The Mechanism of the combination of radix paeoniae rubra-cortex moutan in treating cerebral hemorrhage based on network pharmacology

Objective:To clarify the material basis of Chinese medicine pair“Radix Paeoniae Rubra-Cortex Moutan”(Chishao-Mudanpi)and explore their mechanism in the treatment of ICH with network pharmacology.Methods:The active ingredients contained in Radix Paeoniae Rubra and Cortex Moutan were searched and selected based on the oral bioavailability prediction and drug-likeness prediction from the TCMSP database.Then the targets of cerebral hemorrhage were collected from GeneCards,OMIM,and DrugBank databases.After obtained the intersections of drugs and disease,the active component target disease interactive network diagram was drawn by Cytoscape software.The obtained key targets were uploaded to the STRING database for analysis and construct a PPI network map.GO function enrichment analysis and KEGG analysis were performed on the key target proteins.Results:Collected the active ingredients of Radix Paeoniae 119,Radix Paeoniae 55,including paeoniflorin,baicalin,β-sitosterol,etc.Related drug target protein 1190,ICH disease-related genes 823,"Radix Paeoniae-Radix Paeoniae"and 72 common targets of ICH,mainly acting on Akt1,IL6,VEGFA,CASP3,EGF,involving 133 related signaling pathways such as AGE-RAGE,TNF,IL-17,HIF1,PI3K-Akt.Conclusion:The combination of"Radix Paeoniae Rubra-Cortex Moutan"in the treatment of ICH has the characteristics of multiple pathways and multiple targets,which provides a reference and basis for further molecular biology verification in the future.

Effect of radix paeoniae rubra on expression of p38 MAPK/iNOS/HO-1 in rats with lipopolysaccharide·induced acute lung injury

Objective: To investigate the effect of radix paeoniae rubra (RPR) on expression of p38 mitogen activated protein kinase ( MAPK )/iNOS/HO-1 in rats with lipopolysaccharide-induced acute lung injury and explore the molecular mechanism.Methods: Forty healthy male Wistar rats, weighing 200-250 g, aged 6-8 weeks (mean =7 weeks), provided by the Experimental Center, Medical College, Wuhan University, Wuhan, China, were employed in this study.Under anesthesia with 7% chloraldurat (5 ml/kg body weight) through intraperitoneal injection, the trachea of the rat was exposed and an arterial puncture needle pricked into the trachea via cricothyroid membrane. Then they were randomly divided into five groups: 8 rats receiving 1 ml normal saline through the puncture needle (Group A),8 receiving 1 ml lipopolysaccharide (LPS, 2.5 mg/kg,Group B), 8 receiving LPS and RPR (30 mg/kg, pumped through the femoral vein for 2 hours, Group C ), 8 receiving RPR 2 hours before dripping LPS ( Group D),and 8 receiving hemin (75 μmol/L through intraperitoneal injection) 18 hours before dripping LPS (Group E). After 6 hours of LPS dripping, blood samples were obtained through the carotid artery to perform blood gas analysis,then all the rats were exsanguinated to death and specimens of lung tissues were obtained. The pathomorphological changes of the lung tissues were observed. The expression of p38 MAPK/iNOS/HO-1, the neutrophil ratio, protein content in alveolar irrigating solution and malonaldehyde (MDA) content in the lung tissues were also detected.Results: Compared with Group A, the expression of p38 MAPK, iNOS and HO-1 markedly increased in Groups B, C, D, and E (P ＜0.01). But in Groups C, D, and E,the expression of p38 MAPK and iNOS were significantly lower than that of Group B, while expression of HO-1 was obviously higher than that of Group B ( P ＜ 0.05 ). The protein content, the ratio of neutrophils in bronchoalveolar lavage fluid ( BALF), the content of MDA and the activities of serum NO in Group B were significantly higher than those of Group A ( P ＜ 0.01 ). There was a significant decrease in the level of arterial bicarbonate and partial pressure of oxygen in Group B (P ＜ 0.01). Compared with Group B, these indexes of lung injury were significantly lower while the levels of arterial bicarbonate and partial pressure of oxygen increased significantly in Groups C, D,and E (P ＜ 0.05 or P ＜ 0.01 ). Under light microscope, the pathological changes induced by LPS were significantly attenuated by RPR and hemin.Conclusions: The high expression of MAPK plays an important role in lipopolysaccharide-induced acute lung injury. Protective effect of RPR on lipopolysaccharideinduced acute lung injury may be related to the inhibition of the abnormal high expression of p38 MAPK/iNOS/HO-1.

