delivery system.展开更多
Stroke is the third leading cause of death and the first cause of adult disability in industrial countries [1]. It is charicaterized by hemiplegia, hemianopsia, aphasia, mouth askew and sever sequelae. It is considere...Stroke is the third leading cause of death and the first cause of adult disability in industrial countries [1]. It is charicaterized by hemiplegia, hemianopsia, aphasia, mouth askew and sever sequelae. It is considered that an ischemic disease without any specific treatment method and few effective drugs such as tPA (human tissue-type plasminogen activator) and Edarovone with specific therapeutic window will cause a lot of disadvantages if being used inaccurate. Root of Panax notoginseng (PN) which is one of traditional Chinese medicines (TCMs), was first found in “Shennong’s Classic of Materia Medica” around 200 AD. Panax notogineng saponins(PNS) is a multi-components mixture containing ginseng and saponins as the most important bioactive components which are commonly used in clinical treatment. Also, ginseng and saponins form the main components of many herbal medicines in the market, e.g., Xueshuantong injection [2], Xuesaitong injection [3], Xuesaitong soft capsule [4] and so on. The main monomers of Panax notoginseng saponins (PNS) are Ginsenoside-Rb1, Gensenoside-Rg1, Gensenoside-Re, Gensenoside-Rd and Panax notoginseng saponins-R1 [5]. In this review, we found some important points as well as shortcomings that require special consideration. We therefore highlighted the advances in neuro-protection of PNS and its main monomers in the area of experimental research.展开更多
OBJECTIVE Eurycoma longifolia is a tropical medicinal plant belonging to Simaroubaceae distributed in South East Asia.The aim of this study is to explore the effect and mechanism of E.longifolia stem 70%ethanol extrac...OBJECTIVE Eurycoma longifolia is a tropical medicinal plant belonging to Simaroubaceae distributed in South East Asia.The aim of this study is to explore the effect and mechanism of E.longifolia stem 70%ethanol extract(EL)and its active com⁃poundson uric acid excretion.METHODS Potassium oxonate(PO)induced hyperuricemia rats and adenine-PO induced hyperuricemia mouse model were used to evaluate the effects of EL.Ultra Performance Liquid Chromatography was used to determine the levels of plasma or serum uric acid and creatinine.Hematoxylin-eosin staining was applied to observe kidney pathological changes,Western blot⁃ting was applied to detect protein expression levels of uric acid transporters.Effects of constituents on urate uptake were tested in hU⁃RAT1-expressing HEK293T cells.RESULTS EL significantly reduced serum and plasma uric acid levels at dosages of 100,200 and 400 mg·kg^-1 in hyperuricemia rats and mice,and increased the clearance rate of uric acid and creatinine,improved therenal pathological injury.The protein expression levels of urate reabsorption transporter 1(URAT1)and glucose transporter 9 were down-regulated while sodium-dependent phosphate transporter 1 and ATP-binding cassette transporter G2 were up-regulated in the kidney after EL treat⁃ment.The diterpenes(50μmol·L^-1)isolated from EL showed inhibitory effects on urate uptake in hURAT1-expressing HEK293T cells,and the effect of eurycomanol was further confirmed in vivo.CONCLUSION EL significantly reduced blood uric acid levels and prevented pathological changes of kidney in PO induced hyperuricemia animal model,improved renal urate transports.We partly clarified the mechanism was related to suppressing effect of URAT1 by diterpene in EL.This study is the first to demonstrate that EL plays a role in hyperuricemia by promoting renal uric acid excretion.展开更多
Labrasol, as a non-ionic surfactant, can enhance the permeation and absorption of drugs, and is extensively used in topical, transdermal, and oral pharmaceutical preparations as an emulsifier and absorption enhancer. ...Labrasol, as a non-ionic surfactant, can enhance the permeation and absorption of drugs, and is extensively used in topical, transdermal, and oral pharmaceutical preparations as an emulsifier and absorption enhancer. Recent studies in our laboratory have indicated that labrasol has a strong absorption enhancing effect on different types of drugs in vitro and in vivo. This study was performed to further elucidate the action mechanism of labrasol on the corneal penetration. In this research, the fluorescein sodium, a marker of passive paracellular transport of tight junction, was selected as the model drug to assess the effect of labrasol on in vitro corneal permeability. To investigate the continuous and real-time influence of labrasol on the membrane permeability and integrity, the Ussing chamber system was applied to monitor the electrophysiological parameters. And, furthermore, we elucidated the effect of labrasol on excised cornea at the molecular level by application of RT-PCR, Western blot, and immunohistochemical staining. The results indicated that labrasol obviously enhance the transcorneal permeability of fluorescein sodium, and the enhancement was realized by interacting with and down-regulating the associated proteins, such as Factin, claudin-1 and β-catenin, which were contributed to cell-cell connections, respectively.展开更多
