The Treatment Of Blood Stasis Lung Cancer By JAK1/STAT3 Pathway Was Discussed Based On Blood Circulation And Stasis

Objective To sort out the development of lung cancer syndrome of blood stasis and the method of promoting blood stasis,and to study the method of promoting blood stasis and its representative drugs for blood stasis.To demonstrate the scientific nature of Diterpenoid Tanshinone(DT)in the treatment of lung cancer under the guidance of activating blood circulation and removing blood stasis,and to study the related mechanism of Diterpenoid tanshinone in the treatment of lung cancer through JAK1/STAT3 pathway regulation of apoptosis and angiogenesis.Methods1.Theoretical research:through searching,reading and analyzing ancient and modern literature,combing the development context of lung cancer blood stasis,summarizing and summarizing the pathogenesis and characteristics of lung cancer blood stasis,and analyzing the formation mechanism of lung cancer blood stasis from the perspective of modern medicine;This paper reviews the development process of blood circulation and stasis removal method,lists the representative drugs of blood circulation and stasis,and expounds the scientific evidence of blood stasis in the treatment of lung cancer.2.Experimental studies:1)The results of CCK8 showed that different concentrations of DT could inhibit the activity of lung cancer A549 cells,and the IC50 value was 7.153μg/ml.2)The scratch test results showed that the cell number of 2.5,4.5,6.5μg/mlDT scratches at 12h,24h and 48h was significantly different from that of the blank control group(*P<0.05;***P<0.01;***P<0.001);Transwell migration experiment showed that compared with 0μg/ml DT group,the cell migration number of A549 cells in different concentration DT groups decreased(*P<0.05,**P<0.01,***P<0.001),and the higher the concentration,the less the cell migration number.The number of cell migration decreased significantly in 6.5μg/mlDT group,and the difference was statistically significant(***P<0.001).3)Transwell invasion assay showed that DT inhibited the invasion ability of A549 cells.Compared with 0μg/mlDT group,the invasion number of A549 cells in other DT administration groups was significantly decreased(***P<0.01).In the 6.5μg/mlDT group,the number of cell invasion decreased significantly,and the number of cell invasion was(3473±227.7)per well.4)WesternBlot showed that DT down-regulated JAK1,STAT3,VEGFA and Bcl2 proteins and up-regulated the expression of Bax protein.5)DT had no serious effect on the body weight,liver and kidney weight,tumor weight and tumor volume ratio of nude mice.Low dose of DT reduced tumor weight(**P<0.01),and inhibited tumor volume significantly(***P<0.001).6)Immunohistochemical results showed that the positive expression of VEGFA in cisplatin group,combined group,DT high-dose,mediumdose and low-dose groups was significantly decreased compared with model group(*P<0.05).7)Immunofluorescence results showed that the number of CD31 fluorescence in DT administration.group was significantly reduced compared with model group.8)Elisa results showed that DT could reduce the content of VEGFA and IL-6 in tumor tissues,and the difference was statistically significant(***P<0.001).9)WesternBlot showed that DT could inhibit JAK1,STAT3,VEGFA and Bcl2 proteins and promote the expression of Bax protein in tumor tissues.10)GO functional enrichment analysis results showed that compared with control,It was mainly enriched in cell population proliferation and leukocyte adhesion to vascular endothelial cells cell),the epithelium migration,muscle cell apoptotic process,chronic inflammatory response,etc.Compared with the control group,LH mainly concentrated in positive regulation of epidermal growth factor-activated receptor activity activity,macrophage apoptotic process,blood vessel endothelial cell differentiation differentiation,positive regulation of vasculogenesis,inflammatory cell apoptotic process,etc.;Compared with the control group,SB mainly enriched in the regulation of angiogenesis and the regulation of endothelial cell proliferation proliferation),angiogenesis involved in wound healing,extrinsic apoptotic signaling pathway),vascular wound healing,tumor necrosis factor production,interleukin-1 receptor binding binding),signaling receptor activator activity,etc.11)KEGG enrichment analysis showed that 74 KEGG signaling pathways were enriched in DTM compared with control group(P<0.05).It mainly consisted of Leukocyte transendothelial migration,Ras signaling pathway,and PI3K-Akt signaling pathway pathway),NF-kappa B signaling pathway,etc.LH enriched 185 KEGG signaling pathways compared with control group,It mainly contains JAK-STAT3 signaling pathway,VEGF signaling pathway,Apoptosis and Non-small cell lung cancer lung cancer),etc.The differential genes of SB and control group were enriched in 185 KEGG signaling pathways.It mainly includes JAK-STAT3 signaling pathway,Apoptosis and Leukocyte transendothelial