Mechanism of total glucosides from Chishao(Radix Paeoniae Rubra)on proliferation and apoptosis of hepatocellular carcinoma cells via phosphatase and tensin homolog deleted on chromosome ten/phosphatidylinositol 3-kinase/protein kinase B signaling pathway

OBJECTIVE:To investigate the possible molecular mechanism of total glycosides of Chishao(Radix Paeoniae Rubra)(TG-RPR)on proliferation and apoptosis of hepatocellular carcinoma cells.METHODS:The proliferation of TG-RPR on Hep G2 cells was detected using the 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide(MTT)assay.The apoptosis of Hep G2 cells was measured by annexin V-FITC/double staining.The phosphatase and tensin homolog deleted on chromosome ten(PTEN)/phosphatidylinositol 3-kinase(PI3 K)/protein kinase B(Akt)signaling pathway was evaluated by Western Blot and reverse transcription-polymerase chain reaction(RT-PCR).RESULTS:TG-RPR can up-regulation the expression of pro-apoptotic factors such as PTEN and BCL2-Associated X(Bax),down-regulation the expression of anti-apoptotic factors including B-cell lymphoma-2(Bcl-2),PI3 K,and Akt.CONCLUSION:TG-RPR significantly inhibits the proliferation of Hep G2 cells in a dose-dependent manner and promotes apoptosis.These results demonstrated TG-RPR has significant inhibitory effect on Hep G2 cells.These results identify a critical role of TG-RPR in proliferation and apoptosis of Hep G2 cells via modulating PTEN/PI3 K/Akt signaling pathway.TG-RPR may offer a promise as a potential pharmaceutical therapy for hepatocellular carcinoma.

Moutan Cortex and Paeoniae Radix Rubra reverse high-fat-diet-induced metabolic disorder and restore gut microbiota homeostasis

The present study was designed to investigate the therapeutic effcts of Moutan Cortex(CM,root bark of Paeonia suffruticosa Andr) and Paeoniae Radix Rubra(PR,root of Paeonia veitchii Lynch) on metabolic disorders,focusing on the infuence of CM and PR on the obesity-related gut microbiota homeostasis.The diet-induced obese(DIO) mouse model was used to test the therapeutic effects of CM and PR.The mice were orally administered with CM and PR for 6 weeks,and oral glucose tolerance test(OGTT) and insulin tolerance test(ITT) were performed to evaluate the insulin sensitivity of the mice.Sterol-regulatory element binding proteins(SREBPs) and their target genes were measured by quantitative RT-PCR.High-throughput 16 S ribosomal RNA(16S rR NA) gene sequencing technology was used to determine the composition of gut microbiota,and the metabolites in serum were analyzed by GC-MS.Our results indicated that CM and PR combination alleviated obese and insulin resistance in the DIO mice,leading to increased glucose uptake and gene expression in muscle and liver,and down-regulated SREBPs and their target genes in liver.Interesting,neither the CM-PR extracts,nor the major components of CM and PR did not affect SREBPs activity in cultured cells.Meanwhile,CM and PR significantly modulated the gut microbiota of the high-fat diet(HFD) treated mice,similar to metformin,and CM-PR reversed the overall microbiota composition similar to the normal chow diet(NCD) treated mice.In conclusion,our results provide novel mechanisms of action for the effects of CM and PR in treating DIO-induced dysregulation of sugar and lipid metabolism.

