| Mori Fuctus,the mature fruit of Morus alba L.,has a sweet and warm nature,has a sweet and sour taste,and has the effects of nourishing yin and blood,nourishing liver and kidney,and clearing the liver.Mori Fuctus was listed in Chinese Pharmacopoeia 2015 eidition and the Catalogue of Medicinal Food Raw Materials(2019).Mori Fuctus polysaccharide(MFP)was an important active substance extracted from it.It has various physiological activities such as anti-tumor,anti-oxidation,anti-aging,anti-viral,anti-bacterial,blood sugar lowering,blood lipid lowering and immune regulation.Because of the wide biological activity of polysaccharide components in Mori Fuctus and the huge yield of Mori Fuctus,it has attracted the attention of scientific researchers and is one of the main goals of new drug development.The results of the previous research showed that the MFPs and their derivatives have biological activities such as anti-oxidation,anti-fatigue,and in vitro anti-alcohol dehydrogenase.It is a potential good liver protection natural drug.Previous studies have not explored the protective mechanism of MFPs and their derivatives,which has affected the further development and utilization of MFPs.The changes of cytokines in Kupffer cells were studied by ELISA,and the effects of the MFPs and their derivatives on the immune function of Kupffer cells activated by LPS were obtained.Thus,it could be determined whether MFPs could protect liver through Kupffer cells and lay a foundation for further study on the mechanism of MFPs.In addition,it was found in previous studies that MFPs have certain anticancer activity.Therefore,the author has selected eight kinds of cancer cells that are harmful to human body and higher prevalence rate.MTT method was used to study the inhibitory activity of MFPs and their derivatives on these cancer cells.The dose-effect relationship and time-dependent relationship of MFPs on the target cells were explored.Cell scratch test was used to investigate the inhibitory effect of MFPs on the cell migration of target cells.The experimental results are as follows:1.The extraction method of Kupffer cells was optimized.The length of the perfusion channel was adjusted to 0.9 m,the fully torn liver was digested for 15 minutes,and the extracted cells were purified by Percoll differential centrifugation,which greatly improved the number and the phagocytosis activity of cells.2.Three kinds of concentrations of MFP-30-1 decreased the MDA capacity in the order of 1.0 > 0.5 > 2.0 mg·m L-1.Considering the effect of polysaccharide on the immune activity of Kupffer cells,0.5 mg·m L-1 as the concentrations of MFP for further experiments.In the preliminary screening test,two components of positive drugs(PDTC,DDB)and ten kinds of MFPs can inhibit the increase of MDA levels.3.In the formal test,Kupffer cells were treated with twelve kinds of drugs,the contents of MDA,NO,IL-6,IL-1? and NF-?B and the activity values of TNF-?,i NOS of the the supernatant of Kupffer cells were measured.After treatment of Kupffer cells activated by LPS for 24 hours,the contents of MDA,NO,IL-6,IL-1? and NF-?B,as well as the activity values of TNF-? and i NOS were significantly decreased,which indicated that all the drugs in the 12 groups could inhibit the immune function of Kupffer cells activated by LPS.MFP-70-1 can significantly reduce the increase of MDA,IL-6 and TNF-? caused by LPS(P < 0.01),and has good biological activity.According to the results,MFPs and their derivatives can regulate the immunoregulation of Kupffer cells by regulating the expression of NF-?B signal pathway and down regulating the content of related cytokines.For the seven indicators,the order of decreasing effect of the two positive drugs,ten MFPs and their derivatives on these seven indicators is not the same,which may indicate that 12 groups of drugs regulate Kupffer cell immunity is a complex process,in addition to the NF-?B signal pathway involved in this process,there are other signal pathways involved in this process.4.MFPs and their derivatives have good inhibitory activities on seven kinds of selected cancer cells.The results of MTT showed that MFPs had a certain inhibitory effect on a variety of selected cells(P < 0.05),and MFPs with more than 60% inhibitory activity was selected for follow-up study.Among them,at the concentration of 2 mg·m L-1,the inhibitory activity of MFPs on He La cells was not high,and the inhibition rate was less than 60%.S-MFP-30,MFP-50-2 and MFP-70-2 had excellent inhibitory effects on MKN-45 cells,with inhibition rates of 91.35 ± 0.49%,71.8 ± 0.37% and 72.44 ± 0.44%(P < 0.01),respectively;four components of MFPs showed good inhibitory effects on NCI-H1650 cells,with inhibition rates of S-MFP-30(95.06 ± 0.43%),MFP-50-1(63.69 ± 0.32%),MFP-50-2(73.35 ± 0.49%)and MFP-70-2(82.03 ± 0.32%)(P < 0.01);MFP-30-2,S-MFP-30,MFP-50-2 and MFP-70-2 showed a strong ability to inhibit the proliferation of MCF-7 cells,the inhibition rates were 67.9 ± 0.28%,99.95 ± 0.03%,79.27 ± 0.45% and 74.05 ± 0.44%(P < 0.01),respectively;Six kinds of polysaccharides showed a good ability to inhibit the proliferation of OVCAR-3 cells.The six kinds of polysaccharides and their inhibition rates were MFP-30-2(67.36 ± 1.50%),S-MFP-30(95.78 ± 0.64%),MFP-50-1(70.68 ± 1.21%),MFP-50-2(77.27 ± 0.59%),MFP-70-2(78.76 ± 0.72%)and MFP-90-2(73.91 ± 0.80%),respectively,with significant differences(P < 0.01);among the ten polysaccharides,only S-MFP-30 showed the highest inhibition rate on Hep G-2 cells,with the inhibition rate of 90.32 ± 1.23%(P < 0.01);two kinds of MFPs showed good inhibition on A549 cells,with the inhibition rates of MFP-50-2(78.45 ± 1.33%),MFP-70-2(68.81 ± 1.33%),respectively;finally,the inhibitory effect of the two components of polysaccharide on NCI-H292 cell proliferation was the most obvious,the inhibition rates were: MFP-50-2: 73.91 ± 1.14%,MFP-70-2: 69.73 ± 1.82%(P < 0.01).MTT results showed that the inhibition of MFPs and its derivatives on the target cancer cells showed a significant dose-response,time-effect relationship.With the increase of polysaccharide concentration and time,the inhibition rate of MFPs on the target cells increased significantly.The results of cell scratch test showed that MFPs can significantly inhibit the migration of cancer cells,up to 68.89%.To sum up,this study confirmed that MFPs and their derivatives play the therapeutic role of anti-chemical liver injury by regulating the immune activity of Kupffer cells,providing important scientific basis and data support for it as a potential drug for the treatment of chemical liver injury.What's more,the anti-tumor activity of MFPs and their derivatives were preliminarily screened,and the anti-tumor activity site of MFPs were confirmed,which provided scientific basis and data support for the further development and utilization of anti-tumor activity of MFPs. |
PDF Full Text Request