Protective effects of paeonol on LPS-induced macrophage RAW264.7 injury through TLR4/MAPK/NF-κB pathway

Objective:To study the protective effect of paeonol on LPS-induced inflammatory injury of macrophage RAW264.7 cells and its mechanism.Methods:Macrophage RAW264.7 cells were cultured and divided into blank group,LPS(1μg/mL)group,paeonol(240μmol/mL)group and TAK242(10μmol/mL)group.The cell activity was detected by CCK8 method,the cell morphology was observed by inverted microscope,the contents of GSH and MDA in cell culture medium were determined by colorimetry,the mitochondrial membrane potential was detected by JC-1 method,the expression distribution of F4/80 and p-NF-κB protein was detected by immunofluorescence method,and the expression of TLR4/MAPK/NF-κB related pathway protein was detected by Western blotting.Results:Compared with the blank group,the cell viability induced by 1μg/mL LPS was 0.4972±0.061(P<0.01),which was close to the half inhibition rate.Compared with LPS group,the expression of p-NF-κB protein in 240μmol/mL paeonol pretreated cell group was down-regulated most significantly(P<0.01),and the expression of TLR4 protein was inhibited most significantly in 10μmol/mL TAK242 pretreated cell group.Compared with LPS group(P<0.01),the cell morphology of paeonol group recovered.Decrease MDA content and increase GSH content in cell culture medium(P<0.01),In the results of mitochondrial membrane potential,the red light of paeonol group was significantly enhanced and the green light was significantly weakened(P<0.001).The expression distribution of F4/80 and p-NF-κB protein in paeonol group decreased significantly(P<0.01),and the expressions of TLR4,p-IκB,p-p38,p-JNK and p-NF-κB protein were down-regulated(P<0.05).Conclusion:Paeonol can improve the inflammatory injury of RAW264.7 cells induced by LPS,and its mechanism may be related to TLR4/MAPK/NF-κB pathway.

Coating Process of Paeonol Sustained Release Pills

[Objectives] To study the coating process of paeonol sustained release pills by extrusion-spheronization method taking ethyl cellulose as the coating material. [Methods] Paeonol pills were made by Auari AW-95 Full Automatic Pill Making Machine. Coating of paeonol sustained release pills was prepared by Auari Mini Pill Polishing Machine. The prescription and process factors of paeonol sustained release pills coating were investigated by single factor experiment and orthogonal experiment. The release of paeonol sustained release pills was determined according to the cumulative release curve of paeonol. [Results] The prepared paeonol sustained release pills released slowly within 24 h, and the release rate reached 80% in 12 h. [Conclusions] The prepared paeonol sustained release pills basically meet the 24 h sustained release standard, and can be further developed and applied.

Paeonol inhibits tumor growth in gastric cancer in vitro and in vivo

AIM:To investigate the anti-tumor effects of paeonol in gastric cancer cell proliferation and apoptosis in vitro and in vivo.METHODS:Murine gastric cancer cell line mouse forestomach carcinoma(MFC) or human gastric cancer cell line SGC-7901 was cultured in the presence or absence of paeonol.Cell proliferation was determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay,and cell cycle and apoptosis by flow cytometry and TUNEL staining.Tumor growth after subcutaneous implantation of MFC cells in mice was monitored,and the effects of treatment with paeonol were determined.RESULTS:In vitro,paeonol caused dose-dependent inhibition on cell proliferation and induced apoptosis.Cell cycle analysis revealed a decreased proportion of cells in G0/G1 phase,with arrest at S.Paeonol treatment in gastric cancer cell line MFC and SGC-790 cells significantly reduced the expression of Bcl-2 and increased the expression of Bax in a concentration-related manner.Administration of paeonol to MFC tumor-bearing mice significantly lowered the tumor growth and caused tumor regression.CONCLUSION:Paeonol has signif icantly growth-inhibitory and apoptosis-inducing effects in gastric cancer cells both in vitro and in vivo.

