ABSTRACT
Objective:To analyze the material basis, target and pathway of Chebulae fructus and Margarita in Mongolian medicine Garidi-13 Pill on stroke with the method of network pharmacology and molecular docking technology, so as to better understand its "detoxification" mechanism. Methods:TCMIP and BATMAN-TCM were used to predict the target of Chebulae fructus and Margarita composition, and GeneCards were used to search for the target of stroke. The overlapping targets of the two platforms were imported into the Metascape software for GO biological analysis and KEGG pathway analysis. The molecular docking of key molecules and targets was carried out with LEDOCK. Results:A total of 22 active components were collected and 217 targets related to stroke were predicted. Among them, 1-O-galloyl-glucose, cuprum, ellagicacid, arjungenin and corilagin were the key substances playing the role of "detoxification" of the Chebulae fructus and Margarita; IL6, TNF, HSP90AA1, PTGS2, CASP3, NR1I2, VKORC1 and ATP1A1 were the key targets playing the role of "detoxification" . These targets were significantly enriched in cell response, humoral level regulation, hemostasis, response to steroid hormones, steroid metabolism, coagulation and other biological processes, as well as nitrogen metabolism, IL-17 signaling pathway and other pathways. Molecular docking verified the accuracy of previous prediction results from computer simulation level. Conclusion:The process of Chebulae fructus and Margarita intervening strok is closely related to the elimination of harmful metabolites and calmingthe inflammatory reaction, which was not only consistent with the modern medicine on the pathological process of stroke, but also consistent with the interpretation of "evil and poison" with Mongolian medicine theory.
ABSTRACT
Objective:To study the role of 14-3-3εand Cdc25B in germinal vesicle (GV)-stage arrest of mouse oocytes,and to pay foundation for further study on the molecular mechanism of PKA/Cdc25B/14-3-3εpathway in GV-stage arrest of mouse oocytes.Methods:The eukaryotic expression vectors of pcDNA3.1-ZEO-HA-14-3-3ε, pcDNA3.1-MYC-Cdc25B-WT, pcDNA3.1-MYC-Cdc25B-S321A, and pcDNA3.1-MYC-Cdc25B-S321D were transcribed into mRNA invitro.The mouse GV-stage oocytes were collected after superovulation and divided into no injection group,TE buffer microinjection group,14-3-3εmRNA injection group,14-3-3εmRNAs + Cdc25B-WT mRNA injection group,and 14-3-3εmRNA + Cdc25B-S321A mRNA injection group,14-3-3εmRNA+Cdc25B-S321D mRNA injection group.The protein expression levels of HA-14-3-3εand MYC-Cdc25B and the phosphorylation status of Cdc2-pTyr15 were observed by Western blotting method.The morphological changes and germinal vesicle breakdown (GVDB)rates of mouse oocytes were observed under phase-contrast microscope. Results:None of the oocytes in no injection group, TE buffer microinjection group, 14-3-3εmRNA injection group,14-3-3εmRNA + Cdc25B-WT mRNAs injection group and 14-3-3εmRNA + Cdc25B-S321D mRNA were able to undergo GVBD until at least 20 h after injection (P>0.05 );the GVBD rates of oocytes in 14-3-3εmRNA+Cdc25B-S321A mRNA group at 1 h (5.00%±0.68%),2 h (62.00%±3.56%)and 3 h (100.00%± 0.00%)after injection were significantly higher than those in no injection group and TE buffer injection group (P<0.01);the oocytes in 14-3-3εmRNA+ Cdc25B-Ser321A mRNA group at 20 h (79.00%±2.80%)after injection progressed to MII (P<0.01).Conclusion:14-3-3εcan regulate the transition from GV to GVBD of mouse oocytes by means of phosphorylation and dephosphorylation of S321-Cdc25B.