灵芝康爱芝宝增强环磷酰胺抑制小鼠乳腺癌肺转移的增效减毒作用

Effects of GanoExtra combined with CTX on lung metastasis and immune function in mice

目的探讨灵芝康爱芝宝(GanoExtra)与环磷酰胺(cyclophosphamide,CTX)联用对抑制小鼠乳腺癌肺转移的增效减毒作用以及对免疫功能的影响。方法通过CCK-8实验验证GanoExtra对肿瘤细胞MCF-7和4T1的毒性;尾静脉注射4T1细胞建立小鼠乳腺癌肺转移模型,设立4T1模型组、CTX组、GanoExtra组和GanoExtra+CTX组,另设对照组。21 d后麻醉处死小鼠,记录小鼠体重、肺转移灶数;肺和脾脏系数;测定小鼠脾脏细胞的活性和NK细胞的活性;测定小鼠血样中各细胞数量;流式细胞术测定血样中CD4^(+)和CD8^(+)T细胞的含量;ELISA法检测血清中TNF-α和IL-6含量;HE染色法观察肿瘤及各组织病理形态学变化;CFSE染色测定GanoExtra对CD8^(+)T细胞的增殖作用。结果在体外,GanoExtra可明显抑制MCF-7和4T1肿瘤细胞活性;在乳腺癌肺转移模型中,与模型组相比,GanoExtra组小鼠脾脏系数和血液WBCs含量明显降低,RBCs、NK细胞、脾细胞活性、CD3^(+)和CD8^(+)细胞比例明显升高;治疗结束后,与CTX组相比较,GanoExtra+CTX组肺转移灶数和肺脏系数明显降低,小鼠血液HGB、CD8^(+)细胞、IL-6和TNF-α明显升高。在体外10 mg·L^(-1)的GanoExtra可明显促进CD8^(+)T细胞增殖。结论GanoExtra可以提高CTX对肿瘤转移的抑制作用,减少由于CTX导致的不良反应,具有促进CD8^(+)T细胞增殖以增强机体免疫疗效的保健功能。

Aim To investigate the enhanced efficacy and reduced toxicity of GanoExtra in combination with cyclophosphamide(CTX)on inhibiting lung metastasis and immune function in mice.Methods The CCK-8 method was used to verify the cytotoxic effects of GanoExtra on MCF-7 and 4T1 tumor cells.In vivo experiment,a mouse model of lung metastasis of breast cancer was established by injecting 4T1 tumor cells into the tail vein,which was divided into four groups including 4T1 model group,CTX group,GanoExtra group and GanoExtra+CTX group.The control group was set.After 21 days,the mice were euthanized under anesthesia,and the body weight of the mice was recorded.Average lung index and spleen index were calculated.The mouse spleen lymphocyte transformation experiment was used to determine the activity of spleen cells.The NK cell activity assay was used to determine the activity of NK cells.Blood cells were determined in mouse blood samples.Flow cytometry was used to determine the levels of CD4^(+)and CD8^(+)T cells in blood samples.ELISA was used to measure the levels of TNF-αand IL-6 in serum.HE staining was used to observe the pathological morphological changes in tumors and various tissues;and CFSE staining was used to determine the proliferative effect of GanoExtra on CD8^(+)cells.Results In vitro GanoExtra at 50 mg·L^(-1)significantly inhibited the activity of MCF-7 and 4T1 tumor cells.In the breast cancer pulmonary metastasis model,compared with the model group,the spleen index and blood WBCs content were significantly reduced,while the activity of NK cells,spleen cells,and the proportion of RBCs,CD3^(+)and CD8^(+)T cells in the blood were significantly increased.At the end of the treatment,compared with the CTX group,the number of lung metastases and lung index of the GanoExtra+CTX group were significantly reduced,and the levels of HGB,CD8^(+)cells,IL-6,and TNF-αin the blood of mice were significantly increased.GanoExtra at 10 mg·L^(-1)significantly promoted the proliferation of CD8^(+)T cells in vitro.Conclusions GanoExtra can enhance the inhibitory effect of CTX on tumor metastasis,alleviate adverse reactions such as splenomegaly and pulmonary enlargement caused by CTX,and have a health-promoting function of promoting the proliferation of CD8^(+)T cells to enhance the immune efficacy of the body.