Paeonol attenuates inflammation-mediated neurotoxicity and microglial activation

Chronic activation of microglial cells endangers neuronal survival through the release of various proinflammatory and neurotoxic factors. The root of Paeonia lactiflora Pall has been considered useful for the treatment of various disorders in traditional oriental medicine. Paeonol, found in the root of Paeonia lactiflora Pall, has a wide range of pharmacological functions, including anti-oxidative, anti-inflammatory and neuroprotective activities. The objective of this study was to examine the efficacy of paeonol in the repression of inflammation-induced neurotoxicity and microglial cell activation. Organotypic hippocampal slice cultures and primary microglial cells from rat brain were stimulated with bacterial lipopolysaccharide. Paeonol pretreatment was performed for 30 minutes prior to lipopolysaccharide addition. Cell viability and nitrite （the production of nitric oxide）, tumor necrosis factor-alpha and interleukin-lbeta products were measured after lipopolysaccharide treatment. In organotypic hippocampal slice cultures, paeonol blocked lipopolysaccharide-related hippocampal cell death and inhibited the release of nitrite and interleukin-lbeta. Paeonol was effective in inhibiting nitric oxide release from primary microglial cells. It also reduced the lipopolysaccharide-stimulated release of tumor necrosis factor-alpha and intefleukin-1β from microglial cells. Paeonol possesses neuroprotective activity in a model of inflammation-induced neurotoxicity and reduces the release of neurotoxic and proinflammatory factors in activated microglial cells.

Paeonol attenuated vascular fibrosis through regulating Treg/Th17 balance in gut microbiota-dependent manner

OBJECTIVE Atherosclerosis(AS)is a chronic inflammatory disease characterized by the accumulation of lipids,vascular fibrosis,and inflammation.Paeonol(Pae)is a natural phenolic compounds isolated from a traditional Chinese medicine,Cortex Moutan,which exhibits anti-AS effects.Our previous work demonstrated that gut microbiota plays an important role during AS treatment as it affects the efficacy of Pae.However,the mechanism of Pae in protecting against vascular fibrosis as related to gut microbiota has yet to be elucidated.To investigate the anti-fibrosis effect of Pae on AS mice and demonstrate the underlying gut microbiota-dependent mechanism.METHODS ApoE-/-mice were fed with high-fat-diet(HFD)to replicate the AS model.HE and Masson staining were used to observe the plaque formation and collagen deposition.Gut microbiota alteration and short-chain fatty acids(SCFAs)production were analyzed through 16S rRNA sequencing and LC-MS/MS.The frequency of immune cells in spleen were phenotyped by flow cytometry.The mRNA expression of aortic inflammatory cytokines were detected by qRT-PCR.The protein expression of LOX and fibrosis related indicators were examined by Western blotting.RESULTS Pae restricted the development of AS and collagen deposition.Notably,the anti-fibrosis effect of Pae was achieved by regulating the gut microbiota.16S rRNA sequencing and LC-MS/MS data indicated that the relative abundance of SCFAs-producing bacteria and SCFAs production was increased.Additionally,Pae administration selectively up-regulated the frequency of regulatory T(Treg)cells as well as down-regulated the ratio of T helper type 17(Th17)cells in the spleen of AS mice,improving the Treg/Th17 balance.In addition,as expected,Pae intervention significantly down-regulate the mRNA expression levels of pro-inflammatory cytokines IL^(-1)β,IL-6,TNF-αand IL^(-1)7 in the aorta tissue,up-regulate the levels of anti-inflammatory factor IL^(-1)0,a marker of Treg cells.Finally,Pae′s intervention in the gut microbiota resulted in the restoration of the balance of Treg/Th17,which indirectly down-regulated the protein expression level of LOX and fibrosis-related indicators(MMP-2/9 and collagenⅠ/Ⅲ).CONCLUSION Pae attenuates vascular fibrosis in a gut microbiota-dependent manner.The underlying protective mechanism is associated with the improved Treg/Th17 balance in spleen mediated through the increased microbiota-derived SCFAs production.

