| Colorectal cancer(CRC)is the third most deadly cancer in the world,and patients have developed resistance to chemotherapy drugs such as 5-Fluorouracil(5-Fu),so it is urgent to find new drugs.The first part of this paper aims to study the antitumor activity of the compound 3-O-α-L-arabinopyranosyl-oleanolic acid(3-O-L-AO),which was derived from Pulsatilla cernua(Thunb.)Bercht.ex J.Presl.There are methods and results:CCK-8 assay was used to explore the effect of the compound on the proliferation of CRC cells.It was found that 3-O-L-AO could inhibit the proliferation of LOVO and RKO in a time-dependent and dose-dependent manner;Use Transwell to detect whether the compound affect cells migration.And 3-O-L-AO could significantly inhibit the migration of LOVO;The effects of the compound on the cell cycle and apoptosis of cells were detected by flow cytometry,q RT-PCR and WB.It was found that 3-O-L-AO could significantly block the cell cycle,induce apoptosis,and affect the expression of genes and proteins related to cell cycle and apoptosis;And use DCFH-DA probe to detect the effect of the compound on ROS level.3-O-L-AO could significantly increase ROS level of LOVO and RKO.After that,based on the transcriptome sequencing and the validation of q-RT-PCR and WB,the target gene COL9A3(down-regulated)was screened out.Then,use molecular docking technology to study the interaction mode of the compound and COL9A3.It was found that the interaction between the two was mainly hydrogen bond and hydrophobic force.Finally,We investigate the mechanism of 3-O-L-AO,and we found that 3-O-L-AO influence PI3K-Akt signal pathway,and the phosphorylation of Akt,the key molecule of the pathway,was enhanced after administration.Therefore,The conclusion is as follows: 3-O-L-AO,a small molecular compound derived from P.cernua,can influence the PI3K-Akt signal pathway by down-regulating the expression of COL9A3,and can up-regulating the ROS level of LOVO and RKO,inducing cell cycle arrest and apoptosis,and inhibiting the proliferation and migration of CRC cells.It has the potential to develop a drug for CRC.The traditional Chinese medicinal plant Tripterygium Wilfordii Hook.f.has many clinical effects,but its growth is greatly affected by the environment.The endophytic fungus of plants have the characteristics of widespread distribution,rapid propagation and producing similar active substances with the host,which can make up for the plants,and have attracted the attention of researchers.In this study,we isolate 40 strains of endophytic fungus from Tripterygium Wilfordii from Fujian and Zhejiang.The second part of the article aims to screen the endophytic fungus of Tripterygium Wilfordii with abundant metabolites and antibacterial activity,and then,study the metabolites of the target strains.The methods and results are as follows: use HPLC and TLC to chemically screen the fermentation extracts,the target strain Penicillium sp.LGT-7 with rich and characteristic metabolites was determined.And then,we screen the fermentation medium of LGT-7,it was found that LGT-7 had abundant metabolites in PBD and both Staphylococcus aureus and Escherichia coli showed antibacterial activity.Therefore,it was finally determined to use PDB medium for mass fermentation.After fermentation,the secondary metabolites of LGT-7 were separated and purified by silica gel column chromatography,HPLC and other methods,six compounds were finally obtained.Two compounds were identified by NMR:orsellinaldehyde(1)and 2-[(hydroxy)methyl]-3-methyl-2-cyclopentenone(2),which were firstly isolated from Penicillium.Therefore,the study concluded that the metabolites of the endophytic fungi Penicillium sp.LGT-7 was abundant and had good antibacterial activity.At present,wo got two compounds,which were firstly isolated from Penicillium.It enriched the study of Penicillium fungus,and provided new lead compounds and microbial resources for antibacterial drugs. |
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