Paeoniflorin (PF) is one of the main bioactive components of total glucosides of paeony (TGP) extracted from the root of Paeonia lactiflora Pall. TGP exhibit various biological activities such as improvement in memory...Paeoniflorin (PF) is one of the main bioactive components of total glucosides of paeony (TGP) extracted from the root of Paeonia lactiflora Pall. TGP exhibit various biological activities such as improvement in memory, hepatoprotection, antimutagenic properties and platelet aggregation inhibition. The aim of this paper is to review the pharmacokinetics (PK) of PF as a pure compound and in single or multiple herb(s) of traditional Chinese medicine (TCM) prescriptions. The distribution of PF or PF in TCM fitted one or two compartmental model after oral administration or intravenous injection, respectively. However, PF has a low bioavailability (BA) in rabbit (7.24%) and rat (3.24%) after oral administration. The PK profiles and BA of PF were remarkably improved when co-administered with sinomenine or glycyrrhizin acid. The PK profiles and BA of PF in Radix Paeonia Rubra (RP-R) and Jing-zhi guan-xin were improved, but in co-administration of RP-R with Radix Angelicae Sinensis, the BA was significantly reduced. PK profiles and BA of PF in Shan yao gan-cao tang or Danggui-Shaoyao-San was either remarkably improved or not. However, neither the PK profiles nor the BA of PF in Radix paeonia alba, Huangqin-tang Si ni san or Tang-Min-Ling-Wan was improved. Metabolism in the liver did not play any role in the low oral BA of PF. The low BA was thus attributed to poor permeation due to low lipophilicity, P-glycoprotein mediated efflux, intestinal bacteria and hydrolytic degradation in the intestine by the intestinal brush border lactase phlorizin hydrolase (LPH) and certain esterases. These findings show the in vivo course of PF and provide information on the maximum biological actions of PF that may help traditional Chinese herbal medicinal practitioners.展开更多
Gout is a common of inflammatory arthritis and is caused by the deposition of monosodium urate(MSU)crystals as a result of hyperuricemia(HUA).Although HUA is considered to be the main risk factor for gout,only approxi...Gout is a common of inflammatory arthritis and is caused by the deposition of monosodium urate(MSU)crystals as a result of hyperuricemia(HUA).Although HUA is considered to be the main risk factor for gout,only approximately 10%of the individuals with HUA will eventually experience a gout attack.In this review,we first briefly introduce the development of gout and then summarize several possible reasons for its development.Genetic factors play a more prominent role in gout than in other diseases;functional mutations related to urate control and innate immunity components have been found to be associated with gout.Here,we list some of the most prominent genes involved in the pathogenesis of gout.In joints with MSU deposition,mature macrophages may uptake MSU crystals without causing inflammation,and this helps to maintain joints in an asymptomatic state.As an auxiliary inflammation pathway,the ATP-P2X7R-NLRP3 axis may contribute to the amplification of MSU-induced inflammation to affect the development of gout.Finally,this review summarizes the research progress on natural products that can be used in the treatment of HUA and gout.展开更多
Licorice is one of the oldest herbal medicines for its various ethno pharmacological uses.In both Asian and European countries,it has been recorded for treatment of inflammatory diseases.A large number of ingredients ...Licorice is one of the oldest herbal medicines for its various ethno pharmacological uses.In both Asian and European countries,it has been recorded for treatment of inflammatory diseases.A large number of ingredients have been isolated from licorice,including triterpene saponins and flavonoids,which are normally being considered to be the main biologically active components.In the last decade,licorice has been proved exert anti-diabetic effect in various in vivo and in vitro models of diabetes mellitus.Furthermore,licorice can also antagonize all sorts of diabetes complications,including diabetic nephropathy,atherosclerosis,diabetic retinopathy and neuropathy.Except anti-inflammation,licorice and its active components show anti-diabetic effects by improving insulin resistance and increasing insulin secretion,regulating lipid metabolism,and anti-oxidation.The useful effects of licorice and its active components are due to regulating different pathways and proteins,including NF-κB,AMPK,insulin signaling pathway,MAPK,etc.In this review,we provide an overview of the beneficial effects and related molecular mechanism of licorice and its effective components on improving diabetes and its complications.展开更多
In order to identify the potential nephrotoxic compounds in traditional Chinese medicine Lithospermum erythrorhizon,it was separated into serial fractions according to their polarities.An in vitro method was utilized ...In order to identify the potential nephrotoxic compounds in traditional Chinese medicine Lithospermum erythrorhizon,it was separated into serial fractions according to their polarities.An in vitro method was utilized to determine the nephrotoxicity of these fractions with the help of fluorescence image analysis.As a result,the primary fraction A05 and its secondary fractions C06 "C09 and C12 "C14 were found to have significant toxicity to LLC-PK1 cell line,as determined by the survive rate less than 20% after they were treated with these fractions.These potential nephrotoxic fractions were further analyzed by multistage and high resolution mass spectrometry.The main compounds in these fractions were tentatively identified to be acetylshikonin,isobutyrylshikonin,β,β'-dimethyla-cryloylshikonin,and isovalerylshikonin,which may bring nephrotoxicity.展开更多
Rhizoma Coptidis,a traditional Chinese herbal medicine,has been used for treating diabetes for thousands of years.However,the molecular basis for this action has not been elucidated.In the present study,the effects of...Rhizoma Coptidis,a traditional Chinese herbal medicine,has been used for treating diabetes for thousands of years.However,the molecular basis for this action has not been elucidated.In the present study,the effects of seven main alkaloids of Rhizoma Coptidis on glycometabolism were investigated and the related molecular mechanism of five active compounds on insulin resistant(IR)cell model was explored for the first time.Results showed that berberine,palmatine,epiberberine,columbamine and groenlandicine enhanced glucose consumption in the palmitic acid(PA)-induced IR-HepG2 cells,indicating that these compounds could improve IR.In addition,we found that among these active alkaloids,berberine,columbamine,epiberberine and groenlandicine could inhibit the activation of ERK and p38 pathway,while berberine,columbamine,palmatine and epiberberine could activate AMPK pathway.Moreover,palmatine and columbamine regulated the mRNA expression of GLUT2 to ameliorate IR via activating AMPK and inactivating p38 MAPK signal pathway.To sum up,berberine,columbamine,palmatine,epiberberine and groenlandicine might be the active reagents,which contribute to the glucose lowering effects of Rhizoma Coptidis.展开更多