Protective effect of paeonol on beta-amyloid 25-35- induced toxicity in PC12 cells

BACKGROUND：Paeonol is a primary phenolic component of the Chinese medicinal herb Cortex moutan. Recent studies have shown that paeonol has anti-inflammatory, analgesic, and antioxidative effects as well as a significant cardioprotective effect against myocardial ischemia. OBJECTIVE： To investigate the protective effect of paeonol on β-amyloid 25-35-induced toxicity in PC12 cells and analyze its mechanism of action. DESIGN, TIME AND SETTING： A controlled repeated-measures cell-based study was performed in the Department of Pharmacology of Guangdong Medical College between September 2006 and December 2007. MATERIALS： Paeonol was supplied by Xuancheng Baicao Plant Industry and Trade Company, China. PC12 cells were a kind gift from Dr. Haitao Zhang at Guangdong Medical College. β-amyloid 25-35 was purchased from Sigma Company, USA. Lactate dehydrogenase （LDH） and malondialdehyde （MDA） kits were purchased from Nanjing Jiancheng Bioengineering Research Institute, China. METHODS： PC12 cells were maintained in Dulbecco＇s modified eagle＇s medium （DMEM） supplemented with 100 mL/L heat-inactivated horse serum and 50 mL/L fetal bovine serum at 37 ℃ and cultured in an incubator with 5% CO2. The medium was renewed every other day. Batches of cells were assigned into three groups. （1） Paeonol group： cells were preincubated with different concentrations of paeonol （12, 25 or 50 μmol/L） for one hour and β-amyloid 25-35 was added to the medium; （2） control group： cells were cultured in DMEM supplemented with 100 mL/L heat-inactivated horse serum and 50 mL/L fetal bovine serum; and （3） β-amyloid 25-35 group： β-amyloid 25-35 was added to the medium. MAIN OUTCOME MEASURES： When PC12 cells in each group were cultured for 24 hours, the cell viability was determined using the MTT reduction assay, LDH release into the culture media was measured by 2,4-dinitrophenylhydrazine chromatometry and MDA content was measured using a thiobarbituric acid assay. RESULTS： When PC12 cells were treated withβ-amyloid 25-35 （50 μmol/L） for 24 hours, their viability was significantly lower compared with the control group （P 〈 0.01）. When the cells were treated with paeonol for one hour prior to incubation withβ-amyloid 25-35, their viability was significantly increased compared with theβ-amyloid 25-35 group （P 〈 0.05–0.01）. LDH activity and MDA level in the β-amyloid 25-35 group were significantly increased compared with the control group （P 〈 0.01）. When the cells were treated with different concentrations of paeonol, LDH activity and MDA level in PC12 cells were significantly decreased compared with theβ-amyloid 25-35 group （P 〈 0.01）. CONCLUSION： Paeonol protects PC12 cells againstβ-amyloid 25-35-induced toxicity and the protective effect of paeonol is probably achieved through its antioxidative effects.

Preparation and Crystal Structure of Paeonol

Paeonol was prepared by the extraction method from Moutan Cortex. Its crystal struc- ture was determined by single-crystal X-ray diffraction.The compound crystallizes in the monoclinic sys- tem, space group P21/c with a = 6.724(4), b = 8.792(6), c = 14.689(10) ?, β = 100.138(11)o, V = 854.8(10) ?3, Mr = 166.17, Z = 4, F(000) = 352, Dc = 1.291 g/cm3, μ = 0.097 mm-1, the final R = 0.0454 and wR = 0.0988 for 845 observed reflections with I > 2σ(I).

