益肺逐积方联合环磷酰胺对Lewis肺癌小鼠模型肿瘤体积及免疫功能的影响

Yifeizhuji Decoction Combined with Cyclophosphamide for Reducing the Tumor Volume and Improving the Immunologic Functions in the Lewis Lung Carcinoma Mouse Model

目的观察益肺逐积方联合环磷酰胺(CTX)对Lewis肺癌小鼠模型肿瘤体积、胸腺指数及淋巴细胞百分比的影响,以探讨益肺逐积方对肺癌的治疗及免疫调节作用。方法 2016年4—5月,选取SPF级C57BL/6雄性小鼠55只,其中5只接种小鼠Lewis肺癌细胞制备Lewis瘤细胞悬液,另50只采用随机数字表法分为模型小鼠(40只)和正常小鼠作为正常组(10只),模型小鼠右前肢腋窝皮下接种Lewis瘤细胞悬液,建立Lewis肺癌小鼠模型。造模成功后模型小鼠采用随机数字表法分为益肺逐积方组(10只)、CTX组(10只)、联合用药组(10只)、模型对照组(10只)。益肺逐积方组中药液灌胃14.95 g/kg,CTX组在接种后72 h和10 d腹腔注射CTX 40 mg/kg,联合用药组在CTX组用药基础上中药液灌胃14.95 g/kg,模型对照组和正常组给予与中药液等体积的0.9%氯化钠溶液灌胃,各组均于造模第2天起开始用药,皆1次/d,连续14 d。用药14 d后测量各组小鼠瘤体直径,计算肿瘤体积;称量小鼠体质量、胸腺质量、计算胸腺指数;流式细胞术检测脾细胞CD_4^+T、CD_8^+T、B淋巴细胞百分比。结果益肺逐积方组、联合用药组小鼠肿瘤体积小于模型对照组(P<0.05),联合用药组小鼠肿瘤体积小于CTX组(P<0.05);模型对照组、益肺逐积方组、CTX组及联合用药组体质量小于正常组(P<0.05),益肺逐积方组胸腺质量及胸腺指数大于正常组及CTX组(P<0.05);模型对照组小鼠CD_4^+T、CD_8^+T淋巴细胞百分比小于正常组(P<0.05),益肺逐积方组、CTX组、联合用药组小鼠CD_4^+T、CD_8^+T淋巴细胞百分比大于模型对照组(P<0.05),联合用药组CD_4^+T、CD_8^+T细胞百分比大于CTX组(P<0.05);模型对照组B淋巴细胞百分比大于正常组(P<0.05),益肺逐积方组、CTX组、联合用药组B淋巴细胞百分比低于模型对照组(P<0.05),益肺逐积方组B淋巴细胞百分比大于CTX组(P<0.05)。结论益肺逐积方联合CTX可有效抑制Lewis肺癌肿瘤生长,并通过上调胸腺指数以及CD_4^+T、CD_8^+T、B淋巴细胞百分比,改善CTX的免疫抑制作用。

Objective To explore the role of yifeizhuji decoction(YFZJD)combined with cyclophosphamide(CTX)in the treatment of lung carcinoma and immune regulation based on observing its effects on the tumor volume,thymus indices and percentage of lymphocytes in the Lewis lung cancer mouse model.Methods We conducted this study from April to May 2016.55 male C57BL/6 mice were selected,5 of them were inoculated with Lewis lung cancer cells to prepare tumor cell suspension,other 50 were randomly divided into model group(40)and normal control group(10).Model group mice received subcutaneous inoculation of Lewis tumor cell suspension in the right forelimb armpit to establish the lung cancer mice model.When the models were successfully developed,mice from model group were divided into YFZJD subgroup(10),CTX subgroup(10),YFZJD combined with CTX subgroup(10),model control subgroup(10)by means of random number table.From the second day after modeling,YFZJD subgroup and YFZJD combined with CTX subgroup received a 14-day sequential therapy of intragastric administration of YFZJD(14.95 g/kg per day),and the model control subgroup and the normal control group received a 14-day sequential therapy of intragastric administration of 0.9%sodium chloride solution(with the volume equal to that of YFZJD per day).The CTX subgroup only received and YFZJD combined with CTX subgroup additionally received intraperitoneal injection of CTX(40 mg/kg)at 72 h and on the 10th day after inoculation.At the end of intervention,the tumor diameter was measured and the tumor volume was calculated;the body weight and thymus weight were measured,the thymus indices were calculated;the percentages of CD4+,CD8+T-lymphocytes and B cells were detected by flow cytometry.Results Smaller tumor was found in YFZJD subgroup and YFZJD combined with CTX subgroup instead of the model control subgroup(P<0.05),and in the YFZJD combined with CTX subgroup instead of CTX subgroup(P<0.05).The normal control group had greater body weight compared with other 4 subgroups(P<0.05).The YFZJD subgroup had greater thymus weight and higher levels of thymus indices than the normal control group as well as the CTX subgroup(P<0.05).The percentages of CD4+,CD8+T-lymphocytes in the model control subgroup were less than those of the normal control group as well as those of other 3 subgroups(P<0.05).The YFZJD combined with CTX subgroup owned higher percentages of CD4+,CD8+T-lymphocytes compared with the CTX subgroup(P<0.05).The model control subgroup demonstrated a higher percentage of B cells compared with the normal control group and other 3 subgroups(P<0.05).The percentage of B cells in YFZJD subgroup was higher than that of the CTX subgroup(P<0.05).Conclusion The combined use of YFZJD with CTX can effectively inhibit the growth of Lewis lung cancer tumor and improve the CTX-induced immunosuppression by upregulating the thymus indices as well as the percentages of CD4+and CD8+T-lymphocytes and B cells.