Lianqiao(Forsythia suspensa(Thunb.)Vahl,Abbreviated as“F.suspensa”below)is traditional Chinese medicine in China,widely distributed all over the country and also highly rich in resources.The Northwest Territories is...Lianqiao(Forsythia suspensa(Thunb.)Vahl,Abbreviated as“F.suspensa”below)is traditional Chinese medicine in China,widely distributed all over the country and also highly rich in resources.The Northwest Territories is a genuine producing area.There are many chemical components in F.suspensa,including lignans,phenylethanolic glycosides,flavonoids,C6-C2 natural alcohols and their glycosides,phenolic acids,terpenes and volatile oils.Among them,lignans and phenylethanolic glycosides are the main active components.Based on the review of chemical components or constituents and main pharmacological activities,according to the definition of quality marker,the components of quality marker of F.suspensa were predicted and analyzed from the aspects of plant genetics and chemical composition,traditional efficacy,traditional medicine,new efficacy,blood entering composition,measurable composition,effective composition in different compatibility,and the influence of storage conditions of extract.Further research on its medicinal active ingredients will provide scientific basis for the evaluation of F.suspensa quality.展开更多
Shuxuetong injection composed of leech(Hirudo nipponica Whitman) and earthworm(Pheretima aspergillum) has been used for the clinical treatment of acute stroke for many years in China. However, the precise neuroprotect...Shuxuetong injection composed of leech(Hirudo nipponica Whitman) and earthworm(Pheretima aspergillum) has been used for the clinical treatment of acute stroke for many years in China. However, the precise neuroprotective mechanism of Shuxuetong injection remains poorly understood. Here, cerebral microvascular endothelial cells(bEnd.3) were incubated in glucose-free Dulbecco's modified Eagle's medium containing 95% N_2/5% CO_2 for 6 hours, followed by high-glucose medium containing 95% O_2 and 5% CO_2 for 18 hours to establish an oxygen-glucose deprivation/reperfusion model. This in vitro cell model was administered Shuxuetong injection at 1/32, 1/64, and 1/128 concentrations(diluted 32-, 64-, and 128-times). Cell Counting Kit-8 assay was used to evaluate cell viability. A fluorescence method was used to measure lactate dehydrogenase, and a fluorescence microplate reader used to detect intracellular reactive oxygen species. A fluorescent probe was also used to measure mitochondrial superoxide production. A cell resistance meter was used to measure transepithelial resistance and examine integrity of monolayer cells. The fluorescein isothiocyanate-dextran test was performed to examine blood-brain barrier permeability. Real-time reverse transcription polymerase chain reaction was performed to analyze mRNA expression levels of tumor necrosis factor alpha, interleukin-1β, interleukin-6, and inducible nitric oxide synthase. Western blot assay was performed to analyze expression of caspase-3, intercellular adhesion molecule 1, vascular cell adhesion molecule 1, occludin, vascular endothelial growth factor, cleaved caspase-3, B-cell lymphoma 2, phosphorylated extracellular signal-regulated protein kinase, extracellular signal-regulated protein kinase, nuclear factor-κB p65, I kappa B alpha, phosphorylated I kappa B alpha, I kappa B kinase, phosphorylated I kappa B kinase, claudin-5, and zonula occludens-1. Our results show that Shuxuetong injection increases bEnd.3 cell viability and B-cell lymphoma 2 expression, reduces cleaved caspase-3 expression, inhibits production of reactive oxygen species and mitochondrial superoxide, suppresses expression of tumor necrosis factor alpha, interleukin-1β, interleukin-6, inducible nitric oxide synthase mRNA, intercellular adhesion molecule-1, and vascular cell adhesion molecule-1, markedly increases transepithelial resistance, decreases blood-brain barrier permeability, upregulates claudin-5, occludin, and zonula occludens-1 expression, reduces nuclear factor-κB p65 and vascular endothelial growth factor expression, and reduces I kappa B alpha, extracellular signal-regulated protein kinase 1/2, and I kappa B kinase phosphorylation levels. Overall, these findings suggest that Shuxuetong injection has protective effects on brain microvascular endothelial cells after oxygen-glucose deprivation/reperfusion. Moreover, its protective effect is associated with reduction of mitochondrial superoxide production, inhibition of the inflammatory response, and inhibition of vascular endothelial growth factor, extracellular signal-regulated protein kinase 1/2, and the nuclear factor-κB p65 signaling pathway.展开更多
文摘Metabolomics is a newly developed discipline following genomics,transcriptomics,and proteomics.It is based on the theory of systems biology,supported by modern spectroscopy analysis technology and is analyzed by the external stimulation of organisms.By analyzing the overall changes of metabolites with low molecular weight in organism after being stimulated by the outside world,we can understand the physiological and pathological state of organism,and finally clarify the essence of biological changes of organism.In recent years,metabolomics has developed rapidly.It has been widely used in early toxicity screening of lead compounds,preclinical toxicity of drugs and toxicity evaluation of clinical drugs.The in-depth interpretation of metabolomics results is also developing.This article reviewed the progress of metabolomics technology and its application in the study of toxicity evaluation of traditional Chinese medicine.