Paeonol inhibits NLRP3 mediated inflammatory reaction in rat endothelial cells by elevating hyperlipidemic rats plasma exosomal miRNA-223

OBJECTIVE Atherosclerosis(AS)is featured as a chronic inflammatory disease of vascular stenosis.Paeonol(Pae)is a natural phenolic compounds isolated from a traditional Chinese medicine,Cortex Moutan,which exhibits anti-AS effects in vitro and in vivo.In this study,we aimed to investigate whether the anti-AS efficacy of Pae was regulated through inhibiting NLRP3 inflammasome activityvia elevating hyperlipidemic rats plasma exosomalmicroR⁃NA-223(miR-223).METHODS The Sprague-Dawley rats was induced by a high-fat diet,which was used as AS models.AS aortic pathological morphological in AS mice was examined by HE staining,and serum TC and TG levels were deter⁃mined by automatic chemistry analyzer.Rat aortic endothelial cells(RAECs)were used during the whole study.After oral administration of Pae,we isolated exosomes from hyperlipidemic rats plasma(Pae-Exos)by ultracentrifugation and characterized by transmission electron,nanoparticle tracking analysis,dynamic light scattering and Western blotting.The activity of RAECs was detected by CCK-8 and trypan blue staining method.IL-1βand IL-6 levels were detected by ELISA method.The expression of miR-223 was detected by qPCR,and the expression of NLRP3,ASC,caspase-1,and ICAM-1 was detected by Western blotting.RESULTS In vivo experiments confirmed that Pae could effectively reduce serum TC and TG levels and decrease serum IL-1βand IL-6 levels,which demonstrated that Pae restricted AS develop⁃ment in hyperlipidemia rats.Both CCK-8 and trypan blue staining showed that the survival rate of RAECs in the Pae-Exos co-incubation group was higher than that in the model group.We also confirmed via real-time qPCR that Pae-Exos suppressed the expression of the inflammatory cytokines IL-1βand IL-6.Accordingly,Pae-Exos dose-dependently increased the survival rate of RAECs and reduced inflammatory response.Furthermore,compared with the model group,Pae-Exos more successfully increased the expression of miR-223 and inhibited IL-1βand IL-6 expression,which implied that Pae-exo may inhibited the inflammatory response of RAECs by increasing the content of miR-223.Subse⁃quently,we found that Pae-Exos reduced the expressions of NLRP3,ASC,caspase-1 and ICAM-1,which indicated that Pae-Exos may reduced RAECs inflammation by suppressing NLRP3 signaling pathway via promoting miR-223 expression.CONCLUSION Pae can inhibit the downstream NLRP3 inflammatory corpuscle signaling pathway by increasing the level of miR-223 in plasma Exos of hyperlipidemic rats,providing new insights into the anti-atherosclerosis activity of Pae.

Effects of Paeonol Combined with Panax Notoginseng Saponins on Nrf2/ARE Pathway in Rats with Diabetic Cardiomyopathy

[Objectives]To investigate whether paeonol(Pae)combined with panax notoginseng saponins(PNS)can protect the myocardium of rats with diabetic cardiomyopathy(DCM)through improving the antioxidant capacity of the rats by activating the Nrf2/ARE pathway.[Methods]The rats were fed with high-fat and high-sugar diet for 6 weeks,and combined with intraperitoneal injection of small-dose STZ to build a type II diabetes model;the model rats were randomly divided into a model group,a Pae group of 80 mg/kg,and a PNS group of 100 mg/kg,a Pae 80 mg/kg+PNS 100 mg/kg,and a metformin group 157.5 mg/kg;rats of normal group and model group were injected with an equal volume of sodium carboxymethyl cellulose(1%),10 rats in each group.After 8 weeks,3 rats in the model group were taken for histopathological examination.Changes in myocardial fibrosis and myocardial collagen formation indicated that the building of DCM model was successful.qRT-PCR and Western blot were used to detect the expression levels of Nrf2,HO-1,Col-I,and Col-III in myocardial tissue of each group of rats.[Results]Compared with the normal group,the mRNA and protein expression levels of Nrf2 and HO-1 in myocardial tissue of the DCM group rats were significantly reduced,and the mRNA and protein expression levels of Col-I and Col-III were significantly increased;compared with the DCM group,the mRNA and protein expression levels of Nrf2 and HO-1 in the myocardial tissue of each drug group increased to varying degrees,and the combined drug group increased more significantly than that of the single drug group;the mRNA and protein expression levels of Col-I and Col-III were reduced to varying degrees,and the combined drug group declined more significantly than that of the single drug group.[Conclusions]Paeonol combined with panax notoginseng saponins can up-regulate the expression of Nrf2 and HO-1 in myocardial tissue,inhibit the expression of type I and type III collagen.The mechanism may be related to improving the DCM myocardial fibrosis through activating the Nrf2/ARE pathway.