"Dose-effect-response" Relationships of Paeoniae Radix Rubra on α-Naphthylisothiocyanate-induced Acute Cholestatic Hepatitis in Rats

Objective To investigate the hepatoprotective effects of Paeoniae Radix Rubra(PRR)at different doses against α-naphthylisothiocyanate(α-NIT)-induced acute cholestatic hepatitis in rats.Methods Rats were ig admini-strated with vehicle or PRR[(1,9,18,36,54,72,and 81 g/(kg·d)]3 d before and 2 d afterα-NIT(60 mg/kg)ig administration.The general status of rats,histopathology of liver,serum alanine aminotransaminase,aspartate aminotransaminase,total bilirubin,direct bilirubin,and alkaline phosphatase levels,were observed at respective time points(24 and 48 h)afterα-NIT administration.Using cluster analysis and correspondence analysis,the 'dose-effect-response'relationships of PRR were evaluated.Results The results showed that compared with model group,the serum biochemistry index significantly decreased with the increasing of PRR dosage(P<0.01), and the change and necrosis of hepatic cellula,and inflammatory cell infiltration were gradually alleviated. However,the improvement was not obviously found in the low-dose group[1 g/(kg·d)].The cluster analysis and correspondence analysis results showed that different doses of PRR could significantly ameliorateα-NIT-induced acute cholestatic hepatitis of rats in a dose-dependent manner.Conclusion The experiments show that administration doses of PRR in clinical use should be added properly in order to gain the expectant therapeutic effect,especially in the treatment of heavy acute cholestasis hepatitis.

Investigation on the Mode of Action of the Traditional Chinese Medical Prescription-Yiqihuoxue Formula, an Effective Extravasation Treatment for Cerebral Vascular Microemboli in ApoE-/- mice

Objective: The objective of this study was to investigate the mechanisms underlying anti-embolism and extravasational effects of traditional Chinese medical prescription YiqiHuoxue(YQHX) formula in ApoE-/-mice with cerebral vascular microemboli. Materials and Methods: An ApoE-/-mice model with microemboli was developed by infusing fluorescently labeled heterologous fibrin-rich microparticles into the internal carotid artery of ApoE -/-gene knockout male mice through the common carotid artery. Before microemboli injection, the animals were randomly divided into four groups of 10 animals, treated daily for 6 weeks by intragastric administration: The ApoE-/-control group(physiological saline, 0.2 mL/10 g/d), YQHX group(0.2 ml/10 g/d), clopidogrel group(3 mg/kg/d), and atorvastatin group(3 mg/kg/d);a further group was constituted of normal male C57 BL/6 J mice(with the same genetic background as ApoE-/-mice;normal control group;no treatment;microemboli injection). The mice in each microemboli group were divided into three subgroups, the 2-h, 24-h, and 72-h subgroups, corresponding to the time after microemboli injection. Two hours(or 24 h or 72 h) after microemboli injection, the changes in aortic intima and brain tissue were analyzed by histopathology, the amounts of fluorescent emboli being measured by fluorescence microscopy image analysis. Comparison points included the microemboli induced loss of aorta functions and pathological changes, atherosclerotic plaque, brain ultrastructure and functions, and embolus extravasation. Results: Loss of aorta functions and adverse pathological changes, atherosclerotic plaque, serious damage in brain ultrastructure and functions, and reduced thrombus elimination were obviously serious in microemboli injected ApoE-/-mice. These symptoms were significantly relieved by the YQHX pretreatment:(i) the ratio of thrombus accumulation was increased with a significant decrease in thrombus extravasation in ApoE-/-mice, while YQHX induced an increased thrombus extravasation;(ii) the degree of aortic intimal thickening and brain tissue structural disorders were significantly increased in ApoE-/-mice, but overtly inhibited in the YQHX group;(iii) YQHX restored cell viability and homeostasis in the brain;(iv) YQHX regulated the expression of pro-and anti-inflammatory cytokines in the aorta;and(v) YQHX reduced cortical nerve nuclei pyknosis, edema, liquefaction, and necrosis induced by brain hypoxia, especially in the 24 h and 72 h groups. Conclusions: These findings indicate that the protective effects of YQHX on the brain against microemboli-induced injury may be attributed to the activation of extravasation mechanisms, which are involved in the cerebrovascular injury pathway and constitutively important in the progression of ischemic stroke.