文摘Traditional Chinese medicine (TCM) is a medical system that has collected and summarized abundant clinical experience over its long history of more than 2000 years. However, the frequent occurrence of TCM-induced adverse reactions has hindered the modernization and internationalization of TCM, while attracting increasing attention from around the world. Unlike chemical drugs and biological agents, the difficulties involved in research on the toxicity and safety of TCM mainly include the complexity of its components and the unpredictability of drug–body interactions. Much of TCM, which has overall therapeutic effects, has the typical mechanisms of multiple components, multiple pathways, and multiple targets. While considering the gradualness and unpredictability of TCM toxicity, the ambiguity of toxicants and safe dosage, and individual differences during long-term TCM administration, we have systematically established key techniques for the toxicity assessment of TCM. These techniques mainly include TCM toxicity discovery in an early phase, based on a combination of drug toxicology genomics and metabolomics;methods to identify dose–toxicity relationships in TCM;and integrated techniques for the exploration of TCM interactions, such as fast-screening tests based on drug-metabolizing enzymes and receptor pathways. In particular, we have developed a new technical system for TCM safety evaluation using molecular toxicology, which has been validated well in research on TCM compatibility contraindication, quality control, and allergen discovery. The application of this key technical platform is introduced here in detail. This application includes model organisms, toxicant biomarkers, a magnetic suspension technique, and the application of network toxicology and computational toxicology in research on the toxicity of Fructus toosendan, Semen cassiae, Polygonum multiflorum, and Fructus psoraleae.
基金Supported by National Natural Science Foundation of China,No.81173469 and No.81273891the Key New Drug Creation and Manufacturing Program,No.2012ZX09304007
文摘AIM:To investigate the hepatoprotective effects and mechanisms of an extract of Salvia miltiorrhiza and Carthamus tinctorius in vivo.METHODS:C57BL/6J mice were randomly assigned to five groups and intraperitoneally administered 0.9% saline,Salvia miltiorrhiza and Carthamus tinctorius extract [Danhong injection(DHI),0.75 and 3 g/kg mixed extract] or reduced glutathione for injection(RGI,300 mg/kg) for 30 min before exposure to lipopolysaccharide(LPS,16 mg/kg). After intraperitoneal LPS stimulation for 90 min or 6 h,the mice were sacrificed by ether anaesthesia,and serum and liver samples were collected. Histological analysis(H&E) and terminal deoxynucleotidyl transferase-mediated d UTP nick end-labelling(TUNEL) staining were performed. Alanine transferase(ALT),aspartate transaminase(AST),total bilirubin(TBil),glutathione-S-transferase(GST),malondialdehyde(MDA),tumour necrosis factor(TNF)-α,interleukin(IL)-6,and caspase-3 levels were measured. Bax,Bcl-2,P-IκBα,IκBα,P-NF-κB p65,and NF-κB p65 protein levels were determined by Western blot. TNF-α,IL-6,caspase-3,Bax and Bcl-2 m RNA expression was measured by real-time reverse transcription-polymerase chain reaction(RT-PCR).RESULTS:Hematoxylin-eosin staining and TUNEL results suggested that DHI(3 g/kg) treatment alleviated inflammatory and apoptotic(P < 0.01) injury in the liver of mice. DHI treatment dose-dependently blunted the abnormal changes in biochemical parameters such as ALT(72.53 ± 2.83 for 3 g/kg,P < 0.01),AST(76.97 ± 5.00 for 3 g/kg,P < 0.01),TBil(1.17 ± 0.10 for 3 g/kg,P < 0.01),MDA(0.81 ± 0.36 for 3 g/kg,P < 0.01),and GST(358.86 ± 12.09 for 3 g/kg,P < 0.01). Moreover,DHI(3 g/kg) remarkably decreased LPS-induced protein expression of TNF-α(340.55 ± 10.18 for 3 g/kg,P < 0.01),IL-6(261.34 ± 10.18 for 3 g/kg,P < 0.01),and enzyme activity of caspase-3(0.93 ± 0.029 for 3 g/kg,P < 0.01). The LPS-induced m RNA expression of TNF-α,IL-6 and caspase-3 was also decreased by DHI. Western blot analysis revealed that DHI antagonised LPS-stimulated decrease of Bcl-2 and increase of Bax protein expression. Furthermore,DHI inhibited LPS-induced IκBα and NF-κB p65 phosphorylation.CONCLUSION:DHI may be a multi-function protectant against acute hepatic injury in mice through its antiinflammatory,anti-oxidative and anti-apoptotic activities.