Protective Effect of Paeonol on Diabetic Cardiomyopathy in Rats

[Objectives] The protective effect of paeonol on DCM was further confirmed by observing the changes of myocardial structure and function in rats. [Methods] Rats were fed high-sugar and high-fat diet and injected intraperitoneally with streptozotocin(STZ) for two consecutive days to prepare animal model of type II diabetes. The model rats were randomly divided into model group, low-dose paeonol group(30 mg/kg), middle-dose paeonol group(60 mg/kg), high-dose paeonol group(120 mg/kg) and metformin group(157 mg/kg). The detected indicators included heart weight index(HWI), blood glucose(FBG), body mass, triglyceride(TG), and total cholesterol(TC). Myocardial pathological changes were observed by HE staining. The extent of myocardial fibrosis was observed by Masson staining. [Results] In the model group, the body weight of the rats declined significantly, and the phenomena of overdrinking, overeating and polyuria occurred. Compared with the model group, the body weight of rats in the paeonol treatment groups increased to different extents. Compared with the normal group, the HWI, FBG, LDH, AST, and TG increased in the model group, and decreased in different degrees in the paeonol treatment groups(P<0.05), suggesting that paeonol has certain protective effect on diabetic cardiomyopathy in rats. After HE staining and fixation, the myocardial tissue of rats was observed by optical microscope. The morphology of myocardial cells and myocardial fibers in the normal group were normal;Compared with the normal group, the model group has hypertrophy of myocardial cells, disordered arrangement of myocardial fibers, fiber breakage and dissolution, uneven staining between myocardial cells, and nucleus breakage or even disappearance;Compared with the model group, the pathological changes of myocardium in the paeonol treatment groups and DMBG group have been improved to different degrees. The arrangement of myocardial fibers is regular and the coloration is uniform. The results of Masson staining showed that in the normal group, and there was no significant increase of myocardial collagen fibers in the myocardial interstitium. Compared with the normal group, myocardial cells and interstitial collagen fibers(blue) in the model group were significantly increased. Compared with model group, myocardial collagen fiber hyperplasia in the paeonol treatment groups and DMBG group showed different degrees of improvement, among which the Pae-H group had the least blue bands and the most obvious improvement of collagen fiber. [Conclusions] Paeonol has different degrees of improvement effects on myocardial fibrosis in rats with diabetic cardiomyopathy, suggesting that paeonol has a protective effect on diabetic cardiomyopathy in rats.

Effects of Paeonol on Expression of Type Ⅰ and Type Ⅲ Collagen of High Glucose Induced Proliferation of Cardiac Fibroblasts

[Objectives]To explore the effects of paeonol on the inhibition of myocardial fibrosis in high glucose induced cardiac fibroblasts(CFs).[Methods]Differential adherence method was used to culture the primary CFs of neonatal rats(passage culture);CCK-8 method was used to detect the cell proliferation;MTT assay was used to screen the safe concentration of paeonol;the 2 nd to 3 rd generation CFs were randomly divided into normal group(5.5 mmol/L,expressed in C),high glucose group(30 mmol/L,expressed in HG),paeonol low dose group(Pae-L,17.5 mg/L),and medium dose paeonol group(Pae-M,35 mg/L),paeonol high dose group(Pae-H,70 mg/L);Western Blot method was used to detect the expression of Col-I and Col-III protein.[Results]The extraction of CFs from primary neonatal rats was successful;high glucose(30 mmol/L)induction had a significant proliferation effect on CFs;compared with the normal group,the expressions of COI-I and Col-III protein were increased in the high glucose group(P<0.05);compared with the high glucose group,the expression of COI-I protein was decreased in each treatment group(P<0.05,P<0.01),and the decrease was most significant in the high dose group(P<0.01);the expression of COI-III in the high dose group was decreased and it was statistically significant(P<0.05).[Conclusions]Paeonol significantly inhibited the proliferation of high glucose induced CFs in neonatal rats.This experiment is intended to provide a new experimental basis for the prevention and treatment of diabetic cardiomyopathy(DCM).

