Experimental Study On Lentinan Treating Mice Breast Cancer By Regulating IL-35 And ITr35 Subgroup

Objective:Replicate the tumor-bearing mouse model by inoculating 4T1 breast cancer cells,and observe the changes of IL-35 molecular expression and the relative proportion of iTr35 subgroup in experimental animals.After breast mice lentinan(Lentinan,LNT)drug therapy,breast cancer mice were tested for IL-35 and iTr35subgroups in tumor tissues,peripheral blood and some peripheral lymphoid organs(spleen and lymph nodes),and to explore the therapeutic effect of lentinan on breast cancer and its possible effect mechanism.Methods:A breast cancer 4T1 cell line was subcutaneously inoculated into Balb/c mice to replicate a breast cancer xenograft model in vivo.The test animals were divided into normal control group(NCG),tumor-bearing mice were randomly divided into tumor-bearing control group(TCG),Lentinan treatment group-Low(LTG-L,100mg/kg),Lentinan treatment group-Medium(LTG-M,200 mg/kg)and Lentinan treatment group-High(LTG-H,400 mg/kg),10 rats in each group.The corresponding amount of lentinan was administered to the lentinan administration group,and the NCG and the TCG were given the same amount of normal saline for 21 days,once a day.After the treatment,the test mice were sacrificed by eyeball removal,and peripheral blood and dissected tumor tissue,spleen and lymph tissue were retained.ELISA method was used to detect the levels of IFN-γ,IL-4 and IL-35 in serum;RT-PCR method was used to detect the relative expression level of IL-35 mRNA in tumor and spleen tissue;Western Blot was used to detect the expression of IL-35protein in spleen tissue;The relative proportion of iTr35 subpopulations in peripheral blood,spleen,and lymph node tissue was detected by cytometry.Results:1.The general condition of tumor-bearing mice became worse,the activity was weakened,and the amount of dietary water was reduced.By the fourth week,the weight growth rate was relatively slowed down,and the difference was statistically significant(P<0.01).Histopathological staining of the tumor tissue showed that the nuclei were enlarged,hyperchromatic,of different sizes and disordered arrangement,with heteromorphism,pathological mitosis and neoplastic necrosis.2.The expression of IL-35 in peripheral blood and peripheral lymphoid organs(spleen and lymph nodes)of tumor-bearing mice increased,and the difference was statistically significant(P<0.01 or P<0.001).3.The proportion of CD4~+T cells in peripheral blood,spleen and lymph nodes of tumor-bearing mice decreased,and the relative proportion of iTr35 subpopulation in peripheral blood and spleen also decreased significantly,the difference was statistically significant(P<0.01 or P<0.001),while the relative proportion of iTr35subgroups in lymph nodes increased,and the difference was statistically significant(P<0.01 or P<0.001).4.After the tumor-bearing mice were treated with lentinan,the experimental animals showed that the general condition improved,tumor volume decreased,and the weight was reduced.The effect of the LTG-M was the best,and the difference was statistically significant(P<0.01).5.Compared with the TCG,the expression of IL-35 in peripheral blood of experimental animals in the lentinan administration group decreased,while the level of IFN-γincreased.The LTG-M was the most significant,and the difference was statistically significant(P<0.01);IL-4 expression in the peripheral blood of mice in the drug treatment group also decreased,but the difference was not statistically significant(P>0.05).6.Compared with the TCG,the expressions of p35 and EBI3 in tumor tissue and spleen tissue of the mice treated with lentinan were decreased,especially in the LTG-M,the difference was statistically significant(P<0.05).7.Compared with the TCG,the relative proportion of iTr35 subpopulations in the lymph nodes of the lentinan-administered group decreased,and the effect of the LTG-M was significant,and the difference was statistically significant(P<0.05).Conclusion:1.Subcutaneous injection of 4T1 breast cancer cell line successfully replicated a mouse breast cancer xenograft model.2.The expression levels of IL-35 in peripheral blood,spleen and lymph node tissues of 4T1-cell breast cancer mice increased,and the relative proportion of i Tr35subpopulations in lymph nodes increased.It is speculated that the development of breast cancer is closely related to the suppression of immune function.3.Under the experimental conditions,lentinan has obvious therapeutic effect on 4T1cell transplantation breast cancer,and the best effect is at a medium dose(200 mg/kg).The mechanism may be by reducing tumor tissue and some peripheral lymphoid organs(spleen and lymph nodes).IL-35 expression increases IFN-γsecretion and reduces iTr35 subpopulation formation in lymph nodes.Thus,to a certain extent,the body’s immune function is enhanced,especially cellular immunity is closely related.