基金the National Natural Science Foundation of China(Grant No.81872996)the State Key Research and Development Project(Grant No.2017YFC1702104)+1 种基金the State Key Project for the Creation of Major New Drugs(2018ZX09711001-009-010)the Tianjin Municipal Education Commission Research Project(Grant No.2017ZD07)。
文摘Inherent complexity of plant metabolites necessitates the use of multi-dimensional information to accomplish comprehensive profiling and confirmative identification.A dimension-enhanced strategy,by offline two-dimensional liquid chromatography/ion mobility-quadrupole time-of-flight mass spectrometry(2 D-LC/IM-QTOF-MS)enabling four-dimensional separations(2 D-LC,IM,and MS),is proposed.In combination with in-house database-driven automated peak annotation,this strategy was utilized to characterize ginsenosides simultaneously from white ginseng(WG)and red ginseng(RG).An offline 2 DLC system configuring an Xbridge Amide column and an HSS T3 column showed orthogonality 0.76 in the resolution of ginsenosides.Ginsenoside analysis was performed by data-independent high-definition MSE(HDMSE)in the negative ESI mode on a Vion?IMS-QTOF hybrid high-resolution mass spectrometer,which could better resolve ginsenosides than MSEand directly give the CCS information.An in-house ginsenoside database recording 504 known ginsenosides and 58 reference compounds,was established to assist the identification of ginsenosides.Streamlined workflows,by applying UNIFI?to automatedly annotate the HDMSEdata,were proposed.We could separate and characterize 323 ginsenosides(including 286 from WG and 306 from RG),and 125 thereof may have not been isolated from the Panax genus.The established 2 D-LC/IM-QTOF-HDMSEapproach could also act as a magnifier to probe differentiated components between WG and RG.Compared with conventional approaches,this dimensionenhanced strategy could better resolve coeluting herbal components and more efficiently,more reliably identify the multicomponents,which,we believe,offers more possibilities for the systematic exposure and confirmative identification of plant metabolites.
文摘Baicalein(BE) is one of the main active flavonoids representing the variety of pharmacological effects including anticancer, anti-inflammatory and cardiovascular protective activities, but it's very low solubility, dissolution rate and poor oral absorption limit the therapeutic applications. In this work, a nano-cocrystal strategy was successfully applied to improve the dissolution rate and bioavailability of BE. Baicalein-nicotinamide(BE-NCT) nanococrystals were prepared by high pressure homogenization and evaluated both in vitro and in vivo. Physical characterization results including scanning electron microscopy, dynamic light scattering, powder X-ray diffraction and differential scanning calorimetry demonstrated that BE-NCT nano-cocrystals were changed into amorphous state with mean particle size of 251.53 nm. In the dissolution test, the BE-NCT nano-cocrystals performed 2.17-fold and 2.54-fold enhancement than BE coarse powder in FaSSIF-V2 and FaSSGF. Upon oral administration, the integrated AUC0-t of BE-NCT nano-cocrystals(6.02-fold) was significantly higher than BE coarse powder(1-fold), BE-NCT cocrystals(2.87-fold) and BE nanocrystals(3.32-fold). Compared with BE coarse powder, BE-NCT cocrystals and BE nanocrystals, BENCT nano-cocrystals possessed excellent performance both in vitro and in vivo evaluations.Thus, it can be seen that nano-cocrystal is an appropriate novel strategy for improving dissolution rate and bioavailability of poor soluble natural products such as BE.
文摘OBJECTIVE To highlight the pharmacological effects of rosemary and its active compounds and eluci⁃date its related mechanisms in nonalcoholic fatty liver disease(NAFLD)management both in vitro and in vivo.METHODS In orotic acid induced NAFLD rats model,rats were administrated with 100,200 and 400 mg·kg^-1 rosemary ethanol extract(RO),10,25 and 50 mg·kg^-1 rosemary acid(RA),and 5,10 and 25 mg·kg^-1 carnosic acid(CA)for three weeks respec⁃tively.Sodium oleate induced HepG2 cell model was used to study the regulation effect of rosemary ethanol extract and its main metabolites on fat accumulation.lipid metabolism related gene expression was analyzed by Western blotting and real-time PCR to clarify the specific molecular mechanism of RO,RA and CA in lipid accumulation.RESULTS RO,RA and CA significantly reduced the contents of liver triglyceride(TG),total cholesterol(TC),free fatty acids(FFA)and improved cell hypertrophy,vacuolation,and cell necrosis in liver of orotic acid induced NAFLD model rats.The mecha⁃nism and related pathways of RO and its main metabolites against lipid disorder was related to the up-regulation of the phosphorylation of adenosine 5′-monophosphate(AMP)-activated protein kinase(AMPK)and inhibition of the sterol regu⁃latory element binding protein-1c(SREBP-1c)cracking into the nuclear,following down-regulation of fatty acid synthesis.CONCLUSION The rosemary has effectively function to regulate lipid metabolism through AMPK/SREBP1c signaling pathway.