migration migration,and Platelet activation.Results1.Theoretical research:Blood stasis is the main pathogenic factor for the formation and development of blood stasis syndrome of lung cancer,mainly Qi deficiency,causing blood stasis;Blood stasis as a sign,because of stasis accumulated poison;Due to toxic changes,lung cancer is the key pathogenesis of blood stasis syndrome of lung cancer.The method of promoting blood circulation and removing blood stasis is an important treatment of lung cancer and runs through the whole process of the treatment of blood stasis syndrome of lung cancer.Diterpenoid quinone of Salvia miltiorrhiza is the effective part of the representative medicine Salvia Miltiorrhiza to promote blood circulation and remove blood stasis,which can effectively treat the blood stasis syndrome of lung cancer.2.Experimental study:1)CCK8 results showed that different concentrations of DT could inhibit the activity of lung cancer A549 cells,and the measured IC50 value was 7.153μg/ml;2)The results of scratch experiments showed that the number of cells scratched by DT at 2.5,4.5 and 6.5μg/ml after 12h,24h and 48h had significant differences compared with the blank control group(*P<0.05;***P<0.01;***P<0.001);Transwell migration experiments showed that compared with the Oμg/ml DT group,the number of cell migration after intervention of A549 cells in different concentrations of DT group was reduced(*P<0.05.**P<0.01,***P<0.001),the higher the concentration,the less cell migration,among which the 6.5μg/ml DT group cell migration number decreased significantly,the difference had obvious statistical differences(***P<0.001);D T can inhibit lung cancer A549 cell migration ability 3)Transwell invasion experiments showed that DT can inhibit A549 cell invasion ability,compared with 0μg/ml DT group,the number of A549 cell invasion in the rest of the DT administration group decreased significantly(***P<0.01),among which the number of cell invasions in the 6.5μg/ml DT group decreased significantly,and the number of cell invasions was(3473±227.7)pcs/well;4)Western Blot experiments showed that DT can downregulate JAK1,STAT3,VEGFA,Bcl2 proteins and upregulate the expression of Bax proteins;5)The weight,liver and kidney weight,tumor weight and tumor volume ratio of nude mice showed that DT did not have a serious impact on the liver and kidney of nude mice.The low dose of DT can now reduce tumor weight(**P<0.01)and inhibit tumor volume(****P<0.001);6)the immunohistochemical results showed that the positive expression of VEGFA in the cisplatin group,the combined group,the DT high,medium and low dose groups and the model group were significantly reduced(*P<0.05);7)The immunofluorescence results showed that the number of CD31 fluorescence in the DT administration group was significantly reduced compared with the model group.8)The results of Elisa experimental results showed that DT could reduce the content of VEGFA and IL-6 in tumor tissues,and the difference was statistically significant(***P<0.001);9)Western Blot results showed that DT could inhibit JAK1,STAT3,VEGFA,Bcl2 proteins in tumor tissues and promote the expression of Bax protein.Conclusion1.Deficiency of origin and deficiency of substance are the general pathogenesis of blood stasis syndrome of lung cancer.Lung qi deficiency is the foundation and blood stasis is the standard,and blood stasis is the key factor leading to blood stasis syndrome of lung cancer.According to the key factors of the occurrence of blood stasis syndrome of lung cancer,it is scientific to use the method of promoting blood stasis to prevent and treat the blood stasis syndrome of lung cancer.The clinical application of drugs of promoting blood stasis to treat the blood stasis syndrome of lung cancer does have a certain effect,which provides a theoretical basis for the study of the mechanism of action of DT in the treatment of lung cancer.2.DT can inhibit the activity,proliferation,migration and invasion of lung cancer A549 cells,and it has been verified by in vivo experiments that DT can inhibit the growth of transplanted tumor in nude mice,and Elisa detection of DT can reduce the content of VEGFA and IL-6 in transplanted tumor in nude mice.Immunohistochemistry and immunofluorescence showed that DT could inhibit the expression of VEGFA and CD31 and reduce the microvascular density in transplanted mice.DT detection by Western Blot downregulates JAK1,STAT3,VEGFA,Bcl2 and Bax protein expression.3.DT interferes with gene expression changes in tumor tissue after tumor transplantation in nude mice,and enrichment analysis of transcriptome differential genes GO and KEGG pathway shows that it is closely related to vascular endothelial adhesion factors,platelet activating factors,chronic inflammation,apoptosis,leukocyte adhesion to vascular endothelial cells,leukocyte activation and other processes.DT may promote apoptosis and inhibit angiogenesis by regulating JAK1/STAT3 pathway to treat lung cancer.