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Objective@#To excavate the mechanism of the combination of Radix Ophiopogonis and Schisandra chinensis to treatatherosclerosisbased on network pharmacology to discuss its mechnism.@*Methods@#This paper excavated the associated proteins with Radix Ophiopogonis and Schisandra chinensis from the TCMGeneDIT database, and constructed the multicomponent protein network of Radix Ophiopogonis, Schisandra chinensis and proteins ApoE-/- mice were randomly divided into control group, model group, low, medium, high dose group and atorvastatin calcium group. Except the control group, other groups were fed with H10540 high fat diet for 12 weeks. From the 4th week, the atrovastatin calcium group was given atrovastatin calcium liquid 6 mg/kg by gavage. The low, medium and high dose groups were administed 4.68, 2.34 and 1.17 g/kg, respectively, once a day by gavage for 8 weeks. The oil red staining was applied to observe the pathological changes of atherosclerotic aortic wall. Western blot was subjected to detect the expression change of mitogen activated protein kinases p38 (p38), ATP binding cassette subfamily G member 1 (ABCG1), Toll like receptor 4 (TLR4), heat shock protein 90 alpha family class a member 1 (HSP90AA1), MMP-9 and arachidonate 5-lipoxygenase (ALOX5) in liver tissue, as well as nuclear factor related factor 2 (Nrf2) and heme oxygenase 1 (HO-1) in brain tissue.@*Results@#It was found that eleven components were interacted with 37 proteins, forming a protein interaction network with 48 nodes and 190 boundaries without isolated nodes. Compared to the model group, the level of p-p38/p38 (2.12 ± 0.12, 1.76 ± 0.11, 1.69 ± 0.10 vs. 2.45 ± 0.16), TLR4 (1.98 ± 0.10, 1.64 ± 0.11, 1.55 ± 0.12 vs. 2.68 ± 0.06), HSP90AA1 (1.79 ± 0.10, 1.66 ± 0.09, 1.59 ± 0.11 vs. 2.06 ± 0.07), MMP9 (1.84 ± 0.11, 1.75 ± 0.12, 1.66 ± 0.08 vs. 2.68 ± 0.10) in liver tissue of low, medium and high dose groups significantly decreased, the level of ABCG1 (0.53 ± 0.08, 0.78 ± 0.09, 0.81 ± 0.10 vs. 0.45 ± 0.04), ALOX5 (0.59 ± 0.04, 0.67 ± 0.09, 0.88 ± 0.07 vs. 0.47 ± 0.02) in liver tissue of low, medium and high dose groups significantly increased (P<0.05). The expression of Nrf2 (1.62 ± 0.12, 1.32 ± 0.09, 1.14 ± 0.06 vs. 2.12 ± 0.08) in cytoplasm of brain tissue significantly decreased, and Nrf2 (1.12 ± 0.09, 1.61 ± 0.07, 1.68 ± 0.11 vs. 1.07 ± 0.08) in cell nucleus of brain tissue significantly increased. The expression of HO-1 (1.16 ± 0.09, 1.73 ± 0.11, 1.82 ± 0.08 vs. 1.05 ± 0.04) in brain tissue significantly increased.@*Conclusions@#Network pharmacology and molecular biology were used to elucidate the molecular mechanism of the combination of Schisandra chinensis and Ophiopogon japonicus in the prevention and treatment of atherosclerosis, also to validate the related mechanism via Nrf2 pathway, which provided a reference for the further study of the combined prescription.