文摘Blindness and vision impairment are the most devastating global health problems resulting in a substantial economic and social burden.Delivery of drug to particular parts of the anterior or posterior segment has been a major challenge due to various protective barriers and elimination mechanisms associated with the unique anatomical and physiological nature of the ocular system.Drug administration to the eye by conventional delivery systems results in poor ocular bioavailability(<5%).The designing of a novel approach for a safe,simple,and effective ocular drug delivery is a major concern and requires innovative strategies to combat the problem.Over the past decades,several novel approaches involving different strategies have been developed to improve the ocular delivery system.Among these,the ophthalmic in-situ gel has attained a great attention over the past few years.This review discussed and summarized the recent and the promising research progress of in-situ gelling in ocular drug delivery system.
文摘Stroke is the third leading cause of death and the first cause of adult disability in industrial countries [1]. It is charicaterized by hemiplegia, hemianopsia, aphasia, mouth askew and sever sequelae. It is considered that an ischemic disease without any specific treatment method and few effective drugs such as tPA (human tissue-type plasminogen activator) and Edarovone with specific therapeutic window will cause a lot of disadvantages if being used inaccurate. Root of Panax notoginseng (PN) which is one of traditional Chinese medicines (TCMs), was first found in “Shennong’s Classic of Materia Medica” around 200 AD. Panax notogineng saponins(PNS) is a multi-components mixture containing ginseng and saponins as the most important bioactive components which are commonly used in clinical treatment. Also, ginseng and saponins form the main components of many herbal medicines in the market, e.g., Xueshuantong injection [2], Xuesaitong injection [3], Xuesaitong soft capsule [4] and so on. The main monomers of Panax notoginseng saponins (PNS) are Ginsenoside-Rb1, Gensenoside-Rg1, Gensenoside-Re, Gensenoside-Rd and Panax notoginseng saponins-R1 [5]. In this review, we found some important points as well as shortcomings that require special consideration. We therefore highlighted the advances in neuro-protection of PNS and its main monomers in the area of experimental research.
文摘OBJECTIVE Eurycoma longifolia is a tropical medicinal plant belonging to Simaroubaceae distributed in South East Asia.The aim of this study is to explore the effect and mechanism of E.longifolia stem 70%ethanol extract(EL)and its active com⁃poundson uric acid excretion.METHODS Potassium oxonate(PO)induced hyperuricemia rats and adenine-PO induced hyperuricemia mouse model were used to evaluate the effects of EL.Ultra Performance Liquid Chromatography was used to determine the levels of plasma or serum uric acid and creatinine.Hematoxylin-eosin staining was applied to observe kidney pathological changes,Western blot⁃ting was applied to detect protein expression levels of uric acid transporters.Effects of constituents on urate uptake were tested in hU⁃RAT1-expressing HEK293T cells.RESULTS EL significantly reduced serum and plasma uric acid levels at dosages of 100,200 and 400 mg·kg^-1 in hyperuricemia rats and mice,and increased the clearance rate of uric acid and creatinine,improved therenal pathological injury.The protein expression levels of urate reabsorption transporter 1(URAT1)and glucose transporter 9 were down-regulated while sodium-dependent phosphate transporter 1 and ATP-binding cassette transporter G2 were up-regulated in the kidney after EL treat⁃ment.The diterpenes(50μmol·L^-1)isolated from EL showed inhibitory effects on urate uptake in hURAT1-expressing HEK293T cells,and the effect of eurycomanol was further confirmed in vivo.CONCLUSION EL significantly reduced blood uric acid levels and prevented pathological changes of kidney in PO induced hyperuricemia animal model,improved renal urate transports.We partly clarified the mechanism was related to suppressing effect of URAT1 by diterpene in EL.This study is the first to demonstrate that EL plays a role in hyperuricemia by promoting renal uric acid excretion.
文摘Labrasol, as a non-ionic surfactant, can enhance the permeation and absorption of drugs, and is extensively used in topical, transdermal, and oral pharmaceutical preparations as an emulsifier and absorption enhancer. Recent studies in our laboratory have indicated that labrasol has a strong absorption enhancing effect on different types of drugs in vitro and in vivo. This study was performed to further elucidate the action mechanism of labrasol on the corneal penetration. In this research, the fluorescein sodium, a marker of passive paracellular transport of tight junction, was selected as the model drug to assess the effect of labrasol on in vitro corneal permeability. To investigate the continuous and real-time influence of labrasol on the membrane permeability and integrity, the Ussing chamber system was applied to monitor the electrophysiological parameters. And, furthermore, we elucidated the effect of labrasol on excised cornea at the molecular level by application of RT-PCR, Western blot, and immunohistochemical staining. The results indicated that labrasol obviously enhance the transcorneal permeability of fluorescein sodium, and the enhancement was realized by interacting with and down-regulating the associated proteins, such as Factin, claudin-1 and β-catenin, which were contributed to cell-cell connections, respectively.
