Fuzheng Guben Granule With Chemotherapy In Tumor-bearing Mice Pharmacodynamics And Proteomics Research

Objective:Fuzheng Guben Granule has been used clinically for many years and has a good clinical curative effect.Its efficacy is cooling blood and detoxifying,benefiting Qi and Yin.It is used for the treatment of tumor patients,after chemotherapy,Qi and Yin deficiency and heat syndrome,clinically often combined with chemotherapy drugs.Based on the preliminary results of the pharmacodynamic experiment of Fuzheng Guben Granule on the immune function enhancement of tumor-bearing mice,in order to further deepen its influence on immune and hematopoietic function,on the one hand,it guides clinical rational drug use,and on the other hand,it guides clinical rational drug use.On the one hand,expand its market to deepen its understanding.Methods:1.After receiving the cells,disinfect the appearance of the cell culture flask according to the instructions.After the cells are buffered in the incubator for 1.5hours,after the cells are basically restored,the whole bottle of cells and the culture solution are divided into steps.The batch was centrifuged and changed.After the cells were completely grown,the passage was started.At the time of passage,the centrifuge speed was set to 1000 rpm for 5 min,and all the cell culture liquid was centrifuged to collect the cells for use.2.Dilute the collected S₁₈₀80 ascites cells 5 times with normal saline,and then take1 mL for intraperitoneal injection of mice to observe the growth of ascites.After the mice are dislocated,they are wiped with alcohol cotton balls.Use a 5mL syringe to extract the third generation of ascites at 1000rmp/5min,centrifuge all the ascites,collect S₁₈₀80 cells with a pipette,take out the used cells,and put the rest into the-80 C?refrigerator for use.The ascites tumor cells and physiological saline were mixed and adjusted to a concentration of 1×10⁷ mL^-1,and 0.2 mL was inoculated subcutaneously in the right temporal part of the mouse,and the tumor growth was observed daily.Visual observation and hand touch to determine whether the modeling was successful,all the modeling was successful,and the mortality rate was 0%.3.On the next day,the tumor-bearing mice were randomly divided into 5 groups:model group,chemotherapy group,chemotherapy combined with Fuzheng Guben?low,medium and high?dose groups.No treatment was performed in the normal group.Model group:equal volume of distilled water by intragastric administration;chemotherapy group:intraperitoneal injection of cisplatin(1mg·kg^-1).Chemotherapy?cisplatin injection?+Fuzheng Guben granules?low,medium,high?dose group:intraperitoneal injection of cisplatin injection(1mg·kg^-1)combined with intragastric administration of Fuzheng Guben granules(5.625,11.25,22.5g·kg^-1).After 14 days of continuous administration,the general condition,body weight change,tumor inhibition rate,thymus and spleen index of S180 tumor-bearing mice were observed.The CD4 and CD8 levels in peripheral blood of tumor-bearing mice were detected by flow cytometry./CD8 balance and T cell subsets in the main T cell subsets of cell levels and their balance were observed.Peripheral blood cells were counted using a peripheral blood cell counter,and the number of bone marrow hematopoietic stem cells was measured by electron microscopy on a blood cell count plate,and bone marrow pathology was performed to observe the hematopoietic function.4.Finally,based on TMT labeling,high performance liquid chromatography grading technology and mass spectrometry-based quantitative proteomics technology,the above model group?M?,cisplatin group?S?,cisplatin combined with high-dose group?Fg?The right femur bone marrow protein was detected,and the liquid chromatography-mass spectrometry analysis was performed according to different peptides.The GO database,the Kyoto Gene Encyclopedia database,and the InterPro database were used to perform protein function and enrichment analysis.Related literatures explored its biological significance,identified specific proteins and related pathways that improved the side effects of cisplatin,and further explored the reversal mechanism of Fuzheng Guben Granules on the side effects of tumor-bearing mice after chemotherapy.Result:1.General situation1.1 Except the blank group,all the tumor-bearing mice in each group were successful in modeling and showed different degrees of activity limitation.The density and gloss of the model group did not change significantly with the blank group.The tumor volume of the cisplatin group was significantly reduced.The activity was restricted compared with the model group,but the coat color was dull,dull and the skin was sparse.The combined drug group had bright coat color,firm fur,and the activity was better than the model group.No death occurred during the experiment.2.Main pharmacodynamics2.1 Effects on body weight,tumor weight and tumor inhibition rateThe weight of Fuzheng Guben Granules combined with the chemotherapy group was significantly increased?P<0.01?,and there was significant difference between the model group and the cisplatin group and Fuzheng Guben particles?low,medium?group.Significance?P<0.05 