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Objective:To excavate the mechanism of the combination of Radix Ophiopogonis and Schisandra chinensis to treatatherosclerosisbased on network pharmacology to discuss its mechnism.Methods:This paper excavated the associated proteins with Radix Ophiopogonis and Schisandra chinensis from the TCMGeneDIT database, and constructed the multicomponent protein network of Radix Ophiopogonis, Schisandra chinensis and proteins ApoE-/- mice were randomly divided into control group, model group, low, medium, high dose group and atorvastatin calcium group. Except the control group, other groups were fed with H10540 high fat diet for 12 weeks. From the 4th week, the atrovastatin calcium group was given atrovastatin calcium liquid 6 mg/kg by gavage. The low, medium and high dose groups were administed 4.68, 2.34 and 1.17 g/kg, respectively, once a day by gavage for 8 weeks. The oil red staining was applied to observe the pathological changes of atherosclerotic aortic wall. Western blot was subjected to detect the expression change of mitogen activated protein kinases p38 (p38), ATP binding cassette subfamily G member 1 (ABCG1), Toll like receptor 4 (TLR4), heat shock protein 90 alpha family class a member 1 (HSP90AA1), MMP-9 and arachidonate 5-lipoxygenase (ALOX5) in liver tissue, as well as nuclear factor related factor 2 (Nrf2) and heme oxygenase 1 (HO-1) in brain tissue.Results:It was found that eleven components were interacted with 37 proteins, forming a protein interaction network with 48 nodes and 190 boundaries without isolated nodes. Compared to the model group, the level of p-p38/p38 (2.12 ± 0.12, 1.76 ± 0.11, 1.69 ± 0.10 vs. 2.45 ± 0.16), TLR4 (1.98 ± 0.10, 1.64 ± 0.11, 1.55 ± 0.12 vs. 2.68 ± 0.06), HSP90AA1 (1.79 ± 0.10, 1.66 ± 0.09, 1.59 ± 0.11 vs. 2.06 ± 0.07), MMP9 (1.84 ± 0.11, 1.75 ± 0.12, 1.66 ± 0.08 vs. 2.68 ± 0.10) in liver tissue of low, medium and high dose groups significantly decreased, the level of ABCG1 (0.53 ± 0.08, 0.78 ± 0.09, 0.81 ± 0.10 vs. 0.45 ± 0.04), ALOX5 (0.59 ± 0.04, 0.67 ± 0.09, 0.88 ± 0.07 vs. 0.47 ± 0.02) in liver tissue of low, medium and high dose groups significantly increased ( P<0.05). The expression of Nrf2 (1.62 ± 0.12, 1.32 ± 0.09, 1.14 ± 0.06 vs. 2.12 ± 0.08) in cytoplasm of brain tissue significantly decreased, and Nrf2 (1.12 ± 0.09, 1.61 ± 0.07, 1.68 ± 0.11 vs. 1.07 ± 0.08) in cell nucleus of brain tissue significantly increased. The expression of HO-1 (1.16 ± 0.09, 1.73 ± 0.11, 1.82 ± 0.08 vs. 1.05 ± 0.04) in brain tissue significantly increased. Conclusions:Network pharmacology and molecular biology were used to elucidate the molecular mechanism of the combination of Schisandra chinensis and Ophiopogon japonicus in the prevention and treatment of atherosclerosis, also to validate the related mechanism via Nrf2 pathway, which provided a reference for the further study of the combined prescription.
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Objective To observe the effect of hydroxysafflor yellow A (HSYA) on the protection of blood brain barrier of cerebral ischemia mice, and explore the mechaniam. Methods Seventy-two C57/BL mice were divided into 6 groups: the sham operation group, the cerebral ischemia mice group, the TLR4 blocking group, the TLR4 blocking+cerebral ischemia mice group, the HSYA intervention+cerebral ischemia mice group, HSYA intervention+TLR4 blocking+cerebral ischemia mice group. Cerebral ischemia mice group were subjected to the middle cerebral artery occlusion (MCAO) model, TLR4 blocking was used, while TLR4 blocking was injected TLR4 antibody via right common carotid artery, and HSYA intervention group was injected 2 mg/kg HSYA by tail vein 0.5 h before cerebral ischemia. RT-PCR and western blot were applied to detect the mRNA and protein expression change of Wnt3a and β-catenin in each group. Results Compared with the cerebral ischemia mice group,the expression of TLR4 mRNA(1.63 ± 0.05,1.53 ± 0.04,1.84 ± 0.03 vs. 1.97 ± 0.05) significantly decreased (P<0.05 or P<0.01), the Wnt3a mRNA (0.56 ± 0.01, 0.58 ± 0.01, 0.50 ± 0.04 vs.0.42 ± 0.03),β-catenin mRNA(0.61 ± 0.03,0.74 ± 0.02,0.58 ± 0.04 vs.0.50 ± 0.03),Claudin-5 mRNA (0.54 ± 0.02, 0.58 ± 0.01, 0.47 ± 0.01 vs. 0.46 ± 0.02) mRNA significantly