文摘Paeoniflorin (PF) is one of the main bioactive components of total glucosides of paeony (TGP) extracted from the root of Paeonia lactiflora Pall. TGP exhibit various biological activities such as improvement in memory, hepatoprotection, antimutagenic properties and platelet aggregation inhibition. The aim of this paper is to review the pharmacokinetics (PK) of PF as a pure compound and in single or multiple herb(s) of traditional Chinese medicine (TCM) prescriptions. The distribution of PF or PF in TCM fitted one or two compartmental model after oral administration or intravenous injection, respectively. However, PF has a low bioavailability (BA) in rabbit (7.24%) and rat (3.24%) after oral administration. The PK profiles and BA of PF were remarkably improved when co-administered with sinomenine or glycyrrhizin acid. The PK profiles and BA of PF in Radix Paeonia Rubra (RP-R) and Jing-zhi guan-xin were improved, but in co-administration of RP-R with Radix Angelicae Sinensis, the BA was significantly reduced. PK profiles and BA of PF in Shan yao gan-cao tang or Danggui-Shaoyao-San was either remarkably improved or not. However, neither the PK profiles nor the BA of PF in Radix paeonia alba, Huangqin-tang Si ni san or Tang-Min-Ling-Wan was improved. Metabolism in the liver did not play any role in the low oral BA of PF. The low BA was thus attributed to poor permeation due to low lipophilicity, P-glycoprotein mediated efflux, intestinal bacteria and hydrolytic degradation in the intestine by the intestinal brush border lactase phlorizin hydrolase (LPH) and certain esterases. These findings show the in vivo course of PF and provide information on the maximum biological actions of PF that may help traditional Chinese herbal medicinal practitioners.
基金Important Drug Development Fund,Ministry of Science and Technology of China(2018ZX09735-002)National Natural Science Foundation of China(81173524,81673688)。
文摘Gout is a common of inflammatory arthritis and is caused by the deposition of monosodium urate(MSU)crystals as a result of hyperuricemia(HUA).Although HUA is considered to be the main risk factor for gout,only approximately 10%of the individuals with HUA will eventually experience a gout attack.In this review,we first briefly introduce the development of gout and then summarize several possible reasons for its development.Genetic factors play a more prominent role in gout than in other diseases;functional mutations related to urate control and innate immunity components have been found to be associated with gout.Here,we list some of the most prominent genes involved in the pathogenesis of gout.In joints with MSU deposition,mature macrophages may uptake MSU crystals without causing inflammation,and this helps to maintain joints in an asymptomatic state.As an auxiliary inflammation pathway,the ATP-P2X7R-NLRP3 axis may contribute to the amplification of MSU-induced inflammation to affect the development of gout.Finally,this review summarizes the research progress on natural products that can be used in the treatment of HUA and gout.
基金This work was supported by State Key Program of National Natural Science Foundation of China(Grant number:81430095).
文摘Licorice is one of the oldest herbal medicines for its various ethno pharmacological uses.In both Asian and European countries,it has been recorded for treatment of inflammatory diseases.A large number of ingredients have been isolated from licorice,including triterpene saponins and flavonoids,which are normally being considered to be the main biologically active components.In the last decade,licorice has been proved exert anti-diabetic effect in various in vivo and in vitro models of diabetes mellitus.Furthermore,licorice can also antagonize all sorts of diabetes complications,including diabetic nephropathy,atherosclerosis,diabetic retinopathy and neuropathy.Except anti-inflammation,licorice and its active components show anti-diabetic effects by improving insulin resistance and increasing insulin secretion,regulating lipid metabolism,and anti-oxidation.The useful effects of licorice and its active components are due to regulating different pathways and proteins,including NF-κB,AMPK,insulin signaling pathway,MAPK,etc.In this review,we provide an overview of the beneficial effects and related molecular mechanism of licorice and its effective components on improving diabetes and its complications.
基金Supported by the National Key Scientific and Technological Project of China(No.2009ZX09502-012)the Research Fund for the Doctoral Program of Higher Education of China(No.20090101110126)the Zhejiang Province Science and Technology Plan Project,China(No.2008C23065)
文摘In order to identify the potential nephrotoxic compounds in traditional Chinese medicine Lithospermum erythrorhizon,it was separated into serial fractions according to their polarities.An in vitro method was utilized to determine the nephrotoxicity of these fractions with the help of fluorescence image analysis.As a result,the primary fraction A05 and its secondary fractions C06 "C09 and C12 "C14 were found to have significant toxicity to LLC-PK1 cell line,as determined by the survive rate less than 20% after they were treated with these fractions.These potential nephrotoxic fractions were further analyzed by multistage and high resolution mass spectrometry.The main compounds in these fractions were tentatively identified to be acetylshikonin,isobutyrylshikonin,β,β'-dimethyla-cryloylshikonin,and isovalerylshikonin,which may bring nephrotoxicity.
基金supported by Technology Major Project of China“Key New Drug Creation and Manufacturing Program”(No.2017ZX09301012-001)the National Natural Science Foundation of China(No.20972098,No.81703776 and No.81430095)the National Basic Research Program of China(No.2014CB560706).
文摘Rhizoma Coptidis,a traditional Chinese herbal medicine,has been used for treating diabetes for thousands of years.However,the molecular basis for this action has not been elucidated.In the present study,the effects of seven main alkaloids of Rhizoma Coptidis on glycometabolism were investigated and the related molecular mechanism of five active compounds on insulin resistant(IR)cell model was explored for the first time.Results showed that berberine,palmatine,epiberberine,columbamine and groenlandicine enhanced glucose consumption in the palmitic acid(PA)-induced IR-HepG2 cells,indicating that these compounds could improve IR.In addition,we found that among these active alkaloids,berberine,columbamine,epiberberine and groenlandicine could inhibit the activation of ERK and p38 pathway,while berberine,columbamine,palmatine and epiberberine could activate AMPK pathway.Moreover,palmatine and columbamine regulated the mRNA expression of GLUT2 to ameliorate IR via activating AMPK and inactivating p38 MAPK signal pathway.To sum up,berberine,columbamine,palmatine,epiberberine and groenlandicine might be the active reagents,which contribute to the glucose lowering effects of Rhizoma Coptidis.