or P<0.01?,there was no significant difference in the effect of high dose on tumor weight.2.2 Effect on immune function of tumor-bearing miceCompared alone with the chemotherapy group,the chest?spleen?visceral index of the chemotherapy group combined with Fuzheng Guben granules?medium and high?was significantly increased?P<0.05 or P<0.01?.The level of CD8 in the blood of Fuzheng Guben Granule?medium and high?group combined with chemotherapy was significantly decreased,the difference was statistically significant?P<0.05 or P<0.01?.Fuzheng Guben Granule?medium and high?dose combined with chemotherapy The level of CD4 in the blood of the mice was significantly increased,and the difference was statistically significant?P<0.01?.The combination of Fuzheng Guben Granules?medium and high?combined with chemotherapy can significantly increase the content of Th1 and Th17 in the blood of mice?P<0.05 or P<0.01?.The content of Th17 and Treg in the blood of Fuzheng Guben Granule combined with high dose combined with chemotherapy Significantly decreased?P<0.05?,the combination of Fuzheng Guben granules in combination with chemotherapy can significantly increase the level of Th1/Th2 in the blood of mice?P<0.05 or P<0.01?,and the high dose of Fuzheng Guben granule combined with chemotherapy Th17/Treg levels in the blood of rats increased significantly?P<0.01?.2.3 Effect on hematopoietic function of tumor-bearing miceCompared with the model group,the number of red blood cells in the peripheral blood of the mice in the chemotherapy group was significantly decreased?P<0.05?.Compared with the chemotherapy group,the Fuzheng Guben high-dose group combined with the blood in the peripheral blood of the mice,red,white blood cells,and platelets.The number increased in different degrees?P<0.01?.Compared with the model group,the number of hematopoietic stem cells in the peripheral blood of the mice in the chemotherapy group was significantly decreased?P<0.01?.Compared with the chemotherapy group,the number of hematopoietic stem cells in the peripheral blood of Fuzheng Guben high-dose combined with chemotherapy was significantly increased.?P<0.01?.Compared with the model group,the hematopoietic tissue volume in the bone marrow cavity of the chemotherapy group was significantly decreased?P<0.01?and the fat volume was significantly increased?P<0.01?.Compared with the chemotherapy group,the Fuzheng Guben granules?medium,high?dose group was combined.The hematopoietic tissue volume in the medullary cavity of the chemotherapy mice was significantly increased?P<0.01?;the adipose tissue volume was significantly decreased?P<0.01?.3.Proteomics study of Fuzheng Guben Granules combined with cisplatin on mouse femur bone marrow.A total of 331667 proteins were detected,of which 5954 were qualitative and5466 were quantitative.There were 383 differential proteins in the model group?M?and cisplatin group?S?,and the Fuzheng Guben group?FG?and cisplatin.Group?S?has a total of 104 differential proteins.Bioinformatics analysis showed that the cisplatin group had significant functional differences in the regulation of myofibrillar contraction and composition of chromosomes by atp6V0c,atp6v1g1,atp2a1 and other proteins,and the high-dose combination of Fuzheng Guben.There was a significant functional difference between the platinum group and the cisplatin group in the combination of the mRNA signal sequence complex and the SRP signaling factor,and the induction of apoptotic DNA break repair.The results of mass spectrometry showed that Fuzheng Guben granules can regulate the immune,hematopoietic function by regulating the expression of poll,Timeless,Rps27a,chd2,Hist1h1a,Hist1h1d and other proteins involved in DNA double-strand broken chain repair to reduce the side effects of DNA hydration using cisplatin damage.Conclusion:1.Fuzheng Guben Granules effectively improve the general state of tumor-bearing mice.2.Fuzheng Guben granules?medium,high?dose combined with cisplatin can increase the thymus?spleen?index,CD4+in tumor-bearing mice,and reduce the level of CD8+,which is beneficial to the proliferation of T cell subsets in animals,including Fuzheng Guben Granules.High dose combined with cisplatin group can significantly increase the ratio of Th1/Th2 and Th17/Treg to improve the overall T cell immune function.3.Fuzheng Guben high-dose group combined with chemotherapy cisplatin,the experimental results suggest that Fuzheng Guben granules can significantly reverse the bone marrow suppression caused by cisplatin,increase the number of peripheral blood cells and bone marrow hematopoietic stem cells,and compare the hematopoietic tissue in bone marrow.With the capacity of adipose tissue and pathological section of mouse bone marrow,it was found that the high dose group of Fuzheng Guben Granule combined with cisplatin had the best effect on bone marrow repair.4.The results of proteomic experiments show that Fuzheng Guben Granule can repair the side effects caused by cisplatin by regulating DNA repair protein?poll?,and reduce the regulation of cisplatin on myosin complex by regulating atp6V0c,atp6v1g1,atp2a1 and other proteins.The role of myofibril contraction.