Effects of Paeonol on CRP and NF-κB p65 in Rats with Atherosclerosis

[Objectives] The protective mechanism of paeonol on atherosclerotic vessels was investigated by detecting CRP content in the serum and expression of NF-κB p65 in the aorta of atherosclerotic rats. [Methods] An animal model of AS rats was prepared by high-fat diet combined with intraperitoneal injection of VD3. The rats were randomly divided into the normal group,model group,high-dose paeonol group( 120 mg/kg),medium-dose paeonol group( 60 mg/kg),low-dose paeonol group( 30 mg/kg),and simvastatin group( 10 mg/kg) as the positive control group,and drug intervention was continued for 16 weeks. CRP content in the serum was detected by ELISA method,and the expression level of NF-κB p65 was detected by RT-PCR and Western-blot method. [Results]Compared with the normal group,CRP content in the serum and the expression of NF-κB p65 mRNA and protein in the aorta of rats in the model group significantly increased. Compared with the model group,CRP content in the serum dropped obviously,and the expression of NF-κB p65 mRNA and protein in the aorta of rats significantly reduced in the administration groups. Moreover,the improvement of each indicator in the high-dose paeonol group was obviously better than that of the medium-dose and low-dose groups. [Conclusions] Paeonol may protect atherosclerotic vessels by reducing serum CRP content,inhibiting the expression of NF-κB p65 and reducing inflammation.

Investigation of paeonol-geniposide on acute alcoholic liver injury based on uniform design method

Objective:To explore the optimal ratio and compatibility effect of paeonol-geniposide combination on acute alcoholic liver injury by uniform design.Methods:Lieber-DeCarli alcoholic liquid feed was used to induce acute alcoholic liver injury in mice.Uniform design was used to select the best dosage combination of paeonol and geniposide,and the related indexes of liver injury and oxidative stress were detected by kit.Serum inflammatory factors were detected by ELISA,and the expressions of p38 MAPK,JNK and NF-κB P65 related proteins in liver were detected by Western-blot.Results:The regression equation suggested that paeonol:geniposide=220:20 was the best ratio of paeonol and geniposide to resist alcoholic liver injury.Compared with the model group,the liver injury indexes and oxidation products of the paeonol+geniposide group decreased significantly,the antioxidant activity of liver tissue increased significantly,and the expression levels of p-p38 MAPK,p-JNK and NF-κB P65 protein decreased significantly.Conclusion:The optimal dosage of paeonolgeniposide was effectively optimized by uniform design and pharmacodynamic analysis.The combination of the two drugs could reduce the alcoholic liver injury by reducing the oxidative stress injury and inflammatory response in the liver tissue of mice,and its effect might be related to the targeting of p38 MAPK/JNK/NF-κB channel.

Prescription Design and Preparation Process of Paeonol Bead Popping Gum with Hypoglycemic Effect

[Objectives]To develop a paeonol bead popping gum with hypoglycemic effect.[Methods]The paeonol bead popping gum was prepared by the"two-step method",that is,the pill core was prepared by the guttate pill method,and then the coating was cured by sodium alginate solution and CaCl_(2) solution.The single factor method was used to determine the effects of PEG-4000:paeonol dosage ratio,dropper diameter,condensation time,dropping distance,melting temperature on the comprehensive score of paeonol guttate pill,and the effects of sodium alginate solution concentration,CaCl_(2) solution concentration,number of coating layers,drying time on the comprehensive score of popping gum.Finally,the optimal process was determined and verified by orthogonal experiment method.[Results]When the dosage ratio of PEG-4000:paeonol was 4∶1,the dropper diameter was 4 mm,the condensation time was 5 min,the dropping distance was 6 cm,and the melting temperature was 90℃,the quality of the prepared guttate pill was the optimal.When the concentration of sodium alginate solution was 0.02 g/mL,the concentration of CaCl_(2) solution was 0.25 g/mL,the number of coating layers was 3,and the drying time was 25 min,the appearance and comprehensive score of the obtained popping beads were the optimal.[Conclusions]This study is expected to provide some reference and basis for the development and utilization of hypoglycemic products of traditional Chinese medicine.