increased(P<0.05 or P<0.01), the protein expression of TLR4 (1.73 ± 0.05, 1.57 ± 0.03, 1.79 ± 0.08 vs. 1.89 ± 0.02) significantly decreased (P<0.05 or P<0.01), the protein expression of Wnt3a (0.67 ± 0.03, 0.74 ± 0.03, 0.57 ± 0.01 vs. 0.46 ± 0.01), Occludin(0.66 ± 0.02,0.73 ± 0.02,0.67 ± 0.01 vs.0.53 ± 0.01),Claudin-5(0.71 ± 0.01,0.73 ± 0.01,0.66 ± 0.01 vs. 0.64 ± 0.03) significantly increased (P<0.05 or P<0.01) in the TLR4 blocking+cerebral ischemia mice group, the HSYA intervention+cerebral ischemia mice group, HSYA intervention+TLR4 blocking+cerebral ischemia mice group. Conclusions TLR4 plays a critical regulatory role on the activation of Wnt3a and β-catenin in cerebral ischemic mice model. HSYA could intervene on the tight junction of cerebral ischemic brain through the intervention of Wnt3a and β-catenin, thus exerting the protection for cerebral ischemic brain.
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Aim To investigate the change of miR-124 expression in methamphetamine-induced addiction in PC12 cells and the possible regulatory mechanism that it involves.Methods PC12 cells were randomly divided into 6 groups as follows: control group, methamphetamine group, agomir Negative Control group, miR-124 agomir group, agomir Negative Control+methamphetamine group and miR-124 agomir+methamphetamine group.After the treatment, the total RNA and protein were extracted in PC12 cells.The expression of miR-124 was measured by Real-time PCR and the expression of GluR2 was determined by Western blot in PC12 cells.Results Compared with those in the control group, the expression of miR-124 was remarkably decreased and the expression of GluR2 was significantly increased in the methamphetamine group in PC12 cells.Compared with those in the agomir Negative Control+methamphetamine group, the expression of miR-124 was remarkably increased and the expression of GluR2 was significantly decreased in the miR-124 agomir+methamphetamine group in PC12 cells.Conclusion MiR-124 might involve in methamphetamine-induced addiction in PC12 cells by inhibiting GluR2.
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Objective To compare the therapeutical effect of puerarin, ligustrazine, ginsenoside Rb1, Hydroxysafflor yellow A on cerebral ischemia reperfusion mice. Methods The mice were randomly assigned for sham group, model group, puerarin group, ligustrazine group, ginsenoside Rb1 group, and Hydroxysafflor yellow A group, 24 mice for each group. All the groups were subjected to middle cerebral artery occlusion (MCAO) by 1 h ischemia and 24 h of reperfusion except the sham group. The puerarin, ligustrazine, ginsenoside Rb1, Hydroxysafflor yellow A were administrated by tail vein injection with 3μmol/kg at the onset of 1 h of ischemia. The neurologic deficit score, infarct area calculated by TTC staining, cerebral cortex blood flow monitored by laser doppler flowmetry, NO content measured by chemical colorimetry and western blot were applied to determine the expression for cleaved-caspase-3 and nuclear transcription factor NF-κB for each group. Results Compared with the model group, the infarct area (15.83%± 1.83%, 22.00%± 2.53%, 22.83%± 1.83%, 17.83%± 1.72%vs. 34.67%± 2.66%) in the puerarin group, ligustrazine group, ginsenoside Rb1 group, Hydroxysafflor yellow A group was significantly decreased (P<0.01 or P<0.05);the cerebral cortex blood flow (598.81 ± 9.90 μl/kg?min-1, 614.78 ± 9.20 μl/kg?min-1, 577.83 ± 5.55 μl/kg?min-1, 583.54 ± 7.98 μl/kg?min-1 vs. 548.43 ± 1.97 μl/kg?min-1) significantly increased (P<0.01 or P<0.05);the NO content (17.09 ± 1.18μmol/L, 18.54 ± 0.54μmol/L, 18.17 ± 0.49μmol/L, 15.10 ± 0.73μmol/L vs. 20.63 ± 0.73μmol/L) ignificantly decreased (P<0.01 or P<0.05);the expression of cleaved-caspase-3 (1.02 ± 0.08, 1.12 ± 0.04, 0.87 ± 0.08, 1.07 ± 0.08 vs. 1.30 ± 0.06) and NF-κB p-p65/NF-κB p65 (1.03 ± 0.19, 1.15 ± 0.05, 1.12 ± 0.08, 0.72 ± 0.08 vs. 1.45 ± 0.08) ignificantly decreased (P<0.01 or P<0.05) Conclusions Four Chinese herbal monomers could improve nerve and cerebral dysfunctions and ameliorate ischemia symptoms with varying degrees. The mechanisms were involved with the enhancement of cerebral cortex blood flow and inhibition of cell apoptosis and the activation of inflammatory signaling pathways.