文摘Lianqiao(Forsythia suspensa(Thunb.)Vahl,Abbreviated as“F.suspensa”below)is traditional Chinese medicine in China,widely distributed all over the country and also highly rich in resources.The Northwest Territories is a genuine producing area.There are many chemical components in F.suspensa,including lignans,phenylethanolic glycosides,flavonoids,C6-C2 natural alcohols and their glycosides,phenolic acids,terpenes and volatile oils.Among them,lignans and phenylethanolic glycosides are the main active components.Based on the review of chemical components or constituents and main pharmacological activities,according to the definition of quality marker,the components of quality marker of F.suspensa were predicted and analyzed from the aspects of plant genetics and chemical composition,traditional efficacy,traditional medicine,new efficacy,blood entering composition,measurable composition,effective composition in different compatibility,and the influence of storage conditions of extract.Further research on its medicinal active ingredients will provide scientific basis for the evaluation of F.suspensa quality.
基金supported in part by the National Natural Science Foundation of China,No.81573644(to LMH),81573733(to SWX)the Tianjin 131 Innovative Team Project,China(to HW)+5 种基金the National Major Science and Technology Project of China,No.2012ZX09101201-004(to SWX)the Science and Technology Plan Project of Tianjin of China,No.16PTSYJC00120(to LMH)the Applied Foundation and Frontier Technology Research Program of Tianjin of China(General Project),No.14JCYBJC28900(to SXW)the National International Science and Technology Cooperation Project of China,No.2015DFA30430(to HW)the Key Program of the Natural Science Foundation of Tianjin of China,No.16ICZDJC36300(to HW)the Scientific Research and Technology Development Plan Project of Guangxi Zhuang Autonomous Region of China,No.14125008-2-5(to SXW)
文摘Shuxuetong injection composed of leech(Hirudo nipponica Whitman) and earthworm(Pheretima aspergillum) has been used for the clinical treatment of acute stroke for many years in China. However, the precise neuroprotective mechanism of Shuxuetong injection remains poorly understood. Here, cerebral microvascular endothelial cells(bEnd.3) were incubated in glucose-free Dulbecco's modified Eagle's medium containing 95% N_2/5% CO_2 for 6 hours, followed by high-glucose medium containing 95% O_2 and 5% CO_2 for 18 hours to establish an oxygen-glucose deprivation/reperfusion model. This in vitro cell model was administered Shuxuetong injection at 1/32, 1/64, and 1/128 concentrations(diluted 32-, 64-, and 128-times). Cell Counting Kit-8 assay was used to evaluate cell viability. A fluorescence method was used to measure lactate dehydrogenase, and a fluorescence microplate reader used to detect intracellular reactive oxygen species. A fluorescent probe was also used to measure mitochondrial superoxide production. A cell resistance meter was used to measure transepithelial resistance and examine integrity of monolayer cells. The fluorescein isothiocyanate-dextran test was performed to examine blood-brain barrier permeability. Real-time reverse transcription polymerase chain reaction was performed to analyze mRNA expression levels of tumor necrosis factor alpha, interleukin-1β, interleukin-6, and inducible nitric oxide synthase. Western blot assay was performed to analyze expression of caspase-3, intercellular adhesion molecule 1, vascular cell adhesion molecule 1, occludin, vascular endothelial growth factor, cleaved caspase-3, B-cell lymphoma 2, phosphorylated extracellular signal-regulated protein kinase, extracellular signal-regulated protein kinase, nuclear factor-κB p65, I kappa B alpha, phosphorylated I kappa B alpha, I kappa B kinase, phosphorylated I kappa B kinase, claudin-5, and zonula occludens-1. Our results show that Shuxuetong injection increases bEnd.3 cell viability and B-cell lymphoma 2 expression, reduces cleaved caspase-3 expression, inhibits production of reactive oxygen species and mitochondrial superoxide, suppresses expression of tumor necrosis factor alpha, interleukin-1β, interleukin-6, inducible nitric oxide synthase mRNA, intercellular adhesion molecule-1, and vascular cell adhesion molecule-1, markedly increases transepithelial resistance, decreases blood-brain barrier permeability, upregulates claudin-5, occludin, and zonula occludens-1 expression, reduces nuclear factor-κB p65 and vascular endothelial growth factor expression, and reduces I kappa B alpha, extracellular signal-regulated protein kinase 1/2, and I kappa B kinase phosphorylation levels. Overall, these findings suggest that Shuxuetong injection has protective effects on brain microvascular endothelial cells after oxygen-glucose deprivation/reperfusion. Moreover, its protective effect is associated with reduction of mitochondrial superoxide production, inhibition of the inflammatory response, and inhibition of vascular endothelial growth factor, extracellular signal-regulated protein kinase 1/2, and the nuclear factor-κB p65 signaling pathway.
