Li Kun No.1 Inhibits The Proliferation Of Human Colon Cancer Cell Line HCT-116 And Its Protective Effect On 5-Fu Induced Bone Marrow Suppression Injury In Mice

Objective: To investigate the mechanism of the inhibition of the proliferation of human colon cancer cell line HCT-116 and the protective effect of 5-Fu on bone marrow suppression in mice.Methods:(1)Different concentrations(0,0.25,0.5,1,2,4mg/ml)were used.HCT-116 was detected by human colon cancer cell line.The proliferation of HCT-116 cells was detected by CCK8 method.The apoptosis rate of HCT-116 cells was detected by flow cytometry.The apoptosis-related proteins Bax,Bad and Bcl-were detected by Western blot.2.Akt and caspase3 expression;(2)60 pure male BALB/c mice were randomly divided into normal control group(Control),5-Fu group(Model),and recombinant human granulocyte colony-stimulating factor group(rhG-CSF),li kun No.1 low,medium and high dose group(133,200,300mg/kg).5-Fu(250mg/kg)was injected intraperitoneally to replicate the mouse bone marrow suppression model,and the control group was injected with the same volume of normal saline.On the second day after the injection,different doses of li kun No.1 were administered by gavage,and the normal group and the model group were given.The stomach was given an equal volume of physiological saline,and the positive control group was intraperitoneally injected with rhG-CSF(250?g/kg)for 7 days.The general condition of the mice(hair,diet,activity,diarrhea,etc.)was observed every day.At the end of the experiment,blood was collected from the eyelids,bone marrow smear,and the mice were sacrificed by cervical dislocation.The spleen was dissected and stored in a refrigerator at-80 °C for subsequent experiments.Peripheral blood cells(WBC,PLT,LY,MO%)were counted by automatic blood cell analyzer;serum TNF-? and IL-2 levels were detected by ELISA;JAK2,STAT5,p-JAK2,p-STAT5 were detected by Western blot.p-STAT5 protein expression.Results:(1)Compared with the blank group,2mg.ml-1and 4mg.ml-1 of li kun No.1 significantly inhibited the proliferation of colon cancer cells HCT-116(P<0.05)in a time-and dose-dependent manner.Death(P<0.05),li kun No.1 significantly up-regulated the expression of Bax,Bad,cleaved caspase-3 protein(P<0.05),down-regulated the expression of Bcl-2,Akt,p-Akt protein(P<0.05);(2)Compared with the normal control group,the weight of the mice in the model group decreased significantly(P<0.05),and the symptoms of diarrhea appeared(P<0.05).The results of bone marrow smear showed that the bone marrow hyperplasia of the model group was extremely low,and the periphery The number of white blood cells,platelets,and lymphocytes was significantly decreased(P<0.05),the ratio of monocytes was significantly increased(P<0.05),and the content of TNF-? was significantly increased(P<0.05)IL-2.The content of p-JAK2 and p-STAT5 protein in spleen tissues decreased significantly(P<0.05),and the total protein expression of JAK2 and STAT5 remained unchanged.Compared with the model group,the number of li kun No.1 was low.The weight loss symptoms of the dose group were significantly improved(P<0.05),and the weight of the middle and high dose groups was not Significant improvement(P>0.05),can significantly improve the symptoms of diarrhea caused by 5-Fu,bone marrow smear results show that after giving different doses of li kun No.1,the degree of hyperplasia has improved,the effect is not particularly obvious,blood routine test results It showed that the white blood cells,platelets and lymphocytes of the mice in each dose group were increased in different degrees(P<0.05),the ratio of monocytes decreased(P<0.05),and the mice in each dose group The level of serum TNF-? was significantly decreased(P<0.05),and the level of IL-2 was significantly increased in the middle and high dose groups(P<0.05).The results of Western blot showed that the medium and high doses of li kun No.1 were significantly up-regulated.Expression of p-JAK2 and p-STAT5(P<0.05).Conclusion:(1)Li kun No.1 can inhibit the proliferation of colon cancer cell line HCT-116,possibly by up-regulating Bax,Bad,cleaved caspase-3 protein Bcl-2,Akt,p-Akt protein promotes apoptosis and exerts anti-tumor effect.(2)Li kun No.1 can alleviate the symptoms of diarrhea in mice induced by 5-Fu and alleviate the bone marrow suppression injury caused by 5-Fu.The mechanism may be through up-regulation of spleen cells p-JAK2,p-STAT5.The expression of p-STAT5 protein is reduced by reducing the release of TNF-? and increasing the content of IL-2.