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Objective To investigate the effects and probable mechanism ofShenfu injection on the oxidaitve damage of H2O2-induced PC12 cells.Methods PC12 cells were cultured and exposed to 100μmol/L H2O2 for 1 h to establish the oxidative damage model. The protective effect ofShenfu injection was observed by the cell survival rate measured by colorimetric MTT assay, and the leakage rate of lactic dehydrogense (LDH). Western blot methods were used to detect the expression of NF-κB signaling pathway.Results Compared with the model group,Shenfu injection at 5, 10, 20 ml/L could improve the PC12 cells survival rate (83.11% ± 2.59 %, 87.99% ± 0.59%, 85.26% ± 1.07%vs. 73.82% ± 1.82%;P<0.01 orP<0.05), decrease the LDH leakage rate (32.75% ± 4.10%, 28.52% ± 1.14%, 35.79% ± 1.62%vs. 64.34% ± 3.18%;P<0.01 or P<0.05). Western blot results showed thatShenfu injection could protect the PC12 cells from oxidaitve damage by suppressing the p-p65/p65 (1.30 ± 0.10, 1.17 ± 0.06, 1.37 ± 0.15 vs. 1.70 ± 0.10;P<0.01 orP<0.05), p-IκBα/IκBα (1.07 ± 0.12, 1.00 ± 0.10, 1.03 ± 0.06 vs. 1.17 ± 0.06; P<0.01 orP<0.05).ConclusionShenfu injection has a obvious antioxidant effect on PC12 cells in vitro.
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Aim To investigate the effect of indirubin derivative PHⅡ-7 and TRAIL on proliferation in breast cancer cell MCF-7 and its MDR counterpart MCF-7/ADR and the mechanism.Methods Growth inhibition rate was examined respectively by MTT assay under treatment with TRAIL or PHⅡ-7 or in combination. Cell apoptosis and ROS production were examined by flow cytometry.The change of TRAIL receptors(DR4/DR5 )in mRNA was analysed by realtime PCR.Re-sults IC50 of PHⅡ-7 on MCF-7 and MCF-7/ADR was (4.49 ±1.55 ),(3.44 ±0.90 )μmol · L-1 respec-tively;MDA-MB-231 was TRAIL sensitive cell line, and apparently TRAIL induced apoptosis in MDA-MB-23 1 .Low concentration of PHⅡ-7 in combination with TRAIL could augment TRAIL-induced cytotoxic effect including apoptosis while TRAIL or PHⅡ-7 treatment alone had limited cytotoxity to those cells.Besides, PHⅡ-7 at this concentration had little toxicity to hu-man peripheral blood mononuclear cells even if in com-bination with TRAIL.PHⅡ-7 generated ROS produc-tion inside MCF-7 and MCF-7/ADR cells and up-regu-lated DR4/DR5 expression concentration dependently. Once upon ROS scavenger NAC involved,the effect of TRAIL receptors up-regualtion by expression was abro-gated.Conclusions PHⅡ-7 at low concentration could improve the sensitivities of breast cancer cell MCF-7 and MCF-7/ADR to TRAIL,the mechanism of which may be the ability of ROS production by PHⅡ-7 help up-regulated TRAIL receptor DR4,DR5 .Our re-search set a solid foundation for PHⅡ-7 in combination with TRAIL in future clinical application.