Determination of Paeonol in Detergent by High Performance Liquid Chromatography

By analyzing the ultraviolet absorption spectrum of paeonol, selecting the comparatively good test wavelength and separating the appropriate relative flow detergent, the paeonol content in the detergent was measured. By measuring the repeatability, precision and recovery rate, a method for the determination of paeonol in detergent by high performance liquid chromatography was finally established and verified. The experimental results show that the standard solution is linearly correlated between 0.1 and 20 μg ? mL and the recovery rate is 99% to 101%.

Influence of paeonol combined with γ ray on glioma cell apoptosis and apoptosis gene expressiona

Objective: To study the influence of paeonol combined with γ ray on glioma cell apoptosis and apoptosis gene expression. Methods: Glioma C6 cells were cultured and divided into control group (treated with serum-free DMEM), ray group (exposed to 4 Gy γ ray), Pae group (treated with serum-free DMEM contains 4 μg/mL paeonol) and γ+Pae group (treated with serum-free DMEM contains 4 μg/mL paeonol and exposed to 4 Gy γ ray). The cell proliferation activity value as well as the expression of pro-apoptosis genes and anti-apoptosis genes was measured after the intervention. Results: After 6 h, 12 h, 18 h and 24 h of intervention, cell proliferation activity value of ray group, Pae group and γ+Pae group were greatly lower than that of control group, and cell proliferation activity value of γ+Pae group was obviously lower than that of ray group and Pae group. After 24 h of intervention, p21cip1, TRAIL, LRIG1, NPRL2 and SEPT7 mRNA expression in ray group, Pae group and γ+Pae group were notably higher than those in control group whereas PIAS3, CEACAM1, EZH2, MDM2, c-myc and Survivin mRNA expression were greatly lower than those in control group;p21cip1, TRAIL, LRIG1, NPRL2 and SEPT7 mRNA expression in γ+Pae group were remarkably higher than those in ray group and Pae group whereas PIAS3, CEACAM1, EZH2, MDM2, c-myc and Survivin mRNA expression were greatly lower than those in ray group and Pae group. Conclusion: Paeonol combined with γ ray can be more effective than γ ray alone in inducing the apoptosis of glioma cells, increasing the expression of pro-apoptosis genes and decreasing the expression of anti-apoptosis genes.

Antiproliferation and apoptosis induction of paeonol in HepG_2 cells

AIM: To investigate the antiproliferative effect of paeonol (Pae) used alone or in combination with chemotherapeutic agents [cisplatin (CDDP), doxorubicin (DOX) and 5-fluorouracil (5-FU)] on human hepatoma cell line HepG2 and the possible mechanisms. METHODS: The cytotoxic effect of drugs on HepG2 cells was measured by 3-(4, 5-dimethylthiazol-2- yl)-2, 5-diphenyltetra-zolium bromide (MTT) assay. Morphologic changes were observed by acridine orange (AO) fluorescence staining. Cell cycle and apoptosis rate were detected by flow cytometry (FCM). Drug-drug interactions were analyzed by the coefficient of drug interaction (CDI). RESULTS: Pae (7.81-250 mg/L) had an inhibitory effect on the proliferation of HepG2 cells in a dose-dependent manner, with the IC50 value of (104.77 ± 7.28) mg/L. AO fluorescence staining and FCM assays showed that Pae induced apoptosis and arrested cell cycle at S phase in HepG2 cells. Further, different extent synergisms were observed when Pae (15.63, 31.25, 62.5 mg/L) was combined with CDDP (0.31-2.5 mg/L), DOX (0.16-1.25 mg/L), or 5-FU (12.5-100 mg/L) at appropriate concentrations. The IC50 value of the three drugs decreased dramatically when combined with Pae (P < 0.01). Of the three different combinations, the sensitivity of cells to drugs was considerably different.CONCLUSION: Pae had a significant growth-inhibitory effect on the human hepatoma cell line HepG2, which may be related to apoptosis induction and cell cycle arrest. It also can enhance the cytotoxicity of chemotherapeutic agents on HepG2 cells, and the S phase arrest induced by Pae may be one of the mechanisms of these interactions.

Paeonol reduces microbial metabolite α-hydroxyisobutyric acid to alleviate the ROS/TXNIP/NLRP3 pathway-mediated endothelial inflammation in atherosclerosis mice

Gut microbiota dysbiosis is an avenue for the promotion of atherosclerosis(AS)and this effect is mediated partly via the circulating microbial metabolites.More microbial metabolites related to AS vascular inflammation,and the mechanisms involved need to be clarified urgently.Paeonol(Pae)is an active compound isolated from Paeonia suffruticoas Andr.with anti-As inflammation effect.However,considering the low oral bioavailability of Pae,it is worth exploring the mechanism by which Pae reduces the harmful metabolites of the gut microbiota to alleviate AS.In this study,ApoE--mice were fed a high-fat diet(HFD)to establish an AS model.AS mice were administrated with Pae(200 or 400 mg:kg')by oral gavage and fecal microbiota transplantation(FMT)was conducted.16S rDNA sequencing was performed to investigate the composition of the gut microbiota,while metabolomics analysis was used to identify the metabolites in serum and cecal contents.The results indicated that Pae significantly improved AS by regulating gut microbiota composition and microbiota metabolic profile in AS mice.We also identified a-hydroxyisobutyric acid(HIBA)as a harmful microbial metabolite reduced by Pae.HIBA supplementation in drinking water promoted AS inflammation in AS mice.Furthermore,vascular endothelial cells(VECs)were cultured and stimulated by HIBA.We verified that HIBA stimulation increased intracellular ROS levels,thereby inducing VEC inflammation via the TXNIP/NLRP3 pathway.In sum,Pae reduces the production of the microbial metabolite HIBA,thus alleviating the ROS/TXNIP/NLRP3 pathway-mediated endothelial inflammation in AS.Our study innovatively confirms the mechanism by which Pae reduces the harmful metabolites of gut microbiota to alleviate AS and proposes HIBA as a potential biomarker for AS clinical judgment.

Fluorimetric analysis of paeonol in Chinese herbal medicine Cynanchi Paniculati Radix by aluminum ion-sensitized fluorescence

A novel fluorimetric method for determination of paeonol,an active component of Chinese herbal medicine,is proposed.The method is based on the reaction of paeonol with aluminum ion in pH 4.4 HAc-NaAc buffer to form a fluorescent Al(Ⅲ)-paeonol complex.The maximum excitation wavelength and emission wavelength of the complex were 296 nm and 455 nm,respectively.The fuorescence quantum yield of the complex was determined to be 0.053 at an excitation wavelength of 296 nm.A linear calibration curve covered the concentration range 0.017-1.2μg/mL.The method has been applied to the analysis of paeonol in medicinal crop Cynanchi Paniculati Radix(Xuchangqing),and the results demonstrated that this method can be used for quality evaluation of crude drug Xuchangqing.

Box-Behnken响应面法优选紫斑牡丹中丹皮酚和芍药苷提取工艺

目的优选紫斑牡丹中丹皮酚和芍药苷的提取工艺。方法在单因素试验基础上,以液料比、提取时间和乙醇体积分数为影响因素,以丹皮酚和芍药苷提取率的总权重综合评分为评价指标,采用Box-Behnken响应面法优选最佳提取工艺,并进行验证试验。结果最佳提取工艺为液料比28倍,提取时间150 min,乙醇体积分数52%。此工艺条件下,丹皮酚提取率为0.49%(RSD=1.33%,n=3),芍药苷提取率为5.38%(RSD=1.07%,n=3)。结论该提取工艺稳定、可行,可用于紫斑牡丹中丹皮酚和芍药苷的提取。